#31 - Navdeep Chandel, Ph.D.: metabolism, mitochondria, and metformin in health and disease

Foreign. Hey everyone. Welcome to the Peter Attia Drive. I'm your host, Peter Attia. The Drive is a result of my hunger for optimizing performance, health, longevity, critical thinking, along with a few other obsessions along the way. I've spent the last several years working with some of the most successful, top performing individuals in the world, and this podcast is my attempt to synthesize what I've learned along the way to help you live a higher quality, more fulfilling life. If you enjoy this podcast, you can find more information on today's episode and other topics@peterattiamd.com. Hi everybody. Welcome to today's episode of the Drive. My guest today is my good friend Navdeep, or Nav, as we like to call him. Chandel. Nav is a professor of medicine and cell and molecular biology at Northwestern in Chicago, which is where I actually met him to do this interview. Some of you may recall that name because Nav is one of the cast of characters that I went to Easter island with in the fall of 2016, the other being David Sabatini and Tim Ferriss, and we actually spoke about this stuff at length on a podcast that Tim recorded while we were on Easter Island. Nav's real area of expertise is in the mitochondria and in metabolism, and in fact he wrote a book called Navigating Metabolism, no pun intended, in 2015, which I highly recommend for anybody attending, especially people who a want to understand this stuff and b don't want to have to buy 17 textbooks and get into every unearthly detail. In fact, this book was written more for a general audience than a very specific audience, and I have a copy of it and love it. In this episode we talk about a bunch of stuff, but we talk about what got Nav interested in the mitochondria. We get into ROS or reactive oxygen species, something that I think many of you will have heard of. And and then we talk about some really nuanced stuff, like this stuff about mitochondria being actual signaling organelles and ROS maybe being beneficial for that signaling. In other words, we get into this idea that reactive oxygen species may not be all bad, it might not be a black white thing, which anyone who listened to this podcast realize. We love to explore things that aren't just binary. We talk about antioxidants and whether they're harmful or not harmful, and I suspect that a lot of people will have a point of view on that. And we get into mitochondrial DNA, which in and of itself is super interesting because some of you may already know this, and if you don't, that's fine. You'll learn it on this episode. But the mitochondria have their own DNA, distinct from the cell, and that DNA is of bacterial origin. It's also transmitted to us maternally. So there's a whole bunch of weird stuff going on genetically with the mitochondria that makes it super interesting. We get into a little bit of a discussion around cortisol. Nav is super interested in the role of cortisol in health. And then we talk, of course, about metformin. And I suspect that for some people listening to this, that's the only thing they're going to care about, because these days, everybody's asking me about metformin, which means it's front and center on a lot of people's minds with respect to longevity. Now, we don't even go as deep as we could on metformin. I'm saving that for another guest that's in the queue who I'm not gonna even say too much about that, but there's a lot more coming on Metformin. This, however, will be a great primer for that because you'll certainly understand how metformin works and how it may play a role in longevity. We talk about the role of the mitochondria in cancer, and again, we take a different view here from what you will have heard from other guests, potentially. We talk about nad, and NAD precurs. That, again, is another thing that people, I think, are pretty excited about. At the end, somehow Nav Jiu Jitsu's me and turns the tables on me and asks me the so what should I eat? Question, which anybody who knows me knows I can't stand being asked that question. But because it's Nav, I humored him a little bit. Before we jump into that, please keep in mind we have an email list. Every Sunday morning, I send out an email, and that email has a whole bunch of stuff in it. It's usually pretty short, though, but it's basically something I've read that's interesting that pertains to longevity, science, lipidology, performance, or just something that really interests me. Beyond those things, keep in mind, we put, I would say, an ungodly amount of effort into preparing show notes. And when I say we, I'm using that term very liberally. I actually do none of that, but I have a team that does that, and that is spearheaded by Travis and Bob. So the people who do spend time on those show notes always come back to us with feedback like, this is freaking awesome. Please keep doing this. And our intention is to keep doing that. So give us a reason to keep doing that by going and checking them out, especially when it comes to understanding some of the technical stuff that we talk about here and getting some of the references. And lastly, if you are digging this podcast, please head over to Apple Podcast Reviews and leave us a review. Especially if it's positive, but if it's negative, at least have the courtesy to give us some constructive feedback so we can improve upon whatever it is you don't like. So without anything further to add, please welcome Nav Chandel. Hey, Nav. Good to see you again, man.

Great to see you, Peter. Yeah.

The last time I saw you, me, you and David Sabatini were hanging out near the airport in San Diego playing patty cakes. We were. You guys were drinking some really lame stuff.

Rose. I like sparkling stuff.

So what got you interested in mitochondria?

So when I was in college, I was a math major. I loved math. Right. I probably did math because as an immigrant, you moved from the Himalayas to Miami. Got a funny accent. Math is a universal language, Right. So I became a math geek. But everybody's got to make a living. And so I worked in a laboratory, which was in a hospital, and it was a transplant laboratory. And the biggest thing in transplant is how do you preserve organs. I mean, you have an md. I don't have an md. I think that's a fair statement. And there's this Wisconsin solution that was used to.

We used it liberally.

And so I wanted to make a better version of it as a 19 year old. And if you got to do that, then you got to think about metabolism. The first set of experiments we did, and I went to a Facet meeting, and it's called 3 phosphoglycerate. Protects against anoxia, injury of hepatocytes. Anoxia is low oxygen, you know, the absence of oxygen. And that got me interested in metabolism. And if you're a mathematician and you start working metabolism and eventually mitochondria, it's electrons, it's enzyme kinetics. Right? There's math. And in fact, my PhD is on enzyme kinetics. And so I'm doing a PhD on an enzyme that's very critical for respiration. So, like, the enzyme that uses oxygen in every cell is what I did my PhD on and how that enzyme works under different oxygen levels. Again, lots of math. Michaelis Menten. But much more sophisticated enzyme kinetics. And then after that, the thing that was probably the most influential discovery in the mitochondrial field for me. And I still think this is one of the three greatest discoveries in the mitochondrial field. Of course, it'll be contentious because lots of people might take issue with it. So the first one is Hans Krebs and the TCA cycle. In 37, there's the 60 paper by Peter Mitchell, how youw Make ATP. So those are the two big things, right? Energy, bioenergetics and biosynthesis. Because the Krebs cycle eventually makes carbon molecules, which are the backbone for lipids, nucleotides, amino acids. The third big experiment was. Came out in 1996. So Xiaodong Wang, when he was at Emory, found that the protein that I used every single day to do enzyme kinetics. So I worked on cytochrome C oxidase. That's an enzyme already. It tells you cytochrome C oxidase means that it uses cytochrome C as a substrate.

Let's take a step back for someone who's going to be listening to this, who doesn't know everything about it. Before we get to Huang's experiment, tell us who Hans Krebs, what he did, what the TCA is. A lot of people will think, I kind of remember this from high school biology, but give me a quick refresher course. So I'll prime you a little bit for it. A cell brings glucose in, just to make it easy. It turns glucose through a number of steps, enzymatic steps, into two smaller three carbon units called pyruvate. Assuming the demand for ATP is not extraordinary and there's sufficient cellular oxygen, what would be the most efficient way to get energy out of that?

Pyruvate has two choices. It can either become lactate. And usually that happens when there's not enough oxygen.

Yeah. And the other thing I always like to say to people and. Or when the demand for ATP is exc. Exceptional.

Exceptional, Right. But the other place where it really goes is pyruvate goes into what's called the TCA cycle, and it's really a cycle. So where glycolysis is a series of linear steps, you know, A, B, C, D, E. Then you get to pyruvate. And pyruvate actually goes in a circular.

And this occurs in a part of the cell called the cytoplasm.

The glycolysis part does. But the TCA happens in the mitochondria.

Which we're gonna get to.

So pyruvate to lactate is all in the cytoplasm, but pyruvate then gets imported through a pyruvate mitochondrial pyruvate carrier. And that's important because it could be a target therapeutically in certain diseases. I think most of us appreciate. And this is what you're getting to while you're focusing on pyruvate. Pyruvate has two choices, go to lactate or the mitochondria. So clearly, the pyruvate to lactate is cytoplasmic pyruvate going into the mitochondria and eventually becoming acetyl CoA. That's an important reaction. So where pyruvate goes and how it gets in and out is quite important. And discovery of that transporter gives you a way to maybe target that.

So now pyruvate's in the mitochondria.

It's in the mitochondria, and then it's a 3 carbon, and it becomes acetyl CoA. Acetyl CoA is very important. And we can get into it later because acetyl COA can acetylate to acetylate acetate.

The ATE means you put it on, basically.

Put it on.

Right. ACE is an enzyme. It's the end of an enzyme to take it off.

Yeah, yeah. And then it goes through the TCA.

Cycle, which stands for.

So some people call it the Krebs cycle, because Hans Krebs discovery. So try carboxylic acid cycle. Some people call it the citric acid.

Cycle, so we'll use those interchangeably.

And so the TCA cycle, as it's going in the mitochondria, it generates NADH and fadh. These are these reducing equivalents which then feed electrons to the electron transport chain. And the electron transport chain can pump protons, which is basically like a battery to generate ATP, which is the currency that makes cellular functions work.

So the way I like to explain this to people, because I think it's such. It's just so damn elegant. You look at a piece of food and all you see is this potential energy, you see? And of course, within all the bonds that exist in food, which is where we get our energy from, you have carbon, carbon bonds, carbon hydrogen bonds, carbon, nitrogen bonds, carbon, oxygen, nitrogen, oxygen. I mean, there's only a finite number of covalent bonds that exist, but the two most energy dense, I think, are carbon. Carbon and carbon hydrogen. Correct. Isn't that where the majority of the chemical energy is liberated?

Right. Those electrons have to eventually be accepted by something.

Right.

And oxygen is the final acceptor of all of those.

So in some sense, the whole purpose of eating is to take a potential energy that is in a chemical form, turn it into an electrical potential energy inside the mitochondria, and then eventually pull a little jiu jitsu and turn that back into a chemical electrical energy via the conversion of ADP to ATP. I mean, to me, that's the simplest way to explain this. And.

And it's just like a battery, right?

Yeah, it's kind of amazing.

Well, that's why they call it a power plant. Yeah. What is the, you know, in the public domain, what do people call mitochondria?

Yeah, Power plant to the cell powerhouse.

Power plant.

Right.

And for exactly the elegant explanation you just gave. So we're already talking about the dogma, Right. Which is all about ATP. So if you asked in 1996, what is the major function of mitochondria?

Make ATP.

That's it.

And for good reason. Because let's call a spade a spade. As we know, if you interrupt that cycle for even moments, it's uniformly fatal. Most people are familiar with a toxin called cyanide. Cyanide is fatal at incredibly low concentration. There are only a handful of toxins that are more potent, more lethal at lower concentrations. But. And they all tend to, you know, like Detroit toxin, I think, blocks a sodium channel transport. It might be even more lethal. But why cyanide is so lethal is it disrupts that process you just described.

Indeed. But it's not clear which organ fails right away. So we'll get into that in a second. The genetic experiments don't totally confirm that. Interesting.

I'd like to hear more about this.

Yeah, we're going to get into that in a little bit. So in 1996, if you ask that question, it's all about ATP. And so there is this protein, cytochrome C, that's part of this energy generating system, which is ATP. And Xiaodong Wang and colleagues found that if that protein gets released from mitochondria into the cytoplasm, which had never been detected before, it can rapidly kill a cell. So now the decision for life and death is based on the localization of cytochrome C between the mitochondria and into the cytoplasm. So if it's in the mitochondria, it helps you generate ATP. If it's out, it kills you.

And I assume it only escapes when the mitochondria is under the most intense duress and destruction.

So when a cell is under some sort of toxin, stress could be low oxygen, complete absence of oxygen, could be a growth factor, which is telling the cell to live is not present. Death by neglect, let's call it that. When the cell is being neglected either by nutrition, starvation, complete depletion of nutrients, or complete depletion of growth factors that keep them happy, the positive signals, then cytochrome C gets released. But it's a very profound discovery because this is a protein that we were all convinced had only one role. We call it the day job now make atv. It's got a moonlighting job that sometimes it leaves the mitochondria and doesn't make ATP and bam, it starts a cascade which is called apoptosis, cell death.

I was just about to say, is this part of a programmed cell death as apoptosis?

This is programmed cell death. So here, here I am in 1996, I'm 26 years old, very happy, doing enzyme kinetics, bioenergetics. And I see this, this finding and all of a sudden I stopped caring about ATP, because in part a lot of that, how it works was more or less discovered. I didn't have the insight because obviously why would I be still working on it? I still thought there was more to be discovered, but the reality was I was probably just cleaning up little side things in that field at best. And that's not hopefully to disparage the bioenergetics field. But there's still some important questions there, more at a structural level, and there's some beautiful work there. But you know, what I was doing.

Was the kinetics had been worked out now.

