NAD (nicotinamide adenine dinucleotide), a widely distributed and crucial substance within cells, is molecularly composed of nicotinamide, adenine, and two nucleotides linked by phosphate groups. This unique structure endows NAD with extraordinary biological functions, enabling it to play a vital dual role in cellular life activities—both as an indispensable coenzyme and an important signaling molecule.
Regarding its coenzyme function, NAD exists in two forms: oxidized (NAD⁺) and reduced (NADH), and the two can be reversibly converted. This dynamic balance has a profound impact on cellular metabolism. In key processes of cellular respiration, such as glycolysis, the tricarboxylic acid cycle, and oxidative phosphorylation, NAD⁺, as a redox coenzyme, accepts electrons and hydrogen ions released from metabolites, thus being reduced to NADH. This process resembles a carefully choreographed relay race, with NADH carrying electrons and hydrogen ions to ultimately transfer electrons to the electron transport chain, releasing a large amount of energy in the process and driving ATP synthesis. ATP, as the cell’s “energy currency,” provides the essential energy for various cellular physiological activities. NAD plays a crucial role in this process, serving as an indispensable link in energy metabolism. Studies have shown that under normal physiological conditions, the intracellular NAD⁺/NADH ratio remains within a relatively stable range, and this stability is vital for normal cellular metabolism. An imbalance in this ratio can trigger a series of metabolic disorders, like dominoes, thereby affecting normal cellular function.
NAD is also an essential substrate for many key enzymes, such as the longevity protein Sirtuins and the DNA repair enzyme PARP. Taking the Sirtuins family as an example, they participate in many important intracellular physiological processes, such as gene expression regulation, metabolic regulation, cellular senescence, and apoptosis. The participation of NAD⁺ is a prerequisite for Sirtuins to perform these functions; NAD⁺ acts like a key, activating the “lock” of Sirtuins and unlocking a series of physiological activities. In DNA repair, PARP enzymes use NAD⁺ as a substrate to catalyze the ADP-ribosylation reaction. This reaction acts like a “warning line” around damaged DNA, recruiting other repair proteins to rapidly gather at the damage site and promptly initiating the DNA repair mechanism to ensure genome stability. If NAD⁺ levels are insufficient, the activity of these NAD⁺-dependent enzymes is inhibited, leading to the inability of related physiological processes to proceed normally, thus affecting normal cell function and survival.
The Molecular Engine of DNA Repair
Throughout the life cycle of a cell, DNA constantly faces threats from both endogenous and exogenous factors. Endogenous factors include reactive oxygen species (ROS) produced during cellular metabolism. These highly oxidizing substances are inevitably produced during normal metabolic activities within the cell; they are like “time bombs” within the cell, potentially causing damage to DNA at any time. Exogenous factors are more diverse. High-energy photons in ultraviolet light can directly break the chemical bonds of DNA, leading to changes in DNA structure. Various chemical substances, such as environmental pollutants and carcinogens, can also react chemically with DNA, causing DNA damage. Furthermore, physical factors such as ionizing radiation can also cause severe damage to DNA, leading to various forms of damage, including single-strand or double-strand breaks.
When DNA is damaged, a sophisticated repair mechanism within the cell is activated, with NAD⁺ playing a central role, acting as the “molecular engine” of DNA repair. The poly(ADP-ribose) polymerase (PARP) family plays a crucial role in DNA repair, catalyzing reactions using NAD⁺ as a substrate. Specifically, when single-strand or double-strand breaks occur in DNA, PARP enzymes can quickly identify the damage site, acting like well-trained “scouts” to detect the problem immediately. Then, PARP enzymes use NAD⁺ as a substrate to transfer ADP ribosyl groups to themselves and other repair proteins, a process called ADP ribosylation. This modification process is like attaching “action tags” to the repair proteins, enabling them to cooperate more effectively to complete DNA repair. In this way, PARP enzymes recruit a series of key proteins involved in base excision repair (BER) and nucleotide excision repair (NER) to the damage site. In BER repair, specific enzymes recognize and remove damaged bases, and then, with the help of DNA polymerase and other enzymes, repair is carried out using new bases to rebuild a complete DNA strand. In nucleotide excision repair, a larger area of DNA damage is recognized and removed, and repair is subsequently completed through steps such as DNA synthesis and ligation. All these repair processes rely on the support of NAD⁺, whose presence provides the necessary energy and material basis for the repair process.
If NAD⁺ levels are insufficient, PARP enzyme activity will be significantly inhibited, leading to a decline in DNA repair capacity. This is analogous to a car running out of fuel and unable to drive properly. If DNA damage is not repaired in time, it will accumulate, leading to serious consequences such as gene mutations and chromosomal abnormalities. These alterations in genetic material may trigger cellular carcinogenesis, as gene mutations can cause uncontrolled cell growth and division, gradually transforming cells into cancer cells. Furthermore, the accumulation of DNA damage accelerates the aging process, causing cells to lose their normal function and eventually die. Therefore, maintaining adequate NAD⁺ levels is crucial for ensuring the normal functioning of DNA repair mechanisms, maintaining genome stability, and preventing disease.
The Guardians of Mitochondrial Function
Mitochondria, organelles hailed as the cell’s “energy factory,” are the primary site of aerobic respiration and play a central role in cellular energy metabolism. Through a series of complex biochemical reactions, they convert the chemical energy of nutrients into ATP, a form directly usable by the cell, providing a continuous energy source for various cellular life activities. Mitochondria act like a “power station” within the cell, ensuring a constant and sufficient energy supply for normal physiological functions.
