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Manipulating mitochondrial dynamics

nad nad+ c60 mito fission fusion stearic acid mtdna methylene blue

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#1711 muntjac

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Posted 29 January 2021 - 10:03 PM

Does Turnbuckle explain anywhere the rationale of 3 days of fission and 2 days of fusion? My understanding is that mitophagy and mito biogenesis occur in unison so that your total pool of mitochondria remains constant. As such it doesn't make sense biochemically that increases and fission and fusion can be spilt. If I have any of this incorrect, please do explain why that is the case. Thank you in advance.

 

The supplements would interfere with each other if not split between days.


Edited by muntjac, 29 January 2021 - 10:04 PM.

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#1712 muntjac

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Posted 30 January 2021 - 01:42 AM

Sulforaphane is a Nrf2-independent inhibitor of mitochondrial fission

 


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#1713 nikolay

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Posted 30 January 2021 - 02:50 AM

d-Ribose as a Contributor to Glycated Haemoglobin


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#1714 CuriousMonkey

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Posted 30 January 2021 - 07:30 PM

The supplements would interfere with each other if not split between days.

 

Muntjac,

 

This my point what is the evidence that they would "interfere"? mitophagy and biogenesis occurred in unison, if so any intervention that increased mito biogenesis would in turn be increasing mitophagy otherwise the total pool of mitochondria would not remain constant, which is what has been found to be the case in experiments. I would be highly interested in any evidence to the contrary.


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#1715 muntjac

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Posted 31 January 2021 - 04:02 PM

I might be mistaken, my understanding was that the supplements are up or down-regulating competing aspects of dynamics, and are more efficient when taken separately for that reason.


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#1716 CuriousMonkey

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Posted 31 January 2021 - 08:49 PM

I might be mistaken, my understanding was that the supplements are up or down-regulating competing aspects of dynamics, and are more efficient when taken separately for that reason.

 

https://pubmed.ncbi....h.gov/25896323/

 

"Our findings reveal a homeostatic feedback loop that integrates metabolic signals to coordinate the biogenesis and turnover of mitochondria."

 

So if you take a supplement that is thought to increase mitophagy it will result in an increase in biogenesis because they are coupled. Conversely, if you take a supplement that is thought to increase biogenesis it will increase mitophagy. 

 

I would love to see any evidence to the contrary. 


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#1717 muntjac

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Posted 31 January 2021 - 09:14 PM

I already linked to a study showing that fission can be inhibited by one of the supplements used to promote fusion. 


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#1718 CuriousMonkey

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Posted 31 January 2021 - 09:54 PM

I already linked to a study showing that fission can be inhibited by one of the supplements used to promote fusion. 

 

Muntjac, I'm not a science expert as is the case with most people here so please help me understand this contradictory evidence. Animal studies finding that the fusion/fission dynamics are tightly bound and the cell line study that you reference which contradicts this. Thanks. 


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#1719 Turnbuckle

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Posted 31 January 2021 - 09:56 PM

https://pubmed.ncbi....h.gov/25896323/

 

"Our findings reveal a homeostatic feedback loop that integrates metabolic signals to coordinate the biogenesis and turnover of mitochondria."

 

So if you take a supplement that is thought to increase mitophagy it will result in an increase in biogenesis because they are coupled. Conversely, if you take a supplement that is thought to increase biogenesis it will increase mitophagy. 

 

I would love to see any evidence to the contrary. 

 

 

This from second link in the OP --

 

NAM-induced Decrease of Mitochondrial Content Requires NAM Conversion to NAD+
Fig. 1A shows an example of the change in the mitochondrial content that is commonly observed in various tested human cells (including normal fibroblasts and MCF-7, H460, and HCT116 cancer cell lines) after supplementation of 5 mm NAM in culture media. The mitochondrial content as determined by flow cytometry using two different mitochondrion-specific dyes was substantially decreased for the first 3 days and thereafter remained at a level about 70% that in the untreated cells, as reported previously

 

 


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#1720 muntjac

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Posted 01 February 2021 - 07:36 PM

It's not contradictory, fission and fusion can happen simultaneously while also being inhibited or activated by numerous factors. Search PubMed for Drp1 and OPA-1, these are two of the proteins which regulate fission and fusion, there are hundreds of results:

 

A Disturbance in the Force: Cellular Stress Sensing by the Mitochondrial Network

As a highly dynamic organellar network, mitochondria are maintained as an organellar network by delicately balancing fission and fusion pathways. This homeostatic balance of organellar dynamics is increasingly revealed to play an integral role in sensing cellular stress stimuli. Mitochondrial fission/fusion balance is highly sensitive to perturbations such as loss of bioenergetic function, oxidative stress, and other stimuli, with mechanistic contribution to subsequent cell-wide cascades including inflammation, autophagy, and apoptosis.


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#1721 CuriousMonkey

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Posted 02 February 2021 - 05:33 PM

 

This from second link in the OP --

 

 

 

 

Interesting. Thank you for replying. 

 

https://pubmed.ncbi....h.gov/29185343/

In your protocol you use PQQ to promote fusion. 

"Our results suggest that PQQ-inducible mitochondrial biogenesis can be attributed to activation of the SIRT1/PGC-1α signaling pathway by enhancing cellular NAD+ formation."

 

So supplementing with Nicotinamide increases fission by increasing NAD+ and supplementing with PQQ increases fusion by increasing NAD+.

 

Do I have this right? 


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#1722 Turnbuckle

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Posted 02 February 2021 - 07:18 PM

Interesting. Thank you for replying. 

 

https://pubmed.ncbi....h.gov/29185343/

In your protocol you use PQQ to promote fusion. 

"Our results suggest that PQQ-inducible mitochondrial biogenesis can be attributed to activation of the SIRT1/PGC-1α signaling pathway by enhancing cellular NAD+ formation."

 

So supplementing with Nicotinamide increases fission by increasing NAD+ and supplementing with PQQ increases fusion by increasing NAD+.

 

Do I have this right? 

 

 

Higher NAD+/NADH (oxidized to reduced) ratios induce mito fission and mitophagy (PINK1/Parkin QC requires fission, as do lysosomes) . This ratio can be raised with nicotinamide (N), nicotinamide + ribose (N+R), nicotinamide riboside (NR), and nicotinamide mononucleotide (NMN).

 

Stearic acid induces fusion that overrides fission.

 

PQQ promotes mito biogenesis. 


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#1723 CuriousMonkey

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Posted 02 February 2021 - 10:05 PM

Higher NAD+/NADH (oxidized to reduced) ratios induce mito fission and mitophagy (PINK1/Parkin QC requires fission, as do lysosomes) . This ratio can be raised with nicotinamide (N), nicotinamide + ribose (N+R), nicotinamide riboside (NR), and nicotinamide mononucleotide (NMN).

 

Stearic acid induces fusion that overrides fission.

 

PQQ promotes mito biogenesis. 

 

Turnbuckle,

 

Thank you again for taking the time to reply.

 

https://pubmed.ncbi....h.gov/32977419/

"PQQ pretreatment also increased the ratio of LC3-II/LC3-I and expression of Atg5 in BV2 cells stimulated with rotenone. Additionally, the autophagosome observed by transmission electron microscopy (TEM) and co-localization of mitochondria with lysosomes indicated that mitophagy was induced by PQQ in rotenone-injured BV2 cells, and the PINK1/parkin mediated mitophagy pathway was regulated by PQQ."

 

https://pubmed.ncbi....h.gov/24711540/

"Oral administration of nicotinamide riboside (NR), a vitamin B3 and NAD(+) precursor, was previously shown to boost NAD(+) levels in mice and to induce mitochondrial biogenesis."

 

So supplementing with NR induces mitochondrial biogenesis and supplementing with PQQ induces mitophagy or am I misinterpreting the findings? Thank again for the guidance. 


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#1724 Turnbuckle

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Posted 03 February 2021 - 01:59 AM

Look again at the link in post 1719, taken from the OP. If raising NAD+ with nicotinamide produced any substantial biogenesis, mitochondria mass would increase, not decrease. Also look at the following paper, The NAD+ precursor nicotinamide riboside decreases exercise performance in rats. Again, this points to a decline in mito mass, which is not surprising, as NR is substantially broken down into nicotinamide before it reaches the muscles, so it will behave in the same way.