I mean, some of it had been and not. But this was great because this connected the mitochondria just not for energy. And what the mitochondria does almost in a cell, like in an autonomous way, like it just does ATP. But this says it starts to control cell biology and function, like the decision of life and death. So obviously the first thing we think about is it can't be selected just for death, right? It must be doing something interesting. So let's think about what else it might release. And one of the first things that we thought about, which is something that people had noticed for almost 40, 50 years before they release superoxide and hydrogen peroxide, reactive oxygen species, oxidants, free radicals. And most people in the mid-90s assumed that mitochondria only release this so called cleaning molecule or toxic molecule when the mitochondria is damaged. Maybe in neurodegeneration, maybe in aging, we'll get into this, maybe when a heart failure, when there's low oxygen ischemia. So in other words, it's there almost as A way that the mitochondria, when it's not working well, it just spills out because mitochondria has all these electrons and they have to. They're not working. Those electrons just spill into superoxide and eventually to hydrogen peroxide. And what we thought about is, well, maybe nature could have used that as a signaling molecule, in other words, to dictate cellular function. And so there were some papers just right as we were working that came out and that showed, in fact, H2O2 could be a signaling molecule, but they didn't think it was necessarily mitochondria. So there's a neat little story that there's another system in the cytoplasm that can generate H2O2, and they thought that's all about signaling. Signaling basically means it's making decisions in the cell to die, to live, to proliferate, to grow, if you're an immune cell, to make cytokines, to do inflammation. But the mitochondria only did it when it's not working and it's spilling all these electrons into hydrogen peroxide and it's causing damage. And we actually showed that, in fact, under physiological conditions, you can make hydrogen peroxide, which has a beneficial effect. And so from that point on, the first paper was published in 98, now, almost 20 years, there's my group and lots of other people continue to show that mitochondrial generation of this cleaning molecule can actually cause cellular functions to happen. For example, we've shown our T cells, which are part of the adaptive immune system, right? They fight off pathogens, right? Or innate immune cells as well. Just immunity in general, right? They fight off viral infections, B infections, that H2O2 is used by those immune cells to properly function. And what's the profound implication, which is what you're going to get into is antioxidants. Are they beneficial? Are they harmful? So already, to the audience, it should be obvious, if I'm saying that something you think is a cleaning molecule only then antioxidants are great. You're getting rid of this toxic molecule, cleaning molecule, you're getting rid of this bad stuff. But if it's also there for good purposes, like immune function, it paradoxically could.

Be bad to have an antioxidant in your system at a time when you need enhanced, especially adaptive immunity.

And so one of the clinical trials that came out was in the icu. Sepsis, that's the big disease, which is basically, on a simplistic level, tons of inflammation. And those trials failed. They made them worse. Antioxidant Trials in cancer, again in lung cancer, vitamin E trial, it didn't have a curative effect.

Yeah. Do you think it's overly simplistic to say that there ought to be a balance in the body between pro oxidative and antioxidative stress? And it's never the case that one is absolutely good or absolutely bad. It depends on the state of the organism. So under perhaps a normal state, a more antioxidant, that is, let's reduce the Ross is the right thing to do. But to your point, when the immune system is required to take the front step, cancer and sepsis being two enormous examples that having too much antioxidant property can actually be harmful. And that's a moment when you actually want to be able to inhibit that process.

So one of the interesting antioxidant trials. Now, again, caveat in this experiment, I'm going to tell you, is an exercise experiment. So we all agree that if you do vigorous exercise, it has benefits and you can take a biopsy and look at all the genes that exercise turns on. And these are genes we think are beneficial to the host, to the rest of the body, if you give high doses of antioxidants. So that's the caveat. It's high doses, probably not the doses that most people use. It actually turns off that beneficial response. So in other words, to your point, I think what you were making is that when the system is stressed and the mitochondria integrates that stress, how does it pass that information back to the cell? We think that perhaps it releases hydrogen peroxide. So like, clearly exercise is a good example, right? That's the stress to the system, to your muscle. And how does that muscle then turn on all these genes and blah, blah, blah. We think it does it perhaps by releasing hydrogen peroxide.

Prior to this insight Nav, is it the case that people knew the mitochondria was signaling but assumed there was a different molecule that was used to transmit the message? Or that people didn't actually think the mitochondria were playing any role in signaling. And we're still back in the ATP only paradigm.

They did think about signaling only in the context of pathology. So in other words, so there are these childhood diseases like Lee syndrome, where there's a mitochondrial mutation in a particular protein and there's a devastating disease. Kids don't live that long and clearly that's a mitochondrial mutation and a protein that gets mutated doesn't function properly in the mitochondria and you get a childhood disease. So how does that work. So people say, well, probably ATP or maybe some. Maybe too much. Ross. That's right. So in that context, there was some idea of signaling. Right. Because. But this is saying under physiological conditions, exercise is not pathology, it's physiologic. When you get a pathogen. In other words, when you get a virus or bacteria, that happens, you get a cold, your immune system has to be activated. That's not pathology. That's a good, normal response. So under all those conditions, the mitochondria is playing a signaling role. And so I've coined this term. So there's the mitochondria as powerhouses. So my talks never talk about that. The title of every talk I give all around the world is the same. Not very original anymore. Mitochondria as signaling organelles. And H2O2 is one way, cytochrome C is another way. Right. That's probably maybe the most original because that's for cell death.

By the way, 22 years later, do we still think that that work out of Emory is what has been added to that body of knowledge about the role of. In apoptosis and programmed cell death?

They know the whole pathway, but it.

Is still coming from the single enzyme.

Yeah, yeah, yeah.

Wow.

There are these proteins called Bax and back, that make these pores in the mitochondria. And it releases this 13 kilodalton protein. It just releases that protein right out. And then it binds to a whole bunch of other proteins, Apaf1 and Caspase9, and starts this proteolytic pathway which eventually causes cell death. So they've worked out all the biochemistry.

But the main point remains conceptually.

Yes.

Wow. That's. That's. You don't get that all the time in biology, that 22 years later, the punchline is still the same.

The punchline is the same. I mean, you know, there's a lot of details that I have to. But conceptually. Yeah.

Are there other ways, by the way, that cells can undergo apoptosis?

Yeah, Now. Now there's a whole industry. Every day there's a new version of it. And in fact, we can talk about one new one, which I'm very excited about. So clearly we can show that H2O2 in many contexts will signal for positive responses, immune functions, as we just talked about, the exercise response. But clearly there must be cases where H2O2 or other reactive oxygen species cause cell death. How does that work? And there is a new form of cell death that was just discovered by Brent Stockwell and Scott Dixon at Columbia or so 2011, and it's called ferroptosis and it's basically taking H2O2. If there's free iron, you can make a hydroxyl radical, an oh, and that will then make a lipid hydroperoxide. So if you have polyunsaturated fatty acids, it will basically make it into a lipid hydroperoxide, which can be very toxic. So the bottom line is there are times where hydrogen peroxide with iron and lipids, the three can come together and make something called a lipid hydroperoxide, which will cause cell death and ferroptosis. The good thing is your body's full of an enzyme called GPX4 that gets rid of it all the time. Now, if you don't have that enzyme, you're in big trouble. So there are places where clearly H2O2 is positive and it can become very lethal by making another form of reactive oxygen species. But again, nature has selected, I can't count all of them, but at least 30 enzymes which are constantly mopping up these reactive oxygen species, keeping them quite low so you don't get to those toxic levels.

So when we go back to apoptosis, which I didn't think we were going to even talk about this, but it's so great to be able to bring it up because I think for many people that they're still probably thinking, what are we talking about? Apoptosis. So if a cell undergoes a genetic mutation, nuclear, genetic, we'll come back to mitochondrial genes later. But if a cell undergoes a genetic mutation in the nucleus that's unrecoverable, it will, on a good day, kill itself, it will commit suicide and that would be one of the things that would drive apoptosis.

You're getting to tumor suppression mechanisms.

Yes, exactly. That's where I'm going. Do we know if this mechanism you just described of apoptosis plays a role in the type of cancer apoptosis that we want to see? And if so, the implication is the nuclear genome must be communicating with the mitochondria.

Yes. So there is a drug that targets that. Abbott made a good friend of mine, Steve Fesik, was involved in it, who's now at Vanderbilt. And So in the 90s, they figured out the structure of a particular protein that controls cytochrome C release. And so they've made a drug against that protein and specifically in cancer, to make the cancer cells sensitive to chemotherapy. Basically. Right. So what do cancer cells love to do? They upregulate anti apoptotic proteins. Right. So what does that mean? That means they turn on a whole program which is basically an anti death program.

Right. Protects them from everything.

You just described this release of cytochrome C and all of that st. And so one idea is, why don't we target those anti epoptotic proteins, these anti death proteins? Because if we can target them and prevent them from functioning, then when we give them chemotherapy now, the cells will die a lot quicker. The problem with chemotherapy is all these anti death proteins are there. So a normal cell, like, you know, so doxyrubin is a good case. A normal cell being a heart gets toxicity at the same time you're trying to kill the cancer cell. So it's not selective. It's not selective. Right. And so people have been targeting these anti apoptotic proteins in cancer as a mechanism to make chemotherapy more effective.

Yeah.

So this mechanism, we were talking from 1996, I didn't think we would get into this, but it's been a long time. This is my previous life. I haven't thought about this in a long time. But yeah, it still applies and it's very important for cancer.

So while we're still going on history, I just alluded to something a moment ago that I think is an important point for the, for the listener to understand, which is, you know, everybody knows, or I guess, you know, most people who were thinking about biology would understand that the part of the cell that contains your DNA is called the nucleus. And we've got lots of genes in there, about 20,000 genes. But the mitochondria has genes too. Not that many. What is like 35 or something?

Yeah, it's 37. And there's the key point. There is there's 13 genes that are essential for the respiratory chain to work. And the respiratory chain is where all that oxygen is being consumed. That's the one that makes that energy, that electrochemical energy we talked about that ultimately gets converted to ATP. So the key subunits of the respiratory chain which generates that battery that we've been talking about, it holds onto those 13 genes that are critical for it. So, for example, complex one and is one of those respiratory chains, there's these five complexes. Well, one of the complex has 45 subunits, but a few of them, which means 45 proteins make this huge complex. But a few of them are in the mitochondrial genome. Complex three, very important. My favorite complex. Everybody's got to have a favorite complex. And so my favorite complex.

I see a T shirt here.

Yeah, yeah. So my favorite complex is complex three. And it has one gene that's still in the mitochondria, by the way.

Are you saying that just to be contrarian, because complex one is actually the coolest? Because, like, how can, how can complex one not be your favorite mitochondrial complex?

I'll tell you in a second.

It's just because you're cool and you're, you're too cool for school because you study this. The rest of us who are in the peanut gallery, we, we default into complex one being the coolest.

So you're saying I'm a mitochondrial hipster, right?

I think you are. You're leading the charge on this.

No, but think Complex 3 is very interesting because it has only one subunit is in the mitochondrial encoded by the mitochondrial genome at one point, the mitochondria. So going back in evolution, we think there was an Alpha Proteobacteria and an archaea, probably a methygen. And these two prokaryotes got together and had a symbiotic love affair.

And just to explain to the. So we are eukaryotes.

We are eukaryotes.

And what does that mean again?

We've got a nucleus, got a bunch.

Of organelles, and what's a prokaryote?

It doesn't have those organelles.

Got it. So bacteria.

So you got a bacteria and you got an archaea and they got together. So we think the archaea is kind of where the nucleus came from. And the Alpha Proteobacteria is the modern day mitochondria. And one of the best evidences is. I remember an experiment that I did as a graduate student and was, I took a bacteria that somebody had discovered in the 70s, a beautiful paper in nature. And this bacteria, if you gave it like succinate, like a mitochondrial substrate, and it'll grow on it and it would respire very similar to modern day bacteria. Modern day mitochondria.

Wow. A nice, elegant proof of concept that the mitochondria are basically bacteria. Came from bacteria.

Yeah, they came from bacteria. Yeah. I mean, you know, those who doubt evolution, I always tell them that every organism I know burns glucose very similarly.

It is hard to make that. Yeah. I don't want to go down this path because we'll simply alienate a million people. But it is interesting to think. It strikes me as it's very hard to come up with an alternative explanation for why you would have this effectively foreign DNA inside every cell. And I'm so intrigued by this mitochondrial DNA thing because again, it's such a tiny number of genes on a Relative scale. But yet they're so critical. And to my knowledge, I don't think there's any other organelle that carries its own genes with it, is there?

No, it doesn't. And what's interesting is that it only encodes for like 1% of, you know, there's, I don't know, 1,000 proteins in the mitochondria.

But it's absolutely critical you knock those 37 genes out.

No, you knock out any one of those genes out, you're done. You're done. So why did the mitochondria hold on to those? Right. So it basically gave up. You know, if it has a thousand proteins, it said, you know what, the nuclear genome.

Yeah, you take 990.

Right. I'm gonna hold on. And they're all the critical catalytic subunits and all of this. In other words, they're really essential for the function and I'm gonna hold on to them. It's almost like it doesn't trust its symbiotic partner. Right, right. It's like a love affair. Right? Yeah. I love you, but I'm going to hold onto a few things.

Yeah. And it's very interesting because I believe I could be wrong, but you could make the case and correct me if I'm wrong, that we pay a price for that lack of trust. In many ways, don't you think the nucleus would be a better steward of those genes? Doesn't the nucleus have more ways to protect the genome than the mitochondria? And therefore, don't we run a greater risk of disease when the mitochondria, with its beautiful stewardship over its precious 37 genes, gets under stress?

Yeah. This is a very important point you're making. Basically, we have a lot of DNA repair enzymes, Right. That are all in the nucleus. There's these proteins called histones that cover the double strand DNA. So, you know, and there's many mechanisms to protect our genome. And the mitochondrial DNA is just like this round little circle.

It's so vulnerable.

It's totally vulnerable and is especially vulnerable because the sight of those free radicals we've been talking about is right there.

Exactly. That's the worst part. You put these very exposed, fragile, not protected genes in the presence of a potential toxin.

Right.

I mean, I wish I had more time to think about. I wish I was quicker on my feet. Cause I think I could think of an elegant analogy of how counterintuitive that is. Right. It's like leaving the keys to the kingdom in the hands of the guy next to the. You Know, I can't even think of it. I'm not smart enough. But, like, it doesn't make sense.

It's like a hen house next to where the foxes live.

Yeah, yeah, yeah, that's. That's a better. So. So what's the advantage of it?

One thing I should stress is that potential molecule that could be damaging. Superoxide, hydrogen peroxide, hydroxyl radicals, the mitochondria, where the mitochondrial DNA resides in the mitochondrial matrix. There's tons and tons of antioxidants there. So even though that the DNA itself can be protected, it's protected from a variety of these toxins because there's so many proteins that clean them up. Essentially the highest level of antioxidant activity sitting in the mitochondrial matrix. And I think that's because. To protect the. That. That almost naked DNA.

So one other.

That makes sense.