With cellular aging and the influence of various internal and external environmental factors, mitochondrial function gradually declines, a significant marker of cellular aging. The manifestations of mitochondrial functional decline are diverse, with damage to the mitochondrial membrane structure being a key aspect. The mitochondrial membrane is the essential structural basis for energy conversion and substance transport within mitochondria. When the mitochondrial membrane is damaged, the normal functioning of the electron transport chain is affected. The electron transport chain is a crucial link in ATP production in mitochondria. It consists of a series of protein complexes that transfer electrons from donors such as NADH and FADH₂ to oxygen, while simultaneously using the energy released during electron transport to pump protons out of the inner mitochondrial membrane, forming a proton gradient. The proton gradient acts like an “energy reservoir,” storing a large amount of energy. When protons flow back to the mitochondrial matrix via ATP synthase, they drive ATP synthesis. However, when the mitochondrial membrane is damaged, the efficiency of the electron transport chain decreases significantly, making it difficult to maintain the proton gradient and leading to a sharp decline in ATP production. Furthermore, mitochondrial dysfunction also results in the excessive production of reactive oxygen species (ROS). Under normal circumstances, mitochondria produce small amounts of ROS during energy metabolism, which can participate in some physiological processes as signaling molecules. But when mitochondrial function is impaired, the electron transport chain malfunctions, electron leakage increases, leading to excessive ROS production. Excessive ROS is highly oxidizing and can cause severe oxidative damage to intracellular macromolecules such as DNA, proteins, and lipids, further exacerbating cellular aging and dysfunction.
NAD⁺ plays a crucial role in maintaining mitochondrial function, acting as its “guardian.” It primarily achieves this function by activating mitochondrial deacetylases such as SIRT3. SIRT3 is a deacetylase specifically expressed in mitochondria that recognizes and removes acetyl modifications from certain proteins in mitochondria. These modified proteins are involved in several important physiological processes in mitochondria, such as mitophagy and biosynthesis. When SIRT3 is activated by NAD⁺, it promotes mitophagy. Mitophagy is an important quality control mechanism in the cell, recognizing and clearing damaged mitochondria, acting like a “garbage collector” to remove waste mitochondria and maintain the health and function of the mitochondrial population. Through mitophagy, damaged mitochondria are encapsulated in autophagosomes, then fuse with lysosomes and are degraded by hydrolases in the lysosomes. This process not only clears damaged mitochondria, preventing further production of harmful ROS, but also provides the cell with reusable substances such as amino acids and fatty acids for the synthesis of new mitochondria.
NAD⁺ can also promote mitochondrial biosynthesis by activating SIRT3. Mitochondrial biosynthesis refers to the process by which cells produce new mitochondria, a process crucial for maintaining the balance of mitochondrial numbers and functions within the cell. Under the influence of NAD⁺ and SIRT3, a series of genes involved in mitochondrial biosynthesis are activated. These genes encode proteins involved in mitochondrial assembly, replication, and functional maintenance. By promoting mitochondrial biosynthesis, cells can replenish damaged or aging mitochondria in a timely manner, maintaining the vitality and function of the mitochondrial population. Studies have shown that increasing intracellular NAD⁺ levels and activating deacetylases such as SIRT3 can increase mitochondrial ATP production capacity by more than 40%, significantly improving mitochondrial function. This discovery provides new insights and potential therapeutic targets for treating diseases related to mitochondrial dysfunction, such as neurodegenerative diseases and cardiovascular diseases. Supplementing NAD⁺ or activating its related signaling pathways holds promise for improving mitochondrial function, delaying cellular aging, and enhancing overall health.
NAD’s Four Core Functions in Regulating Cell Repair
NAD’s Four Core Functions in Regulating Cell Repair(I) Combating Oxidative Stress: A Synergistic Network for Scavenging Free Radicals
Free radicals are inevitably generated during normal cellular metabolism. These free radicals act like “troublemakers” within the cell, constantly threatening cellular health. Among them, reactive oxygen species (ROS) are a class of highly oxidizing free radicals, mainly including hydroxyl radicals (・OH), hydrogen peroxide (H₂O₂), and superoxide anions (O₂⁻). When cells are stimulated by external environmental factors, such as ultraviolet radiation, chemical pollution, ionizing radiation, or when there is an imbalance in intracellular metabolism, the production of ROS increases significantly, leading to oxidative stress. Oxidative stress can cause severe damage to intracellular biomolecules, such as lipids, proteins, and DNA. In terms of lipids, ROS can trigger lipid peroxidation, leading to damage to the structure and function of the cell membrane. The cell membrane is a barrier between the cell and the external environment, and its integrity is crucial for the normal physiological function of the cell. When lipid peroxidation occurs in the cell membrane, membrane fluidity and permeability change, affecting the exchange of substances and signal transduction between the intracellular and extracellular spaces. Regarding proteins, reactive oxygen species (ROS) oxidize amino acid residues, leading to alterations in protein structure and function. Proteins are the main molecules performing various physiological functions within the cell; once their structure and function are damaged, normal cellular physiological activities are affected. Regarding DNA, ROS can cause DNA strand breaks, base modifications, and gene mutations, seriously threatening genome stability.
