1033 replies to this topic

[Castiel]

It's downregulated with enough exposure. So no, it isn't possible to overdose vitamin D through sun-exposure. Also the abilty to synthesize it in the skin is very individual and lowers with age.


I needed about 8.000 IU/d to keep my serum-levels at 70 ng/ml for the last 12 years. Additionally increased full-body sun-exposure up to 340 hrs per year for the last 8. Though sun-exposure did have other beneficial effects by other means, it didn't really further budge my serum levels up at all.

See the table in post 4 of this thread: https://www.longecit...nal-remissions/

 

Interesting, 

 

Bowles suggested lifeguards can have 125~ng/ml iirc.  he also suggests 100~ng/ml are not overdose levels, iirc. 

 

Maybe you're in a more northern or southern latitude where the sun angle doesn't allow as much production.

 

Somewhere nearer to the equator like Florida probably would get better results.

 

Was checking effects of D on epigenetic aging, and found this article.  Though it deals with deficiency and mild supplementation, it sounds promising.

 

 

Results

Fifty-one participants (aged 26.1 ± 9.3 years, 16% are male) were included in the study. After the adjustment of multi-covariates, vitamin D3 supplementation of 4,000 IU/d was associated with 1.85 years decrease in Horvath epigenetic aging compared with placebo (p value = .046), and 2,000 IU/d was associated with 1.90 years decrease in Hannum epigenetic aging (p value = .044). Serum 25(OH)D concentrations were significantly associated with decreased Horvath ∆Age only (p values = .002), regardless of treatments.

Conclusions

Our results suggest that vitamin D supplementation may slow down Horvath epigenetic aging. But the effect on Hannum epigenetic aging is not conclusive. Large-scale and longer duration clinical trials are needed to replicate the findings.

https://www.ncbi.nlm...les/PMC6612014/

Edited by Castiel, 26 May 2021 - 03:31 PM.

[pamojja]

Maybe you're in a more northern or southern latitude where the sun angle doesn't allow as much production.

200 hrs of those 340 in a year (8 years in a row) were on a South Indian beach. The remaining 140 hrs is about the maximum one can get a whole year in central Europe.

Bowles suggested lifeguards can have 125~ng/ml iirc.

Well, if lifeguard for a living 48 weeks per year 40 hours each ~1900 hrs sun-exposure a year!

Edited by pamojja, 26 May 2021 - 04:17 PM.

[Castiel]

200 hrs of those 340 in a year (8 years in a row) were on a South Indian beach. The remaining 140 hrs is about the maximum one can get a whole year in central Europe.


Well, if lifeguard for a living 48 weeks per year 40 hours each ~1900 hrs sun-exposure a year!

 

South India looks like that place at least doesn't look too far from equator.   A question would be pollution in that area.  I know that some places can have little pollution others like China at least you can't even see the sun during the day at times.   

 

In any case wonder how much aging downregulates, and sun exposure downregulates production.   If Bowles Hypothesis of 125-150ng/ml being healthy is true,  I would assume the body can generate that much if you have ample exposure like farmers or hunter gatherers.

[pamojja]

The South-Indian beach I've been too had a constant breeze from the Arabian Sea, and very clear skies almost all the time.

If Bowles Hypothesis of 125-150ng/ml being healthy is true, I would assume the body can generate that much if you have ample exposure like farmers or hunter gatherers.

Well, my last remission occured after overshooting to 135 ng/ml one time.

However, how ready are you really to give up your living and lifestyle for becoming a farmer or hunter/gatherer?

If I would be you I would more realistically try with cheapest supplements first. And get as much of whole-body sun-exposure you can get in your situation.

Edited by pamojja, 26 May 2021 - 04:49 PM.

[Castiel]

The South-Indian beach I've been too had a constant breeze from the Arabian Sea, and very clear skies almost all the time.


Well, my last remission occured after overshooting to 135 ng/ml one time.

However, how ready are you really to give up your living and lifestyle for becoming a farmer or hunter/gatherer?

If I would be you I would more realistically try with cheapest supplements first. And get as much of whole-body sun-exposure you can get in your situation.

 

Yeah, I'm thinking it might be good to go to the beach a bit more often, not something too crazy.   Sure that won't bring 125ng/ml to 150ng/ml but it should give a nice boost to D status.

 

Another thing that can affect D generation on the sun is skin color.   The whiter the skin the easier it is to produce D, but also the easier it is to get burnt.   The darker the skin the harder it is to get sun burnt, but also the harder it is to produce D.

[Gediminas Jesinas]

BTW when it comes to vitamin D it seems the sun is best source, as it is basically free.

 

https://www.nordicna...n-to-vitamin-d/

 

If that is so a couple of hours at the beach should easily generate 50,000 to 100,000 iu for free.    Perhaps simply going to the beach a few times a month, could provide megadose vitamin D without risk(though I think it would be wise to avoid facial exposure to sun as face has lots of muscles and sun damage will result in massive wrinkling unlike the middle of a limb or the chest that basically has no major movement causing wrinkling).    I'm not sure how it is that vitamin D supplementation increases calcium, but I would presume it is by aiding absorption when taken together with calcium through the digestive track.   I assume sun exposure does not carry that risk.   Otherwise lifeguards and farmers would suffer from serious calcium issues.

 

Yes, Sun is free but a lot of UV and blue light damages skin. Probably good idea is to get exposed only to UV-B for most efficiency, UV-A is only good for nitrous oxide and skin tanning but penetrates deeper.

 

Pineal gland calcification as well neural damage, accelerated aging can be caused by fluoride. I suspect I have now insomnia problems due to pineal gland calcification since I had stupid habit in childhood eating toothpaste. In Xray tomography I can clearly see high density point in the pineal gland.  I'm not sure why it's still legal to add large amounts of it into toothpaste or drinking water. Isn't there a way to decalcify pineal gland? I could only find treatments that sound very pseudoscientific even on Google Scholar such as drinking vinegar.

