Dr. Guido Kroemer on Autophagy, Caloric Restriction Mimetics, Fasting & Protein Acetylation

Dr. Guido Kroemer on Autophagy, Caloric Restriction Mimetics, Fasting & Protein Acetylation

Hello, everyone. Today I’m
sitting here with Dr. Guido Kroemer, who is a professor at the University of
Paris Descartes. He is a cell biologist who has made major contributions to
understanding the role mitochondria play in cell death. He has also published
numerous publications in the fields of cell biology, cancer biology,
immunology, aging, and autophagy. The latter of which, I suspect, we’re
probably going to talk a lot about today because it’s such
an important topic. So, autophagy is a
spectacular phenomenon in cell biology, one that you can see with your eyes
because cells become vacuolated when this process is induced. So you can see
it by light microscopy and better, of course, by electron microscopy. It is
a process that consists in sequestering portions of the cytoplasm of the cell
and then digesting them to recycle the material and to degrade macromolecules
into micromolecules, metabolites, and to allow rebuilding the
structures that have been destroyed. So technically, it works in the sense
that the mouth that is involved in this self-digestion process is the
autophagosome. So, it will sequester portions of the cell in the cytoplasm.
It can be an entire organelles including mitochondria, it can be protein
aggregates, it can be bacteria or viruses that invade the cell, and this
autophagosome, once it closes, will fuse with the lysosomes, which is the sort of
stomach of the cell. And in the lysosome, it has been fusing with the autophagosome,
which is then called Autophagolysosome, the luminal content
will then be digested. So you mentioned the digesting
of these multiple organelles or mitochondria, but also protein
aggregates, and viruses, bacteria, pieces of chromatin, and things all seem
to be sort of things that at least in the sense, if you’re looking at the
aggregates, and you know, damage that occurs in a cell, seems to be something
that’s associated with aging in general. So it’s sort of like, kind of seems like
it’s getting rid of all these damaging, potentially damaging, not just aging
but also obviously, an infection. But this process is getting rid of these
damaging, potentially damaging molecules and aggregates and
mitochondria which are defective. What is the actual goal of autophagy is?
Like you said, is to get rid of these defective things to
then provide energy? Well, it’s actually
interesting to look at the history of autophagy. The name comes from “autos
phagy” in Greek which means “self-eating.” And it’s actually a linguistic invitation
to think about self-mutilation and self-destruction and cell death. And
actually, the phenomenon is observed mostly in the context of stress.
So cells, when they are stressed, will often undergo an autophagic reaction
which occurs before the cells die. And so, this chronology of the phenomenon
has been also an invitation to think that autophagy is a mechanism that leads to
cell death until it has been understood that inhibition of autophagy, which can
only be achieved in a specific way by genetic tricks, will actually sensitize
cells to cell death induction. And so, this means that autophagy is a
means of adaptation to stress and a technique of the cell to avoid cell death.
So, the primary goal of autophagy is adaptation to changing conditions
and adaptation to external stress, and at the end, avoidance of the
unwarranted demise of the cell. You bring up so many important
points that I’d love to touch on. The first is the external signals
that are actually causing autophagy. You mentioned it’s a response, it’s
a generalized response to stress. Like, you know, mostly probably a hormetic
type of stress, but even…this is another question I’ll have for you later, the
differentiating between the type of stress that can cause autophagy versus
actually pushing it over to cell death. But in terms of the actual external
signals that…the main ones that we know about that cause autophagy, a lot of
them have to do with nutrient sensing. Exactly. So,
perhaps the most physiological way, to induce autophagy that is
phylogenetically conserved from yeast to primitive animals to ourselves is
nutrient deprivation, starvation, hunger. And so, the idea is that a
cell that is deprived from its energetic supply, which can be the absence of
nutrients or the absence of growth factors that are required for these nutrients to
be transported from the outside world into the intercellular space or the absence
of oxygen, all these factors can induce autophagy, and the cells actually will
destroy its bioenergetic reserves, which are macromolecules,
proteins, lipids, and ribonucleic acids
to generate energy. In terms of the energy part, I
was reading about like three really major pathways that seem to lead to autophagy.
One, being the actual energetic charge of the cell, like ATP status. When that
lowers, that activates the AMP Kinase pathway. And then the amino acid sensing
pathway, which then when you don’t have enough amino acids, that can basically
inhibit mTOR. And then there’s a third one, the protein assimilation pathway,
which I’m not as familiar with essentially how that activates autophagy. So, it’s actually
extremely easy. When you think about basic biochemistry, one of the central
metabolites is Acetyl-CoA. And so, the cytosolic pool of acetyl-CoA
determines the level of protein acetylation for the simple reason that
acetyltransferases, which you see acetyl, moiety of Acetyl-CoA, to transfer
it on lysine residues in proteins. Acetyltransferases are having a low
affinity for Acetyl-CoA as compared to kinases which have a high affinity for
ATP. So if you vary the ATP concentration in the cell, it has little impact
on phosphorylation reactions. But if you vary acetyl concentrations,
it has a major impact on the acetylation level of cellular proteins. So
that’s a major difference. And so, since Acetyl-CoA is built in the
degradation of glucose, a glycolytic pathway, from pure beta or in the
catabolism of branched amino acids, as well as a final product of
beta-oxidation. All major nutrients are actually supplying Acetyl-CoA as an end
product. And taking away glucose or amino acids or fatty acids will cause a
reduction in the Acetyl-CoA pool which is important to note, it is a cytosolic acid
Acetyl-Co-A pool that is accounting for autophagy regulation. And this reduction
of Acetyl-CoA in the cytosol will cause deacetylation at the end of cellular
proteins, hundreds of different proteins, and hence, a sort of multi punked
induction of major subpathways of the apoptotic process. Autophagy is
actually a very complicated process that involves dozens, perhaps hundreds, of
different proteins and is regulated by hundreds, perhaps thousands, of additional
proteins. And so, this common regulation by acetylation is very efficient in
stimulating the autophagic pathway. As a side effect of deacetylation
reactions, you usually also observe from the inhibition of mTOR and the activation
of AMP Kinase. So, everything comes together at the end. There’s no
exclusivity for one or the other pathway. They are connected. Really? So, changing the
acetylation status of proteins affects mTOR and AMP Kinase? Yes. Indirectly,
we don’t know how this actually works in molecular details. Because that was sort of, my
next question was I know that if you take cultured cells in a dish and you
just…you’re doing some, you know, specific nutrient withdraw, you withdraw
amino acids when you withdraw glucose, or you withdraw glutamine, you can induce
autophagy. But in the whole organism, for example, in mice and in humans,
ultimately, you know, can you just limit your protein intake for a week and induce
autophagy even in the presence of a normally caloric diet where you’re
still getting enough energy. That’s a good question.
We have never tested to selective completion of one or the other nutrient,
I suppose that this would work, because protein depletion may affect
a newer endocrine factors like insulin growth factor that, at the
end, will, due to its depletion, decrease the transport of glucose into the
cells. And hence, stimulate autophagy. But it has not been tested thoroughly in
mice. What we usually do is we starve mice and sometimes human volunteers completely
from any kind of caloric uptake, and in this case, we do see, at the whole
body level, that in all major cell types, perhaps with the exception of the brain
that is somehow buffered against this effect, protein deacetylation
occurs mostly in the cytoplasm. And that’s a biomarker
of autophagy, you would say? Well, it’s too early to
say that it is a surrogate or proxy of autophagy. So far, we have not been
able to dissociate the two phenomenon, autophagy and protein deacetylation,
in the response to nutrients. However, when you induce autophagy by
pharmacological tricks, such as cell permeable peptides that
dissociates an inhibitory interaction between a Golgi protein and Beclin 1. You
can induce autophagy without that protein, deacetylation would occur before. And
similarly, when you give chemical inhibitors of mTOR like rapamycin or
the rapalogs, there’s also no protein deacetylation. So, you can induce
autophagy without protein deacetylation which means that the proxy would be