Yeah, no, it really makes sense. Nav. Thank you. Because I've. This has been on my mind a lot lately because I saw a paper recently, I may have even sent it to you, about another hypothesis around inflammation, the effect that inflammation can have on the mitochondria. And the mitochondria starts to shed its DNA, which actually kicks off an immune response as it exits the cell. And that's what got me thinking about this. I was like, wait a minute. That's a really good point. That must happen an awful lot. And actually, what did they peg it to? They pegged it to hypercortisolemia, which we're gonna come back to, because you're kind of a cortisol guy, too. When it's all said and done, you're a cortisophile, if such a word exists.

No. You know, in the hormone world, I call people like you insulin profits and many of your other former guests. I'm the cortisol prophet. Right. I think that's the missing link for a lot of stuff without any real data. But it's just my own intuition.

Well, these data that I saw actually suggested that the hypercorticolemia, not just cortisol, but other glucocorticoids and including other hormones from the adrenal glands, could really become toxic to the mitochondria at high enough doses. And it was basically jettisoning broken strands of mitochondrial DNA that basically kicked off immune responses and sort of you had these inflammatory responses that resulted from an immune response to mitochondrial DNA being damaged by cortisol. So bringing it back to kind of dinner table trivia, the other thing about mitochondrial DNA that's interesting is it comes from one parent. So, yeah. Tell us what that's all about and why that's the case and what the.

Well, we don't. Yeah. I mean, again, two big questions in the field. One is why we still keep those genes. We just went over like, you know, it's kind of like it doesn't trust. And then the other one is why does it come from the molecules?

And there's an implication there, which is you have far less dilution by generation.

You do, but why did nature select that? Right.

I'm asking you.

I don't, you know, I don't have a good explanation.

Even teleologically. You don't have a good explanation?

I really don't. We think more about the function of mitochondria. I mean, there's a whole group of people who think about the bottleneck of mitochondrial DNA being passed on, but I actually, I don't have a good explanation to it. It's a fascinating. I think lots of people have very interesting ideas around it. But to be honest with you, why we continue to have those genes and why does it come from the mom? I think these are still outstanding questions in the field. Something that my lab doesn't work on. And as you know, Peter, I don't comment on things that I don't work on just because you can speculate, but you want to have good data. But I do want to talk about one aspect of mitochondrial DNA, and this is in my wheelhouse, which has nothing to do with those two questions, which is there's a third role going back to signaling. So if you think that H2O2. So what does mitochondria dump into the cytoplasm? ATP for energy, we think H2O2 for signaling, to do immune responses, exercise, et cetera. What else could it release? Well, one of the things it could release is mitochondrial DNA, which would then kick off an immune response. The only thing about that hypothesis that I struggle with is how do you release mitochondrial DNA in a physiological way without not releasing cytochrome C?

So you're saying maybe all the damage, that's. Well, okay, I'll give you an answer. I'm making this up now, but maybe you are also releasing cytochrome C. The cytochrome C results in the apoptotic death of the cell. But the DNA gets into the plasma, which is where the immune system begins to recognize it. So you could still have apoptosis at the cellular level, but globally. Right. So locally, apoptosis, globally, you have the immune response.

Right. So it depends on that cell dying.

Yes. Which makes sense.

That's okay.

Yeah. You're saying. I don't see a way that that could happen with an intact cell just willy nilly passing off its mitochondrial DNA. That makes sense.

Yeah. So that's. And people have sort of proposed that it could do that and maybe it can. I'm open to the idea, but someone's got to show me how you selectively release some mitochondrial DNA without releasing everything else, you know, and where H2O2 and ATP get released in a much more.

Benign fashion without ATP has an active transporter.

It has an active transport.

H2O2 goes by diffusion.

We think so probably. Maybe VDAC channels, voltage dependent anion channels could maybe release the superoxide which then gets converted quickly to H2O2 right outside the mitochondria. So again, the mechanisms of. You know, it's kind of like water, right. Before the aquaporins, we just thought water, just water.

Yep. Diffusion freely.

Now there's active transport. Actually, Pioneer was at your institute at Hopkins, right? Got the Nobel prize for the aquaporins.

I didn't know that.

Yeah. And so when we think about signaling, the simple idea is what gets released? ATP. What gets released without cytochrome C getting released. That's the key. Right. Because cytochrome C gets released.

It doesn't matter.

Doesn't matter. Everything is. The cells are on its way to die. So what gets released? So we know hydrogen peroxide, ATP and metabolites. Metabolites are always being released. So citrate.

Well, CO2.

CO2, yeah.

For obvious reasons.

But citrate. Right. So citrate is very interesting molecule. Citrate gets exported from the TCA cycle into the cytoplasm where it can get broken down back into acetyl CoA, which can then be a primer for making new lipids, new fatty acids. Right. But also that acetyl COA can cause acetylation reactions like on histones to control gene expression. The so called chromatin modifications, meaning it.

Goes back to the nucleus.

Yeah. So, but that's quite. So Citrix could be a signaling molecule, right?

Well, if it's doing what you just described, it would be.

It would be. Right. And so there's a bunch of. So in my wheelhouse, the ones that I like to think about, H2O2 and TCA cycle metabolites that get released and they do, they're in constant flux between the cytoplasm and the TCA cycle. And they can control gene expression through chromatin modifications, through histone Modifications.

So, because we're going to come back to this through a totally different lens, I want to also have you and or me explain somewhat to the listener what this idea means of histone deacetylation. And basically because those terms I think, are largely for foreign to even reasonably informed folks. But I think the reasonably informed folk will understand what epigenome means, what a modification of a gene means, and how genes are potentially silenced or upregulated. Is that happening often in the mitochondrial DNA as well, or are they pretty much just on their own?

They don't have.

So what transcription factors tell those genes when to turn on again?

They're all nuclear encoded and they have to be.

Oh my God, this is so staggeringly inefficient. There has to be a reason for this that hasn't. Yeah.

Why? You know, I mean, I know you're fascinated by mitochondrial DNA and I don't have good answers for this. That's why we didn't want to talk about this, which is, you know, why does it continue to have those genes?

I'm hoping there's a college student out there or a graduate student out there who's thinking like, I want to understand what could be the reason for this. Because if you can find, in my sort of somewhat simplistic way of thinking about problems, I think, think when you look at something in nature and you don't have a clue why it's occurring, if you could get a clue why it's occurring, you will unlock a whole bunch of other knowledge as well. That might not be true, but that's like kind of a working hypothesis. And in this case, think of like, you're one of the world's experts on this topic and yet you're acknowledging there are so many fundamental, obvious questions like a high school biology student could ask. The questions I'm asking on this topic, these are not like super nuanced questions and yet the field doesn't know the answer. That's really interesting.

To be fair, there are people who think a lot about this and they have opinions on it. But what I'm just simply saying is what is well established is that mitochondria generate ATP and they generate metabolites for growth. And all of this stuff, including my signaling hypothesis, you know, it's still a work in progress, like everything we've talked about, just full disclosure to the audience. It's a work in progress. So if you're a high school student, come on and join the party. There's a lot to be discovered here.

Well, that's the beauty. Right. It's not like we're ever going to run out of questions that need to be answered.

And I think the difficulty in the field historically has been the bias about ATP and thinking about energy and solving that problem. And the great biochemists did that. They figured out how we make ATP fundamental to life. They figured out how we make the metabolites and TCA cycle. But now is getting much more challenging because those same processes can control gene expression. Well, how does that work? I don't. I mean, I mean I have some reasonable hypotheses we're testing, but I'm very careful, as you know, to not give a strong opinion on a work in progress.

So let's shift gears a little bit to talk about some other broad mitochondrial questions because I do think that people today, and maybe it's just the bias I experience because of what I'm looking for. So this might not be the. But it seems that the interest in mitochondria has exploded. I think people are realizing there's a lot going on here. It's more than we realize. Mitochondrial function is now a term people use all the time. But they're not just talk. I don't think they're just talking about oxidative phosphorylation. I think they're talking about broader things. And when we talk about aging, we talk about something's changing in the mitochondria as we age. When you think of hallmarks of aging, we can debate the merits of some of them, but something's different in the mitochondria of an 80 year old versus an 8 year old. What are some of those changes?

Well, so this is going to get contentious now because the data suggests that you have a decrease in mitochondrial DNA. Some of that mitochondrial DNA has deletions that the capacity to do maximal ATP generation goes down. Oxidative phosphorylation, the key there is maximal. So one of the perplexing things, and this is really perplexing for me a much more. So I don't think much about mitochondrial DNA and that's why I didn't have good answers for that. But this is the one that really is, and this is fundamental to the aging. When you're born, let's say you have 100% capacity and then as you age that capacity, if you're giving 100% mitochondria, you get an A plus 100%. How far do you go as you.

Age just to make sure I understand what you're talking about? Are you saying Amount of ATP generated per mole of oxygen. As a metric.

As a metric we could use. ATP generation is a simple one. Okay, so it's ability to.

For every mole of oxygen you generate X mole of ATP and whatever your maximum is, it declines.

And you can burn it through fat, glucose, carbohydrates, proteins. It's working at its maximal efficiency. It's everything's fine. And that efficiency now declines with age. The question is, is it ever rate limiting? So you and I, I mean again, these are loose terms at any given point are using maybe 10 to 20% of our maximal activity.

Hey, speak for yourself, dude. I mean, I do wind sprints every day, man.

Okay, so when you, when you do that, you might go up to 40 or 50%. I didn't realize that you don't hit that. And so when we knock out a protein in the mitochondria and we knock out it completely from 100% to 0%, pathology happens. We go from 100 to 50%. We never see any pathology.

Even under stress?

Even under stress. If anything, they behave better. So.

Oh, this is interesting.

Yeah, this is very interesting. Right. And so then when I look at the data on aging, sometimes, you know, in some tissues it goes down by 50% of maximal, maybe 70%, 80%. But is that ever rate limiting?

Well, if what you said, if what you said is true, then it would not be rate limited within a certain band.

So I am on. And this full disclosure, again, I am in way out in outer space on this idea. I don't think mitochondrial function, and if you don't have a disease, let's be clear, normal aging we're talking about, right? You don't have cardiovascular disease. You didn't have, you know, you're just sort of, you're pretty healthy. And I don't think a healthy heart, you know, is really limited for mitochondrial function. The implication there is that most people think, again, this is like dogma, like with antioxidants. Right. They're good for you. That mitochondria are declining. Let's give supplements that boost mitochondrial function. Is that a fair statement?

Heck yeah.

I think there's no evidence to support that.

What is the most popular of said supplements? Would that MITOQ be a popular supplement?

No, that's an mitochondrial targeted antioxidant. So we can talk about mitoq in a second. Actually, we should maybe talk about mitoq. It's quite fascinating.

Yeah, let's go back to your question.

What is. Well, no one's actually has, you know, people have Been trying to have these, what's called mitochondrial biogenesis activator, something that will make more mitochondria and I'm not quite sure what supplement people use that they think is the best one but I would argue the opposite which is that they're not rate limiting and if anything maybe you can decrease mitochondrial function in certain tissues a little bit to activate stress responses which will then fight off if you do get a disease. And this is going to go into.

Metformin which I was just. How do you read my mind? I didn't even start mouthing the word yet. Why did you know I was going to bring up Metformin?

Because we think metformin is a weak mitochondrial complex. One inhibitor.

Yes.

Another which is part of the respiratory chain.

That's why complex one is my favorite complex.

That's why your complex one is your favorite.

But I'm a poser. I'm a mitochondrial poser. So. Okay, because what I was going to actually ask you was on the heels of that when you give metformin you inhibit complex one, you are now reducing mitochondrial function. Mitochondrial function. And if what you're saying is correct, you would need a lot of metformin to generate actual etc toxicity.

Right.

Do we ever see that?

No. I mean you do if you go.

To certain doses but not physiologically. We can't do we the anti diabetic.

Dosing that is given to people. I mean there's some toxicity can happen due to certain patient populations. But it's a very safe drug. Right. It's used by almost what 300 million people now? It's estimated to be used by half a billion people as the diabetes epidemic explodes in China and India. So no one quite understands how metformin works. We think it's, you know, it has three effects clearly it lowers glucose production in the liver, it has some anti inflammatory effects and it has some anti cancer effects.

Well let's talk more specifically about it. We were joking around when we were at Easter island that our next trip actually needs to be to France to see the goats, to see the lilac. Where from where wasn't that in France where they. Where that medicine Metformin came from. So two sentences on how metformin was discovered.

They noticed these goats that were eating were pretty healthy.

I love this, I love these stories. But you know yesterday I was talking to Ted Schaefer about goats as well. So I love that the two Northwestern guys, the only two podcasts that will ever have goats are going to be these ones.

So what's Interesting about metformin, I think, is it? It got approved as an antidiabetic drug. People went back and looked at people who were taking different diabetes and epidemiologically found that there was a lower rates of prostate cancer. So you're probably talking to Ted Schaefer about this, right? And they're lower in breast cancer. So then people started investigating it as an anti cancer drug. Then some people started noticing, wow, it has anti inflammatory effects. I've talked to our friend David Sabuttini that isn't it interesting that rapamycin, anti inflammatory, somewhat anti cancer, promotes metabolic health. So how does that all work? Well, he'll argue it's all mtor. I said, well, I would argue how can metformin do three very disparate effects. Anti diabetic, anti cancer, anti inflammatory, just like rapamycin would. It must be hitting a node that's very important for the cell. So metformin doesn't hit.

MTOR does though. AMPK does.

The AMPK, but that's due to first hitting mitochondrial complex 1 and then activating AMPK, which can repress MTOR. But the analogy that I'm using basically is MTOR, we know, is the master of the universe. So is mitochondria. Right. So if you inhibit mitochondria, not to the point where you cause toxicity just enough, you can activate a variety of pathways which can promote, have anti cancer, anti diabetic and anti inflammatory effects.

Now, metformin is somewhat tissue specific. It seems to have a preference for the hepatocyte.

