1033 replies to this topic

[QuestforLife]

The inflammation side effect at the sight of old injury is curious. Old injuries often contain large concentrations of senescent cells. GDF11 does have a senolytic effect, so perhaps the inflammation you feel is related to the senescent cells and the process of rejuvenation.

Have you considered continuing with daily AKG dosing, but reducing your AKG dose to a lower level?

I'm not the first to noticed the inflammatory effects of GDF11; it is part of the TGF-B family after all.

I restarted AKG@900mg + Berberine@500mg /day and after almost 3 weeks break took a small (0.02ng) GDF11 dose this morning. No flare up of the achilles tendon so far.

[Castiel]

It does look promising, especially as Asc2P is cell permeable. The 130uM used does look like a silly concentration - I've seen similar in Vit E studies showing promising anti-senescent effects - but nevertheless I may switch from the ester-Vit C I currently use.

Note Vit C also activates the TET enzymes and therefore helps with demethylation.

See: Reprogramming the Epigenome With Vitamin C

https://doi.org/10.3...cell.2019.00128
 

 Asc2p seems to have benefits, I saw one brand on amazon.   But not sure, how trusty it is.   Also I'll try doing further research to see if there have been animal trials.    Guessing by the fact it probably has been in the market for a while, probably no major short term downsides occur at a frequent enough rate to remove product from store.   But would like to check for animal trials at least or human trials to see if there is longer term proven benefit.

 

Also I wouldn't drop ester c, Ascorbyl palmitate or ester C was shown to strongly inhibit hyaluronidase,  an enzyme that breaks down hyaluronan.   IIRC, such breakdown enzymes increase with age, and are part of the reason we look older, they deplete hyaluronan from tissues most likely.   Other enzymes probably increase breaking down collagen and elastin too.

 

Also there was this study

 

 

International research group finds 17 different genetic interventions that extend life.

Researchers in Moscow and Harvard have been able to identify a group of genes that, in response to a variety of interactions, exhibited activity associated with Longevity. This meant they could be categorised as biomarkers of lifespan extension.

Longevity.Technology: Rather than work on specific interventions, these researchers decided to breakdown the actual molecular processes involved, leading to a better understanding of how Longevity can be achieved on a molecular level.

Research has discovered numerous interventions that can extend the lifespan of a whole variety of organisms from yeast to worms to mammals. These life-extending interventions include chemicals like rapamycin (a drug found originally in Easter Island bacteria), behaviours such as fasting (calorie restriction promotes autophagy, a process whereby the body cleans out and regenerates its cells) or genetic interventions like mutations (when growth hormones are disrupted, lifespan can be extended [1]).

Lead researcher Alexander Tyshkovskiy explained: “In our lab, we subjected mice of different sexes and ages to 8 longevity interventions and analyzed gene expression changes induced by these treatments. Although in general the effects produced by individual treatments turned out to be rather specific, a certain group of genes changed its expression in a similar way in response to different lifespan-extending intervention [2].”

Once the team had established these “longevity signatures,” and determined both general and specific transcriptomic details of lifespan extension, they looked for different interventions in order to identify new “lifespan-extending candidates [3]”

 

 

https://longevity.te...ity-discovered/

 

 

 

Several pharmacological, dietary, and genetic interventions that increase mammalian lifespan are known, but general principles of lifespan extension remain unclear. Here, we performed RNA sequencing (RNA-seq) analyses of mice subjected to 8 longevity interventions. 

We further applied the discovered longevity signatures to identify new lifespan-extending candidates, such as chronic hypoxia, KU-0063794, and ascorbyl-palmitate. 

https://www.scienced...550413119303729

 

They sought genes that were activated in common among 8 lifespan increasing interventions and based on that have found promising new longevity extending interventions, one of which is ascorbyl-palmitate also known as ester C.

 

So it seems both ester C and asc2p might have roles to play.   Unless ascorbyl palmitate was tested for slowing down telomere shortening.

 

 

While we are on it, also found out that white tea inhibits collagenase and elastase.

 

 

White Tea Could Keep You Healthy And Looking Young

 

 

Next time you're making a cup of tea, new research shows it might be wise to opt for a white tea if you want to reduce your risk of cancer, rheumatoid arthritis or even just age-associated wrinkles. Researchers tested the health properties of 21 plant and herb extracts. They discovered all of the plants tested had some potential benefits, but were intrigued to find white tea considerably outperformed all of them.

The researchers were blown away by exactly how well the white tea had performed. “We were testing very small amounts far less than you would find in a drink,” Professor Naughton, one of the country’s leading specialists on inflammation, said. “The early indicators are that white tea reduces the risk of inflammation which is characteristic of rheumatoid arthritis and some cancers as well as wrinkles.”

https://www.scienced...90810085312.htm

 

 

That too me suggests that the combination of white tea with ester C could have strong skin antiaging effects.   As with these 3 you inhibit collagenase elastase and hyaluronidase.

[Castiel]

A search of PubMed using the text phrase "Asc2P telomere" turns up the paper below.  It says that Asc2G (Asc-2-O-alpha-glucoside) is more effective than Asc2P (Asc-2-O-phosphate) in slowing shortening of telomeres.  And most importantly, regular plain old Asc (ascorbate) does not slow shortening of telomeres at all.  Interestingly, a dilute solution of hydrogen peroxide worked better than any form of ascorbate at slowing telomeric shortening, presumably because of a hormetic effect.

 

Interesting.   But I've heard that vitamin c at high enough dose switches from antioxidant to oxidant by production of hydrogen peroxide, some of it breaks down into hydrogen peroxide.

 

 

 

Cancer researchers have homed in on how high-dose vitamin C kills cancer cells. Vitamin C breaks down to generate hydrogen peroxide, which can damage tissue and DNA. The new study shows that tumor cells with low levels of catalase enzyme activity are much less capable of removing hydrogen peroxide than normal cells, and are more susceptible to damage and death when they are exposed to high doses of vitamin C.  Why high-dose vitamin C kills cancer cells: Low levels of catalase enzyme make cancer cells vulnerable to high-dose vitamin C -- ScienceDaily

 

That suggests that higher doses of vitamin c might produce the same as dilute hydrogen peroxide solution.   The question is what would be optimal dose, as you don't want excessive hydrogen peroxide.

 

Also I've tried very very high megadose vitamin c in the past(about 1gram every few hours), and had very strong autoimmune reaction, drugs I was given couldn't deal with it, but I managed to remove it with a megadose of fish oil(several gram omega 3).

 

I'm now at a few grams of vitamin c, and have no issues.    Though sometimes and some people can get a bit of joint pain with at least some forms of vitamin c at doses as low as 500mg.  The joint pain usually goes away when the dose is lowered.

 

Could be the vast anti inflammatory number of plants I eat, that allow me to get away with multigram vitamin c.   Since I suspect the joint pain could be a result of increased immune activation.    I will add ester C to my regimen, and might add asc2p, but need to check, as again only saw one brand, and want some research on how its done in animals as well as brand reputability and if multiple brands are also selling it.

[Castiel]

will say according to a controversial researcher called Bill Sardi some research has suggested that oral doses of some forms of vitamin C can actually reach blood levels seen in cancer killing studies. 

Researchers Achieve Cancer-Killing Effect With Oral-Dose Vitamin C (knowledgeofhealth.com)

 

Another of his articles points that a study suggested iron inhibits vitamin c hydrogen peroxide cancer killing activity, and that iron chelators might be beneficial for such therapies.

