1028 replies to this topic

[Castiel]

I wonder why he takes it in the morning with yogurt, and 'breaks' his fast.  Than never made sense to me.

 

What I don't know if Sinclair knows, is that milk at least reduces absorption of chocolate, tea, coffee, and potentially other beneficial plant compounds absorption, iirc.   While fat may aid absorption it appears some proteins, probably casein, designed to interfere with plant substance absorption to protect developing organisms taking milk from external plant compounds.    Now I don't know to what extent this affects yogurt, as the bacteria might have degraded some of the problematic compounds in milk.  

 

I think it is best to take resveratrol with extra virgin olive oil or coconut fat or grass fed butter.    As for fast, I've read of Valter Longo's research on fast mimicking diets.     From what I gather MTOR mostly activated by aminos, particularly many of the amino types found in animal based protein sources.    A low calorie low plant based protein diet, is essentially practically equivalent to a fast, while being far safer, and probably able to be handled for longer terms but that remains to be tested.    There are many benefits to low protein low calorie diet over normal fasting, you can take high fiber which has benefits, you can also take and easily absorb micronutrients and not get into vitamin, mineral or electrolyte deficiencies.   Also many plant compounds, such as coffee, matcha, amla, black elderberry, are godlike super compounds you can be ingesting in such diets.    As for aminos, you can take collagen along with plant based protein sources, as that is glycine rich source that helps clear away methionine, it also helps the body build collagen.   And  IIRC glycine one of the critical components needed to build glutathione.

 

 

Moreover, supplemental glycine has been reported to increase tissue glutathione levels in several animal studies. Dietary Glycine Is Rate-Limiting for Glutathione Synthesis and May Have Broad Potential for Health Protection (nih.gov)

[QuestforLife]

Finding the culprit: the hormones required for sexual maturity may be the trigger that starts aging via downregulation of TET2

 

 

The above figure by Blagosklonny is highly relevant to our discussion of hormones, (methylation) and aging.

He describes the figure as follows:

Why men age faster but reproduce longer than women: mTOR and evolutionary perspectives

by Blagosklonny

doi: 10.18632/aging.100149

The menstrual cycle is regulated by interplay of negative and positive feedback loops. The hypothalamus stimulates the pituitary gland to secrete Follicle-Stimulating Hormone (FSH), which in turn stimulates follicles in the ovaries. Follicles maturate and secrete estrogens. Estrogens inhibit the hypothalamus, decreasing secretion of FSH (a negative feedback loop). In turn, FSH stimulates ovarian follicles, which produce estrogens, which in turn inhibit FSH production. Also, estrogens stimulate secretion of Lutenizing Hormone (LH). LH in turn causes ovulation. So for the normal menstrual cycle, the hypothalamus should have a narrow range of sensitivity to estrogens. Both too high and too low sensitivities are not compatible with menstrual cycles. In comparison, regulation of reproduction in men is simpler. There is a gradual decrease in fertility in men too (analogous to pre-menopause), although this usually does not result in testicular failure during a man's lifetime.

 

 

You could argue, and I would, that the real cause of aging in this scenario is the limited number of activatable follicles and egg cells therein. And this links aging back to the telomeres of oocytes, which are finite. Indeed, we see numerous issues in older mother relate to short telomeres in eggs (can discuss another time as it is a big subject).

But as we know the same thing occurs with men (albeit slower), with supposedly immortal spermatozoa. I have discussed this before, in post #474, and I won’t go any further into it here, other than to say aging also affects the male reproduction system in a way compatible with the idea that sperm are failing to differentiate properly.

From Post #474: Spermatogonial stem cells are already immortal, but another paper [10] shows that a similar process is occurring (although there is no evidence at this time that this is mediated by increased methylation); with age sperm cell progenitors reduce mitochondria, increase glycolysis and self-renew at the cost of differentiation and testicular atrophy (ouch!). [10] source: www.pnas.org/cgi/doi/10.1073/pnas.1904980116

 

How does this all link up to hormones and aging? It is possible that the feedback mechanisms that time and trigger puberty also lead to aging, but on a less well timed basis, as suggested by Blagosklonny with menopause. Blagoskonny clearly explains that an initial small amount of LH and FSH from the hypothalamus and the pituitary gland activate a small amount of estrogen from the follicles, which is sufficient to shut down LH and FSH. Later the hypothalamus and the pituitary get resistant to estrogen and the loop escalates until there is sufficient LH/FSH to release an egg (and menarche begins). The large quantity of estrogen released with the egg knocks LH/FSH right down and the build-up begins again, cumulating a month later with another egg being released. This continues until the eggs all run out, and even though LH/FSH stays high from then on, no eggs are released and the system is broken.

 

Note: there are reports melatonin and NAD+ boosters can restart menarche; this fits with my previous post linking increased oxidative stress to methylation and blocked differentiation (in this case oocytes). There might be some egg cells still remaining after menopause, but they can’t be made to mature whilst oxidative stress is high.  I also do believe there is a hard limit here, but I’ll discuss that on request.