So it gets into the kidney and the liver. I think we've talked about this before about mtor. Right. So MTOR and rapamycin, Right. Why wouldn't you use rapamycin? Well, it might get in everywhere. I think David's argued and you've argued, wouldn't it be great if you can get metformin to go to the liver, to the liver, but not to the skeletal muscle? So metformin already has a little bit of that property. It only gets selectively into. It doesn't get into the heart that well. Right. So it doesn't infect your heart function, not directly. It affects your liver, your kidney. It actually accumulates quite a bit in the gut. And some people get diarrhea with metformin. And so some people think metformin is affecting your microbiome. And there's a huge literature now thinking that some of how it's having its effects. So I think the liver. So I think there's three places that are important the liver. And that can account for some of shutting down the glucose production and having the so called anti diabetic effect the colon and affecting your microbiome. And I think the immune cells, I think that's the big one we're missing. So that's the one that I'm very fascinated. In other words, metformin getting into your macrophages or your probably maybe not your T cells but at least some of your immune cells that might be causing high levels of inflammation. And you know, if you look at the three drugs that people like to use, aspirin, metformin and statins globally use combined, maybe what a billion people.

Probably more, probably more.

The Venn diagram where they all overlap is inflammation.

So let's talk a little bit about the anti inflammatory properties of metformin because the first thing is the one I guess we would understand the most, which is you've alluded to this, but I just, I want to orient the listener a little bit. The mitochondria, as you said, have these five complexes. Each of them have multiple subunits. And what happens is these are basically the chains between the inner mitochondrial membrane and the inner part of the mitochondria where these reducing agents like nad, nadh, NADP and NADPH are transferring the electrons and building up that gradient. So by the time you get to the end of this thing, you've got so much potential energy and all of those electrons and then you run that transfer of phosphates from ADP to ATP and everybody wins the game. So this is essential, this electron transport chain, like messing with that, probably not a good idea. Metformin comes along and it blocks complex I. Now it doesn't block it completely, it blocks it partially. Now complex I, the chemical reaction that's occurring on the inner part of the mitochondrial, inner part of the mitochondrial is the transfer of NADH to nad. Now we're going to come back to nad, but for totally unrelated reasons. When you do that, what is that telling the cell by inhibiting that the cell's readout is what physiologically three things.

First, that battery that you're talking about that generates to make ATP less charge, less charge. So then what happens? ATP goes down, ADP goes up. There's a kinase called AMPK kinase. AMPK is the AMP kinase. Hence ampk, it gets activated. And in part that that enzyme is.

Activated is a signal that says I'm not fed enough.

Right, Right. And one of the major things it does is it promotes autophagy.

My favorite word.

Your favorite word, Right. It's one of the dominant things it does.

That's why when AMPK gets activated, we get another little benefit, a la rapamycin's effect on mtor, which is it says, hey, man, I'm telling you, from a glycolytic standpoint, or from an OX phos standpoint, energy is low, shut things down, right? Nutrients are scarce, right?

And this is happening in the liver. So, for example, glucose production that happens in the liver starts to shut down in part or lipogenesis, making new lipids in the liver. And that's why, like, for fatty liver, it might have some benefits. So that's one. The other thing is what you alluded to. NAD to nadh. Sorry, nad.

Nad, Right.

To making nad. So that ratio also gets transmitted back to the cell.

And what's that signal? How does the low NAD to NADH get transmitted?

So the biggest one is. Is lactate to pyruvate, right? So lactate to pyruvate is a source of. So there's many ways you can feed pyruvate. So we talked about glucose to pyruvate, right? So glucose to pyruvate uses NAD to nadh, and usually pyruvate to lactate will go.

Go ahead and explain what I'm doing.

So I have to tell you, and I think we're both. Because we're not on camera, one of the difficulties of really getting into the nitty gritty of metabolism is it's so much easier to write it in diagrams, right? When you write it in the diagram in simplistic ways, it's just like.

It's easier.

It's just easier, right?

I mean, it's just so Nav's laughing at me because I'm closing my eyes.

Drawing it, and as I'm saying it, you know, nad. No, it's the nadh. I mean, I'm getting confused.

I promise you this. This will be one of those episodes where the. The show notes will be handy because we'll have all the diagrams.

Well, they can actually. You know what they can really do, right?

There's a book, I've heard.

Yeah, there's a book. You want to give a plug. This is a good moment.

I'm happy to give a plug. So for all you metabolite lovers out there, Nav wrote a book called Navigating Metabolism. Wa wa wa. And I actually picked up a copy as soon as we got back from Easter Island. In fact, I probably ordered it from the airport in Santiago. And it's a fantastic resource, and we will absolutely be sure. To link to that. I would say it is. And I'm not just saying this because you're sitting here, but if you are a person who's interested in this area, but you're not going to devote your life to it, it's a fantastic. It's the one book you need to get. Obviously, if you're someone who's doing a postdoc in Nav's lab, it's something you need to read, but it's not going to be sufficient to get you to the next level of understanding. But for the knuckle draggers amongst us, you can get pretty far on understanding this stuff through Nav's book. And it's actually a pretty quick read. It's not like reading Stryer's Biochemistry, where.

That'S a very good book, by the way. Stryer was my professor, as is the Leninger book.

Yes. So anyway, yeah, little digression, which is to acknowledge this is hard, apologies for it, but I think this topic is so important and I just know I get asked about this stuff all the time. I'm on a personal level, professional level, so interested in this topic that you just have to pay the price. Like, you have to be willing to get into the details. And the reason is we're going to talk about other things. You know, people are, if I get. If I had a dollar for every time I've been asked, should I be taking, you know, nr, nmn and should I be going to a clinic where they do IV nad and all these things? If you want to be able to think through those things and read the papers that are asking those questions, you have to understand how this stuff works. There's no shortcut.

Right.

So unfortunately, we have to continue doing this the way we're doing it. And that might mean that I have to close my eyes and pretend I'm drawing complex one.

Right. So I guess we should just talk about NAD then. Right? NAD and nadh.

I sort of interrupted you, though. Let's go back to metformin.

Right. So what metformin doesn't allow is it starts to weakly inhibit complex one. So your NADH to NAD is going to be slowed down and how that gets transmitted to the rest of the cell is quite. It's not fully understood. So I have many ideas around this and we can talk about one of them later because it has to do with neurodegeneration potentially. But the big thing is that NADH to NAD ratio is very important. And one of the important things is that when Lactate, which can come from the muscle, the liver takes it up and the lactate becomes pyruvate and that can then eventually become glucose. As gluconeogenesis says it needs nad. But if you have metformin, you don't have as much nad. So lactate to pyruvate slows down and therefore you don't make as much glucose. And that's another reason why metformin has its anti diabetic properties.

That just gave me an interesting idea. We have a friend in common, Josh Rabinowitz at Princeton, who's a classmate of mine in medical school, a colleague, brilliant metabolism scientist, off the charts, incredible guy. He had a paper that came out, I've talked about it on the podcast, very briefly. I actually want to interview Josh and I just have to drag my ass down to Princeton or he has to drag his up to New York City. But this paper in cell metabolism, which we'll be sure to link to, took orally administered NR or nmn, both precursors to nad. And it showed that the liver could take those up in significant quantities, combine them with tryptophan, and make lots of intrahepatic nad. But none of it made it into the cell. So the NAD wasn't making into the cell, and the NR and the NMN were not being taken up by cells other than hepatocytes. But what you just said made me think of something. If the liver in the presence of NR and NMN is making a lot of nad, that means it's making lots of substrate to enzymatically force gluconeogenesis. Or is that never rate limited and this becomes irrelevant?

The latter, what you just said.

So we're never too low on nad?

No, no, we are. We are the maximal amounts. And so there's two things about nad. One is just the quantity of NAD which then is utilized by sirtuins, parp, a variety of other reactions that are important biologically, especially the sirtuins, which are NAD dependent. But that's just simply nad. What I'm talking about is the ratio of NAD to nadh. And so these supplements, I don't think they don't drastically change that redox ratio of NAD to nadh. It's just the absolute amount of NAD which is then utilized is by sirtuins and parp.

And there's something to be said about this because people talk about how NAD ratios decline in the mitochondria as we age. Does that.

Is that rate limiting?

Yeah, exactly. Does it matter if it affects the enzymatic chain at complex one comes back.

To is a 50% decline in NAD rate limiting for complex one activity.

Because that would mirror what metformin is doing. Metformin is lowering the ratio of NAD to nadh, which would seem to parallel what we're told happens when we age. Yeah, that's a bit counterintuitive.

Exactly. Bingo. So this is my argument. Most of the people say you got to boost your mitochondria because NAD is declining, the respiratory chain is declining, mitochondrial rate limiting. But how does that jive with this metformin inhibiting complex one theory then? If you think that mitochondria and NAD and everything around mitochondria is declining, you want to boost them if you think they're declining. And maybe it's adaptive, that's why it is declining, which is my favorite theory.

Wow, that's a little out there.

That's way out there that this is. There's a reason. And if you can then give something like. And it's never rate limiting, really, at least for normal physiology, not maybe for stress. And then if you can give something like metformin, you can now stress out that mitochondria at times and turn on some adaptive responses. So that's a different theory. Right. So the best evidence for it really is we have to really nail down whether all of these effects of metformin happen by complex 1 inhibition. There's wide disagreement in the literature. So just to be clear, all these roles of metformin for cancer, for diabetes, for inflammation, does it require complex 1 inhibition? So how would you test that?

Well, it's hard to have a complex one knockout.

Exactly.

Because that's incompatible with life. So that would be the obvious answer, that delta.

Well, so I would argue for any drug, the best experiment is to make a mutant of that particular complex that doesn't bind metformin. Bingo. Yeah. So I'm not a good structural biologist, but we did something really clever. We noticed that the yeast has a protein, single protein, which will catalyze NADH to nad.

Okay.

What complex one does, in part.

Yes, exactly.

It doesn't proton pump, which means it doesn't contribute to ATP generation. But we have engineered cells and mice to get rid of complex I and put back this yeast, complex 1, which is refractory to metformin, but asked this.

Question, but the phenotype of that cell is what is its electron transport chain doing if it's basically losing anything at complex one? Basically, complex two through five still can't work.

No, no, no. So what this protein, this yeast Complex one homologe. Oh.

It does everything except the electron transport transportation. Sorry, it doesn't do the. The proton pump. Yeah, okay. Got it. So you basically reduced your battery charge a little bit, but you haven't interrupted the electron transport.

Right.

Oh, that's elegant.

Thank you.

That's very elegant.

It's clever. Yeah, yeah.

This is about the geekiest moment right now. This is like. This is.

Yeah, yeah. It's one of those aha things.

Right.

It's clever, right.

Very clever.

Very clever. Right. So you put. So the first experiment we did. Did is we did the cancer experiment. So people had noticed, at least in laboratory settings, if you give metformin, you can reduce tumor burden in mice. You know, the classic sort of experiments. So what we did is we took those cancer cells and put back the yeast version.

But I want to say something here before you say. You tell us what happened, which is in fairness.

Yeah.

And we did a lit review of this in 2014. So it's very dated. I know what's going to happen. A bunch of people are going to say, peter, can you please link to it? It's an internal document. I may link to it. I got to go back and look and see how ridiculous it is, but. Because it's now four years old. But it was not clear at the time of this review if the anti cancer benefits, which seemed real, were either due to the inhibition of Complex 1 or due to some other mechanism by which AMPK was activated. And to my knowledge, that is still not clear.

Oh, it's clear.

You're going to tell me it's clear for cancer. Okay, keep going.

So I'll tell you the experiment in cancers and then I'll tell you. For diabetes, there's main effect anti diabetes. I think that's still up in the air.

Okay.

For inflammation, I think there's some strong evidence for complex one as well. So the simple experiment we did was we said, so let's put back in cancer cells that human cancer cells. We put it in a mouse and it grows rapidly. Right. And you can do it in colon cancer, breast cancer, lung cancer cells. Typical cancer biology experiments. And we put the yeast complex one in. Okay. So when the yeast complex one is not there, metformin decreases tumor burden. If the yeast complex one is there. It does. Metformin can't bind to it. So the mitochondria still continues to work, and voila, the tumors don't go down.

So what you demonstrated through that experiment, assuming we're not being fooled by some other artifact which is Always possible is that when you prevent metformin from this one particular issue, which is binding to complex one and inhibiting that its anti cancer properties cease to exist.

Yeah.

Did you assess the effect in that setting on ampk? How much was AMPK activity upregulated?

Yeah. So we don't think the anti cancer effects are due to ampk.

Fair. But do you have an answer to the question, do you know what happened in that setting?

Yes. So we.

Because they should go down a little bit but not off.

Yeah, yeah. So we didn't think about that. We looked at other properties like ned, nedh ratio, which we think is the more important.

And how much did the nad?

Oh, yeah, no. So we could show clearly you could shut it down. Yeah, yeah. With metformin you could decrease it. When the yeast complex, you can recover that ratio, the NAD to NADH ratio and all the metabolomics that go with it. So when you inhibit complex 1 by metformin, the TCA cycle slows down and you can capture that by mass spec. Classic, what's called metabolomics, which is basically looking at metabolite profiling. And what's cool about that is there's a very good scientist. Give him a shout out. Jason Locasal at Duke University, again, kind of a younger version of Josh Rabinowitz. And what Jason hooked up with the University of Chicago ovarian cancer, very famous ovarian cancer, Dr. Ernst L' Engle and his fellow Iris Romero at that point. And they were giving metformin to patients and then they gave these biopsies to Jason and Jason could detect TCA cycle. He could see if he could ask two questions. Did the metformin get actually into the tumor?

Yes.

Yes. And the second one was if our mechanism of complex I that we showed is correct, then the TCA cycle metabolites should be altered and they warrant those human cancer. This is cell metabolism paper. People can link in.

What year was that?

So we published our metformin paper in 014. I think Jason's paper was in O16. I think we did the simple, elegant experiment which shows the necessity of complex one inhibition. But I think Jason did the. As close as you can, if that mechanism is correct in the humans, he did the best, the next best thing you could do, which is show that TCs cycle metabolism.

