1028 replies to this topic

[Castiel]

Some future additions to the protocol

 

Telomerase Activation
A recent paper has found some fairly potent telomerase activators, related but not the same as cycloastragenol – namely various triterpenes, the most effective being a 95% purity extract of the triterpenes in Gotu Kola, at peak effectiveness providing about 17% of HELA in immune cells (compared to about 3% for TA-65). This was at a very low concentration (0.02mg/ml). Ten times the concentration was slightly less effective, whilst 100 times the concentration gave almost no telomerase benefit (without causing any outright harm). Given that the strongest triterpene content Gotu Kola supplement I can find has 20% triterpenes, getting the same results in vivo seems plausible. I couldn’t find any bioavailability or half-life information for humans, but on the bright side, various health benefits for Gotu Kola are reported by users on Amazon. The study also confirms the small but apparently robust benefits of Vitamin D for telomerase. [doi:  10.3892/mmr.2019.10614].
Stem Cell mobilisation
AFA (Aphanizomenon Flos-Aquae), Fuciodan and Sea Buckthorn Berry extract all have some evidence for increasing the release of stem cells from the bone marrow [DOI: 10.1016/j.carrev.2007.03.004, 10.1016/j.exphem.2007.02.009, http://dx.doi.org/10.2147/CIA.s186893]. Ordinarily this would be done to increase healing after injury, or for general health, for the former there is some evidence of effectiveness [DOI: 10.19080/NTAB.2017.01.555564, DOI: 10.15406/mojcsr.2015.02.00023]. I think combining this with the Statin-Sartan Protocol, which is hypothesized to partially block differentiation whilst selecting for the expansion of the smallest, more multipotent cells, would be highly synergistic. AFA seems to be the most effective of the proposed substances, and this supplement also has fucoidan (https://www.stemenhance.co.uk/cerule-stemenhance-ultra).

 

I found two supplements in amazon from reputable brands, one with 30% triterpenes in phytosome(60mg per pill) around 13$ for 60 pills(Swanson phytosome gotu kola), and one without phytosomes but 35% triterpenes(257mg per pill) 30$ 30 pills(Life extension arterial protect)

 

Also

 

 

During prolonged treatment, especially with higher doses, the metabolism of active constituents slows down and can produce toxicity, so it was suggested that this pharmacokinetic phenomena should be considered during pharmacotherapy for effective and safe treatment[78]. The use of CA for more than 6 weeks is not recommended in the literature. People taking the herb for an extended period of time (up to 6 weeks) should take a 2-week break before taking the herb again. The standardized CA extracts and asiaticoside were well tolerated in experimental animals especially by oral route. Asiaticoside did not show any sign of toxicity up to the dose of 1 mg/kg after oral administration, whereas the toxic dose by intramuscular application reported for mice and rabbits was 40-50 mg/kg[79].

Pharmacological Review on Centella asiatica: A Potential Herbal Cure-all (nih.gov)

 

It appears that gotu kola needs to be cycled and can't be taken continually

Edited by Castiel, 03 January 2021 - 03:48 AM.

[QuestforLife]

The problem is IIRC Bill Andrews suggested the average telomere length when you're under 20 is already enough to produce a cancer that can grow big enough to kill you even without telomerase.
According to several sources, one of the things that's downregulated is the rate of protein turnover with aging. Reduced protein turnover leads in turn to the accumulation of damaged proteins and cellular dysfunction. This reduction in protein replacement rate is reversed with restored telomere length.

The cells are not adapting to a bad situation, they're just purposely making everything worse, they're downregulating mitochondrial maintenance, protein maintenance, and in the case of senescent cells releasing protumor formation and proinflammatory toxic chemicals to their surrounding.

(PDF) Protein Turnover in Aging and Longevity (researchgate.net)

The cell is actively trying to worsen the health of the organism and lead to it being eaten or killed by disease

It's like some companies and their product's designed with specific lifetime, designed to wear out within a specific period, to be replaced by newer models. The rate of evolution is increased by killing animals and replacing them with a new wave of gene mixed babies. But signal that food is scarce, the babies might be dying, and the organism will postpone the end of its shelf life, calorie restriction.

Yes, Bill Andrews has said that even in aged individuals telomere length is sufficient to cause cancer. I would like to see more data on that. But based on the reading I've done (there is a reference to this a few posts up, I'll link to it later when I'm not using my phone) aged individuals have SOME cells with long telomeres and some cells with short telomeres, just like young individuals. But in the old MORE cells have short telomeres and LESS cells have long telomeres. It is important to look not at individual cell telomere length but at the distribution. If you look at a telomere length result from Lifelength you can see what I mean. So that means that just because the old have SOME cells that have long enough telomeres to form a large enough lump to turn malignant doesn't mean this is likely due to their smaller number. It is far more likely such individuals will escape cancer until a large number of cells are becoming arrested simultaneously due replicative senescence. Hence it still looks to me like a balance of risks.

But like I said, more data could settle this, and the jury is still out on whether this is due to group selection for a given species aging rate, or a genuine effort to minimise cancer in the young balanced against aging in the old.

I'm in total agreement with you on protein turnover, it IS function of telomere length (or rather a function of relative telomere length compared to the telomere length in the young). Taking rapamycin or starving yourself is just a way to make the old cells act a little bit more like young cells.

Edited by QuestforLife, 03 January 2021 - 08:18 AM.

[QuestforLife]

I found two supplements in amazon from reputable brands, one with 30% triterpenes in phytosome(60mg per pill) around 13$ for 60 pills(Swanson phytosome gotu kola), and one without phytosomes but 35% triterpenes(257mg per pill) 30$ 30 pills(Life extension arterial protect)

Also
Pharmacological Review on Centella asiatica: A Potential Herbal Cure-all (nih.gov)

It appears that gotu kola needs to be cycled and can't be taken continually

Interesting that it builds up in the system. Even if you're not worried about toxicity you'd want to take a break to stay in the optimal range for telomerase activation. I must admit I didn't like that in the paper I referenced a larger dose produced less telomerase activation. Makes dosing tricky. An alternative might be just to use a triterpene supplement with a lower asiaticoside content and use that continuously.

[Castiel]

Yes, Bill Andrews has said that even in aged individuals telomere length is sufficient to cause cancer. I would like to see more data on that. But based on the reading I've done (there is a reference to this a few posts up, I'll link to it later when I'm not using my phone) aged individuals have SOME cells with long telomeres and some cells with short telomeres, just like young individuals. But in the old MORE cells have short telomeres and LESS cells have long telomeres. It is important to look not at individual cell telomere length but at the distribution. If you look at a telomere length result from Lifelength you can see what I mean. So that means that just because the old have SOME cells that have long enough telomeres to form a large enough lump to turn malignant doesn't mean this is likely due to their smaller number. It is far more likely such individuals will escape cancer until a large number of cells are becoming arrested simultaneously due replicative senescence. Hence it still looks to me like a balance of risks.

But like I said, more data could settle this, and the jury is still out on whether this is due to group selection for a given species aging rate, or a genuine effort to minimise cancer in the young balanced against aging in the old.

I'm in total agreement with you on protein turnover, it IS function of telomere length (or rather a function of relative telomere length compared to the telomere length in the young). Taking rapamycin or starving yourself is just a way to make the old cells act a little bit more like young cells.

  What about the mice, which were engineered to have significantly longer telomeres from birth having less cancer rate and longer lifespan and greater health?   Mice tend to be more cancer prone than humans, giving them longer telomeres from birth rather than increasing their cancer rate reduced it.

