Today, we have an interview with Dr. Michael Bonkowski, an expert on NAD+ biology and aging from the David Sinclair Lab, Harvard Medical School.
Michael Bonkowski aims to advance our understanding of the links between metabolism, aging, and age-associated diseases. He has published 35 peer-reviewed journal articles and has conducted multiple successful longevity studies. In Dr. David Sinclair’s lab, his research efforts are focused on the role of nutrient sensors’ regulation of endocrine signaling and aging in the mouse. He is also working on direct and indirect ways to drive the activity of these nutrient sensors by using dietary manipulations, small molecules, and chemical treatments.
Michael is trained as a pharmacologist, physiologist, and animal scientist. Some of his areas of expertise include animal physiology, genetics, glucose, and insulin homeostasis, metabolism, assay development, protein biochemistry, and transmission electron microscopy imaging.
As we are currently hosting the NAD+ Mouse Project on LIfespan.io, which includes Michael on the research team, now is an ideal time to talk to him about his work and why the project is important for aging research.
For those new to the topic, can you tell us what NAD+ is?
NAD+ is a bioactive metabolite. It is well known for its role as a high-energy hydride carrier between glucose and mitochondria. It is involved in over 500 enzymatic and biochemical reactions, with multiple research groups reporting an age-related decline. NAD+ is also studied for its role in the body as a resistance sensor against diseases of aging.
Some people simplify the role of NAD+ in metabolism as just being part of nutrient sensing and often see it as only being a caloric restriction mimetic. NAD+ has a lot more roles in metabolism than this; could you tell us what some of them are?
NAD+ is most well-known for its role in redox metabolism, as a cosubstrate for enzymatic reactions and as a downstream metabolite of Vitamin B3 (a.k.a. niacin or nicotinic acid). Its levels do go up during calorie restriction and exercise, which are currently the best-known interventions for slowing aging.
More information can be learned by looking at these reviews by our group and others: Canto et al. 2015, PMID: 26118927; Verdin, 2015 PMID 29031725; Bonkowski & Sinclair 2016, PMID 27552971; Rajman et al. 2018, PMID 29514064.
There are a number of what we call NAD+ precursors, including NR, niacin, and NMN. Can you explain how they are different from each other and what makes NMN potentially the most interesting in regards to NAD+ repletion?
There are many metabolic routes by which NAD+ is produced. The body can make it de novo from tryptophan or from external vitamins, such as niacin or NR. Most importantly, free nicotinamide in our cells, the product of NAD+ hydrolysis, can be remade into NAD+ through the salvage pathway. In this pathway, NMN is the direct precursor to NAD+ – just one enzymatic step away. In our lab, we found that giving NMN exogenously can raise NAD+ in most of the tissues that we have tested (liver, muscle, kidney, immune cells, and blood).
Why use precursors; why not just introduce NAD+ into the bloodstream for the cells to uptake?
We find that administering NMN can significantly increase cellular and tissue pools of NAD+.
NAD+ is known to facilitate DNA repair by preventing PARP1 and DBC1 binding; can you explain a little bit more about how this happens?
Our lab was the first to show that NAD+ is involved in mediating protein-protein interactions between DBC1 and PARP1. With age-associated declines in NAD+, we find that DBC1 is bound to and inhibits PARP1. Increasing NAD+ levels through exogenous NMN supplementation decreases this interaction and restores PARP1 activity and DNA repair (Li, Bonkowski & Sinclair et al. 2017, PMID:28336669).
Does NAD+ also influence mitochondrial DNA (mtDNA) repair by this same PARP1/DBC1 mechanism?
We have shown that NAD+ in mitochondria is important for cell survival. While PARP1 is present in the mitochondria, we have not found evidence that DBC1 is present in the mitochondria.
Has anyone looked at how NAD+ repletion affects mitochondria; does it help to improve their function and/or reverse aberrant age-related dysfunction?