NAD⁺ plays a crucial role in combating oxidative stress. It enhances the cell’s antioxidant defense system by regulating the Nrf2 signaling pathway and activating the expression of a series of antioxidant enzymes. Nrf2 is a transcription factor that plays a central regulatory role in the cell’s antioxidant stress response. Under normal conditions, Nrf2 binds to the Keap1 protein and is inactive. When cells are stimulated by oxidative stress, ROS oxidize cysteine residues in the Keap1 protein, causing Nrf2 to dissociate from Keap1. After dissociation, Nrf2 enters the cell nucleus and binds to antioxidant response elements (AREs), initiating the transcription and expression of a series of antioxidant enzyme genes, such as glutathione synthase and heme oxygenase-1 (HO-1). Glutathione synthase catalyzes the synthesis of glutathione (GSH), an important intracellular antioxidant that can directly scavenge reactive oxygen species (ROS) or reduce H₂O₂ to water by participating in the glutathione peroxidase (GPx) reaction, thereby mitigating oxidative stress-induced cell damage. Heme oxygenase-1 (HO-1) catalyzes the breakdown of heme into carbon monoxide (CO), biliverdin, and iron ions. Biliverdin can be further reduced to bilirubin, and these products all possess antioxidant properties. CO can reduce oxidative stress-induced cell damage by regulating intracellular signaling pathways, inhibiting inflammatory responses and apoptosis. Bilirubin is a potent antioxidant that can scavenge ROS and protect cells from oxidative damage.
Maintaining a high NAD⁺/NADH ratio is also an important cellular strategy against oxidative stress. A dynamic balance exists between NAD⁺ and NADH within the cell, and this balance significantly impacts cellular metabolism and antioxidant capacity. A decreased NAD⁺/NADH ratio indicates an enhanced reducing state within the cell, leading to increased ROS production and exacerbating oxidative stress. Conversely, maintaining a high NAD⁺/NADH ratio promotes the pentose phosphate pathway (PPP). PPP is a crucial intracellular metabolic pathway that produces NADPH, an important reducing equivalent that provides electrons for intracellular antioxidant responses and participates in maintaining redox balance. In PPP, glucose-6-phosphate is catalyzed by glucose-6-phosphate dehydrogenase (G6PD) to produce 6-phosphogluconolactone, simultaneously generating NADPH. NADPH can reduce oxidized glutathione (GSSG) to reduced glutathione (GSH), thereby maintaining intracellular GSH levels and enhancing cellular antioxidant capacity. NADPH can also directly participate in the scavenging of reactive oxygen species (ROS), such as reducing H₂O₂ to water under the action of glutathione peroxidase (GPx), or decomposing H₂O₂ into water and oxygen under the action of catalase (CAT).
A large body of research data has fully demonstrated the significant effects of NAD⁺ in combating oxidative stress. In an experiment on aging mice, supplementation with NAD⁺ precursors successfully increased NAD⁺ levels in the mice. The results showed a significant reduction in intracellular ROS levels, a marked decrease in lipid peroxidation, and effective inhibition of oxidative damage to proteins and DNA. Compared with the control group, the liver tissue of mice supplemented with NAD⁺ showed a 30% increase in GSH content and a 50% upregulation of HO-1 expression, indicating that NAD⁺ enhances cellular antioxidant defense by activating the expression of antioxidant enzymes. In another in vitro study on human cells, NAD⁺ supplementation restored the intracellular NAD⁺/NADH ratio to normal after oxidative stress stimulation, thereby promoting PPP and increasing NADPH production. By detecting intracellular ROS levels and antioxidant enzyme activities, it was found that in NAD⁺-supplemented cells, ROS levels decreased by 40%, and the activities of G6PD and GPx increased by 25% and 35%, respectively, further confirming the crucial role of NAD⁺ in combating oxidative stress.
(II) Delaying Cellular Senescence: Dual Regulation by Telomeres and Epigenetics
Cellular senescence is a complex process precisely regulated by multiple factors. It resembles the “old age” stage in a cell’s life cycle, accompanied by a series of significant physiological changes. Morphologically, senescent cells gradually enlarge and flatten, losing their original normal shape and structure. Internal organelles, such as mitochondria and endoplasmic reticulum, also show signs of functional decline. As the cell’s “energy factory,” mitochondria experience a decrease in membrane potential and ATP production capacity in senescent cells, leading to insufficient cellular energy supply. The endoplasmic reticulum exhibits stress responses, affecting protein synthesis, folding, and transport. At the molecular level, the gene expression patterns of senescent cells undergo significant changes. Some genes related to cell proliferation and metabolism are downregulated, while genes related to senescence are upregulated. Senescent cells also secrete a series of cytokines, chemokines, and proteases, forming a senescence-associated secretory phenotype (SASP). These substances affect surrounding cells and tissues, triggering chronic inflammatory responses and further accelerating the senescence process of cells and tissues.
Telomeres, special structures at the ends of chromosomes, act like “protective caps” for the chromosomes, playing a crucial role in maintaining their stability and integrity. They consist of a repetitive DNA sequence and related proteins. During cell division, telomeres gradually shorten as chromosomes replicate. This is because DNA polymerase cannot completely replicate the sequences at the ends of chromosomes during DNA replication, resulting in the loss of a small segment of telomere with each cell division. When telomeres shorten to a certain extent, the cell enters a senescent state and stops dividing. This is analogous to the plastic caps at the ends of shoelaces; if they gradually wear down and shorten, the shoelaces become prone to coming undone. Similarly, chromosomes become unstable due to excessively short telomeres, leading to cellular senescence. Telomerase is an enzyme that can lengthen telomeres. It consists of RNA and protein, with the RNA portion serving as a template to guide the synthesis of telomere DNA sequences. In normal somatic cells, telomerase activity is strictly regulated and usually at a low level, causing telomeres to continuously shorten with cell division. However, in some stem cells and cancer cells, telomerase activity is high, maintaining telomere length and allowing the cell to continue dividing.
