#299 ‒ Optimizing muscle protein synthesis: the crucial impact of protein quality and quantity, and the key role of resistance training | Luc van Loon, Ph.D.

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My guest this week is Professor Luke Van Loon. Luke is a Professor of Physiology and exercise and the head of the M3 research unit, which is a part of the Department of Human Biology at the Faculty of Health, Medicine and Life Sciences at Maastricht University. He is internationally renowned for his research that is focused on skeletal muscle metabolism in humans, and he is focused on four main fields of interest which include skeletal muscle metabolism, exercise metabolism, sports and clinical nutrition and aging. I first came across Luke when I saw a video from a lecture he gave many years ago, and it's not common that I'm watching a video of somebody talking about protein where I'm actually stopping and watching it for great lengths because I'm actually learning something. So I immediately became hooked, became more and more familiar with his work, and ultimately wanted to have him on the podcast. Luke received his Ph.D. from Maastricht University in the Department of Human Biology, subsequently did an internship at the Department of Kinesiology and Health Education right here in Texas at the University of Austin, though long before I got here. And then he did a couple of postdoctoral fellowships, one in Australia and one back at Maastricht University. He is also the Associate Editor of the International Journal of Sports and Exercise Metabolism and is on the editorial Board of the European Journal of Sports Science. In this episode, we talk about the role of insulin and glucose for endurance exercising. We talk about the role of protein in all of this. We speak about how different types of proteins in different forms will foster muscle protein synthesis in different rates. So we talk specifically, of course, about the absorbability, digestibility, amino acid quality and other features there. Of course, we talk again in length about some of the different types of protein. First of all, the difference between animal sources and plant sources, but using probably more helpful designations than that, we get into even specific types of proteins. So for example, even if you're talking about milk based protein, what are the differences between whey and casein? And we also talk about the use case, if at all, for collagen protein. Talk about how protein digestion is impacted not just by the type of food, but even by the preparation of the food. And we cover the relationship between activity, lean mass building muscle and protein from resistance training and the role that protein specifically plays in that in terms of timing and type of protein. So without further delay, please enjoy my.

Conversation with Luke Van Loon.

Luke, thank you so much for being here. I know it's evening time for you and as we were talking a moment ago, I think there's a decent chance we're not going to get through all of this. You laughed when you saw my agenda of topics. So maybe we will have a little Austin reunion. We'll do the next one in person here in Austin. But let's give folks a little bit of a bit of your background. You had quite a storied resume. Can you give us a little bit of the highlight?

Yeah. As many people I didn't know where to go to college. As most of us exercise physiologists, we're all field athletes. So we want to know how our genetics can be actually just compensated for by science. And so I wanted to do movement sciences. So I studied movement sciences here in Maastricht, the Netherlands. After that I went to Austin, Texas for my master internship to work with Jack Wilmore, who people in exercise physiology will obviously know at UT. After that I finished did my PhD in Maastricht again. Then after that I went to Melbourne to work with Mark Hargraves, also well known to most people in this field, spent some time in Melbourne and then came back and actually did the rest of my career here in the Netherlands. So from assistant Prof. To associate and to Prof. Here. And that's already been like, I don't know, 13, 14, 15 years ago today.

We'Re going to not talk so much about the movement stuff, although that would be obviously very interesting and our audience, I think, would appreciate that as much as what we're about to talk about. But we're going to talk more on the nutrition side of things. We're going to talk specifically and in great detail about protein. But that's, if I'm reading your CV correctly, not necessarily the first foray into nutrition that you took, correct?

No, that's correct. So my main interest was in the beginning, and also my PhD was in fuel selection. So substrate selection during exercise. And of course, most of that stuff is done during endurance type exercise. And if you're thinking about substrates, you of course are not thinking about protein, which is, from a quantitative point of view, is not a very good substrate. So it's carbohydrate, fat metabolism. And most important or most interesting thing for me at that time was that they had set up a stable isotope research facility here so you could track metabolites. The first things that that lab did is actually measuring carbohydrate oxidation rates. So you would simply throw in some stable isotope labeled carbohydrates in your drink and by simply the oxidation of those carbohydrates, and you would actually expire, exhale 13 CO2, because then if it's labeled with 13C, you get 13 CO2 after you oxidize it. And by simply acquiring your expired breath, you can actually calculate how much of your energy came from a sports drink. Now, that was so cool for me at that time.

Just to be clear, you would also need to know the amount of oxygen consumption to be sure that that CO2 came from glucose and not from fat, correct?

Yes. So it's a combination of indirect kilometry. So that's total oxygen uptake, total carbon dioxide production combined gives you total energy expenditure and also total oxidation of carbohydrates and fats. And then because you know what purpose percentage of the carbohydrates derived CO2 was expired by 13 CO2, you could calculate back how much of the carbohydrate is coming from your drink. And that is cool, because then you see how much of the drink are you actually using.

Now, it's interesting, Luke, that there's a method obviously, that uses deuterated water and C13 doubly labeled water to measure energy expenditure in a free living environment over a long period of time. Is the method you're referring to only using labeled carbon and therefore only suitable for a short period of time to evaluate glucose oxidation. Or do you also label oxygen and potentially get energy expenditure over a long period of time?

If you use doubly labeled water, you can actually do that over several days. You need several days in order to do that. So this is really a way of you're getting carbohydrates in, you start oxidizing them. That CO2 is also mixed in your blood. So it takes a while before the expiration of labeled Carbon is also a good proxy for the amount of carbohydrates that you're oxidizing. So it takes about two hours. Then you have a nice steady state. That's why we're measuring endurance athletes, of course. And then for one or two hours, you can measure the oxidation rate. So that's only for exercise trials, for example.

And did this tend to be more accurate than the estimate you would get of carbohydrate oxidation rate just using the indirect calorimetry and the Weir equation? Or was the point here to determine not total carbohydrate metabolism, but specifically how much is coming from the drink?

Exactly, exactly. So the basis is still in the red kilometry, so respiratory exchange ratio. And then from the carbohydrate oxidation, you can see how much of that carbohydrate is coming from your drink.

Got it. Versus glycogen, for example.

Exactly. So you have your glycogen coming from your liver, you have your glycogen coming from your muscle. You can actually detect that with an intravenous glucose tracer with a different label. And then you have your 13C label in your drink. So you can actually see all three substrates. So you can see what is coming from the drink, what is coming from your muscle glycogen, and what is total coming from your plasma glucose, which includes, of course, the drink. But you can subtract that from total, and then you have all three different sources. And of course, that was not enough for us. So we also wanted to know what fatty acids are being used from your intermuscular triglycerides. So the fat inside your muscle fibers and what has come actually from the fat tissue that is releasing fatty acids and then transported to the circulation and taken up in the muscle, because that is also an important topic for athletes, but as we will probably touch upon later also, for example, diabetes patients.

Well, it's not going to be possible for me to just leave that one alone, Luke, and jump into protein, because I got to understand, especially for my own selfish needs, as I'm preparing for kind of a long distance endurance thing. I thought my long endurance days were behind me, but I do have one more very long endurance event that's going to take about 20 hours coming up in a few months. And it's been so long since I've thought about how to prepare energetically for something like this. And I really believe, at least based on my history, that nutrition can make or break you in these 20 hour marathon events. So tell me some of the things that you learned in studying, even though I know you are only studying it only, you know, over two hours. Hours. But what did you learn about, for example, the rate of carbohydrate metabolism, muscle glycogen, liver glycogen in the steady state.

So I thought we already had quite.

A list, but I think just added something to it.

No. So one of the most important things is basically all the glucose that is extra coming in your circulation will be used for oxidation during exercise. So a continuous supply of glucose coming from your gut is a good idea to, to save your liver glycogen, to keep your liver glycogen intact, or at least not to deplete it too fast. And so if you can maximize that. And of course we know that with glucose or glucose polymers, that is about at a rate of 1 to 1.1 grams per minute, so about 60 to 70 grams per hour. However, it can be a little bit higher if you actually add some fructose, because fructose requires a different transporter. So if you combine both glucose or glucose polymers with fructose, you actually can get a little bit higher, but that's only for really the high end athletes that can actually gain that high energy expenditure. Then you can maybe upcode to 1.3, 1.4 grams per minute. And so that would be the ideal from the carbohydrate point of things. But of course, if you're doing a 20 hour endurance event and you're well trained, you'll actually be using quite a lot of fat because that's what you're training for. And in that case you require your fat stores in your muscle as well as in your. Yeah, I mean, you already have enough fat stores in your fat tissue, of course, because theoretically, if you're 70 kilogram weighing lean men, you can run for almost. I think so. So would it be probably three days or something like that, 24 hours? Of course that is only theoretical, but it's obvious that the amount of fat inside your skeletal muscle tissue is also very relevant because, I mean, you know, you appreciate that your muscle glycogen storage increases as you become a better endurance athlete or better athlete on itself. Same goes for intramuscular triglycerides.

Just to clarify one thing, Luke, because people who listen to this podcast may remember, gosh, it was a couple of years ago, I had Gerald Shulman on from Yale. Gerry's one of the world's experts on insulin resistance, and we spoke there about the mechanism by which intracellular fat, or it was actually diac Glyceride, not triacylglyceride, within the cell, was one of the hallmarks of insulin resistance. But I think you're talking about something different here. Are you talking about triglyceride between the muscle cells and not in?

No, this is actually. I mean, you will actually appreciate that as a physician as well. So a lot of the clinical work is done by clinicians and people that focus on the clinical work. The exercise physiologists focus on the exercise work and sports or sports supplements. You do notice that a lot of work that has been done in the two fields does not get used by either side. So when I was doing substrate metabolism, muscle and the selection and how that is organized and regulated, first I was interested, of course, in carbohydrates, because during moderate to high intensity exercise, from a quantitative point of view, that's the most important substrate source. And so as exercise intensity increases, you depend more on your endogenous carbohydrate stores. But what you can see is doing moderate intensity exercise and lower much of the fat is actually being used by athletes. And that increases also during high intensity exercise as you become a better athlete. So as a good athlete, you get the same exercise intensity, even the same relative exercise intensity, you can oxidize more fat, and that is the benefit because you need less of your limited stores of carbohydrates. Now, the body of an athlete or the muscle of an athlete adapts to store more glycogen, but it also adapts to store more fat inside muscle fibers. And that's myocellar lipid. So that's really inside the cell lipid droplets that you see in the muscle fibers. So in these days, I was actually cutting muscle biopsies up, looking under the microscope, and if you actually see them, you see those individual fibers, a lot of lipid droplets. It looks like you're looking at a soup from the top and you see those little droplets floating around. Now that is a storage that is ample, used by athletes. And Hans Hopler in those days was one of the first to do electron microscopy pictures of that. And if you then look inside one of those lipid droplets, you see that on most of those lipid droplets, there's a mitochondrion attached to it. It's like a backpack. It's like the mitochondria is the backpack of a big lipid droplet. And so we started, and I think we were one of the first to start measuring how much of that lipid inside the muscle, inside the muscle fibers is being used during exercise. And we did that using those stable isotopes indirectly, but also by taking muscle biopsies before and after exercise and looking at the number and the size of these small lipid droplets, and that goes down. So athletes use a lot of the intramyocellar lipid for their energy provision, and particularly in the beginning of exercise. And then you think like, hey, this makes sense. When you start exercising, all your lipolysis in your fat tissue takes time to get that process going. Perfusion lipolysis.

Yeah, this is almost immediate access. Prime the pump. Yeah. So just to be clear, I was never aware of this, that you could actually see mitochondria on the fat droplets within the cell of the muscle. Would this be a fundamental distinguishing feature between the muscle cell of an athlete and the muscle cell of somebody with type 2 diabetes, which will also have fat within the muscle? Will it simply not have the mitochondria and therefore not be turning to it as an energy source out of the gate?

Very likely. I've never seen electron microscope pictures of that because it's difficult to quantitate that because there's so many lipid droplets. So you can't make it quantitatively. But we actually, and exactly what you said. There were also other people in our university and our department looking at diabetics. And then I started to be interested in, hey, what's the difference? Because if you compare an endurance trained athlete and a diabetic, they both have huge intermoceller lipids. And the difference is the athlete uses it, depletes it, builds it up again, but the diabetic doesn't. And if we actually exercise the diabetics, they were not really using that intramyocellar lipid. And that makes perfect sense because why? Because the blood is full with free fatty acids. So the muscle is not going to use those limited stores in the muscle because it's constantly getting free fatty acids from the circulation. So in the diabetics it's a permanent storage with very little turnover, while in the athletes, it's a substrate source that is constantly being used. And so the use, the turnover of that substrate is the important factor driving towards insulin resistance or actually insulin sensitivity. Yeah.

I remember reading a paper 12 years ago that very erroneously came to the conclusion that athletes on a ketogenic diet who had high amounts of intramyocellular lipids were insulin resistant. And I was shocked because I couldn't believe they could fail to look at turnover rate and actually differentiate a static from a dynamic process. This is obviously a More important example and a more relevant example.