So clinically it begs a question, is that working? Because hepatic glucose output is going down and insulin is going down and presumably IGF is going down. If insulin is going down, IGF BP3 should go up. Insulin should go down even if there's no change in the amino acids. Those things should all. If you had a little like on off switch, more glucose or less, less is better. More insulin or less, less is better. More igf, more or less. For cancer, for cancer, all of those things would move in the right direction if hepatic glucose output went down. Down. So do we believe that that is the vehicle through which that transduction is becoming clinically relevant?

Right.

Or do we believe that Somehow inhibiting complex 1 in a cancer cell is deleterious to a cancer cell?

I think both mechanisms are working in concert. And so clearly metformin, as you pointed out, lowers glucose, insulin, igf, and insulin and IGF in certain tumors can be a mitogen or something that promotes cancer proliferation.

Right. It seems that about 2/3 of cancers seem sensitive to insulin and IGF.

So that mechanism is still in play. But what we showed is it's equally plausible for cancers that have transporters of metformin. And they're called organic cation transporters, not every tumor has it. And that's why metformin doesn't work clinically as a great anti cancer agent, because lots of tumors just don't have them. But if they do have them, they'll take them up, they'll inhibit complex I and that will have anti cancer effects right into the cancer cell. The reason that's important again is our work genetically has shown when we knock out complex one or three, tumors don't grow. If we give metformin, we can show its anti cancer effects are due to complex one inhibition. So if that's right, then could we design Complex 1 inhibitors?

Wait, let me ask you a question. Sorry to interrupt. Nav. When you and what you just said a moment ago. When you inhibit complex three and tumors don't grow, that's you have to inhibit complex 3 in a tumor cell or in a hepatocyte in a tumor cell and the hepatocytes are normal.

Well, the way we, these are genetically engineered where the complex 3 or 1 is only lost in the cancer cell.

So this is the next experiment you have to do. You have to be able to separate. Listen to me telling you the experiment you have to do. It's important, I think, to separate out how much of this is tumor specific versus global metabolic. And the reason is the implications are profound, not just for other therapeutics, but frankly for a more fundamental question was what the hell should people be eating right? If inasmuch as you believe that nutrition can impact cancer therapy, the answer to this question is relevant.

So we have now generated an unpublished work. I don't know. Can you talk about unpublished work on a podcast? I don't know.

It depends. When's it going to be published?

Not for a while, but it doesn't matter. So we've generated a mouse that contains the yeast complex one in the liver.

Ah, okay. So that this is. This is where you'll be able to do the experiment. Because there's really a two by two that needs to be done here.

I think they're both working in concert. But the major thing isn't that metformin may have some anti cancer effects. But what it's led to, because of our work and others, is the idea that maybe we should target mitochondria inhibition for cancer therapy.

Now that's a little counterintuitive.

Yeah. So this is again, I mean, I'm sure the audience is like, everything this guy says is contrarian. So let's just turn him off now. But you know, our data is very clear. Mitochondria are necessary. Mitochondrial function is necessary for tumor genesis.

All right, so let's take a step back and explain to the listener who hasn't heard what you just said, why that is gonna rock some people's world.

I think you've talked about in some other podcasts. So there was a observation made in the 1920s by a gentleman named Otto Warburg, one of the giants actually trained. Hans Krebs, for example, he won a Nobel Prize in 1931 or 32, basically for discovering an enzyme for respiration. And so he loved measuring respiration, but he did it in cancer. He did it in normal cells. And he noticed that cancer cells, at least on the benchtop, not in vivo, not in a real tumor, not in humans just taking tissues out, that they made a lot of lactate and they didn't consume as much oxygen. And this didn't make sense to him because he's like, there's plenty of oxygen, why should it do that? And it led him to think about perhaps that maybe the mitochondria being suppressed in cancer. And this led to this long, long, long dogma that a very elegant, simple theory. Normal cells use a lot of mitochondria and mitochondrial ATP. Very little glycolysis. So in other words, very little glucose to lactate, tons of oxygen. Your heart does it, your brain does it, but when you become cancerous, the mitochondria sort of shuts off and you upregulate tons of lactate. And you can see that huge uptake of glucose uptake by FDG PET that the clinicians do. And you can see lots of lactate. And that was this theory. And then you target glycolysis for cancer because it'll specifically hit cancer cells because they're so glycolytic and spared all the other cells like your heart, your brain, your liver, because they're all mitochondrial dependent for energy. Keep it simple.

Right, Right.

So far so good.

Absolutely. So, and then, I mean, just to add to that story about 10 years ago, and I've talked about this paper on the podcast, but Thompson and Mel, Matt Vadahan was the lead author. But so.

So Vander Heyden saw him last night in Madison.

He was here last night.

He was in Madison.

Oh, oh, you were not sure. So Matthew and Lou Cantley and Craig Thompson wrote this paper in science almost exactly 10 years ago, and it proposed a different reason for the observation. So it said the observation that Warburg put forth is what you just described, but maybe a different reason is that the cancer cell is not optimizing for energetics because that was always viewed as an energetically very inefficient and wasteful thing to do. But the argument they put forth was, well, it's not doing it for energetic reasons, it's doing it for growth reasons. It's doing that to get the throughput of building blocks for cells. So same observation, different explanation.

The one thing I will say is, for whatever reason, and including that beautiful review in science, which the part that people don't highlight is what is the mitochondria really doing? So people sometimes assume, oh, yeah, all that glycolysis and it's for biomass and building blocks. Oh, yeah. The mitochondria has a negligible contribution. It's just sort of in the background. It's the potato, as we call it.

In the Sabatini world.

Yeah, yeah, right. It's just, it's a bystander. And we did a simple genetic experiment, said, let's just test this. So we're going to take in a mouse with an intact immune system. We gave the mouse, made it have poor mouse got lung adenocarcinoma, Lung cancer. Right. The biggest cancer, the most prevalent cancer in the world, obviously, due to smoking. And we genetically knocked out the respiratory chain so it can't respire. So now it's 100% glycolysis. It's exactly what a tumor that Warburg would love. Do you make bigger tumors? According to him? Yes, if it's all about glycolysis. Or do you make smaller tumors. We made very smaller tumors. Little, little, tiny little tumors, which told us that mitochondria are necessary. Our mitochondrial respiration is necessary for tumor genesis.

Van der Heyden's hypothesis would explain that. That because if you knock out the mitochondria, you don't have the biomass through.

But he didn't say that.

No, no, no, I know, I know, but I'm extrapolating.

Yeah, it's consistent with that. Yeah. No, Matt and I completely agree. It's just the way that review was written 10 years ago. It was more glycolytic centric. So I will give a cheap plug to a review that I wrote with Ralph DeBradines.

It's a very elegant review, by the.

Way, 2016, where we updated this. It's called Fundamentals of Cancer Metabolism. And it's really simple. It just says if you go back to your biochemistry books and you ask, how do you make a nucleotide? Let's keep that simple. You need to make new DNA because cancers proliferate. You make one to two daughter cells to four, and there you go. So where does that nucleotide. Let's look at the structure. It has a ribose as a backbone that comes from glucose. Bingo. It needs a variety of nitrogen atoms put on it. Where does that all come from? Some of it comes from like a aspartate and glutamine. Where do they all come from?

They can come from glucose and they're amino acids, but.

They're amino acids, but they'll come from mitochondria. Right. So in other words, they both are one of my favorite words. They're both necessary for tumor genesis, but neither is sufficient.

Sufficient. Can we just pause for a moment? Unnecessary but not sufficient. If there's one thing that I loved in medical school, when you were doing the basic science classes before you got into the clinic. Clinic. It's that when you were doing your physiology classes and your molecular biology and things like that. These professors, they were so great at explaining the importance of very elegant experiments that can demonstrate whether something is necessary but not sufficient. Sufficient but not necessary. Neither necessary nor sufficient. All these other things. Something happens in medical school when you leave the classroom and you start to go into clinical medicine, people start to forget that logic.

I would say the real logic. People forget. Everybody does. We're all guilty of this. And this is what you and I have talked extensively about. Correlation versus causality. So, you know, my daughter can bitch all she wants about how her papa hasn't taught her any math, but her papa has Taught her one thing, correlation versus causality.

But this goes even deeper than that. Right?

You gotta start there. Oh, yeah, fundamentally. But I think people. I mean, this is. And right now, we even in my world, where people see like mitochondrial function go up or down. So, like, take your aging one.

Yes.

Mitochondrial function goes down. So that's a correlation with aging. That doesn't mean that decrease in mitochondrial function causes aging or drives aging. It's just a correlation. It could be adaptive or maladaptive.

That's right. I mean, that's. So, see, to me, the second order point is the right point. The obvious point is, is you can't infer causality. But the second order point to that is if there is causality, it doesn't tell you if it's adaptive or maladaptive. That's the nuance. And this idea of necessary but not sufficient to me, is very important in biology because you can have things that are causal, necessary, but not sufficient.

I just told you one.

Exactly. Causal, sufficient, but not necessary. And most of all, causal, neither necessary nor sufficient. Patient. And people love to dismiss those things. I'll give you an example. Smoking and lung cancer.

There's never been a trial.

I'm going to argue smoking is causal with lung cancer, just as I'll argue that smoking is causal with cardiovascular disease through different mechanisms. Endothelial dysfunction in the latter. But no one in their right mind would say that smoking is either necessary nor sufficient. Lots of smokers don't get lung cancer. Lots of smokers don't get heart disease. Lots of people who get heart disease and lung cancer don't smoke. So isn't it interesting that you can have something that is neither necessary nor sufficient and yet can play a causal role in a disease? And again, people might be listening, going, what the hell is he making such a big deal out of this for? I make a big deal out of this for. Because when you get to complex diseases like cancer and Alzheimer's disease and atherosclerosis, it is very unlikely you will find something that is necessary and sufficient. They're very rare to find those exceptions. These are such multifaceted diseases. And there are so many different ways to skin a cat. You know, when you talk about cardiovascular disease, you've got like four completely different things that have to be going on to cause this disease. You have to have lipoproteins trafficking the sterols into the subendothelial space. You have to have the endothelial dysfunction to enable that to get in there and get retained. You have to have the inflammatory response. You take one of those things away, you change the dynamic of the disease. And your other point's a great one. Even if you can infer causality, it's not entirely clear what's adaptive and what's maladaptive.

So our experiments really, which showed there's mitochondria's necessary tumor genesis. So it's interesting, that paper you referred to. Vanderheim and I were graduate students together and postdoc. When I was starting my postdoc, he was sort of finishing his graduate training with Craig Thompson, and we were in the same lab and we actually worked together. So he's an old friend of mine. And obviously Craig's a former mentor and I like Luke Handley a lot. But that paper, when it came out in 2009, right around at that point when it came out in Science, we had sent our paper to Science and showing that mitochondria necessary tumor genesis. They didn't send it out. They said, oh, come on, this is.

No.

So then we sent it to 11 other journals. No editor sent it out. Cancer Cells, Cell, Nature, Nature Medicine, you know, Nature Cell Biology, Journal of Clinical Medicine. Nobody sent it out because the review didn't say mitochondria are not functional. It just didn't mention anything about mitochondria. It sort of just provided an explanation of the glucose, like glucose to lactate. Why do tumors show that? And it's for the biomass, not the energy. Right. That was the point. And it didn't say it was sort of agnostic about mitochondria. Right. In the review, if you go back. But people misinterpret that and saying, oh, yeah, mitochondria are not necessary. It's all about glycolysis. Let's target glycolysis. So eventually, the paper did get published in pnas, and I think that sort of of started, I would argue, the revolution of looking at mitochondria and cancer. Other people were doing it as well, but I sort of became a preacher, just pointing it out. Look, if you inhibit the respiratory chain, you can decrease tumor genesis 10 years later, to the best of my knowledge. And this is really important because it goes back to the clinic. As far as I know, there are no clinical drugs in the clinic that are necessarily targeting glycolysis. Right now, there are potentially two drugs that target the mitochondria.

What do they target specifically?

So one of them is a complex one inhibitor that the M folks at MD Anderson have generated, and they just published two papers in Nature Medicine, which.

Is different from fenformin?

They're fenformin. Yeah, they're different from metformin fenformin. They're much stronger. They know the binding side by the way, they use the same yeast, NDI one complex one that doesn't bind bind to metformin, but doesn't bind to their drug as well to show test it.

So how will they prevent toxicity?

Yes, yes. So now they're doing a trial in aml. And so this is the million dollar question, and can you find a therapeutic window where maybe the drug gets taken up preferentially by just because of the properties by leukemic cells or prostate or lung cancer cells, but spares the brain, the heart and other organs where you could have toxicity? So that's the big issue. The good news is if you inhibit complex one, it will decrease tumor genesis. The bad news is it might kill you. Right. So we gotta find that window. And does it also work well with immunotherapy? That's an open question. Right. Which is the new kid on the block. Like does it help synergize with immunotherapy or does it prevent the immune function? So all of this has to be worked out. But my point is that there's some space in this area. The other one is a drug by Rafael Pharmaceuticals. And full disclosure, I sit on their scientific advisory board.

Thank you.

And I'm not pitching anything here simply. In fact, they were already doing this stuff. We're just sort of giving them some, you know, my little biological insight, which is I'm trying to provide today. But they've already done a clinical trial and it was, and I'll send it to you, it was published in Lansand Oncology and it was in pancreatic cancer. Just a safety trial.

Phase one.

Phase one. And it targets alpha keto TCA cycle enzymes. And so it's preventing the TCA cycle from functioning to build that biomass.

And did they generate any data on the tissue specificity of that agent?

Not yet. So this is the kind of stuff that needs to be done to try that drug and do the kind of experiments where you can see TCA cycle metabolites changing pre and post drug treatment of a particular tumor and then and correlating that with success of remission and you know, all the usual parameters. Now what's interesting about that drug again is why would that be safe, any of this? The only way these drugs can be safe is they somehow preferentially are getting.

More into the tumors or they're just so weak that they're not bringing you below the threshold of shutting off the tca.