 

Could the aging of the female reproductive system be extended to the whole organism? I believe so. The following paper offers a tantalising hint:

 

 

An epigenetic switch repressing Tet1 in gonadotropes activates the reproductive axis

Source: https://www.pnas.org...nt/114/38/10131

We present an epigenetic switch in the central control of reproduction as a truncated TET1, expressed in proliferating gonadotrope-precursor cells, which inhibits Lhb (=luteinizing hormone) expression and so must be repressed for reproductive development. Expression of this TET1 isoform is regulated by cis-elements mediating effects of gonadal steroids and PKA, and also a potentially methylated distal enhancer. As this isoform appears more common than the canonical TET1 in other differentiated tissues, our study has broader functional implications outside of the reproductive axis. Furthermore, our findings support the idea that distinct genomic regions are used at different developmental stages or in different tissues, and that a particular sequence can be part of the primary transcript in some tissues or an enhancer RNA in others.

 

 

The paper gives more detail: reproductive cells upregulate LH as TET1 is downregulated (as an intentional programmed part of maturity of reproductive cells; note TET1 is normally only expressed in embryonic cells, TET2/3 is expressed other tissues). But they also show other non-reproductive differentiated cells have an isoform of TET1 expressed (an alternatively spliced version), which keeps LH suppressed in those cells. This can be overcome by a range of sex hormones or by methylation of TET2, which keeps the TET1 promoter turned on by removing its methylation.

 

It all comes back to those pesky TETs! Methylation of TET2 not only messes up things like GDF11 expression and oxytocin receptors and (possibly) mitochondrial health, but also causes random tissues in the body to turn on LH. We know LH is an oxidative hormone, and this will cause further downregulation of TET2 and ratchet the feedback loop to more methylation and more LH and less TET2. Where does this all start? Given we need LH (and FSH if you’re female) to reach sexual maturity, it is possible that this LH/FSH ‘leaks’ and gradually turns up methylation across all body tissues. And it is not as if we can do without these hormones; without FSH/LH women would release no eggs, and presumably (I need to look into this (*)) without LH men would not mature sperm cells. The signal to reproduce is also the signal for aging (on a much slower timescale) with women getting a surge of aging with their period each month, turning up to a constant surge of aging after menopause, and men getting a constant aging signal, which likewise ratchets up with LH in late middle age.

 

 

Source: https://doctorlib.in...ology-2/91.html

Note how there is a spike in the gonadotrophins in the fetus; perhaps related to the downregulation of TET1 and the creation of sex organs. The next time the hormones are that high triggers sexual maturity, thereafter levels are constant (averaging the monthly female cycle), until late life increases them yet further.  

 

(*) Early research on male fertility initiation and aging

Gonaldotropins are required for spermarche (start of sperm production), see https://pubmed.ncbi....ih.gov/2492750/
But rising luteinizing hormone also causes loss of quality of sperm in later life, see;
https://bmcurol.biom...894-020-00674-7

Edited by QuestforLife, 19 July 2021 - 12:11 PM.

[Castiel]

Finding the culprit: the hormones required for sexual maturity may be the trigger that starts aging via downregulation of TET2

 

 

The above figure by Blagosklonny is highly relevant to our discussion of hormones, (methylation) and aging.

He describes the figure as follows:

 

 

You could argue, and I would, that the real cause of aging in this scenario is the limited number of activatable follicles and egg cells therein. And this links aging back to the telomeres of oocytes, which are finite. Indeed, we see numerous issues in older mother relate to short telomeres in eggs (can discuss another time as it is a big subject).

But as we know the same thing occurs with men (albeit slower), with supposedly immortal spermatozoa. I have discussed this before, in post #474, and I won’t go any further into it here, other than to say aging also affects the male reproduction system in a way compatible with the idea that sperm are failing to differentiate properly.

 

How does this all link up to hormones and aging? It is possible that the feedback mechanisms that time and trigger puberty also lead to aging, but on a less well timed basis, as suggested by Blagosklonny with menopause. Blagoskonny clearly explains that an initial small amount of LH and FSH from the hypothalamus and the pituitary gland activate a small amount of estrogen from the follicles, which is sufficient to shut down LH and FSH. Later the hypothalamus and the pituitary get resistant to estrogen and the loop escalates until there is sufficient LH/FSH to release an egg (and menarche begins). The large quantity of estrogen released with the egg knocks LH/FSH right down and the build-up begins again, cumulating a month later with another egg being released. This continues until the eggs all run out, and even though LH/FSH stays high from then on, no eggs are released and the system is broken.

 

Note: there are reports melatonin and NAD+ boosters can restart menarche; this fits with my previous post linking increased oxidative stress to methylation and blocked differentiation (in this case oocytes). There might be some egg cells still remaining after menopause, but they can’t be made to mature whilst oxidative stress is high.  I also do believe there is a hard limit here, but I’ll discuss that on request.

 

Could the aging of the female reproductive system be extended to the whole organism? I believe so. The following paper offers a tantalising hint:

 

 

 

 

The paper gives more detail: reproductive cells upregulate LH as TET1 is downregulated (as an intentional programmed part of maturity of reproductive cells; note TET1 is normally only expressed in embryonic cells, TET2/3 is expressed other tissues). But they also show other non-reproductive differentiated cells have an isoform of TET1 expressed (an alternatively spliced version), which keeps LH suppressed in those cells. This can be overcome by a range of sex hormones or by methylation of TET2, which keeps the TET1 promoter turned on by removing its methylation.