Yeah, One of the fun things that we actually did is we saw that athletes in the beginning use a lot of intramyocellular lipids, but in the second stage of their endurance exercise they stopped using them. And then we thought like, strange, why is that? But it actually happens when you're free fatty acids in the circulation go up. And it makes sense, of course, by the time that your adipose tissue is constantly actually feeding free fatty acids, you're not using that storage depot anymore in the muscle. So as soon as free fatty acids went up, the use of intermyocellolipids went down. And then we thought like, hey, what would happen if we reduce the free fatty acids in diabetics during exercise? And we actually gave them a Sipomox. So it's something that blocks adipose tissue lipolysis. So in the diabetics the free fatty acids would not go up anymore, they would actually go down and they suddenly started using intermyocell lipids. And that improved post exercise insulin sensitivity. So it's a very nice way to stimulate the turnover of the intermyocell lipids. And it's probably one of the mechanisms that makes insulin sensitivity improve after exercise.

What role does insulin play in accessing the intramyocellular fat store? So obviously insulin plays, along with maybe hormone sensitive lipase, the most important role in determining the rate of esterification versus lipolysis in the fat cell. And obviously we know the role insulin plays for glucose into the muscle cell. Obviously there is also non insulin dependent glucose uptake. But where does insulin if at all factor into both the athlete and the diabetic with respect to how fat is getting into and out of the muscle or into the muscle and then oxidized?

So insulin of course also stimulates fatty acid uptake. But if we talk in an exercise setting, insulin doesn't play a whole important role anymore during and immediately after exercise because the stimulation of your AMPK pathway also stimulates glucose uptake. As you know that GLUT4 transporters are actually translocating to the outer membrane. Insulin dependent or exercise dependence, Two separate pathways for fatty acids. It's even a lot more difficult because the free fatty acids, a lot of it is actually facilitated diffusion.

Yeah, I was about to say because the membrane is fat soluble. I was going to ask you how. I assumed it was just going to diffuse through with a gradient.

A very chance finding. We were just a lucky finding on the side of the study when we were measuring depletion of Intramyocell lipids in the leg in a non invasive way using Mrs. Magnetic resonance spectroscopy. We did that in the leg, but we just for fun, we also measured the arm. And of course you're not cycling with your arm. We did an MRI before and after exercise. And of course the fat in the muscle went down in the legs, but it actually increased in the arm. So simply the greater turnover, the greater flux of free fatty acids in inactive muscle, it actually increased the amount of intermyocellipids. A lot of it is not regulated. Yeah, the muscle is not that intelligent. The muscle just takes what it can get. It's very opportunistic.

Yeah. This is incredibly fascinating. What do we know about the endurance athlete's capacity to consume fat and during a steady state exercise? So for example, if a person is doing. Again, I'll just be selfish and use my own example. Right. If I'm doing a 20 hour activity and let's just say energy needs are going to be relatively low because the energy expenditure is. Let's just say we're requiring 500 kilo calories per hour. So we could, you know, almost meet that through ingested glucose. That would be 1.25 grams per hour. But you know, truthfully that would be sort of punishing on a GI system for 20 hours. So you'd want to sort of potentially maybe limit that to 30 grams per hour of carbohydrate, which would meet about half those needs. Would you say, look, you're very likely going to need nothing else because the remainder of that will be met through endogenous fat stores and a little bit of endogenous glycogen. Or would you say no, you could prime that with a little bit of additional substrate that is presumably fatty acid.

Yeah, I mean that's a difficult question, of course, because nobody's done a lot of work in long term, I mean that long term endurance type activities. But after I've done, and this is always fun, if you do nutrition research first, you start with carbohydrate related research because I thought the be all and end all of everything is liver and muscle glycogen. But then you start looking at, oh, the intramuscular triglycerides are an important substrate source as well. What happens if you deplete them after a few hours of exercise and how fast do you replete them? Because if you were to do a next session with depleted intermuscular triglycerides, you wouldn't have that prime that you're referring to. So repleting that is also important. And then we realize that only carbohydrates is not enough. You also need fat. And of course now that I'm actually doing a lot of research in protein, we now know that we also need protein. And then we're back to food. But that's another discussion.

Let's talk about that. Let's talk about your transition. I could sit here and pick your brain all day on the carbon and fat issue, but how did your own interests transition you towards protein metabolism?

So just going slightly back to your last question because I think you're asking also a practical question for yourself. So when we saw that fat was important in the muscle, we started also looking at post exercise fat depletion. So you have enough fat to provide you enough fatty acids during that long the 20 hour endurance event. But I think you should not start doing that with too low of an intramuscular fat level. And when we started talking with ultra endurance athletes, they were actually proving me right. But even though there's no data, they were saying like, look, I can't really perform in several day events, so multi day events if I don't eat enough fat. And that's maybe because of the intermised cell lipid storage that you want to replete between several days and it takes one or two days to replete it. So if you do more of these 20 day events back to back, then the fat repletion is important for a single event. I would think it wouldn't be that important.

Do you have a sense we do tend to quantify glycogen capacity? We would say maybe a healthy, athletic, reasonably sized male would have 100 grams of glycogen storage capacity in the liver, 3 to 350 grams in the muscle. I'm sure there are athletes even higher. Do you have a sense of what those numbers are like in the muscle for how many grams of fat could be stored in the muscle of a healthy athlete?

No, it was way less than that. I think there were estimates and now I have to rethink because I'm going back 20 years. I think it was also around like 100, 200 grams.

But that's still a lot because of the caloric density of fat being higher. I mean that's functionally as many calories as you would have from glycogen.

Yes, but I think the calories are not important. The most important but the priming dose to actually cover from activating the adipose tissue towards having enough free fatty acids in the circulation. And that takes about half an hour, 45 minutes. And that's where it's important.

Last question on this topic. I know I said we would leave it, but this just reminded me of something else, which is what does the window of time look like in which an athlete has the greatest opportunity to replenish intramuscular fat stores and intramuscular glycogen? Presumably it's a window following exercise. How long does that window remain open?

The first, I mean they don't know exactly, but within about the first four or five hours we see that insulin, that glucose uptake is less or almost not insulin dependent. So basically the more carbohydrates you ingest, the more it goes into the muscle. But then as your glycogen levels increase, then you get a break on glycogen deposition. So glycogen storage is inhibited simply by its content. So it's a self limiting process, which is great because if you don't, you actually have a muscle disease. So the first few hours, all the GLUT4 transporters are in the sarcolemma. So all the glucose that comes into circulation gets sucked into the muscle, meaning.

Just the contraction of the muscle alone during exercise is getting that GLUT4 transporter into the membrane without the need of insulin. And it's open field running for glucose coming into the cell.

Exactly. And that's one of the reasons why exercise is so good for maintaining glucose homeostasis. It's not only the total amount of muscle, but especially the way you use the muscle. I mean, the easiest way to actually cruise through your or glucosturrence test is before you go to your gp, you actually run for two hours because then you'll have the lowest OGTT that you'll ever see.

Yeah, I did that actually once just as a test. I wanted to see how insulin sensitive I could make my muscles before a 2 hour OGTT. And I don't remember the exact results, but it was astonishing. Glucose at the start is maybe 80 milligrams per deciliter, 30 minutes in an hour and two hours later it never went above 100 milligrams per deciliter in glucose, which is. But also to your point about the insulin independence, I don't think my insulin went above 11. Even a fasting insulin above 11 is not uncommon. So yeah, that's a very interesting experiment. What about the fat window, is that also about a four hour window for.

Those of you interested in that insulin effect, I mean that goes up to 24 to even. People have shown 48 hours after. And that is the reason why nowadays for exercise for diabetics, they say 150 minutes of exercise throughout the week. I would think that's a minimum. That's the advice. But at least every other day, and at least every other day is because every session improves your glucose homeostasis for up to 24 hours. So if that exercise is actually every other day, you maximize benefit from the exercise with each and every meal that you ingest afterwards. For fatty acids, it's actually less difficult because it doesn't seem to be tightly regulated because of the facilitated diffusion of your fatty acids. But of course, if you're an athlete and you're consuming a huge load of carbohydrates, especially also during exercise, of course you're going to limit your lipolysis and then you have less free fatty acids. So the only thing that we really did there doesn't seem to be a short frame, it's just constantly going. It seemed like up to about 48 hours the fatty acids for the intermocell lipid was almost at the same level again.

And just to be clear, are you saying that most of the filling of the muscle with fat will come from endogenous fat stores via lipolysis, or is there a role for exogenous I. E. Dietary fat in the post exercise phase to boost that further? And I understand your point, but I want to make sure the listener understands that the relationship between carbohydrate ingestion and lipolysis is very clear. The more carbohydrate you ingest and the higher insulin goes, the more you inhibit lipolysis, which is the breakdown of fat from the stored fat cell. But with that explanation out of the way, does my question make sense for you about the source of the fat?

Yes. So in a well trained athlete, about 50% of the fat oxidation during exercise comes from your intermuscular triglycerides and the other 50% comes from free fatty acids released from your adipose tissue transported to your blood. Taken up. Now the only studies that we've done here was with fat supplements during exercise because there's not much reason to take fatty acids because there's enough fatty acid being released from adipose tissue. We did play around and that was Oscar Jurgen Derp actually playing around with medium chain triglycerides. So medium chain triglycerides are actually smaller fatty acids. The tail is less long and they can be actually transported directly into the mitochondria. So they don't need the CPT1 facility of transport. So we thought it was nice to get a faster oxidation labeled substrates. They show that the medium chain triglycerides, even during high intensity exercise are really oxidized. And the idea was that could save carbohydrate use so that you can maximize fat use during high intensity exercise and therefore spare glycogen and improve performance. Now, basically it worked with mcts, but then we tried to see whether we could make it quantitatively interesting and then almost everybody got diarrhea.

Yeah. I was about to say the big challenge with getting sufficient enough MCT volume is the gastrointestinal distress. And I think for most people at least, if consuming it in a pure oil form, 30cc is really pushing it. I mean, when I was hard on MCT oil back in the day, this is 12 years ago, I could do 2 tablespoons of MCT. So whatever, how many ever milliliters that is. That said, and I haven't done this, but I do see other products out there that are somehow lawfulizing the mct. They're almost putting it into a powder form. Have you seen any of these things?

Yeah, I've seen suggestions that there's a lot of MCTs or MCT like products that presumably have less issues on the gastrointestinal tract. Yeah, I haven't used them and I haven't used them in research. Still. I would contemplate like in what amounts do they actually contribute substantially so that it improves performance. And I think as long as you can actually reach your goals with carbohydrates, why would you play around? But yeah, is there a role for.

MCT oil immediately following exercise as a very quick rapid source of fatty acid in the muscle? Or are you saying no because right after exercise your insulin is so low, just let the lipolysis fill the tank. Why would you bother adding we all want to lose a little bit of adipose tissue anyway, right?

Yeah, I think so. I agree. I'm a nerd, of course. So I love to see what is happening, but I don't think it has any relevance in practice.

Okay, so with that very interesting detour out of the way, let's talk about how your professional interests pivoted from this obviously interesting and relevant field to another.

I think one of the interesting things was when I saw they called that the athlete's paradox. High intramyocellular fatty acids in high intramyocellular lipid stores in diabetic and obese people and also in athletes. So what is the difference between them? So I started doing some diabetes related research and exercise work in diabetics to improve insulin sensitivity and improve substrate metabolism. And then we were taking muscle biopsies. My MENTOR Back then, Dr. Hans Kaiser, he was actually teaching me how to take muscle biopsies. And he's a physician, so he was taking biopsies. And what we often saw is that when you took a biopsy from an athlete, it's like a chunk of muscle sitting there, like a piece of good steak. But then if we took biopsies from the diabetics, often sedentary people, it was like a blob. It wasn't standing up. It didn't have any structure. It was just like liquid. And then we always saw that, and we were discussing that, and it was obvious that diabetes or a more sedentary lifestyle comes with changes in muscle quality. And of course, if you start thinking about muscle quality, you start thinking about protein synthesis and protein metabolism. And because I was working with older people, because the diabetics were older than my athletes, this is much more interesting than the athletes. What is happening with that muscle. And then you automatically go from only substrate to protein metabolism. And, yeah, that's where I got stuck. And still haven't figured everything out, or actually, I have more questions than I ever had before.

Well, I think that's the mark of a really fun place to be, truthfully. Let's give folks some basic background. Let's not say any more about carbs and fats for the moment, because we could spend hours just defining them. But I do think people at a high level clearly understand that if you include ethanol, there are four major macronutrients. But let's talk about this one called protein. What is protein? What are the building blocks? What's it made up of? What defines a protein? Which ones can we make? Which ones can we not? I'll let you run with that in any way you see fit.

Protein is one of the three micronutrients. Protein consists of amino acids. Amino acids are the building blocks of protein. And so they're also the building blocks of our cells because all our tissues is mainly protein. Of course, we have, say, about 20, or, depends how you define it, 20 or 22amino acids, of which nine are essential. The rest is non essential. The definition of essential is always confusing to my students because essential means we can't make ourselves. So there's no endogenous synthesis in our bodies in humans, whereas the non essential, you can actually to some extent synthesize. Now, I'm probably mucking it up for everybody, because what I always say to the students, that doesn't mean that your non essential amino acids are not essential in your diet. And then they never differentiate between them anymore. Because as far as I know, I reckon that it's almost impossible to synthesize those non essentials. That is something that is still a topic that I would like to address. But that's difficult in humans, of course, because you can't simply just throw away all the protein and start feeding amino acids.