 

Given this was done early on most cells would have longer telomeres.

[Castiel]

Interesting that it builds up in the system. Even if you're not worried about toxicity you'd want to take a break to stay in the optimal range for telomerase activation. I must admit I didn't like that in the paper I referenced a larger dose produced less telomerase activation. Makes dosing tricky. An alternative might be just to use a triterpene supplement with a lower asiaticoside content and use that continuously.

 

We would have to see what percent standardized the experiments with goku kola have used.

 

In any case I would be cautious of using it continuously.

 

Even a month long seems like it might be pushing it.    I would probably take for two weeks and then a two week break.

 

Also it would be interesting to see how much HeLa telomerase affects telomeres, yes they are immortal, but is 100% of HeLa telomerase activity needed for immortalization?   Or is their telomerase activity overshooting the needed activity value?

 

639.full.pdf (aacrjournals.org)

 

It appears 50% telomerase inhibition of HeLa is enough to cause cell cycle arrest.   

 

But there is also the issue of how rapidly do HeLa cells divide

 

 

HeLa cells are rapidly dividing cancer cells, and the number of chromosomes varied during cancer formation and cell culture.-wiki

 

Would cells with slower rate of division be able to make due with lower level of telomerase activity and be immortalized?

 

In any case I'd be interested to see what Bill Andrews comments on the paper describing gotu kola and other telomerase activators.      There were other molecules with various levels of telomerase activation, and seeing what's different between the molecules might give ideas to promising changes to further increase telomerase activation.

[Castiel]

 

 

 Stabilization of telomeric G4 leads to telomere dysfunctions demonstrated by telomere shortening or damage, resulting in genome instability and apoptosis. Chemical compounds targeting G4 structures have been reported to induce telomere disturbance and tumor suppression... Emodin was identified as one of the best candidates, showing a great G4-binding potential. Subsequently, we confirmed that emodin could stabilize G4 structures in vitro and trigger telomere dysfunctions including fragile telomeres, telomere loss, and telomeric DNA damage. 

Combined treatment with emodin and a telomerase inhibitor induces significant telomere damage/dysfunction and cell death | Cell Death & Disease (nature.com)

 

Hmmm, that doesn't sound good for low purity resveratrol supplements.  

 

It seems high purity or emodin free is the way to go.

[QuestforLife]

Also it would be interesting to see how much HeLa telomerase affects telomeres, yes they are immortal, but is 100% of HeLa telomerase activity needed for immortalization? Or is their telomerase activity overshooting the needed activity value?

639.full.pdf (aacrjournals.org)

It appears 50% telomerase inhibition of HeLa is enough to cause cell cycle arrest.

But there is also the issue of how rapidly do HeLa cells divide

Would cells with slower rate of division be able to make due with lower level of telomerase activity and be immortalized?

In any case I'd be interested to see what Bill Andrews comments on the paper describing gotu kola and other telomerase activators. There were other molecules with various levels of telomerase activation, and seeing what's different between the molecules might give ideas to promising changes to further increase telomerase activation.

Have you seen this paper before?

https://pubmed.ncbi....h.gov/31761987/

'Effect of Peptide AEDG on Telomere Length and Mitotic Index of PHA-Stimulated Human Blood Lymphocytes'

Fig1 shows some T cells cultured with epitalon got longer telomeres, and some got shorter. Looking at Table 1 you can see this was due to the change in mitosis. The fastest dividing samples tended to shorten telomere length, even with epitalon.

So sometimes if a telomerase activator increases mitosis, it can decrease telomere length. You see the same with ROCK inhibitors. They allow continued division, but often you even see a decline in telomere length before stabilisation because of the renewed division.

Similarly, bone marrow transplant patients often have dramatic telomere shortening as the hematopoietic system is reconstituted, only for it to recover later. This suggests to me that Hematopoietic Stem Cells have some level of active telomerase, but not enough for the fastest division.

HELA might indeed be an upper limit to what is required to prevent telomere shortening in most tissues.

Bill Andrews has said many odd things about telomerase activtors, including: cycloastragenol doesn't activate telomerase so TA-65 but be something else and Epitalon doesn't lengthen telomeres. I assume it must be something to do with his assay. I can't believe he would say such things for financial gain.

I've never heard him say anything about asiaticoside.

Edited by QuestforLife, 04 January 2021 - 09:04 AM.

[Castiel]

Have you seen this paper before?

https://pubmed.ncbi....h.gov/31761987/

'Effect of Peptide AEDG on Telomere Length and Mitotic Index of PHA-Stimulated Human Blood Lymphocytes'

Fig1 shows some T cells cultured with epitalon got longer telomeres, and some got shorter. Looking at Table 1 you can see this was due to the change in mitosis. The fastest dividing samples tended to shorten telomere length, even with epitalon.

So sometimes if a telomerase activator increases mitosis, it can decrease telomere length. You see the same with ROCK inhibitors. They allow continued division, but often you even see a decline in telomere length before stabilisation because of the renewed division.

Similarly, bone marrow transplant patients often have dramatic telomere shortening as the hematopoietic system is reconstituted, only for it to recover later. This suggests to me that Hematopoietic Stem Cells have some level of active telomerase, but not enough for the fastest division.

HELA might indeed be an upper limit to what is required to prevent telomere shortening in most tissues.

Bill Andrews has said many odd things about telomerase activtors, including: cycloastragenol doesn't activate telomerase so TA-65 but be something else and Epitalon doesn't lengthen telomeres. I assume it must be something to do with his assay. I can't believe he would say such things for financial gain.

I've never heard him say anything about asiaticoside.

 

 

I hadn't seen that paper before.

 

Regards Bill Andrews, I wonder what sort of assay he's doing.   Hopefully he's not testing against the actual telomerase molecule with a snippet of telomeres.     Many molecules might alter cellular regulation such that another molecule is the one that actually activates telomerase.

[QuestforLife]

  What about the mice, which were engineered to have significantly longer telomeres from birth having less cancer rate and longer lifespan and greater health?   Mice tend to be more cancer prone than humans, giving them longer telomeres from birth rather than increasing their cancer rate reduced it.

 

Given this was done early on most cells would have longer telomeres.

 

The Blasco paper is one of the most important and interesting papers I've read in a long time. 

 

https://www.nature.c...467-019-12664-x

 

Mice with hyper-long telomeres show less metabolic aging and longer lifespans

 

 Short telomeres trigger age-related pathologies and shorter lifespans in mice and humans. In the past, we generated mouse embryonic (ES) cells with longer telomeres than normal (hyper-long telomeres) in the absence of genetic manipulations, which contributed to all mouse tissues. To address whether hyper-long telomeres have deleterious effects, we generated mice in which 100% of their cells are derived from hyper-long telomere ES cells. We observe that these mice have longer telomeres and less DNA damage with aging. Hyper-long telomere mice are lean and show low cholesterol and LDL levels, as well as improved glucose and insulin tolerance. Hyper-long telomere mice also have less incidence of cancer and an increased longevity. These findings demonstrate that longer telomeres than normal in a given species are not deleterious but instead, show beneficial effects.

 

They basically cultured ESCs for longer than would normally be the case in the womb, and because telomerase is so active in such cells, it resulted in an entire body with longer telomeres. It avoided any confounding effects with gene therapy or telomerase activators. Of note it  showed that longer telomeres led to lower DNA damage (and presumably mutations), which is interesting given longer telomeres enable more mitosis, and more mitosis generally leads to more mutations. 