imperfect. So we do have a system to measure autophagy which is relatively easy
to be used in experimental systems which is the study of the redistribution of LC3
and other members of the same family that are usually diffusely distributed all
over the cell, mostly in the cytosol, and then we’ll aggregate or redistribute
towards autophagosomes and autolysosomes. So they acquire a punctate distribution
small dots in the cytoplasm and these dots can be seen by fluorescence microscopy if
LC3 is labeled by immunofluorescence or when it is fused with green fluorescent
protein or similar biosensors. And so, in humans, the only accessible
cell type is the circulating white blood cell, the leucocyte. So we can draw blood
and determine by immunofluorescence the redistribution of LC3 from a diffuse
to a punctate from a pattern. And this is then a sort of detection of
autophagy that can be applied to human beings as well. That seems like it’s kind
of complicated though for your standard clinic to be able to use
immunofluorescence to look at some leukocytes, circulating leukocytes, right?
I mean that’s more… Well, you need some
technology. Especially, ordinary cytofluorometry cannot be used for this
kind of approach, because the standard cytofluorometer just measures an intensity
of fluorescence signal per cell not its subcellular distribution. So there are
cytofluorometers that take pictures of the cells that are flying in front of the
detector, and using these pictures and analyzing them by image analysis
software, allows them to quantitate the redistribution of LC3
to autophagosomes. So there is hope for a
non-invasive clinically relevant biomarker for autophagy but there still, it seems
like there needs to be more work done before that actually happen…before
I can go to my doctor and say, “I did a four-day fast. I’d like to
see if I’ve activated autophagy. Can you please draw some blood.” Right?
We’re not quite there yet. Yeah, it would be
wonderful to have the reward of measuring autophagy as a result of fasting
and to get an objective incentive as a biomarker for doing that. Right. So it kind of brings up
another question I had which related to when you started talking about how you
can fast, and fasting in organisms like rodents and also in some human
volunteers does induce autophagy. And the question that I had for you
is, like, I’ve talked with Dr. Valter Longo, he was on the podcast, and
he talked quite a bit about his research on prolonged fasting in both rodents and
also in humans and how the prolonged fast, at least, in rodents is 48 hours,
which in humans is around 4 days, 4 to 5 days. And that was able to very
robustly, not only activate autophagy, but also cell death, and that was
followed by a regeneration period. But, the question is do we know what the
minimum amount of fasting time is for humans or rodents that can activate
autophagy? So for example, when I’m not pregnant, I usually followed
a very time restricted eating schedule where I like to eat all of my food within
at least 10 hours, and then I fast for 14 hours every night. Some people do even
more strict. They eat within 8 hours and they fast for 16 hours. Does that
16-hour fast induce any autophagy in any of our tissues? Is there
any evidence, do we know? We don’t know. So, Craig
Thompson published a paper on circadian variations in hepatic autophagy.
So you know that mice don’t eat during the day and they eat during the night, and
so, the entire cycle is inversed. And he observed that as a result
of not eating during the day, there was more autophagy in the liver.
So this result is intriguing. It has not been, to my knowledge,
extrapolated to other organs. And it still certainly requires
more profound studies. Okay.
When you say during… So what we did on
circulating leukocytes is that we needed to wait for three or four days to
see a massive induction of autophagy. There’s a fundamental difference
between rodents and humans, and so, the two days that you have been alluding
to cause a 20% weight loss in mice, that are, at this time point, at the verge
of death. Another day would potentially kill them. And so, 20% is a lot,
so imagine this for yourself. In two days. In four days, a human
being only loses one to two percent of his or her weight. Is that because they have
a higher metabolism, rodents do, or…? Yeah, it’s certainly
linked to the change in the surface volume ratio that is classically associated
with an accelerated metabolism. Yeah, okay. That sort of…
So we don’t really know to what extent autophagy can be occurring in a shorter
intermittent fast. There’s some hope that it does. I mean, I know, for example, you
mentioned IGF-1 and how IGF-1…lowering IGF-1 is important for inducing autophagy
because in the whole mTOR pathway and so on. But I do know that the half-life
of IGF-1 is, in a serum at least, is around 12 hours. So, the question
becomes well, okay, if you start to lower IGF-1, after 12 hours, is do you still
need more to occur, like more ATP depletion, more…what…you know, what is
it that needs to happen to actually send a signal to the cells to go, “Oh, I’m
stressed. I need to start eating my whatever organelle or damaged proteins
or something.” So is that something that people are currently investigating? Like,
the minimum amount of time that it would sort of take to induce, for fasting
at least, to induce autophagy. It’s an extremely
interesting question that is easy to be answered in rodents, and difficult in
humans. Because it may be easy to find a volunteer who fasts and allows
for regular blood drawing, but it will be very difficult to find a
volunteer who fasts and allows for liver, muscle, or skin biopsies. Right, right. It kind of
reminds me there was a study I was actually reading the other day that was
done in the Caloric Restricted Society. You know, there’s a group of people
that are out there practicing caloric restriction which typically is eating
around, what, 30% less food than you normally would eat or something like that.
A lot more difficult for people to maintain, I think, than intermittent
fasting is, but there was a study that was published. And these individuals had been
doing caloric restriction for about six years, plus or minus. I’m sure you’ve seen
the study. But they did muscle biopsies on them and they measured LC3. They
measured some of the biomarkers of autophagy, I think Beclin and some other
things. And then they measured heat shock proteins, which are also a stress
response. And it was, you know, like in some cases, the heat shock
protein, HSP70, was elevated by 12 fold compared to age-matched, lean controls
that eat more of a Western-type diet. But you know, the fact of the matter
is that they did do a muscle biopsy. Autophagy was activated. You know,
the stress response pathway, in general, was activated. But six years
of doing caloric restriction is not very sustainable for the majority of
the population in the, you know, at least in the United States
and in Western world. Actually, in mice, you
can obtain exactly the same longevity extension that you would obtain, the 30%
of caloric restriction by intermittent fasting. So, it’s logistically much more
difficult. Imagine, you have to weigh, for each mouse, the amount of food that
they would eat normally, subtract 30%, put it in the cage, individual cages
because calorically restricted mice tend to eat each other. Oh, wow. Yeah, they become
aggressive because they are hungry. They
become cannibals. Yeah, they become
cannibals. So, it is logistically much more simple to take out the food from the
cage completely, and to put back the food on the next day. So it’s one day without
any food, and another day with normal nutrition. And at the end, so, you have
an oscillation of the weight of the mice, 10% every day. These oscillations tend to
become smaller because the mice somehow adapt to this sort of stress, but the
final result is that the intermittently fasted mouse has the same weight as a
normally fat mouse. A difference with the calorically restricted mouse, it weighs
also 20% to 30% less. And in spite of this difference in the body weight,
intermittent fasting allows for a lifetime expansion in the same way as does caloric
restriction. So, one can also consider that this may be more amusing to have, if
I was a mouse, I would probably prefer the intermittent regimen, because it means
satisfaction during one day and dissatisfaction on the other day,
but not permanent dissatisfaction. Most people prefer
doing intermittent fasting. I mean, it’ll be very interesting to see more
studies come out on, you know, the translation of this to humans and,
as you mentioned, you needed three days of…was it a water fast they did? Was it
a complete fast or they had coffee or…? Coffee. Tea. Okay. No food. No sugar,
no milk. And water. Okay. So three days was enough
to, at least, show signs of autophagy in circulating leukocytes. And
Valter’s work has shown, you know, four to five days and he’s done. You know,
he’s got his fasting and then he’s got the fasting-mimicking diet into… Some
also hints that that also is enough. So, that’s sort of encouraging. It would
be more encouraging to have like a 24-hour fast or 48-hour. I mean, that’s
so much easier to do in general. But the other thing that induces