The NAD⁺-dependent SIRT1 protein plays a crucial role in delaying cellular senescence. It primarily regulates the stability of the telomerase-associated protein TERT through deacetylation, thereby indirectly affecting telomere length. Specifically, SIRT1 recognizes and binds to TERT, removing the acetyl group from its lysine residues. This deacetylation enhances TERT stability, inhibits its degradation rate by the proteasome, and thus increases telomerase activity. Studies have shown that under sufficient NAD⁺ conditions, SIRT1’s deacetylation of TERT is enhanced, enabling telomerase to more effectively lengthen telomeres and delay cellular senescence. Experimental data show that by increasing intracellular NAD⁺ levels and activating SIRT1, the rate of telomere shortening can be slowed by approximately 30%, meaning that cell lifespan is effectively extended and the senescence process is significantly slowed.
In addition to regulating telomeres, the Sirtuins family also delays cellular senescence through epigenetic regulatory mechanisms. Epigenetics refers to the regulation of gene expression without altering the DNA sequence, primarily including DNA methylation and histone modifications. During cellular senescence, epigenetic states undergo significant changes, affecting gene expression and consequently leading to cellular functional decline. Several members of the Sirtuins family, such as SIRT1, SIRT2, and SIRT6, can influence gene expression by regulating histone acetylation levels and reshaping chromatin structure. For example, SIRT1 can deacetylate specific lysine residues on histone H3 and H4, making chromatin structure more compact and inhibiting the expression of certain genes related to cellular senescence. Simultaneously, SIRT1 can activate genes related to cellular metabolism and oxidative stress, enhancing cellular function and resilience. Regarding DNA methylation, senescent cells often exhibit abnormal DNA methylation patterns, with increased methylation levels in the promoter regions of some genes, leading to gene silencing. The Sirtuins family can maintain the balance of DNA methylation and correct aging-related DNA methylation abnormalities by regulating the activity of DNA methyltransferases and demethylases, thereby delaying cellular senescence. Treatment of senescent cells with Sirtuins activators revealed a significant improvement in senescence-related phenotypes, a reduction in cell cycle arrest, and a significant decrease in SASP secretion. This fully demonstrates the important role of the Sirtuins family in epigenetic regulation of delaying cellular senescence.
(III) Precise Regulation of Apoptosis: The Molecular Switch for Life and Death Balance
Apoptosis, also known as programmed cell death, is a genetically controlled process of active cell death. Like a “countdown timer” for cellular life, it plays a crucial role in maintaining normal development and homeostasis. In the development of multicellular organisms, apoptosis participates in organ formation and shaping. For example, during embryonic development, the formation of fingers and toes involves the removal of excess cells between the fingers through apoptosis, thus shaping normal limb form. In the immune system, apoptosis eliminates cells infected by pathogens, cancerous cells, or those with abnormal function, ensuring the normal function of the immune system. Abnormalities in the apoptosis process can lead to a series of serious diseases. Excessive apoptosis can cause tissue and organ atrophy and functional decline. For example, massive neuronal apoptosis in neurodegenerative diseases can lead to cognitive impairment and motor incoordination. Conversely, insufficient apoptosis can lead to abnormal cell proliferation, increasing the risk of cancer. Cancer cells often evade the regulation of apoptosis, thus proliferating indefinitely and forming tumors.
In the regulatory network of apoptosis, NAD⁺ plays a central role, acting as a “molecular switch” for cellular life-and-death balance. DNA damage is one of the important signals that trigger apoptosis. When cells are stimulated by factors such as ultraviolet radiation, ionizing radiation, and chemicals, DNA damage occurs. At this time, NAD⁺ plays a crucial regulatory role in the choice between DNA repair and apoptosis. The poly(ADP-ribose) polymerase (PARP) family is an important participant in DNA damage repair. When single-strand or double-strand breaks occur in DNA, PARP enzymes can rapidly recognize the damage site and catalyze the reaction using NAD⁺ as a substrate. PARP enzymes transfer ADP ribose groups to themselves and other repair proteins, forming poly(ADP-ribose) chains; this process is called ADP ribosylation. Through ADP ribosylation modification, PARP enzymes can recruit other DNA repair proteins to the damage site, initiating DNA repair mechanisms such as base excision repair (BER) and nucleotide excision repair (NER). In this process, NAD⁺ provides the necessary substrate for PARP enzyme activity, ensuring the smooth progress of DNA repair. If the DNA damage is mild, with the support of NAD⁺, PARP enzymes can effectively repair DNA damage, restoring cells to a normal state and preventing apoptosis.