Yeah. And then maybe they combine. Well. So I can tell you, if you give the standard care of therapy, cisplatin, one of the chemotherapies or targeted therapies like BRAF inhibitors, is what happens. As, you know, the primary tumor sort of debulks, right? It slows down, and slowly you get resistance, and then it comes back during that slow resistance phase. At that point, they're really dependent on mitochondrial function, so there might be a window to attack it with, like, BRAF inhibitors or cisplatin. And again, like everything else, they're gonna have to find that sweet spot. That's.

See, to me, that's where I think this has to go. I mean, I've always found immunotherapy to be the most elegant of all approaches to cancer. I'm highly biased because that's what I studied. But ultimately, I think the Swiss cheese approach has to be the approach too, which is, why would we only take one modality of therapy? If you're David and you're trying to slay Goliath in the fairy tale, one stone to the head does it. But in the real world, to take a Goliath down, I think you need to go to bang him at the knees, and when he bends over and complains about it, bang him in the other knee, and when he's complaining about that, whack him in the hamstring. In other words, you've got to be able to do successive blows to a vulnerable cell. And that's going to be when is it most dependent on the mitochondria? Okay, well, bang. Now, you hit with that therapy, and then all of a sudden, to your point earlier, maybe at some point you begin to weaken it even you make it more identifiable from an immune perspective, and bang, that's when you would hit with the immunotherapy.

The last point I really like, you know, would the immune system recognize that tumor better if the mitochondria of that tumor was not working properly?

Yeah, see, that's.

I mean, I maybe release that mitochondrial DNA for example, as it's dying.

As long as it were specific. Right? As long as it could. It didn't create a diffuse immune reaction, but instead a localized. Allowed the body to say, that's not itself.

Right.

That's. You know, and I haven't. I haven't done a podcast yet on immunotherapy. I want to have Steve Rosenberg on to talk about this because, I mean, who better to talk about this with. But anyway, so let's talk about another issue in cancer, right? Which is, I mean, I think everybody agrees that most cancers involve somatic mutations. You know, there are very few cancers that involve germline mutations. Those are the exceptions. But the general one is these are acquired mutations. Now, some have argued, and this is a very minority opinion, but some have argued that the somatic mutations of the nuclear genome are actually the result of the mitochondrial dysfunction. I think the majority would argue, no, it's the other way around, that the nuclear genomic mutations, somatic, are actually what leads to ultimately whatever's happening in the mitochondria that may be dysfunctional, may be maladaptive, may be adaptive. You would be in the latter camp, correct?

Completely, 100%. And again, this is why these clinical trials are really important. This is why the metformin trial is important, right? So I've told you two things. I've told you three points that are quite contrarian. We started with antioxidants that there's no evidence that antioxidants in large scale, the dietary antioxidants, just to be clear, vitamin E, vitamin C, have had any benefit to mankind and womankind, right? For human health and disease. It just hasn't worked out. So either we haven't built the right antioxidants or the theory that Ross and oxidants are bad. That theory is off. And I would argue that theory is off because if anything, normally we use oxidants as signaling molecules. The second thing I told you about is the fact that during aging, yeah, mitochondria decline, but that correlates and it could be adaptive. Very contrarian point of view. If anything, if you gave an agent that decreases mitochondrial function like metformin, that that could be a good anti aging therapy, right? It's not turning on mitochondria, but turning off mitochondria. And by the way, Andy Dillon, a good friend of mine I was there with yesterday who's done beautiful work on worms. Clearly in the worm when you decrease complex one or three, you live longer as a worm.

Yeah, I don't know Andy, but David has spoken so highly of him. David Sabatini, you have as well. I need to meet Andy and hopefully interview him at some point and talk about all this stuff. Of course, it could be going back to your point that the inhibition of complex 1 which inhibits mitochondrial function inside of a non toxic range might be, might not actually be part of why metformin makes you live longer. That might just be a. It survives despite that not because of that, the organism.

Yeah. I would argue that, you know, metformin being anti inflammatory, and I want to.

Come back to anti inflammatory.

We'll come back to that. But so the third point is that glycolysis is necessary, but so is mitochondria. That cancer cells use a robust mitochondrial function. And if that function doesn't work, the TCA cycle or the respiratory chain doesn't work. In most cancers, you don't get tumors. Now, there are these rare cancers where it has a TCA cycle mutation. And so this is another sort of logic point. Right. So they look at that rare phenomenon, the exception to the rule. And some people say, oh, there are these rare cancers that have a TCA cycle mutation. Aha. Therefore, cancers have TCA cycle mutations. Oh, come on. This is illogical. It's the exception to the rule. Most cancers, at least that we've studied, whether it's in cell culture, in mouse models, and my good friend Ralph Debradines, he's doing tracing experiments like Joshua Binowitz has done as well. They can clearly show the TCS cycle is quite robust. And so you know, why those cancers can arise is an interesting question. But they're the exception to the rule. So I'm in that camp.

Well, and the other, I guess.

And if I'm right, by the way, then these drugs, if they make it in the clinic and really make a difference, voila.

The other thing that I think would favor the genomic argument is a lot of the viral research, because when we see viral vectors driving cancer, it. It's presumably nuclear DNA. I'm not aware. Are there viruses that are causing cancer through infecting mitochondrial DNA?

I don't believe that. I think it's all nuclear, personally.

So that would be another.

Yeah. If anything, you know. So renal cell carcinomas are really. So you know these oncocytomas.

Yep.

You know what they are benign tumors, essentially, if you look at. In renal cancer, and just for the.

Listener to understand, meaning they still grow in a somewhat unregulated way, but they don't have metastatic potential. So you're not going to die from these things. But they're sort of ben growing tumors.

Yeah, Even those. The major mutation that they have is in the respiratory chain. So in other words, again, they're very rare, but sometimes you do get complex one loss, but you don't get an aggressive tumor. You get this benign tumor at best. Right. So again, it's almost a barrier to progression. And that's from human genetic data. So I'm in the camp. But, you know, ultimately we can do all these cute little mouse experiments and the data is very clear in our hands and Ralph's hand and Josh's hands and lots of people's hands that mitochondria necessary for tumor genesis, TCA cycle activity. The ultimate proof is, well, let's inhibit the TCA cycle for prostate cancer, for colon cancer, for pancreatic, and does it make a difference with immunotherapy or chemotherapy? So we'll see. And by the way, the same one goes to metformin and which is that if metformin really does work as a, potentially as an anti aging strategy, and we can show that those effects are due to complex one inhibition, then it's hard to think why that, you know, that this idea that mitochondria rate limiting or declining to a point during aging, that's injurious, that idea can't hold up because you're giving essentially a weak complex 1 inhibitor to turn on stress responses, which means you must have enough mitochondrial activity as you age. Age is down, but not gone, right?

Right.

Right enough to not be rate limiting. And of course, the antioxidant one, we've already won that battle, right, because the trials have all failed.

And for the person listening to this, who's scratching their head and confused and says, does this mean I shouldn't be taking my vitamin C or my vitamin E or my whole foods proprietary antioxidant blend of blueberry skin, I think you're right. The answer is pretty clear that, that the harm of taking those things might not be great, but the benefit seems to be negligible to nowhere.

That's a fair statement.

Are there benefits, you think, to the natural quantities of antioxidants we consume in our food? So, for example, berries do contain lots of antioxidants. People love to talk about those benefits without saying, go up and take, you know, ground berry capsules or something like that. Do you still think there is a benefit in having. In other words, if a berry doesn't give you benefit in the antioxidant, there's not a hell of a lot of benefit in it because it's basically just a vehicle to deliver fructose, which I could argue you don't need any fructose in your life and it's a vehicle to deliver glucose, but you can get glucose in better forms or more of it elsewhere. So is there some other benefit?

So I don't know much about berries. So the best one is vitamin C, right? Like, how much should you take okay.

So let's talk about an orange. Is there some benefit in eating an orange?

Yes.

Okay, so what's the benefit in eating an orange from an antioxidant standpoint?

Yes. So there are enzymes that control DNA methylation and other reactions. So simply that to maintain proper function of gene expression, so your genes turn on and off properly, there are enzymes that are dependent on vitamin C. And you basically need about an orange a day or a glass of orange juice, you know, Please, please, please, not orange juice.

Please not encourage people.

Not orange juice. Fine, fine, fine. But, but, you know, but. But what I mean is you don't need to take 10 oranges, right. An orange a day is enough, right. To get you. Give you enough vitamin C to make those reactions work properly. And one or two oranges, but you don't need to then go and take 400.

That has nothing to do with the antioxidant properties of.

No, no, no, no, no. Nothing to do with Ross. And all of this stuff we talked about. It has to do with running some enzymes that are important for controlling gene expression. In other words, genes that have to get turned on and off are dependent on those. And of course, there are some people who might not be getting enough vitamin C due to their diet. So that's fine.

Pretty hard to do it.

Pretty hard to do it. I mean, citric acid, we have it a lot of places.

Would you be comfortable speculating that a cancer patient in particular should avoid antioxidants?

No, no, I don't think. I don't think the dietary.

Well, sorry, sorry, let me rephrase my question. I don't mean through dietary means, but through supplemental means. Like, if any patient could actually be harmed by an antioxidant, could it be a cancer patient?

Well, one of the trials, the lung cancer trial, argued that, I think was a vitamin E trial, and they did worse. And in mice, you can recapitulate that. So why is that? It's not clear. So I personally don't take any of these supplements because I think I've got a reasonable diet just like you.

Now, why don't you take metformin?

I will not take anything unless it's done any drug, unless it's gone through a rigorous clinical trial. That's just my own bias. Now, some people would say, okay, okay, you're aging now, so you're gonna. By the time they do a trial, you may not be around, so why not take a chance, right? Lots of people argue this about NAD boosting pills, right? That you should just take NAD Boosting pill by the time people do a clinical trial and all of that will take years and years and years. And for those people who are already later in life, you know, go ahead and take metformin.

And so, so putting your bias aside, because inherent in your bias is an assumption which is the risk of taking it is greater than the benefit of not taking it. So either you don't think the benefit is that much or you think the risk is maybe greater than some do. And you're certainly somebody who's in a position to evaluate both. So tell me where it fails. Is it a not enough benefit or a too much risk problem?

Probably. I'm not convinced about the how much benefit for someone like me who exercises especially, right? I mean the best effect of metformin and it's still its anti diabetic effect. And you and I both know you lift weights, you run, you're active, you sort of mimic the effects of metformin in many ways, activates ampk, you get the muscle benefits. And so why should I take that? Now you could say, well, maybe as an anti cancer prevention agent, maybe the data as an anti cancer. We'll see where that pans out. There's a large scale trial that's going to come out.

Wait, you're not talking about tame, are you?

No, not tame is the anti aging trial. That won't be done for another five years.

Will that ever be done?

Yeah, I think so. I think so. But if you're healthy and you're active, it's hard to see why you would take it. Now of course you could argue that as you've aged, you've gotten some indication that things aren't working as well and therefore you should help take it as a complement whatever loss you've had. Maybe you're a little diabetic. Maybe. The one place I'm rethinking about metformin a lot is whether it's a mild anti inflammatory agent. So in other words, it sort of keeps inflammation down all the time. And whether the effect of that over 20, 30 years, if you know, you and I are sort of in our 40s and that would have a benefit.

20, 30 years later, what's the mechanism by which. I'm glad you brought that up because I forgot to revisit it.

And you know, one of my favorite trials is the Cantos trial where they basically targeted IL1 Beta Pro inflammatory agent directly. And it didn't change the lipid profile.

But it reduced cardiomy.

Exactly. Which tells you that inflammation is.

Yeah, and there's going to be another trial announced very soon that I think will show similar results using low dose and mesatrexate. Of course, I could be proved wrong, and maybe that's not. I don't know what the trial's gonna show, but that's the hypothesis. Let's take as a fact, just for the sake of time, that lowering inflammation has wonderful benefits. What's the mechanism by which metformin will reduce inflammation?

So we think that reactive oxygen species, the free radicals, can serve as signaling molecules to activate cytokines. And metformin by inhibiting the respiratory chain, which is a major site of those reactive oxygen species, decreases reactive oxygen species and decreases cytokine in production. And again, a little bit, right. It's a weak inhibitor of the respiratory chain. So if you knock out the respiratory chain completely, you can never turn on the cytokine.

You've got a bigger problem, you got.

A bigger problem, you get a bacterial infection, et cetera. So this is just again, dampening it enough that if you get an infection, you can still respond, but just keeping the set point where you're at a little bit lower and whether that has good effects over 30 years, 20 years of keeping inflammation down. And then, you know, there may be some benefit to that. So then one other place you might want to do it is if you live in Beijing or in Delhi, because pollution increases inflammation, that's well known. It increases IL6. Right. So in those sort of places, taking something that might decrease inflammation might, might be helpful.

And how robust are the data on the immune modulating or inflammatory modulating benefits of metformin? Is that relatively. It's not as strong as the other stuff we've talked about, is it?

No, no, I think, I think it's not that many. There's not that many papers on it. It's, you know, know to be determined. But it's something that we're thinking a lot about as a mechanism of why it might have anti aging properties.

Well, look, speaking of anti aging, let's go back to nad, because we sort of skirted around a little bit. Obviously, one of the most popular types of supplements being offered on the market today, and there are several of them are supplements that are aimed at delivering precursors to NAD production. So again, the logic here is it's generally well regarded that cells can't take up free nad. They have to make their own nad. Josh and Rabinowitz and colleagues actually published a paper in June of this year that demonstrated that the NAD must be made in the cytoplasm not in the mitochondria and it's actually transported from the cytoplasm to the mitochondria.

And these supplements mainly went to the liver first.

Well, even. Even before that. But just to explain the logic. The logic is you can make NAD from NR or nmn and NR and NMN exist in an equilibrium, if I'm.

Not mistaken, and the de novo pathway, which is from tryptophan.

That's right.

As well.