autophagy, you’re mentioning the stress response and oxidative oxygen, and it
sort of reminded me of exercise and how exercise also induces autophagy. I’ve
seen some studies where in humans, they’ve looked at muscle, skeletal muscle,
and how aerobic exercise and eccentric and concentric exercise all can activate
autophagy in skeletal muscle. Do you know if it activates autophagy
in multiple tissues? Exercise? That’s something
that we have not studied. So, it is known that endurance training is
particularly efficient in mice to induce autophagy and that it mediates
anti-obesity and anti-diabetic effects that are depending, in a way, on autophagy
induction, because genetic modifications of the process that leads to
autophagy induction, it’s inhibition, specifically by exercise, can prevent
these anti-diabetic effects. Really?
Yeah. Oh, I didn’t know that the
role of the exercise in preventing diabetes was shown to be dependent
on autophagy to some degree. That’s very interesting. So, do you
think that has to do with the liver, and the like pancreas,
somewhere… I mean, is it known? To know this in detail,
it would be necessary to inhibit autophagy specifically in different
tissues and, to my knowledge, this has not been done yet. Okay. So do you think fasting
while you’re like exercising in a fasted state… Now, that’s another thing
that’s…do you think that would be important? Or do we… I mean, I’ve seen
some studies in mice where they claim it is, but mice have a very high metabolism.
And so, there’s a synergy there. But when you look in humans, it’s not so
important. Like, the exercise can still induce autophagy in skeletal muscle in
humans even without being in a fasted state, but the question is like
will you synergize more and… You can
speculate, but we don’t know. I’m giving you a lot of ideas
here. So maybe we can kind of talk, shift a little bit into the general role
that autophagy plays in some of these age-related diseases, like
neurodegenerative disease, cardiovascular disease, and cancer. Talk a little
bit about the microautophagy, or is that what you call it, like,
when you’re talking about the specific degradation of organelles like
mitochondria or protein aggregates? So normally, when
we refer to autophagy, we talk about macroautophagy, which is the phenomenon
that you can see easily by microscopy. Because of the formation of the
autophagosomes that are big enough to be seen by conventional microscopy, face
contrast, and especially of course when you enhance a solution by
immunofluorescence or similar technologies. So, there are other types
of autophagy that are less well studied. Like, chaperone-mediated autophagy or
microautophagy where basically proteins or portions of the cytosol are
introduced directly into lysosomes. So you don’t need the mouth of the process
or autophagosome, you just need the lysosome. And they are much less studied.
And then there’s a special case among different kinds of macroautophagy. So
to be very simple in the dichotomy, there is the case that autophagy is
dictated by general stress or general absence of neutrons, which means
that it is dictated by demand. So the cell needs to eat some portions
of itself to adapt to nutrient stress, and the other kind of autophagy is
dictated by the offers. So a damaged organelle will change the composition of
its surface in a way that it is decorated by signals for stimulating its
engulfment by the autophagosome. And so, it’s another kind of
autophagy that then can be specific. Specific for organelles of different types
like mitochondria, and it is called mitophagy, or for peroxisomes, and it is
called pexophagy, for the endoplasmic reticulum, and it’s called reticulophagy,
specific for ribosomes, ribophagy. Perfect, yes. And specific for
viruses, and it is called virophagy. And the two processes may also interact
in a way. So when you stimulate general autophagy by activating the nutrient
sensors, AMP kinase, inhibition of mTOR, or by provoking deacetylation, then you
increase the demand, and the autophagy machinery actually prefers in a way to
sequester and to destroy those organelles that are already slightly marked for
destruction. The protein aggregates that are not yet harmful enough to emit
a signal per se but they are there. And so it’s a sort of preferential
cleaning of the slightly damaged and slightly aging portions of the cell. And
this may actually explain why stimulation of autophagy in cells, when they are
monocellular organisms or at the organismal level at different organs,
can be a sort of device against aging. Wow, that was very beautiful
explanation. It actually answered a question I was going to ask you which
was, you know, the difference between the signal, for example, nutrient generalized
autophagy, when you have the nutrient sensing stress that even that can, to some
degree, selectively degrade mitochondria, for example, but the actual signal that
really does activate mitophagy…when you’re talking about mitophagy,
it’s a little different, right? It’s the actual mitochondrial
damage, the membrane potential… Yes.so when a
mitochondrion is suboptimal in its function, it will decrease its
mitochondrial transmembrane potential. And this is a signal to activate enzymes
on the surface of the mitochondria that cause ubiquity relation, recruitment of
autophagy adapters, and leads at the end to autophagy because of the
organelles or the organelle in a way offers itself, it proclaims its
sacrifice by autophagy. And so, of course, this is not an all-or-nothing
phenomenon. So mitochondria can be aging in the cell, and as they age, they
gradually decrease the performance and the mitochondrial transmembrane potential.
So, those mitochondria that are most dysfunctional, they will be eaten first
if you increase the demand for autophagy. That is very cool. And if you
are selectively degrading these damaged mitochondria, which are you know, or aged
which are damaged, do they get replaced by new mitochondria? Is that a signal
for mitochondrial biogenesis? Yes. So in C.elegans,
this was a study that actually the whole turnover of mitochondria is regulated.
So, there’s a sort of coupling between mitophagy and
mitochondrial biogenesis. That’s good to know. So it’s very clever,
how the system has been designed. It’s great. So it’s not like
you’re losing…you’re not losing the pool of mitochondria. You’re effectively
losing the defective pool, and you’re almost making younger
mitochondria. If you’re going to make a new mitochondria, then it’s going to
be young and fresh and not damaged. So it’s very elegant
way to sort of replenish your mitochondrial
population, it seems. So we have to make the
difference between homeostatic conditions and, for instance, cellular
differentiation when cells change their metabolic program. So the easiest example
is yeast that you suddenly place in the glucose-containing medium to allow for the
fermentation of glucose in wine or beer production. So these yeast cells don’t
eat much oxidative phosphorylation, and they essentially rely during the
process on glycolysis. So they adapt to this change by destroying most of
their mitochondria, by mitophagy. And this makes actually a metabolic
adaptation of the yeast cell efficient. Do you have similar examples in the
embryonic development of the retina for retinal ganglion cells or the
differentiation of macrophages from so-called M0 to M1 macrophages, in
which the cells change from oxidative phosphorylation respiration to an
essentially glycolytic metabolism that is coupled to mitophagy. And so, inhibition
of mitophagy actually avoids the differentiation process in both
examples that I just gave to you. That’s really interesting. So
obviously, these processes are not just as a stress response, they’re
part of development as well. They can be used
in multiple different instances. Very interesting. And
in the case of mitophagy here, it’s also it plays an important role
in the prevention of neurodegenerative diseases. Correct? Yes. So, most known
neurodegenerative diseases are either caused by the aggregation of poorly built
protein cell that somehow create protein aggregates that are toxic for the cell.
Or they can also be caused by septal deficiencies in the autophagic and
lysosomal machineries that lead to the accumulation of unfolded proteins at
the end. And so, either the excessive production of unfolded proteins or
their reduced removal causes to a slow accumulation of these toxic
protein aggregates. Remember that neurodegenerative diseases are slow
processes in most cases that manifests with old age. And so, one strategy
to treat neurodegeneration, at least theoretically, is to
increase autophagic turnover. And so, one technique is actually then to
stimulate general autophagy by increasing the demand, by starvation, or by
biochemical trickster that substitute for starvation, and to reduce
the protein aggregates that are the cause of the disease. So these protein aggregates
like amyloid beta plaques in Alzheimer’s disease or alpha-synuclein in
Parkinson’s disease. So, basically, the clearing out of those protein
aggregates obviously would play an important role not only in prevention
but presumably also, to some degree, in help with the treatment. Of course,
that’s you know, an speculation, but… And then the mitochondria, the one