However, if the DNA damage is too severe, exceeding the cell’s repair capacity, excessive PARP enzymes will be continuously activated, consuming large amounts of intracellular NAD⁺. When NAD⁺ levels drop sharply, a series of apoptosis signaling pathways are triggered, among which the mitochondrial apoptosis pathway is one of the important pathways of apoptosis. Under normal circumstances, the outer membrane of mitochondria contains some anti-apoptotic proteins, such as Bcl-2 and Bcl-xL from the Bcl-2 family, which can maintain the stability of the mitochondrial membrane and inhibit apoptosis. When NAD⁺ is depleted, it leads to a decrease in mitochondrial membrane potential and an increase in permeability, prompting mitochondria to release apoptosis factors such as cytochrome C into the cytoplasm. Cytochrome C binds to Apaf-1 (Apaf-1) to form an apoptotic body, recruiting and activating caspase-9 from the caspase family, which in turn activates the downstream caspase cascade, ultimately leading to apoptosis. NAD⁺ also regulates apoptosis by modulating the balance of the Bcl-2/Bax protein family. Bax is a pro-apoptotic protein; when cells are stimulated by apoptotic signals, Bax translocates from the cytoplasm to the mitochondrial membrane, forming pores that increase mitochondrial membrane permeability and release apoptotic factors. Bcl-2, on the other hand, is an anti-apoptotic protein that interacts with Bax, inhibiting its pro-apoptotic effects. NAD⁺ can maintain this balance by activating Sirtuins family proteins, regulating the expression and activity of Bcl-2 and Bax. Under conditions of sufficient NAD⁺, Sirtuins can promote Bcl-2 expression and inhibit Bax activity, thereby suppressing apoptosis. Conversely, when NAD⁺ levels decrease, Bax activity increases, Bcl-2 expression is inhibited, and the tendency for apoptosis increases. This finely regulated mechanism ensures that cells can make appropriate life-or-death decisions when faced with varying degrees of DNA damage, maintaining the health and stability of the cell population.
(IV) Metabolic Reprogramming: Repairing the Energy Supply Network
Cellular metabolism, as the foundation of cellular life activities, is a complex and orderly network of chemical reactions. It involves multiple aspects such as the synthesis and decomposition of intracellular substances and the production and utilization of energy, acting like a “chemical factory” inside the cell, continuously providing material and energy support for various physiological activities. Among the many pathways of cellular metabolism, glycolysis, the tricarboxylic acid cycle, and oxidative phosphorylation are the core links in energy metabolism. Glycolysis is the process of breaking down glucose into pyruvate. It takes place in the cytoplasm and, although it produces relatively little energy, it is fast enough to provide a certain amount of energy to the cell in a short time. During strenuous exercise, muscle cells need to quickly obtain energy, so glycolysis accelerates, producing lactic acid and causing muscle soreness. The tricarboxylic acid cycle, under aerobic conditions, further oxidizes and decomposes pyruvate, producing large amounts of energy carriers NADH and FADH₂, while releasing carbon dioxide. Oxidative phosphorylation utilizes NADH and FADH₂ to transfer electrons in the electron transport chain, generating a proton gradient that drives ATP synthesis. This is the primary way cells produce energy, providing a large amount of ATP. NAD⁺, as a key coenzyme in cellular metabolism, plays an indispensable role in these core metabolic pathways. During glycolysis, NAD⁺ accepts hydrogen and electrons from the oxidation of glyceraldehyde-3-phosphate, being reduced to NADH, while simultaneously promoting the production of 1,3-bisphosphoglycerate, thus facilitating glycolysis. In the tricarboxylic acid cycle, NAD⁺ participates in multiple reactions, accepting hydrogen and electrons from the oxidation of metabolites such as isocitrate, α-ketoglutarate, and malic acid, being reduced to NADH. The electrons carried by these NADH molecules enter the electron transport chain, participating in oxidative phosphorylation and generating a large amount of ATP. In short, NAD⁺ acts as an “energy carrier” in cellular energy metabolism, ensuring the efficient operation of energy metabolic pathways and providing cells with sufficient energy.
As we age and are affected by various diseases, cellular metabolic functions gradually become disordered. This is analogous to a chemical plant that has been running for many years; as equipment ages, production efficiency declines, and various malfunctions occur. In patients with metabolic syndrome, insulin resistance is common, leading to decreased cellular sensitivity to insulin, impaired glucose uptake and utilization, and elevated blood sugar levels. Simultaneously, lipid metabolism also becomes abnormal, with elevated levels of triglycerides and low-density lipoprotein cholesterol (LDL-C). Fat accumulates in tissues such as the liver and muscles, causing fatty liver disease, atherosclerosis, and other conditions. These metabolic disorders not only affect normal cellular function but also further burden the body, triggering a series of chronic diseases.
NAD⁺ supplementation has been proven to be an effective strategy for improving cellular metabolic disorders and repairing energy supply networks. Studies have found that supplementing with NAD⁺ precursors, such as nicotinamide mononucleotide (NMN) and nicotinamide nucleoside (NR), can increase intracellular NAD⁺ levels, thereby activating a series of enzymes and signaling pathways related to metabolic regulation. NAD⁺ can activate the SIRT1 protein, which deacetylates key transcription factors regulating metabolism, such as peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α), enhancing mitochondrial biosynthesis and function and improving cellular energy metabolism efficiency. NAD⁺ supplementation can also regulate the insulin signaling pathway, enhancing insulin sensitivity and promoting glucose uptake and utilization. In a clinical study of patients with metabolic syndrome, NMN supplementation significantly improved insulin resistance, increased insulin sensitivity by approximately 25%, and effectively controlled blood glucose levels. Triglyceride and LDL-C levels also decreased by 15% and 10%, respectively. This indicates that NAD⁺ supplementation can effectively improve metabolic disorders in patients with metabolic syndrome, restore cellular energy homeostasis, and provide new ideas and methods for treating metabolic diseases.