So by giving these precursors, the cells can take those up and presumably make NAD there and get more of that NAD into the mitochondria where presumably it. I think the main argument, if I'm not mistaken, is actually not around the. Etc. The alkaline transport chain, but more around having them as cofactors for the sirtuins. Because the sirtuins of course played these two roles of acetylating, deacetylating as gene regulators. They're basically turning on and off genes. So I think the thinking is, and again, I don't want to speak out of turn, this is not my area of expertise. But more NAD should be an important CO factor for sirtuins, which are a NAD dependent histone deacetylases. Hw, did I get that story mostly right? Okay.

I mean that's the simplest hypothesis. NAD levels decline in aging, you lose sirtuin activity declines, which is not good.

Right. Because you now lose the ability to control genes, to control gene expression either on or off.

So you boost the NAD as it's declining and you get a little bit increase in search and activity.

Right, right. Which so totally makes sense conceptually.

So I think what Josh did is a great experiment. He basically asked when you take these supplements, where do they go? And a lot of it goes to the liver. Eventually it makes its way into other tissues because there was this simple idea like it's going to get into the brain easily, it's going to go to your heart, it's going to go everywhere, and it's going to do exactly what you said and therefore have all these magical properties. I think the place where NAD supplements and metformin start to cross talk is two places. The first is it goes to the liver. Liver. So it might be having some metabolically healthy effects on the liver, like metformin and similar to like what metformin does potentially. It's a hypothesis, by the way. The second one is I think it gets into immune cells.

You think that nr. Let's just make it simple and talk about nr because that's the preparation that's more commercially available. You think that NR is being taken up by immune cells?

Potentially.

But wouldn't Josh's paper contradict that?

I don't remember if they looked at all the immune cells.

Well, I don't think they did. But isn't the takeaway from Josh's paper that the first pass effect is so significant that all of the NR was getting taken up by the liver?

But you know, it still circulates in the blood, Right? And your immune cells are in the blood. So I don't know. And by the way, in Josh's paper, he also said that the liver, once it makes a downstream products of nr, it distributes it back into circulation to the rest.

What did it deliver? Remind me, was it delivering nad? No, no, it was delivering what downstream product?

I think it was nam. Okay, Yeah, I don't remember, bro.

And can NAM be taken up by the other cells? And can NAM be worked? Can NAM work its way back?

I don't, I don't remember.

Put you on the spot with something that's not your world, but.

Well, no, it's not. You know, again, pathways talk to Josh. But. But I think that the more simpler point is, is what you. What I think Josh's paper is getting at and what you're getting at is really this stuff gets everywhere and has magical properties. Is. And he starts to argue, well, it goes on to the liver or only to certain tissues, is that. I'm just arguing that a tissue, if you can call it, is not really, but a compartment that we don't think about enough of, whether it's with metformin or nad. Supplements are the inflammatory cells.

So what would an inflammatory cell, how would it benefit from having more nad?

Well, I mean, again, for the same reasons.

Right?

The sirtuin.

The sirtuin reason.

Yeah, yeah. Or some other NAD dependent process that in an immune cell that might be important. Important. And by the way, there is an enzyme that gets rid of NAD. It's an NADase, basically CD38. It's most abundant on immune cells. Yeah, yeah. So there's an immune connection.

I didn't realize there was much CD38 off immune cells.

I mean, depends.

Yeah, yeah. No, I mean, I'm not disagreeing with you. I just didn't know.

Again, this is a little bit out of my wheelhouse, but I'm just speculating that there might be a connection between these supplements as basically working as mild anti inflammatory agents.

Now, of course, the other way that these things are typically delivered is through intravenous nad. Which says, hey, you don't need to make it. We'll give it to you. We know you can't take NAD orally, so you have to do it intravenously.

There. I think it went everywhere in that paper.

Right, but does it get into the cells? Is there a cell that can take up nad?

I don't think so, but again, I haven't.

I don't remember.

Josh, you should. This is this I ask for the next podcast. We have stayed away from NAD biology. It's been quite a contentious field. As you know, Josh and others are not doing really nice experiments. We've avoided nad. What we haven't avoided is NAD to NADH ratio. Right. Because that's linked to complex one function.

That's what matters.

An NAD to NADH ratio, to the best of my knowledge, isn't controlling sirtuins or we don't have great evidence for that. NAD itself might be. But what NAD and the nadh ratio is doing biologically or how does a cell process that ratio is right now, probably the thing that keeps me up at night the most. My favorite new theory of life as we know it, which is tied to that ratio.

Now, speaking of supplements, you alluded to one earlier. I alluded to it called mitoq. I'm getting a lot of questions from patients about this. Should I be taking this? Should I be taking this? Should I be taking this? Can you tell us what it is?

It's basically CoQ. It's CoQ 10. So people take lots of CoQ.

What does MitoQ differ from the regular CoQ 10?

It has a cation attached to it. And because mitochondria pumping those hydrogen ions, they're quite negative in charge, like a battery positive. And so it will take a molecule that is very positively charged.

So by putting a cation on it which is positively charged, you increase its affinity for the cell.

Is that it into the mitochondria?

Into the mitochondria.

Right. The problem is the therapeutic window is very tight on that because when we give mito Q, we can shut off a lot of Ross production and all those beneficial stuff. Stuff gone. So you put it on stem cell. Stem cells don't renew. You put, you know, you put on immune cells as they don't get activated. And so I think again, antioxidants get tough because they have normal biological roles.

And is CoQ considered an antioxidant? Coenzyme Q, the reduced form of it is.

Which is ubiquinol. Yeah.

Yes. And do you think so therefore.

But they're hard by the way. You know this super, super hydrophobic, right?

Yeah. They're most, most commercially available preparations. Don't even seem to have any bioavailability. They don't. You, you can take a ton of them and you can't measure it in the blood. But there are potent ones that make their way into the blood and you can measure those levels. I think the question is, is there benefit in that? I, I again your view is no.

My view would be no.

And your view is it could be harmful.

I think a lot of these antioxidants have poor availability.

So, so when you know in the Peter Attia 2x2 is on the X axis you think about harm and on the Y axis you think about benefit. So and I, but to simplify it, even though these are continuous variables, you, you go with two categories. So on the X axis which is harm, you think about picking something up in front of a tricycle versus picking something up in front of a train. Obviously one has more much dire, much more dire consequences. And then in the benefit it's picking up, you know, a bitcoin versus a quarter. And so do you view most of these antioxidants CoQ10M as picking up quarters in front of tricycles where the upside if they work is probably not that big, but the downside is also probably not that big?

Yeah.

And do you put metformin in that category or do you think metformin has more potential? Has more potential but you still don't take it?

I still don't take it.

So you think metformin is potentially picking up a bitcoin in front of a truck?

Maybe. Time will tell. Time will tell. I mean, you know, people are now doing clinical trials with metform for anti cancer. I mean, I mean clearly it's still one of the first line anti diabetic drugs and people are now running them through inflammation models. We've done some interesting work around pollution and metformin which I cannot comment on. So there's a lot of interest beyond the anti diabetic effects of metformin and we'll see how it plays out. From our point of view, we want to really nail down is it by inhibiting mitochondrial complex 1?

Yeah, that's super elegant stuff. So you got to go back and do the experiment we talked about with these.

We're making, you know, we've made mice and all of this. So we're doing that just to finish time is probably, we've probably gone over as always. But so in my world Those ROS from mitochondria are beneficial and you don't. You know, there's not. I'm not sure if there's a window where antioxidants get in to really scavenge them. So they're. I don't consider them harmful, Peter. Right. For that reason. So generally, Ross, are good. Again, very contrarian. But this is where the data is taking us. So when is metabolism bad? So my favorite new theory, which. This is what I'm really excited about, and I'm hoping somebody will give me lots and lots of money to test this, because it's way out there. So if you think back about what causes pathologies like neurodegeneration, even diabetes, the big idea for 20 years has been that proteins get misfolded or they aggregate. The idea of what's called proteotoxicity, let's clean up bad proteins. So now they've done some trials in Alzheimer's and Parkinson's. It hasn't quite worked out, but, you know, again, maybe they caught them too late.

Yeah, that's my argument. That's the.

Yeah, yeah. So I don't have a problem with the theory. I think it's a nice theory. I still think, you know, proteoxicity is a real phenomenon. It causes diseases, all that good stuff. But what if. Which is not a mutually exclusive idea. What if there's metabolite toxicity? What does that mean? That means that certain metabolites that are normally found and at low levels and they do normal functions, if they rise, they can incur pathology. So what's the evidence for that? Well, this is where inborn errors of metabolism come in. So, unfortunately, people have genetic mutations and metabolic genes, and those pathways get altered and some metabolite increases or decreases, and that causes major pathology. So we know that metabolites are at a certain threshold are sufficient to cause pathology based on inborn errors of metabolism. And so why couldn't it be that in Alzheimer's we have a particular metabolite or metabolites that increase due to the tau and all the amyloid plaques that people talk about, and those then are causing neurodegeneration?

Again, not mutually exclusive.

Not mutually exclusive. It's just a different way of thinking about it. So one way you test it is, you know, someone's got to give me money to do this. But you start, start screening metabolites in mice and rats and people and see if you see signatures. And if you see certain signatures and you say, so I have one right now that I'm Very interested in. It's called L2 Hydroxyglutarate, L2AG. And I know that if a human has a mutation in a pathway that can't get rid of L2AG and it starts to accumulate, they get neuropathology. So could L2AG be elevated in Parkinson's and in Alzheimer's? What's so cool about L2AG? And this is going to wrap everything up from the start, is if mitochondria are not working, are not functional, the respiratory chain is not working, then NAD goes down and NADH goes up and it will trigger L2HG. That's the key trigger. When NADH goes up and NAD goes down, L2AG gets made.

So is this why you don't take metformin?

No, because metformin is a weak.

So this ratio has to change a lot.

This ratio has to change a lot.

So, for example, are there physiologic conditions under which that ratio changes?

Hypoxia. Aha. And under hypoxia, it's been shown that L2Hg levels actually increase and they can then function as a signaling molecule. So here is more where the organelle mitochondria is completely dysfunctional. So complex one loss has been correlated with Parkinson's. Dopaminergic neurons not functioning. And so is L2AG elevated there?

So have you looked at, you know, the other patient population you should study this in would be patients who undergo circulatory arrest in cardiac bypass. So you're going to see or even frankly just bypass because they're so hypoxic. Yeah, yeah. It'd be interesting to see, you know, quote unquote, this idea of pump head. Can it be explained through any of this stuff?

Well, so one other thing, another thing that I like about this is that's an acute event, I think we don't see it acutely. So you inhibit complex one and the nadh goes up, nad goes down. A real severe inhibition. It takes a long time for that to accumulate.

How long?

In cell culture with complex one, not 24 hours. Couple of days. And, you know, so you could imagine. Let's just play fantasy here. You get loss of complex one slowly in a dopaminergic neuron, which causes results in Parkinson's. And then slowly, over years, you get this accumulation of this particular metabolite, and that could then cause the pathology. Again, I'm very interested in just testing the broad idea that metabolites can cause pathologies like metabolite toxicity, kind of like protexic. Say probably wrong, but at least it's original.

Well, I mean, it's just, I guess what the thing that would concern me is the ubiquity of the potential signaling molecules. And trying to identify like what the patterns are can. That's a. I mean, look, there are no problems worth solving that are easy, so. But boy way you have so many variables in so many directions. You don't just have the number of metabolomics, you have the time series in which they occur relative to an insult and then the amount of exposure of each that's necessary to drive the disease. It could be so easy to miss something with all of those variables. Right?

Absolutely. And that's why we're being very biased in going after this one.

Yeah.

Keep it simple. Test that one. But of course, we could totally miss it because there could be five other metabolites that might go up that might be causing the pathology in synchrony. Right. But at least this one is tied to mitochondrial function. So when mitochondria are really dysfunctional, so we started with the powerhouse. People would say, well, mitochondrial dysfunction. ATP goes down. Okay, I would say mitochondrial dysfunction. Guess what? L2AG goes up this two hydroxyglut. So.

Interesting.

So it's a new way to think about mito dysfunction that I'm very interested in pursuing.

So last thing I just wrote down when we were talking earlier, and I want to come back to it, is talk to me a little bit about cortisol and your views on it. How does cortisol interact with the mitochondria?

Yeah, well, I don't know. I see another one that I would love to work on. So it's just a weird observation. I don't know if you have this. I mean, I think maybe in our circles we have a lot of kind of type A personalities who exercise vigorously, they watch their diet, they do all this sort of stuff.

I don't know anybody that does that.

No, you don't know anybody?

No one.

Nobody. But I always wonder if they're stressing themselves out by being so careful about everything, you know? And you know me, I am the opposite. You've seen me eat and drink.

Yeah, but you're pretty fastidious with your exercise. You play soccer every day.

I do. And I don't over overeat and I do time feedings. Right. I still fast 12 to 15 hours most days. Closer to 15 hours. No, I do watch what I mean. But what I'm saying is I know people who get very, very regimented about these things and it's like the marathon runners who. You know the old line that they die, don't run marathons.

I was with a friend last night and he was joking about this exact concept. And he was talking about his. He's an orthopedic surgeon. He was talking about his part partner or someone he knows. And he was like, the guy is so into yoga, but it's become. He's like, the way he described it was really funny. He's like. He's stuck in traffic and he's like, I gotta get to my fucking yoga class.

Yes, exactly.

And it's like, yeah, no, of course there's the irony to that, right?

So, you know, your insulin levels might be fine and everything's fine, but, you know, what's your cortisol levels? So I'm fascinated by cortisol. In fact, I'm fascinated by. I would love for you to basically develop a very simple test that you can sell at Walgreens where you take a prick of blood and you do it as often as you want. And you tell me my thyroid and hormones, you tell me my insulin, my glucagon, my estrogen, my testosterone, my dopamine, serotonin, and obviously cortisol. Right. These five or six, seven things that I just said, because it's about a lot of biology or physiology can be explained.

Yeah, we just. We're not going to be able to get it that way because cortisol is mostly bound to albumin and cortisol binding proteins. So it's the free cortisol that exerts its metabolic effects and its physiologic effects. And most of it's not free. So you can only measure the free cortisol in saliva and urine. And then the other, you know, of those other ones.