I was thinking about with mitophagy, was the role, at least some of the
proteins that are involved in that, like, the PINK/Parkin, and how they seem
to be important for Parkinson’s disease. Is that accurate? Yeah. So the
PINK/Parkin pathway is one pathway among others, that allows for marking
mitochondria that are damaged for destruction. And so, inhibition of this
pathway leads to the accumulation of malfunctioning mitochondria with major
consequences for the cell that harbours cells’ mitochondria because all of a
sudden bioenergetic metabolism becomes inefficient, reactive oxygen species
are produced, and as you know, mitochondria are latent bombs in the sense
that they enclose potentially dangerous proteins that once released will
activate the apoptotic machinery and cause cellular suicide. Yeah. So, that’s the question.
Do we know the threshold for the stress threshold for, you know, activating
autophagy, and when that pushes the mitochondria then to permeabilize
and cause cell death? Like, where, for example, with Valter’s
work in mice, he had done 48-hour fasts and there was both autophagy and
massive apoptosis occurring. So, is it just the intensity of the
signal that can then say, “Okay, autophagy is not going to work here.
We got to die.” Or do we know? Well, autophagy in used
in most cell types, while apoptosis is occurring in selected cell types.
So what Volter has been observing, if I remember well, is destruction of
leukocytes, right, white blood cells, which are very easily to be rebuilt. And
so, the loss of 50% or 75% of leukocytes can be easily repaired in a few days. And
it is a way to adapt the repertoire of immune cells to changing circumstances.
It is a way also to inhibit unwarranted inflammatory reactions. So depending on
the context, induction of autophagy can be actually a subtle way to avoid excessive
inflammation. One example is the so-called sickness response. So, a cat or a dog or
a human being or a mouse that is sick, that has a bacterial infection, will
hide away, avoid light and noise, and will not eat. It’s a classical
phylogenetically conserved reaction in most cases of bacterial infection. And
so this phenomenon leads to changes in the metabolism. Ketone in the production of
ketone bodies, the reduction of glucose levels, presumably also induction of
autophagy, and altogether these mechanisms avoid excessive inflammation that may be
lethal. So Aslan Medzhitov published a paper in cell last year showing that
force-feeding mice or just increasing the glucose levels to a normal concentration
was sufficient to make bacterial infection that otherwise would have been
able to cope with lethal. Wow. So, I know in humans too.
And we have a bacterial infection, for example, a stomach virus or
something that’s bacterial of origin, you don’t eat as well. So, it sounds like
it’s sort of a protective mechanism. It is. That’s really interesting. I
didn’t know that. It’s very interesting. I want to kind of move on to cancer, just
for time purposes. So cancer is another sort of very, it’s been, in regards to
autophagy, something that I’ve always sort of been unsure about, because it’s
very clear to me that preventing the accumulation of damage, you know, pieces
of nucleic acid and pieces of chromatin and all sorts of things that can cause
inflammation by having damaged proteins around and things like that. Obviously,
clearing those out would be very important for preventing cancer. But when it comes
to treating cancer, it’s not as clear. There seems to be… I mean, for example,
you know, there’s a very classic drug out there, chloroquine, right, that inhibits
autophagy that’s used to kill cancer cells. But… Well, it’s not exactly
[inaudible] or chloroquine is a lysosomal inhibitor. It’s a molecule that,
due to its charge, will specifically enrich in the membranes of lysosomes, and
then causes lysosomal membrane damage, potentially also inhibition of autophagy.
But it fundamentally also liberates the potentially toxic content of lysosomes
into the cytosolic space.and so, there are a few reports around showing
that inhibition of autophagy is not the sole mechanism by which chloroquine
can mediate such a toxic effects. Okay. Well,
that’s good to know. The other thing that is
important to notice is that chloroquine and hydroxychloroquine, which are
antimalarial agents that have been used for a long period, and also actually used
for the treatment of rheumatoid arthritis because they have anti-inflammatory
properties. Only introduced into clinical trials, most in combination with
chemotherapy or radiotherapy to treat cancer. And those clinical trials,
so far, are not convincing. Okay. So what about
the fact that some cancer cells do activate autophagy? So one relatively general
mechanism may be that early during oncogenesis, the deletion of tumor
suppressor genes or the activation of oncogenes leads to autophagy suppression.
So there are several examples for this. And it is part of the process that
leads to cellular transformation, because autophagy is a homeostatic
mechanism that, if inhibited, favors genomic instability and
malignant transformation of the cells. So there are examples on the literature
also that dived inhibition of autophagy is sufficient to cause oncogenesis, in
particular, in the context of leukemia. And so, later on, when the cells strive
and adapt to an ever more hostile microenvironment, hostile because there’s
too little vascularization for the expanding cancer cells, so initially there
are hypoxic areas, there’s no normal tissue architecture, so the cells are
usually undernourished. The doctor may apply some chemotherapeutic agent
which is an additional stress. So there are internal and external stress
pathways that the cell has to cope with. And it is an advantage for the cancer
cells to reactivate the autophagic process. And so, it has been proposed that
inhibition of autophagy would be a way to make the cancer cells more fragile and
vulnerable to therapeutic intervention by chemotherapy, radiotherapy, targeted
therapies. The problem is that nothing is simple in oncology and that cancer is
not just a cell-autonomous disease. It is more. It is not just that one
cell has become wild term and has been accumulating genetic and epigenetic
changes that make it selfish. No, a cancer cell will only survive if
it escapes from amino surveillance. So the immune system, fortunately for us,
is usually very efficient in eliminating aberrant sells, premalignant cells,
and the initial cancer cells. And actually, the inhibition of autophagy
that occurs during early oncogenesis maybe also a way for the cancer cells
to hide from the immune system. And so, it is complex. It’s immunology,
multiple different players come into action. Autophagy, for instance, is
required for stressed cells to release ATP into the microenvironment. You know,
of course, ATP is the most important, energy-rich metabolite in the cell. It’s
like the equivalent of the dollar for bioenergy clinics in the economy. And ATP,
when it appears all of a sudden outside of the cell, is considered as
non-physiological. It is a dangerous signal. It is perceived by so-called
purinergic receptors that are present, among other cell types, on leukocytes,
and particularly myeloid cells. And a cell that undergoes autophagy may,
especially, when this occurs before cell death, release ATP to attract myeloid
cells into its proximity and to start an immune response against tumor antigens
in the context of the initial oncogenic events. And so, autophagy is required for
some steps of the immunosurveillance process. And it is exactly this process
that makes cancer therapies efficient. So, in contrast to the official dogma
that has been en vogue for several decades, chemotherapy is not just killing
the cancer cells as if we used an antibiotic that specifically paralyzes
the metabolism of bacteria. No. It is true that chemotherapy induces
cancer cell death, but the important point is that chemotherapy must provoke this
cell death in a way that it later leads to an immune response. And so, if you have
a long-term effect of chemotherapy, for years or decades, that continues
beyond removal of the drug, it is due to an anti-cancer immune
response. And so, since this is so important, the capacity of the
chemotherapeutic agent to induce autophagy is actually required for the
long-term efficacy of the treatment. I did not know that. I had no
idea that the induction of autophagy would stimulate the immune system through
this extracellular ATP mechanism. And how that is, I mean, obviously the
immune system is extremely important for killing cancer cells. But that’s very
cool, and probably leads to the next topic on some of your work with the fasting,
so-called fasting mimetics like spermidine, hydroxycitrate that you’ve
done. Maybe can you kind of just briefly explain… I’ll start with spermidine.
What is spermidine? What does it do? Well, it was first
start to explain what are these fasting mimetics is as you say and
caloric restriction mimetics as we say. So the CRM’s, caloric restriction
mimetics, are actually inducing the same biochemical changes in the cells
as would do starvation or fasting. So, we have been discussing on the
importance of acetyl-CoA and protein deacetylation resulting from the
inhibition of Acetyl-CoA in the context of fasting. And caloric restriction mimetics,