And maybe we'll just also just so people kind of get a sense of these things. They're called amino acids because they have a very clear structure. And I have to be honest with you, my biochemistry is so rusty, but I sort of remember them having. Was it. Did they have a carboxyl head and a nitrous tail with a carbon in between and then organic acid? Yeah, yeah. So it's an organic acid with this nitrous tail. So nitrogen is the big piece here that we don't see in carbohydrates and fat. Carbohydrates and fat are basically carbon, hydrogen, oxygen. Here you have carbon, hydrogen, oxygen, but you also have this big piece of nitrogen. So I'm sure we will talk about nitrogen balance and then you have other things thrown in there. Methionine has a little sulfur in it and things like that as well. But I guess the point is biochemically they all have a similar backbone. But what differentiates the 20 or 22 of them is kind of the what's in the middle. That's the special sauce for each one. And that's what differentiates them.

That's what differentiates them. Yeah, that also defines their individual characteristics. Now we see that amino acids are important as our own building blocks, of course, and so that's why we need to consume protein, because they provide us with those building blocks. But what is interesting, and we'll come to that later, is that those amino acids are more than building blocks. They're also signaling molecules that directly stimulate muscle protein synthesis. So they directly activate the MTOR pathway driving muscle protein synthesis. And so it's interesting and I always explain it to first year students. You have a building site where you actually have bricks being delivered and the bricks themselves pick up the phone and call the bricklayers to come over. So it's really amazing how that works. So simply eating, and I'll come to that later, stimulates your muscle protein synthesis. Now, of course, I'm a muscle physiologist, so most of my work is on muscle. We've been starting to work also on different organs, because this is of interest and there's not a lot of data there available. But what we do need to know is that you're synthesizing about 300 grams of protein on a daily basis. That includes tissues, that includes hormones, that includes enzymes, blood, proteins, everything together. It's about average estimate, about 300 grams. As most of the listeners will consider is that they are consuming about say 70 to up to 100 grams of protein per day. That means that for 70 grams, for one gram a day, for 70 kilogram weighing men, that would be your recycling. Two hundred and thirty grams. And this is something that people don't realize. So yes, we consume foods and we need those building blocks. But you're also constantly using amino acids that are being released from the breakdown of tissues. And then you use them again, you're making 300 grams, you're only ingesting 70 grams. So that means on a 24 hour basis you're recycling 230 grams of amino acids from your own body. So you're very sustainable.

Let's talk a little bit about the use case for it. Right? So 300 grams being utilized per day. Let's talk about how that differs in an active person, a person who's strength training, a person who's doing steady state cardio training, a person who's not training. Let's just use those three examples.

I don't think anybody would have data on that because this is on the whole body level. So organs, tissues, the guts, everything. And I think if we're asking these questions, we have to just move back to a muscle centric perspective.

There's a question I want to ask before all of this, which is a technical question that explains to people the methods that you use to actually do these types of measurements. You already talked a little bit about one isotope method with respect to carbohydrate metabolism. Can you explain the technical tools that permit this type of study for amino acids?

I'll keep it very simple. You have stable isotopes. So everybody knows from high school, or most of you will know from high school, that in chemistry you have carbon 12 or carbon is weight of 12 in nature. There's also carbon 13 and carbon 13 means it has an extra nutrient in its core. It's stable, doesn't fall apart. You also have 14. Carbon that's unstable, falls apart and you add radioactivity and that's why it's a radioactive isotope. Now you can actually purchase carbon 13 labeled metabolites. And so you can also buy carbon 13C carbon 13 labeled phenylalanine, for example. Nowadays we often use phenylalanine as the tracer because it's an essential amino acid that is hardly metabolized in muscle or not metabolized in muscle, and it's not as volatile as, for example, leucine. And leucine, of course, has a lot of other effects. So you want to have a nice stable amino acid that is a good representation of other essential amino acids. So if you actually buy 13C labeled phenylalanine, you can dilute it in a bag of saline, infuse it in the body, and then you actually have like 5% of all your phenylalanine floating around in your circulation. And then you take a muscle biopsy. Then you take the muscle out of the biopsy, you take the protein out of the biopsy. You can even separate mitochondrial protein or myofibrillar protein, so that's actin and myosine, for example. And then hydrolyze that protein back to its free amino acids, and then use mass spectrometry to look at how many of those amino acids in the muscle protein are coming from the carbon 13C labeled phenylalanine. Now, a few hours later, you take a new biopsy and you see how much more of that 13C phenylanine is built into the muscle protein. If you divide that by the availability and the circulation and the time between the two biopsies, you have a fractional synthetic rate, generally expressed in percent per hour. Now, on a daily basis, that's about 1 to 2% per day. So it's always confusing to, or confusing. It's always magic. To me, that means if all the proteins in your muscle are generic, which they aren't, of course, but assume an average, then they are actually completely refurbished in 50 to 100 days. So I always ask people to look at their own arm and realize that in 50 to 100 days they have a new arm. It's completely refurbished.

That is amazing.

So the individual proteins in the muscle are broken down and build up again. And we can measure that by infusing those labeled amino acids and simply measuring their synthesis rates.

First of all, that is an absolutely amazing idea that is, of course, self evident based on the nature of what we know about protein turnover, but when phrased that way is simply remarkable. Right? In other words, we don't take it as so earth shattering that every three months our entire hematopoietic system turns over. I mean, we know this, we know that every 90 days a red blood cell is gone and it's replaced with a new one. Effectively. The entire red blood cell architecture in my body today is completely new from what it was three months ago. But when you talk about it through the lens of muscle, it's much more complicated. I mean, it's just much more difficult to wrap your mind around. Obviously one of the questions is, but I don't notice any difference what preserves the architecture. This gets to what is being replaced and what is technically not being replaced. How does the shape change so that we don't look different, so that we're not a constantly morphing creature?

So you have more questions than I can ever answer. But that is also. And we'll come back to that later, I hope. So we started also measuring turnover rate of tissues other than muscle. And then it gets even more freaky. And then you start really thinking like, how is this regulated? Let's not do that because it will be confusing. But we've also measured protein turnover in the brain, in the human brain. And then if you see those turnover rates, then you have different questions. But let's not go there. Let's think for the muscle for now. So it means that in order to keep the muscle and maintain muscle mass, you need to stimulate it because the breakdown will continue. And so in order to maintain your muscle as it is, you actually have anabolic stimuli that stimulate the synthesis. And then we come back to nutrition and exercise because it's food intake, and particularly of course protein intake and exercise that stimulate muscle protein synthesis. How these two stimuli know which proteins to build and in what structure. Oh man, that is just an enigma. I always show a picture of Lance Armstrong and I don't know who am I showing as bodybuilder. Actually that changes every so many years. One of the Mr. Universes. And so I showed them that these two guys have huge protein synthesis after an exercise session, but the phenotypic response is completely different. And so with resistance type exercise, you build a lot of myofibrillar protein, while the endurance athlete will build a lot of mitochondrial proteins. So how the body knows by the recruitment of your fibers which proteins should be synthesized? I mean, we know a lot of molecular pathways, of course, but how it's directly regulated, I mean, first we thought we had everything by measuring MRNA's, then we thought we had everything by measuring Western blots, protein content and then protein phosphorylation. And then we went to transcriptomics and then we suddenly knew that we had post translational modification and then we had mRNA Splicing, it's getting complex and complex and complex and complex.

Let's use that example, right? So we'll put up a world class endurance athlete against a world class bodybuilder. And let's even simplify the equation and take drugs out of it. So let's even suggest, let's talk about not the best bodybuilder because obviously they are using high amounts of anabolic steroids, but a world class natural bodybuilder who still by any metric is enormous, and a world class endurance athlete. The question, of course is how does the body know, in the case of the bodybuilder, to build myofibrillar protein, meaning to disproportionately build that. Whereas in the endurance athlete, you're disproportionately building mitochondrial protein. Now, the obvious answer to me seems to be the training stimulus. The bodybuilder is pushing enormous amounts of weight repeatedly within the confines of certain reps in certain sets. And the endurance athlete is never, ever stressing the muscle at a single rep. If you think about it, an endurance cyclist might be 80 to 100, 80 to 100 cadence. And they'll do that. They'll do hundreds of thousands of those repetitions. But none of them are as hard as the reps that the bodybuilder is doing for six to 12 reps. So that's an obvious difference. Is the question that we don't understand how that is translating a signal into the mps.

Yes. I mean, we don't know. And if I would actually pose this question to a second or third year student, they would actually just move away from the real answer by going towards saying like, hey, it's different fibers. And of course that's part of the answer, because a resistance type athlete will more likely recruit the type 2 fibers. The type 2 fibers are more likely to build in mass. If we see people doing resistance training, it's mainly, the type 2 fibers that get bigger. We see that with the bodybuilders as well. Huge type 2 fibers. The opposite happens when we get older. You get smaller type 2 fibers actually. And that is where most of the muscle loss with aging actually comes from. Well, the endurance athletes, we don't see much happening with the size of the fibers. We just see greater capillary density, we see more mitochondria, higher density of mitochondria, sub sarcomal mitochondria. So the adaptive response is completely different. And part of it is because of which fibers you recruit. But it's not the complete answer, because if you do. For example, you probably noticed the discussion about high reps, low reps, low weight, high weight. There's now very nice studies that show if you do high reps with a low weight, if you do the two exhaustion, you also start using the type two fibers and then you also get hypertrophy. And that's a nice way, especially in rehabilitation, that you can actually drive those type 2 fibers without putting too much effort on the. Just operated, the hip or the leg, whatever, just operated on. So it's not the complete answer. There's more going. But what is exactly the signal, the molecular signal from the tension on the muscle towards the synthesis of the specific sets of proteins, there's enormous amounts of pathways involved. And I'm not sure whether we'll ever completely know what is happening now and where that selection is going to be.

And that's actually, it's just so fascinating that we can't understand the cascade that goes from stimulus to muscle protein synthesis. Would it be reasonable to assume that the bodybuilder has more upregulation of MTOR than the endurance athlete? Or is that even a stretch in terms of an assumption, given that it's difficult to measure that?

I mean, we know also now that a lot of the hypertrophy is actually mtor. The stimulation muscle protein synthesis can be mto M dependent. There's so many parallel pathways that are driving responses. And like I said, for example, if you ingest protein, you stimulate MTOR signaling, you stimulate MRNA translation initiation and you get greater protein synthesis. But it actually is not continuously stimulated if you do not provide all the building blocks. So if we, for example, after exercise or even at rest, we provide people with the branch chain amino acids. So mainly leucine is driving that anabolic response. It's a signaling response. You stimulate muscle protein synthesis and for up to two hours. But if we give the same amount of leucine on this case branch chain amino acids in a similar amount of milk protein, so containing the same amount of branch in amino acids plus all the other amino acids we see, that response is actually sustained over more prolonged period of time. So it's not only the stimulation, it's the stimulation in combination with the right amount of building blocks at the right amount of time. How I always tell it to the students, you have a parking lot where you want to park as many cars as possible. You can put the light on green to drive in, but if there's no cars, nothing's going to happen. And so it's a combination of the right signaling Responses, plus the availability of your substrate.

I want to come back to this in detail because it's very relevant. It's also there was a recent study that came out that I believe you were an author on, if I'm not mistaken. Maybe I am mistaken. But the study looked at the duration of muscle protein synthesis following the digestion of different types of amino acids. So we come to that. But let's take a step back for a moment and make sure we at least cover the basics of how protein is even digested. And I want to talk about three types. So one, I want to talk about a whole meal. So a meal of protein that often comes with fat. So let's talk about eating a steak. And they want to ask how that differs from two different types of milk based proteins, whey protein and casein. So I want to understand what is actually happening from a digestive standpoint and how the thing that you actually consume and put in your mouth turns into building blocks that presumably are being absorbed somewhere in the ileum or jejunum.

So I'll just first take protein and then go to the specific protein sources or meals. So protein is basically ingested, goes to your stomach acid, acid is added to it. So you have the first basically start of the digestion. You actually can get clotting of your protein due to the acids. We'll come to that later. If we compare casein versus whey, then it goes to the duodenum, then you have all your enzymes being unloaded on your protein. If they can get access to it. You actually get the free amino acids. A lot of people have suggestions that also oligopeptides, di and tripeptides, so small proteins composed of a few amino acids. There are transporters in the gut that would allow them to actually enter the intestinal wall or the intestinal cells. We think from a quantitative point of view it's hardly relevant, but it's possible. The amino acids are then part of them are actually incorporated into intestinal protein. Because the gut also has a very rapid protein, much faster than muscle. So part of it, it actually remains in the intestinal tissues as protein. Some of it is released on the other side of the intestine in the portal vein. The portal vein transports it to the liver. The liver can do something with the amino acids if it wants to make proteins, but most of it is actually released in the circulation and there it can basically be funded to all those different tissues taken up and used for muscle protein synthesis. We don't have a real storage depot. There's a small free amino acid pool. If you compare it with the intact protein pool, they always say it's very small in muscle. But acutely, after a meal, it can actually vary a lot so that it's a buffer, a temporary buffer. But we don't really have real storage sites for amino acids. So that's the whole pathway.

Let's finish one thing there, actually, Luke, because that's an important point you raise, which is unlike fat, which can be stored in unlimited quantities, and glucose, which can initially be stored in glycogen, although that's a relatively finite store, and then eventually can be stored as fat through de novo lipogenesis. What exactly happens to excess protein if we consume it? And let's just use an extreme example, right? Like I sit you down and give you a 200 gram protein meal, make it whatever number you want, such that the point is you clearly have excess amino acids. Once fully digested, what do you do? What does your body do with those?