 

That said I find Bill Sardi controversial given some of his antiaging theories, and also because at least that website, which may be his seems to have some seemingly conspiratorial takes on a few subjects.   So some of his stuff may be questionable, good to check his sources to verify his claims.

 

here is info from one of the source studies he cites

 

 

Conclusions. Since a single oral dose can produce plasma levels in excess of 400 µm L−1, pharmacokinetic theory suggests that repeated doses could sustain levels well above the formerly assumed maximum. These results have implications for the use of ascorbate, as a nutrient and as a drug. For example, a short in vitro treatment of human Burkitt's lymphoma cells with ascorbate, at 400 µm L−1, has been shown to result in ∼50% cancer cell death. Using frequent oral doses, an equivalent plasma level could be sustained indefinitely. Thus, oral vitamin C has potential for use as a non‐toxic, sustainable, therapeutic agent. Further research into the experimental and therapeutic aspects of high, frequent, oral doses of ascorbic acid either alone or (for cancer therapy) in combination with synergistic substances, such as alpha‐lipoic acid, copper or vitamin K3, is needed urgently. https://www.tandfonl...90840802305423#

 

That higher blood level reaching cancer killing blood levels(which are said to likely work by producing hydrogen peroxide) was achieved using forms of liposomal vitamin C.

 

Need to do more research to see at what doses or blood levels it starts producing hydrogen peroxide, at what amounts, and compare the levels to the concentration achieved in the study were such was said to be stronger preserver of telomeres.   That would allow optimal level or dose to be achieved.

[Castiel]

more research

 

Ascorbyl palmitate is also marketed as vitamin C ester," which should not be confused with Ester-C® (see above).

https://lpi.oregonst...plemental-forms

 

it seems the other promising compound ascorbyl palmitate is a vitamin c ester but not ester-c brand.

 

edit:

 

 

Ascorbyl palmitate

Ascorbyl palmitate is a fat-soluble antioxidant used to increase the shelf life of vegetable oils and potato chips (13). It is an amphipathic molecule, meaning one end is water-soluble and the other end is fat-soluble. This dual solubility allows it to be incorporated into cell membranes. When incorporated into the cell membranes of human red blood cells, ascorbyl palmitate has been found to protect them from oxidative damage and to protect α-tocopherol (a fat-soluble antioxidant) from oxidation by free radicals (14). However, the protective effects of ascorbyl palmitate on cell membranes have only been demonstrated in the test tube. Taking ascorbyl palmitate orally probably doesn't result in any significant incorporation into cell membranes because most of it appears to be hydrolyzed (broken apart into palmitate and ascorbic acid) in the human digestive tract before it is absorbed. The ascorbic acid released by the hydrolysis of ascorbyl palmitate appears to be as bioavailable as ascorbic acid alone (15). The presence of ascorbyl palmitate in oral supplements contributes to the ascorbic acid content of the supplement and probably helps protect fat-soluble antioxidants in the supplement. The roles of vitamin C in promoting collagen synthesis and as an antioxidant have generated interest in its use on the skin (see the article, Vitamin C and Skin Health). Ascorbyl palmitate is frequently used in topical preparations because it is more stable than some aqueous (water-soluble) forms of vitamin C (16). Ascorbyl palmitate is also marketed as vitamin C ester," which should not be confused with Ester-C® (see above).

That suggests that most of it is probably broken in the digestive track, but it seems probable some of it makes it intact into the blood.   Though not sure if enough to be effective.

 

edit2:

Saw that some liposomal forms have ascorbyl palmitate, not sure, but the liposomal delivery might allow more to get intact into the blood.

 

edit3:

 

The "Fat Soluble" Vitamin C Ester Myth

A perfect example of a product taking advantage of the "liposomal" term is a type of vitamin C  a  vitamin C ester . Usually labeled as "fat soluble" vitamin C, esters like Ascorbyl Palmitate are marketed as a “liposomal” but will never result in forming an actual "liposome". https://coremedscien...somal-vitamin-c

 

That article suggests that the liposomal forms using ascorbyl palmitate do not truly form liposomes, so might not have enhanced absorption

 

edit 4:

Also same article

 

 

Unlike natural vitamin C (Ascorbic Acid), Ascorbyl Palmitate may actually be toxic to skin cells damaged by UV exposure according to one study. 

Need to check doses here too, seems some doses might be toxic to skin, thankfully most of it is broken down by digestion, hopefully cells in digestive tracks aren't as sensitive.     Need more research need to check if toxicity doses are higher or lower than doses suggesting potential longevity or lifespan increase and doses necessary to inhibit hyaluronidase.

Edited by Castiel, 07 April 2021 - 10:20 PM.

[QuestforLife]

Need to do more research to see at what doses or blood levels it starts producing hydrogen peroxide, at what amounts, and compare the levels to the concentration achieved in the study were such was said to be stronger preserver of telomeres. That would allow optimal level or dose to be achieved.

In the paper you posted: 'AGE-DEPENDENT TELOMERE SHORTENING IS SLOWED DOWN
BY ENRICHMENT OF INTRACELLULAR VITAMIN C VIA
SUPPRESSION OF OXIDATIVE STRESS'

Endothelial cells - which line the blood vessels and will highly likely be affected by oxidative stress and the presence or absence of Vit C and H2O2 in the blood - lost telomeres at a greater rate with only 0.1-1uM concentration of H2O2 (see Fig 2).

The beneficial effects from 20uM of H2O2 in the paper posted by JamesPaul 'Slow-Down of Age-Dependent Telomere Shortening
Is Executed in Human Skin Keratinocytes by
Hormesis-Like-Effects of Trace Hydrogen Peroxide
or by Anti-Oxidative Effects of Pro-Vitamin C
in Common Concurrently With Reduction of
Intracellular Oxidative Stress' was found as the title states in keratinocytes.

There is no way you are going to have a higher concentration of H2O2 in keratinocytes than endothelial cells via an oral dose. Although if Vit C is preferentially sucked into the skin it is plausible it could then make H2O2. But I don't think that's relevant to this discussion.

Edited by QuestforLife, 07 April 2021 - 09:34 PM.

[Castiel]

In the paper you posted: 'AGE-DEPENDENT TELOMERE SHORTENING IS SLOWED DOWN
BY ENRICHMENT OF INTRACELLULAR VITAMIN C VIA
SUPPRESSION OF OXIDATIVE STRESS'

Endothelial cells - which line the blood vessels and will highly likely be affected by oxidative stress and the presence or absence of Vit C and H2O2 in the blood - lost telomeres at a greater rate with only 0.1-1uM concentration of H2O2 (see Fig 2).

The beneficial effects from 20uM of H2O2 in the paper posted by JamesPaul 'Slow-Down of Age-Dependent Telomere Shortening
Is Executed in Human Skin Keratinocytes by
Hormesis-Like-Effects of Trace Hydrogen Peroxide
or by Anti-Oxidative Effects of Pro-Vitamin C
in Common Concurrently With Reduction of
Intracellular Oxidative Stress' was found as the title states in keratinocytes.

There is no way you are going to have a higher concentration in keratinocytes than endothelial cells via an oral dose. So I don't think that's relevant to this discussion.

 

interesting had only read the abstracts, didn't notice that.