 

It all comes back to those pesky TETs! Methylation of TET2 not only messes up things like GDF11 expression and oxytocin receptors and (possibly) mitochondrial health, but also causes random tissues in the body to turn on LH. We know LH is an oxidative hormone, and this will cause further downregulation of TET2 and ratchet the feedback loop to more methylation and more LH and less TET2. Where does this all start? Given we need LH (and FSH if you’re female) to reach sexual maturity, it is possible that this LH/FSH ‘leaks’ and gradually turns up methylation across all body tissues. And it is not as if we can do without these hormones; without FSH/LH women would release no eggs, and presumably (I need to look into this (*)) without LH men would not mature sperm cells. The signal to reproduce is also the signal for aging (on a much slower timescale) with women getting a surge of aging with their period each month, turning up to a constant surge of aging after menopause, and men getting a constant aging signal, which likewise ratchets up with LH in late middle age.

 

 

Source: https://doctorlib.in...ology-2/91.html

Note how there is a spike in the gonadotrophins in the fetus; perhaps related to the downregulation of TET1 and the creation of sex organs. The next time the hormones are that high triggers sexual maturity, thereafter levels are constant (averaging the monthly female cycle), until late life increases them yet further.  

 

(*) Early research on male fertility initiation and aging

Gonaldotropins are required for spermarche (start of sperm production), see https://pubmed.ncbi....ih.gov/2492750/
But rising luteinizing hormone also causes loss of quality of sperm in later life, see;
https://bmcurol.biom...894-020-00674-7

It is still not fully confirmed but high melatonin appears to act as contraceptive in both men and women.   It reduces sperm quality and number, iirc.   And some say resembles signals in animals with temporary reproductive periods during the year, where their reproduction is shutdown for most of the year.

 

Pierpaoli considered Melatonin to be part of if not one of the vital signals of the aging program.   Perhaps the issue with Pierpaoli, is that he wasnt bold enough to megadose.

 

We need more experiments with megadose melatonin, to see what effects it has.  It is a potent antioxidant and activates many other endogenous antioxidants, it also activates nrf2, iirc a master detox antioxidant gene that in turn activates hundreds of protective genes.

 

Melatonin also affects LH FSH levels, iirc, but judging by Bowles comments I think not to a sufficient extent.    Sadly these hormones are not based on cholesterol, so plummeting cholesterol which is dietarily achievable won't affect their levels, or so it'd seem.    Low protein is associated with longer lifespan, low protein would shut down mtor, and might also affect the body at a hormonal level.   Things like melatonin do not seem to keep dropping if the body believes nutrients are scarce, it goes into repair and defense mode.

[QuestforLife]

Crazy hypothesis:
Age-related methylation of differentiation gene promoters is actually protective against stem cell exhaustion (due to telomere loss, secondary to telomerase silencing on differentiation). It is intentional, but not an aging program, it is a self preservation program (the body reducing its
self-replacement rate).

Why have I come up with this now? Because these genes are clearly programmed to turn off. It's either programmed aging or a programmed response to something else. How so? A subset of genes are methylated with age across every species studied (Horvath
pre-print,https://www.biorxiv....1.18.426733v1).Why does this matter? Because the genes are in different places and even on different chromosomes for different species. For example, take the gene LHFPL4, a top result for hypermethylation from Horvath's pan species clock. It's on human chromosome 3, but mouse chromosome 6. Let's take another:Pax2. It's on human chromosome 10 but mouse chromosome 19. Need I go on? That means
this is a targeted, programmed process. If the same locations (but different genes), i.e. near telomeres or near centromeres were getting methylated, then I could imagine a mechanistic, non-programmed explanation. But if that were the case then different types of animals would experience different consequences (aging symptoms). But we know they don't.

And even a Blagoskonny pleiotropic argument that aging is an accidental continuation of thegrowth program doesn't work. Because developmental genes are what is required for growth and they are being turned down, not up, in age.

It's either programmed aging or some other programmed response. Today's hypothesis is that it is a programmed response to telomere loss.
How could we prove or disprove my hypothesis that blocking differentiation is a response to telomere loss? Well there is what appears to be contradictory data. Horvath found telomerase immortalised cells continued aging according to his earlier clocks, driven by the fact they lived long enough to acquire more methylation changes (though weren't impeded in their replication; possibly because culturing selected for cells that could continue to divide in spite of methylation).

On the face of it that seems to contradict my hypothesis. More telomerase should not cause more blocking of differentiation! But I wonder what he would find if Horvath used his latest pan-species clock on the same cells? I predict that in telomerase immortalised cells he would not find the age related methylation of the differentiation gene promoters he found across many
species.

If proven true then it would be very strong evidence that telomeres are the lynchpin of aging. Indeed I argued something very similar with Turnbuckle years ago, when he suggested lengthening telomeres would cause aging because they would prevent cellular replacement from epigenetically younger stem cells. Indeed he and I both found telomerase activators did increase our methylation age. Yet I responded that there were two completely different
scenarios that could lead to such epigenetic aging. The first would be his delayed cellular replacement, which I say is actually an illusion and not aging. The second actually is aging, when old people slow their cellular turnover due to telomere shortening in their stem cell
reserve.

Let me know If you think the idea is crazy or not.

Attached Thumbnails

• 

Edited by QuestforLife, 21 July 2021 - 08:59 AM.

[QuestforLife]

It appears my prediction has been disproved.

Even if not tumorogenic, telomerase immortalised cells still acquire methylation on differentiation gene promoters; this is part of how they maximise proliferation in cell culture.