NAD Supplementation Strategies: From Natural Sources to Precision Intervention
(I) Efficient Conversion Pathways of Precursor Substances
Since NAD⁺ cannot be directly absorbed orally, the key to NAD⁺ supplementation lies in ingesting its precursor substances, which are then metabolized into NAD⁺ in the body, thereby increasing NAD⁺ levels. Among many precursor substances, NMN (nicotinamide mononucleotide) stands out due to its unique advantages. NMN has a relatively small molecular weight of only 334 Da, which gives it excellent cell penetration capabilities. It can easily bypass the NAMPT rate-limiting enzyme and, with the help of the Slc12a8 transporter, directly enter the cell through what seems like a “fast track.” Once inside the cell, NMN is rapidly converted into NAD⁺. This conversion process is highly efficient and direct, significantly increasing the efficiency of NMN to NAD⁺ conversion compared to other precursor substances, reaching up to 60%. Numerous clinical studies have also provided solid data support for the high efficiency of NMN. In a clinical trial involving healthy adults, participants took 250mg of NMN orally daily for four weeks. After this continuous supplementation, their blood NAD⁺ levels significantly increased, rising by as much as 50%. This result vividly demonstrates the remarkable effect of NMN in increasing NAD⁺ levels and lays a solid foundation for its application in the health field.
NR (nicotinamide riboside), another important NAD⁺ precursor, also has a unique metabolic pathway and advantages. NR mainly enters cells via the ENT1 transporter, which acts like a “dedicated entrance” for the cell, precisely recognizing and guiding NR into the cell. Once inside the cell, NR undergoes phosphorylation under the catalysis of the NRK enzyme, converting into NMN. NMN then undergoes further metabolic processes, ultimately generating NAD⁺. Although the conversion pathway of NR has an additional step compared to NMN, it has unique advantages in certain aspects. For people with sensitive digestive systems, NR may be a better choice. Due to its unique chemical structure and metabolic pathways, NR is relatively more stable in the gastrointestinal tract, reducing irritation and the risk of gastrointestinal discomfort. Studies have also found that combining NR with NMN produces a synergistic effect. This combination can increase sirtuin activity by 35%. As proteins closely related to cellular aging and metabolic regulation, increased sirtuins activity signifies further optimization of cellular metabolic function and a more effective slowing of the aging process, bringing more comprehensive benefits to human health.
Besides synthetic precursor supplements, some natural foods also contain trace amounts of NMN, providing a natural way to supplement NAD⁺. Avocados, a fruit rich in various nutrients, contain approximately 1.8 mg of NMN per 100g; edamame, a common legume, contains approximately 1.5 mg of NMN per 100g; and broccoli, a nutrient-rich vegetable, contains approximately 0.8 mg of NMN per 100g. While these natural foods contain NMN, the content is relatively low. To achieve an effective dose to increase NAD⁺ levels through diet, assuming a daily effective dose of 200-300mg of NMN, one would need to consume 1-2kg of foods like avocados, edamame, or broccoli daily, which is practically impossible to achieve. Furthermore, during cooking, NMN in these foods may degrade due to high temperatures, further reducing its effective content. Therefore, although natural foods provide a way to supplement NMN, relying solely on food to increase NAD⁺ levels has limited practical value. More efficient methods, such as prodrug supplements, are needed to meet the body’s NAD⁺ requirements.
(II) Individualized Supplementation Plans
In the process of NAD⁺ supplementation, individualized design is crucial. Different individuals have different physical conditions and needs, therefore, targeted supplementation plans are necessary. For those focused on preventative health, their bodies are relatively healthy, but with age and various daily stresses, they may gradually show signs of aging, such as increased fatigue and decreased sleep quality. For this group, the recommended NAD⁺ supplementation dose is 200-300 mg/day. Taking it after breakfast is recommended because the body’s metabolism gradually becomes more active after breakfast, allowing for better absorption and utilization of NAD⁺. The effectiveness of supplementation can be monitored by changes in fatigue and improvements in sleep quality. If, after a period of supplementation, fatigue is significantly reduced and sleep quality improves (e.g., falling asleep faster, sleeping more soundly, and waking up less frequently at night), the supplementation plan may be effective.
For individuals with metabolic disorders, such as those suffering from metabolic syndrome or insulin resistance, their metabolic functions are disordered, resulting in abnormal blood sugar and lipid levels and decreased insulin sensitivity. This group requires higher doses of NAD⁺ supplementation, with a recommended dose of 400-600 mg/day. To maximize the effectiveness of NAD⁺, the daily dose should be divided into two doses taken with meals. Taking it with meals reduces gastrointestinal irritation, and the presence of food helps promote NAD⁺ absorption. Regarding monitoring indicators, close attention should be paid to changes in blood glucose and blood lipids. Fasting blood glucose, postprandial blood glucose, and four blood lipid parameters (total cholesterol, triglycerides, LDL cholesterol, and HDL cholesterol) can be measured regularly to assess the effectiveness of NAD⁺ supplementation. HOMA-IR (Homologous Body Mass Index for Insulin Resistance) can also be measured, as this index more accurately reflects the degree of insulin resistance. If, after NAD⁺ supplementation, blood glucose and blood lipid levels gradually normalize, and the HOMA-IR index decreases, it indicates that NAD⁺ supplementation has a positive effect on improving metabolic abnormalities.