 

 

 

Now that info is curious will recheck the numbers but why would keratinocytes have reduction of telomere shortening with 20uM h202 which seems 20-200x higher than the 0.1-1uM concentration that it seems the other paper suggests increase telomere shortening in endothelial cells?  

 

edit: if different cells do indeed have such diverse responses to hydrogen peroxide, it seems like inducing a general small amount of hydrogen peroxide might not be usable in practice.

 

edit2:  Note my previous post had some significant edits, multiple ones, interesting at least regards one of the promising vitamin C compounds, the vitamin c ester ascorbyl palmitate

Edited by Castiel, 07 April 2021 - 10:21 PM.

[Advocatus Diaboli]

Removed duplicate reference.

Edited by Advocatus Diaboli, 07 April 2021 - 10:52 PM.

[Castiel]

interesting had only read the abstracts, didn't notice that.

 

 

 

Now that info is curious will recheck the numbers but why would keratinocytes have reduction of telomere shortening with 20uM h202 which seems 20-200x higher than the 0.1-1uM concentration that it seems the other paper suggests increase telomere shortening in endothelial cells?  

 

edit: if different cells do indeed have such diverse responses to hydrogen peroxide, it seems like inducing a general small amount of hydrogen peroxide might not be usable in practice.

 

 

 

edit time ran out but I will check the papers another possibility is that one is measuring intracellular h2o2 and the other is measuring medium h2o2.   If that's not the case and those are the numbers their responses to h2o2 levels seem highly different.

[Castiel]

Skimmed the article on potential interventions with similar genetic signatures shown by multiple lifespan extending intervention.

 

Very long article, a treasure trove of data.   Interestingly the commonly activated pathways were even able to predict which mice strains would live longer without any intervention, suggesting they are indeed linked to longevity itself.

 

 

This is the part on ascorbyl palmitate

 

For the second approach, to identify candidate lifespan-extending drugs, we utilized the CMap platform (Lamb et al., 2006, Subramanian et al., 2017). It contains gene expression profiles of different human cell lines subjected to >1,500 compounds and allows searching for perturbagens producing gene expression changes similar to the pattern of interest. To identify drugs with significant longevity effects, we ranked them based on their association with gene signature related to maximum lifespan (Figure 6A). We then chose four compounds from the top of the ranking and applied them to UM-HET3 male mice for 1 month (Table S1E). These drugs included two mTOR inhibitors KU-0063794 (García-Martínez et al., 2009) and AZD8055 (Chresta et al., 2010), antioxidant ascorbyl-palmitate (Cort, 1974), and antihypertensive agent rilmenidine (Mpoy et al., 1988).

We performed RNA-seq of the liver samples from mice subjected to the drugs, together with the corresponding controls. To check if the hits predicted with human cell lines are reproduced in mouse tissues, we calculated gene expression responses to each of these drugs and passed them to the association test (Figure 7E). In agreement with the predictions, all compounds demonstrated positive associations with the common gene signature across lifespan-extending interventions (adjusted p value < 0.077). Moreover, KU-0063794 and ascorbyl-palmitate showed a consistent positive association with all longevity signatures, except for rapamycin (adjusted p value < 0.055).

https://www.scienced...550413119303729

The dose of ascorbyl palmitate given to the mice seems to be parts per million, since they mention mice I assume this is digestion and probably oral.   Maybe somewhere they specify more.   But if it is oral it seems that either mice have different digestion properties for ascorbyl palmitate than humans(given humans are said to break down most of it in digestive track), or if not even very small amounts of ascorbyl palmitate reaching the rest of the body have longevity promoting effects.

 

[Castiel]

Ok had time to reread abstract on h2o2 keratinocytes and skimmed a bit of the articles

 

From the reading it suggests to me they both are extracellular h2o2. 

 

suggesting asc2p might be a better avenue than h2o2 as regards telomere shortening.

 

Will have to check brands, and presence of animal or human trials.

 

As for ascorbyl palmitate sounds intriguing but haven't found enough information.   Several brands that are reputable sell it.   Will also have to check if there are animal or human trials.  seems like it might increase lipid peroxidation, perhaps ascorbyl palmitate works in low doses as a hormetic?  Although it also has some antioxidant activity.

 

Another thing is that perhaps asc2p might have similar longevity and hyaluronidase inhibition effects as ascorbyl palmitate.  And it appears to be more stable and probably absorbed in greater amounts.  But don't think such has been investigated.

[Castiel]

Continued the vitamin C research.

 

And found this interesting quote in an interview with Sardi

 

 

Sardi: Irwin Stone noted that animals that naturally secrete vitamin C live 8-12 times beyond their age of physical maturity whereas animals like guinea pigs, primate monkeys and fruit bats that have the same GULOP gene mutation live only 2-3 times beyond their age of maturation. If vitamin C synthesis could be naturally restored, using these figures, humans would then be expected to live to 144-216 years.

The Amazing Health and Substantial Longevity Benefits of Restoring the Grossly Inadequate Levels of Vitamin C in Humans to Normal Mammalian Levels | WholeFoods Magazine

 

That sounds very promising if it is true, if it holds across multiple species.   Not sure about other species, need to check what is being used as age of physical maturation for humans too.    Height stops increasing at age 18-20, but brain development is said to continue till about age 25, iirc.

 

As mentioned previously this study in mice seems to suggest high vitamin c can restore lifespan of an artificial induction of gulop mutation back to that of an animal without the mutation.

Serum vitamin C levels modulate the lifespan and endoplasmic reticulum stress response pathways in mice synthesizing a nonfunctional mutant WRN protein (nih.gov)

[Castiel]

Continued vitamin c research and found some more cool info, and also came up with an interesting hypothesis.

 

First hydroxytyrosol one of the main polyphenols of extra virgin olive oil is said might increase vitamin c levels, some say even without additional supplementation

 

https://www.research...volunteer_trial

 

According to Sardi and other sources, iirc, despite some point mutations causing premature termination codons rendering genes nonfunctional, substances have been found that can allow early stopcodon readthrough, essentially partially restoring the function of a mutated gene.  Certain antibiotics, and resveratrol have shown this functionality for certain genes.   The rumor is that the way that hydroxytyrosol in extra virgin olive oil increases vitamin C might be by partially restoring the vitamin c gulo gene functionality in humans and allowing the final protein or final step in the vitamin c production cascade to be completed.

 

Another interesting article was this

https://nutrafol.com...or-hair-health/

 

One of their links shows that a version of vitamin c called asc 2p which I think is the one mentioned earlier with regards to reducing telomere shortening, might also have potential in hair growth at least according to animal studies.

https://applepoly.co...hair_growth.pdf

 

The article claims DHT activates dkk1 which some call the baldness protein, supposedly vitamin C can stop production of the downstream dkk1 protein.

 

Searching for that on google, suggested that asc 2p is the substance that is capable of blocking dkk1

https://pubmed.ncbi....h.gov/20701628/

 

Interesting, need to check if regular vitamin c might have similar functionality or not.   That sounds good to help preserve hair density.

 

Also this is the hypothesis

 

Will add have come up with a hypothesis on why CR might have had less benefits on primates.

Most animals synthesize vitamin c. Primates have a defect that they dont. Heard, but need to confirm, that animals that produce vitamin c live 8-12x their age of maturity also heard animals that dont only live 2-3x age of maturity. Also heard when an animal is stressed vitamin c production significantly increases. I suspect CR being a constant strong stressor might be drastically increasing vitamin c production, though need to check literature to see if anyone has checked.

