2523 replies to this topic

[anon2099]

Thanks Turnbuckle. 

 

Your latest protocol update shows you've switched back to using stearic acid instead of GMS. Is there a particular issue [other than high BP] with using GMS or is it purely for including dihydromyricetin/lecithin alongside with stearic acid in a single package?

 

Is the astragalus powder you are using standardized to certain amount of astragalosides, what would be the ideal astragaloside content?

 

If rat metabolism is 6 times faster than human, would it make sense to water fast for 5-6 days then break the fast with this protocol? Or would this be unwise due to lack of nutrients, despite of amino acid supplementation, and ultimately causing cell arrest?

 

 

Edited by anon2099, 03 April 2022 - 11:13 PM.

[Kentavr]

Thanks Turnbuckle. 

 

Your latest protocol update shows you've switched back to using stearic acid instead of GMS. Is there a particular issue [other than high BP] with using GMS or is it purely for including dihydromyricetin/lecithin alongside with stearic acid in a single package?

 

Is the astragalus powder you are using standardized to certain amount of astragalosides, what would be the ideal astragaloside content?

 

If rat metabolism is 6 times faster than human, would it make sense to water fast for 5-6 days then break the fast with this protocol? Or would this be unwise due to lack of nutrients, despite of amino acid supplementation, and ultimately causing cell arrest?

 

As far as I know, fasting for 5-6 days increases the risk of death because there may not be enough potassium in the body. 

Also, I have read that a very low calorie diet (500-700 kcal) is very similar to starvation, and the risks of dying are much lower.

I suppose it is possible to follow a very strict diet for 5-6 days and still eat fruits such as kiwi and apricots to keep the potassium in the body.

[Turnbuckle]

 

If rat metabolism is 6 times faster than human, would it make sense to water fast for 5-6 days then break the fast with this protocol? Or would this be unwise due to lack of nutrients, despite of amino acid supplementation, and ultimately causing cell arrest?

 

Since the rats were fasted overnight, that might translate to 2-3 days of fasting by a human. Does fasting make sense with this protocol? Not for most Americans, surely!

[QuestforLife]

Since the rats were fasted overnight, that might translate to 2-3 days of fasting by a human. Does fasting make sense with this protocol? Not for most Americans, surely!

 

mTOR inhibition might be an alternative to a fast. It worked in planarians.

 

Downregulation of mTOR Signaling Increases Stem Cell Population Telomere Length during Starvation of Immortal Planarians

 

To test this theory, we performed RNAi experiments for the main components of mTOR signaling followed by FISH for smedwi-1

and TelQ-FISH. Planarians were either injected with Smed-tor double-stranded RNA (dsRNA) (to inactivate the mTOR pathway) or with Smed-smg-1 dsRNA (to overactivate the mTOR pathway). Downregulation of tor further enhanced the effect of starvation on stem cell telomere length when compared with control injected worms (Figures 6C–6E). We also noticed that downregulation of tor is able to increase the maximum telomere length in stem cells.

 

https://doi.org/10.1...mcr.2019.06.005

 

Note that this worked on the pluripotent stem cells present in planaria. Presumably in humans it would benefit the VSELs this protocol is hypothesised to rely on.

[Turnbuckle]

mTOR inhibition might be an alternative to a fast. It worked in planarians.

 

 

Downregulation of mTOR Signaling Increases Stem Cell Population Telomere Length during Starvation of Immortal Planarians

 

Note that this worked on the pluripotent stem cells present in planaria. Presumably in humans it would benefit the VSELs this protocol is hypothesised to rely on.

 

 

I'm not sure about this, as mTOR inhibitors are known to be cytostatic, inhibiting proliferation.

[nadaepeu]

A new article "Multi-omic rejuvenation of human cells by maturation phase transient reprogramming"

 

https://elifescience...s/71624#content

[Turnbuckle]

A new article "Multi-omic rejuvenation of human cells by maturation phase transient reprogramming"

 

https://elifescience...s/71624#content

 

This is hard way of doing things --

 

A very recent publication by Gill D et al. available on bioRxiv claims to have developed an unprecedented transcription factor rejuvenation technology to rejuvenate human fibroblasts for as much as 30 years (as measured by a novel transcriptome clock) [206]. A technology called maturation phase transient reprogramming (MPTR) involves transfection of a polycistronic cassette with OCT4, SOX2, KLF4, c-MYC, and GFP genes under the control of a doxycycline promoter. Genes were expressed ectopically until maturation was reached, after which expression was turned off. MPTR has been shown to significantly rejuvenate all measured molecular signs of aging such as the epigenome, including H3K9me3 histone methylation levels and the DNA methylation aging clock without loss of cell identity.

https://www.mdpi.com.../11/11/1050/htm

 

 

They've achieved a reversal of "as much as 30 years" with cells in a dish using a genetic treatment via a retrovirus vector, whereas I've shown a reversal of 28 years with oral supplements.