For individuals with age-related diseases, such as neurodegenerative diseases and cardiovascular diseases, their bodies experience accelerated cellular damage and aging due to the effects of these diseases, often resulting in lower NAD⁺ levels. These individuals require NAD⁺ supplementation under the strict guidance of a physician, with a recommended dosage of 800-1000 mg/day. Because their conditions are complex and their physical circumstances vary, supplementation must be carried out under the supervision of a physician to ensure safety and effectiveness. The physician will develop a personalized supplementation plan based on the patient’s specific condition, physical indicators, and medication regimen. In terms of monitoring indicators, in addition to focusing on routine physiological indicators, it is crucial to monitor the levels of NAD⁺ metabolites (such as NADH). As a reduced form of NAD⁺, changes in NADH levels reflect the metabolism of NAD⁺ in the body. By monitoring the levels of NADH and other metabolites, physicians can promptly understand the effectiveness of NAD⁺ supplementation and adjust the dosage and regimen to achieve the best therapeutic effect.
(III) Safety and Potential Risks
When using NAD⁺ supplements, safety and potential risks must be fully considered, especially for certain populations. Patients with autoimmune diseases should avoid NAD⁺ supplements because NAD⁺ participates in multiple cell signaling pathways in the body and may activate inflammatory pathways. For patients with autoimmune diseases, their immune systems are already abnormally active, with excessive inflammatory responses. Supplementing with NAD⁺ may further exacerbate the inflammatory response, leading to a worsening of the condition. Patients with rheumatoid arthritis, whose joints are already eroded by inflammation, may experience more severe joint inflammation, increased pain and swelling, affecting their quality of life and joint function. Pregnant women should also avoid NAD⁺ supplements. Although current research on the effects of NAD⁺ on fetal development is insufficient, to ensure the healthy development of the fetus, pregnant women should not risk using it. The fetus is in a rapid growth and development stage in the womb and is very sensitive to external factors; any uncertain factors may pose potential risks to fetal development. Patients taking metformin should exercise special caution when using NAD⁺ supplements, and it is recommended to take them at least 4 hours apart. Metformin is a commonly used medication for treating type 2 diabetes, and it may interact with NAD⁺, affecting the efficacy of the medication or increasing the risk of adverse reactions. Taking them at least 4 hours apart can reduce the possibility of such interactions to some extent, ensuring the safety and effectiveness of the medication.
NAD⁺ supplements may also interact with other medications, which requires attention. When NAD⁺ supplements are used in combination with aspirin, they may enhance the inhibitory effect of PARP. Aspirin is a commonly used antiplatelet drug, while PARP plays an important role in DNA repair. Enhanced PARP inhibition may affect blood clotting function and increase the risk of bleeding. Therefore, when using NAD⁺ supplements and aspirin simultaneously, it is recommended to closely monitor coagulation function, such as regularly checking platelet counts and prothrombin time, to promptly detect and manage any potential coagulation abnormalities. When NAD⁺ supplements are taken concurrently with statins, liver enzyme (ALT/AST) levels should be monitored. Statins are commonly used lipid-lowering drugs, primarily lowering blood lipid levels by inhibiting cholesterol synthesis in the liver. However, NAD⁺ supplements may affect the metabolism of statins in the liver, leading to elevated liver enzyme levels, indicating potential liver damage. Therefore, when using these two medications simultaneously, liver enzyme levels should be monitored regularly. If elevated liver enzymes are detected, the medication dosage should be adjusted promptly or appropriate treatment measures should be taken to protect liver function.
Cutting-Edge Research and Clinical Translation Breakthroughs
(I) New Targets for Intervention in Age-Related Diseases
In the field of research on age-related diseases, NAD⁺ has shown great potential, becoming a new target for intervention in these diseases.
Neurodegenerative diseases, such as Alzheimer’s disease and Parkinson’s disease, seriously threaten the health and quality of life of the elderly. Taking Alzheimer’s disease as an example, an experiment conducted by Harvard Medical School achieved remarkable results. In Alzheimer’s disease model mice, supplementation with NAD⁺ precursors significantly reduced β-amyloid protein deposition in the mouse brain by 40%. Abnormal deposition of β-amyloid protein is one of the important pathological features of Alzheimer’s disease; it forms plaques in the brain, destroys the connections between neurons, and leads to cognitive dysfunction. Supplementation with NAD⁺ precursors effectively reduced this deposition, bringing new hope for the treatment of Alzheimer’s disease. Supplementation with NAD⁺ precursors also increased synaptic density in the mouse brain by 25%. Synapses are key structures for information transmission between neurons. Increased synaptic density indicates improved communication between neurons, contributing to the restoration of normal brain function. Further mechanistic studies have revealed that this process is closely related to SIRT1-mediated dephosphorylation of Tau protein. Hyperphosphorylation of Tau protein leads to the formation of neurofibrillary tangles, a pathological feature of Alzheimer’s disease. NAD⁺ promotes Tau protein dephosphorylation by activating SIRT1, thereby reducing the formation of neurofibrillary tangles, protecting neurons from damage, and improving cognitive function.
Cardiovascular disease is one of the major health threats worldwide, and NAD⁺ also plays an important role in cardiovascular protection. A clinical study in Japan confirmed this, in which patients with coronary heart disease supplemented with NMN at a dose of 500 mg/day for six consecutive months. The results showed a significant improvement in vascular endothelial function, with the FMD value, reflecting vascular endothelial function, increasing by 18%. Vascular endothelial cells are a layer of cells lining the inner wall of blood vessels, crucial for maintaining normal vascular function. When vascular endothelial function is impaired, it leads to abnormal vasoconstriction and vasodilation, increasing the risk of cardiovascular disease. NMN supplementation improves vascular endothelial function, meaning improved vascular health and better maintenance of normal blood circulation. Patients experienced a 22% decrease in serum inflammatory factors such as IL-6. Inflammation plays a crucial role in the development of cardiovascular disease; elevated inflammatory factors exacerbate inflammation in the blood vessel walls, promoting the formation and development of atherosclerosis. NMN supplementation reduces inflammatory factor levels, alleviating inflammation and thus lowering the risk of plaque rupture and cardiovascular events.