Very high vitamin c might be the missing ingredient that yielded subpar cr results in primates

 

[QuestforLife]

 

First hydroxytyrosol one of the main polyphenols of extra virgin olive oil is said might increase vitamin c levels, some say even without additional supplementation

 

According to Sardi and other sources, iirc, despite some point mutations causing premature termination codons rendering genes nonfunctional, substances have been found that can allow early stopcodon readthrough, essentially partially restoring the function of a mutated gene.  Certain antibiotics, and resveratrol have shown this functionality for certain genes.   The rumor is that the way that hydroxytyrosol in extra virgin olive oil increases vitamin C might be by partially restoring the vitamin c gulo gene functionality in humans and allowing the final protein or final step in the vitamin c production cascade to be completed.

 

 

 

Hydroxytyrosol is claimed as some sort of magic ingredient to increase Vit C levels, but actually it is probably just preserving Vit C, by virtue of being a strong anti-oxidant in its own right.

Like others on this site I’ve done some crazy experiments, including 30-60g Glycine/day experiment. Previously I’d been judging different Vit C sources by testing my urine content with a reagent strip. During the high dose Glycine experiment (which presumably boosts glutathione) the Vitamin C levels in my urine hit the maximum measurable level immediately on urinating (100mg/dl). And that is 12 hours after taking 500mg of Vitamin C (palmitate). I’d never seen my urine turn the strip that bright yellow that quickly before, even with much higher doses of Vitamin C. My body was disposing of Vitamin C like it had no need of it!

But when we start talking about Bill Sardi it is a sign we might have gone too deeply down a Vitamin C rabbit warren.

To finish I’ll say this - Asc2p tends to be the choice of researchers when growing stem cells in vitro, as mentioned here:

The Influence of Cell Culture Density on the Cytotoxicity of Adipose-Derived Stem Cells Induced by L-Ascorbic Acid-2-Phosphate

https://www.nature.c...598-019-56875-0

Ascorbic acid-2-phosphate (A2-P) is an oxidation-resistant derivative of ascorbic acid that has been widely employed in culturing adipose-derived stem cells (ASCs) for faster expansion and cell sheet formation.

 

This is because according to many sources, it is resistant to oxidation (more stable).

 

According to the first paper posted by you, ‘AGE-DEPENDENT TELOMERE SHORTENING IS SLOWED DOWN BY ENRICHMENT OF INTRACELLULAR VITAMIN C VIA SUPPRESSION OF OXIDATIVE STRESS’ the intra cellular concentration of vitamin C was increased 4x over plain old vanilla Vitamin C (ascorbic acid).

 

So in vitro Asc2P is superior. The question of whether it is better in vivo is open. According to the link you sent from the Linus Pauling Institute no Vitamin C is any much better than any other for bioavailability, but I note they haven’t included Asc2p in their analysis.

 

To conclude, my suggestion for telomeres is the following: try Asc2p plus another antioxidant like Hydroxytyrosol. Use a reagent strip to test them alone and together and compare to other Vitamin C sources. 

[QuestforLife]

So what is the most powerful telomerase activator anyway?

1.TAM818 @ 16% of that produced by HELA cells according to mRNA assay for HTERT gene expression (according to Bill Andrews); or
2. A triterpene rich fraction including Asiatic acid/asiaticoside @17% of HELA, according to this paper (REF1: DOI: 10.3892/mmr.2019.10614), measured by TRAP assay (TERT protein); or
3. The tetrapeptide Epitalon @ 100% HELA according to this paper (REF2: DOI: 10.1023/a:1025493705728) measured again by TRAP (protein) assay (and also immunohistological staining using antibodies, which also detects the protein)

 

What are we to make of these various claims? Who has the strongest telomerase activator really?
REF1 has not been replicated yet, though asiaticoside is known to be anti inflammatory (SMAD 7 activator, which downregulates TGF-B), is known to be beneficial for skin, and I have heard of one Longecity user who got longer telomeres apparently from taking gotu kola. 
REF2 has not been replicated outside Russia, and in fact Bill Andrews states in a video interview that epitalon does not activate the Telomerase gene, in repeated experiments using his mRNA assay. It does not seem to be in question that epitalon increases endogenous production of melatonin, a powerful endogenous anti-oxidant, or that epitalon can increase telomere lengths in people (although I should note, even antioxidants can do that – REF4: DOI: 10.3892/ijmm.2019.4191, also here: https://www.certifie...cts-telos95.php)
Bill Andrews developed his assay after many years of painstaking manual tests until he found a hit and then used that as a positive control to create an automated assay. No one else can replicate his findings. But he has had several university departments independently confirm that Sierra Sciences’ first mRNA TERT hit was also positive in a TRAP assay.

 

This brings us to an interesting point. For activation of TERT you’d expect increased mRNA transcription, as measured by Bill, followed by translation into the telomerase protein, as measured by everyone else. Bill Andrews has stated sometimes they’d get a hit with the mRNA but not with the subsequent TRAP assay, possibly because some compounds both activate the gene and downregulate translation.
Bill Andrews’ TERT mRNA assay is proprietary and has been refined to be highly sensitive.  Almost everyone else is relying on the TRAP assay. Is there anything wrong with that? One might think that if mRNA is a hit, TRAP might not be if the protein is not made for some reason. But if TRAP is a hit, then surely mRNA must also be a hit? Is there any circumstance where this is not the case?

 

Before we answer that question there is some housekeeping to do.

How accurate is a TRAP assay? I am no expert, but I’d say looking at this paper (REF5: https://pubmed.ncbi....h.gov/27182535/) that it can’t be very accurate. It relies on seeing how many telomere repeats are added to an artificial substrate, and then using PCR to amplify those repeats into something sufficiently long we can measure and compare. Presumably, the number of amplification cycles is critical and the same for all samples. But clearly one could just increase the number of cycles and make it appear you had a massive hit. I’d hazard a guess that there is at least some doubt over the TRAP assay. I’d also point out that References 1-3 have not be replicated (to my knowledge), which adds weight to my argument.

 

There are also more legitimate reasons why the TRAP assay might give you a hit without activating transcription from the gene. Any ‘spare’ telomerase mRNA could be translated into the telomerase protein in the right circumstances. The fact telomerase can be detected at all in human cells even though the gene is normally repressed shows that this is plausible. Bill Andrews has stated that during DNA replication any repression is temporarily removed. So, we’d expect some low level telomerase to be detectable in culture where cells are continually dividing. As we’ve discussed recently, antioxidants can help cells divide for longer – and some papers show this is due to a higher telomerase level being preserved for longer. This shows that at least at low levels, there is some flex in what you can do with a fixed level of telomerase. The same might be true one step earlier: the process of making telomerase from the mRNA. I’ve seen various studies showing Astralagus, Vitamin D, Broccoli extract, Rhodolia, Resveratrol, etc., all producing telomerase to some degree above the background level. But still at a very low level. I do not have access to Bill Andrews’ lab to know whether this is via mRNA upregulation or by increasing the amount of translation to the active protein form. But it might explain transient telomerase expression. I’ve read papers for example where resveratrol transiently caused telomerase expression in culture. Why transiently? It might be because resveratrol is one of those substances that uses up the already available gene product to make the protein. But once that mRNA is used up, translation fails. So again, I’d hazard a guess to say many purported telomerase activators have little effect on increasing gene expression.