We used Weinberg’s classical transformation system to study the evolution of DNA methylation and gene expression patterns. Early passage (EP) BJ fibroblast cells were sequentially infected with the human telomerase catalytic subunit (hTERT), the simian virus 40 large T antigen (SV40) and the H-Ras oncoprotein (H-rasV12) to establish the immortalization-transformation lineage and, as control, with empty vectors (EV) for the above. These latter cells also serve, to measure cells entry into senescence.As previously shown, cells expressing hTERT and SV40 large T antigen become immortalized but are not tumorigenic...

...a set of 917 genes with hypermethylated promoters, relative to EP cells, in any of the three transformation replicates (HRAS-Specific Methylated, HSM) are most enriched for a group of normally inducible development and differentiation regulators....

...In contrast to the above genes, the bulk of 491 near-senescent and senescent-specific methylated (SSM) genes are enriched for processes involved in positive regulation of biosynthetic and metabolic processes. The silencing or prevention of induction of these genes may then be related to a slowing of metabolism as cells enter senescence...

...We find that near-senescent cells, while they can be forced to escape senescence to become immortalized, are not only resistant to transformation but continue to harbor the majority of DNA methylation patterns seen in senescent cells. When the same regimen used for achieving tumor progression is applied to the near-senescent cells, with good resultant protein expression of telomerase (Near-Sen-hTERT), SV40 (Near-Sen-SV40), and H-rasV12 (Near-Sen-HRAS), the growth rate of these cells increases with introduction of each of the above genes. Yet, none of these cell populations, even those with the final addition of H-rasV12 (Near-Sen-HRAS), showed anchorage independent growth in soft agar, which is a hallmark of H-rasV12 transformed EP BJ-fibroblasts. Furthermore, at both genome-wide and promoter CpGI regions, the immortalized cells generated from the Near-Sen cells maintain the DNA methylation patterns of the near-senescent and the senescent cells. doi:10.1016/j.ccell.2018.01.008.

My interpretation: cells with downregulated telomerase acquire a methylation pattern that turns down the turnover of biomolecules and metabolism and this state is resistant to tumorgenic transformation, even after immortalisation with telomerase.

Younger cells that are immortalised gradually acquire a methylation pattern that maximised growth and minimised differentiation genes, and this is even stronger with infection with simian virus and with oncogene activation.

Conclusions? Cells with highly active telomerase must be tightly controlled metabolically or they acquire a pre-cancerous, undifferentiated phenotype, as I've outlined before when describing the 'selfish cell'. Differentiated cells lose telomerase so will eventually become senescent. With age it appears we become less differentiated rather than more senescent, as you'd expect.

Based on these (admittedly in vitro) findings, supplying our stem cells with large quantities of telomerase might not be the cure all I've hoped. But using telomerase on cells approaching senescence might well be very useful. This paper also highlights the importance of forcing differentiation as both an anti aging and anti cancer treatment.

So what about all this being programmed aging? These results suggest it is stochastic (random) methylation, the being acted on by selection pressure, I.e. the cells that divide most predominate. And we end up being made of these cells. Whether or not there are other factors in the body, where we have these various effects downregulating demethylases, I don't know...

[Neil R]

Progeria patients have normal epigenetic ages but accelerated telomere shortening. Epigenetic aging is pretty irrelevant and exaggerated in my opinion.

[QuestforLife]

Progeria patients have normal epigenetic ages but accelerated telomere shortening. Epigenetic aging is pretty irrelevant and exaggerated in my opinion.

I tend to agree with you.

Are there eggs left after menopause that could be made to release? Probably. And are there stem cells in the body at large that could be made to differentiate and improve the aged body? Most likely. But isn't the real reason women run out of eggs and the body runs out of stem cells that they have a limited number of possible divisions, with those left at the end the hardest to stimulate?

We are back to the idea that this refusal to differentiate is all self-protective; a preserving of what is left. I just wasn't able to prove it using telomerase immortalised cell culture.

[Castiel]

Progeria patients have normal epigenetic ages but accelerated telomere shortening. Epigenetic aging is pretty irrelevant and exaggerated in my opinion.

 

I've heard iirc that telomerase restores youthful gene expression with the exception of few dozen abnormally expressed genes.  It also rejuvenates cells and the  tissues they produce.

 

All negligible senescence species either have telomerase or alternate means of lengthening telomeres.

 

That said it does seem that changes to gene expression do result from telomerase activation and telomere lengthening.

Edited by Castiel, 23 July 2021 - 10:13 AM.

[QuestforLife]

I've heard iirc that telomerase restores youthful gene expression with the exception of few dozen abnormally expressed genes. It also rejuvenates cells and the tissues they produce.

All negligible senescence species either have telomerase or alternate means of lengthening telomeres.

That said it does seem that changes to gene expression do result from telomerase activation and telomere lengthening.

It's not that telomerase causes gene expression changes that result in more methylation, it is that cell lines that survive for longer will have selection pressure on them. And random methylation on their gene promoters will result in cells with slightly different gene expression profiles. It turns out that cells that self renew and don't differentiate are selected for in this situation (like people who have more babies dominanting the future gene pool). This happens in immortalised cells in a dish, and possibly in the body as well in stem cell niches (like sperm cells, skin, etc.).