similar induced deacetylation reactions to stimulate autophagy. And this can be
actually achieved in three different ways. First, you simply inhibit the generation
of Acetyl-CoA. The enzyme that generates Acetyl-CoA in our cells, the most
important one for the cytosolic pool, is ATP citrate lyase and hydroxycitrate
or pharmacological compounds that inhibit this enzyme cause Acetyl-CoA depletion,
deacetylation, and autophagy. And you can have the same effect by
inhibiting the protein acetyltransferases, some of them have been identified. Like,
EP300 which appears extremely important for autophagy regulation, and specific
inhibitors of EP300 such as spermidine and natural compound or C646 which is a
pharmacological compound specifically designed for this function. They can
also cause deacetylation and autophagy. And finally, it is possible to activate
deacetylases or enzymes that remove acetyl groups from proteins and cause
hypoacetylation and autophagy. And one example that is well known is resveratrol
contained in red wine that induces autophagy through this pathway. So,
all these agents, caloric restriction mimetics, have different molecular
targets, but it activate autophagy by a final common pathway. The protein
acetylation seems like that. Yes, exactly. Okay. So, with some of the
major ones that you’ve worked with, spermidine. I’ve read quite a bit about
spermidine. I know it’s found in high concentrations in natto, the Japanese
fermented soybean that doesn’t taste like great. But I’ve seen studies about aging,
you know, giving it to even aging mice or something, can then extend their lifespan.
Is that true? So spermidine, to come to
the source of spermidine is contained in the nuclei of all kind of cells. So, in
the nucleoid of bacteria but also the nuclei from yeast cells or from
plant cells or animal cells. So, all food items that contain nuclei cells,
are actually containing spermidine. Also, there are large variations in
the content. So we have to know, on this spermidine, it also is volatile
and accounts for the smell of sperm. So, it is frequently found in food items
that have some kind of smell like natto or durian fruit or fermented cheese, when it
is generated from non pasteurized sources and very rich in bacteria and fungi that
are contributing to the fermentation process, which is, of course, smelly
cheese. And it’s also quite abundant in some vegetables and food where the scent
is more agreeable to most people because it is complex to other molecules that
reduce its volatility. So spermidine has the capacity to induce autophagy
when it is taken up with fruit or with the drinking water when we take mice.
It can also be injected, of course. It is produced by our
microbiota. So, one-third of the spermidine in our body is probably
produced in the intestine, and you can manipulate a microbiome to increase
its production of polyamines, including spermidine. Through what?
Probiotics? Or through… Yes. So you know what
strains of bacteria… ? Yeah, there is a Japanese
group that has been publishing that specific bacteria overproducing polyamines
can be used to reduce the development of colon cancer or to reduce aging. Wow, fascinating. That’s very
interesting. And you’ve shown with the spermidine, I know we have a limited time
here, with the spermidine that’s been shown to… Was it spermidine or
hydroxycitric, I think, that was shown to synergize with, like you were mentioning
before, the chemotherapeutic… Yeah,
both of them actually. Both? Okay. So the mechanism is that
when you combine chemotherapy with caloric restriction mimetics, all the caloric
restriction mimetics that I mentioned, including spermidine and hydroxycitrate
and resveratrol, will enhance the anti-cancer immune response that
makes the therapy durable. So, we have been able to show that inhibition
of autophagy in the malignant cells or destruction of the extracellular ATP that
is released as a result of autophagy is sufficient to abolish the favorable
interaction between caloric restriction mimetics and chemotherapy. And similarly,
actually, it is sufficient to remove T-cells from the system. And you will lose
any kind of tumor growth reduction induced by chemotherapy combined with caloric
restriction mimetics as it proves that the cellular immune response is actually
decisive for therapeutic outcome. Wow, that’s really quite
promising, I think, for you know at least in the clinic, if you can somehow test
whether or not this caloric restriction or fasting mimetics work in conjunction
with some of these immunotherapies, that would be fantastic. But I want to ask
you one last thing too about some of these fasting mimetics. Like, if I were to
just supplement with hydroxycitric, for example, or if there were spermidine
supplement, and I was still eating a normal, you know, healthy diet, but not
caloric restricted and not fasting. Do you think that would be
sufficient to induce autophagy? Well, I can
respond for mice cell… Okay.
How about in mice? …that this is certainly
the case. I don’t know about humans because we have no clinical
studies in this field. So in mice, it does? It does. And in mice,
you actually can give a combination of high-fat diets, that usually would cause
obesity, with spermidine to reduce weight gain through mechanisms that we don’t
understand and that we believe to be autophagy-dependent but have to
elucidate in some molecular detail. That’s very cool. So I’m going
to look up those strains of bacteria, you know, to see that can I eat fermented
foods to increase that population and get more spermidine. You know, things like
that would be very interesting to me as useful little tools. Do you have
any practices that you do? Do you, for example, do intermittent
fasting or any type of fasting or time-restricted eating? So, usually in my
ordinary life, when I’m in Paris and working, I only have one meal per day.
So I have dinner with cheese and wine containing spermidine resveratrol. It’s an
ideal combination because inhibiting the acetyltransferase and activating the
deacetylase will have, of course, a synergistic effect.
Oh, you do? Well, if
you can show this in mice and this is an excuse for me to… Yeah, wine and
cheese go great together. …profit from a banquet
combination. And I do also practice some fasting twice per year also when I
don’t eat anything for five days. Oh, wow. So you do
a prolonged fast twice a year. Well, it’s not so
prolonged, so you know that Gandhi, for instance, has been doing fasting
exercises for 20 days or more, which is the time at which a normal
individual could have some long-term consequences on the health. So, 20 days
is some period that usually is could be easily supported by a healthy
middle-aged individual that has no underlying pathologies. I have a couple of friends
that have done. One of them is used to very prolonged fast, like 20 days. He’s
morbidly obese, and he’s lost now like 200 pounds over the course of a year by doing
just several rounds of these prolonged fast. But he’s got a lot of fat, you know,
to supply energy. I’m not sure I would still subject myself to a 20-day water
fast, but it’s cool to know that you do these five-day water fast, twice a
year, and eating one meal a day. Yeah,
and doing exercise. And doing exercise.
And when you’re eating… Sorry, throughout the day, when you’re
not eating, do you drink coffee? Yes. And there’s polyphenols
in coffee that actually… Yes, we published a study
in mice giving them a non-toxic dose of caffeinated or decaffeinated coffee
with a drinking water continuously. And we could show that this was
magnificently inducing autophagy. Like caffeine
independent fasting. Right, totally independent
caffeine, so just the polyphenols. And this was like, I think, I read your
study, it was like after four hours or something in the mice. So, in
humans, potentially, maybe the fasting plus the coffee… Well, coffee abuse has
been linked to major health-promoting effects. So, most cardiovascular and
neurodegenerative diseases are actually reduced in heavy coffee drinkers as an
independent link between lifestyle and pathology. So, of course, it’s an
association that obviously can be criticized because it’s just a study in
which you find a statistical correlation. It would be much more interesting
to do a randomized clinical trial on coffee intake. More interesting
and more expensive, yeah. Yeah.
Absolutely. Great. So fasting,
coffee, wine and cheese, one meal, exercise, you’re doing it all.
Awesome. Well, really enjoyed this conversation,
Guido. If people want to find you, you have a lab website, kroemerlab.com.
That’s K-R-O-E-M-E-R lab, L-A-B .com. So, thank you again for this wonderful,
very illuminating conversation. It’s a pleasure. I hope you enjoyed
learning about all things autophagy, both macro and micro from Dr. Guido
Kroemer. Dr. Kroemer in my mind is the consummate expert in this field, and also
is amazingly prolific with somewhere in the neighborhood of a thousand
publications to his name. Quite the feat to say the least, and all of the more
reason why it was an enormous privilege to get to talk to him. Do you love these geeky, deep biology
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100 Replies to “Dr. Guido Kroemer on Autophagy, Caloric Restriction Mimetics, Fasting & Protein Acetylation”