Yeah. So first the amino acids, you said that we have no storage like the glucose stored as glycogen. The fatty acids are stored as fat, but the amino acids are stored as protein. But is it a real storage depot? Now, if you ask me and yourself, we hope we're not losing muscle, so we say we don't have a storage depot, but if we end up in a concentration camp, then we're actually quite happy that we have a storage depot of amino acids in the form of muscle. So it is a storage depot, but hope that we're not using it. So if we get excess protein, we thought, I mean, in the literature a lot say that everything that you can't immediately process is being oxidized. Now, that study by Johan Tr that you just referred to that we published a few weeks ago, we showed that with 100 gram oxidation, at least in the first 12 hours after ingestion, is very overestimated. But I mean, over the long run, if you keep continuing eating more protein and in a setting of over calories, you simply store the protein as fat.

What'S the pathway that that happens? Because it's so funny, when I think back to biochemistry classes more than 25 years ago, what I vaguely remember was amino acids could undergo gluconeogenesis to become glucose and then either glycogen or fat. Is that what you're referring to?

Yes.

Okay.

And then it's different for different amino acids. And of course it's a very inefficient way. So that is also one of the reasons why people tend to get less fat accumulation if they overeat in the form of protein. Besides the satiety effect.

Right. Because there's a thermogenic and thermodynamic loss or use of energy just in the metabolism of protein. That itself is obviously beneficial if your goal is to store less energy.

Correct. Now, I have to say something about the techniques that we use because we were interested. I mean, we've done protein synthesis measurements all the time, but at some point, exactly what you were saying. I wanted to know more about the digestion and the absorption prior to stimulating muscle protein synthesis. And in order to understand that you need those stabilized traces not only in an infusate, but you would like to have those labeled amino acids in the food as protein. Now you can't purchase those. So about 15 to 20 years ago, we worked with Eat Marie in France because they already made intrinsically labeled protein. So what they did, they infused a cow with those labeled amino acids. And so the cow integrates those labeled amino acids in the milk and then you can actually use the milk in clinical experiments. Now, we wanted to take it one step further because we wanted to have the number of labeled amino acids in the milk so high, which is very expensive to actually see the digestion and absorption, but even the incorporation in the muscle. So we spent more than €50,000 of tracer. And the first time that's pretty challenging because you actually put it in a cow, you infuse it in the cow and you just hope you you're ever going to see that money back. We put labeled phenylalanine in a cow, the cow made milk, we extracted the milk, we got the protein powder and we used it in clinical experiments to assess digestion, absorption, release, extraction over the leg and incorporation in the muscle. Now, if you see this, I mean, we can now go into the quantitative measure because we actually did this to see the difference between Casey in a way, we'll come to that later, but simply that I bought traces isotopes in the US they send it over. I had it in a small jar on my desk. Then I went to wagening it to in our first in France actually, and we've done it a few times since. You actually infuse it in a cow, you actually get the milk, you bring the milk to a factory to extract the milk protein. About a year later, that same amphenylalanine is now integrated in milk protein on my desk. The same whole molecule that came from the US to my desk, from the US to France, in a cow, then from the cow's milk on my desk again. Then it went into an older subject in a study. We took a muscle biopsy. Now, year later, I have a muscle biopsy on my desk that has that same full amino acid integrated in the muscle. And I actually dissolve it, bring it to the mass spec analysis, and I have that same amino acid in my hand again. That amino acid has been around for four years and went from a lab in the US Through a cow, through a cow's blood protein, through the milk protein, to my desk, back into somebody else's leg. That's amazing.

And just give me a sense of yield. When you made the casein or the whey protein from the cow's milk, what fraction of the phenylalanine was actually labeled with the isotope?

We got 25% of the trace, so we found back in the milk. And so you can imagine as a Dutch person, I'm supposed to be cheap. So losing 75% of that money in a cow is not something that you can sleep, sleep on. So we butchered the cow.

How much of the phenylalanine did you find in the meat of the cow then?

Exactly. So this was really funny because we actually butchered the cow. So we got. Of course it gets difficult because you have to multiply it by total amount of meat. And if you butcher a cow, you don't get all the meat. But we sampled all the organs, and that was a starting point for us to start measuring turnover of other organs in humans. But what was interesting is that different muscles had different enrichment, but it was all ballpark the same. We never calculated whether we actually came back to 75%, but it must have been very close. But we use that meat. And that will come back to your other question, to see whether there's a difference in the digestion and absorption when you ingest meat as a steak, or if that same steak was first put in a meat grinder and you have basically minced meat. Huge difference. With minced meat, you get more rapid digestion and absorption, which should also expedite your muscle protein synthesis, because the more rapid absorption, the greater and faster the release of leucine, the greater the stimulation of muscle protein synthesis. Now, that was a fun study for us to do. And when you were writing it down, I was trying to find in the literature some suggestion that the minced meat versus steak. There was some data to suggest that it improves, accelerates digestion and absorption, potentially improves anabolic response responses. We came up of a study that was done in the 60s that showed that this was epidemiology and has its limitations, that people that have their. Older people that still have their own teeth generally also have their muscle still intact. So that was a nice link to say that your mom has been right on a lot of things because, I mean, I spent probably like 30 million euros of research money in the last 30 years to provide clinical proof that my mom was right on a lot of things.

Meaning when she said, chew your food, chew your food.

But even something as simple as sitting upright, huge effect on digestion and absorption. So all of these things come back. So chewing is a factor that drives the anabolic response to feeding. And so it's really funny then if you go to mixed meals, what you just said, if there's macronutrients coming together, a lot of energy will actually reduce gastric emptying or slow down gastric empty, that is one factor. But the matrix of food, the chewing of the food, all of these, whether you cook it or not, even that is an important one. We'll come to that later as well. I mean, there's so many topics here. We did a study with eating eggs that were raw or cooked. Huge effect on digestion and absorption. I actually tried to have Sylvester alone on that paper, but he never responded.

Unfortunately, Rocky's protein Shakespeare, we basically said.

Was Rocky right or wrong? Because in the first movie, we actually used part of his script. He's ingesting those raw eggs. Would it not be better to actually cook those eggs because you have more rapid digestion and absorption?

I think of how many raw eggs I consumed as a child because I watched Rocky. It's insane how I didn't get salmonella. It's a miracle. Anyway, so it was. Let's go back to the example of the steak versus the ground beef. So ISO quantities, so 50 grams of protein in a steak that obviously will have some fat with it. In fact, let's make them not just isocaloric and isoprotein, but everything about them is the same. So same amount of fat, same amount of whatever one is ground, one is not. Talk me through the kinetics of those two in an individual.

Yeah, so I think the most important factor is, of course, that all the enzymes that are being released in the duodenum and also part of it. Also the gastric emptying, of course, is accelerated with your minced meat, with your ground beef, so you have more rapid gastric emptying. The acid has already had more space to actually have its effect. Faster gastric emptying than it gets into the duodenum you don't have those big chunks of meat, but you actually have those enzymes that can actually attach everywhere much easier so you get more rapid digestion and absorption. It's a little bit similar to for example, casein away. Everybody knows now, everybody, I mean all the athletes that say we need whey protein, but that's because micelle casein, the other 80% in milk, 20 whey, 80% casein. That was actually the first study. That's actually the study why we did the intrinsically labeled protein. We got the milk processed the milk down to micellar casein and down to whey. Because for all those years, especially the companies was telling me that pre digested protein, the hydrolyzed protein, is more rapidly digested and absorbed. And I said, where's the proof? And they said like, yes, this in vitro essay shows. I said in vitro essay doesn't mean anything to me. In vitro essays, you have all these tubes and glass things and, and then they scream for digestion and absorption. But to me, the human in vivo system is much more complex because all those tubes and glass cylinders are actually alive in a human. So I want to assess this in a human in vivo setting. And then they asked me like, how would you want to do it? And then I said like, yeah, I would infuse a cow and then this and that. And then they looked at me like I was completely crazy. Took me seven years until they funded it. And then we got the milk processed micellar casein and whey and we also got the micellar casein and we pre digested it with enzymes. So you get a casein hydrolysate. Now we've all checked them for digestion and absorption and then you see that the hydrolyzed casein and also the whey protein are much more rapidly digested and absorbed than the micellar casein because the micellar casein in the acidity of the gut starts coagulating like spoiled milk or something like that. And so the gastric emptying is probably reduced or slowed down, but also the capacity of the enzymes to basically start digesting the foods. And so that's how we now know that rapidly digestible protein is more likely to stimulate muscle protein synthesis to a greater extent. So there's two characteristics of a protein that is the digestion of it and the rate of digestion. But then you would expect that a pre digested casein would do exactly the same as a whey protein, but it doesn't just as rapidly digested and absorbed, but whey protein still has a greater anabolic response. And of course that might have something to do with the amino acid composition. And we know that whey protein has a higher leucine content. And so if we add, for example, free leucine to casein, we get a greater anabolic response. So obviously the amount of leucine is important factor. So in a nutshell, two characteristics of a protein and their capacity to stimulate muscle protein synthesis is digestibility, the rate of digestion and the amino acid composition, with particularly the amount of leucine in the protein. And anything that slows down the gastric emptying or the release of leucine in circulation will likely also slow down the anabolic response.

So let's unpack that a bit because there's a lot there. We're kind of collapsing digestibility and rate of digestion into one feature. Although can't they be disentangled a little bit? Because the example of the whole meat versus the mince meat, presumably they have similar digestibility, they have just as much sinew, they have just as much fat. But one has been effectively broken down and it would have a higher rate of digestibility, correct?

No, I think with cooked meat the digestibility is the same. It will take a little bit longer for the steak than for the minced.

Sorry, I mean, the rate of digestion, wouldn't that be quicker?

Yes, yeah, yes, yes, yes, exactly. I mean, if you're ever in for sabbatical and you want to do a few years of research in Maastricht, please come over because these are all the same questions that I had and there's so many more. I mean, we as nutrition scientists, we love to work with a single ingredient because otherwise we don't know what is going on. So most of the work that we've done in the past is actually comparing protein X with protein Y and then with protein, I mean, protein isolate or protein concentrate, so protein powders. And that's why a lot of people in the public media just of course read or like to see our studies, because it's about the powders they eat. But of course, normal people don't eat protein powders, protein isolates or concentrates. We eat foods. Now, if you go back to the literature and looking at how many studies have assessed the anabolic response to eating foods, that is very slim because, I mean, nobody's interested in foods. I mean, it's often like, is protein X versus this? Because if you start having meals, everything comes together. Plant based proteins, animal based proteins, the processing of the protein, whether it's heated or not heated, whether it's cut fine, minced, not minced, chewing well, not whether it's combined, how much on the plate, how many fibers. All of these things together actually determine the anabolic response to feeding. And then the digestibility becomes most important. I think 99% of our studies, the digestibility is not an issue because we use extracted proteins. Also when we go to plant based proteins, and that's I guess, also a topic that you want to discuss. Most of the plant based proteins that we have assessed are plant based proteins, and I prefer to call them plant derived proteins that have been extracted from the whole foods. And then the digestibility, it's often still a little less than animal derived proteins, but it's not a major issue anymore. It's a major issue if it comes in the form of whole foods.

All right, let's go through this in some detail because this is super fascinating. But before we do, I want to really ask the question of how much does it clinically matter the rate at which muscle protein synthesis is stimulized? Because what I'm hearing so far, and I want to try to summarize this for both myself and the listener, is that you really have three factors that are going to determine the rate at which muscle protein synthesis takes place. You have the digestibility of the thing that you are ingesting. And this could be the difference between a plant derived protein and an animal derived protein. This could be the difference between a cooked protein versus a not cooked protein. And this could be the difference between two different types of proteins, period. You then have the rate at which this protein is digested. You can picture a curve with the x axis being time and the Y axis being digestion. The rate of digestion will speak to the shape of that curve. And again, I think there are probably many things that impact that. But the obvious examples you've given here would be how the food is cut, and it could be ground beef versus whole beef, how well the person chews. All of those things impact the rate. And then the third factor is the composition of the amino acids itself. You've alluded to leucine many times. We talk a lot about leucine on this podcast because of course it is the number one and the most potent MTOR stimulator. But obviously there are other proteins as well. Lysine, methionine, these are probably more pro anabolic. And therefore the concentration or distribution of amino acids plays a role. Now let's go back to the macro question, which is a person is looking to Maximize, optimize, maintain, increase lean body mass. This should be a goal of everybody, right? It doesn't matter if you're a 75 year old woman or a 25 year old man, the maintenance of lean body mass, if not the increase in lean body mass, is a very important pillar of living a long healthy life. So if that is true, then presumably anything we can do nutritionally, we'll talk about training later, but anything we can do nutritionally to maximize or increase muscle protein synthesis should be good. But the one thing that we're missing is the window over which that happens. So is the goal always to maximize that for the highest peak independent of the base, or do we want more of a time released effect where we say, look, we'll take a lower rate or a lower peak muscle protein synthesis, but I'd like to sustain that for many, many hours. You're laughing, so I've touched a nerve, which means I'm probably asking a question. You are asking, but I'll let you go from there.