That doesn't mean we should abandon telomerase, as human cells are deficient in it leading to shortening telomeres. And my experience is that it has mitochondrial benefits also. But anything we do to help cells survive gives them more chance to become 'selfish'.

So we need to also make sure we can remove methylation on differentiation gene promoters. Some combination of AKG, Vit A/C and GDF11 seems like a good recipe for this.

I attach some pics from https://doi.org/10.1...pnas.1608679113
Showing the effectiveness of Vit A alone and with the antioxidant Vit C in reducing methylation, increasing hydroxy-methylation and increasing the TET demethylases.

Attached Thumbnails

• 
• 

Edited by QuestforLife, 23 July 2021 - 11:14 AM.

[Hebbeh]

https://www.scienced...10723105258.htm

"A research team headed by Jiyue Zhu, a professor in the College of Pharmacy and Pharmaceutical Sciences, recently identified a DNA region known as VNTR2-1 that appears to drive the activity of the telomerase gene, which has been shown to prevent aging in certain types of cells. The study was published in the journal Proceedings of the National Academy of Sciences (PNAS)."

"Zhu said that his team's latest finding that VNTR2-1 helps to drive the activity of the telomerase gene is especially notable because of the type of DNA sequence it represents.

"Almost 50% of our genome consists of repetitive DNA that does not code for protein," Zhu said. "These DNA sequences tend to be considered as 'junk DNA' or dark matters in our genome, and they are difficult to study. Our study describes that one of those units actually has a function in that it enhances the activity of the telomerase gene."

Rest at link

[QuestforLife]

https://www.scienced...10723105258.htm

"A research team headed by Jiyue Zhu, a professor in the College of Pharmacy and Pharmaceutical Sciences, recently identified a DNA region known as VNTR2-1 that appears to drive the activity of the telomerase gene, which has been shown to prevent aging in certain types of cells. The study was published in the journal Proceedings of the National Academy of Sciences (PNAS)."

Polymorphic tandem DNA repeats activate the human telomerase reverse transcriptase gene

https://doi.org/10.1...pnas.2019043118

Telomerase regulation

I haven't been able to get the full text yet, but from what I've been able to read this is an interesting study that advances our understanding of how the telomerase gene activity is turned up or down..

One of the interesting things about the telomerase gene is that if you add it to human cells using gene therapy (as a section of bacterial DNA including HTERT), it behaves quite differently to how the native HTERT gene. What I mean by this is that it is easily turned on. But the native HTERT gene is controlled not only by the sections of DNA near to it (cis), but also by regulatory regions much further away (trans). I believe this is what they mean by helix-curl-helix regulation. This is when another distant section of the DNA helix is curled around and comes close to the gene in question, influencing it's expression. Shay and Wright did lots of work showing that long telomeres looping back to HTERT were one (but not the only) element limiting telomerase expression.

The hTERT gene is ~ 1.2 Mb from the human chromosome 5p end. We observed that when telomeres are long hTERT gene expression is repressed and a probe next to the 5p telomere and the hTERT locus are spatially co-localized. When telomeres are short at least one of the hTERT alleles is spatially separated from the telomere, developing more active histone marks and changes in DNA methylation in the hTERT promoter region. Source: https://doi.org/10.1...iff.2017.11.005

In this recent study they found an element called VNTR2-1 that actually increases telomerase expression. It's a non-coding (doesn't make proteins) region of DNA. People with more repeats of this sequence have higher telomerase expression apparently; I don't know by how much until I read the full paper. It is interesting that people could inherit more or less of a non coding region of DNA and this would possibly influence telomere length. To be honest I really doubt VNTR-2 will be very substantial in terms of telomere length but we'll see. I think mostly our TL is just inherited directly from the average of our parents' sperm and egg, with a set amount added during the embryonic stage when telomerase is turned up to Max. Then various genes and lifestyle factors and our physical size has a smaller impact on individual variation in TL.

Edited by QuestforLife, 25 July 2021 - 08:17 AM.

[QuestforLife]

Summing up the twin evils of aging

This is how the body of all mammals is guaranteed to age:

It either activates telomerase for indefinite cellular renewal and inadvertently selects for pre-cancerous cells that are 'selfish', i.e. would rather self renew than renew the body

OR

It turns off telomerase with differentiation, and condemns it's tissues to eventual atrophy.

Or you can get some combination of the two.

All it took to guarantee mammal aging was to have methylation of gene promoters occur at slightly higher rates than demethylation. This is probably guaranteed by the necessity to mature sex organs, a process than requires downregulation of TETs in gonadotropes (see previous posts).

You can make the clock tick more slowly, but you can't stop it.

Until now. When we can both activate telomerase and increase demethylation to force differentiation.

Looking at the two problems in more detail:

Telomere loss means slower cellular replacement and finally proliferative failure. Slowing down of internal cellular functions, including metabolism as a consequence of telomere shortening, is mediated by programmed and predictable methylation of gene promoters. This is cellular aging.

Stochastic (random) methylation of gene promoters leads to cellular selection for those cells most able to self-replicate, and this has the effect of reducing useful differentiation and replacement of tissues, as more selfish stem cells start to become dominant. As well as contributing to bodywide aging, this is also a driver of cancer.

Addressing these two problems is difficult, but possible.

Addressing cellular survival via telomerase protects telomeres and mitochondria. I believe this will also improve autophagy and waste disposal. The most noticeable effect of increasing telomerase is improved energy and rapid recovery from exercise.