 

As for my previous post about the mTor inhibitors --

 

Despite the limited clinical success of the first-generation drugs, rapamycin and its analogs (which are known as rapalogs)...

Both animal and clinical studies have shown that rapalogs are primarily cytostatic

https://www.ncbi.nlm...les/PMC3069769/

 

 

Cytostatic means it inhibits proliferation. And while rapalogs may inhibit proliferation of stem cells, they can promote cancer cell stemness --

 

Here, we describe a novel H-ras specific feedback, which leads to an inadvertent rapalog induced activation of tumorigenicity 

https://pubmed.ncbi....h.gov/28562352/

 

 

So to use mTor inhibitors to achieve fusion (doubtful) or to increase telomerase (not generally desirable in this protocol, anyway) is not advised.

Edited by Turnbuckle, 08 April 2022 - 11:13 AM.

[QuestforLife]

 

As for my previous post about the mTor inhibitors --

 

 

Cytostatic means it inhibits proliferation. And while rapalogs may inhibit proliferation of stem cells, they can promote cancer cell stemness --

 

 

So to use mTor inhibitors to achieve fusion (doubtful) or to increase telomerase (not generally desirable in this protocol, anyway) is not advised.

 

What do you think about glucosamine?

 

As would be expected by shifting metabolism away from sugars, glucosamine dose dependently increases fatty acid oxidation (see ref 1)

 

Anything that competes with sugar uptake would be expected to increase mito response to stearic acid; note diabetics with impaired glucose response had a better (more fused) response to stearic acid than healthy subjects (see ref 2)

 

NAG (N-acetyl-glucosamine) in particular has been shown to have benefits for stem cells through increasing SAM/SAH ratio (see ref 3).

I expect this is connected to starvation/feeding cycles whereby lack of food stimulates fatty acid oxidation from stores and during this period stem cells renew rather than supplying the body. The stem cell protocol already uses SAMe; adding NAG may further improve the SAM/SAH ratio by reducing conversion of the former to the latter.

 

[1] https://journals.plo...al.pone.0018417
[2]https://doi.org/10.1...467-018-05614-6
[3]https://www.pnas.org...pnas.1915582117

[Turnbuckle]

What do you think about glucosamine?

 

 

 

Are you not getting enough action from the recommended fusion agents? In any case, glucosamine is known to increase stem cell activity in vitro, and might be of interest in diabetes.

 

Glucosamine is known to be a structural polysaccharide found in chitosan or chitin in the exoskeleton of crustaceans and arthropods, and glucosamine sulphate is the sulphate derivative of the natural amino monosaccharide glucosamine. Glucosamine plays a role in the formation and function of tendon, skin, bone, heart valves, blood vessels, articular surfaces, and mucus secretions of the digestive and respiratory tracts (16). In addition, glucosamine has been reported to increase migration and proliferation of stem cells. Glucosamine enhances the differentiation of cartilage from stem cells (17,18). In this study, we investigated whether addition of glucosamine into the differentiation medium of hADSCs for differentiation into IPCs is beneficial for improvement of generation of functional IPCs...

 

In conclusion, we found that glucosamine supplementation promotes the differentiation of hADSCs into glucose-responsive insulin secretory cells through the increase of mRNA levels of beta-cell specific genes, including insulin, PDX1 and NGN3. In addition, glucosamine supplementation also increases mRNA expression levels of genes such as Syt4, Gck and Glut2 in differentiated IPCs. These results indicate that glucosamine might be beneficial for the differentiation of hADSCs into functional beta-cells.

 

https://atm.amegroup...view/40524/html

 

 

Edited by Turnbuckle, 11 April 2022 - 07:21 PM.

[DJSwarm]

They've achieved a reversal of "as much as 30 years" with cells in a dish using a genetic treatment via a retrovirus vector, whereas I've shown a reversal of 28 years with oral supplements.