 

But there are so many unanswered questions. 

 

Did the longer telomeres allow more cell divisions and this is what allowed a longer, healthier life? Or was it simply the longer telomeres than gave the benefits (such as lower mutations) and the cell divisions were roughly the same? We know the hyperlong telomere mice had less fat, and tended to be slightly smaller, so perhaps the latter answer is correct. 

 

Did the mice telomeres shorten longitudinally at the same rate as a 'normal' lab mouse with (relatively) shorter telomeres? Did they suffer from the same type of diseases as 'normal' mice, but just at a later date (we only know cancer incidence was lower).

 

'Normal' lab mice have freakishly long telomeres compared to wild mice, and yet this doesn't seem to benefit lifespan. So there are lots of unanswered questions. Hopefully Blasco and co are on this, and we'll get a lot of very long standing questions about telomeres answered. 

[Castiel]

The Blasco paper is one of the most important and interesting papers I've read in a long time. 

 

https://www.nature.c...467-019-12664-x

 

Mice with hyper-long telomeres show less metabolic aging and longer lifespans

 

 

 

 

They basically cultured ESCs for longer than would normally be the case in the womb, and because telomerase is so active in such cells, it resulted in an entire body with longer telomeres. It avoided any confounding effects with gene therapy or telomerase activators. Of note it  showed that longer telomeres led to lower DNA damage (and presumably mutations), which is interesting given longer telomeres enable more mitosis, and more mitosis generally leads to more mutations. 

 

But there are so many unanswered questions. 

 

Did the longer telomeres allow more cell divisions and this is what allowed a longer, healthier life? Or was it simply the longer telomeres than gave the benefits (such as lower mutations) and the cell divisions were roughly the same? We know the hyperlong telomere mice had less fat, and tended to be slightly smaller, so perhaps the latter answer is correct. 

 

Did the mice telomeres shorten longitudinally at the same rate as a 'normal' lab mouse with (relatively) shorter telomeres? Did they suffer from the same type of diseases as 'normal' mice, but just at a later date (we only know cancer incidence was lower).

 

'Normal' lab mice have freakishly long telomeres compared to wild mice, and yet this doesn't seem to benefit lifespan. So there are lots of unanswered questions. Hopefully Blasco and co are on this, and we'll get a lot of very long standing questions about telomeres answered. 

 

Normal mice have very long telomeres, but humans reduce telomeres at 70Kb per division while mice reduce telomeres at 7000Kb per division, iirc.  Their rate of telomere shortening is far far greater, and telomere shortening has effects on gene expression long before they become critically short.   And rate of telomere shortening as you well know is correlated with maximum species lifespan.

 

We know of the experiment were mice were allowed to suffer from telomere shortening, and how activating telomerase restored health.

Telomerase reverses ageing process

 

 

Dramatic rejuvenation of prematurely aged mice hints at potential therapy.

Mice engineered to lack the enzyme, called telomerase, become prematurely decrepit. But they bounced back to health when the enzyme was replaced. The finding, published online today in Nature¹, hints that some disorders characterized by early ageing could be treated by boosting telomerase activity.

 

Telomerase reverses ageing process : Nature News

 

 

It seems another experiment conducted later showed that inducing telomerase in adult and in old mice improved health, fitness, longevity without increasing cancer rate either.

 

 

A major goal in aging research is to improve health during aging. In the case of mice, genetic manipulations that shorten or lengthen telomeres result, respectively, in decreased or increased longevity. Based on this, we have tested the effects of a telomerase gene therapy in adult (1 year of age) and old (2 years of age) mice. Treatment of 1- and 2-year old mice with an adeno associated virus (AAV) of wide tropism expressing mouse TERT had remarkable beneficial effects on health and fitness, including insulin sensitivity, osteoporosis, neuromuscular coordination and several molecular biomarkers of aging. Importantly, telomerase-treated mice did not develop more cancer than their control littermates, suggesting that the known tumorigenic activity of telomerase is severely decreased when expressed in adult or old organisms using AAV vectors. Finally, telomerase-treated mice, both at 1-year and at 2-year of age, had an increase in median lifespan of 24 and 13%, respectively. These beneficial effects were not observed with a catalytically inactive TERT, demonstrating that they require telomerase activity. Together, these results constitute a proof-of-principle of a role of TERT in delaying physiological aging and extending longevity in normal mice through a telomerase-based treatment, and demonstrate the feasibility of anti-aging gene therapy.

 

Telomerase gene therapy in adult and old mice delays aging and increases longevity without increasing cancer (nih.gov)

[QuestforLife]

Normal mice have very long telomeres, but humans reduce telomeres at 70Kb per division while mice reduce telomeres at 7000Kb per division, iirc.  Their rate of telomere shortening is far far greater, and telomere shortening has effects on gene expression long before they become critically short.   And rate of telomere shortening as you well know is correlated with maximum species lifespan.

 

 

 

Yes, this is akin to having an inflatable life raft - having a higher telomerase level means you you top up the air faster, but if there are lots of holes in the dingy you're still going to end up losing air fast.

 

So mice are pumping up their life raft continuously, but still losing 'air'  100x faster than humans, who barely top up their air at all. It is likely that having lots of holes in the dingy is the equivalent of 'premature' senescence inducers like ROS (from various causes). It is not clear what the proportion is in humans between premature senescence and that induced by telomere shortening . But in any cases premature senescence will have a knock on effect on telomere length as well, as it will necessitate further division in the cells still capable of doing so. And of course this is even further complicated by the fact that senescent cells that persist can then arrest the division of nearby otherwise healthy cells whatever their telomere length. So I ultimately think that telomere shortening cannot be solved by telomere lengthening only (or at least not very efficiently), but should be paired with a treatment or compensation for premature senescence. But I digress. 

 

I think the reason mice suffer ill health with this loss of telomere length ( or 'air' in our analogy), is because multiple genes are affected in their expression with the configuration of the chromosome, which is altered depending on the length of the telomere. Even if the telomere never gets anywhere near senescent length, gene expression is nonetheless altered as the few genes affected by the shortening telomere then go on to influence the expression of many more. In this way the whole orchestra gradually loses its togetherness and the symphony becomes noise.

 

Long-range telomere regulation of gene expression: Telomere looping and telomere position effect over long distances (TPE-OLD)

 

This paper focusses on how telomere length regulates the expression of telomerase itself (so it is a self control mechanism, effectively) 

 

 The human cellular reverse transcriptase, telomerase, is very tightly regulated in large long-lived species. Telomerase is expressed during early human fetal development, is turned off in most adult tissues, and then becomes reactivated in almost all human cancers. However, the exact mechanism regulating these switches in expression are not known. We recently described a phenomenon where genes are regulated by telomere length dependent loops (telomere position effects over long distances; TPE-OLD). The hTERT gene is ~ 1.2Mb 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. These findings have implications for how cells turn off telomerase when telomeres are long during fetal development and how cancer cells reactivate telomerase in cells that have short telomeres. In addition to TPE-OLD, in proliferating stem cells such as activated T-lymphocytes, telomerase can be reversibly activated and silenced by telomere looping. In telomerase positive cancer cells that are induced to differentiate and downregulate telomerase, telomere looping correlates with silencing of the hTERT gene. These studies and others support a role of telomeres in regulating gene expression via telomere looping that may involve interactions with internal telomeric sequences (ITS). In addition to telomere looping, TPE-OLD may be one mechanism of how cells time changes in physiology without initiating a DNA damage response.