 

This is all ground plowed years ago. So tell me what you goal is in asking these questions. Is this protocol something you intend to try, or do you have some other objective?


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#1725 Kimer Med

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Posted 06 February 2021 - 08:36 PM

I just finished reading all 58 pages of this thread. I recall reading it back in 2017/2018, so it was interesting to see how things have progressed since then. I'm convinced from other work I've done that "mitochondrial disease" (a poorly defined term) is very common. Very promising therapy!

 

One question keeps ringing through my mind, though: Rather than using stearic acid supplements (that are often mixed with palmitic acid), why not just consume foods that contain stearic acid? Getting 10g in food wouldn't be very difficult. Are the supplements intended for people who are on restricted diets? Or is there some benefit to taking a large bolus of stearic acid that's not mixed with the other things that come with food? Along similar lines, taking care to avoid high-stearic acid foods on fission days also seems important.

 

I'm also wondering if there are any lab tests that can be used to help track progress. It might be interesting to look at before-and-after organic acids, for example, which show the concentration and distribution of the intermediate products of the citric acid cycle. In conventional Functional Medicine, maldistributions in that area are normally attributed to nutrient deficiencies, but it occurs to me that the reason may in fact be mtDNA damage.



#1726 Turnbuckle

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Posted 06 February 2021 - 09:25 PM

I just finished reading all 58 pages of this thread. I recall reading it back in 2017/2018, so it was interesting to see how things have progressed since then. I'm convinced from other work I've done that "mitochondrial disease" (a poorly defined term) is very common. Very promising therapy!

 

One question keeps ringing through my mind, though: Rather than using stearic acid supplements (that are often mixed with palmitic acid), why not just consume foods that contain stearic acid? Getting 10g in food wouldn't be very difficult. Are the supplements intended for people who are on restricted diets? Or is there some benefit to taking a large bolus of stearic acid that's not mixed with the other things that come with food? Along similar lines, taking care to avoid high-stearic acid foods on fission days also seems important.

 

I'm also wondering if there are any lab tests that can be used to help track progress. It might be interesting to look at before-and-after organic acids, for example, which show the concentration and distribution of the intermediate products of the citric acid cycle. In conventional Functional Medicine, maldistributions in that area are normally attributed to nutrient deficiencies, but it occurs to me that the reason may in fact be mtDNA damage.

 

All 58 pages! You should get a prize. 

 

I will be posting an update of a much improved protocol around the end of this month. It will be considerably simplified, and will also have a very simple method for tracking your progress.


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#1727 userCK

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Posted 06 February 2021 - 09:56 PM

 

 

If you believe you have a great deal of damage, you might start off with only one day of fission/mitophagy. 

 

 

Is this what you meant? I mean, if I've a great deal of damage, shouldn't I be doing fission for more days, rather than less?



#1728 userCK

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Posted 07 February 2021 - 07:39 AM

I started the Mito Fission protocol today. I am being a bit paranoid here but I think many supplement brands under-dose their products so I nearly doubled dosage of most things because I took 2 different brands for each. @Turnbuckle, I hope that still leads to fission?

 

Nictoinamide 2g & Ribose 2grams:

-- I took Thorne Niacinmine 1.5grams (500mg x3) + Niagen NR 1.5g (300mg x 5) + Jarrow's D-Ribose 2grams (1g x 2) + Carl's Niacin 150mg (50mg x 3, causes flush)

(had strong flush)

 

Jiaogulan leaf 1 gram-

I could only find one brand, Nature Restore, so I took 1.2 grams (200mg x 6) incase it is underodsed

 

Apigenin 100mg:

Could only find one brand (Swanson) so I took Swanson Apigenin 150mg (50mg x3)

 

Fistein 100mg:

I took 200mg (Doctor's best 100mg + Peak Performance 100mg)

 

I also took a spoon (maybe 5 grams) of Coconut Oil incase some require a fat source to dissolve.

 

Am I doing it right @Turnbuckle?

 

 

BTW for fusion days, I don't have Stearic Acid, but I purchased Food Grade Mango Butter so I'd be taking that instead.

