Increasing NAD+ Levels Restores Mitochondrial Homeostasis through the SIRT1-HIF-1α-c-Myc Pathway

CR is known to delay numerous diseases of aging in mammals, including cancer and type 2 diabetes. Interestingly, CR (30%–40% instituted at 6 weeks) completely prevented the decline in VHL and the increase in HIF-1α that occurs in ad-libitum (AL)-fed 22-month-old mice (Figure 7A). The observed decreases in NAD+ and ATP levels, COX activity, mtDNA, and mitochondrially encoded OXPHOS components with age were also prevented by CR (Figures 7B–7D, S6A, and S6B). Unlike the accumulation of mutations in mtDNA, the pathway that we describe here should be rapidly reversible. Treatment of 22-month-old mice for 1 week with NMN, a precursor to NAD+ that increases NAD+ levels in vivo (Yoshino et al., 2011), reversed the decline in VHL and accumulation of HIF-1α (Figures 7E and 7F); reduced lactate levels; and increased ATP, COX activity, and mitochondrially encoded OXPHOS transcripts (Figures 7G–7I and S6D). In EglN1 and SIRT1 iKO mice, however, NMN failed to induce mitochondrially encoded genes or to raise ATP levels (Figures 7J–7L). Knockdown of NMNAT1 also prevented NMN from inducing mitochondrially encoded OXPHOS genes (Figure 7M), consistent with nuclear NAD+ being a key regulatory molecule. The SIRT1 iKO and the 22-month-old mice had increased levels of markers of muscle atrophy and inflammation compared to young WT mice, along with impaired insulin signaling and insulin-stimulated glucose uptake (Figures S1G–S1J and S6E–S6H). Strikingly, treatment of old mice with NMN reversed all of these biochemical aspects of aging and switched gastrocnemius muscle to a more oxidative fiber type (Figures S6E–S6H). However, we did not observe an improvement in muscle strength (data not shown), indicating that 1 week of treatment might not be sufficient to reverse whole-organism aging and that longer treatments might be required.

Discussion

Impairment in mitochondrial homeostasis is one of the hallmarks of aging that may underlie common age-related diseases (Lanza and Nair, 2010, Satoh et al., 2013). Despite its importance, there is still controversy as to why mitochondrial homeostasis is disrupted with age and whether this process can be slowed or even reversed. Here, we present evidence for a PGC-1α/β-independent pathway that ensures OXPHOS function and maintenance of mitochondrial homeostasis (Figure 7N). During aging, however, decline in nuclear energetic state or NAD+ levels reduces the activity of SIRT1 in the nucleus, causing VHL levels to decline and HIF-1α to be stabilized. This program, which likely evolved to modulate mitochondrial metabolism in response to changes in energy supply, becomes chronically activated in old mice, inducing a pseudohypoxic state that disrupts OXPHOS, a phenomenon that is consistent with antagonistic pleiotropy (Williams and Day, 2003).