This is awesome. I mean, these are all the questions that the field has and that we all have. So it becomes much more difficult because one factor we didn't discuss is the amount of protein, because the amount of protein also has an effect, of course, on the curve and you can compensate with the amount. We'll probably come back to that as well. But yeah, there's estimations up to 2 to 3 grams of leucine in a meal, we'll actually induce a rapid increase in circulating leucine and that will stimulate muscle protein synthesis. And if you have enough building blocks available, you have an anabolic response for at least up to five hours. So this has led most of us in the field to believe that it's best to ingest. At least now we're talking about healthy people. If you become older, you become less resistant to the anabolic properties of amino acids. Anabolic resistance probably also something that we come back to later. But if you're a healthy active male or female, 20 grams of protein is assumed to be the optimal amount to maximize muscle protein synthesis for up to four to five hours after a meal. And this is the reason why in so many packages, you now see 20 grams. Everywhere you see 20 grams. Now, that comes from studies showing that ingestion of 40 grams of protein does not result in greater muscle protein synthesis rates than 20 grams of protein. So it's assumed that 20 gram is the optimal amount in healthy people. And of course, we always have these discussions. If you're 120 kilogram you probably need more. If you're 50 kilogram, you probably need less. Yes, but you can't do a study with every individual in the world. But 20 grams. Now, if you assume that you will have an anabolic response to each main meal, then the advice is there to ingest 20 grams of protein with each main meal. Now if you're becoming older, there's suggestions you need more to compensate for that. But let's stick now with healthy people, that's 20, 20, 20, possibly also an evening protein snack. 20. And then you already have 80 grams of protein, which for most people is already more than 1 to 1.1 grams protein per day. Now, we are people that believe that you actually could use more protein, which automatically happens when you become active, because when you become, you eat more and then you easily eat more than 1.1. So it's a non discussion, but. So the idea is distribution of protein, that every meal is an anabolic response.

Just one question before you go further. This 20 grams is the maximum amount of protein you need to get maximum protein synthesis. Muscle protein synthesis was based on what type of protein?

That was based on milk and egg. So there was a study on milk and there was a study on an egg.

Okay, so it's whole food. It's not using just whey or just casein?

No, it's not whole foods. This is a protein concentrate. So egg protein concentrate and milk protein concentrate. So that's in a very important point because as I said, if your digestibility is not 100%, which in a meal is never, of course, and you have a delay in digestion and absorption, then 20 grams could be suggested to be a minimum.

Yeah. So in other words, what I'm hearing here is for people like me, who don't really like shakes that much, I prefer to eat food as opposed to drink shakes. I do drink them because sometimes it's convenient. But if you're gonna try to get this through food, by definition you're gonna be working with an inferior protein from the standpoint of speed in terms of digestibility and rate of digestion. So you might need a protein meal that's 30 grams or more. And then if you're 90 kilos, you're probably on the bigger end of that anyway. You might be 40 grams that you need per meal of real food. Is that kind of how you would think about that?

Yes. And so automatically also get to that, because we always have those discussions, of course. Because if I say that you need more protein, you ingest more protein, and they look at me like I'm advocating a high protein diet. But these discussions are of course always a little bit weird. If you have a 65 kilogram weighing Tour de France cyclist and he only consumes a very little amount of protein in the form of only 10% of his energy intake, he's still ingesting probably more than 2 1/2 grams of protein per kg body mass per day, even though he absolutely doesn't need it for his limited amount of muscle. The more active you become and the more healthy you are, the more protein you consume. We always have. I mean, World health organization says 0.8 grams per kilogram body mass per day, but yes, maybe you can survive on that, that's fine. But it's certainly, in my beliefs, not optimal. But whether it's optimal or not, it's a non discussion. Every healthy, relatively sedentary person, I mean, whether it's here in England or the us healthy people, nothing strange and not complete. Recreational athletes already consume between 1.1 and 1.3 grams of protein per kilograms per day. And they're not even trying to consume more protein.

We're typically pushing our patients much higher than that. We're asking patients to be closer to 1.6 grams per kilo per day. And truthfully, if we're really trying to put muscle on people and their training volume is high, we're closer to 2 grams per kilo.

It's great if you can reach that. I mean, here in the hospital, I mean, we're doing a lot, because I do a lot of clinical work. So we evaluated protein intake in the elective hip and knee surgery patients. So the older population, but these are not sick, they're coming in for new knee or new hip. So this is a new generation of patients that we get, of course, because in the past that was associated with a lot of pain. But these are patients that want to go skiing with the grandchildren again. So they want the new hip or new knee. So they come in, they're very healthy. But in the four days that they're in the hospital to go back to your advice, Aspen guidelines, For example, between 1.2 and 1.5 grams of protein per kilogram body mass per day. That's the clinical advice for recovering patients. You know what they actually so in our hospital, so what they actually get, what they're being delivered is 0.8 grams.

How much lean mass do they lose in four days of inactivity?

We'll come back to that. So they get 0.8. And you would say like, oh, give them more. They consume 0.5. So the healthy patients in the hospital here, the healthy patients go for elective surgery in those four days, they consume 0.5 grams, which is one third of the clinical guidelines for the advised amount, and they lose about 1.4 kilograms.

Wow. So they'll lose more than three pounds of lean mass in four days. And I've talked about this extensively on the podcast about the asymmetry in how quickly you lose versus how long it takes to get back. And of course, we talk about this a lot in the context of falls that result in fractures. Fractures. Because here it's an even more vulnerable population for many reasons. First, unlike the elective hip replacement and the elective knee replacement, whose patients are generally very healthy and able to get back to PT quite quickly, especially in the hips, the people who are falling and fracturing femur and neck, neca femur and hip, they're typically not as healthy and they're much longer to get back to recovery. And they're potentially bedridden for much longer periods of time. And oftentimes they never make it back to the level of muscle mass, strength and function prior to. In fact, I'm not sure if you're aware of this statistic. You probably are, but Adam Cohen on this podcast mentioned that, you know, we talk a lot about the 15 to 30% of people over 65 who will die within a year of a hip fracture. What we don't talk a lot about, of the people who don't die, call it the 70 to 85% who don't die within a year, 50% of them never regain their same function pre fall. In many ways, I find that statistic even more profound.

I was giving lectures for a group of older people, general public here, and I basically mentioned those numbers, not thinking about basically what I was saying. So half the audience turned white when I said that, because of course, there were a lot of people that actually broke their hip. But we took biopsies from patients coming in with a hip fracture after falling, and we compared that muscle with aged matched people that didn't have a hip fracture, didn't have a fall. And we compared it with young women. Most women are the ones with hip fractures because the men have already passed away. But you actually saw that the size of the type 2 fibers was tremendously smaller in the women with the hip fracture than the ones that didn't have the hip fracture. Even though we matched for almost everything, lifestyle and everything.

What's very interesting there is the causality could be in reverse. I think you could also make a very compelling case that having smaller type 2 fibers, which means having less power, having less force generating capacity, would make you more susceptible to a fall. In fact, I wouldn't be surprised if there's a bi directional association. So lower type 2 muscle fiber size and density, much more predisposition to a fall. Once you are inactive, you now experience even greater atrophy, which as you pointed out earlier, atrophy of the type to a muscle fiber is, I would argue it's should be described as one of the hallmarks of aging. Right up there with decreased mitochondrial functioning, increased senescent cells, all of the things we typically think of with aging, we should really be adding atrophy of type 2 fibers.

And that's why maintaining muscle there is more than maintaining muscle. It's also especially maintaining the type 2 fiber size. Because what we now often see with patients getting back, and especially now with COVID we are sending people home much more fast because we don't want them to get a bacteria or anything like. And we don't want them to attract Covid, so we send them home earlier. Now what happens when a lot of older people go home? Even if it's not a hip fracture, but an elective knee or hip surgery, Everybody means well. So children, grandchildren. Well, actually the first thing that they will do while you're in the hospital, they'll put your bed downstairs into the living room. So those people will never walk stairs again. First of all, they're afraid of walking stairs. They won't walk stairs again anymore because everything is put down. So they never recruit their type 2 fibers anymore. They might go for walks, but you don't recruit those type 2 fibers. So you need to stand up from the toilet several times in a row. You need to do or if possible, resistance training. You need to have somebody do resistance training with you or walking stairs. You need to recruit those type 2 fibers because otherwise it just goes down. I mean, we now believe that age related muscle loss, I mean it's a demographic, but it's not a physiological process of a slow decline in muscle. We now believe it's short, successive periods of reduced physical activity that are actually experienced after which they don't fully regain their muscle. And that adds up in the last two decades of life. And that seems to be contributing to the muscle loss that we see in those demographics.

Let's talk about that again because I think that is such an important point that I want to make sure not a single person listening to us Missed that point. Let me restate it. And I want you to clarify. We have long looked at data. These are all sorts of data. These are anthropometric data. These are functional data where we put activity monitors on people. And there is an unambiguous and clear decline in spontaneous activity, deliberate activity, and lean mass in the aging individual. And the decline starts somewhat slowly and by about the seventh decade, starts to accelerate by the eighth decade when a person is in their 70s, the decline is so rapid that it appears almost irreversible. We would typically talk about this as an inevitability of aging. Hey, that's just the physiology of what happens to the muscle. But a minute ago you said something entirely different, which suggests that it is not inevitable and that it is not a continuous slope of decline that reflects some physiologic process within the atrophying muscle. But instead it's a series of discrete declines, each one precipitated by a period of inactivity, some of them perhaps deliberate, meaning, hey, I'm going on vacation for a week and I'm just going to sit on the beach and do nothing. Some of them forced upon us by injury or illness. First of all, did I get it right? Is that what you basically said?

Yes, and I think everybody has that observation. Think of parents or grandparents that you saw in the last, say, 10 years before they died. Everybody will say it started with that urethra infection, it started with that surgery on the hip. And then you actually have all these little episodes. And then you saw that happening. And that's exactly what is happening. But this, of course, muscle centric. So I'm talking about muscle loss. Of course, if we're talking about cardiovascular disease and progression, that's a different story. But for the muscle loss is not something we don't believe anymore that it's in an individual. There's not a gradual loss over time, because in the individual level it can be completely different. The demographics show this because as we age, there are more people in that situation where they have short, successive periods of reduced physical activity. And I think one of the first period people to actually publish this was Douglas Penn Jones, who unfortunately passed away himself at a way too young age. But he called that the catabolic crisis model.

What I find interesting about that is you mentioned that maybe this doesn't apply to a decline in cardiorespiratory fitness or cardiovascular function, all these other things. Maybe. But I would argue that a muscle centric view could potentially be the most important view, because when your Movement stops, everything else deteriorates with it. People say, well, what would that have to do with heart health or brain health? I would argue it has everything to do with heart health or brain health. When a person becomes sedentary, everything deteriorates in its wake. Of course your cardiovascular system will deteriorate at an accelerated rate. Of course, we know unequivocally your brain will erode at an accelerated rate when you become inactive. And of course we know that the quality of your life, your happiness, your well being will deteriorate as you become inactive. And so you could make a very compelling case. I believe that a muscle centric view might be the most important view.

I agree. And that sometimes it's like talking to a mirror when you respond. We've done bed rest studies in healthy people because we often use healthy young people as a model to see what inactivity does. And besides that 1.4kg of muscle that we see disappear in a week, or if we immobilize a leg, almost 220 grams of muscle that is lost in a week, we also see a massive decline in oxidative capacity. We see a decline in insulin sensitivity. So all of these markers that we have for cardiovascular metabolic health go down in a single week of inactivity. So it's completely right. But of course, in one person it might be the cardiovascular decline that is driving the muscle loss, while in the other person, it's the muscle loss that's driving the cardiovascular decline.

Yeah, I'll make another shameless plug for an idea that we talk about a lot on this podcast, which is this idea of the centenarian decathlon, this idea that the best way to avoid this fate when you're in your middle age, when you're young, whenever, is to be very deliberate and specific about the type of training and of course, nutrition that is necessary to have the most physically robust final decade of life, what we call the marginal decade. And so if you train with great specificity to be very active and very independent and free of pain and all of those things in the last decade of your life, by definition, you're going to be doing a lot of varied forms of exercise, and you're going to have to be supporting that nutritionally to get there. I want to go back to kind of one last housekeeping issue on the protein side, which was the difference between whey and casein protein in identical amounts relative to the parameters you've spoken about digestibility, speed of digestion and amino acid composition. What are the differences and where do you see the use cases clinically between them.

So the digestibility is the same. It's 100%. So there's no differences there. But if you ingest casein versus. And that's micellar casein versus whey protein, the whey protein is much more rapidly digested and absorbed. You see a greater spike in your essential amino acid concentrations in your blood, including leucine, and you get a greater stimulation of muscle protein synthesis as a result. However, the longer you measure, the more the casein catches up on the way. That's also something that you have to realize.

Yeah, tell us about that study that just came out a few weeks ago, which seemed to challenge a lot of the conventional thinking around the amount of usable substrate that one could ingest. So traditional thinking had been, boy, it's awfully hard to use more than 40 to 50 grams of protein in a meal for muscle protein synthesis. And if I recall, you had three groups in this study. A group that was given 0 protein, 25 grams of protein, and 100 grams of protein. That was all casein. Correct.

That was milk protein concentrates.

It was a milk concentrate. So that means equal parts casein and whey, or what was the breakdown?

80, 20.

80, 20.

In favor of which casein is always 80 and whey is 20 in milk. Okay, so if you have milk, about 80% is casein, 20% is whey.

Got it. And then tell us what you saw in that study over a long enough period of time.