 


Edited by userCK, 07 February 2021 - 07:40 AM.


#1729 userCK

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Posted 07 February 2021 - 09:42 AM

Also, for food, I've been on keto like diet and I hope that doesn't interfere with fission or fusion. My normal diet is:

Several Eggs, Steak, Bacon, Salmon filet, Boiled Broccoli (contains Sulforaphane) with coconut oil, mixed vegetables in soup, and a couple of cups of coffee, green tea, BBQ Chicken or Pan Fried Chicken (no breading).

And lots of spiced-dishes like Thai curries (chicken or beef), Indian curries (chicken).

 

 


Edited by userCK, 07 February 2021 - 09:46 AM.


#1730 stephen_b

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Posted 08 February 2021 - 10:19 PM

Is this what you meant? I mean, if I've a great deal of damage, shouldn't I be doing fission for more days, rather than less?

 

No, the full protocol was quite uncomfortable for some people with a lot of mitochondrial damage (due to ciprofloxacin for example), and the recommendation was to ease into it. 



#1731 kurt9

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Posted 08 February 2021 - 11:37 PM

Apparently I did not have a lot of mitochondrial damage when I did it last year since I was not that uncomfortable with it. However, I did manage to give myself the 'rona the first time around when I flew back from business travel on a Saturday, felt a little run down, and dived right into the fission part of the protocol the following Monday. I came down with the ''rona on that Tuesday. 

 

You definitely want to do the social distancing thing while doing this protocol. 



#1732 userCK

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Posted 09 February 2021 - 05:51 AM

No, the full protocol was quite uncomfortable for some people with a lot of mitochondrial damage (due to ciprofloxacin for example), and the recommendation was to ease into it. 

 

Interesting. I don't see ciprofloxacin in the protocol though.

 

Today is Day 3 of the Fission protocol for me and it has been comfortable for me eventhough I've been taking higher-dosage of everything. So, perhaps, I don't have as much Mito damage. Nonetheless, I plan to do a 3 more cycles of this and then move-on to Stem Cell Renewal with C60 protocol, also by Turnbuckle.

 

 

Apparently I did not have a lot of mitochondrial damage when I did it last year since I was not that uncomfortable with it. However, I did manage to give myself the 'rona the first time around when I flew back from business travel on a Saturday, felt a little run down, and dived right into the fission part of the protocol the following Monday. I came down with the ''rona on that Tuesday. 

 

You definitely want to do the social distancing thing while doing this protocol. 

wow! Hope you recovered without any lasting side-effects/symptoms?

 


Edited by userCK, 09 February 2021 - 05:52 AM.


#1733 PAMPAGUY

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Posted 09 February 2021 - 06:11 AM

From what I gather, GTP is synthesized during the Krebs cycle AKA the citric acid cycle (CAC) AKA TCA cycle (tricarboxylic acid cycle). (I have no idea why we have 3 names for the same cycle.)

https://biology.stac...nesis-tca-cycle

 

What feeds the Krebs cycle? Acetyl-CoA

https://en.wikipedia...tric_acid_cycle

 

How does the body produce Acetyl-CoA? Glycolysis and Beta Oxidation

https://en.wikipedia...wiki/Acetyl-CoA

 

Glycolysis requires glucose, and beta oxidation requires fatty acids.

 

So, I'm going to state somewhat the obvious here... it looks like a key component to optimal mitochondrial function is supplying the optimal fuel supply (GTP), and that requires glucose and fatty acids, e.g. stearic acid. Glycolysis and beta oxidation then creates the fuel (Acetyl-CoA) that feeds the generation of GTP.

 

One way to get glucose in your body is to eat it, of course... However, we all know that high spikes in blood glucose levels cause insulin to spike, and that, in general, causes bad stuff to happen, like insulin resistance (type 2 diabetes), or sugar crashes. Clearly, then, the best way to boost glucose levels is with foods that have a low glycemic index, such as apples or whole grain breads.

 

Therefore, in order to have optimal mitochondrial function (read optimal fission and fusion cycles etc.), we also need to eat low glycemic indexed fruits etc. during fatty acid intake and particularly in between meals, like maybe mid-morning and mid-afternoon. So, if we are eating stearic acid for fusion promotion, for instance, we probably need to eat some fruit with it (etc.).