[Andey]

What do you mean by real? I didn't notice any androgen drop when I was keto. 

 

 

 

  My testosterone was way higher normal range half a year after I started keto (I became more aggressive too). Unfortunately it was a lot of polyunsaturated fats, so since then I changed diet composition few times, all keto but different fat ratios and fat types and my T stays as it was before keto.

 

 

 BTW  I ve tried to take Vit C few times a day as Castiel suggested and the difference in skin quality is noticeable. Probably the most profound difference from a supplement that I ve seen. I use time released version from Swanson.

[Castiel]

Btw checked some of Pierpaoli's websites, as I knew he'd talked about blood levels with supplementation, and found the following

 

 

Why is 1 milligram enough?

Melatonin does not have a quantitative pharmacological value, as has been demonstrated in hundreds of scientific journals, but is only necessary to “rest” the pineal gland. The “physiological” (dose-dependent) effect of taking Melatonin is obtained within the range of physiological values of the endogenous plasma melatonin (50-200 pg / ml) with equal doses, in a young healthy adult, at 0.1-0.3 mg. It has been proven that doses of 0.3 - 0.5 mg are already effective and are able to perfectly imitate a youthful nocturnal peak. Administering 0.3 mg doses of Melatonin to adult males will result in blood levels close to physiological levels. A 0.3 mg dose of Melatonin is in fact sufficient to increase the level of Melatonin in blood about 120 picograms per milliliter, a level that corresponds to the nocturnal peak of melatonin in youth.

It should, however, be considered that individual needs may vary according to differences in metabolism. Precisely because everybody has a different intestinal absorption, I prudentially suggest taking 1 to 3 mg. There is no risk of overdose. Melatonin is water-soluble, and the excess quantity is eliminated by kidneys in the urine. During a clinical study, a daily dose of 6 grams (or 6,000 times higher than recommended) was given to some women for a long period of time without the appearance of any side effects. In fact, as we have said, a dose of 0.3 - 1.0 mg Melatonin Zinc-Selenium tablets is normally sufficient to return the nocturnal peak of melatonin to youthful levels and to saturate the organism during the night.-bioclock.it

 

If I'm not mistaken that suggests that 300mcg to 500mcg should yield close to normal peak melatonin levels.   A bit higher might be needed if there are intestinal issues regards absorption of melatonin.

 

But the other studies linked earlier suggested that 3mg seemed to have an effect lowering LH, while 6mg did not and 500mcg had marginal effects on LH.

 

Also would need to know what minimal dose to rejuvenate thymus is.   Was the dose used in rodent experiments the minimum?  I'd need to check the paper to see if they tried lower doses.

[aribadabar]

 

Bowles suggested lifeguards can have 125~ng/ml iirc.  he also suggests 100~ng/ml are not overdose levels, iirc. 

 

If you have not noticed already, Bowles likes to megadose everything and claim that megadosing is the cause for some cure/improvement.

One has to temper his shouting and use common sense and some saner doses.

Edited by aribadabar, 27 May 2021 - 03:15 PM.

[QuestforLife]

Isn't there a way to decalcify pineal gland? I could only find treatments that sound very pseudoscientific even on Google Scholar such as drinking vinegar.

 

Possibly melatonin

 

Melatonin rejuvenates degenerated thymus and redresses peripheral immune functions in aged mice

The effect of melatonin on age-related thymic involution and peripheral immune dysfunctions was investigated. Exogenous

melatonin was administered through the drinking water (15 mg/ml) of 22-month-old female C57BL mice for 60 consecutive days.
Our results show that melatonin distinctly reversed the age-related thymic involution as revealed by the notable increase of thymus
weight, total number of thymocytes and percentage of thymocytes at G2/S phases. More strikingly, spleen weight, total number of
splenocytes and some peripheral immune capacity such as mitogen responsiveness and NK cell activity were also significantly
recovered by 60 days of melatonin application in aged mice. Our findings demonstrate that even when the melatonin
supplementation begins late in life, the age-related thymic involution and peripheral immune dysfunctions can be restored at
least partially in old mice. 

doi:10.1016/S0165-2478(03)00068-3

 

you could try an equivalent dose of melatonin before bed (to avoid daytime sleepiness).

 

According to the following paper (https://www.ncbi.nlm...les/PMC4804402/) we can calculate a Human Equivalent Dose (HED).

 

15ug/ml melatonin in the water

x9ml (max consumed by mice per day, from google search)

/12 = HED/kg

x your mass in kg

For me it comes out at about 1mg of melatonin/day.

 

Try this for a while (maybe 6 months?) and get your calcification checked again. 

[Castiel]

Possibly melatonin

 

 

 

 

you could try an equivalent dose of melatonin before bed (to avoid daytime sleepiness).

 

According to the following paper (https://www.ncbi.nlm...les/PMC4804402/) we can calculate a Human Equivalent Dose (HED).

 

15ug/ml melatonin in the water

x9ml (max consumed by mice per day, from google search)

/12 = HED/kg

x your mass in kg

For me it comes out at about 1mg of melatonin/day.

 

Try this for a while (maybe 6 months?) and get your calcification checked again. 

your source mentions thymus, you sure it also targets pineal?  

If you have not noticed already, Bowles likes to megadose everything and claim that megadosing is the cause for some cure/improvement.

One has to temper his shouting and use common sense and some saner doses.

 

Yeah sometimes megadose is kind of sensible like vitamin C, but I'm not sure megadose is needed for most stuff.

 

If RDA for magnesium was 800mg previously and was truly lowered to 400mg just to lower food stamps costs to the gov, and they were considering lowering to 200mg than that too may be something that should have a higher dose.

