Present investigations and attempts to influence nicotinamide adenine dinucleotide (NAD+) metabolism in aging might be viewed as the direct descendant of the heavily hyped sirtuin research of a nearly decade ago. We can check the boxes for (a) mechanisms linked to mitochondrial activity, (b) supplements claimed to adjust age-related changes in those mechanisms, and (c) many of the same people in the scientific community being involved. At the end of the day this may well arrive at the same destination as that sirtuin development, which is to say nothing of any practical use to improve human longevity, but at least the outcome of an incrementally greater understanding of this narrow section of mammalian cellular metabolism.
The data for benefits to result from some of the presently available supplements that might increase NAD+ levels is admittedly considerably better than was the case for tinkering with sirtuins, but that is nonetheless a low bar to pass. Even so, some of these approaches clearly produce the same old story of unreliably, tiny effects that tend to vanish given more care and more rigorous studies. The best outcome we could hope for in the near future is a modestly useful exercise mimetic that can help the unfit to evade some fraction of the consequences of their lack of fitness. Rigorous data has yet to arrive, however, and it could all still come to nothing much yet. That best outcome is still not rejuvenation in any meaningful sense. It is tinkering with the damaged machinery, hoping to get a few extra percentage points of operational capacity out of it, without actually trying to fix the breakages.
Does loss of NAD+ and related decline in mitochondrial function affect the processes that cause cells to become senescent, and thereafter linger and cause harm to surrounding tissues? Since mitochondria are central to most aspects of cellular health, by virtue of providing the chemical energy stores needed for the cell to run at all, and are also central to the processes of programmed cell death that determine whether senescent cells self-destruct or linger, this seems plausible. The details matter, however: what is the size of the effect, in comparison to, say, the decline of the immune system, or other factors in age-related mitochondrial dysfunction, in determining how many senescent cells are present in a tissue? How does it vary between tissues? Further, the methodology used here to reduce NAD+ levels may or may not be a good substitute for what takes place in aging; as a general rule, few such models are, and it is always a question of whether or not the differences happen to be large enough or relevant enough to be a problem for the present area of interest.
The retinal pigment epithelium (RPE) performs numerous functions essential to normal retinal health and function. RPE is implicated directly and prominently in the pathogenesis of most degenerative diseases of the retina, including age-related macular degeneration (AMD), the leading cause of blindness among persons aged 60 and above worldwide. AMD is a complex multifactorial disease and, as its name implies, age is a primary risk factor for its development.
Interestingly however, most available experimental models and related studies have focused more heavily on identifying, understanding, and limiting secondary consequences of aging and related RPE dysfunction (e.g., increased oxidative stress, inflammation, altered cholesterol metabolism) as opposed to targeting directly factors, such as energy deprivation, that precipitate accelerated aging at a cellular level. The consequence of the latter is an imbalance in homeostatic processes and subsequent damage, as shown in many specific cell types. This is the premise of a number of recent studies including the present investigation in which we focused on nicotinamide adenine dinucleotide (NAD+) and factors governing its bioavailability in relation to the overall impact on RPE viability.
NAD+, a central metabolic cofactor, plays a critical role in regulating cellular metabolism and energy homeostasis. The ratio of NAD+ to NADH (oxidized to reduced NAD+) regulates the activity of various enzymes essential to metabolic pathways including glycolysis, the Kreb's cycle, and fatty acid oxidation. There is a wealth of clinical and experimental data stemming from studies of other primary diseases of aging demonstrating clearly a generalized decline in the availability of NAD+ in association with increased age and the related reduction in the activity of a number of downstream metabolic pathways that contribute to the development and progression of degenerative processes.
Neuronal cells and tissues appear to be especially sensitive in this regard. Importantly, the aforementioned studies additionally suggest that age-related degenerative processes might be prevented or at the least, the consequences thereof lessened in severity by therapies that boost NAD+. In the present investigation we focused on evaluating the impact of NAD+ and factors that regulate its availability on RPE viability both in vivo and in vitro. Cellular senescence is a common consequence of aging hence, the decline in NAD+ in RPE and the associated upregulated expression of markers of senescence that we observed was not totally surprising. Though there has been some debate over whether dysfunction occurs first in the RPE or in the overlying photoreceptors, the contribution of senescence-associated RPE damage to age-related RPE dysfunction is undeniable.
Here, using adult C57BL/6J mice across a broad range of ages (2-18 months), we first confirmed that NAD+ levels decline significantly in association with increased age as has been reported to occur in other retinal and non-retinal cell types. Our related evaluation of enzymes that drive key steps in NAD+ biosynthesis revealed NAMPT as the enzyme principally responsible for maintaining adequate NAD+ levels in RPE. This is congruent with recent work by others demonstrating that NAMPT-mediated NAD+ biosynthesis is essential for proper visual function. We used the compound to optimize a cell culture model system that allowed us to simulate and study the impact of decreased NAMPT expression and related NAD+ availability on RPE cell viability relevant to aging. Our studies in the human RPE cell line ARPE-19 demonstrated an increase in RPE cell senescence in conjunction with reduced NAMPT and NAD+ availability as indicated by analyses of the expression of senescence markers.
Our present data demonstrating an age-dependent decline in NAMPT expression and in turn, NAD+ generation in RPE which ultimately promotes RPE senescence supports strongly the rationale for enhancing NAMPT expression and associated NAD+ generation therapeutically. Indeed, such therapies may represent a viable strategy for preventing and treating RPE and consequent photoreceptor damage in aging/AMD and broadly, in other degenerative retinal diseases in which RPE is prominently affected.