 

See REF3: A critical role of nicotinamide phosphoribosyltransferase in human telomerase reverse transcriptase induction by resveratrol in aortic smooth muscle cells

Here, we report that resveratrol activates human nicotinamide phosphoribosyltransferase (NAMPT), SIRT4 and telomerase reverse transcriptase (hTERT) in human aortic smooth muscle cells. Similar observations were obtained in resveratrol treated C57BL/6J mouse heart and liver tissues. 

See Fig 1: https://www.ncbi.nlm...port=objectonly

 

Note here they claim Resveratrol DOES increase mRNA levels, although Bill Andrews has said he has not been able to replicate this (personal communication), although he did say he takes it anyway ‘just in case’.

 

Although to some the argument that most ‘telomerase activators’ might not actually be increasing gene expression could be depressing, it also offers us an opportunity. Because if for example, Epitalon is actually only upregulating translation (and I do not know if this is the case or not), then it means we have a way of enhancing the effects of a genuine telomerase activator like TAM818. That way we’d be increasing mRNA from the gene and also upregulating its translation into telomerase. Doing this we might make the most telomerase possible. I am speculating now, but I find it interesting that the pathway described in the paper above (REF3) involves NAMPT, so likely influences the oxidative/reductive balance of the cell, so may be influencing telomerase via the anti-oxidant effect. The oxidative/reductive balance of cells also influence circadian rhythm (as anyone who has taken NMN can attest). Taking this another step, epitalon dosing increases melatonin production, and melatonin is a very important endogenous antioxidant also involved in regulating senescence through multiple pathways, and the circadian rhythm. Melatonin will be explored in a future post.

Edited by QuestforLife, 08 April 2021 - 03:25 PM.

[Castiel]

 

There are also more legitimate reasons why the TRAP assay might give you a hit without activating transcription from the gene. Any ‘spare’ telomerase mRNA could be translated into the telomerase protein in the right circumstances. The fact telomerase can be detected at all in human cells even though the gene is normally repressed shows that this is plausible. Bill Andrews has stated that during DNA replication any repression is temporarily removed. So, we’d expect some low level telomerase to be detectable in culture where cells are continually dividing. As we’ve discussed recently, antioxidants can help cells divide for longer – and some papers show this is due to a higher telomerase level being preserved for longer. This shows that at least at low levels, there is some flex in what you can do with a fixed level of telomerase. The same might be true one step earlier: the process of making telomerase from the mRNA. I’ve seen various studies showing Astralagus, Vitamin D, Broccoli extract, Rhodolia, Resveratrol, etc., all producing telomerase to some degree above the background level. But still at a very low level. I do not have access to Bill Andrews’ lab to know whether this is via mRNA upregulation or by increasing the amount of translation to the active protein form. But it might explain transient telomerase expression. I’ve read papers for example where resveratrol transiently caused telomerase expression in culture. Why transiently? It might be because resveratrol is one of those substances that uses up the already available gene product to make the protein. But once that mRNA is used up, translation fails. So again, I’d hazard a guess to say many purported telomerase activators have little effect on increasing gene expression.

 

See REF3: A critical role of nicotinamide phosphoribosyltransferase in human telomerase reverse transcriptase induction by resveratrol in aortic smooth muscle cells

See Fig 1: https://www.ncbi.nlm...port=objectonly

 

Note here they claim Resveratrol DOES increase mRNA levels, although Bill Andrews has said he has not been able to replicate this (personal communication), although he did say he takes it anyway ‘just in case’.

 

 

 

Didn't we have the other article in the resveratrol subsection that showed resveratrol doubled the telomere length of near senescent cells?

 

IIRC humans lose about 70 base pairs, senescent cells have about 5Kb, so near senescent cell telomeres should be even more.  If you double near senescent cell telomere that is 5Kb gain, which at a loss of 70 base pairs per year seems like you fully rejuvenate the cell, at least in terms of telomeres.

 

 

 

"When I saw some of the cells in the culture dish rejuvenating I couldn't believe it. These old cells were looking like young cells. It was like magic," she said. "I repeated the experiments several times and in each case the cells rejuvenated. I am very excited by the implications and potential for this research."

 Old human cells rejuvenated in breakthrough discovery on aging -- ScienceDaily

 

Eva Latorre, Vishal C. Birar, Angela N. Sheerin, J. Charles C. Jeynes, Amy Hooper, Helen R. Dawe, David Melzer, Lynne S. Cox, Richard G. A. Faragher, Elizabeth L. Ostler, Lorna W. Harries. Small molecule modulation of splicing factor expression is associated with rescue from cellular senescence. BMC Cell Biology, 2017

Edited by Castiel, 08 April 2021 - 09:04 PM.

[QuestforLife]

 Old human cells rejuvenated in breakthrough discovery on aging -- ScienceDaily

 

Eva Latorre, Vishal C. Birar, Angela N. Sheerin, J. Charles C. Jeynes, Amy Hooper, Helen R. Dawe, David Melzer, Lynne S. Cox, Richard G. A. Faragher, Elizabeth L. Ostler, Lorna W. Harries. Small molecule modulation of splicing factor expression is associated with rescue from cellular senescence. BMC Cell Biology, 2017

 Yes, you're right - in vitro resveratrol can do something to the telomere maintenance genes via splicing factors (I think it may have been HnRNP) and cause a big increase in telomere length. I did loads of research into this near the start of this thread, and found others papers with similar hints of telomerase activity from resveratrol. It is very odd though - it doesn't always work. It could be a very narrow concentration range, or require the telomere to be very short already (like in the Latorre study) or require some other unknown factor the researchers weren't cognizant of to be present as well. It might well be that being a HDAC inhibitor (which makes the genome more accessible) means resveratrol would pair well with other substances. But I hit a dead end with resveratrol research. Plus it is quite a toxic substance and taking large doses made me feel ill. 

[Believer]

I feel stupid asking these questions, seems the answer should be simple but I can't figure it out

If something was able to match HELA cells how would that show in a telomere length test before and after using the product?

Would a cell go from 10,000 base pairs in telomere length to 15,000 after one cell division or would it go from those 10k to 10.1k as (it seems) that products like epitalon can achieve?

 

If supplements like cycloastragenol and others are too mild for cell immortality, why do the studies show a net increase in telomere length in cell culture and in human trials?

[QuestforLife]

I feel stupid asking these questions, seems the answer should be simple but I can't figure it out
If something was able to match HELA cells how would that show in a telomere length test before and after using the product?
Would a cell go from 10,000 base pairs in telomere length to 15,000 after one cell division or would it go from those 10k to 10.1k as (it seems) that products like epitalon can achieve?

If supplements like cycloastragenol and others are too mild for cell immortality, why do the studies show a net increase in telomere length in cell culture and in human trials?

Not a stupid question at all.

Telomere length (TL) is measured as a snapshot in white blood cells (WBCs). Even a weak Telomerase activator or a powerful antioxidant can appear to increase the TL of WBCs because they are replaced every week or so by fresh WBCs from their underlying progenitors, who naturally have longer telomeres. You haven't actually increased the TL of a given WBC, just improved conditions slightly, enough for a new WBC to have slightly longer TL in the same situation than the one it replaced. Conversely, if you catch a virus, WBCs will divide faster and TL will get shorter on average. But the underlying picture hasn't changed much - you are still losing TL overall (in the stem cell compartment), just at a slightly slower rate.