 

D maybe 50,000 iu to 100,000 iu is a bit crazy.   But I think higher doses of 2000 to 3000iu might not be a bad idea.

[QuestforLife]

your source mentions thymus, you sure it also targets pineal?  

 

No, I'm not sure, but it seems an experiment worth trying.

 

An alternative is epitalon, which targets the pineal gland to produce melatonin. 

 

 Pineal peptides restore the age-related disturbances in hormonal functions of the pineal gland and the pancreas

The purpose of this research was to study age-related changes in functioning of pineal and pancreatic glands of non-human primates, rhesus monkeys, and to elucidate the possibility of their corrections with the help of epitalon, a synthetic analogue of the pharmacopoeia drug epithalamin. In old (20-27 years) animals, the basal plasma levels of glucose and insulin were found to be higher, while the night melatonin level was lower in comparison with (6-8 years) young animals...The epitalon administration to old monkeys caused the decrease in the basal levels of glucose and insulin and the increase in the basal night melatonin level. Additionally, in the case of old monkeys, epitalon decreased the area under the plasma glucose response curve, markedly increased the glucose 'disappearance' rate and normalized the plasma insulin dynamics in response to glucose administration. DOI: 10.1016/j.exger.2004.10.004

 

If melatonin falls with age because of calcification of the pineal, and exogeneous epitalon increases the release of melatonin from the pineal, then it is a reasonable hypothesis epitalon is decalcifying the pineal gland. It may be operating via another mechanism unrelated to calcification, but this is testable via a computed tomography scan.

[aribadabar]

Yeah sometimes megadose is kind of sensible like vitamin C, but I'm not sure megadose is needed for most stuff.

If RDA for magnesium was 800mg previously and was truly lowered to 400mg just to lower food stamps costs to the gov, and they were considering lowering to 200mg than that too may be something that should have a higher dose.

D maybe 50,000 iu to 100,000 iu is a bit crazy. But I think higher doses of 2000 to 3000iu might not be a bad idea.

Yes, I can get behind 500-750mg Mg and 3000-5000IU of D3 as RDA as I am using them myself and will continue to do so.

Edited by aribadabar, 28 May 2021 - 12:39 PM.

[QuestforLife]

Hyperfunctional telomerase

 

How do we increase telomere length, rather than increasing mitosis?*

 

This paper suggests that providing T cells with additional telomerase doesn't necessarily increase telomere length, if those cells use it for greater division.

 

 

Effect of Peptide AEDG on Telomere Length and Mitotic Index of PHA-Stimulated Human Blood Lymphocytes
We studied the effect of peptide AEDG (ed: Epitalon) on telomere length and mitotic index of PHA-stimulated blood lymphocytes from (a) young (18-22 years, N=5) and (b) middle-aged (49-54 years, N=6) Men.

Source: https://www.research...ation/337492728

 

I've taken the liberty of plotting the change in proliferative index (PI) versus the change in telomere length (TL) resulting from the addition of epitalon, as the paper's authors seemed to have missed the connection. The correlation between the two is about 0.45, which is pretty strong when you consider that we are not even taking into account the absolute telomere length, with short telomeres tending on average to get longer, and long telomeres more likely not to benefit or to get shorter. Note that the one outlier (number 6), in the top right of my attached chart, who got longer telomeres AND increased the proliferation of their lymphocytes, had the shortest telomeres to start with. 

 

(See attached chart)

 

This further paper also suggests MSCs derived from dental pulp can actually increase their TL when they are growth inhibited, either by contact inhibition or recovery from cryopreservation.

 

 

The Effect of Cultivation Passaging on the Relative Telomere Length and Proliferation Capacity of Dental Pulp Stem Cells

We documented the telomere attrition with increasing passaging. The shorter the relative telomere length, the lower reached population doublings, and longer population doubling time were observed at the end of the cultivation. We observed the telomere prolongation in DPSCs (Dental Pulp Stem Cell) cultivated for two weeks with no passaging in the added subsequent study. We concluded that excessive proliferation demands on DPSCs during in vitro cultivation result in telomere attrition. We opened the theory that the telomerase might be more efficient during cell cultivation with no passaging. This observation could help in preserving the telomere length during ex vivo DPSC expansion.

Source: https://www.mdpi.com...8-273X/11/3/464

 

 

 

Note the increase in TL in 2/3 cell lines cultivated but not passaged for 2 weeks. After 3 weeks telomeres shorten once more; the authors speculate 'the relative telomere length attrition correlated with the cell viability reduction. We assumed that DPSCs had to proliferate to replace dead cells that were washed out during medium exchange', which seems plausible.

 

Now that we know telomere length can be increased when telomerase is supplied, so long as proliferation isn't increased, the next question is how do we do it? The obvious way is to use an mTOR inhibitor like rapamycin or everolimus. Taking a telomerase activator with an mTOR inhibitor (possibly also with a fast) should result in longer telomeres all around. The exact protocol remains to be worked out.

 

*[This is not to suggest increased mitosis is not beneficial in some cases, but I have an intuition stem cell pools could considerably recover from the depletion that comes with aging, if only given a break from the demands of constant mitosis. Note this belief is partially inspired by the aging of mice related to the rapid shortening of their telomeres, even though their telomeres do not seem to ever reach the crisis of telomere induced senescence.]

Attached Thumbnails

• 

Edited by QuestforLife, 07 June 2021 - 09:33 AM.