 

Source: https://www.ncbi.nlm...les/PMC5826875/

Edited by QuestforLife, 06 January 2021 - 03:05 PM.

[Castiel]

Yes, this is akin to having an inflatable life raft - having a higher telomerase level means you you top up the air faster, but if there are lots of holes in the dingy you're still going to end up losing air fast.

 

So mice are pumping up their life raft continuously, but still losing 'air'  100x faster than humans, who barely top up their air at all. It is likely that having lots of holes in the dingy is the equivalent of 'premature' senescence inducers like ROS (from various causes). It is not clear what the proportion is in humans between premature senescence and that induced by telomere shortening . But in any cases premature senescence will have a knock on effect on telomere length as well, as it will necessitate further division in the cells still capable of doing so. And of course this is even further complicated by the fact that senescent cells that persist can then arrest the division of nearby otherwise healthy cells whatever their telomere length. So I ultimately think that telomere shortening cannot be solved by telomere lengthening only (or at least not very efficiently), but should be paired with a treatment or compensation for premature senescence. But I digress. 

 

I think the reason mice suffer ill health with this loss of telomere length ( or 'air' in our analogy), is because multiple genes are affected in their expression with the configuration of the chromosome, which is altered depending on the length of the telomere. Even if the telomere never gets anywhere near senescent length, gene expression is nonetheless altered as the few genes affected by the shortening telomere then go on to influence the expression of many more. In this way the whole orchestra gradually loses its togetherness and the symphony becomes noise.

 

Long-range telomere regulation of gene expression: Telomere looping and telomere position effect over long distances (TPE-OLD)

 

This paper focusses on how telomere length regulates the expression of telomerase itself (so it is a self control mechanism, effectively) 

 

 

Source: https://www.ncbi.nlm...les/PMC5826875/

 

The fact senescent cells can be made regardless of telomere length due to damage, means they will likely accumulate and do harm even if telomerase positive.

 

I think cancer immunity is possible to evolve, some humans have it, and the lack of telomerase is just another thing to stop biological immortality from occurring from some accidental mutation.   Many biologically immortal species are telomerase positive, so I do have doubts that telomerase is problematic in the right context.

[aribadabar]

Normal mice have very long telomeres, but humans reduce telomeres at 70Kb per division while mice reduce telomeres at 7000Kb per division, iirc. 

 

I have seen this 70 kb mentioned often but why these supposedly faster TRF losers have shown only ~28bp /yr loss?

[QuestforLife]

I have seen this 70 kb mentioned often but why these supposedly faster TRF losers have shown only ~28bp /yr loss?

 

You are right, it warrants further investigation.

 

For example this paper from 2013 shows an attrition rate of 26bp/yr for leukocytes, 24bp/yr for muscle cells, 23bp/yr for skin and 25bp/yr for fat (i.e. an equivalent rate). This paper from 2020 establishes that age is the biggest factor for relative telomere change between people, though the strength of the correlation varies between tissues as does the rate. Eyeballing Fig 3c for leukocytes the relative length changes by 0.3 over 50 years or 6% per year (between ages 20 and 70). The question is: what is leukocyte TL at age 20? My Lifelength TL report claims 12kb to be average@age 20, which indicates a 72bp/year loss (0.006x12,000).

 

72bp/yr also matches up better to losing around 25% of TL by mean life expectancy and 50% of TL by max life expectancy, see Blasco, here

Which at 72bp/year suggests mean life expectancy of a human with 12kbp @age 20 would be ~62 years and max LS would be ~103 years. The fact we can add around 20 years to both these figures can probably be put down to medical care keeping people alive who otherwise would have died.

 

So why do some studies come back with ~25bp/year? A couple of educated guesses: small sample size, especially at older ages means you get an underestimate of TL attrition from young to old; the fact that correlation between TL and age is not that high (even though it is the single biggest contributor) because of other confounding factors like variations in inherited length, lifestyle/inflammation, various SNPs that influence TL maintenance through telomerase. Also, measurement error can cause the contribution of age to TL to be underestimated. This means errors in fitting the curve can easily give you serious errors in the gradient. A prime example is Fig A from your reference, how do you fit a line of best fit to that?

Edited by QuestforLife, 08 January 2021 - 12:10 PM.

[aribadabar]

You are right, it warrants further investigation.

 

For example this paper from 2013 shows an attrition rate of 26bp/yr for leukocytes, 24bp/yr for muscle cells, 23bp/yr for skin and 25bp/yr for fat (i.e. an equivalent rate). This paper from 2020 establishes that age is the biggest factor for relative telomere change between people, though the strength of the correlation varies between tissues as does the rate. Eyeballing Fig 3c for leukocytes the relative length changes by 0.3 over 50 years or 6% per year (between ages 20 and 70). The question is: what is leukocyte TL at age 20? My Lifelength TL report claims 12kb to be average@age 20, which indicates a 72bp/year loss (0.006x12,000).

 

72bp/yr also matches up better to losing around 25% of TL by mean life expectancy and 50% of TL by max life expectancy, see Blasco, here

Which at 72bp/year suggests mean life expectancy of a human with 12kbp @age 20 would be ~62 years and max LS would be ~103 years. The fact we can add around 20 years to both these figures can probably be put down to medical care keeping people alive who otherwise would have died.

 

So why do some studies come back with ~25bp/year? A couple of educated guesses: small sample size, especially at older ages means you get an underestimate of TL attrition from young to old; the fact that correlation between TL and age is not that high (even though it is the single biggest contributor) because of other confounding factors like variations in inherited length, lifestyle/inflammation, various SNPs that influence TL maintenance through telomerase. Also, measurement error can cause the contribution of age to TL to be underestimated. This means errors in fitting the curve can easily give you serious errors in the gradient. A prime example is Fig A from your reference, how do you fit a line of best fit to that?

 

Excellent points!

 

Minor calc note : 6% is 0.06 (not 0.006) so that should be 720bp/yr!

My guess is that the TL attrition is not linear and accelerates with age ( or , as you mentioned earlier, the shorter and shorter telomeres trigger profound differences in gene expression thus same or similar shortening leads to bigger and bigger changes as the age progresses).

Edited by aribadabar, 08 January 2021 - 01:32 PM.

[QuestforLife]

Excellent points!

 

Minor calc note : 6% is 0.06 (not 0.006) so that should be 720bp/yr!

My guess is that the TL attrition is not linear and accelerates with age ( or , as you mentioned earlier, the shorter and shorter telomeres trigger profound differences in gene expression thus same or similar shortening leads to bigger and bigger changes as the age progresses).

 

The calculation is not wrong, but I should have written 0.6%/year.

 

Losing 0.3 of TL (compared to age 20) over the 50 year age gap of the study subjects is 0.3/50 = 0.006 loss per year (or 0.6%, sorry for the confusion).

 

If 1 is TL@age 20 = 12,000bp, then 0.006 x 12,000 = 72bp loss/year.