So the theory goes that even a weak telomerase activator and an antioxidant (which doesn't downregulate endogenous production so end up being counterproductive) will extend lifespan, even though all it is doing is slowing the rate of telomere loss in stem cells.

HELA is used as a positive control as they have the lowest telerase activity of cancer cells known to be sufficient to sustain indefinite division. If you had a way of conferring this level of telomerase activity on your cells, they wouldn't necessarily get longer telomeres, they might just divide faster, if that is what the body needed. At this point we don't know exactly whether greater telomerase would benefit us by giving us faster turnover of skin, the immune system, etc., or whether the benefit comes from having longer telomere in and of themselves - perhaps from more youthful gene expression. As telomerase is known to preferentially lengthen the shortest telomeres, this suggests to me that these are what is preventing division not only in themselves, but perhaps in other cells nearby too. So we shouldn't discount the benefits of faster turnover of cells.

But to my mind a super high dose of telomerase, much more than HELA, such as from mRNA treatment, or a gene therapy, applied once, and restoring all your telomeres to youthful (long) length, with telomerase activity then ceasing - would be the ideal treatment.

Edit - just noticed you asked about cell culture. Telomerase activators have not been shown to lengthen telomeres in culture, only extend the number of passages achieved before senescence, as you'd expect.

Edited by QuestforLife, 09 April 2021 - 02:54 PM.

[Castiel]

 Yes, you're right - in vitro resveratrol can do something to the telomere maintenance genes via splicing factors (I think it may have been HnRNP) and cause a big increase in telomere length. I did loads of research into this near the start of this thread, and found others papers with similar hints of telomerase activity from resveratrol. It is very odd though - it doesn't always work. It could be a very narrow concentration range, or require the telomere to be very short already (like in the Latorre study) or require some other unknown factor the researchers weren't cognizant of to be present as well. It might well be that being a HDAC inhibitor (which makes the genome more accessible) means resveratrol would pair well with other substances. But I hit a dead end with resveratrol research. Plus it is quite a toxic substance and taking large doses made me feel ill. 

 

I don't take large quantities long term because I've heard it could cause issues.   But for very short term of a 1-2 days, I didn't get any uncomfort from a few gram high purity dose.    Of course very high purity is needed, otherwise you're gonna get issues due to emodin(also emodin is said to destabilize telomeres, so no good, also a bit of fiber helps counteract digestive issues from very small emodin content).    Given the information from blood levels attained from several brands, I should have been able to sustain the experimental values for 10+hours, which should have been enough to see some kind of result.   Also will add that the resveratrol telomerase connection may or may not be from splicing factor, some data in some experiments suggests sirtuin 4 is activated and can lengthen telomeres, in which case NAD+ may be needed, I'm not sure if I took that into account on earlier experiments.   On mice the older the mice the less telomerase activity was seen from resveratrol, which I've suspected is due to reduced NAD+ levels.  I think I'll try it again resveratrol but this time with several glasses of chamomile throughout the day and several spoons of parsley, + CR.   That should keep NAD+ at higher levels and may help with results.

 

I also did the fisetin megadose senescent cell clearance experiment at another time.

 

I barely have any gray hairs, and several of the few have turned black, but not all.    Skin wise I'm pretty youthful, so can't tell with that(have even been asked for ID at times).    Was doing a bit of gotu kola extract + low dose resveratrol for a bit.   Right now I'm planning on seeing what the asc2p does, not sure on dosing.   Will try 250mg, at first to see how it goes.   Being pretty potent and being able to affect hair growth, interfere with the actions of dht, it might solve the few remaining gray hairs.

 

I'll try asc2p at 250mg for a few weeks, if don't notice any reversal on roots, will try upping it to 500mg.  Then I'll try seeing what happens if I use asc2p + gotu kola + low dose resveratrol.(I'll try these while on some level of CR, which should increase NAD+ and a lot of longevity factors too.)

 

I hope asc2p is not broken down in digestive track, and is absorbed.   Would be nice to see if an experiment has been done on its absorption profile.

 

 

 

Telomerase activators have not been shown to lengthen telomeres in culture, only extend the number of passages achieved before senescence, as you'd expect.

But isn't resveratrol an exception, weren't the rejuvenated human cells in culture?   They said telomeres doubled in length, and went from looking near senescent to youthful looking.

Edited by Castiel, 10 April 2021 - 01:20 AM.

[QuestforLife]

Yes you got me there Castile - resveratrol does appear to lengthen the telomeres in an arrested cell culture. I await further confirmation from the team of the mechanism; their later work looked at hydrogen sulphide donors and did not achieve telomere elongation.

Yes, I've discussed the NAMP-SIRT4-TERT link ad naseum before. It drove me mad and I never got anywhere. Nor has any other paper ever replicated those findings.

Just to give you a flavour of how infuriating resveratrol is, look at this Jerry Shay paper from 2008.

Immortalization of epithelial progenitor cells mediated by resveratrol

doi: 10.1038/sj.onc.1210886

In the present work, we show that resveratrol activates, while progesterone inactivates, continuous telomerase activity within 24 h in subpopulations of human Li–Fraumeni syndrome-derived breast epithelial cells. Resveratrol results in immortalization of mixed progenitor cells with mutant p53, but not human epithelial cells with wild type p53. Our results demonstrate the potential for renewing progenitor cells with mutant p53 to immortalize after continuous telomerase expression when exposed to certain environmental compounds.

According to this work, low concentrations of resveratrol actual IMMORTALISED a subset of cells that had damaged P53.

Not something you want. Resveratrol is just bizarre. Until I know more about it, I'm steering clear. But keep me posted on your experiments.

[QuestforLife]

What is the best antioxidant to use to help our telomeres?

First, some background.

In vitro senescence is primarily caused by oxidative stress due to the atmospheric oxygen levels being used.

Nevertheless, the same occurs at physiological oxygen levels, just more slowly, where the effect is relevant to human tissues.

There is interplay between replicative senescence caused by short telomeres and stress induced senescence caused by ROS.

I evidence these statements below.
But first...

The case for Melatonin
According to the following paper, conducted mostly in vitro, senescing mesenchymal stem cells sabotage mitophagy to induce arrest [1].

Melatonin suppresses senescence‐derived mitochondrial dysfunction in mesenchymal stem cells via the HSPA1L–mitophagy pathway

https://doi.org/10.1111/acel.13111

In senescent MSCs (late passage), replicative senescence decreased mitophagy by inhibiting mitofission, resulting in the augmentation of mitochondrial dysfunction. Treatment with melatonin rescued replicative senescence by enhancing mitophagy and mitochondrial function through upregulation of heat shock 70 kDa protein 1L (HSPA1L).

Melatonin upregulates mitophagy via a heat shock protein as well as complex IV and endogenous antioxidants to reverse senescence. In the paper they claim MSCs are undergoing 'replicative' senescence and this is rescued by melatonin. Indeed, melatonin restored various checkpoint and proliferation markers and reduced markers of senescence. But they didn’t actually multiply cells beyond passage 9 (when they originally arrested) and they didn’t measure telomeres. It is a starling omission in an otherwise brilliant paper. They did show that ischemic hind legs of mice were (mostly) saved from apoptosis by an infusion of Passage 2 (young) MSCs and Passage 9 (old) MSCs with Melatonin but not with Passage 9 (old) MSCs without Melatonin. Nevertheless, I think the passage 9 (old) MSCs mostly senesced via ROS induced stress, not short telomeres.