[Castiel]

From the earlier linked paper on membrane composition and lifespan

 

 

A decreased rate of lipid peroxidation can come about via a decrease in the availability of peroxidizable fatty acids, and it has been proposed that such membrane alteration is the basis of the protective effect of CR (382). Laganiere and Yu (188) first showed CR altered the fatty acid composition of liver mitochondrial and microsomal membranes in rats such that they became less susceptible to peroxidative damage. Since this seminal observation, others have reported similar CR-induced changes for other tissues (53, 190, 201, 268, 270, 346) as well as for different classes of liver phospholipids (54, 168). All of these studies involved long-term CR in rats with the shortest period examined being 10-wk CR (54). A study of CR in mice (99) reports similar changes in the fatty acid composition of phospholipids from liver, heart, kidney, and brain, as well as liver and muscle mitochondria, with the changes in this study being manifest following 1 mo of CR (the earliest period sampled). Recently, it has been found that CR treatments as low as 8.5 and 25% result in a decreased PI of membrane lipids and lipoxidative damage to proteins (Pamplona, unpublished results). The effect was especially pronounced when specific components of the diet (protein and methionine) were restricted (9, 313).

Life and Death: Metabolic Rate, Membrane Composition, and Life Span of Animals | Physiological Reviews (physiology.org)

 

CR appears to alter membrane composition, it is conceivable that CR mimetics(some of which are senolytics and others are telomerase activators too) also might alter the membrane composition reducing membrane peroxidation index(which should vastly lower mitochondrial damage, dna damage, and protein damage)

 

 

 

 it is only recently that we have become aware that cell composition (and specifically membrane composition) varies in a systematic manner between species. This fact was not known when the connection between the speed of metabolism and a species longevity was first recognized. We have suggested here that membrane fatty acid composition may be a fundamental property determining maximum longevity.

When the fact that fatty acids differ dramatically in their susceptibility to peroxidative damage is combined with species variation in membrane composition, the link between body size, metabolic rate, and longevity becomes more apparent. In this contribution, we have discussed evidence that the fatty acid composition of membranes can potentially explain 1) the shorter longevity of small mammals compared with larger mammals, 2) the exceptional longevity of naked mole-rats compared with similar-sized mice, 3) the extended longevity of wild-derived lines of mice compared with laboratory mice, 4) the longer life spans of birds compared with similar-sized mammals, 5) the extended longevity of rodents caused by calorie restriction, 6) the longevity difference between workers and queens in honeybees, and 7) also suggested it may be an explanation for the exceptional longevity of our own species, Homo sapiens.-same paper

 

Ways to alter membrane composition seem like another promising avenue for antiaging interventions.   But the membrane composition is under tight genetic control, dietary intake of different types of fats is not usually a way to alter these, iirc, you need to target the genetic pathways themselves.

 

CR appears to alter membrane composition, they suggest lower insulin also might alter membrane composition(this can be achieved by increasing insulin sensitivity and reducing simple carbohydrates in the diet, and replacing them with complex carbs.  Also thinking metformin might help by increasing insulin sensitivity.  And perhaps Amla as that is said to lower blood sugar levels, and probably also increases insulin sensitivity ), they say CR lowers growth hormone and that might also affect membrane composition, but is that true?  As fasting is said to increase growth hormone, so I'd need to verify that

[QuestforLife]

From the earlier linked paper on membrane composition and lifespan

Life and Death: Metabolic Rate, Membrane Composition, and Life Span of Animals | Physiological Reviews (physiology.org)

 

 

 

Do you have any reason to believe this is an important part of aging in humans? It almost certainly is in mice, but I'm not concerned with mouse aging.

 

Murine cells do not senescence at low O2 levels, whereas human cells do. Conversely, human cells require more telomerase to continue proliferation, murine cells do not.

 

In the case of mice, their cellular lifespan appears to be driven by DNA damage from ROS, possibly influenced by the ease of peroxidation of their lipid membranes. In the case of humans, cellular lifespan is controlled by telomere shortening. If human cell membranes were a limiting factor, then telomerase would not be sufficient to keep cells proliferating without lowering ROS.   

 

 

 Most mammalian cells do not divide indefinitely, owing to a process termed replicative senescence. In human cells, replicative senescence is caused by telomere shortening, but murine cells senesce despite having long stable telomeres1. Here, we show that the phenotypes of senescent human fibroblasts and mouse embryonic fibroblasts (MEFs) differ under standard culture conditions, which include 20% oxygen. MEFs did not senesce in physiological (3%) oxygen levels, but underwent a spontaneous event that allowed indefinite proliferation in 20% oxygen. The proliferation and cytogenetic profiles of DNA repair-deficient MEFs suggested that DNA damage limits MEF proliferation in 20% oxygen. Indeed, MEFs accumulated more DNA damage in 20% oxygen than 3% oxygen, and more damage than human fibroblasts in 20% oxygen. Our results identify oxygen sensitivity as a critical difference between mouse and human cells, explaining their proliferative differences in culture, and possibly their different rates of cancer and ageing.  source: https://www.ncbi.nlm...les/PMC4940195/

 

My guess would be humans make the most of very short telomeres by keeping ROS very low, and our membranes mostly saturated. 

Edited by QuestforLife, 08 June 2021 - 01:12 PM.

[Castiel]

Do you have any reason to believe this is an important part of aging in humans? It almost certainly is in mice, but I'm not concerned with mouse aging.

 

Murine cells do not senescence at low O2 levels, whereas human cells do. Conversely, human cells require more telomerase to continue proliferation, murine cells do not.

 

In the case of mice, their cellular lifespan appears to be driven by DNA damage from ROS, possibly influenced by the ease of peroxidation of their lipid membranes. In the case of humans, cellular lifespan is controlled by telomere shortening. If human cell membranes were a limiting factor, then telomerase would not be sufficient to keep cells proliferating without lowering ROS.   

 

 

 

My guess would be humans make the most of very short telomeres by keeping ROS very low, and our membranes mostly saturated. 

