NAD+ metabolism: pathophysiologic mechanisms and therapeutic potential

Abstract

Nicotinamide adenine dinucleotide (NAD⁺) and its metabolites function as critical regulators to maintain physiologic processes, enabling the plastic cells to adapt to environmental changes including nutrient perturbation, genotoxic factors, circadian disorder, infection, inflammation and xenobiotics. These effects are mainly achieved by the driving effect of NAD⁺ on metabolic pathways as enzyme cofactors transferring hydrogen in oxidation-reduction reactions. Besides, multiple NAD⁺-dependent enzymes are involved in physiology either by post-synthesis chemical modification of DNA, RNA and proteins, or releasing second messenger cyclic ADP-ribose (cADPR) and NAADP⁺. Prolonged disequilibrium of NAD⁺ metabolism disturbs the physiological functions, resulting in diseases including metabolic diseases, cancer, aging and neurodegeneration disorder. In this review, we summarize recent advances in our understanding of the molecular mechanisms of NAD⁺-regulated physiological responses to stresses, the contribution of NAD⁺ deficiency to various diseases via manipulating cellular communication networks and the potential new avenues for therapeutic intervention.

Fig. 1

Overview of the NAD⁺ metabolism and its physiological function. Mammalian cells can synthesize NAD⁺ de novo from tryptophan by the kynurenine pathway or from NA by the Preiss‐Handler pathway, while most NAD⁺ is recycled via salvage pathways from nicotinamide (NAM), a by-production of NAD⁺-consuming reactions. NAD⁺ can be reduced into NADH in the metabolic processes, including glycolysis, fatty acid oxidation and the TCA cycle. NADH, in turn, drives the generation of ATP via OXPHOS, the production of lactic acid from pyruvate, and the desaturation of PUFAs. NADPH plays an essential role in antioxidant defense and regulates cellular signaling via NADPH oxidases (NOXs). Moreover, NAD⁺ is found to decorate various RNAs in different organisms as nucleotide analog and serves as an alternative adenylation donor for DNA ligation in NHEJ repair. NAD⁺ also acts as a co-substrate for a wide variety of enzymes, including PARPs, sirtuins, CD38/CD157 and SARM1, impacting metabolism, genomic stability, gene expression, inflammation, circadian rhythm and stress resistance. Using NAD⁺ as a co-substrate, both PARPs and sirtuins regulate their target molecules, generating NAM as a by-product. The CD38/CD157 and SARM1 also catalyze NAD⁺ to NAM, producing ADPR and cADPR. Abbreviations: IDOs, indoleamine 2,3-dioxygenase; QA, quinolinic acid; NAMN, nicotinate mononucleotide; QPRT, quinolinate phosphoribosyl-transferase; NAPRT, nicotinic acid phosphoribosyltransferase; NMNATs, nicotinamide mononucleotide adenylyl transferases; NADSYN, NAD synthase; NR, nicotinamide riboside; Trp, tryptophan; NADKs, NAD⁺ kinases; PARPs, poly (ADP-ribose) polymerases; NNT, nicotinamide nucleotide transhydrogenase; TDO, tryptophan 2,3-dioxygenase; SARM1, sterile alpha and TIR motif containing 1; NNMT, Nicotinamide N-methyltransferase; NMN, nicotinamide mononucleotide; PUFAs, polyunsaturated fatty acids; NAM, nicotinamide

Fig. 2

Subcellular equilibrium of NAD⁺. The NAD⁺ homeostasis is maintained by the biosynthesis, consumption and recycling in differentiate subcellular compartments including the cytosol, the nucleus and the mitochondria. NAD⁺ precursors including Trp, NA, NR, NMN and NAM are metabolized into NAD⁺ via Preiss-Handler pathway, de novo pathway and salvage pathway, respectively. NAD⁺ can receive hydride to yield the reduced form NADH in the metabolic processes including glycolysis, FAO, and the TCA cycle. NADH provides an electron pair to drive the mitochondrial OXPHOS for the generation of ATP and the conversion of lactic acid to pyruvate. The cytosolic and mitochondrial NADH is exchanged through the malate-aspartate shuttle and glycerol-3-phosphate shuttle, while the cytosolic and mitochondrial NADPH is exchanged by the isocitrate-a-KG shuttle. NAD⁺ can also be phosphorylated into NADP⁺ by NAD⁺ kinases including nicotinamide nucleotide transhydrogenase (NNT) and NAD kinases (NADKs). Cytosolic NADP⁺ is reduced into NADPH by G6PD and 6PGD in the pentose phosphate pathway, and by ME1 in the conversion of malate to pyruvate. Mitochondrial NADPH is produced by IDH2, GLUD, NNT and ME3. The NADPH is required for the activation of NOXs and the synthesis of palmitate. Abbreviation: α-KGDH, alpha-ketoglutarate dehydrogenase; GLUD, glutamate dehydrogenase; NNT, nicotinamide nucleotide transhydrogenase; G3PDH, glyceraldehyde 3-phosphate dehydrogenase; 6PGD, 6-phosphogluconate dehydrogenase; G6PD, glucose-6-phosphate dehydrogenase; GPx, glutathione peroxidases; IDH1/2, isocitrate dehydrogenase 1 and 2; MDH, malate dehydrogenase; ME1/3, malic enzyme; NADK, NAD⁺ kinase; NOXs, NADPH oxidases; OXPHOS, oxidative phosphorylation; PPP, pentose phosphate pathway; PRx, peroxiredoxin; SDH, succinate dehydrogenase; SOD1-3, superoxide dismutase type 1-3; TCA cycle, tricarboxylic acid cycle; GSH, Glutathione; LDH, Lactate dehydrogenase