But this then leads to longer surviving cell lines and eventually selection for selfish cells that don't differentiate.

We therefore need to drive back methylation of differentiation gene promoters and there are several ways to do this. When done successfully this leads to improved differentiation into tissues (more rapid and noticeable than via telomerase upregulation) and often noticeable age reversal. For example better skin and faster reactions.

Continued telomerase activation then makes these new improved tissues last longer.

A final possibility is slowing down cellular processes via MTOR inhibition, which reduces the rate of telomere loss and the rate of undesired methylation (both the major aging pathways). But it is unknown how much MTOR can be slowed down before the body encounters serious problems, for example in fighting off common infections.

[kurt9]

I'm wondering if a protocol combining Turbuckle's stem cell/senolytic protocol with something like this one if it could be targeted for the stem cell populations only. It seems to me that such a combination ought to be quite powerful.

[dlewis1453]

Until now. When we can both activate telomerase and increase demethylation to force differentiation.

 

 

Do you anticipate that some type of cycling between telomerase and demethylation protocols will be necessary? Or could we do both simultaneously? 

 

Some compounds, such as GDF11, appear to be effective at both. 

[QuestforLife]

I'm wondering if a protocol combining Turbuckle's stem cell/senolytic protocol with something like this one if it could be targeted for the stem cell populations only. It seems to me that such a combination ought to be quite powerful.

It's possible.

I tried Turnbuckle's mito fission protocol years ago and it caused me to rip a tendon and a ligament, so I'd say it's effective at reducing mitochondrial numbers. This was before he added AKG.

I also tried Turnbuckle's Stem Cell Protocol when he first started that thread. Taking C60 and stearic acid certainly caused fatigue, but it had no effect on my epigenetic age. But I was only 40 at the time; perhaps it is effective for older folks with more depleted stem cell numbers. Again, this is before he added AKG to that protocol.

I mention this because I used AKG from April 2020; I believe I was the first on Longecity to verify it could reduce epigenetic age. I was certainly the first to show you didn't need to use the Rejuvant version (I used Kirkman).

[QuestforLife]

Do you anticipate that some type of cycling between telomerase and demethylation protocols will be necessary? Or could we do both simultaneously?

Some compounds, such as GDF11, appear to be effective at both.

I don't believe you need to cycle between the two treatments. This is based on my recent results showing epigenetic age was not increased when I added TAM-818, when I continued using AKG.

https://www.longecit...-20#entry907470

You might want to take breaks however; AKG seems to have deleterious effects after a while.

I haven't seen further reductions in epigenetic age adding GDF11 to AKG, but I've no doubt it works having experienced some noticeable improvements (see my report on the subject). Ideally I'd like to be able to just occasionally take GDF11 and not need AKG, if testing showed it was as effective at upregulating TET (papers suggest it should be). I'm less convinced GDF11 is a powerful telomerase activator; it only seemed to upregulate TERC not TERT in the paper I reviewed (which might be a good thing if you want to restrict its effects to already telomerase positive cells).

[dlewis1453]

 I'm less convinced GDF11 is a powerful telomerase activator; it only seemed to upregulate TERC not TERT in the paper I reviewed (which might be a good thing if you want to restrict its effects to already telomerase positive cells).

 

You touched on what was to be my next point...the benefits/detriments of increasing telomerase in all cells vs only increasing telomerase in stem cells. 

[dlewis1453]

Stochastic (random) methylation of gene promoters leads to cellular selection for those cells most able to self-replicate, and this has the effect of reducing useful differentiation and replacement of tissues, as more selfish stem cells start to become dominant. As well as contributing to bodywide aging, this is also a driver of cancer.
 

 

In addition to domination of the stem cell niches by selfish stem cells, isn't there also decline in the total number of stem cells in these niches due to increasing inflammation in the body driven by increasing numbers of senescent cells in a viscous cycle?

 

So the degradation of the stem cell niches is two-fold - 1. selfish stem cells take over the stem cell niche leading to loss of differentiation ability, which contributes to build up of senescent cells; and 2. increasing body-wide inflammation driven by senescent cells harms the stem cell niches and leads to a decrease of stem cell number. 

[QuestforLife]

In addition to domination of the stem cell niches by selfish stem cells, isn't there also decline in the total number of stem cells in these niches due to increasing inflammation in the body driven by increasing numbers of senescent cells in a viscous cycle?

So the degradation of the stem cell niches is two-fold - 1. selfish stem cells take over the stem cell niche leading to loss of differentiation ability, which contributes to build up of senescent cells; and 2. increasing body-wide inflammation driven by senescent cells harms the stem cell niches and leads to a decrease of stem cell number.

Both sides of aging (telomeres and methylation of differentiation promoters) can cause senescent cells and it is not clear which primarily drives their accumulation. It may be tissue dependent.

For example (non selfish) stem cells can suffer from short telomeres and this can lead to a reduction in the rate of tissue replacement and the build up of senescent cells. Or selfish stem cells can develop that deprive tissues of reinforcements by not differentiating, again leading to senescent cells.

Note that in either case senolytics force differentiation by deleting senescent cells, but this may not be helpful in the long run, given such differentiation must come from stem cells that either have short telomeres themselves or a pre-cancerous methylation profile.