Well, you know what I'm saying. I would love to have those hormones at my disposal. Is my testosterone too low? My estrogen too high? This too high? Insulin. You know what I mean? Just having that data points all the time. And then you could sort of modify your diet and your exercise. But I think the cortisol one is. I pay more attention today to stress than anything else. I'll be honest with you. I mean, I still exercise and I watch what I eat. Those things seem now intuitive because I've done it for so long. But I've been wondering if myself and many of my colleagues, especially because we fly, give talks, you have grand pressures. A teenager, daughter. She's lovely. Lots of different pressures we all have. And if that somehow is being manifested. Metabolically through cortisol to the mitochondria, just like we think about insulin. Right. So that's all. That's the only reason I want to talk about cortisol.

No, I think I agree with that wholeheartedly. I think certainly in the last three years, as I've dug my heels into it, I think hypercortisolemia is a problem. And I think I wish people would think of these hormones through more broad endocrinologic terms. It's very easy for people to think of hypo and hyperthyroidism. We accept those as states. You can be euthyroid or you can have too much or you can have too little. And yet people have such a hard time thinking of insulin in those terms. You can have too much, you can have too little. There's a range in which this hormone makes sense. And cortisol is probably equally important, if not more important, in terms of the damage that can be done, especially from too much with respect to everything from blood pressure, which would then impact the endothelium. What it does in terms of inhibiting melatonin secretion in the brain. And melatonin obviously plays an immediate role in terms of sleep, but also plays an indirect role in terms of neuroregeneration. And that says nothing about what we just talked about, which was the role that cortisol metrics may even play in the mitochondria, which I'm just learning about literally in the past couple of months. So I don't disagree. I think the challenge, in many ways, for anyone listening to this, if we're going to be brutally honest, I think for many people, it's easier to control what they eat, how they exercise and exert discipline around taking medications, taking supplements. But in many ways, one of the hardest things to control is our response to stress. And I think that's an important distinction to make. I don't think there's any, anything that's particularly troubling with being in stressful situations. I think the difference is less about the situation you're in and more about the response you have to it. And that's probably where the greatest differences lie between people, is there are some people who can be in relatively low stress situations, and yet they're sort of, they're not reacting well to it, they're not coping with it well. And there are others who can be.

They have different set points where they become again, from maybe.

I mean, I, I guess I just don't understand enough of this stuff. I mean, I, I, I think.

But it's I don't, you know, I don't hear too many people talk about it.

I don't know, I think it's, I.

Think no, people talk about stress, but, but sort of like we talk about insulin all the time and glucose levels and for men, testosterone.

You mean sort of in longevity circles.

Yeah, in longevity circles. Like, you know, is that a variable we're missing? You know.

No, I, I, I agree. You're right.

You know what? Itching, cortisol.

Well, part of it is we don't have a target for it. Right. No one's thinking about pharmacologic ways to manipulate this. And we don't have great, obvious ways to curb our behaviors. Like meditation probably is the single most valuable thing I've ever found to help regulate this. But you also don't have the ability to measure cortisol levels that easily. Every time you want to do one of these tests, you're collecting urine over the course of a day and doing a bunch of other things. So it's involved. You don't have the glass of wine. Yeah. You know, it's really funny. I mean, I think there was a paper that came out probably about three months ago that looked at, basically the punchline of the paper was, look at any dose. Alcohol is toxic.

But, you know, if you look at those events, it's like 950. Yeah, yeah, yeah, 954.

The point of it is there's no dose of ethanol where the ethanol becomes valuable, but the toxicity takes a while to kick in. So, you know, for some people, a glass a day seems perfectly reasonable. There's no toxicity. But the flip side of it is, and this is where I kind of try to have this discussion with every patient is, look, I'm not going to tell somebody not to drink. I mean, I'm not going to tell myself not to drink. I probably have four drinks a week and, you know, I pick and choose my shots. You know, I have this rule called don't drink on airplanes because the alcohol on airplanes sucks. So I'm not drinking alcohol just for the sake of drinking alcohol. But if you're sitting there and the alcohol is really great, and it's something you really. The downside of the ethanol, the hepatic toxicity of the ethanol can be offset by the emotional benefit that could come from the enjoyment of having that glass of wine with your buddy.

That brings me to another one of those things. We should always measure alt, you know, the liver enzymes.

How well your liver, how's your ALT this morning, by the way?

It's pretty good.

Yeah.

I haven't checked it and so I think we're almost at the end.

Yeah.

So I have one question for you to sort of wrap it up.

Wait, I didn't think that was part of the rules.

Well, it is, because I'll tell you why. So when I interact outside of the scientific circles, if I'm at a dinner party, if I'm at a bar, if I'm on an airplane, and whoever I'm engaging with, they ask me like, what do you do? Now if I say I am, technically my title is professor of medicine and cell biology. If you say that, they think, oh, you teach something. And I do indirectly. But what we do is research. So if I just say I do research or if I say I'm a scientist, they go, oh, that's nice. But minute I say I'm a metabolism scientist, it's like, aha. Like they light up and they want the next question, which is what should I eat? You got it.

But wait, can I ask you a question given that you know that that's going to happen? When you go to parties, do you go out of your way to make sure that you don't prime people for that question or do you enjoy being asked that question?

I think I enjoyed being asked that question initially as a way to tell people, look, science is cool, metabolism is cool. You know what I mean? Sort of not think there's this image that people have about scientists. As you can see, I'm pretty flamboyant, so I figure, you know, something that they can relate to as a common language. But now I regret it because all I hear about is what should I eat? What should I eat?

Oh yeah. So here, let me give you a piece of advice on this, Nev. So first of all, I have the same problem whenever I'm in a situation. Could be a wedding, could be a funeral, could be a party, it doesn't matter. So I've learned that there are two. I have two go to things that I tell people I do for a living. And I know enough about each of them that I can almost never get called out. And the good news is both of these generate almost no follow up questions. Now the difference is you're an extrovert and I'm an introvert. So you at a party would like to talk to people. I, on the other hand, don't. I don't like to go to parties. But if I'm dragged to a party, I love to go to party. I don't, I don't want to be at parties. I don't want to be at happy hours. I don't want to be around anybody. It can except two of my friends at a time sort of thing. So I just tell people I shouldn't admit this now because now if somebody hears this, they'll know my trick. All right, I'm not gonna say it. Oh, I'm not gonna say it.

You're gonna leave them hanging?

I'm gonna leave them hanging. But I have two awesome alter egos that whenever I met and I. In fact, I busted one out last night. I was at a dinner thing last night. I didn't know people, and there are a lot of doctors there, and there was a lot of butt sniffing, which always happens at doctor parties where everybody wants to sniff everybody else's butt. You know, it's like kind of the dogs, you know, walking around sniffing each other's butts. And there was a lot of, what do you do? Oh, I'm the chairman of this, and I'm the chairman of that, and I. I am the head of this and blah, blah. Then they looked at me, what do you do? And I just said, I'm. And I said it, and it was amazing. The crickets are chirping. Everybody is like, they don't know what to say. And they said collectively nothing. And then the discussion just went elsewhere, and it was awesome. I didn't have to talk about anything.

Right. So I do give an answer, and I want you to tell me if this is the right answer or the wrong answer. I still liked, and I know it got debunked a little bit. I still like the Mediterranean diet. I mean, not. I mean, with curry. Okay. But generally that kind of diet with nuts and avocados.

Well, when you say debunked, I mean, I'm not even sure I would agree on either. I don't think it has.

But you know what I mean by that.

What you're referring to is the predimed study, which I'm guessing many people listening to this will know what that is, but in case somebody's not, we'll certainly link to it. But this was a study that randomized something in the neighborhood of 7,500 patients. Although we'll come back to the word randomization. I'll put a little asterisk beside that into three groups. So about 2,500 patients per group. And they were randomized in a one to one to one fashion between a Mediterranean diet that was high in extra virgin olive oil oil, a Mediterranean diet that was high in nuts and a low fat diet. And this was a primary prevention study, which makes it a very difficult study to do, especially with nutritional therapeutics. And the study was stopped early. It was stopped at about 4.7 years, if my memory serves correctly, because both Mediterranean arms were outperforming the low fat arm. Now, I used to view that as one of the more interesting studies ever done in nutrition because nutrition studies generally suck. It did have one major criticism that didn't get any attention at the time of the initial publication, which I think was 2014 maybe, and that was the performance bias. So the groups that were getting olive oil and nuts had those products sent to them. The low fat group, to my knowledge, did not receive anything, didn't get food given to them. Well, that creates the potential for a difference in behavior. And that's problematic. That's very problematic in clinical trials where you can't blind anyone. But the more recent issue, the one that I think you're referring to, is some irregularities popped up in their randomization. So some people doing post hoc analyses found, hey, these numbers don't make sense. It's very improbable that these people were all randomly assigned. And I believe, because it's been a while since I read the correction, that what they identified was that a number of those patients were not randomized correctly, for example, and it wasn't nefarious, but it was done through convenience. So. So if you had a husband, wife, team in the study, they were immediately put on the same diet, which, by the way, is logical. That's actually a better study design. But you have to then change the statistics to accommodate for that because you have no longer randomized each of those individuals, they would be considered one randomization, not two. To the best of my knowledge. Even when you take into account those changes or those inconsistencies or those methodologic failures, I don't believe it changed the results or the outcome of predimed. So we're still back to the initial limitation of were those representative diets and were those subjects the victim, for lack of a better word, but the victims of a performance bias? So all that said, look, I think the Mediterranean diet, which is unfortunately not a very descriptive term because what the hell is a Mediterranean diet? Is that what people eat in Italy, Egypt, Greece, Spain? Like, I mean, so I guess the only opposition I would take to the concept is I don't, I like to be more specific in my description of the diet.

What I dislike is high protein or a high carb or a high Fat diet, but people love. So the high protein people like is they want to look like a South beach model, essentially, as far as I can tell, because you can lose weight. You know, the high carb. We know you talk about it all the time, why that's bad. And you know, sort of the ketogenic diet, I mean it has some, some benefits maybe for the brain and other systems, but you know, it does make you insulin resistant. I mean there's data in mice.

Well, I would take issue with those data though. Right. So are there data that ketogenic diet make mice insulin resistant?

I don't think there's hundreds of studies in mice that people have done. And so my favorite one, I think you should, I sent it to you where they looked at a whole bunch of diets and essentially the best one was sort of a 1/3, 1/3, like 20% protein. Because we can both agree if there's too much protein, your MTOR might be quite active if you need enough for your muscles. But this is again, not in the elderly, so again not with the disease. This is just primary prevention that we're talking about in healthy sort of 40 somethings. Let's start with. So it was relatively not high in protein and it had about 40, 50% carbs almost, I think. And this was a mice study. And then the rest was sort of the good fat, you know, avocados, nuts and stuff like that. And so I sort of liked that diet because it was a pretty good study in cell metabolism published on, on these mice. And, and to me, intuitively, some of this makes sense. And so, you know, keeping protein not too high because you want to keep.

Is that the paper that Simpson was the last author on? Yeah, Stephen Simpson, I think is maybe there was like. He's Australian.

Yes, there was 25. Again, I don't think there's a clear answer to this, but you know, rather.

Than answer the question, let me tell you my two cents on this topic. One, I don't have a lot of interest in mouse studies for human nutrition. I struggle with them because I think there are so many other issues going on and it's very hard to make that that's a fair point. The second thing is I always want to be sure that I'm distinguishing between short term insulin resistance and long term insulin resistance. So I think you're right. In the short run, ketogenic diets in a non trivial subset of people generate profound insulin resistance in the muscle. Again, I don't even know what insulin resistance means. If we're Going to be truthful, like if we were going to put me in the confession booth, I don't have a goddamn clue what that term even means. It's so ubiquitous. Does it mean the failure of one type of cell but not another type of cell to respond to insulin signaling? I mean, all of these things, but I think I know what you mean. You can see huge elevations in glucose and insulin and basically a complete refusal of the muscle to accept glucose in someone on a ketogenic diet when they first encounter a carbohydrate. But I think it's generally also regarded that after about three days of carbohydrate refeeding, that effect goes away. And that that effect is sort of a physiologic response to an individual who's been so carbohydrate deprived that their muscles are basically saying, any glucose in the system we're going to preferentially save for the brain, since we now have all the fatty acids and beta hydroxybutyrate in the world we need as metabolic substrate. So the short answer is, I don't know. I'm. These days I find myself far more interested in fixating less on the exact amount of this micronutrient or this macronutrient or that micronutrient, and more on the complete deprivation of calories for more prolonged periods of time. So, you know, people who are used to following me these days, I'm spending much more time thinking about fasting than I am sort of sticking on one diet and sticking to it.

Yeah. So I basically eat a third of my calories, probably from fat, carbs and protein. And then the other thing is just the 15 hour fast every day.

But if you're getting a third from protein, that's probably quite a bit.

Probably. I think probably have to lower that.

You don't have to check your Torah.

Well, maybe third is too much, but I eat a fair amount of protein.

Probably at the parties. Tell them you're a math professor, they won't ask you any more questions.

I don't remember any math and we'll leave it at that.

Yeah, but hopefully they won't ask you the questions.

No, I'm an extrovert and I like talking about metabolism. I just, I don't have a good answer on the diet. I have lots of answers on mitochondria and all that.

But then just tell them you're. Tell them you're a mitochondrial expert, but don't use the word metabolism.

But then people really look at you.

Like, oh, anyways, hey, man, thank you so much for for taking the time to talk about all this stuff. I had written down a bunch of things I wanted to talk about and I think we actually got through like half of them. Oh only okay, well, but that's the nature of this stuff. It's so fun. There's so many rabbit holes to go down. In particular, I really loved the double double click deep dive on Metformin, which is something I think a lot about myself.

Hopefully once we have some some results and there's more clinical trials and come back and we can really talk about some more answers.

All right man. Thanks.

Thank you.

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