    • 00:03:22 – How genetically inhibiting autophagy actually increases cellular sensitivity to death and how this was counterintuitive because autophagy was thought, at one time, as a type of activity predominantly concerned with actually initiating cell death.
    • 00:04:44 – The external signals causing autophagy.
    • 00:05:29 – The role of growth factors in transporting nutrients from the outside world into the intracellular space.
    • 00:05:48 – Autophagy as a process that destroys bioenergetic macromolecular reserves including proteins, lipids, and nucleic acids to generate energy.
    • 00:06:28 – How declines in the ATP (energetic) status of the cell trigger autophagy by increasing the activity of a pathway known as AMP Kinase.
    • 00:06:36 – How reductions in the cytosolic pool of acetyl-CoA as a consequence of reductions in glycolysis, amino acid catabolism, or beta-oxidation, ultimately result in the de-acetylation of hundreds of cellular proteins involved in autophagy. Nutrient deprivation → ↑ Protein Deacetylation (↓ cytosolic Acetyl CoA) + ↓ mTOR + ↑ AMP Kinase → Autophagy
    • 00:09:37 – The role of the inhibition of mTOR and activation of AMP Kinase in cellular autophagy. mTOR is a pathway robustly activated by IGF-1 and associated with increases in cell growth, proliferation, motility and protein synthesis. AMP Kinase, on the other hand, is a pathway important in regulating cellular energy homeostasis by inhibiting synthesis of fatty acids and triglycerides and activating fatty acid uptake and beta-oxidation in the liver.
    • 00:09:37 – The indirect relationship between protein acetylation status (and cytosolic acetyl CoA availability) and the suppression of mTOR and activation of AMP Kinase in cellular autophagy.
    • 00:10:23 – The possibility of using selective nutrient restriction as an autophagy-inducer instead of more generalized nutrient deprivation or fasting.
    • 00:11:11 – Cytoplasmic protein deacetylation as a potential surrogate marker for fasting-induced autophagy (still undergoing validation) but possibly not other forms of autophagy, such as the kind pharmacologically induced by interaction with a protein involved in vesicle-trafficking processes called Beclin 1.
    • 00:12:53 – How a protein called LC3 associate with structures called autophagosomes to facilitate autophagy in response to deacetylation it undergoes. Note: this sirt1-mediated deacetylation of LC3 is induced as an important response to cell starvation.
    • 00:14:22 – The special flow cytometry needed in order to measure some of the proteins associated with the activation of autophagosomes in autophagy.
    • 00:15:04 – The desirability of being able to know and test whether or not your fasting is triggering robust autophagy or not.
    • 00:16:30 – The minimum amount of fasting necessary to activate autophagy.
    • 00:16:56 – Whether or not time-restricted eating or 16:8 intermittent fasting reliably induces autophagy in any of our tissues.
    • 00:17:57 – The important differences between prolonged fasting in humans and the rodent animal models used in studies.
    • 00:19:03 – The ~12-hour half-life of IGF-1 and whether or not that has any implications for the potential of autophagy in shorter duration fasts.
    • 00:20:12 – The effects of long-term caloric restriction on markers of cellular autophagy in humans.
    • 00:21:32 – The effect of an every-other-day eating pattern in rodents and how this pattern actually mimics the longevity producing effects of caloric restriction but with the advantage of stabilizing long-term at a more normal body weight. (Note: Dr. Kroemer is quick to point out that this type of intermittent fasting is more dramatic than it would be in humans because of differences in metabolism… the animals actually experience a 10% oscillation in body weight from this pattern!)
    • 00:24:35 – The effect of exercise (especially endurance exercise) on autophagy in muscle tissue.
    • 00:25:08 – The role of autophagy as a mediator of the anti-obesity and anti-diabetic effects of endurance exercise.
    • 00:26:58 – The differences between macroautophagy, microautophagy, and chaperone-mediated autophagy.
    • 00:28:03 – The difference between autophagy that is dictated by demand (nutrient stress) versus autophagy that occurs as the need arises to recycle damaged organelles.
    • 00:28:40 – How damaged organelles change the composition of their surfaces in order to decorate them with signals for the stimulation of their engulfment by the autophagosome.
    • 00:29:07 – The many, many names autophagy has when we are talking about it in the context of specific macromolecular structures and organelles (e.g. mitophagy for mitochondria, pexophagy for peroxisomes, reticulophagy for endoplasmic reticulum, ribophagy for ribosomes, and virophagy for viruses).
    • 00:29:35 – How autophagy from nutrient deprivation still prefers to first recycle organelles that have been slightly marked… in other words, that are already aged or slightly damaged.
    • 00:31:15 – How mitochondrial use ubiquitination, a process which occurs when these organelles begin to lose their membrane potential, in order to signal damage and to ensure preferential targeting by the autophagic machinery. ↓ mitochondrial transmembrane potential → ↑ ubiquitination → ↑ mitophagy
    • 00:32:42 – The coordinated manner in which mitophagy and mitochondrial biogenesis act together in a closed feedback loop in order to preserve mitochondrial quality while preserving the total pool of functioning mitochondria.
    • 00:33:34 – The changes that can occur in total mitochondrial pool as cells adapt to take on new metabolic profiles through a mitophagy-mediated process. This can occur as cells differentiate into new cell types that are specialized for glycolytic energy generation.
    • 00:35:13 – The role autophagy plays in the prevention of neurodegenerative diseases caused by protein aggregates.
    • 00:37:18 – How autosomal recessive mutations (where both parents must contribute a defective gene for PD to arise in the offspring) in a kinase protein called PINK1 disrupts its ability to recruit a protein called Parkin that mediates the targeting of mitochondria for mitophagy. ↓ PINK1’s kinase activity → ↓ parkin recruitment → ↓ mitophagy → ↑ accumulation of damaged mitochondria
    • 00:38:01 – Mitochondrial membrane permeabilization as a death signal due to the release of dangerous proteins contained in the mitochondria.
    • 00:39:05 – Dr. Valter Longo’s research demonstrating a cycle of prolonged fasting followed by a refeeding can recycle 28% of the white blood cells, and reduce the severity of an autoimmune disease (multiple sclerosis).
    • 00:39:52 – The evolutionarily-conserved sickness response (food avoidance) as a way of beneficially altering immune responses through through altered metabolism and autophagy.
    • 00:41:02 – The surprising increase in lethality that happens when mice are force fed (glucose in this case) while exhibiting sickness response from a bacterial challenge.
    • 00:42:27 – The effects of an anti-malarial known as chloroquine which has some cytotoxic effects in cancer cells, but is actually (surprisingly) an inhibitor of autophagy.
    • 00:43:12 – Studies of the lysosomal disrupter and anti-malarial chloroquine in combination with chemotherapy in the treatment of cancer.
    • 00:44:04 – The differential roles autophagy plays in the progression of cancer in pre-malignant cells, when it is suppressed, versus in malignant cells, when it is sometimes used as an adaptation helpful to the survival of malignant cells experiencing environmental stress. ↓ tumor suppressor gene activity → ↓ autophagy → survival of pre-malignant cells → ↑ autophagy as a malignant adaptation
    • 00:44:49 – How inhibition of autophagy by itself is sufficient to induce oncogenesis, particularly in leukemia.
    • 00:48:10 – How cells undergoing autophagy can release ATP into the extracellular space where it can function as a signal that recruits and activates immune cells against tumor antigens through the activities of purinergic receptors. ↑ autophagy → ↑ extracellular ATP → activation of purinergic receptor-mediated immunosurveillance
    • 00:48:49 – The importance of the triggering of this immunosurveilance system as part of the cell death associated with chemotherapy.
    • 00:50:27 – The class of compounds known as “caloric restriction mimetics” that affect autophagy by perturbing various pathways in such a way as to reductions in cytosolic acetyl CoA and protein deacetylation in a manner similar to nutrient deprivation. Examples include: hydroxycitrate (inhibits ATP citrate lyase), spermidine (inhibits Ep300, a protein acetyltransferase), and resveratrol (activates deacetylases).
    • 00:55:27 – How intestinal bacteria may produce as much as 1/3rd of the body’s spermidine and how this production rate can be manipulated by probiotic and dietary interventions.
    • 00:56:17 – How the ability of caloric restriction mimetics (CRMs) to induce a type of autophagy that provokes immunosurveillance potentially offers an opportunity for synergy when used in combination with chemotherapeutic agents in the treatment of cancer.
    • 00:56:56 – How the inhibition of autophagy in malignant cells or destruction of the extracellular ATP released by cells undergoing autophagy is able to abolish the favorable interaction between caloric restriction mimetic drugs and chemotherapy.
    • 00:58:02 – The tendency for dietary consumption of caloric restriction mimetics, particularly hydroxy citrate, to induce autophagy and reduce weight gain in mice fed an obesogenic diet.
    • 00:59:17 – Dr. Kroemer’s personal intermittent and prolonged fasting practices and his habit of consuming foods that may contain some natural quantities of caloric restriction mimicking compounds.
    • 01:01:40 – The autophagy-enhancing effect of coffee (with and without caffeine).