For the purposes of this protocol,I don't care which cause is primary in senescent cell accumulation. Telomerase and demethylation will solve both. I don't think we need to worry overly about lengthening telomeres in non-stem cells. The main reason for worrying about cells hanging about too long is methylation of promoters leading to a pre cancerous phenotype. And we're dealing with that with demethylators.

[dlewis1453]

The main reason for worrying about cells hanging about too long is methylation of promoters leading to a pre cancerous phenotype. And we're dealing with that with demethylators.

 

Thanks for the thoughtful analysis. Regarding demethylators, my main question is - how many sites need to be demethylated, and when will we know then we have demethylated enough? Certain sites are key, and demethylating those should lead to downstream effects. The primary demethylators I have seen you discuss recently are AKG, GDF11, and rapamycin, and all three are now the subject of a considerable amount of research showing their applicability to extending life. 

 

Some other less well known demethylators are GHK, which seems to primarily effect the skin and respiratory system, and Compound H, which increases Klotho expression. 

 

https://www.research...NA_Repair_Genes

 

https://pubmed.ncbi....h.gov/28657902/

[QuestforLife]

Thanks for the thoughtful analysis. Regarding demethylators, my main question is - how many sites need to be demethylated, and when will we know then we have demethylated enough?

Compound H also seems to work by upregulating demethylation, at least according to your reference, although they didn't specifically look at the TETs.

So I don't think we need to identify other compounds besides the ones we already know about to upregulate demethylation. Once it's upregulated, the various promoters that have been shut down (GDF11, Klotho, Oxytocin Receptors, TET1) will gradually be demethylated and turn back on.

It is also possible to downregulate methylation, but I'm not sure how safe or effective it is in comparison to turning up the TETs.

As for how will we know when enough (demethylation) is enough, I go off side effects. You'll know what I mean when you try this stuff.

[Castiel]

I think eventually we can akin to blood stem cell transplants, transplant genetically modified negligible senescence stem cells with the ability to dissolve scar tissue and replace it with young as well as the ability to regenerate the central nervous system and other organs.   It'd be a cell therapy with genetically modified super cells, able to cleanse molecular garbage and replace existing cells,  in theory sufficiently advanced synthetic biology and they could even use dna information to repair mutations in the nucleus of neurons via temporary fusions.   

[dlewis1453]

So I don't think we need to identify other compounds besides the ones we already know about to upregulate demethylation. Once it's upregulated, the various promoters that have been shut down (GDF11, Klotho, Oxytocin Receptors, TET1) will gradually be demethylated and turn back on.
 

 

Assuming you follow this protocol diligently for decades into the future, how do you expect (or hope) to see your aging (or younging) play out? We cant know for sure, of course, but do you have a theory or a picture in your imagination of what state you could be in in say 20 years? 

[QuestforLife]

I think eventually we can akin to blood stem cell transplants, transplant genetically modified negligible senescence stem cells with the ability to dissolve scar tissue and replace it with young as well as the ability to regenerate the central nervous system and other organs.

But you'd still have to make sure these cells didn't become 'selfish' through selection of random epimutations. They made immortal HSCs in mice before (they knocked out the de Novo methyl transferase DNMT3a; the stem cells' telomeres never shortened probably because they never differentiated). Unfortunately these immortal stem cells never made any blood cells for the mice they were transplanted into.

Dnmt3a loss of function in hematopoietic stem cells (HSCs) skews divisions toward self-renewal at the expense of differentiation. Moreover, DNMT3A mutations can be detected in the blood of aging individuals, indicating that mutant cells outcompete normal HSCs over time. It is important to understand how these mutations provide a competitive advantage to HSCs. Here we show that Dnmt3a-null HSCs can regenerate over at least 12 transplant generations in mice, far exceeding the lifespan of normal HSCs. Doi: 10.1016/j.celrep.2018.03.025

This is an example of how a systemic signal (methylation) can cause aging in the body through paradoxically immortalising a cell. So it might not be possible to cure aging just through super cell transplants. But the bright side is we might not need them.

[QuestforLife]

Assuming you follow this protocol diligently for decades into the future, how do you expect (or hope) to see your aging (or younging) play out? We cant know for sure, of course, but do you have a theory or a picture in your imagination of what state you could be in in say 20 years?

What is new about this protocol is that no one knew before that stopping telomere shortening would throw you on the other horn of aging (selfish cells). And vice versa, that taking a demethylator would stop selection of cells with methylation of important promoters, but would cause eventual stem cell exhaustion via telomere loss. (The exact balance depending on the mammal species in question and how hard you push aging in one direction).

For mice for example, both telomerase gene therapy (https://www.ncbi.nlm...#__ffn_sectitle), or AKG supplementation (https://www.scienced...550413120304174) extends life and healthspan.

How much might be gained by combining both treatments? Or in finding more effective ways of lengthening telomeres and making stem cells differentiate? Only time will tell. But given what we currently have, I expect we should be able to hover around a healthy middle age for a considerably extended duration. I am not sure what will be required to dramatically reverse age back into our twenties. I think we'll need telomerase temporarily elevated to super high levels, which isn't currently possible outside of gene therapy.

[iftikhargul]

A newbie in this discussion and would like to ask a few questions. I am an engineer and not a biologist, so please forgive my understanding if it is not up to a biologists standard. Here is my understanding of this subject matter from this discussion and also from some external readings

 

  1: Telomere length may not be the most important factor as it has been hyped so far as for as the rejuvenation is concerned. So, investment in lengthing the telomeres may have marginal gains and other options may produce better returns for the time being till this aspect of rejuvenation technology can be managed better.