 

 

DNA damage will also cause cells to senesce even with long telomeres.    Membrane peroxidation index, is kept lower in longer lived species and EVEN LOWER STILL in negligible senescence species.   Perhaps human membranes are already good enough for negligible senescence for our given metabolism, I'm not sure.   But a big problem is it appears part of the aging program's mechanism is to alter membrane composition such that membranes are more prone to oxidative damage, which is like an autocatalytic positive feedback reactive chemical chain reaction.    This along with decreased nad, decreased endogenous antioxidants, and decreased protein turn over, is only going to make the situation worse.   

 

I do believe we must keep the membrane peroxidation index as low as in youth or lower as CR seems to do.    Certainly worsening of membrane peroxidation index with age related changes in membrane composition is probably contributing to the accelerated decay of humans.

Edited by Castiel, 08 June 2021 - 02:05 PM.

[QuestforLife]

We should be aiming for mouse cell levels of telomerase, not HELA levels

 

Most mammalian cells do not divide indefinitely, owing to a process termed replicative senescence. In human cells, replicative senescence is caused by telomere shortening, but murine cells senesce despite having long stable telomeres1. Here, we show that the phenotypes of senescent human fibroblasts and mouse embryonic fibroblasts (MEFs) differ under standard culture conditions, which include 20% oxygen. MEFs did not senesce in physiological (3%) oxygen levels, but underwent a spontaneous event that allowed indefinite proliferation in 20% oxygen. The proliferation and cytogenetic profiles of DNA repair-deficient MEFs suggested that DNA damage limits MEF proliferation in 20% oxygen. Indeed, MEFs accumulated more DNA damage in 20% oxygen than 3% oxygen, and more damage than human fibroblasts in 20% oxygen. Our results identify oxygen sensitivity as a critical difference between mouse and human cells, explaining their proliferative differences in culture, and possibly their different rates of cancer and ageing.  source: https://www.ncbi.nlm...les/PMC4940195/

 

This paper really got me thinking. 

 

Mouse cells senesce quickly at 20% oxygen but not at all at 3% (roughly physiological levels). Human cells senesce more slowly than mouse cells at 20% due to better defence against ROS, but unlike mouse cells do (eventually) senesce at 3% oxygen levels, due to lack of telomerase. The difference between 20% and 3% oxygen acts as growth inhibition (see my recent post on epitalon and T cells). The normal state of most human cells is (near) hypoxia (1-6%).

 

 

 Most mammalian tissue cells experience oxygen partial pressures in vivo equivalent to 1–6% O2 (i.e., physioxia). In standard cell culture, however, headspace O2 levels are usually not actively regulated and under these conditions are ~18%. This drives hyperoxia in cell culture media that can affect a wide variety of cellular activities and may compromise the ability of in vitro models to reproduce in vivo biology. source: https://www.hindawi....l/2018/8238459/

 

 

Bill Andrews uses HELA cancer cell telomerase levels as his benchmark for stopping aging in human cells. But given the above, I think a better benchmark would be differentiated mouse cells. But finding out what these telomerase levels are has not been easy. Eventually I found a thread of evidence linking HELA and Mouse telomerase levels.

 

 

FIG 1.Telomerase activity in human (cancer) cell lines:

Source: https://www.nature.c...rticles/1206468

 

 

Note that HELA levels are slightly above U937 (leukaemia cells line) telomerase levels. 

 

 

Fig 2. Bottom Left: TRAP Assay(telomerase protein level) of HELA and K562 cells

Source: https://www.cell.com...t/S0092-8674(00)80538-3

 

 

Note that HELA levels are slightly less than K562 (another cancerous leukaemia line) telomerase levels.

 

Next I found a link showing the telomerase level of K562 and HL60 (yet another leukaemia cells line) is the same:

 

 

Fig 3. mRNA TERT (top left) and protein telomerase (top right) white panels for HL60 and K562 cell lines

 

 

 

source: https://bmccancer.bi...471-2407-11-512

 

 

Finally I tied it all together by finding a paper comparing HL60, U937 and both differentiated and undifferentiated mouse cells:

 

 

Fig 4. TOP LEFT: mRNA TERT levels for HL60 and U937 (cancer), H9 (human stem) and Mouse (stem white and grey differentiated) cells

 

source: https://academic.oup.../8/2618/2410230

 

 

This last paper is interesting in its own right, showing how human stem (H9) or cancer cell lines (HL60 and U937) lose most (by a factor of 37-1000) or all of their telomerase expression when they differentiate, whereas mouse cells by comparison drop only by a factor of 5-10. 

 

But for our purposes we can now link HELA telomerase levels to the telomerase levels of U937 or HL60 (via a comparison with K562) cells to that of differentiated mouse cells. From Fig 1 we can see HELA telomerase levels are a little above that of U937 and from Fig 2 a little below that of K562 and (through Fig 3) HL60 cells. For the sake of argument this puts HELA at about the same level as Human embryonic stem cells (H9) or undifferentiated mouse cells (M1 or F2-1 types) in Fig 4. This is 5 or 6 times higher than differentiated mouse cells (grey bars in same image). 

 

Thanks for sticking with me. I did find other references confirming the comparations in telomerase levels I have shown above. I am reasonably happy I'm not cherry picking my data. 

 

If the above is even vaguely accurate, then for human cells in 3% oxygen, we can reduce our telomerase requirement by 5 or 6 times. If TAM-818, Bill Andrew's telomerase activator, is 16% of HELA (as he states), then it is 83-94% of differentiated mouse cells' telomerase levels (depending on whether you are talking about M1 or F2-1 cells; it's even better for F2-9 cells). This is very close to what we need to stop aging.

 

You might argue the assumption all our tissues are at 3% oxygen is not valid; I quoted a reference stating 1-6% above. I plan to make up that shortfall by periodically increasing MTOR inhibition, which is (in my opinion) the main effect of hypoxia.  

