LONGECITY

 

 

SIRT1-dependent restoration of NAD+ homeostasis after increased extracellular NAD+ exposure

 

 

Open AccessDOI:https://doi.org/10.1...jbc.2021.100855

 

 

Abstract

 

 

In the last several years, nicotinamide adenine dinucleotide (NAD+) supplementationhas emerged as an innovative and safe therapeutic strategy for a wide spectrum of disorders, including diabetes and neuropathy. However, critical questions remain as to how NAD+ and its precursors are taken up by cells, as well as the effects of long-lasting intracellular NAD+ (iNAD+) increases. Here, we investigated the kinetics of iNAD+ levels in different cell types challenged with prolonged exposure to extracellular NAD+ (eNAD+). Surprisingly, we found that after the initial increase, iNAD+ contents decreased back to control levels (iNAD+ resetting). Focusing our attention on Hela cells, we found that oxygen and adenosine triphosphate (ATP) consumption ocurred with similar temporal kinetics following eNAD+ exposure. Using [3H]NAD+ and [14C]NAD+, we determined that NAD+ resetting was not due to increased dinucleotide extrusion, but rather to reduced uptake of cleaved NAD+ products. Indeed, eNAD+ exposure reduced expression of the ectonucleotidase CD73, the nicotinamide adenine mononucleotide (NMN) transporter Slc12a8, and the nicotinamide riboside kinase NRK1. Interestingly, silencing the NAD+-sensor enzyme SIRT1 prevented eNAD+-dependent transcriptional repression of CD73, Slc12a8, and NRK1, as well as iNAD+ resetting. Our findings provide the first evidence for a SIRT1-mediated homeostatic response aimed at maintaining physiological iNAD+ levels in conditions of excess eNAD+ availability. These data may be of relevance for therapies designed to support the NAD+ metabolome via extracellular supplementation of the dinucleotide or its precursors.

 

 

full study at link 

 

https://www.jbc.org/...3RtYWlsLmNvbSJ9

Here is ABS comment on this study 

 

https://alivebyscien...3RtYWlsLmNvbSJ9

 

 

 

They found that iNAD+ levels reached the maximum level after 8 hours of continued exposure, after which those pathways were all downregulated.

 

Surprisingly, iNAD+ levels returned back to baseline levels at 24 hours, even though  eNAD+ levels were still high.

In effect, when given a good supply of eNAD+, cells used all pathways to take in more NAD+.  Once iNAD+ was sufficient, pathways for importation of more NAD+ were shut down.

 

So this study would suggest that supplementing pure NAD+ may actually be beneficial I guess. 

 

Also the study has not date on it? Weird. 

Edited by Phoebus, 18 June 2021 - 01:30 PM.

I find this study really because I have been taking ABS's sublingual NAD+ and find it to be a great supplement. And folks kept saying that you could not absorb endogenous NAD because it doesn't penetrate the cell, but this study shows it does. 

 

So if this is the case, then what would be the advantage to taking NMN vs NAD+? It seems there would be no advantage at all. 

 

and this from the study

 

 

 

Interestingly, silencing the NAD+-sensor enzyme SIRT1 prevented eNAD+-dependent transcriptional repression of CD73, Slc12a8, and NRK1, as well as iNAD+ resetting. Our findings provide the first evidence for a SIRT1-mediated homeostatic response aimed at maintaining physiological iNAD+ levels in conditions of excess eNAD+ availability. These data may be of relevance for therapies designed to support the NAD+ metabolome via extracellular supplementation of the dinucleotide or its precursors.

 

so if resveratrol activates SIRT1, then taking some along with your chosen NAD boosting supplementing might be a good idea? Is that what you get from that? 

Edited by Phoebus, 18 June 2021 - 06:20 PM.

I find this study really because I have been taking ABS's sublingual NAD+ and find it to be a great supplement. And folks kept saying that you could not absorb endogenous NAD because it doesn't penetrate the cell, but this study shows it does.

So if this is the case, then what would be the advantage to taking NMN vs NAD+? It seems there would be no advantage at all.

and this from the study


so if resveratrol activates SIRT1, then taking some along with your chosen NAD boosting supplementing might be a good idea? Is that what you get from that?

On the contrary

NAD+ doesn’t enter cells directly. NAD+ degrades into NMn, NR, NAM before entering cells.

NAD+ doesn’t enter cells directly. NAD+ degrades into NMn, NR, NAM before entering cells.

NAD+ doesn’t enter cells directly. NAD+ degrades into NMn, NR, NAM before entering cells.

 

The study specifically says that all three thing happen when eNAD+ is present 

 

 

 

Regardless of the role of CD73 and Slc12a8 in eNAD+ uptake, our data also suggest that eNAD+ can cross the plasma membrane uncleaved. Indeed, in the presence of an extracellular concentration of AMPCP leading to an almost complete CD73 inhibition (see Fig. 2H), eNAD+ can still prompt significant iNAD+ increases. Furthermore, silencing of NAMPT or NRK1 did not reduce the iNAD+ increase under eNAD exposure, suggesting that part of NAD+ was not cleaved. This is in keeping with prior findings on the ability of eNAD+ to trigger iNAD+ increases without evidence for extracellular hydrolysis (32, 33, 35, 36). 

 

CD73 is what converts NAD into NMN and then NR. So even when CD73 is silenced and thus eNAD cannot be converted to NMN, iNAD still increases in the presence of eNAD. The only conclusion to be reached is that NAD is in fact crossing the membrane intact via an unidentified transporter. Remember, the SLC transporter for NMN was only recently discovered. Obviously there could be other transporters out there. 