 

  2: Telomerase may have other usefull functions and it's elevation may have better outcome at the risk of cancer. So, once again caution is advised and may again be not worth the trouble untill better options are available in future.

 

Having said that, here are some more items of interest,

 

  3:   "Selfish stem cells" accumulate with age, specially in bone marrow. Their total numbers increase and they are generally descibed as LT-HSC cells in literature. So, with age, we have more stem cells in our bodies than what we had when we were younger.

 

 4: So, what follows from item 3 is that we may not need stem cells from outside source if we could trigger these stem cells somehow to start repair and contribute towards rejuvenation. In other words, immortality may already exist in our bodies but these cells become dormant and can not be activated.

 

 5: In case of item 4, what causes tthem to become useless and getting them back into action may be the best option to imptove lifespan and rejuvenation.. Is the cause epigentic, or defects and waht genes are triggered to stop these cells from further differentiation. May be a holly garilof rejuvenation if it can be cracked.  May be they can be eliminated and only truely functional cells can be allowed to propogate and exist in our bone marrows and other tissues. May be a stemolytic  ( my own word,not literature based word) therapy as well as senolytic therapy will help in this regard.

 

A response will be appreciated.

 

 

[QuestforLife]

Hi iftikhargul

This thread is full of posts with evidence for the importance of telomeres in aging. I am on holiday with only my old smart phone, so I do not propose to summarise that all again here. But on my profile page, or regularly in the thread, I provide Summary posts with links to all the most important posts I and others have made over the years on the subject of telomeres and aging. The most recent 'Summary' was in Post #602.

The popularity of various 'Theories of Aging' waxes and wanes and is often unconnected from the evidence.

If you look up the page a little at Post #642 'Summing up the Twin Evils of Aging' (on 25th July) you can see my take on the interplay between telomere shortening on differentiation, methylation of differentiation gene promoters, and cancer and aging.

As you say, there is evidence 'selfish' stem cells accumulate with age, for no benefit to the body, and my Post and others prior to it explains why this is so.

It is my assertion that telomerase is a useful treatment for aging, as it allows those cells that do differentiate to last longer and go further, using both telomere and mitochondrial mechanisms. This is probably not sufficient against aging on its own however, as over time this allows random methylation of such cells lines to build up and be selected for, resulting in more 'selfish cells'. Hence the requirement for demethylators. This is what allows such 'selfish' cells to become 'good' again, and is behind the success of GDF11 treatment and AKG trials.

But again, making such stem cells differentiate will turn telomerase down, so eventually aging us through telomere shortening. Hence we can't forget about telomeres.

Of course I don't know the exact balance of aging in each species or even each tissue between both these two mechanisms. And short telomeres with age can be due to either mechanism (straight telomere shortening through replication or selfish stem cell accumulation and failure to differentiate). But so far I've found telomerase activation and upregulating the TET enzymes (2+3) to be personally beneficial both subjectively and through various biomarkers in have monitored and explained in the thread.

I will talk about cancer more in the future. I expect this treatment to be cancer preventative too, by forcing differentiation.

Edited by QuestforLife, 07 August 2021 - 07:38 AM.

[iftikhargul]

Hi QuestforLife,

 

Appreciate your response. Enjoy your holidays. Expect to learn more from your posts and need to study more the summary material.  Obviously, I hastily summarised the conclusions thinking that in order of prefrence, the stem cells provide first order benefits while the telomeres provide second order benefits, based on the fact that they do have longer telomeres before they differenciate. Obviously for longevity purpose they are coupled but only a matter of where one should put one,s limited resources in terms of  immediate benefits. Do need to study more of your posts before selecting the protocol, specially if ithe supplements are hard to secure. However, I do believe that non-functional stem cells either need to be removed or corrected because they do limit rejuvenation options and consume resources.  Have fun.  and I will  read your excellent posts in future when you come back.

[QuestforLife]

I'm happy to reply to posts on hols. I just can't put in all the links, cross references etc. that I normally do. One thing I will say,briefly. Having long telomeres in one functional stem cell or more functional stem cells with shorter telomeres amounts to the same thing in terms of the number of useful cells that can be provided to the body. Does that make sense?

This also maps to our treatment choices. Imagine only 10% of your hematopoietic stem cells were functional and capable of differentiating. Then if you provided additional telomerase such that those HSCs could complete an additional division, this would double the number of blood cells they could supply. Or if you reduced the methylation on differentiation gene promoters on those same cells you might double the number of functional HSCs from 10 to 20%. This would have an identical effect to the telomerase treatment in terms of the number of useful cells supplied to the body (a doubling). If you applied both treatments you would X4 the cells you could supply the body. Obviously this is a gross simplification but you get the idea.

Edited by QuestforLife, 07 August 2021 - 08:26 PM.

[iftikhargul]

Yes, it makes great sense with assumptions if one can push 10% functional stem cells to 40% by using appropriate demethylating agents and telomerase and it can have great impact on one,s health in old age ( >60 yrs) . I am also interested in non-functional parasitic selfish stem cells that if eliminated/activated/fixed would create a better working environment for the body. Obviously, it is a different subject matter and not the main subject of your thread and needs it's own thread. Thanks again for clarification.