Robust Mouse Rejuvenation Project

[John_Hemming]

I was experimenting with cholecalciferol and I took 12,000 and 24,000 iu (on two separate occasions. This disrupted my sleep for a day or so. In fact even 6,000 was a bit high. Hence I have obtained some 25OHD (Dedrogyl) and I use that to bring my D3 up every so often. On a daily basis I take 3,000 which I find my body can process without side effects. My initial conclusion is that cholecalciferol itself is mildly toxic until processed into 25OHD.

The background is that I wanted to get an idea as to how quickly my body converts large doses of cholecalciferol into 25OHD and how long the benefit lasted (which I measured by the impact on my sleep).

[L_H]

Why such a high dose?
I’ve received my rapamycin today and it came with a vitamin d3 ‘Bomb’ cholecalcierol sixty thousand iu
is this to boost immunity whilst on rapa?

[John_Hemming]

The reason I took 12,000 and 24,000 was to see what happened. I think of the metabolism as a system in flux rather than something static. To me it is obvious that there would be a limit on processing from D3 into 25OHD. I wanted to see what that limit was.

I am continually experimenting with all sorts of metabolic processes.

I do check first that it is unlikely that any serious harm will be done, but I don’t get stressed with side effects. I can always stop what I am doing.

I now have a good vitamin D protocol which enables me to keep my 25OHD levels up around the top of normal. If I stopped my weekly blood tests I would have some idea what to do to achieve that.

[AlexKChen]

Are they going to transcend statistical significance? If you just double a SINGLE mouse’s lifespan, you don’t need N = 30 - the result is so obvious

[Ilya_M]

First survival curves

Aubrey de Grey

[RapAdmin]

An updated on the Robust Mouse Rejuvenation project (from Andrew Steele at the Dublin Longevity Summit):

Current drugs/therapies being tested:

RMR Version 2 planned?

Discussion / coverage by Andrew Steele:

https://twitter.com/statto/status/1692812476725793084

[RapAdmin]

A new update from Aubrey:

[Ilya_M]

An update.

[RapAdmin]

Its a little hard to figure out some of them, but rapamycin and HSCs seem to be doing extremely well, as well as the “all”.

Here is the legend to better understand it all:

Robust Mouse Rejuvenation - Study 1 — LEV Foundation

[Ilya_M]

Looks like HSCs work much better with males than with females. But it definitely very reassuring that “all” works the best for both.
Aubrey in his interviews mentioned the costs of the experiment, but I’ve forgotten. Do you know by any chance?

[RapAdmin]

About $3 Million.

Source: Robust Mouse Rejuvenation Study Started | NextBigFuture.com

[DeStrider]

Actually looks like Rapamycin is the best if you consider both genders.

[jakexb]

Am I misunderstanding or is HSCs just straight up infusing stem cells into the mouse? I’m shocked that is doing anything at all, much less going head to head with rapamycin.

[RapAdmin]

From the study design link I posted earlier:

Hematopoietic Stem Cell Transplant

Rationale: A reduced regenerative capacity of stem cells is widely believed to contribute to age-related morbidity and functional tissue decline. Numerous studies have shown stem cell transplantation to have rejuvenating effects in mouse tissues, and several additionally show a lifespan extension effect [PMID 32012439, 31031800].

Design: The study design is largely (but see below) based on the protocol utilized by Guderyon et al. 2020 [PMID 32012439] and consists of mobilizing the recipient bone marrow niche followed by transplant of lineage-depleted hematopoietic stem cells (HSCT).

Bone Marrow: We have opted to use lineage-depleted bone marrow HSCs as opposed to additional selection for and expansion of long-term self-renewing HSCs. This was on the basis that 1) prior lifespan studies used whole or lineage-depleted bone marrow, which may include MSCs or other beneficial cells promoting engraftment, and 2) expansion protocols have not been extensively validated.

Mobilization Protocol: We will follow the mobilization protocol for recipient mice as outlined in Guderyon 2020, consisting of G-CSF + AMD3100. Although other newer mobilization reagents require fewer injections, efficacy is not well established; the current protocol is most common for chemical mobilization, has been used in aged mice, and remains the current standard of care for mobilizing HSCs from human BM donors.

# of cells: Each administration will consist of ~2x10⁷ HSCs, derived from 8 sex-matched C57Bl/6J PEP-BOY (CD45.2) donor mice per injection. This number is four-fold greater than the Guderyon study, however in line with other HSCT studies [PMID 31031800, 18491294]. Additionally, it is demonstrated that % donor cell engraftment is largely based on competition with mobilized recipient cells; thus, increasing the donor to recipient ratio in the blood may allow for increased engraftment with each administration allowing for fewer transplants overall and thus less stress on the animals.

Number of Transplant Injections: While the lifespan cohort of the Guderyon study received a total of 8 HSCTs, each round administered only 5x10⁶ lineage-negative (long-term repopulating) donor cells. We wish to minimize the number of administrations for each mouse. Therefore, we will perform 2 rounds of HSCT, a month apart, and then assess the % donor-derived cells (CD45.2) in peripheral blood after 4-8 weeks (timing to coincide with planned blood draw). Whether or not additional rounds of HSCT are performed (for a given experimental group) will be decided based on percentage engraftment in recipient mice.

[jakexb]

Wow, I would not have seen that one coming as a contender

[misterios]

1. We can see clear gender differences and this is something we have also seen when ITP (Intervention Testing Program) has tested different compounds in mice. It’s interesting that we see differences in stem cell therapy but not gene therapy. Does anyone know what could be the cause why we see such a big gender difference in stem cell therapy?

2. Rapamycin seems to play a key role in several of the top synergistic cocktail combinations in the top list. This is also well aligned with my assumption why Rapamycin should be a base ingredient in the powerful longevity cocktail we will engineer to extend maximum lifespan. My guess is therefore that the best longevity cocktails will be the ones which will have effects on both slowed and reversed aging.

3. It’s very interesting that the best intervention is when all interventions were combined but it’s also interesting that female mice get the same effect if they only take Rapamycin. Let’s see how things develop in the upcoming months!
   By @KristerKauppi

[Lost]

Heh. I don’t know how to interpret that. The all interventions except gene therapy survival is terrible in males, but the other things don’t agree. So presumably one needs to dig deeper into the error ranges.

[Ilya_M]

Second study overview

Robust Mouse Rejuvenation - Study 2 — LEV Foundation

[RapAdmin]

Baseline treatments: Combination therapies are only valuable if their benefit exceeds that of the best known alternative. To date, the most effective rejuvenation treatments are rapamycin, caloric restriction, and exercise. We carefully considered these in the context of RMR1, opting to include rapamycin as one of the four interventions for comparison. For RMR2, we are considering giving rapamycin to ALL the animals in all treatment groups. This would allow us to gauge the efficacy of other rejuvenation interventions when the overall damage burden is already slightly lowered.

[DeStrider]

Actually, that’s a pretty smart way of doing this. Maybe they should administer Rapa and have the mice exercise and then gauge the effectiveness of treatments.