 

You may be right that it is not exactly linear, i.e. if turnover falls then TL attrition should also fall - but one would expect this to be significant  only late in life

[QuestforLife]

More on the Selfish Cell theory of aging

 

Aging may be due to the ‘locking in’ of cells whose plasticity is required for regeneration and repair. This appears to be caused by the very signals that trigger that repair. For example muscle satellite cells undergo DNA damage in order to differentiate. This is a ‘sacrifice’ made for the good of the body. But with aging we see more and more cells locked in such a state [1]. One might wonder why the body would have such a flaw, but this is answered by considering the alternative - if cells were too resistant to these signals, then they would not differentiate and we would not develop in the first place. This would result in a failed fetus if it occurred from conception. Or else if it developed gradually during life as a result of natural selection of those stem cells that are more resistant to differentiation, this would result in aging. Therefore aging is the result of the Selfish Cell. A cell such as this has acquired immortality for itself, for example through greater telomerase activation in that cell, which we might think beneficial - but this is a misinterpretation, as the stem cell has kept this ability only because it is not subject to the normal controls that occur upon differentiation (i.e. loss or downregulation of telomerase expression). An example of this might be the natural selection in the bone marrow of cells that are not helpful to the human body. Indeed, experimenters have done this for themselves with mice stem cells, and serial transplantation revealed that such cells are immortal, but do not differentiate into the required red blood progenitors (for example), and hence the recipient mice die unless they have sufficient ‘normal’ stem cells that are willing to differentiate [2]. We also see this in the skin, where the stem cells that divide symmetrically come to dominate the epidermis, but this is bad for the skin as they are more stress resistant and less likely to (divide asymmetrically and) differentiate into the required cells to renew the upper layer of the skin [3].

 

[1] Caspase 3/caspase-activated DNase promote cell differentiation by inducing DNA strand breaks, https://www.pnas.org...tent/107/9/4230

[2] Loss of Dnmt3a Immortalizes Hematopoietic Stem Cells In Vivo, https://www.ncbi.nlm...les/PMC5908249/

[3] Stem cell competition orchestrates skin homeostasis and ageing, https://www.nature.c...586-019-1085-7 

Edited by QuestforLife, 15 January 2021 - 10:04 AM.

[Castiel]

More on the Selfish Cell theory of aging

 

Aging may be due to the ‘locking in’ of cells whose plasticity is required for regeneration and repair. This appears to be caused by the very signals that trigger that repair. For example muscle satellite cells undergo DNA damage in order to differentiate. This is a ‘sacrifice’ made for the good of the body. But with aging we see more and more cells locked in such a state [1]. One might wonder why the body would have such a flaw, but this is answered by considering the alternative - if cells were too resistant to these signals, then they would not differentiate and we would not develop in the first place. This would result in a failed fetus if it occurred from conception. Or else if it developed gradually during life as a result of natural selection of those stem cells that are more resistant to differentiation, this would result in aging. Therefore aging is the result of the Selfish Cell. A cell such as this has acquired immortality for itself, for example through greater telomerase activation in that cell, which we might think beneficial - but this is a misinterpretation, as the stem cell has kept this ability only because it is not subject to the normal controls that occur upon differentiation (i.e. loss or downregulation of telomerase expression). An example of this might be the natural selection in the bone marrow of cells that are not helpful to the human body. Indeed, experimenters have done this for themselves with mice stem cells, and serial transplantation revealed that such cells are immortal, but do not differentiate into the required red blood progenitors (for example), and hence the recipient mice die unless they have sufficient ‘normal’ stem cells that are willing to differentiate [2]. We also see this in the skin, where the stem cells that divide symmetrically come to dominate the epidermis, but this is bad for the skin as they are more stress resistant and less likely to (divide asymmetrically and) differentiate into the required cells to renew the upper layer of the skin [3].

 

[1] Caspase 3/caspase-activated DNase promote cell differentiation by inducing DNA strand breaks, https://www.pnas.org...tent/107/9/4230

[2] Loss of Dnmt3a Immortalizes Hematopoietic Stem Cells In Vivo, https://www.ncbi.nlm...les/PMC5908249/

[3] Stem cell competition orchestrates skin homeostasis and ageing, https://www.nature.c...586-019-1085-7 

 

Whatever the mechanism I think programmed aging is likely the real reason why most of it happens.

 

Have you read the comments of Harold Lawrence Katcher in Josh Mitteldorf's blog?   IIRC he claims by isolating substances from young blood and injecting them into old mice he has been able to fully reset epigenetic age.

[QuestforLife]

Comments on Hallmarks of heterochronic parabiosis

Aging (or old blood) affects the gene expression levels of different cells differently (they have different ‘Differentially Expressed Genes’ (DAG)), but there is a lot of agreement between the cell-specific DEGs for both aging and old blood. For some cells, the agreement is very high indeed (~0.6-0.84 for Endothelial Cells, Hematopoietic Stem Cells, Mesenchymal Stem Cells in Adipose Tissue) suggesting that these cells in particular primarily age extrinsically (i.e. via factors in the blood).
Note this does not mean other cells do not age intrinsically, of course.

Later on in the paper they talk about how some cells age in co-ordination with the tissue in which they reside, whereas other cell types ignore the tissue they’re in and instead age more in step with the tissue they come from (i.e. some bone marrow cells).
By contrast to the cell-type specific changes with age (or old blood), the rejuvenation provided by young blood seems to act on different cells in mostly the same way, via the mitochondrial pathways of the electron transport chain*.

*Weird exception that may prove the rule: brain endothelial cells (BECs) have been observed to undergo increased expression of electron transport chain genes with age. This effect is replicated by exposure to aged mouse plasma and reversed by exposure to young mouse plasma in vivo (ref: https://www.cell.com...81?showall=true Brain Endothelial Cells Are Exquisite Sensors of Age-Related Circulatory Cues).
My note: this suggests the solution to the general downregulation of the electron transport chain is not merely ETC upregulation, as this would be counterproductive to BECs, but instead a single or multiple corrections in the blood that either up- or down- regulates ETC function towards a youthful state as required through some indirect mechanism.

Some further thoughts on the problem: given we know the alteration of ETC function is via extrinsic factors in the blood, it would probably be a mistake to think mitochondria can be fixed via some mitochondrial specific molecule such as ALA or Q10. It makes more sense to assume there is some sort of problem in circulation such that mitochondria are not getting enough of something they need or are getting too much or something they do not need. So for example increasing circulation might give the mitochondria more of whatever it is they need that is present in the blood, or breaking down some sort of blood borne toxin might be the key to avoiding overloading the mitochondria with that toxic compound. This marks a significant change in the approach to reversing aging in that we are treating the blood and the circulation first and the cell second. It bears some similarity to the approach of Janic et al. in their short term statin-sartan treatment for pre-clinical atherosclerosis, discussed at length in this thread (for those that haven’t read it already it is one of the keystones of this thread: https://pubmed.ncbi.....gov/26214555/) .

One other thing that occurs to me from the paper when the Conboys replaced half the blood plasma with saline plus albumin and produced significant rejuvenation (https://www.ncbi.nlm...les/PMC7288913/), is that we mustn’t dismiss the importance of those blood proteins. For example, albumin, quite apart from its liver functions is an antioxidant buffer in the blood (https://pubmed.ncbi....h.gov/21113488/), which should help mitochondria, particularly when in the procedure you are replacing partly oxidised with fully reduced albumin. Fibrinogen is an acknowledged 'damage associated molecular pattern protein' (DAMP) (https://journals.lww..._Induced.7.aspx) and could also impede rejuvenation via decreased blood flow when it causes blockages in capillaries (that are not severe enough for a stroke). Transferrin moves iron, of which the main intracellular recipient is mitochondria, and intracellular transferrin is known to increase in Parkinson’s Disease (https://www.scienced...357272519300433), which also implicates it in mitochondrial dysfunction. Finally, immunoglobulins – the immune system is known to react to pieces of mitochondrial DNA in the blood (https://pubmed.ncbi....h.gov/29125070/), leading to the rise in age related ‘sterile’ inflammation. At the very least these arguments suggest blood protein removal should be eliminated as a cause of aging in these parabiosis studies.