 

Now, the body can store glucose. Perhaps, then this is unnecessary. However, consider the mid-afternoon slump of which many of us are likely familiar. Low blood glucose levels may be partially responsible for the typical mid-afternoon slump. The body doesn't have enough glucose to power the Krebs cycle, and that means low energy levels in general. Maybe this means, generally, a mid-afternoon slump is proof our body has not stored enough glucose to operate at 100%.

New protocol for clearing senescent cells from Dr. Green, only using Dasatinib and Fistein.    https://senolyticstreatment.com/


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#1734 kurt9

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Posted 09 February 2021 - 04:42 PM

Interesting. I don't see ciprofloxacin in the protocol though.

 

wow! Hope you recovered without any lasting side-effects/symptoms?

 

No, I've had no long term symptoms. I actually had the 'rona a second time (late September) because (I suspect) I was taking Circumin when I didn't need it and it suppressed my immune system. I did suffer some head hair loss. But i think that was due mostly to excessive Vitamin A intake, which I stopped in September.


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#1735 Rich D

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Posted 09 February 2021 - 11:38 PM

Also, for food, I've been on keto like diet and I hope that doesn't interfere with fission or fusion. My normal diet is:

Several Eggs, Steak, Bacon, Salmon filet, Boiled Broccoli (contains Sulforaphane) with coconut oil, mixed vegetables in soup, and a couple of cups of coffee, green tea, BBQ Chicken or Pan Fried Chicken (no breading).

And lots of spiced-dishes like Thai curries (chicken or beef), Indian curries (chicken).

 

I'm under the impression that you would be better off on a low protein diet to support fission and a high protein diet to support fusion. Therefore I eat a vegan/vegetarian diet on fission days only.



#1736 aribadabar

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Posted 10 February 2021 - 12:18 AM

Interesting. I don't see ciprofloxacin in the protocol though.

 

Cipro is mentioned as a drug whose possible prior intake may have cause mito damage. It is NOT to be used as part of the protocol.


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#1737 userCK

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Posted 10 February 2021 - 09:18 AM

I'm under the impression that you would be better off on a low protein diet to support fission and a high protein diet to support fusion. Therefore I eat a vegan/vegetarian diet on fission days only.

Interesting. 

 

I read the thread again and it seems to me if you take 2g of Niacinmide + 2g of Riboside (in my case, I doubled the dosage to nearly 4gram each), your body goes in fission. So, eating protein (egg, steak, chicken etc which contain Leucine) wouldn't matter.

 

When you take Stearic Acid (in my case, Mango Butter 20grams), your body would go in fusion and at this time proteins (like Leucine) will matter as it'd cause Biogensis.

 

Cipro is mentioned as a drug whose possible prior intake may have cause mito damage. It is NOT to be used as part of the protocol.

Aah, I see. Yes, I've taken antibiotics in the past; and also taken Statin (Crestor 20mg) for nearly a year.

 

I am on Day 1 of Fusion today (did 3 days of fission):

- Ate 20 grams of Mango Butter, I just mixed it in boiling water and drank it slowly.

- 2 caps of BroccoMax

will wait 3 hours before adding other supplements (leucine etc)

 

so far feel nothing - neither on fission days nor today upon starting fusion. Perhaps Statin (Crestor) did not destroy my mitochondria as much as I suspected it might have had!

 

Nonetheless, I plan to do a few more cycles. Then move on to Stemcell renewal protocol. I'm convinced demethylation of epigenome is going to cure me of Post Finasteride Syndrome! 



#1738 userCK

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Posted 10 February 2021 - 03:28 PM

UPDATE -

 

Didn't feel anything (weakness, lethargy or anything of that sort) in fission days. But today after perhaps 6 hours of starting Fusion, started to feel extreme lethargy. TB mentioned during fusion Leucine etc could get used-up. So, I did this:

- ate some chicken

- ate 6.5 grams of L-Leucine capsules (source naturals)

- 30mg PQQ, 25mg hydroxytyrosol

and lethargy went away!

Also, drank a cup of my evening black coffee, as I usually do. But lethargy went away even before drinking coffee.