  2. 21:26 Autophagy activated at 6 years of 30% calorie restriction…I thought autophagy was a regular thing for any healthy person to clean up the body.

  3. Interesting, he's also eating once a day – just like Dr. Nun Amen Ra who claims to strongly induce autophagy by fasting 23h every day. I'm on his regiment for about 3 months now, but only recently with caloric restriction and I do notice several positive effects.

  4. I really appreciate your interview technique – you let the guest talk, and then summarize and help expand the guest's points. Thank you!

  5. Could it be that in humans, when you actually feel hungry that autophagy is occurring. In people of course who are already slim and have done fasting so are used to it..

  6. I am learning a lot, I fast 40 hrs and eat 8 hrs, and clinical signs are all good and is backed by real science, not pseudoscience that is very common in internet

  7. at minute 7 the text says „low amino acid levels activate mTOR“ – which is wrong. it should read „inhibit“. of course, dr patrick knows this …… and stated it correctly in the interview

  8. Not sure if this was asked before, but have you changed your point of view on Coffee and fasting and autophagy? I am curious!

  9. so i guess when i comes to introduce autophagy as fast as possible it is perfect to be on a ketogenic diet, where the glyogen fuel is already cut off. so i guess it is just down to the aminoacid pool to get empty so that the autophagy starts.
    in january i did one meal a day on keto (being on keto for a year) and i lost 1kg of weight per week without being hungry and i think a also got a little bit younger. but in february i boosted it by sometimes eating every other day or even every third day and sometimes also doing a dry fast for two or three days. i don't do much exercise but my running performance significantly improved in that month (without doing much exercise in between!)
    i do not plan to eat more then one meal a day for the next half year. and i hope i can do more three day (dry) fasts. the outcome is fantastic. my skin gets much better and my energy level also gets better.
    well i have to do that anyway because i lost 25kg of weight in the last year, completely reversed my diabetes typ 2 and i still need to again lose 25kg to get to my ideal goal weight and have nice 6-pack abs.
    and i guess the autophagy is a good way of tightening my skin. i think i am on the right track here, because i would just do carbohydrate calorie restriction with three meals a day i would be hungry and therefor angry all day and at the end i would look like an empty old bag instead of a fresh tight young boy 🙂

  10. 52:43 – so if I've got this right… If I stop eating for 2 weeks, run a marathon & then neck a celebratory bottle of red at the finish line then my autophagy levels will reach heights never before seen & I'll live forever?

  11. It's fantastic and I couldn't be more thankful that you,special person, share with the whole community such infinitetly valuable informations. It's remarcable that you share whole interviews full of awesome infos,my deepest regards for all your work and for sharing it with us. I wish you all the best!

  12. Yes, protein depletion has been tested on humans and protein deprivation has been practising for many years, but for the past 25 years in very large numbers at the sanatoriums of Dr Emilova in Bulgaria. Hundreds of thousands of people have been there fasting under doctors supervision and the data shows improving the health of all of them. You say here, we don't know. Yes, we do know. Its documented. And fasting could be done not only for more than 24 hours, but as long as 30 days eating fruits only. My own experience from it is also amazing. Of course fasting should always be done under doctors supervision as some conditions could get worse.

  13. What about practising both intermittent fasting and caloric restriction? E.g., consuming under two thousand calories in a four hour window? Would there be multiplicative benefit or am I wasting my time?

  14. Rare to find a raw , unfiltered podcast that focuses on the science , without dumbing it down to pander to the audience. I didn't understand a lot of the cellular mechanisms entirely , but the numerous papers being cited, motivate me enough to find out. Dr Guido seems like on of the most knowledgeable persons I've seen on your podcast, its impressive to hear the breadth of his knowledge.

    P.S Spermidine in cheese in synergy with resveratrol in wine , would be interesting to see if this is a reason behind the French longevity paradox.

  15. Legendary as always, keep up the awesome work Rhonda! Really appreciate your non biased, scientific approach to the information you share. You seem to have constant access to the latest scientific evidence. An idea I randomly just had was to have a "literature review webpage" of sorts, whereby you post 100 words or less (for example) on the strongest scientific evidence for various topics such as autophagy, fasting, caloric intake, CRM's, etc with a focus on what the current optimal approach looks like based on the latest evidence. It would be amazing to have a constantly updated reference guide (maybe every 3-6 months an update) where people can go to see an unbiased literature review essentially. But a lot shorter than a typical review. Hope this is an interesting concept for you. Cheers from NZ! Rock on 🙂

  16. I wish polyamines such as putrescine, spermine and spermidine were available as nutritional supplements. It's odd that they're not. I've seen spermine advertised on Amazon, but it's as a laboratory product, not a product intended for human consumption.

  17. Thank you for such an insightful interview and information!
    Durian was mentioned. Just curious how this "king of fruit" can influence our health. I know it's very complex and rich food but that agent producing strong smell as I've heard in this interview could have some benefits (or not). I'm sorry that I didn't get it clear from this video.

  18. The 'M' in MToR is 'Mammalian'; it accounts for insulin and such in critters with complex cells, as opposed to yeast and such.