Edited by QuestforLife, 11 June 2021 - 09:24 AM.

[QuestforLife]

Human and Mouse embryonic stem cells have similar levels of telomerase activity to one another and to HELA cells.

 

Expression of TERT, TERF1, and TERF2 genes in both mouse ES (D3 cell line) and human ES (H1 cell line) was consistent with high telomerase activity measured with TRAP assay. Telomerase activity in both ES cell lines was as high as in HeLa tumor cells.

(See panel E, bottom right

source: https://www.scienced...012160604000193

 

Given telomerase activity is partially lost during differentiation in the case of mice cells, this is further proof that HELA telomerase levels are higher than those in differentiated mouse tissues, (which can nevertheless proliferate indefinitely in 3% oxygen).

 

 We show that the expression of mTert is very closely linked to telomerase activity and that both are substantially reduced upon differentiation of ES cells into more committed lineages... source: https://www.scienced...925477300004238

 

[Castiel]

 

If the above is even vaguely accurate, then for human cells in 3% oxygen, we can reduce our telomerase requirement by 5 or 6 times. If TAM-818, Bill Andrew's telomerase activator, is 16% of HELA (as he states), then it is 83-94% of differentiated mouse cells' telomerase levels (depending on whether you are talking about M1 or F2-1 cells; it's even better for F2-9 cells). This is very close to what we need to stop aging.

 

You might argue the assumption all our tissues are at 3% oxygen is not valid; I quoted a reference stating 1-6% above. I plan to make up that shortfall by periodically increasing MTOR inhibition, which is (in my opinion) the main effect of hypoxia.  

 

 

Quick Google Searching for TAM-818 only found the serum.  Is this not sold as supplement?  Are the ingredients known if not sold as supplement?    Sounds very interesting, and if your analysis is correct it might reach adequate levels if combined with some of the other telomerase activators mentioned in this thread.

 

What's the link to MTOR and telomeres?  I assume since it is a nutrient sensor less activation would lead to reduced rate of replication, hence reduced loss of telomere length, was that mentioned here or previously and I missed it?  

 

As for MTOR inhibition, I think glycine supplementation and periodic fasting mimicking diet might do the trick.    I know that low protein appears to have benefits in several species.  Of course some periods of moderate protein are probably needed to keep decent muscle mass.   It is still unknown what the maximum safe frequency of fasting mimicking periods is.   But during both fasting mimicking periods and during total fasting mtor activity should be vastly reduced.

 

 

Such studies in insects and mice indicate that animals with ad libitum access to low-protein, high-carbohydrate diets have longest lifespans. Remarkably, the optimum content and ratio of dietary protein to carbohydrates for ageing in experimental animals are almost identical to those in the traditional diets of the long-lived people on the island of Okinawa...The longest living people are the residents of the Japanese island of Okinawa, who have as many as five times more centenarians than other developed nations

 

 Branched chain amino acids are potent activators of mTOR, and this result provided a simple mechanism linking low-protein intake, low-circulating branched chain amino acids and inactivation of mTOR. Intriguingly, the regulation of mTOR was not only influenced by branched chain amino acids, but also by the interaction between glucose and branched chain amino acids, providing a clue as to why the ratio of dietary carbohydrates and protein influences ageing. It is of note that elevated branched chain amino acids are a marker of diabetes mellitus in humans [34], but on the other hand, supplementation with branched chain amino acids has been reported to increase lifespan in mice [35] and nematode worms [36], presumably despite activation of mTOR.

New Horizons: Dietary protein, ageing and the Okinawan ratio - PubMed (nih.gov)

 

I will say that I'm not sure whether the fats used in laboratory animal experiments are healthy fats.  I mean really doubt a high fat diet based on healthy monounsaturated fats sourced from extra virgin olive oil avocados and macadamia would somehow fare worse than a high carb diet.   That said I think one of the more beneficial aspects of high carb, is if it is complex carbs with lot of soluble fiber, they get broken down by bacteria into healthy short chain fatty acids like butyrate.

 

Maybe there's a study comparing high healthy fat vs high carb diet, that would be good to find.

 

edit:

As the above study comments, and the following adds on too low protein appears not optimal long term at least if coupled with other sources of calories.   The okinawan ratio is around 10% of calories being protein, iirc.

 

 

 

We previously demonstrated that feeding a 2% protein AIN-76 diet ad libitum for 14 days resulted in substantial clinical and biochemical changes including weight loss, hypoglycemia, hypoalbuminemia, higher levels of plasma cytokines, oxidative stress in the liver, and activation of inflammatory signaling to interleukin (IL)-6, as compared with a 20% protein diet. In the present study, 54 rats were randomly given a standard rat chow diet ad libitum, or a 25% or 50% reduction of this intake for 14 days. The results showed that weight gain was less in the 25% food-restricted group and halted in the 50% group as compared with the control group. Unlike protein restriction, neither level of food restriction altered plasma levels of albumin and glucose, the hepatic protein abundance of signal transducers and activators of transcriptions and of mitogen-activated protein kinases, or the hepatic contents of total glutathione and malondialdehyde. The intracellular signaling in response to IL-6 stimulation was also well maintained. However, both levels of food restriction elevated IL-1 and corticosterone in plasma, did not alter ghrelin, and decreased plasma levels of free fatty acids. Because these latter 3 markers were not examined previously, 20 rats were fed an AIN-76 diet, either with 20% or 2% protein, ad libitum for 14 days. The 2% protein diet significantly decreased plasma levels of free fatty acids and increased ghrelin and corticosterone as compared with the 20% protein diet. Thus, food restriction, where all essential nutrients are reduced in proportion, is a physiologic stress that, while limiting growth, does not activate or impair the systemic inflammatory response, whereas a very low protein diet with little change in energy intake has a substantial impact on systemic inflammation, body composition, and growth.