 

28 years reversal is a pretty significant reversal. Are you seeing macromorphological evidence of this?

 

If we met; would I mistake you for a young man?

[Turnbuckle]

28 years reversal is a pretty significant reversal. Are you seeing macromorphological evidence of this?

 

If we met; would I mistake you for a young man?

 

Yes. There wouldn't be much point if it wasn't a real reversal. 

[Kentavr]

Yes. There wouldn't be much point if it wasn't a real reversal. 

 

Turnbuckle, what is your pressure?

Could you write down what your pressures have been in the last month?

[Empiricus]

I just got my second TruMe report back.

 

First test date 2021-08-04

Biological age: 40.7 years

Chronological age: 57.5 years

Delta: 16.8 years

 

Second test date 2022-03-06

Biological age: 37.8 years

Chronological age: 58.1 years

Delta: 20.3 years

 

I started the stem cell protocol rounds on 2020-09-04. I did 11 rounds before my first test and 8 rounds between the 1st and 2nd tests. I kept up with the protocol updates along the way.

 

 

Nice work there. What is your source for the c60?  

 

By the way, do you still take thiamine? Did you take it along with the protocol?  Did you ever figure out whether it potentiates c60oo?  

Edited by Empiricus, 17 April 2022 - 02:30 PM.

[stephen_b]

I'm using PureC60OliveOil that I get off of Amazon. I still take thiamine on occasion, but I'm not sure if it helps running or potentiates C60.

[Empiricus]

Yes. There wouldn't be much point if it wasn't a real reversal. 

 

If you were to discontinue the protocol, do you believe your biological age would reverse quickly or remain stable for several years?  Is there any reason to believe taking extended breaks from the protocol might be more beneficial than doing it frequently?  Do you believe as your real age increases the frequency will need to increase, or is biological age the determining factor?  

 

The protocol is intended for geriatrics. Once a former geriatric's biological age is, say, less than 50 years, are they not too young for the protocol?    

[Turnbuckle]

If you were to discontinue the protocol, do you believe your biological age would reverse quickly or remain stable for several years?  Is there any reason to believe taking extended breaks from the protocol might be more beneficial than doing it frequently?  Do you believe as your real age increases the frequency will need to increase, or is biological age the determining factor?  

 

The protocol is intended for geriatrics. Once a former geriatric's biological age is, say, less than 50 years, are they not too young for the protocol?    

 

 

My epigenetic age is now 42, but I'm continuing, mainly to investigate new tweaks to the process (especially in the neural area, where I recently made a breakthrough). Fifty years is arbitrary. I originally suggested this protocol for geriatrics because they have less to lose should there be some unexpected untoward effects over the long term. It's possible. I've used C60 for ten years, and the first six did show negative effects. With C60 alone, the initial excellent results turned around after a year or two and disappeared, leaving me in some ways worse off. This has not happened with my protocol, however.  Not yet. If I stopped now, there might be a bounce-forward by a few years as part of the effect lies in the more ephemeral TACs, and then I would expect the aging rate to continue at the normal rate.

 

Also note that aging is a mosaic process. Some organs age much more rapidly than others, and in those organs expect the reversal to be equally rapid (so long as there are still SCs to promote). Reversal will depend on the rate of cellular turnover (or in some cases, nuclear turnover). The lifespan of skeletal muscle cells, for instance, is said to be 10-16 years, and the turnover rate is known to decline with chronological age -- presumably reflecting a lack of sufficient replacement stem cells (or satellite cells in the case of muscle tissue). Skin cell turnover is very rapid -- 4-6 weeks -- so a more youthful appearance will be recovered first, but doesn't reflect the age of the whole body.

[QuestforLife]

Also note that aging is a mosaic process. Some organs age much more rapidly than others, and in those organs expect the reversal to be equally rapid (so long as there are still SCs to promote). Reversal will depend on the rate of cellular turnover (or in some cases, nuclear turnover). The lifespan of skeletal muscle cells, for instance, is said to be 10-16 years, and the turnover rate is known to decline with chronological age -- presumably reflecting a lack of sufficient replacement stem cells (or satellite cells in the case of muscle tissue). Skin cell turnover is very rapid -- 4-6 weeks -- so a more youthful appearance will be recovered first, but doesn't reflect the age of the whole body.