So where does this leave telomeres as a valid point of intervention in the aging process? This paper is particularly interesting as it shows how complex the age-related changes in genes are. Different genes are up or down regulated depending on tissue and cell-type. Much like the real situation with telomeres. At any given point there are cells with nice long telomeres and cells with short telomeres, sometimes living right alongside each other. Some tissues seem to maintain telomeres better than others. Even in a very old person there are cells with long telomeres that you can extract and passage in the lab. Sometimes a cell has long telomeres but is rendered functionally arrested because of mitochondrial damage. All these arguments have been used to dismiss telomeres as an important cause of aging. But the real picture is much more complex.

Now that we can see the body has a kind of aging imposed on much of it by systemic factors, it is possible to imagine that the machine just needs an oil change to make it as good as new. That would be wonderful. And indeed, this would not contradict the telomere story. How many times have I posted about ‘conditional reprogramming’ whereby old somatic cells with the right culture conditions would revert to a progenitor state and proliferate endlessly so long as the culture conditions were maintained? (for recap: https://linkinghub.e...2944013005944).Note that these culture conditions included telomerase. Perhaps young blood does something similar? Indeed, it is a central tenet of this thread that the statin-sartan treatment of Janic et al is doing just that. This is a throwback to the pre-Hayflick days when scientists though cells were immortal; it was just the body that aged. Perhaps they were not so wrong after all. With the right culture conditions.

I've been commenting on Josh's site for years and as you can see Castiel, I discussed heterochronic parabiosis recently.

It remains to be seen how much of aging is due to systemic factors (i.e. cell extrinsic) and how much is due to actual cellular aging.

The argument of Katcher and others go that human aging isn't due to cellular aging exactly - and the Selfish Cell idea agrees, in that individual cells should die so the body can live.

It strikes me that if we were to try and design a long lasting multicellular body but didn't want individual cells to be immortal, setting up a hierarchy of cells from pluripotent right down to tissue specific cells is exactly what you'd do. Inevitably though with so many moving parts you'd get problems, such as the divergence of cellular and bodily interests I've outlined. Restoring a youthful systemic milleau in the blood might be a way to reset the calibration and get all the cells working properly together again. For example, factors like GDF11 might get 'locked in' progenitor cells differentiating again. Then we'd just be left with any intrinsic cellular aging, such as DNA mutations.

Edited by QuestforLife, 16 January 2021 - 09:36 AM.

[QuestforLife]

Pioglitazone (PG) can increase subcutaneous fat whilst decreasing visceral fat [1], which could provide many health benefits [2]. But PG is positively associated with bladder cancer risk [3].

Due to the mechanism of action of PG being forced proliferation of small, new adipocytes [4], I wondered if the elevated cancer risk might be secondary to attrition of telomeres. Short telomeres are independently associated with bladder cancer risk [5].

I found a study that showed that PG was useful for patients with depression, but that successful treatment was correlated with telomere length (TL) and possibly mediated via an increase in insulin sensitivity. This is what you would expect if the improvement in symptoms is caused by having more small, functional adipocytes to take up glucose [6].

This suggests to me that the necessary adipocyte proliferation can only occur successfully (and safely) if the TL of the underlying progenitors is sufficient. Note Leukocyte (blood or buccal) TL is reasonable proxy for TL in other tissues [7] so can be used to estimate the TL of progenitors.

What would the protocol be should one want to try it?
PG dose and duration as per study [1] and [4] respectively; 45mg/day for 12 weeks
Adjuvant to stop Bladder Cancer risk: telomerase activator. Choice asiaticoside or epitalon [8]; also the short duration of the trial.
Expected outcome: an increase in body weight of a couple of kilos, but with the increase coming from subcutaneous fat only.

 

References:
[1] https://pubmed.ncbi....h.gov/15562376/
https://diabetes.dia.../50/8/1863.long
[2] https://www.ncbi.nlm...les/PMC4092104/
[3] https://pubmed.ncbi....h.gov/30800561/
https://pubmed.ncbi....h.gov/29476615/
[4] https://onlinelibrar...38/oby.2009.380
[5] https://pubmed.ncbi....h.gov/17416776/
https://pubmed.ncbi....h.gov/15746160/
[6] https://www.ncbi.nlm...les/PMC5068869/
[7] https://science.scie...9/6509/eaaz6876
[8] https://www.ncbi.nlm...les/PMC6755196/
https://pubmed.ncbi....h.gov/12937682/
https://pubmed.ncbi....h.gov/15455129/

[QuestforLife]

I finally got the results back for my Sept TruMe test.

 

Chronological Age: 41.8

Biological Age:  36.7

 

There has been an improvement. It is not clear whether this is simply because I just took AKG for longer, or because of anything else. After considering adding berberine, ALA and carnitine at the half way stage, I decided not to give my body any help with fatty acid processing and instead just fed it more long chain fatty acids. I did this by eating a lot of cheddar and butter. The butter I clarified by gently heating it, and drained off the fat leaving the milk proteins in the pan. I then added stearic acid to the butter fat to make it more saturated. I used this to butter bread, butter pasta, cook pancakes, roast potatoes, bake cookies, etc. I found it quite palatable and it had the advantage of allowing me to eat copious amounts of carbs without putting on weight. Perhaps this did help my stem cell populations as I discussed in post 275#

 

I experienced increasing fatigue from the 4 1/2 month point. I also retrospectively noticed a considerable reduction in weight lifting strength between months 4 and 6. Again, I do not know if this was due to the AKG or the increased saturated fat intake. I quit AKG when I submitted my second test at the 6 month point. Since then I have continued eating the fatty diet. I have also upped my protein intake and my strength has started to recover. I have experienced fatigue at times from the diet that is ameliorated by carnitine and milk thistle. 

 

It would be helpful to do further testing but TruMe testing has slowed to a crawl due to the lockdown.

 

My original plan was to add back telomerase activators in addition to an effective epigenetic age reversal strategy to see if I could have the best of both worlds. Again that will depend on TruMe being able to accept and process samples relatively quickly. 

 

Latest TruAge results from Jan 2021

 

Chronological Age: 42.1

Biological Age 35.5

Delta 6.6 years

 

Biological age continued to fall from 36.7 to 35.5 years despite 4 months passing between tests.

 

How do I feel? Fine, nothing significant to report. The first traces of white in my beard continues to very slowly advance. The good news is that I have no fatigue this time and my weight lifting has not been affected like last time. 

 

What was different? I stopped my enhanced saturated fat diet in October. I am guessing this was responsible for the fatigue. I had a break from AKG from the start of Sept to mid November 2020. Then I started off on only 600mg/day AKG (kirkman labs salt) rather than previous 900mg/day, but this time alongside 500mg berberine. I reasoned AMPK activation would increase isocitrate dehydrogenase , which should increase the production of AKG in the krebs cycle. Interestingly, berberine caused no weight loss, which it has in the past. I did increase my AKG dose to 900mg/day from mid-December.

 

And telomerase activators? This is the most exciting result. I used asiaticoside from the start of November. My dose varied from 1-3 tablets of 120mg gotu kola/day (from Nootropics). They seemed to have the highest concentration of asiaticoside I could find and were able to supply me with a good certificate proving very low heavy meals content. This telomerase activator dose clearly did not interfere with continued improvement in my biological age as measured by methylation, which did occur when I used epitalon.