Comparison of the effects of food versus protein restriction on selected nutritional and inflammatory markers in rats (nih.gov)

Edited by Castiel, 11 June 2021 - 10:24 AM.

[QuestforLife]

Quick Google Searching for TAM-818 only found the serum.  Is this not sold as supplement?  Are the ingredients known if not sold as supplement?    Sounds very interesting, and if your analysis is correct it might reach adequate levels if combined with some of the other telomerase activators mentioned in this thread.

 

 

 

In answer to you first question, TAM-818 is available here as a softgel, but it is very expensive, even for 3 months supply.

 

https://defytime.com...-care-capsules/

 

A cheaper alternative might be gotu kola, if you believe this study where an asiaticoside (08ATGTLF) extract achieved 17% HELA telomerase levels in T cells. 

 

 

 

Discovery of potent telomerase activators: Unfolding new therapeutic and anti-aging perspectives

source: https://www.ncbi.nlm...les/PMC6755196/

 

 

 

 

 

What's the link to MTOR and telomeres?  I assume since it is a nutrient sensor less activation would lead to reduced rate of replication, hence reduced loss of telomere length, was that mentioned here or previously and I missed it?

 

Spot on; the idea is to get more telomere lengthening if proliferation is reduced, as discussed in Post 584.

Edited by QuestforLife, 11 June 2021 - 10:16 AM.

[Castiel]

 

Spot on; the idea is to get more telomere lengthening if proliferation is reduced, as discussed in Post 584.

That's what worries me about retinol skin creams, iirc, they increase rate of skin cell replication.   I think they might allow improved appearance in the short term but at the cost of accelerated aging.  I always tell people to avoid creams and serums containing it.

Edited by Castiel, 11 June 2021 - 10:30 AM.

[QuestforLife]

Response to the Thought-Provoking Critique of Hyperfunction Theory by Aubrey de Grey
https://doi.org/10.1089/rej.2021.0018

This reference shows an intriguing disagreement between Aubrey de Grey and Blagoskonny discussing whether rapamycin is in fact increasing the rate of damage repair (de Grey's viewpoint) or not, instead possibly reducing it but nevertheless increasing lifespan because the proximal cause of aging is not molecular damage (Blagoskonny's standpoint). Aubrey makes the strong point that if molecular damage had no impact on aging (I take telomere shortening as my example here) then random mutations would eventually cause damage accumulation (i.e. telomere shortening) to become relevant to aging.

I generally support Blagoskonny's point of view, as there is clearly much random damage even in the bodies of the young that has nothing to do with aging. But I don't agree with the dichotomy. Take for example telomere shortening as 'damage'. MTOR drives cellular proliferation. This drives telomere shortening. Telomere shortening then slows proliferation, which increases susceptibility to geroconversion- both in the proliferating cells and in the non-dividing cells they support.

This highlights a problem with the binary approach of many scientists to aging. For example, cellular senescence: a cell can either divide or it can't. But actually all cells are somewhere on a spectrum of readily dividing upon stimulation, to dividing only slowly with much stimulation, right to not being able to divide at all. This can be seen in the decline in the gradient of the number of dividing (and re-plated) cells in a culture. The gradient reduces significantly before it goes completely flat. This is the whole point. We don't just die, we 'age' for a protracted period first.

In this respect I disagree with Blagoskonny when he says telomere shortening has nothing to do with aging. Telomeres don't need to shorten all the way to replicative senescence in order to cause problems. Cells only need to become 'senescent enough' (with senescence on a sliding scale as described above).

For Blagoskonny's argument, see:

https://doi.org/10.18632/aging.202674
DNA- and telomere-damage does not limit lifespan: evidence from rapamycin

This isn't only a question for dividing cells. Post mitotic cells, those that could divide but don't because their function is to remain in place and do a job, are similarly affected by the scheme I outline. Hypertrophy of cardiomyocytes may not only be caused by direct MTOR stimulation, but also by the slowing of the proliferation of endothelial cells (via MTOR driven telomere shortening). The accumulation of toxic amyloids around neurons similarly need not only be the direct result of hyperactive growth signalling to the neurons themselves, but also the attrition of the neuron supporting glial cells via telomere shortening.

So here we have a lovely synthesis of Blagoskonny's hyperfunction theory of aging with the telomere shortening theory of aging. Blagoskonny's strategy of slowing the highway speed car to a rate appropriate for residential areas, is analogous to slowing the rate of telomere shortening because much less telomere is remaining (to the old).

But it is also a dynamic problem. As we have seen from my recent posts a certain quantity of telomerase protein may be sufficient to support cellular division at a certain rate but no higher, (analogous to indefinite proliferation in 3% oxygen but senescence at 20% oxygen).

It may even be possible to regrow telomeres, if we increase telomerase expression at the same time as we reduce proliferation. We may be able not only to slow aging,as Blagoskonny has indicated MTOR inhibition should do, but perhaps even reverse it.

[QuestforLife]

Latest Methylation Epigenetic Age Results

 

Long time readers of this thread may remember my plan to decrease methylation age AND increase telomere length simultaneously, as espoused in post #266

 

This was motivated by the fact epitalon increased my telomere length but eliminated the previous improvement in methylation age I gained using the statin-sartan protocol (late 2019 - early 2020).

 

I identified AKG as an important demethylase cofactor and verified that it did indeed decrease epigenetic methylation age after about 6 months of use. I then began to reintroduce telomerase activators. I first used a strong asiaticoside extract of gotu kola, then an epitalon spray and finally in the last testing period introduced Bill Andrews TAM818 aging care capsules. 