Fig. 3

NAD⁺ metabolism controls the redox homeostasis. ROS could be produced from either metabolic reaction in mitochondria, such as OXPHOS, or from a range of cytosolic enzymes, including NOXs, XO, LOX, CYPs, all of which need the NADH/NADPH serving as the electron donor. To maintain the redox homeostasis, both enzymatic and non-enzymatic antioxidant system components exhibit their effects in coordination with each other to contract with the ROS. GSH, the most abundant of non-enzymatic antioxidants, is synthesized from glutamate, cysteine and glycine catalyzed by two consecutive cytosolic enzymes, GCL and GS. Importantly, NADPH serves as the reductive power for ROS-detoxifying enzymes including glutathione reductases (GR) and thioredoxin reductases (TrxR) to maintain the reduced forms of GSH and Trx (SH)₂ in response to ROS produced from mitochondria or NOXs. Abbreviations: 6PGD, 6-phosphogluconate dehydrogenase; CYPs, Cytochromes P450; G6PD, glucose-6-phosphate dehydrogenase; GCL; GR, glutathione reductases; GS; LOX; NAD, nicotinamide adenine dinucleotide; NOXs, NADPH oxidases; NADPH, nicotinamide adenine dinucleotide phosphate; OXPHOS, oxidative phosphorylation; PRx, peroxiredoxin; GPx, glutathione peroxidases; SOD1/2, superoxide dismutase 1 and 2; Trx, thioredoxin; TrxR, thioredoxin reductases; XO, xanthine oxidase

Fig. 4

NAD⁺ serves as a pivotal regulator of gene expression. NAD⁺ and its metabolites are used as substrates and cofactors for reactions that coordinate genomic stability, epigenetic status and RNA processing through NAD⁺-dependent enzymes. NAD⁺-dependent histone-deacetylases, especially SIRT1, possess deacetylase activities on multiple transcription coactivators as well as histones, resulting in epigenome remodeling. The lower activity of sirtuins upon lower level of NAD⁺ may contribute to histone hyperacetylation and aberrant gene transcription. Using NAD⁺ as a (ADP)-ribose donor, PARPs mediate ADP-ribosylation on itself or on a variety of nuclear target proteins such as topoisomerases, DNA polymerases, histones and DNA ligases, playing roles in genome stability and gene regulation, from chromatin to RNA biology. Recently, it has been found that NAD⁺ can also serve as a nucleotide analog in DNA ligation and RNA capping in response to stresses. Abbreviations: CTCF, CCCTC-binding factor

Fig. 5

Physiological actions of NAD⁺ in the host response to infection. Microbial infection, including viruses and bacteria, causes oxidative stress that has a critical effect on both the microbe and host cells. The production of ROS from NOXs depending on NADPH termed respiratory burst is a powerful antimicrobial weapon and a major component of the innate immune defense against bacterial and fungal infections. Meanwhile, oxidative stress causes the host DNA damage that enhances the consumption of NAD⁺ by elevated PARPs. The intracellular NAD⁺ can also be reduced by activation of CD38 that is required for the inflammation against infection. The NAD⁺ deficiency therefore might not be able to support the clearance of microbial by autophagy or phagolysosome, the innate immune and inflammation response. Abbreviations: EBV, Epstein-Barr virus; HCV, hepatitis C virus; HRV, human rhinovirus; HRSV, human orthopneumovirus; iNOS, inducible nitric oxide synthase; ISGs, interferon-stimulated genes; IV, Influenza virus; KSHV, Kaposi’s sarcoma-associated herpesvirus; MPO, myeloperoxidase; Mtb, Mycobacterium tuberculosis; NOXs, NADPH oxidases