 

It may well be that asiaticoside is a much weaker telomerase activator than epitalon, but I've been unable to get a telomere test due to the continued lockdown in the UK. 

[dlewis1453]

This is a great result! Almost seven years of epigenetic age reversal for someone healthy in his early 40s is very impressive. I'm glad to see that your improvements made with AKG were durable, despite taking a break from AKG in the fall.

 

I'm very curious to see how low AKG can take you, and also how much length your telomeres have gained once you are finally able to test those again. 

 

Regarding the slow greying of your beard - my guess is that there would be a significant lag between epigenetic reprogramming of existing cells and visual changes. For example, people taking gdf11 in Steve's group and also on the GDF11 Facebook page have reported grey hair reversal, but it takes about a year of GDF11 supplementation before that happens. 

Edited by dlewis1453, 09 February 2021 - 05:51 PM.

[aribadabar]

Regarding the slow greying of your beard - my guess is that there would be a significant lag between epigenetic reprogramming of existing cells and visual changes. For example, people taking gdf11 in Steve's group and also on the GDF11 Facebook page have reported grey hair reversal, but it takes about a year of GDF11 supplementation before that happens. 

 

GDF-11 does NOT universally reverse hair graying - I have been taking it for a few years now without anything to show for it in the hair colour dept. That being said, I believe there is a strong genetic component in my case as I have been greying since my mid-20s.

[JamesPaul]

Latest TruAge results from Jan 2021

 

Chronological Age: 42.1

Biological Age 35.5

Delta 6.6 years

 

Biological age continued to fall from 36.7 to 35.5 years despite 4 months passing between tests.

 

... I stopped my enhanced saturated fat diet in October. I am guessing this was responsible for the fatigue. I had a break from AKG from the start of Sept to mid November 2020. Then I started off on only 600mg/day AKG (kirkman labs salt) rather than previous 900mg/day, but this time alongside 500mg berberine. I reasoned AMPK activation would increase isocitrate dehydrogenase , which should increase the production of AKG in the krebs cycle. Interestingly, berberine caused no weight loss, which it has in the past. I did increase my AKG dose to 900mg/day from mid-December.

 

And telomerase activators? This is the most exciting result. I used asiaticoside from the start of November. My dose varied from 1-3 tablets of 120mg gotu kola/day (from Nootropics)...

Thank you much for sharing these positive results with the protocol you developed!

A couple questions.  Since stearic acid promotes mitochondrial fusion, was the reason for fatigue that the mitochondrial quality control function was not being performed?

Also, this article

https://www.longevit...tive-therapies/

states "...researchers found that when CMA [chaperone-mediated autophagy] activity is minimised, the stem cells can maintain high levels of alpha-ketoglutarate, a metabolite that is vital for reinforcing a cell’s pluripotent state. When the cell begins the differentiation process, a reduction in Oct4 and Sox2 trigger the upregulation of CMA, which in turn degrades key enzymes responsible for the production of alpha-ketoglutarate...The chain reaction results in a reduction in alpha-ketoglutarate levels and an increase in other cellular activities that encourage cell differentiation."

That seems to mean, or could mean, that high levels of AKG inhibit stem cell differentiation, with the result that cells are not replaced as readily at their end of useful life, which would be undesirable on a long-term basis.

[Update:  I'm uncertain about this.  The paper https://pubmed.ncbi....h.gov/32652733/ says "An increase of α-ketoglutarate (α-KG), a cofactor for histone and DNA demethylases, triggers multilineage differentiation in human embryonic stem cells (hESCs)."

On the other hand, the paper https://pubmed.ncbi....h.gov/25487152/ states "...In vitro, supplementation with cell-permeable αKG directly supports ES-cell self-renewal while cell-permeable succinate promotes differentiation. This work reveals that intracellular αKG/succinate levels can contribute to the maintenance of cellular identity and have a mechanistic role in the transcriptional and epigenetic state of stem cells."

Also, the paper https://pubmed.ncbi....h.gov/27476977/ states “Moderate knockdown (KD) of Psat1 [phosphoserine aminotransferase 1] in mESCs lowered DNA 5'-hydroxymethylcytosine (5'-hmC) and increased histone methylation levels by downregulating permissive amounts of α-KG, ultimately accelerating differentiation.”] 

So I think that means that both stearic acid and AKG should be cycled.  Do you have any insight into how long optimal "consume" and "don't consume" periods should be?

 

I sent an email to Nootropics.com in the US and they replied they don't currently sell gotu kola.  Did you mean NootropicsDepot.com?  They do carry gotu kola in both tablet and powder form. 

 

 

 

Edited by JamesPaul, 13 February 2021 - 08:25 PM.

[JamesPaul]

I tried to shorten above post but permission was denied.  I tried to replace the text that begins "Also, this article..." with the following:

 

Also, some articles appear to state that high levels of alpha-ketoglutarate promote stem cell self-renewal or alternatively promote differentiation.  Either one could be undesirable if sustained over a long period of time.

 

The paper https://pubmed.ncbi....h.gov/32652733/ says "An increase of α-ketoglutarate (α-KG), a cofactor for histone and DNA demethylases, triggers multilineage differentiation in human embryonic stem cells (hESCs)."

On the other hand, the paper https://pubmed.ncbi....h.gov/25487152/ states "...In vitro, supplementation with cell-permeable αKG directly supports ES-cell self-renewal while cell-permeable succinate promotes differentiation. This work reveals that intracellular αKG/succinate levels can contribute to the maintenance of cellular identity and have a mechanistic role in the transcriptional and epigenetic state of stem cells."

Also, the paper https://pubmed.ncbi....h.gov/27476977/ states “Moderate knockdown (KD) of Psat1 [phosphoserine aminotransferase 1] in mESCs lowered DNA 5'-hydroxymethylcytosine (5'-hmC) and increased histone methylation levels by downregulating permissive amounts of α-KG, ultimately accelerating differentiation.”]

Sorry for the excessive length of the post.
 

Edited by JamesPaul, 13 February 2021 - 08:35 PM.

[QuestforLife]

I tried to shorten above post but permission was denied.  I tried to replace the text that begins "Also, this article..." with the following:

 

Also, some articles appear to state that high levels of alpha-ketoglutarate promote stem cell self-renewal or alternatively promote differentiation.  Either one could be undesirable if sustained over a long period of time.

 

The paper https://pubmed.ncbi....h.gov/32652733/ says "An increase of α-ketoglutarate (α-KG), a cofactor for histone and DNA demethylases, triggers multilineage differentiation in human embryonic stem cells (hESCs)."

On the other hand, the paper https://pubmed.ncbi....h.gov/25487152/ states "...In vitro, supplementation with cell-permeable αKG directly supports ES-cell self-renewal while cell-permeable succinate promotes differentiation. This work reveals that intracellular αKG/succinate levels can contribute to the maintenance of cellular identity and have a mechanistic role in the transcriptional and epigenetic state of stem cells."

Also, the paper https://pubmed.ncbi....h.gov/27476977/ states “Moderate knockdown (KD) of Psat1 [phosphoserine aminotransferase 1] in mESCs lowered DNA 5'-hydroxymethylcytosine (5'-hmC) and increased histone methylation levels by downregulating permissive amounts of α-KG, ultimately accelerating differentiation.”]

Sorry for the excessive length of the post.
 