 

I am happy to report that TAM-818 has not resulted in an increase in my methylation age. You can see all my results since I started using Trume Age attached. Note, I started AKG 3 months before the start of the plot in April 2020. The only period in which methylation age increased was when I used AKG only intermittently. It was also only a small increase. Since then and since starting TAM-818, my methylation age has continued to improve (see attached).

 

It appears that AKG is the only factor that has had much effect on my methylation age. I have been using 900mg/day. In the last period I also used 400mg berberine. Thanks to aribadabar for pointing out the the CEO of Ponce de Leon (maker of Rejuvant) states berberine and AKG are contraindicated and cancelled out the benefits to lifespan in mice. I certainly found that AKG stopped all weight loss, which I'd normally expect from berberine; not suprising given AKG is effectively an energy source for mitochondria. But given my results, berberine has not stopped AKG reducing methylation age. 

 

 

Warning: I do find AKG reduces my sex drive and after a long time my energy levels. I am currently on a multi month break from AKG.  I took at 2 month break between Sept and Nov 2020, and restarted for a month before my Jan 2021 test, and still got an improvement. My guess is AKG is something that just needs topping up, and perhaps should not be taken chronically.

 

 

Attached Thumbnails

• 

[dlewis1453]

Great results! A shame that AKG has those negative side effects over time but at least we can cycle it.

 

How do you feel/look subjectively? Are you still taking GDF11 and noticing biomarker improvements. 

[kurt9]

Great results! A shame that AKG has those negative side effects over time but at least we can cycle it.

 

AKG has these side effects because it is doing mitochondrial fission. Have a look at:

 

Manipulating mitochondrial dynamics - Page 65 - NAD+ - LONGECITY - Page 65

[kurt9]

AKG has these side effects because it is doing mitochondrial fission. Have a look at:

 

Manipulating mitochondrial dynamics - Page 65 - NAD+ - LONGECITY - Page 65

 

I just checked it out again. AKG is used for both fission and fusion. So this might not be the reason for the side effects.

[QuestforLife]

I just checked it out again. AKG is used for both fission and fusion. So this might not be the reason for the side effects.

 

Actually I think AKG has been slowly damaging my liver. My ALT levels were gradually rising from normal to just above normal by April, and then rocketed to double that value in June. Since that time I stopped AKG and it's fallen back towards the normal range. I don't absolutely know 100% it was the AKG, but it is my prime suspect. AKG is cleared rapidly by the liver (according to the video clip above). I wonder if given the effect of demethylases on reducing differentiation, and the highly proliferative nature of the liver, if AKG might also be impeding liver regeneration this way also.

Edited by QuestforLife, 22 June 2021 - 08:53 AM.

[QuestforLife]

Great results! A shame that AKG has those negative side effects over time but at least we can cycle it.

 

How do you feel/look subjectively? Are you still taking GDF11 and noticing biomarker improvements. 

 

My GDF11 results kind of levelled out at around the values you could see I achieved in post #444 and didn't continue to improve any more. But all the values they reached were very healthy and to be honest I wouldn't want BP to go lower . Because hayfever has now destroyed any hope I had of accurately monitoring HRV I've had to abandon that biomarker for now, but I still check my reaction times and if they start to drift higher for too many days in a row I take another dose. This works out to be every 2-3 weeks.

 

After about 3 weeks of taking TAM818 aging care capsules I noticed my energy levels had improved. I started noticing recovery from exercise was much quicker, whether weights or cardio. Instead of a 3 day period between lifting weights I needed only a day. This is similar to what I experienced when injecting epitalon for a 10 day cycle (I believe there is literature supporting this effect with epitalon,) but the effect has sustained over months (unlike after the epitalon cycle is over). 

 

In terms of appearance I feel like I've paused in my mid to late thirties. A few greys hairs in my beard (when I grow it) are not spreading and I've a full head of hair with zero grey. I have a few deep horizontal lines on my forehead due to my strong brow line, and in the early morning light you can notice loss of subcutaneous fat* under the eyes. Any more wrinkles that were forming seem to have stopped or are fading slowly but the deep ones look here to stay. I'd love to have nice full (under)eye sockets again. I seem to remember liposomal Vitamin C helped me here before, so I may give it another go.  

 

* I now have a Renpho scales that tell you your percentage of muscle, fat type, etc. And interestingly, despite still looking reasonably slim, I have a huge quantity of subcutaneous fat (~24%). Obese levels according to the scales. But my visceral fat is only 8% (normal). Years ago when I tried pioglitazone I got much higher fat levels too. I suspect as GDF11 is injected into subcutaneous fat, it is activating adipose derived mesenchymal stem cells, which are predisposed to generate more fat cells. This is not necessarily all bad, as it helps to keep glucose levels down. And these stem cells probably also generate some endothelial cells, otherwise GDF wouldn't improve BP. But it is a pity that my new subcutaneous fat hasn't made it to my face, otherwise I really could pass for being in my twenties again.

Edited by QuestforLife, 22 June 2021 - 08:58 AM.

[kurt9]

From reading and agreeing with "Turbuckle's" approach to live extension as well as my personal experiences, I am coming to the conclusion that we do not want to take things like NAD+ boosters, AKG, and the like on a semi-permanent basis. I think we want to take these things only as a part of a short-term or periodic protocol, and not take them the rest of the time. I think this is especially true for senolytics and NAD+. My own experience with NAD+ over a three year period is that I developed a persistent cough that was annoying to my wife and other people around me. It turned out that a lot of the cause of it was elevated histamine levels that was caused, in part, by NAD+ compounds as well as Zinc. I stopped taking both of these and my cough largely went away. I also think senolytics can be too much of a good thing. Your body needs a certain amount of inflammation signaling for both wound healing as well as immune defense system. I've used senolytics in the past (Fisetin and Quercetin), but only for short-term protocols. My experience with Curcumin also reinforced my beliefs about this.