Many groups have looked at the effects of NAD+ repletion on mitochondria function and biogenesis. These studies have reported that administering NAD+ boosting compounds increases mitochondrial NAD+ altering activity and biogenesis (reviewed in Canto et al. 2015, PMID: 26118927). Work from our lab has found that with the age-associated waning of NAD+ levels, there is miscommunication between the nucleus and mitochondria. We also found that supplementing NMN to raise NAD+ could restore this communication breakdown, leading to a youthful phenotype (Gomes & Sinclair et al. 2013 PMID: 24360282).
NAD+ and SIRT1 were shown to mediate crosstalk with muscle tissue and blood vessels earlier this year. Could we potentially boost NAD+ levels to combat age-related frailty and sarcopenia?
We are very excited about this study. We found that advanced aged mice received a benefit in muscle vascularization and running endurance from consuming NMN in their drinking water. NAD+ may or may not have a role for other muscle pathologies, such as sarcopenia.
What types of mice are you using in the experiment?
For our advanced age longevity, behavior and cognition experiments, we are using the C57BL/6 mouse line, which is used frequently in the aging field. We chose to start treating these mice at an advanced age (20 months = ~50 in human years) to circumvent developmental issues associated with using transgenic or knockout mouse lines. In addition, we will use our progeroid mouse model for cognitive and behavioral testing to accelerate results.
So, you are using both normal mice and a progeric mouse that experiences “accelerated aging”. How is your progeric model different to the progeric mouse strains already available, or how is it a closer analog to regular aging?
Our progeroid model is an accelerated aging model driven by induced changes to the epigenome (ICE). This model has many changes similar to human aging, and we will use these mice to address certain behavioral or physiological questions to accelerate data collection.
Some people criticize the use of progeric mice as not really representing normal aging; how do the modified mice you are using address some of those concerns?
The ICE mouse is a novel and, we believe, an accurate model of aging that is based on accelerating an underlying cause of aging.
You are conducting a lifespan study with normally aging mice, which is a world first for NMN, but does NAD+ work the same way in mouse cells as it does in human cells?
While there are indeed many differences between mice and humans. NAD+ is a common metabolite across all of life, and at the biochemical level, its functions are similar between species.
Conducting a lifespan study is difficult with all kinds of potential confounding factors that could affect the data, such as diet, environment, and so on. What are the biggest challenges for you and your team conducting a lifespan study?
We have conducted multiple longevity studies in mice and have experience in conducting these types of experiments. We house mice in a completely controlled environment and put a lot of thought into the design of the studies to reduce the confounding factors to an absolute minimum. We have already started a pilot study that is in progress and seeks funds to execute and complete the comprehensive experiments.
How large is your lifespan mouse cohort, and how did you determine that these amounts would be statistically sound and avoid the statistical “noise” some studies fall foul of?
We plan to use approximately 40 mice per group. Based on our statistical power analysis, with this N value, we are powered to detect small differences in average and maximal lifespan. In addition, we will conduct behavioral and cognitive tests.
Given that NMN is already in human trials, what could this experiment on Lifespan.io add to our knowledge of NAD+ and aging?
There are phase 1 studies underway at Metrobiotech, and David Sinclair is one of the founders of this company. If we wish to know if NMN has a longevity benefit and improves many age-related diseases, we can have that answer using this mouse study in less than two years. Currently, there are no human trials in the U.S. looking at aging or longevity in response to dietary NAD+ supplementation.
Regarding the NAD+ Mouse Project on Lifespan.io, why does the lab need to do this when it is part of a prestigious University?
Conducting research is very costly. We rely on grants and donations just like the majority of University labs. We are performing comprehensive tests and appreciate any monetary contribution allowing us to complete this important study.
Will the data be open access?
We would aim to pre-publish in biorxiv then publish in a peer-reviewed journal following that. The pre-published data would be open access which anyone can read though it will not have gone through the peer review process at this point.
We would like to thank Michael for taking the time to speak with us about his work and if you would like to learn more about the NAD+ Mouse Project check out their project page over at Lifespan.io.