  19. Fantastic interview, even more remarkable when you think that English is the second language from Dr Kroemer! He speaks better English than most of us native speakers.
    I find that the people with the most knowledge and intelligence are very calm and don't let their emotions get involved with their answers.
    I didn't understand everything he was talking about but bit by bit, I am getting very interested in fasting and autophagy. When a man like Dr.Kroemer is on OMAD plus prolonged fasting, not to mention wine and cheese! That is good enough for me
    Thanks and keep them coming!

  20. There is a type of natto that tastes perfectly fine, even good; it is black soybean natto. I don't know whether it is nutritionally different from regular natto; I'd be curious to know. It is the same mould that makes it, so probably it has nattokinase like normal natto.

  21. The treatment of animals in experiments is just so wrong, thankfully we have organisations like Animal Free Research UK changing things. Mice eating each other because they are so hungry will be a thing of the past soon, and every cage will be empty.

  22. So….we now know…that beyond the Lysosome…that we've known about for 30 years…that we have an endometabolic mechanism to "clean ourselves out; intracellularly for our recapturing of said proteins/amino acids for the resequestering of our own reuse.

    This explains alot,….as related to our ability to fast for many days, weeks,…and even months when "food" is not available..or for our own 'desire' to readapt our inherent cellular mechanism back what may be stated as a "younger state". But…

    There's "no gain without pain" for most people. This appears to essentially be a good way to move into better health by individuals with the proper "notion, and then motivation and discipline". In other words….."go into a FASTing state,…and letting the bodies do their' work to clean, rebuild other "sound cells", and move onto a higher state of functioning.

    Finally, this pathway,….totally understood or not….appears to be of major interest to all athletes,….individuals trying to lose weight…or just get "reset to a better health standing"…without drugs or medical intervention; the costs, toxic side effects,…..time waste, or arguments with other medical individuals who have a vested interest into "the economic realities of massive drug interventions that deal only with symtomologies alone…and do nothing to reset our cell function".

    Many drugs will become obsolete with this realization!

  23. How intelligent is this guy! Wow, a knockout interview. Also, I can tell you took the time to edit the closed captioning, which isn't easy or quick. Wanted to express my appreciation for that, because having captions on helps me concentrate and retain information. Thank you for producing all these high quality, informative videos.

  24. Another GREAT video, thank you! Could you please keep us up to date on your research on probiotics and Spermidine? https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0023652

  25. Wow ..that was an amazing interview. I think I'm going to have to listen a few more times to absorb all this great information.

  26. IT is funny to me when yu were interviewing dr. Longo who actually could answer your questions on when autophagy began, you did not listen to him and now you are asking a man who has not done that perticular research. You also ask him if he knew what stimulated the onset of autphagy. Dr. Longo answered those quetions. You might want t watch his videos when reviewed by people who let him talk more. HE said that in order for the autophagy to begin first the liver must mark the cells to be eaten which occurs after all glucose has been depleted. Once all glucose is removed which typically occurs after day 2 then on day 3 the liver marks the cells as keepers or eaters… then when all of this is marked cells is completed the body begins to eat both fat and the marked cells.

  27. 26:20. Interesting you ask about the Exercise while Fasting. I have been doing 16 hour fasting and find it great for energy (Mental) and weight loss (Fat). Personally I find it really efficient to exercise at the 14th hour for about 30-45 minute run and then eat around the 15-16 hr to break the fast. I no longer have a 3pm low energy and have been loosing weight this way. 🙂

  28. I don't get how everyone's talking about caloric restriction and fasting, but never goes into how some of us should SURVIVE a lifestyle like that. If I restrict my caloric intake by even 25%, I'm gonna lose weight and eventually… well, die?!

  29. Rhonda, I haven't heard you speak yet about the carnivore diet. I am doing carnivore plus warrior. I call it wolverine diet. What is your take on it?

  30. Dr. Kroemer is a well established scientist and also has published in purinergic signaling in HIV among many other topics. It is good to see him after reading many of his papers. Thank you for posting this video. Marc C. E. Wagner http://www.marccewagner.com/

  31. Excellent information! I would like to point out that the Resveratrol is found in 🍇 grapeskins, which is why it shows up in red, not white, wine 🍷, thus, the risks of alcohol may be avoided. Carry on! 🐶

  32. I listened twice and rewound, several complex sections and was still left a bit confused at the end. But in another Dr. Patrick video, they noted even black coffee starts metabolic processing, so it isn't truly fasting. Anyone comment on this distinction please?

  33. My thinking is that Dr. Patrick is pretty humble.
    I feel fortunate every time I watch one of her videos
    on whatever subject I have ‘searched out.’
    She’s like a modern-day Leonardo DV that we
    get to watch on you.tube🇺🇸

  34. Nicely done, even though most has gone over my uneducated head love how you add illustration and definitions et al.Having to educate myself having been recently diagnosed with cll and given NO advice on exercise, diet,just wait and see is the standard. Keep up your good work.Thanx

  35. Its very difficult for me to cognitively induce or restrict mitosis on trillions of my cells. How they heck am I to now induce all the different types of autophagy? I won't have time to eat anything, now or later. By the time I get this figured out, the next phase of published research will contradict all this.
    Call me when they make a pill

  36. I consider myself very fortunate to be living in an age when this type of information is so readily available to the common man.

  37. Indeed a very enlightening explapnation of very complex subject. now I begin to understand why in some occasions we tend lose appetite or avoid foods when we are not well.

  38. This was an amazing presentation!! It really helped connect the dots on how these mechanisms can dictate the outcomes in health and disease. I would certainly be interested in your thoughts on his personal regimen of prolonged fasting, exercise and diet. You are doing a great service to all of us.

  39. Top 10, or even 5 here Rhonda, this was an exceptional interview! There is beauty in science, you know how to waltz with intellects and match them step for step. There were so many "connect the dots" moments for me in this one, I'm sharing it far and wide, and coming back for the encore. Bravo, and many thanks to Guido for coming on, he clearly appreciated your considerate, intelligent interview style. So grateful for your work, thank you.

  40. I found this interview extremely interesting, esp., about the importance of autophagy in using the immune system in the long term suppression of a cancer.

  41. No amount of fasting will make this woman smarter. Concentrate on breeding and do no tire your brain with science.

  42. Hi Dr Patrick

    I was wondering what you think of the 'Snake Diet' which includes an intermittent fasting lifestyle. Is it safe to drink water with these ingredients while fasting? Will true autophagy occur? The creator of the diet says you lose too many electrolytes while fasting for longer periods like 48,72, or more hours. He says you need to replace lost salts and potassium while fasting or you will feel shaky.


    Per 2 Liter H20:
    (Evian bottled water recommended because of PH)

    *drink less than 2L per day of the bottled water mixture*

    Potassium Chloride Powder = 2 tsp
    (4700 mg) —(No Salt brand or Windsor salt free)
    Himalayan Pink Salt = 1 tsp
    (2,000 mg/L)
    Organic lemon Juice – 8 Tbs per day
    Organic apple cider vinegar – 4 Tbs per day

  43. in her new book how to starve cancer jane maclland says autophagy can be a fuel for cancer and needs to be stopped as it acts as fuel for the cancer cell at certain  periods of the process. The cancer cell uses autophagy when arginine  is deprived it used glutimine

  44. I had a hip replacement surgery 18 days ago and i maby got an infection in the SCAR / wond . I was looking for information about fasting and bacteria infection , and this was realy helpfull , i did not feel i wantedto eat anything and i gues that is a sign that i do have an infection !? I did get a antibiotika just in case and i am on it right now, i gues it willalso help to fast a feew days on top of that … thank you for a great video , i just love this , so to the point 🙂

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