So this was a study where we use that intrinsically labeled protein combined with intravenous infusions of amino acids, and that allows us not only to measure muscle protein synthesis, but also how much protein is actually released in the circulation from what you ingest. And so we could make a full quantitative assessment of what is happening. Now, previously, like I said, most of our studies, people like to work on office hours. So you have two hours of a run in period, maybe an hour of an intervention, and then four to six hours of postprandial. So after a meal measuring of muscle protein synthesis, those are long days. So most of the studies so far show that 20 grams of protein does not do better than 40 grams of protein to stimulate protein synthesis for up to four to six hours, because that's typically what we do. Why do we always measure four to six hours? That's all the labs doing this kind of work is because that's the time between two meals. So that makes sense. So the 40 grams always, if you see the studies and a lot of other labs, it looks like a greater response than the 20 grams, but it's not significantly different. And what we saw with the intrinsic label protein, that a lot of the protein is not digested yet, or at least not released in the circulation. And the longer you measure, the more of that protein still coming in. Same with what I said. The casein is catching up on the way because a lot of the casing is not absorbed yet. So what Johan wanted to do, and this was crazy because he wanted to ingest 100 grams of protein, to be honest, he's so crazy, he actually wanted to have more treatments. And I said it's not going to happen because that's way too expensive. And this was already very expensive. So we did a zero, 25 is what is being advised, give or take, and then the 100 gram as an absolutely unpractical amount. And so we actually measured, if I'm correct, at 4, 8 and 12 hours after the ingestion. And so what you saw is that if you measure for an extended period of time, that 100 gram is still increasing muscle protein synthesis for a longer period of time, resulting in a greater response over time. So the suggestion that you can't get a greater response with more protein is biased. It's biased because you don't measure long enough. Now, the problem is this is a study that everybody, at least in my field, needed to see so to better understand what is happening. But it doesn't necessarily translate. And of course, that's the problem now with papers. People start calling me, like, interview this, interview that, like, should I eat one meal a day? No, that's not what we're saying. I still think that it's good to have a distribution of protein throughout the day, that every meal has an anabolic response. But it doesn't mean if you have small meals that you're going to disappear in three weeks. So the body adapts, and if the body has a huge meal, it's still being used. Makes sense, because if you have it, we also wrote it in a discussion. If you have a snake or a crocodile that once in so many days devours a goat or chicken or whatever, then it can actually see that animal actually stuck in the gut for weeks, and he's actually consuming it. Now, to some extent, humans can also do that. So it just shows you that the response, there's no limitation in the response, and it can actually range longer than what is previously expected.

Do you think if you did that experiment, but instead of using a milk concentrate, which was majority casein, you used a Pure whey isolate. Do you believe that you would have still seen muscle protein synthesis at the 12 hour mark or do you think that that the rapid digestibility of whey would have changed the kinetics significantly and you would have been largely done at the four to six hour mark?

I think the difference between the 25 and the hundred grams might have been smaller, but it would still be evident because then the duration of the extension is smaller, of course with the whey than with micellar casein. But the comparison with milk is very small because I mean milk still is very rapid. I mean it's always funny that people say like this is slow or this is fast.

It's all relative.

Yeah, it's all very fast. It's all concentrated proteins. If you compare it with full meals, you get a completely different picture. And that's what Philippe Pinkertz in our group did. He compared the real meal, a vegan meal versus a typical Dutch, and not even know whether it's still allowed to say that typical meal was actually in Holland, vegetables, potatoes and a piece of meat. Now we made those two plates having exactly the same energy and exactly the same amount of protein. But now we also have that digestibility factor in there because we weren't using powders in a drink. We actually have a full meal, a plate with all vegan whole foods and a plate with a piece of meat and potatoes and some veggies.

And was the vegan plate cooked protein or was it raw?

Yes, was cooked protein, but it has a little bit more fiber. The plate is also a little bit bigger because the energy density is of course smaller. And then everything comes together also with the digestibility. And then you actually saw no measurable anabolic response with the vegan meal and you saw a massive anabolic response, muscle, protein, synthetic response with the omnivorous meal. And that shows you that, I mean, and that doesn't mean before everybody just stack us again, that I say that you shouldn't be eating vegan. But the point is that an omnivores meal with animal derived proteins has a greater response when you compare it in an amount, the same amount matter, the same amount of protein. So quality counts. Yeah.

Let's go back to what you talked about with respect to. We completely talk about the relative speed between whey and casein, but let's put those in the context of a meal. So what does the kinetics curve for acacian shake look like compared to ground beef or whole steak? Is it significantly faster?

No, that's similar. Cooked beef is still very fast. So if you have cooked meat or concentrated protein isolate from an animal source, it's all very rapidly digested and absorbed with peak amino acid levels I think between 30 and say 75 minutes. If you have a full meal with potatoes, veggies, fibers and in it, it will take two to three hours longer.

From a very selfish standpoint, I want to ask you a question. So I always do my strength training in the morning. Many days I just don't either not that hungry or I kind of run out of time before I need to jump into stuff. And I'm not always diligent about eating or consuming protein right after that workout. Now truthfully, in part it's because. Well, I want to ask you this question. Let's just assume two scenarios. Scenario one is finish exercising, finish strength training, consume nothing but water, coffee for another three or four hours before consuming a high protein meal versus finish exercising and immediately consume 25 grams of whey protein in a shake with nothing else in it, just whey protein in water, you're getting 100 calories, chug it down and then eat that protein rich meal of 40 grams four hours later. Significant difference in muscle protein synthesis between these two over the course of days. Or does it all come out in the wash? Where assuming you get ISO amounts of protein throughout the day, does it matter on 24 hours?

Definitely there's a difference. For example, if you ingest protein immediately after exercise, you get at least for the first five hours, you definitely have a much greater response because exercise makes the muscle more sensitive to the anabolic response to food intake. However, what people forget is that your response to breakfast the next day is still increased and probably also your lunch and dinner the next day. So if you do an exercise session today and you get all freaked out that you didn't get a milkshake after your session, don't worry because the next day all your three meals are going to be greater responses because of the exercise. So I get so many times the question like how important is the meal before or after the exercise? And I say if you do a consistent training, then there's always three meals before your session and after your session because you train every day. So then every meal is still responding to the previous exercise session. Now is there a benefit of immediately after exercise versus a few hours later? We've done that study, so not sure whether we touched the topic as well. Pre sleep protein feeding, that's a topic that we've been working a lot on also on patients. So we gave people, I think it was even 60 gram. Really proof principle study exercise in the Evening and then 40 or 60, I don't remember, we gave protein then and then we measured the response the next morning after breakfast, after 20 grams of protein in the morning. So what I was thinking, like if there is a window of opportunity, then if you give 60 grams of protein immediately after exercise, maybe you shorten the window of opportunity. So if you already provided the exercise after the session, you don't respond as well to the next dinner, the next morning.

What I'm basically hearing you say is if you have a hundred units of response in you, the timing of your meal might not impact the total amount of response. It just determines when that response occurs. If I slug that 25 grams of whey protein as I'm walking out of the gym, I will get more of my hundred units of response then, but I will get less of it for five hours later when I have a big protein lunch.

I think that's a much clearer explanation then. So that was basically the design of the study and it fits with what I've learned, of course, for glycogen, because you ingest carbohydrates immediately after exercise, so you expedite accelerate glycogen resynthesis. But if you don't have to exercise until two days later, it doesn't make a difference because then you have a full 100% recovery of your glycogen.

Which again is very different from the way I grew up. I mean, when I grew up as an endurance athlete could all be wives tales. But the traditional thinking was you need to be mainlining carbohydrates the second you get off the bike or out of the water or whatever, because you have this very small glycogen window where for an hour you're going to maximally assimilate. And it might be the case that, well, it's true your maximum assimilation would occur in that window, but you will still incorporate carbohydrate into glycogen later. It just might not occur at as high a rate. Is that kind of the same situation.

If you're exercising every Saturday? It doesn't make a difference. Louise Burke has Clearly shown within 24 hours your glycogen are back to normal. If you are in the Tour de France and you have to excel every day, you're going to miss the third day of the Tour if you don't start taking in carbohydrates after the session. Now going back to so that study protein ingestion after an exercise session before them in the evening, does that impact your response to morning? And so the other trial was no protein after the exercise and doing the same thing in the morning. I thought that the response to breakfast will be reduced if you already ingested that 60 grams of protein in the evening. It didn't. The responses were exactly the same to my surprise. So the responses to breakfast exactly the same. So net, I must say that the people ingested the 60 grams prior to sleep had a benefit in that time frame. Whether it's caught up later on 24, 48 to 72 hours, I don't know.

In other words. Yeah, I was going to ask you that. You only did this for one dinner, breakfast and sleep session.

Yeah, so. So these studies. So the studies with infusion of traces are almost always limited to about 12 to 24 hours. Why? Because you have these turnover of the tissues. So at some point your tracer will become available from the breakdown and then you're measuring tracer recycling. It's different techniques and we can go into that with D2O. There's different techniques to counter that. But if there's acute tracer infusion studies, you're limited to 12 to up to 24 hours. But so far, long term training studies basically have shown that protein supplementation can further increase gains in muscle mass and muscle strength. That evidence is there. The evidence becomes smaller when people eat more protein and it gets stronger. If you look at people that do not consume enough protein, I mean we had a black and white response to frail elderly, six months of training. If we didn't provide them additional protein, they didn't gain more muscle. So the more frail people are, the more important the amount of protein gets. But it's also because they're not consuming a lot.

I want to actually come back to the elderly in some detail, but I do want to put a bow on this topic here, which is we don't really know over the long term, meaning over months and years if there is a benefit to consuming a very highly digestible, rapidly absorbed, good quality amino acid composition protein in the immediate aftermath following exercise, it sounds like it's still unclear if there could be a net benefit. So if a person listening to this says, look, I want everything in my favor to maximize my odds of optimization and maximization of muscle mass, I'm going to consume 25 grams of whey protein following every workout. Again, from a caloric perspective, it's irrelevant. It's a hundred calories, right? It's like less than 5% of your daily intake. But it would arguably Be the most most efficient way to deliver amino acids. Is there any reason not to do that? Let's just start with that. Is there any downside of doing that as opposed to going through the hassle of consuming a meal when you finish your training?

No. On the individual? Absolutely not. The only problem that I do have is that if you advocate too much to use like protein supplements and stuff like that, people stop thinking about their foods. So we've had people coming in here saying like, oh, I put a lot of interest in my nutrition and my diet. And then I asked them, so how do you do it? Yeah, I take 29 supplements. Just first think about your nutrition and if every meal contains good solid foods and with ample, ample protein, if you then on top of that, decide that it's easier, more practical to take away protein supplements after training session, be my guest. But if all the rest is crap, then please do not even consider those whey protein supplements, because first think about your nutrition. So I had a lot of people in my life asking me, like, how important is it whether I take my protein shake before or after the training session? But I had never somebody that came up to me like, look, how important is it if I skip one training session or I miss one training session? Consistent training is the benefit. Consistent training so that every meal is a greater impact on your muscle protein synthesis. It's the same. The questions. I always get, like interviews on the radio and they say, look, what should we eat in order to lose weight? And I have only one response. It's less.

Yeah, exactly.

But that's people, it's easier to drink a whey protein supplement than actually just leave the house at 6 o' clock in the morning and do an extra session of training.

Yeah, let's talk a little bit about eating less. Because again, people who listen to this podcast are very familiar with the way I think about this, but I'll just sort of explain it to you very, very quickly. I sort of break it down into three strategies to go about eating less. So strategy one is a very deliberate strategy called caloric restriction. So it's the only way that you directly go about eating less, which is clearly what bodybuilders do, where they're tracking every macronutrient, every calorie, they are counting them, and they are shrinking that volume of calories lower and lower and lower to reach an energy imbalance that is sufficient for the amount of fat loss that they're trying to produce. Again, you could argue that this is the most flexible way to go about weight Loss because it is agnostic to when you eat or what the actual constitutive elements are of the diet. It's really just paying attention to the energy balance or imbalance. The second way that you can go about doing this is indirectly through dietary restriction. You talked about a vegan diet. Hey, you're taking a lot of things out of the diet and many of them are energy dense. So there's a very good chance you're going to lose weight just on the basis of the restriction. Similarly, with a ketogenic diet or something of that nature, you're going to really generally eat a lot less. And it's that effect of the diet on perhaps your appetite and food choices that's going to result in lower energy intake and therefore energy imbalance. The third technique, also indirect, is time restriction, which people call intermittent fasting. Here you're going to make a larger and larger window of non eating and by extension a smaller and smaller window of eating that eventually results in a caloric deficit. Now we have historically meaning in our practice cautioned people about excessive use of time restriction. If people fit a certain demographic, and that is you are obviously overnourished, which is our way of describing you have excess adiposity, you have high visceral fat, all of these things, you're metabolically unhealthy, so you're overnourished. We need to make you less nourished. But if you're also under muscled, I get very worried about excessive time restriction because with calorie restriction comes protein restriction and with protein restriction comes not just a reduction in mass, which on some level is the goal, but a disproportionate loss of lean mass. So I want to pause there. Does this resonate with you?

All of these trade offs fully agree on that. The third one is of course the time restricted feeding or intermittent fasting. It works for people, but it doesn't work in a scientific setting because now there's a lot of studies coming out. We've also done a study on that. So if you standardize the nutrition so the caloric content of the diet, you actually see exactly the same fat loss or weight loss with the intermittent or the time restricted feeding versus the same feeding, but then X percent of less caloric intake doesn't make any difference.

But let's just make sure people understand what you just said. And I want to make sure I understand what you just said. You're saying that if you normalize for calories across the course of the day versus in a shrunk window. If it's the same number of people calories, there's no weight loss.