 

We tend to look for papers that confirm our hypothesis, so it is good to see various sides of the argument. But after a long time in the anti-aging community, you realise most theoretical papers are speculation only. Testing biomarkers is the only thing that really has any validity. Once we have something that works, we can fill in later what the 'mechanism' is.  From that point of view I have found an intervention that significantly reduces methylation age, even when including a purported telomerase activator. I have not noticed any improvements in physical biomarkers measured however (BP, HRV, React time, pulse, etc.) It will be interesting to see if there have been any changes in my blood profiles, when I test that later in the year. Having said all that, taking a break is definitely advised, if only as a precaution against impurities. I took a long break from AKG  (2 1/2 months), but you could equally break for a few days a week. 

[capob]

I found two supplements in amazon from reputable brands, one with 30% triterpenes in phytosome(60mg per pill) around 13$ for 60 pills(Swanson phytosome gotu kola), and one without phytosomes but 35% triterpenes(257mg per pill) 30$ 30 pills(Life extension arterial protect)

 

Also

Pharmacological Review on Centella asiatica: A Potential Herbal Cure-all (nih.gov)

 

It appears that gotu kola needs to be cycled and can't be taken continually

 

 

No.

 

@:study1 [reference 78]

Grimaldi, R., De Ponti, F., D’Angelo, L., Caravaggi, M., Guidi, G., Lecchini, S., … Crema, A. (1990). Pharmacokinetics of the total triterpenic fraction of Centella asiatica after single and multiple administrations to healthy volunteers. A new assay for asiatic acid. Journal of Ethnopharmacology, 28(2), 235–241. doi:10.1016/0378-8741(90)90033-p 

 

The dose target of between .2mcg/ml and .02mcg/ml blood concentration would yield ~12mg - 1.2mg of triterpenes.  

Study1 :

-  chronic 60mg: max 1.5mcg/ml

-  chronic 30mg: max 1mcg/ml

-  single 60mg: max 1mcg/ml

-  single 30mg: max .5mcg/ml

 

Study 1:

1.  it takes longer than 24hr to clear 120mg of triterpenes (indicated by asiatic acid)

2.  it takes less than 24hr to clear 60mg of triterpenes 

 

Since target dose is well below "build up" dose, no cycling indicated.  Further, the "build up" in reference 78 can largely be explained by dose overlap (the final 60mg dose in the chronic group was probably taken 12 hours after the  previous 60mg dose)

 

Looking up triterpene content in extracts, I estimate 10mg/ml on the high side [Triterpene Composition and Bioactivities of Centella asiatica](https://www.mdpi.com...9/16/2/1310/htm) - unless explicit isolation.

Assuming ~1g/ml, the standard dosage on supplemental gotu kola is presented as 1g, which fits nicely into the recommendable amount based on my numbers.

 

I wonder if the bottle I bought 4 years ago is still good...

[QuestforLife]

 

No.

 

 

Study 1:

1.  it takes longer than 24hr to clear 120mg of triterpenes (indicated by asiatic acid)

2.  it takes less than 24hr to clear 60mg of triterpenes 

 

Since target dose is well below "build up" dose, no cycling indicated.  Further, the "build up" in reference 78 can largely be explained by dose overlap (the final 60mg dose in the chronic group was probably taken 12 hours after the  previous 60mg dose)

 

Looking up triterpene content in extracts, I estimate 10mg/ml on the high side [Triterpene Composition and Bioactivities of Centella asiatica](https://www.mdpi.com...9/16/2/1310/htm) - unless explicit isolation.

Assuming ~1g/ml, the standard dosage on supplemental gotu kola is presented as 1g, which fits nicely into the recommendable amount based on my numbers.

 

I wonder if the bottle I bought 4 years ago is still good...

 

 

Regarding the build up of asiatic acid in the blood, the following study sheds some light. 

 

Rush, W. R., Murray, G. R., & Graham, D. J. M. (1993). The comparative steady-state bioavailability of the active ingredients of madecassol. European Journal of Drug Metabolism and Pharmacokinetics, 18(4), 323–326. doi:10.1007/bf03190180

 

They used 24mg of asiaticoside twice a day and got a fairly consistent plasma level after 10 days of dosing (~ 0.05ug/ml), which is right in the range we want.  I attach Fig 2 from the paper (look at the triangles).

 

The Gotu Kola Extract I use (https://nootropicsde...coated-tablets/) claims 35-45% triterpenes, with 15-20% asiaticoside (the batch test confirmed 43% triterpenes), which with a 120mg tablet roughly matches the 24mg of asiaticoside given in this study. Therefore taking a tablet first and last thing should do the job. 

 

I must admit however that I am unsure of the effects of the other triterpene contents of these tablets. 

Attached Thumbnails

• 

[capob]

Regarding the build up of asiatic acid in the blood, the following study sheds some light. 

 

Rush, W. R., Murray, G. R., & Graham, D. J. M. (1993). The comparative steady-state bioavailability of the active ingredients of madecassol. European Journal of Drug Metabolism and Pharmacokinetics, 18(4), 323–326. doi:10.1007/bf03190180

 

They used 24mg of asiaticoside twice a day and got a fairly consistent plasma level after 10 days of dosing (~ 0.05ug/ml), which is right in the range we want.  I attach Fig 2 from the paper (look at the triangles).

 

The Gotu Kola Extract I use (https://nootropicsde...coated-tablets/) claims 35-45% triterpenes, with 15-20% asiaticoside (the batch test confirmed 43% triterpenes), which with a 120mg tablet roughly matches the 24mg of asiaticoside given in this study. Therefore taking a tablet first and last thing should do the job. 

 

I must admit however that I am unsure of the effects of the other triterpene contents of these tablets. 

 

Odd study.

 

1.  Not 24mg twice daily.  From the study,

"Asiatic acid and asiaticoside were administered as two 6 mg or two 12 mg capsules, respectively"

 

2. Results were odd

- asiatic acid group, blood concentration went up a 2nd time despite no additional dose, and no perceivable reason for why

- ^ this also occurred for the asiaticoside group.  Although a decrease in decline rate could be explained by delayed conversion of asiaticode to asiatic acid, a rise in blood concentration could not be so explained.

- the peak after dosing does not correspond with the other study.  Considering the 30mg triterpene dose probably roughly matches the 12mg asiaticode dose, the 12mg dose shows neither the height in peak nor the rate of rise that would be expected.

 

I did expect, from the other study, that 

1. Asiaticoside to asiatic acid was inhibited more highly at lower concentrations of asiatocide. 

2. Slow asiaticoside to asiatic acid, particularly at the tail end, was likely also due to latent asiaticoside build up that is released over time (accumulation in fat?)

 

Given my expectations from the other study, I could explain the 5hr oddity by an increase in metabolism at the 5 hour mark, perhaps brought on lunch and insulin spike.  But, I can't explain the lack of matching peaks, unless the 30mg triterpene dose in the other study had much more that contributed to the resulting asiatic acid content than ~12-15mg of asiaticoside, or, perhaps, there was something delaying the 12mg dose absorption.  Regardless,  your dose is too high.

 

Additionally, I doubt the effort should be to maintain elevated telomerase activity routinely, which is why I mentioned the standard dose (~12mg triterpenes) is recommendable (which will spike blood levels to acceptable range and then fall off to zero over 24hrs)

 

Edited by capob, 16 February 2021 - 01:37 PM.

[Gediminas Jesinas]

Important thing to note that centella asiatica compounds may have poor solubility in water so ethanol tincture has to be prepared. But perhaps there are alternatives to increase bioavailability? How much adult male would have to consume gotu kola powder to achieve telomere lengthening?