 

Of course you (like everyone else) are measuring leukocytes - turned over in 6 days(!), which is no doubt flattering your results. 

 

The same goes in reverse for those whose epigenetic age suddenly rises - it is likely just reflective of changes in leukocytes and would have to be sustained for significant time to be representative of the body as a whole.  

[Empiricus]

Also note that aging is a mosaic process. Some organs age much more rapidly than others, and in those organs expect the reversal to be equally rapid (so long as there are still SCs to promote). Reversal will depend on the rate of cellular turnover (or in some cases, nuclear turnover). The lifespan of skeletal muscle cells, for instance, is said to be 10-16 years, and the turnover rate is known to decline with chronological age -- presumably reflecting a lack of sufficient replacement stem cells (or satellite cells in the case of muscle tissue). Skin cell turnover is very rapid -- 4-6 weeks -- so a more youthful appearance will be recovered first, but doesn't reflect the age of the whole body.

 

Seems like a strong argument for continuing your protocol even once a youthful biological age is attained, as the biological age measure won't be speaking to the status of organs with long lifespans.   

[Turnbuckle]

Of course you (like everyone else) are measuring leukocytes - turned over in 6 days(!), which is no doubt flattering your results. 

 

 

 

 

Are you sure? Spit has a variable mixture of leukocytes and epithelial cells, and to get a consistent reading, you have to select out one or the other. I can't find any information on which one TruMe selects for. But it does make a difference, as they age at different rates. As long as you select for just one, the variability will be much less and you can make comparisons.

 

In conclusion, we found that saliva and buccal swab samples almost always contain a mix of leukocytes and epithelial cell in a wide range of proportions, especially in saliva. 

https://www.ncbi.nlm...les/PMC5932057/

 

 

In most tissues, the source of cells are TACs, and the aging occurs there. Leukocytes have a short life, but their methylation pattern reflects the aging of the source cells. 

 

It has been assumed that the slow rate of healing in aging epidermis is due to slowing of the epidermal stem cell proliferative rate. In this issue, Charruyer et al. report that this may not be the case. Using a long-term repopulating model, they demonstrate that epidermal stem cell kinetics are maintained. Instead, it is the compensatory action of the transit-amplifying (TA) cells that changes in aging skin and thus bears responsibility for slowed wound healing.

https://pubmed.ncbi....h.gov/19826444/

 

 

Edited by Turnbuckle, 28 April 2022 - 08:44 PM.

[QuestforLife]

Are you sure? Spit has a variable mixture of leukocytes and epithelial cells, and to get a consistent reading, you have to select out one or the other. I can't find any information on which one TruMe selects for. But it does make a difference, as they age at different rates. As long as you select for just one, the variability will be much less and you can make comparisons.

 

 

In most tissues, the source of cells are TACs, and the aging occurs there. Leukocytes have a short life, but their methylation pattern reflects the aging of the source cells. 

 

I've asked TruMe. 

 

In any case it would be an interesting experiment to carry out - check the methylation state of 2 cell types with different turnover rate and see what lag there would be on any intervention and if it was proportional to turnover time.

[QuestforLife]

I've asked TruMe.

In any case it would be an interesting experiment to carry out - check the methylation state of 2 cell types with different turnover rate and see what lag there would be on any intervention and if it was proportional to turnover time.

Their reply:

We do not separate by cell type. However, as saliva has 70% leukocytes, it would be safe to say that majority of DNA comes from this cell type.

[Turnbuckle]

This is surely a matter of cost, but a pure sample of spit ought be less problematic than a buccal swab, if you don't select for a given cell type. Nevertheless, it's a source of noise.

Edited by Turnbuckle, 30 April 2022 - 08:25 PM.

[QuestforLife]

Their reply:

And here is the response from MyDNAge regarding their blood test, which many of us have used:

We isolate the DNA from the white blood cells in our blood sample kits. Please let me know if you have any further questions.

Unsurprising that in the case of blood, white blood cells are the way to go. These will be leukocytes but also granulocytes, so there will be some variation in division rate within this sample (there will be many more divisions between blood stem cells and leukocytes than granulocytes).