There is precedent for various antioxidant compounds to enable more passages without increasing telomerase. Interestingly, the two most obvious aren’t direct antioxidants, but (similarly to melatonin) act on mitochondria to reduce ROS endogenously.

In vitro mitophagy enhancer nicotinamide significantly increases the passages required for a culture to senesce. Investigations show that this is caused by keeping ROS from rising [2]. A similar effect has been shown using methylene blue (MB). The mechanism is via hormesis reducing ROS via increased efficiency of mitochondria [3]. Importantly this work was replicated at physiological oxygen levels [4]; the benefits of MB on the culture were reduced, but still significant, showing this is applicable to human tissues. Eventually real replicative senescence occurs (via short telomeres) and this causes a rapid rise in ROS followed by senescence regardless of the presence of nicotinamide [2] or MB.

Going back to melatonin, the following study shows it also increases the growth of colonies of reprogrammed somatic cells (using Somatic nuclear Transfer). Remarkably the mechanism was found to be telomere elongation via the inhibition of repressive histone (3) modifications [5]. But these cells are reprogrammed via somatic nuclear transfer, and animals cloned this way are known to have abnormal telomeres. But in these cells at least, melatonin is actually restoring telomeres.

Melatonin is produced endogenously, but it declines with age; it can be taken directly, but is also upregulated by taking epitalon. It is a powerful antioxidant both directly and via its action on mitochondria, for which it uses a transporter to accumulate within. Melatonin may even lengthen telomeres in some stem cells.

In conclusion, if you are looking for an antioxidant to help your telomeres, look no further than melatonin.

[1] Melatonin suppresses senescence‐derived mitochondrial dysfunction in mesenchymal stem cells via the HSPA1L–mitophagy pathway,
https://doi.org/10.1111/acel.13111

[2] Nicotinamide extends replicative lifespan of human cells
https://doi.org/10.1...26.2006.00234.x

[3] Combined activation of the energy and cellular-defense pathways may explain the potent anti-senescence activity of methylene blue
https://doi.org/10.1...dox.2015.09.004

[4] Methylene blue delays cellular senescence and enhanced key mitochondrial biochemical pathways
doi: 10.1096/fj.07-9610com

[5] Inhibiting repressive epigenetic modification promotes telomere rejuvenation in somatic cell reprogramming
https://doi.org/10.1096/fj.201901486RR

[QuestforLife]

Age Related Methylation

 

An important paper of years past concerned the epigenetic regulation of the nuclear encoded, mitochondrially important, glycine gene in human fibroblasts. Downregulation of this SHMT2 gene with age downregulates the manufacture of new mitochondrial DNA [1].

 

As discussed more recently, reduced regulation of the GDF11 promoter by the demethylase TET2 [2] in mesenchymal (and neural) stem cells drives loss of GDF11 and the renewal capacity of these stem cells.

 

Recently I have been researching Oxytocin as another putative rejuvenation molecule and again found the same pattern regarding methylation changes. Keratinocytes (skin cells) in older women (but not men) suffer from methylation of the Oxytocin receptor. [3] In their experiment, keratinocytes cultured to replicative senescence were harvested for their culture medium (inflammatory cytokines; the SASP), which was then used to induce senescence in separate cultures of otherwise functional, non-senescent keratinocytes. Oxytocin was able to reverse the induced senescence in keratinocytes from young but not old female donors and the reason for the failure in older donor cells was found to be methylation of the oxytocin receptor. This was then overcome using a demethylating agent. From this study it stands to reason that oxytocin could rejuvenate skin cells so long as sufficient oxytocin receptors remain in those cells, otherwise a demethylase cofactor might be required. I note that the Conboys’ study using oxytocin (alone) used only male mice [4], though the focus was muscle for which I do not have methylation data. It is interesting also that the Conboys in a later paper used a ALK5 (tgf-b) inhibitor along with oxytocin to treat aging in mice [5], so perhaps they are concerned that the benefits of oxytocin might be blocked.  In the case of oxytocin and skin cells, I do find it suggestive that specifically women suffer from age related methylation of oxytocin receptors and that this could potentially stand in the way of skin rejuvenation via oxytocin.

 

Moving back to the general theme, it appears that methylation (and potentially other epigenetic mechanisms) are directly causal in age related decline of important genes like GDF11 and Oxytocin (or their receptors). It is not clear how well these sites are captured in the various epigenetic aging clocks that are available. It is also not clear what is driving this errant methylation(*); given methylation is an active process what exactly is causing it to become over-active? It is an interesting question. It does appear that methylation rather than demethylation is driving these issues, this is backed up by Horvath’s latest pan species clock [6] with de novo methylation the main culprit [7].      

 

(*) this paper [8], referenced on this thread previously implies the de novo methylation enzyme Dnmt3a is important for differentiation, with loss in Hematopoetic Stem Cells immortalising those cells, but preventing them differentiating to be used by the body. 

 

References

[1] Epigenetic regulation of the nuclear-coded GCAT and SHMT2 genes confers human age-associated mitochondrial respiration defects, DOI: 10.1038/srep10434
[2] Mutual regulation between GDF11 and TET2 prevents senescence of 2 mesenchymal stem cells, https://doi.org/10.1...20.03.30.008722
[3] Oxytocin alleviates cellular senescence through oxytocin receptor-mediated extracellular signal-regulated kinase/Nrf2 signalling, DOI 10.1111/bjd.17824
[4] Oxytocin is an age-specific circulating hormone that is necessary for muscle maintenance and regeneration, DOI: 10.1038/ncomms5082
[5] Rejuvenation of brain, liver and muscle by simultaneous pharmacological modulation of two signaling determinants, that change in opposite directions with age, doi: 10.18632/aging.102148
[6] Reversing age: dual species measurement of epigenetic age with a single clock, doi: https://doi.org/10.1...20.05.07.082917
[7] Age and gender affect DNMT3a and DNMT3b expression in human liver, DOI 10.1007/s10565-007-9035-9

[8] Loss of Dnmt3a Immortalizes Hematopoietic Stem Cells In Vivo, doi:10.1016/j.celrep.2018.03.025.

Edited by QuestforLife, 20 April 2021 - 11:16 AM.

[QuestforLife]

Continued...From the above post, it appears that there is a conflict between the need for stem cells to replenish the tissues of the body (differentiation; dependent on de novo methylation), and the need for stem cells to self-renew to maintain their own population and their capability to replenish tissues in the future (proliferation; note reprogramming has been shown to be dependent on demethylases [9]). This balance is partly maintained through the day-night cycle and the eating-fasting cycles and homeostasis is marginally improved  by various healthy lifestyles.

 

In reference [7] above, we saw that Hematopoietic stem cells can become immortal (at least in mice) with the simple loss of the de novo methylase Dnmt3a, but that this comes at the cost of haematopoiesis (production of new blood cells). 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!). We have already seen the same rough outline of this process in epidermal progenitors [11], mentioned in this thread many times, whereby symmetrical division (proliferation) wins out over differentiation leading to atrophy of the upper skin layer.

 
One might consider this in the light of my Selfish Cell lives longer theory, whereby Hematopoietic Stem Cells or Epidermal Skin cells or Spermatogonial Stem cells are losing telomeres and their ability to proliferate. A subset of such cells under selection pressure overcomes this epigenetically, but at the cost of differentiation ability.