Exactly. There's less weight loss. So if you have 25% energy restriction independent of the time frame, you're going to lose the same amount of fat mass. Now for me, intermittent feeding or time restricted feeding works because I go to the university, spend most of my day here, have no time to eat, running back and forth and then I come home, I sit behind a computer doing all my emails and revisions of manuscripts and I start eating way too much and also crap food. So I think more than 70% of my food intake, energy intake is between 7 or 8 and 12 o' clock at night. If I do time restricted feeding from say 10 to 6, I will lose weight because I wouldn't have time to eat that amount of calories in that time frame. And that's good for a lot of people because they actually change their homeostasis and for a few weeks they lose a lot of of fat mass and then they start eating differently and then they gain weight again. So sometimes it's easy to change your routine and your bad nutritional habits, but the time of feeding is not a metabolic effect to lose more weight.

Yeah, so what we like to do to try to get around the effect of people losing too much lean mass. And again, I find this to be particularly important in women who want to lose weight using a time restricted feeding approach, but who frankly already have an ALMI or an FFMI that's in the bottom 20%. Just for folks listening, these are two different ways that we measure and quantify lean mass on people. So this is people that are coming in with, they're under muscled as we describe them. We will say, look, if you want to eat between 2pm and 7pm, that's your feeding window. You have a five hour window to eat. So 19 hours of not eating, five hours of eating, we find that women can't. And again, I say this because it is mostly women that experience this, but I think it could be true for anybody. It's very difficult to consume your total amount of protein if we're trying to get you to 1.6 or 1.8 grams per kilogram in a five hour period. Furthermore, you have that 19 hour window where you're missing one of the major inputs to muscle protein synthesis. So a workaround is, hey, during that period of time you're going to have two shakes that are virtually no calories, but are going to give you 50 grams of protein, a 25 gram whey protein shake Again, just mixed with water at 8 o' clock in the morning and again at 11 o' clock in the morning. And then at 2 o' clock you're going to eat a meal and at 7 o' clock you're going to eat a meal. So you're going to get your total amount of protein. And yeah, you've sort of cheated on your time restricted feeding because you've had 200 calories outside of it. But let's be honest, the purpose of this is caloric restriction. That 200 is relatively small compared to what you would have consumed throughout the day. So again, long question, I apologize. It's as much for the listener as it is for you. Would you take that approach as a better way to tackle 2 simultaneous goals? Lose fat mass, preserve or gain lean mass simultaneously.

That would be the main goal and I would completely agree. But I miss one factor. I'm 100% sure that you actually added the effect.

Of course, the training effect.

Yes, the resistance training. So even if you have, I mean old studies show that if you have a caloric restriction, you lose fat free mass, you lose muscle mass. But if you do twice a week a resistance training session, even in a caloric deficit, you don't lose muscle mass. So you can prevent the muscle mass with simply two sessions of resistance type exercise a week, preferably more. But of course that's the minimal. So besides that protein, it's the exercise that makes you respond way more to the same or less amount of protein that you ingest.

And in the scenario I described where you're going to have that person have a protein shake at, at 8 o' clock in the morning and 11 o' clock in the morning and then eat two meals between say 2 and 7, it's probably not that important where the training session goes within that day at this point. I mean, I think for many people it's just convenient to do the exercise in the morning. And I agree of course that that's half the battle is you have to have the training stimulus to produce the effect.

I mean that's what people always ask us, like should we ingest the protein immediately after or an hour after exercise? Does it make a difference? No, not really. Because every meal following your training session will have a greater response and so also off before the next session there will again three meals or four meals. So it's a continuous consistent effect of training to make you respond better to the same amount of protein. And also in the hospital this is essential, simply feeding people more protein because the deficient and losing muscle is not the only solution. Actually, if you make them a little bit more active between meals, every meal has more effect. If we do a little bit of exercise, more of the meal will actually be converted to muscle. So using that intrinsic label protein, I always have three different settings in my lectures. The first one is this. Intrinsically, protein shows you, beyond any discussion, you are what you eat. In fact, you are what you just ate. Now, funny enough, if you eat that same protein after you've done a little exercise, more of that protein is converted to muscle. So if you're physically active, you are more of what you just ate. Now, every athlete is using this, and every coach knows this, but we hardly use it in medical care. And so it gets worse, because then you come to the third part of the lecture where I show you that with physical inactivity, you become anabolically resistance. So if you become physically less active, you are less of what you just ate. Now, the problem is, when you become sick or ill or you get surgery, you have two issues. You exercise less or you become less physically active, but you also eat less. So it's a double whammy downwards. And that's what makes us so susceptible and vulnerable to a short period of inactivity or sickness.

The term anabolic resistance is intuitive because obviously people understand what insulin resistance is. But with insulin resistance, we can actually really explain mechanistically what's happening. I referred to Jerry Shulman. He provided a very, very elegant explanation of what's happening intracellularly, what the triglyceride, or diacylglyceride, if I'm not mistaken, is doing in the cell and how it's impeding the signal transduction to move the GLUT4 transporter up to the translocated site to bring more glucose in, et cetera.

I haven't followed that discussion, but I mean, it's still a discussion. Which fatty acid intermediate is actually causing the whole signaling? Is it the storage of the fat? Is the inflammation that you get from the storage of fat? Is it diacylglycerol? Is it. Are the ceramides the fatty acyl coa? I mean, yes, but it's at least the inability to process that fat in the muscle is inducing insulin resistance.

So the question now is, do we have the same level of detail around what is actually inducing anabolic resistance? Which is a topic that I don't think people are as familiar with, but unfortunately need to be as familiar with. Given its prevalence under two conditions, inactivity.

And aging, I think that's the million dollar question. I think the first time that anabolic resistance was published as the name was coined, was by the people in Dundee, Mike Rennie, the late Mike Rennie. So he did a study where he provided essential amino acids to people, young and elderly. And when they provided a greater amount of essential amino acids, you saw in the young people a greater muscle protein synthetic response. As we discussed before, he got the greatest response, the highest muscle protein synthesis following the provision of 10 grams of essential amino acids, which is perfect because 10 grams of essential amino acid translates to about 20 grams of protein. So it nicely fits with those other studies. If he gave 20 grams of essential amino acids, there was no significant further increase over the next few hours. So that was normal response. And in line with everything that we discussed before, if he did the same thing with the older population, he saw that that increase in muscle protein synthesis was less steep and also leveled off more rapidly. So that's how he coined anabolic resistance. And that was I think 2005. Now it took our lab almost 10 years to verify or to confirm those data because we wanted to show the same thing with a less lab based approach, not essential, with that 20 grams of intrinsically labeled protein. And when we actually finally managed to do that study over a long period of time, we saw that basal protein synthesis is not different between young and elderly. So if they're relatively active, basal protein synthesis, if there's a difference, it's actually higher in elderly than in young. But the response to the 20 grams of protein is less in the older population. So the same amount of protein that was ingested did not lead to the same amount of protein synthesis. And with the intrinsically of a protein, we could show that less of the ingested protein was converted to muscle. So that is also anabolic resistance in a more single meal like type of approach. Now the big question is, and this is now a lot of people are focusing on anabolic resistance. What is causing anabolic resistance? That could be digestion, that could be absorption, that could be. And that's what they call splegnic sequestration, the uptake of amino acids. And what happens between taking up in the guts and releasing in the circulation. And of course, if scientists use difficult names, always beware, it generally means that they don't know what they're talking about. So when we go say splegnic sequestration, everybody says like okay, but it just simply means we don't know to what extent the amino acids that were actually going to the portal vein or actually being absorbed in the intestinal tissues and not being released in the circulation. That's splengnic sequestration. Where is it? Is it in the liver? Is it still in the intestine? Is it taken to other tissues in between the portal vein like lymphatic system? We don't know exactly. And then the release in the circulation. But then insulin also plays a role, because if there's not enough insulin, and insulin is not stimulatory, but permissive is how much of the tissues are being perfected so they get blood. Because if the blood doesn't perfuse the muscle, those free amino acids are never going to be seen by the muscle. Then we have the uptake in the muscle and then you have those signaling responses, MTOR pathway in the muscle. On all these levels, anabolic resistance can reside, so it's impossible to find it. And so a lot of people are focusing on all these different areas. And then the problem is, and this is the problem with every study in where we actually look at aging. We compare young and older people. We don't follow the same person for 40 or 50 years because that doesn't work for studies and it takes too long. That's why we use all those animal species or any organisms like the C. Elegans or whatever. If we compare young and old, we're not comparing the same person. We're actually comparing lifestyle comorbidity, pharmacological interventions that they have, food intake, all of these things. But I just said that physical activity makes the muscle more sensitive to the anabolic properties of amino acids. Does it make perfect sense to say that less physical activity makes the muscle less sensitive to amino acids? So the only way to study this, and this is what we do, is we immobilize young people. So we put one leg in a cast for a week. And then if you didn't believe the story about 1 to 2% turnover of muscle, you know, because even after a week, when the cast comes off, you don't need an MRI scan to see which leg had been in the cast, because you actually see the muscle as the loss of muscle in that leg.

How much do you have to pay a young person to subject themselves to a week in a cast?

Oh, to be honest, that is actually. So one week is, I mean, six weeks will be difficult, Especially here with health sciences and medicine. A lot of the students, I mean, are active, but one week they actually love the experiment. I mean, now they can choose the color of the cost as well.

So I think of some of the crazy Experiments I got paid to do while I was in medical school. At the time, what I would do for a thousand dollars was. I won't admit it publicly. Have you been able to study this longitudinally in a mouse, for example?

No. No. So we only do animal stuff, human studies in vivo. I hardly do any. I mean, I do some animal work with collaborators.

Is anybody studying this? Because again, you raise a good point, which is without a longitudinal assessment, which would not be possible in humans, we really are stuck without understanding the nuances. Of course, in humans you can do a crossover between active and inactive, and that might provide an answer to the question. But as I listen to this list of differences in absorption, circulation, the Spliknik sequestration, which is a. I've never considered that there. Perfusion related. Makes total sense. Uptake in the muscle, lower MTOR activity. We know MTOR activity is lower in the elderly. I mean, it could be any of the above. It could be all of the above.

And what do I think that they're really important? I don't think so.

Yeah, at the end of the day, we know what you need to do. Control what you can control. Be more active, consume more protein.

But. So if we take that previously immobilized leg and we give that person an amount of protein, I see a 35% difference between the lag that was previously immobilized and the leg that was not immobilized. So there's a 35% anabolic resistance after one week of inactivity. That is much more than we see as a difference between the young and the older. So with one week of inactivity, I can make a young leg or young muscle respond completely, like a senescent muscle. And now the fun thing, if I actually take an older person and I do some exercise and a given protein, I see completely normal response. If I take a biopsy of an older person, I see smaller type 2 fibers. If I train that person for three months, the type 2 fiber is bigger than the type 1 fiber. And I don't see any difference in the response to a younger person. And I can say the same thing about satellite cells. So muscle on itself is actually doesn't seem to get that old. It actually still responds completely normal. And I can normalize for age by physical activity.

Yeah, I was about to say the two examples you gave there completely changed the discussion again, I think it's worth restating them because you're saying so many important things. I just want to make sure people are not missing these. Right. So you Took a group of young, healthy people who presumably have lots of anabolic capacity. You put them in a cast for a week, and you immediately demonstrate upon removing the cast that the leg that has been immobile for a week is 35% less responsive to protein assimilation than the other leg. That's an anabolic resistance factor of 35%, which you also pointed out is far greater than what you see in an aged individual.

It depends whether that age, an active older or normal, or if that older person is lying in bed for three weeks with COVID We're talking about other situation, but yeah, but then the other.

Thing you said was you can take an aged individual who might have again, 20 or 30% less anabolic response to protein than a young person, but if you exercise them, you can bring them up to the same level as a younger person. These two factors suggest that activity might be the main determinant of anabolic resistance. And an aging individual or age within an individual is simply a proxy for activity.

And so it's more difficult for older people to maybe do the same training loads. But the muscle itself is still responsive. So the good news is you don't have to start exercising when you're 40 to actually have good muscle when you're 80. You can still do that at 70. Now it's better to do it at 40 as well. But at any age, and that's also 100 plus, the muscle is still very responsive to physical activity and it has nothing to do with hormones. And that is also something that I get so fed up with. We've just finished a study with prostate cancer patients. Now, you know what happens when they get antigen deficiency therapy? They lose a lot of muscle mass, they gain a lot of fat mass. They have an increased risk of becoming diabetes, developing cardiovascular disease. Besides the effects that they become a little bit lethargic. We did training with them resistance training two to three times a week. The ADT had no effect on the muscle mass. They actually gained muscle mass. So despite the fact that they had no testosterone, they were basically chemically castrated two to three times per week. Resistance training increased muscle mass, increased muscle strength, and attenuated fat mass gain. It's that easy. It's ridiculous that people go on ADT and are not immediately getting resistance training in addition to it because it prevents all the negative side effects.