[lost69]

Turnbuckle

hi i see this protocol made amazing results, i stopped it in 2020 due to humeral head osteonecrosis after an accident while swimming.the osteonecrosis area started receding on MRI in 2021 thanks to teriparatide treatment/local stemcells activation by reac frequencies treatment and i am fully asymptomatic now so doctors urged me to restart swimming as soon as possible.so i was thinking to restart this protocol too.vision, energy and grey hair have started to worsen, nothing like baseline before antiaging protocols but i feel aging has restarted

 

i see there are new ingredients like dihydromyricetin and sunflower lecithin, are these very important?can i just make stearic acid powder by a mixer and blend it with oat milk and banana or strawberries to improve flavor like i did previosuly?or foods mixed with stearic acid are important to get best results?

 

thank you

 

[Nate-2004]

Turnbuckle where are you sourcing your C60 oil these days are you still making it yourself? I remember you were doing MCT oil for a while instead of olive oil, which precludes the theory about polyphenols being the key. Also where do you source the stearic acid?

Edited by Nate-2004, 05 May 2022 - 05:47 PM.

[Turnbuckle]

Turnbuckle

hi i see this protocol made amazing results, i stopped it in 2020 due to humeral head osteonecrosis after an accident while swimming.the osteonecrosis area started receding on MRI in 2021 thanks to teriparatide treatment/local stemcells activation by reac frequencies treatment and i am fully asymptomatic now so doctors urged me to restart swimming as soon as possible.so i was thinking to restart this protocol too.vision, energy and grey hair have started to worsen, nothing like baseline before antiaging protocols but i feel aging has restarted

 

i see there are new ingredients like dihydromyricetin and sunflower lecithin, are these very important?can i just make stearic acid powder by a mixer and blend it with oat milk and banana or strawberries to improve flavor like i did previosuly?or foods mixed with stearic acid are important to get best results?

 

thank you

 

 

If you use only stearic acid for fusion, you will eventually deplete stem cell pools in the brain, as stearic acid does not cross the BBB.

 

Turnbuckle where are you sourcing your C60 oil these days are you still making it yourself? I remember you were doing MCT oil for a while instead of olive oil, which precludes the theory about polyphenols being the key. Also where do you source the stearic acid?

 

While the olive-oil-only rats lived 15% longer in the original trials, neither polyphenols nor the antioxidant properties of C60 are key. The researchers made a wrong assumption. You can use either MCT oil or olive oil, but olive oil takes longer to be absorbed, and that's a good thing if you are taking the components together. It gives fusion a head start. And while I have enough in my freezer to last many years, I recently bought a bottle of C60 in olive oil from SES, and it seemed fine. Also note that the concentration will be different. C60 is more soluble in olive oil than in MCT oil.

[stephen_b]

If you use only stearic acid for fusion, you will eventually deplete stem cell pools in the brain, as stearic acid does not cross the BBB.

 

What is the alternative?

[Turnbuckle]

What is the alternative?

 

See the last update: https://www.longecit...ndpost&p=914801

 

The stearic acid brownie may be replaced by a couple of grams of GMS, but the dihydromyricetin could be used alone. Still, I prefer to use more than one fusion agent.

[lost69]

If you use only stearic acid for fusion, you will eventually deplete stem cell pools in the brain, as stearic acid does not cross the BBB.

 

 

While the olive-oil-only rats lived 15% longer in the original trials, neither polyphenols nor the antioxidant properties of C60 are key. The researchers made a wrong assumption. You can use either MCT oil or olive oil, but olive oil takes longer to be absorbed, and that's a good thing if you are taking the components together. It gives fusion a head start. And while I have enough in my freezer to last many years, I recently bought a bottle of C60 in olive oil from SES, and it seemed fine. Also note that the concentration will be different. C60 is more soluble in olive oil than in MCT oil.

 

i have prostaphane pills i can use to cross BBB, they have
10 mg of free and stabilised sulforaphane [CH3-SO-(CH2)4-NCS] derived from
220 mg broccoli seeds (Brassica Oleracea) for one capsule

https://www.prostaph...om/prostaphane/

 

how much should i take of these caps?

 

i will add dihydromyricetin and sunflower lecithin to prostaphane pills anyway

 

thank you
 

Edited by lost69, 06 May 2022 - 10:56 AM.

[Danniel]

If you use only stearic acid for fusion, you will eventually deplete stem cell pools in the brain, as stearic acid does not cross the BBB.

 

Supplementing with 100mg sulforaphane at T+30 would fix that, as sulforaphane does cross the BBB.

Edited by Danniel, 07 May 2022 - 03:14 PM.