Fig. 6

NAD⁺ deficits in aging-associated dysfunction and cancer. Environmental stimuli, including nutrient perturbation, infection, radiation and inflammation, induce oxidative stress, which causes the damage of cellular biomolecules and organelles. NAD⁺ and its metabolites function as crucial regulators to maintain cellular redox homeostasis through replenishing the reducing power or modulating the activity of NAD⁺-consuming enzymes including sirtuins and PARPs. However, disequilibrium of NAD⁺ metabolism could disturb physiological processes, including mitochondria function, circadian rhythm, inflammation, DNA repair and metabolism, leading to aging-associated dysfunction and cancer. NAD⁺ levels could be augmented by dietary NAD⁺ precursor, inhibitors of NAD⁺-consuming enzymes, caloric restriction and exercise. NAD⁺ boosters restore the bioenergetics, redox balance and signaling pathways, ameliorating the adverse effects of pathophysiological conditions, including infection, aging and cancer. Abbreviations: 2-HG, 2-hydroxyglutarate; α-KG, α-ketoglutarate; CCGs, clock-controlled genes; FOXO1, Forkhead Box O1; GSH, Glutathione; IDH1^Mt, mutant isocitrate dehydrogenase 1; NOXs, NADPH oxidase; PER2, period circadian clock 2; PPP, pentose phosphate pathway; PGC-1α, peroxisome proliferator-activated receptor-gamma coactivator alpha; ROS, reactive oxygen species; OXPHOS, Oxidative phosphorylation; TCA cycle, tricarboxylic acid cycle

Fig. 7

Pathophysiological role of NAD⁺ disarrangement in metabolic diseases. a The liver is a master organ of NAD⁺ metabolism and may facilitate the NAD⁺ biosynthesis in other tissues. NAD⁺ metabolism plays a critical role in the lipid metabolism through modulating the activity of sirtuins. The reduced NAMPT expression and NAD⁺ levels contribute to the development of NAFLD through manipulating dysmetabolic imbalance, hepatic energy homeostasis, glucose homeostasis, hepatic inflammation and insulin resistance. b Decreased NAD⁺/NADH ratio by the mismatch between NADH production and oxidation inhibits the activity of sirtuins in the failing heart. Elevated protein acetylation weakens the energy metabolism through negative feedback to OXPHOS and substrate metabolism, impairing antioxidant defense and sensitizing the mPTP to ROS or calcium. c The deduced NAD⁺ levels in kidney are attributed to the decreased expression of enzymes in NAD⁺ de novo synthesis and increased consumption by DNA damage activated PARPs. NAD⁺ depletion inhibits the SIRT1/PGC1α mediated mitochondrial quality control, ATP production and NAD⁺ de novo biosynthesis. The phosphorylation of NAD⁺ to NADP⁺ enhances the antioxidant defense against oxidant stress. NAD⁺-dependent defect in FAO results in intracellular lipid accumulation. In addition, the defected FAO and increased desaturation of PUFAs to HUFAs due to NAD⁺ deficiency and impaired mitochondrial function result in the accumulation of HUFA-containing triglycerides and cellular lipid in renal tubular cells. d The insulin secretion is adjusted by the dynamic glucose concentration in blood. As a master regulator of insulin secretion, glucose is metabolized via the glycolysis and TCA cycle to produce NADH and ATP. The increased NADH and ATP induces the closure of ATP-dependent K⁺ channels, the opening of voltage-gated L-type Ca²⁺ channels, the raising of cytosolic Ca²⁺ and culminating in insulin secretion in pancreatic β-cells. The activity of mitochondrial shuttles including the glycerophosphate and malate/aspartate shuttles allows the reoxidation of cytosolic NADH into NAD⁺, which is required for maintenance of the glycolysis. Purple representants the downregulated proteins or activated biological functions, while brown labels the upregulated proteins and repressed physiological activities. Abbreviations: ACMSD, alpha-amino-beta-carboxy-muconate-semialdehyde decarboxylase; AR, Aldose reductase; ETC, electron transport chain; Grxs, glutaredoxins; HUFAs, highly unsaturated fatty acids; KMO, kynurenine 3-monooxygenase; FAO, fatty acid oxidation; PUFAs, polyunsaturated fatty acids; SDH, Sorbitol dehydrogenase; Trxs, thioredoxins. 3-HK, 3-hydroxykynurenine

Fig. 8

Linkages between NAD⁺ depletion and neurodegenerative disorders. Most neurodegenerative disorders, including axonal degeneration, Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (PD) and Amyotrophic lateral sclerosis (ALS), are associated with mitochondrial dysfunction, lowered antioxidant capacity and heightened mitophagy, all of which are converged into the age-related NAD⁺ depletion induced by either enhanced consumption or impaired biosynthesis. These neural pathologies can be rescued by NAD⁺ boosting. Purple representants the downregulated proteins or activated biological functions, while brown labels the upregulated proteins and repressed physiological activities in neurodegenerative disorders. Abbreviations: mHtt, mutant Huntingtin; Aβ, amyloid beta; NFTs, neurofibrillary tangles; 3-HAA, 3-hydroxyanthranilic acid; QA, quinolinic acid; WldS, slow Wallerian degeneration; 3-HK, 3-hydroxykynurenine