 

 

 


Edited by userCK, 10 February 2021 - 03:29 PM.

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#1739 Turnbuckle

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Posted 17 February 2021 - 01:45 PM

An updated Mito protocol

 

The previous protocol can be found at post #1366

 

 

Background:

 

Previously I posted methods of cycling mitochondrial morphology to clean up defective mtDNA, which eliminated mutations via the PINK1/Parkin QC process. The normal QC process can detect mutated mtDNA genes during fission as all mito genes are critical and thus the mito membrane potential goes to zero if just one is defective. Greatly magnifying fission and fusion with supplements will aid that process. But there is another source of mitochondrial damage that isn’t so easily eliminated — epigenetic damage. Like nDNA, mtDNA also picks up aberrant methylation with age. This methylation degrades ATP production, but the QC process doesn’t catch it unless the problem is addressed at a critical time, like during biogenesis. If a mitochondrion with one loop of methylated mtDNA runs out of enzymes while involved with replication, then membrane potential may dip to zero and it will get labeled for recycling. Thanks to methylation, it won’t have as much enzyme reserves as other mitochondria, so it will be preferentially targeted. Also, biogenesis is the best time to demethylate mtDNA as methyltransferase can’t operate while there is only one strand.

 

Until recently, mtDNA wasn’t even known to have methylation, and researchers are still confused as to why it is there. Some speak of mtDNA hypermethylation like it is bad while normal methylation has some purpose.

 

See, for instance: Hypermethylation of mitochondrial DNA in vascular smooth muscle cells impairs cell contractility

 

I don’t agree. I say all mtDNA methylation is bad. Methylated mtDNA mooches enzymes off other mtDNA, and because they don’t produce as much ATP they don’t produce as much ROS, and thus have a survival advantage as they are less prone to mutation. Eventually the cell will become full of moochers and result in fatigue and many other problems of aging.

 

So I say get rid of them all, mutations and methylation alike.

 

The new protocol:

 

This new procedure is much simplified. It requires only two doses, Mito1 and Mito2, which are alternated on a daily basis.

 

Mito1 (fission)

● NAM+R, 1 g of each

● AKG, 1 g

● PQQ, 20 mg

 

Mito2 (fusion)

● GMS, 1 g

● AKG, 1 g

● PQQ, 20 mg

 

NAM+R (nicotinamide plus ribose) is a fission promoter, GMS (glycerol monostearate) is a fusion promoter, AKG (alpha-ketoglutarate) is a demethylase promoter, and PQQ is a biogenesis promoter. All of these are fast acting.

 

A two week experiment using reps to failure:

 

Warm water was sufficient to dissolve everything, but the PQQ was taken in a capsule to insure that the other ingredients got a slight head start (probably unnecessary).

 

Mito1 and Mito2 were taken on alternating days. Each dose was taken in the evening and reps of dumbbell curls to failure counted first thing in the morning — five or six hours after dosing — using the same arm.

 

My hypothesis was that the number of reps would reflect mito damage. With mito fusion, enzymes are shared, thus ATP production and reps would be maximum. With fission, methylated (or otherwise damaged) mtDNA produce less ATP and reps would be minimum. The difference would reflect average damage, and if the treatment worked, the difference should decline. If all damage was removed, then the difference should go to zero.

 

Which in fact it did. See the plot below. The y-axis shows the reps and % difference, while the x-axis shows days. The curve labeled baseline is without any treatment, and likely reflects the normal intermediate situation with mito morphology in a dynamic state. It is stable at 16 reps. The upper fusion curve is relatively flat and higher than baseline as expected, while the lower fission curve is lower than baseline, but rises to meet the fusion curve after about two weeks, and stays there. Thus the percent difference goes to zero.

 

Results:

 

Improvement in running endurance, reduced hunger, and reduced need for hypertension medication.

 

 

 

Attached Files


Edited by Turnbuckle, 17 February 2021 - 01:56 PM.

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#1740 Blueflash

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Posted 17 February 2021 - 11:40 PM

To say that you're an asset to this community would be selling you short. Even if the protocol doesn't add one day to my life but instead increases healthspan, that's a very big deal. Thanks for all that you do.
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