With age, women face numerous challenges to their reproductive health, particularly fertility issues in older women. Clinical trials in older women have shown significant effects from NAD⁺ supplementation. After NAD⁺ supplementation, the mitochondrial membrane potential of oocytes increased by 30%. Mitochondria are the cell’s “energy factories,” crucial for oocyte development and function. Increased mitochondrial membrane potential indicates improved mitochondrial function, providing more energy to oocytes and contributing to better oocyte quality. The chromosomal abnormality rate decreased by 28%. Chromosomal abnormalities are a major cause of infertility and miscarriage in older women. NAD⁺ supplementation effectively reduced the chromosomal abnormality rate and improved embryo quality. The blastocyst formation rate increased from 35% to 52%. The blastocyst is an important stage in embryonic development, and the improved blastocyst formation rate significantly improved the success rate of assisted reproduction, bringing hope to older women who wish to have children.
(II) Technological Innovation and Product Iteration
To further improve the efficacy and safety of NAD⁺ supplementation, researchers have made continuous efforts in technological innovation and product iteration, achieving a series of significant advancements.
In targeted delivery systems, the application of liposome encapsulation technology has brought a revolutionary breakthrough to NMN absorption. Traditional NMN is easily degraded by stomach acid after entering the body, resulting in a low absorption rate in the small intestine, only about 30%. Liposome-encapsulated NMN effectively resists stomach acid degradation, acting like a “protective layer” that allows it to reach the small intestine smoothly and be absorbed. This technology significantly increases the absorption rate of NMN in the small intestine to 85%, greatly improving its bioavailability. Lipo-NMN, developed by MIT, is a representative product of this technology. It not only improves the absorption rate but also shortens the time to peak blood concentration to 30 minutes. This means that Lipo-NMN can be absorbed and exert its effects more quickly, bringing users more immediate health benefits.
Synergistic effects in compound formulations are also a key direction in current NAD⁺ product development. By adding other synergistic ingredients, the efficacy of NAD⁺ can be further enhanced while reducing the incidence of side effects. Coenzyme Q10 is an important component of the mitochondrial respiratory chain, improving its efficiency and enhancing cellular energy metabolism. Combining coenzyme Q10 with NAD⁺ can make cellular energy production more efficient and further improve mitochondrial function. Alpha-lipoic acid is a powerful antioxidant that enhances antioxidant synergy, working with NAD⁺ to scavenge free radicals within cells and reduce oxidative stress damage. Verification by a Science sub-journal showed that compound formulations containing coenzyme Q10 and alpha-lipoic acid can increase NAD⁺ efficacy by 40% while reducing the incidence of side effects by 60%. This compound formulation of NAD⁺ products provides users with more comprehensive and efficient health protection, better meeting the needs of different populations.
(III) Controversies and Future Directions
While NAD⁺ has demonstrated significant potential in cell repair and intervention for age-related diseases, current research remains controversial, pointing the way for future investigations.
Current research focuses on the long-term safety of NAD⁺ supplementation, with renal burden and carcinogenic risk being key concerns. After NAD⁺ supplements enter the body, they undergo a series of metabolic processes, which may place a burden on organs such as the kidneys. Due to individual differences, tolerance and metabolic capacity to NAD⁺ supplements vary among individuals; therefore, whether long-term NAD⁺ supplementation will adversely affect kidney function requires further research and observation. Regarding the carcinogenic risk of NAD⁺ supplementation, although there is currently no conclusive evidence that it directly causes cancer, some studies suggest that NAD⁺-involved cellular metabolism and signaling pathways may be related to cancer development. Therefore, more long-term, large-scale clinical studies are needed to evaluate the safety of NAD⁺ supplementation and provide a solid theoretical basis for its widespread clinical application.
The differences in NAD⁺ metabolism across different tissues and organs are also an important direction for future research. A recent study published in *Nature Metabolism* in 2024 revealed significant differences in the responses of the liver and muscle tissue to NAD⁺ precursors. The liver, a vital metabolic organ, plays a crucial role in the synthesis, metabolism, and regulation of NAD⁺. Muscle tissue, primarily responsible for movement and energy expenditure, has different NAD⁺ requirements and metabolic pathways compared to the liver. This tissue-specific difference suggests the need to develop tissue-specific delivery technologies for precise anti-aging. Precisely delivering NAD⁺ precursors to specific tissues and organs can not only improve treatment efficacy but also reduce unnecessary impacts on other tissues and lower the risk of side effects. This will provide more precise and effective strategies for treating age-related diseases, further advancing NAD⁺-related research and applications.
From Molecular Mechanisms to Precision Health Management
NAD⁺, as a core hub for cell repair, has value far exceeding that of a single “anti-aging ingredient.” It constructs a cellular-level protective network through multiple pathways, including energy metabolism, DNA repair, and oxidative stress. Scientific NAD⁺ supplementation needs to be combined with individual genotype (such as NAMPT gene polymorphism), metabolic indicators, and health goals, forming a synergistic effect with dietary control (low-GI diet) and regular exercise (150 minutes of resistance training per week). With advancements in delivery technology and precision medicine, NAD⁺ may become an important biological target for delaying aging and preventing chronic diseases, ushering in a new era of “proactive anti-aging.”