You've now hit on the second point that I think is really frustrating and difficult to hear. And I really hope that there are people out there in a position to do something about this. Right. Which is look, there's such a disconnect in medicine when it comes from doing really wonderful things to then missing the boat on the support system that's needed. So the two examples here is the one you just gave. You know, does it make sense to put a man on androgen deprivation therapy? Yes, it does. Under certain circumstances. If a man has metastatic or inoperable high grade prostate cancer, he needs to be on androgen deprivation therapy. And as you pointed out, yeah, as you pointed out, he is being chemically castrated. And I have seen many of these men and they are quite miserable because they are losing lots of muscle mass, they are gaining lots of fat, and they are becoming metabolically profoundly unhealthy. And yet you would think that for every oncologist that puts a patient or every urologist who manages a patient through androgen deprivation therapy, it should be as important that prescribed alongside that androgen deprivation therapy is a resistance training program and a proper diet around high protein intake. And I would say, look, you should be resistance training four or five times a week. I mean, this is the most potent thing you have to counterbalance that. And the other example you gave that's just infuriating is to think of all of these sedentary, hospitalized patients who are basically being protein restricted. Again, we talk so much in the United States, and I'm sure it's true in Europe, about the cost of healthcare and the burden of healthcare and the burden of the sick individual and how so many dollars of a person's healthcare allotment are spent at the end of life when the quality is so low. And yet the system itself is broken in that it doesn't even understand the basic fundamentals of exercise and nutrition. And the funny thing about this that isn't really funny is this is what I think gives much of the population a total lack of trust in the medical establishment because they see, hey, you guys don't know about exercise and nutrition, so why should I believe you on these other things where you actually do have an authority? And I feel like I've seen an acceleration of this over the past few years. Covid clearly didn't help. We don't have to talk about why I think COVID policy eroded a lot of trust, but I think there's something even deeper than that, which is just a general belief that the medical system doesn't communicate through a strong position of knowledge when it comes to the real way to use exercise and nutrition as medical interventions.

I think everybody knows that lifestyle is important, but they sometimes just do not Realize how relatively easy it is to do it. And so we should have scientists and clinicians talk more. And it's changing, but it's very going very slow. Give you a nice example. I mean, when you're in the hospital, you get your nice meal at around five or six o' clock, if you're lucky, and then the next morning at say 8 o' clock or 9 o' clock in the morning. So you spent almost 9 and 6 of 15 hours of fasting. 15 hours of fasting when you're at the risk of losing muscle and you're not eating in those 15 hours, that's two. But not a single athlete in the world will do that. So we were wondering, like if you provide protein prior to sleep, does your gut actually process it while you sleep? Now that sounds easier said than done. So we called some of our subjects and actually they call us especially the elderly. They're really great to work with. It's amazing. So they call us and say, like, look, I haven't been to your lab in the last two years. Don't you have a study running that I can participate in? I would say like, oh, must the Jansse come over. What we'll do is we'll take a muscle biopsy, then we'll take a nasogastric tube, put a tube down your nose into your guts. We'll make you sleep in the hospital overnight. At 2 o' clock in the morning, we will tiptoe into the lab, push 40 grams of preheated intrinsic labeled protein in your gut without waking you up. And in the morning I will wake you up with a muscle biopsy. And then they say, where can I sign up? These guys are really amazing. But we did this. And what happens if while they're sleeping and you provide the protein in the guts, it's rapidly digested and absorbed just as easily as in the morning. So there's no restriction there. And they actually synthesize muscle protein at night. Next morning we see the incorporation in the muscle. So when we published that study, it was really weird. I got phone calls from coaches all over the world asking me where they could buy those nasogastric juice. And then I had to explain. I said, this is a proof of principle study to show you that the gut functions while we sleep. But if you want to do something, just give people a protein rich snack between dinner and going to bed because that will expedite reconditioning, help you with your recovery from training. And more importantly, it will might help older people or people in the hospital to attenuate muscle loss while in the hospital. Now that's part one. Second part is you actually start doing that with a protein, small protein snack in the evening. Done that as well. Then the next step is, does it actually work in hospital? Because if you provide a protein rich snack, do people still eat the same the next morning? Now that's already difficult because then in the hospital this is actually not the invasive work that we do. I just wanted to have people bring around cheese cubes in the evening in the hospital and see how it affected 24 hour food intake in patients. It went up by 20%. So that is stuff that we don't even manage with supplements because people don't eat them. They get a supplement of the meal, it's just thrown away. So bring people a nice protein rich snack in the evening, they increase protein intake by 20%. It's that easy.

I want to talk a little bit more about protein supplements. We've talked extensively about the milk protein isolates, whey and casein. I get asked questions all the time about this and unlike you, I don't know the answer. So I'm kind of like, you know, my answer is I don't think that matters. But, but for example, my wife's friends always ask me about collagen. Hey, should I be consuming this collagen or that collagen or this collagen? And my response is, I don't think that matters. I frankly think you're just better off consuming a high quality protein that has a balance of all of the amino acids. But tell folks first of all what collagen is and then maybe answer my question about whether or not there's a unique benefit to consuming collagen as a supplement.

John was thinking like I should do less research. I don't get all these questions. So collagen is a protein that is pretty prevalent in your body because it's the main protein that is, it's a structural protein. So collagen is in your cartilage, your bone, your tendons, your ligaments and all of that. And it's also in muscle, very small amount, relatively. And it's important to transfer the force of your muscle towards your tendons. So even in the muscle, all of your contractile proteins need to be linked to collagen in order or connective tissue proteins in order to transfer the force. Now a lot of people are in the market, you see now ingest collagen supplements because it helps you with strength, force, skin, bone, ligaments and stuff like that. We're interested in because it's a very nice source of glycine and Proline. About 50% of your collagen is glycine and proline. So it's a poor protein. From a total perspective, it's not as balanced as an animal derived meat protein or milk protein, but it contains a lot of glycine and proline. So what you could say is that, hey, your ligaments, your cartilage, your bone also contains a lot of glycine and proline. And so it's a good source of these two amino acids. Makes perfect sense. So the story makes sense, but it's a little bit like if you eat something that you need, it's going to be better for you. But the question is, do we already get enough glycine and protein in our diet and is additional via collagen of additional value? Now that is something that we don't know. We have been starting to look at this and so we've been ingesting collagen and whey protein after exercise. And then we look at myofibrillar protein synthesis, but also muscle connective protein synthesis. And I hope that doesn't go too fast. Exercise stimulates both myofibrillar as well as muscle connective protein synthesis. So the adaptive response in muscle is both connective proteins as well as myofibrillar proteins. Now, if you ingest protein, it further increases the response to exercise and you see greater myofibrillar protein synthesis. However, the ingestion of dairy protein or protein does not seem to increase muscle connective protein synthesis rates. So so far, at least for up to six hours after exercise, your contractile muscle response to protein in addition to exercise, but your muscle connective does not. Now, we've tried that also with collagen and we do not see a greater increase in connective tissue protein synthesis rates in muscle. So either it is not happening in the first five hours and the exercise is already a stimulus enough and the response is later on, or there's enough glycine and proline in dairy protein.

How much are you seeing in the muscle connective tissue response to dairy? I was under the impression, based on what you said, that it was virtually none. Is it some but just pales in comparison to the exercise.

It pales in comparison to the exercise. Exactly.

Okay.

No significant decrease in muscle connective protein synthesis in addition to the exercise effect.

Have you done this activity or this experiment without exercise? Because obviously exercise is such a potent stimulus that as you said, it might be dwarfing what we see. Have you done that experiment?

Yes, we also done it without exercise, and it does not seem to be responsive to nutrition. But I have to make an exception in that study by y that huge amount of protein, that 100 grams, we suddenly see it.

Interesting. So in that study where you gave the massive dose of protein and those were not exercising patients over a longer period of time.

Yeah, but that was exercise as well.

I see. So it seems that based on those data, there is no benefit in both myofibrillar or muscle connective tissue protein synthesis using collagen versus using whey or casein. So if a person is taking a collagen protein because they believe that it will disproportionately help them increase the strength of their connective tissue, the data would say that that is not correct, at least in the presence of an exercising individual.

Yes.

Over five hours, I guess, to be fully complete.

Yes.

Okay.

So from a practice translation, I still hold an option open for ligaments, tendons, bone, cartilage. Because when I look at muscle connective protein, the fraction that we actually take out of the muscle contains only a few percent of collagen. So the muscle doesn't contain a lot of collagen. So the question is, is it not more important for tendons, ligaments, bone, and cartilage? I wouldn't say say that it's not working there. So again, we are also athletes, we're also patients, and we're also scientists. So as a scientist, I haven't seen evidence that it actually leads to greater connective tissue protein synthesis rates. But if I would actually break my hip or I would actually have a major issue with my knee, and I'm recovering and rehabilitating, I think I would take both a protein supplement that also has a little bit of collagen in order to be sure that I get enough glycine and proline.

The other point to consider is if there are people listening to this who are just using collagen as their supplemental protein source, they're undoubtedly compromising myofibrillar muscle protein synthesis because as you said, they're basically just getting a lot of proline and glycine, and they're probably really missing out on leucine, lysine, methionine, and the other amino acids that are far more potent.

It's a very low quality protein from the perspective amino acid balance. Yes, but it's a nice source of glycine and proline. But if you take enough protein, probably the glycine and proline is already sufficiently available in your diet. However, if you have major issues with ligaments, tendons or other almost purely collagen based structures, it might be a benefit. I'm not throwing that away. That is something that we still want to look at for the next few years, but the jury is still out on that.

Yeah. Let's talk a little bit about something you said earlier, which was what some targets might be for a meal. So you mentioned 2 to 3 grams of leucine in a meal. If you really, really want to kick muscle protein synthesis into overdrive, do you have any other rules of thumb around specific amino acids? One of the things we do try to talk to our patients about, especially patients who are plant based, is rather than just have them worry about the different bioavailability of this protein versus that protein, is just sort of focus on how much leucine, lysine, methionine, you're getting across the course of a day or even in the course of a meal. But I'd like to hear your guidelines around around that.

So many of the plant derived proteins are low in lysine and or methionine. So that is always a discussion, of course. Now if you eat a lot of meat alternatives, often these meat alternatives lack or have a low amount of lysine or methionine. So a lot of these projects are spiked or fortified with those individual amino acids. However, if you eat a well balanced meal, you typically have different plant based proteins in your meal that often compensates low lysine or low methionine. For example, one protein has high lysine and the other one has relatively high methionine. So they compensate. So that's why also your mom would say have a diet that is balanced with a lot of different sources, then the problems become smaller and you don't have to expect huge issues. Certainly not if you're consuming enough protein because you can compensate for lesser quality by greater quantity. And that's also the confusion with for example, the Game Changers documentary that everybody seems to have seen. If you have a huge football player and is consuming a massive amount of food, I couldn't even care less where the protein comes from because simply by the mass of protein, he or she already compensates for lesser quality by the simple mass, the large amount of protein. I mean the hype is also here. I got a phone call from somebody in hospital saying like, look, what do you think? Should we actually get only plant based foods for our patients? And I'm thinking like, oh no, because quality becomes important. When you actually have low quantity of food. So people that due to cancer or pain eat less, don't give them a high plant based foods. It sounds like you're doing a good thing, but you're not helping them at that stage in life.

Yeah, that's very interesting. Right. You would think that the most vulnerable people, the smallest people, the people eating the least, the people who are in this case greatest at risk for loss of lean mass, need to disproportionately focus on the highest quality sources of protein.

Everybody means well and especially that's also the communication between clinical care and science that often you think like, oh, a plant based diet is healthier. Yes, it's healthier if you're over consuming energy. And so if you need a more plant based diet will allow you to eat less energy, become less obese or less overweight, but it doesn't mean it's good for everybody necessarily.

Luke, I want to sort of, before we wrap up, ask you one more question to think about until we we meet again, hopefully in person here in Austin over the next 12 to 24 months. What is the single most interesting question you would like to explore in your lab?

I don't think I'm going to answer it in a year. I'll give this as a kickoff. We'll start over with that one. We measured protein synthesis in brain tissue in humans. People that have severe epilepsy, they get surgery in the brain. So the skull is lifted. These people were great. They actually wanted to participate in research. So we infused labeled amino acid tracer and we actually got part of their neocortex and we measured the synthesis rate of these amino acids incorporation in the human brain throughout surgery. Turnover rate of the human brain is almost three times as high as muscle. And of course there's slowly turning over proteins and fast turning proteins in your brain, but generically on an average level you can actually translate that into you have a new brain in about three weeks. Why do you still think you're you?

I mean that's an interesting question.

Yeah.

Do all amino acids cross the blood brain barrier?

To be honest, I think so. But there might be differences on the large neutral amino acid transporters basically for all amino acids.

So do you have any sense of which amino acids are disproportionately used by the cortex?

No. So you're asking now? I mean I'm not a brain physiologist so we did measure, I mean we published this in Brain, I think about two years ago. I think the amino acid composition, if I remember correctly of the brain was not that much different from muscle. Wow. But the turnover of your brain, and of course we know this, the brain has plasticity as well, just as your liver, just as and now we're also looking at tumors. So everything is growing and breaking down at an immense rate and I can't grasp my head around it because it means that everything is just, just doing this and you don't notice it. Everything seems solid, but it's constantly broken down and build up again. The amino acids in your brain now might be in your toe tomorrow morning. How does the body do this and how do all these organs communicate? It's really amazing.

Well, I think it is safe to say we won't have an answer to that question the next time we meet, but I think what we will have are some more questions and a deeper dive on this. So Luke, thanks very much for staying up late to talk with us this evening, your time. Look forward to hopefully seeing you in person. And maybe you'll wear some orange for me next time in support of your amazing countryman, Max Verstappen.

I will do so. Thanks for having me.

It was fun talking thank you for.

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