 

[9] Reversal of ageing- and injury-induced vision loss by Tet-dependent epigenetic reprogramming, https://doi.org/10.1101/710210

[10] Aging of spermatogonial stem cells by Jnk-mediated glycolysis activation, https://doi.org/10.1...pnas.1904980116
[11] Stem cell competition orchestrates skin homeostasis and ageing, https://doi.org/10.1...1586-019-1085-7

[yz69]

Interesting! Any idea what can we do about it?

[QuestforLife]

Interesting! Any idea what can we do about it?

In my Selfish Cell lives Longer hypothesis, a population of say, stem cells (for example)has a natural variation in epigenetic state. Some cells will naturally be more responsive to differentiation signals, and others will be less likely to differentiate and more likely to self renew. Over time the cells that self renew (and not replenish tissues) will come to predominate. Therefore tissues will atrophy.

The following paper from 2018 presents some evidence for this hypothesis.

DNA Methylation Patterns Separate Senescence
from Transformation Potential and Indicate
Cancer Risk

https://doi.org/10.1...ell.2018.01.008

We show that transformation-associated
methylation changes arise stochastically and independently of programmatic changes during senescence.
Promoter hypermethylation events in transformation involve primarily pro-survival and developmental genes,
similarly modified in primary tumors. Senescence-associated hypermethylation mainly involves metabolic
regulators and appears early in proliferating ‘‘near-senescent’’ cells, which can be immortalized but are re-
fractory to transformation. Importantly, a subset of transformation-associated hypermethylated develop-
mental genes exhibits highest methylation gains at all age-associated cancer risk states across tissue types.
These epigenetic changes favoring cell self-renewal and survival, arising during tissue aging, are fundamen-
tally important for stratifying cancer risk and concepts for cancer prevention.

In this paper they looked at cell culture and found that the methylation changes that are associated with cellular senescence (telomere loss) are programmed and predictable and distinct from the methylation changes the occurred to cell lines that had been immortalised, or immortalised and then also transformed into cancer cells. Then the methylation changes were random, with all lines diverging from one another, except in the case of (expression of) genes that were selected for by the high growth environment, namely developmental genes. Interestingly, it was this latter cancer related signal rather than the senescent one that was prevelant in the tissues taken from older humans.

This 'selfish cell' phenotype is not sufficient but predisposes cells to cancerous transformation. So in this sense aging and cancer are part of the same process. And ironically the cellular senescence experienced in the wider body is the consequence of selection pressure on stem cells selecting for those who can avoid cellular senescence via increased self renewal (and reduced differentiation).

What can we do?

Some ideas spring to mind. I speculate somewhat.

If stem cells had longer telomeres they'd have no need to resort to methylation mediated changes to survive so that selection pressure would be removed.

I also note that in the study when they immortalised cells already near senescence they were unable to then turn them cancerous and their methylation pattern remained constant with continued passaging. So those signals that turn down metabolism seem to guard against the stochastic changes that can lead to selection for the selfish cell and eventually cancer. So rapamycin or everolimus might fit the bill.

Also these changes are driven by methylation, so demethylation seems like a easy choice. AKG is a no-brainer.

Finally anything that encourages differentiation should help push those reticent selfish cells into action. Not sure what to suggest for that. Maybe stem cell stimulants.

Edited by QuestforLife, 21 April 2021 - 05:48 PM.

[naxleo]

Hi, I have a question which is partly related to the last posts in this thread.

 

Which are the effects of S-adenosylmethionine (SAM-e) on methylation age?

 

Are you aware of any study investigating the relation? Do you have any guess on it?

 

I am uncertain whether to supplement with SAM-e, and in case its timing.

[QuestforLife]

 

Which are the effects of S-adenosylmethionine (SAM-e) on methylation age?

 

 

I have not tried SAM-e. 

Some people do find it a helpful supplement. It would be interesting to find out its effects on epigenetic age as measured by methylation changes.

The deleterious changes I have highlighted with GDF11 and Oxytocin Receptor downregulation involve increased methylation.

Therefore I would recommend supplements that reduce methylation rather than add to it. 

[QuestforLife]

AKG and Telomere Length

 

Limiting the activity of de novo methyl transferases is required for telomere lengthening in mouse embryonic stem cells [1]. TET demethylases are required for reprogramming of somatic cells [2]. Alpha Ketoglurate is a cofactor for the TET enzymes [3] and appears to reverse epigenetic age in humans [numerous sources]. 

Following this line of evidence, if there is regulation of telomere length via methylation, can AKG increase telomere length?

 

The following paper offers some encouraging evidence:

α-ketoglutarate delays age-related fertility decline in mammals
https://doi.org/10.1111/acel.13291

 

 

The results showed that the telomere length was significantly shorter in the ovaries of 14‐month‐old mice than that in the 8‐week‐old young mice (p < 0.05) (Figure 6a). However, the telomere length in 14‐month‐old mice treated with α‐KG was significantly longer than that in their age‐matched control mice (p < 0.05) (Figure 6a). Consistently, the telomerase activity in 14‐month‐old mice treated with α‐KG was also slightly higher than that in their age‐matched controls but it did not achieve significant difference (p > 0.05) (Figure 6b). Terc, Tert, and Sirt6 play an important role in telomere length. The results showed that the relative expression of Sirt6, Tert, and Terc was significantly down‐regulated in the ovaries of 14‐month‐old females compared to young ones, while these reductions were significantly recovered with α‐KG administration compared to their age‐matched mice (p < 0.05)

 

TERC (the RNA template copied by telomerase to extend the telomere) was upregulated, as was the mRNA of telomerase itself, but curiously the expected increase in the telomerase protein did not reach statistical significance. Nevertheless, telomere length in the ovaries of aged mice were significantly longer when taking AKG than for controls. Perhaps like with GDF11 the increase in TERC is sufficient to elongate telomeres on its own. We've also seen in the past that whilst telomerase (TERT) may be able to confer immortality to cell lines, TERC upregulation is required to lengthen telomeres [4]. This gives me some confidence that this effect may translate to humans. 

 

 

 

 

[1] DNA methyltransferases control telomere length and telomere recombination in mammalian cells, DOI: 10.1038/ncb1386
[2] Reversal of ageing- and injury-induced vision loss by Tet-dependent epigenetic reprogramming, https://doi.org/10.1101/710210
[3] α-Ketoglutarate Promotes Pancreatic Progenitor-Like Cell Proliferation, doi:10.3390/ijms19040943

[4] Small-Molecule PAPD5 Inhibitors Restore Telomerase Activity in Patient Stem Cells, DOI:https://doi.org/10.1...tem.2020.03.016

[aribadabar]

I have not tried SAM-e.
Some people do find it a helpful supplement. It would be interesting to find out its effects on epigenetic age as measured by methylation changes.
The deleterious changes I have highlighted with GDF11 and Oxytocin Receptor downregulation involve increased methylation.
Therefore I would recommend supplements that reduce methylation rather than add to it.

If you have impaired methylation cycle due to genetic abnormality (MTHFR) that doesn't seem health-protective. SAMe and other methyl donors like TMG, P5P and Methylfolate help in such cases. So if someone is having such genetic profile, I'd focus more on addressing that before worrying about DNA demethylation as too much Hcy will most probably kill you first.