 

Either that or there is yet another NAD metabolite (beyond NMN and NR) that does not depend on the CD37 pathway and has yet to be identified and that metabolite crosses the membrane and increases iNAD levels. This seems highly unlikely to me, although I could be wrong. 

 

 

Now think of this. When you take NAD+ it can enter the cell three different ways - 1. directly, 2. by being converted to NMN and then entering, or 3. by being further converted to NR and then entering. This seems like a massive advantage over taking NMN which can only enter the cell by one method. 

Edited by Phoebus, 20 June 2021 - 03:16 PM.

The study specifically says that all three thing happen when eNAD+ is present 

 

 

CD73 is what converts NAD into NMN and then NR. So even when CD73 is silenced and thus eNAD cannot be converted to NMN, iNAD still increases in the presence of eNAD. The only conclusion to be reached is that NAD is in fact crossing the membrane intact via an unidentified transporter. Remember, the SLC transporter for NMN was only recently discovered. Obviously there could be other transporters out there. 

 

Either that or there is yet another NAD metabolite (beyond NMN and NR) that does not depend on the CD37 pathway and has yet to be identified and that metabolite crosses the membrane and increases iNAD levels. This seems highly unlikely to me, although I could be wrong. 

 

 

Now think of this. When you take NAD+ it can enter the cell three different ways - 1. directly, 2. by being converted to NMN and then entering, or 3. by being further converted to NR and then entering. This seems like a massive advantage over taking NMN which can only enter the cell by one method. 

 

 

I don't see any proof that it is a "massive advantage", as they don't say how much NAD+ makes it directly into cells.  But yeah, they do say it is not zero.  So NAD+ really should be investigated more as a supplement, but doubt that will happen as the big money is betting on NR or NMN.

 

For sure, this disputes the talking points that NAD+ is trapped and useless, therefore we should take NMN or NR.

 

What I found interesting is they found slc12a8 is important for uptake of NMN to cells, without conversion to NR.  I thought slc12a8 was not really used, but they clearly find otherwise.

Strange how Sinclair, who holds multiple NMN patents, and Brenner, who holds NR patents, are both silent on this study 

 

Odd!

 

Very strange!

 

Wonder why that might be? 

 

hmmmmmm.......

NAD+ doesn’t enter cells directly. NAD+ degrades into NMn, NR, NAM before entering cells.

 

I really wish they hadnt segregated the other thread to the vendor section, but as discussed before, Alive by Science uses liposomes which carries the NAD+ into the cells. So it doesnt matter. - LINK

 

 

 

Levy: Basically, the liposome permits a deep intracellular delivery of its contents via three mechanisms. (Please see figure 4.) When dealing with payloads comprised of nutrients like vitamin C, liposome-encapsulated delivery offers advantages that are not even offered by intravenous delivery, long considered the “gold standard” for nutrient and supplement bioavailability.

Any delivery system that can achieve delivery of its payload (in this case vitamin C) into the cytoplasm, or even more deeply into the mitochondria, endosomes, or nuclei, is nearly always the most desirable way to put that payload into the body, especially without the consumption of energy, or with the relatively minimal consumption of energy.

 

"so if resveratrol activates SIRT1, then taking some along with your chosen NAD boosting supplementing might be a good idea? Is that what you get from that?"  Thorne(pharmaceutical grade vitamin comp) also uses Resveratrol in their NR supplement.  Alive By Science includes it in their NAD+ liposome preparation.  I take an additional micro version of Resveratrol from Revgenetics when doing the NAD+

But I found your own experience you posted in the other thread to be best sign of this products effectiveness(Alive by Nature NAD+ liposomal).

Edited by Gal220, 05 July 2021 - 10:05 PM.

I really wish they hadnt segregated the other thread to the vendor section, but as discussed before, Alive by Science uses liposomes which carries the NAD+ into the cells. So it doesnt matter. - LINK

 

 

 

 

"so if resveratrol activates SIRT1, then taking some along with your chosen NAD boosting supplementing might be a good idea? Is that what you get from that?"  Thorne(pharmaceutical grade vitamin comp) also uses Resveratrol in their NR supplement.  Alive By Science includes it in their NAD+ liposome preparation.  I take an additional micro version of Resveratrol from Revgenetics when doing the NAD+

But I found your own experience you posted in the other thread to be best sign of this products effectiveness(Alive by Nature NAD+ liposomal).

 

I could totally be off base here, because I'm a little out of my element, but based on reading "silencing the NAD+-sensor enzyme SIRT1 prevented eNAD+-dependent transcriptional repression of CD73, Slc12a8, and NRK1, as well as iNAD+ resetting" above that says to me just the opposite, that taking resveratrol "can activate the activities of sirtuins including surtuin-1 (SIRT1)."  Therefore, activating SIRT1 through resveratrol and other supplements helps to reach faster homestasis and reduction of NAD+ back to baseline levels, which is exactly what we don't want.  However I doubt it's healthy to silence SIRT1. 

 

I think the main thing we need to be doing is a protocol like 4 days on 3 days off to help avoid reaching a tolerance where we get nothing out of NAD.  When I first started taking NR it was amazing for the first 6-8 weeks or so, then I definitely reached a tolerance stage where I got little benefit in terms of energy.  I've gone long periods without taking it before taking it again, and I just feel virtually nothing now out of any NR supplement.   And I'm convinced it did help me recover from my gastrointestinal problems I was having several years ago.

Edited by Reven, 16 July 2021 - 01:54 AM.

Taking NR, NMN, NAM long term will cause negative feed back loop. Taking breaks and taking Taurine will help.