Course outline
Module 1
Module 1: What NAD+ Is and Why It Matters
What NAD+ Is and Why It Matters
Knowledge check
Q1. What does NAD+ stand for?
- A. Nicotinamide adenine dinucleotide ✓
- B. Nicotinic acid diphosphate
- C. Nucleic acid dehydrogenase
- D. Nicotinamide ascorbate dinucleotide
Why: NAD+ stands for nicotinamide adenine dinucleotide, a coenzyme found in every living cell.
Q2. What is the primary role of NAD+ in cells?
- A. Storing genetic information
- B. Building cell membranes
- C. Carrying electrons in energy-producing reactions ✓
- D. Transporting oxygen in the blood
Why: NAD+ acts as an electron carrier, shuttling electrons during metabolic reactions such as glycolysis and the citric acid cycle to help produce ATP.
Q3. Which of the following processes is NAD+ directly involved in?
- A. Photosynthesis in plant chloroplasts only
- B. Cellular respiration ✓
- C. Muscle contraction through myosin binding
- D. Synthesis of red blood cells in bone marrow
Why: NAD+ is essential for cellular respiration, where it is reduced to NADH while helping break down nutrients to release energy.
Q4. As people age, what generally happens to NAD+ levels in the body?
- A. They rise steadily throughout adulthood
- B. They remain exactly constant
- C. They tend to decline ✓
- D. They only change during childhood
Why: Research suggests NAD+ levels tend to decline with age in many tissues, which is one reason it has become a focus in ageing research.
Q5. Why is NAD+ of particular interest to researchers studying ageing?
- A. It directly reverses all forms of cell damage
- B. It is needed by enzymes such as sirtuins and PARPs that regulate DNA repair and cellular stress responses ✓
- C. It replaces the need for oxygen in the body
- D. It is the only molecule that stores energy in cells
Why: NAD+ is a required substrate for sirtuins and PARP enzymes, which play roles in DNA repair, gene regulation, and stress resistance, linking NAD+ availability to healthy ageing processes.
Module 2
Module 2: NAD+ Decline Across the Lifespan
NAD+ Decline Across the Lifespan
Knowledge check
Q1. What is NAD+ primarily involved in within cells?
- A. Carrying oxygen through the bloodstream
- B. Transferring electrons in metabolic reactions such as energy production ✓
- C. Building the structural proteins of muscle fibres
- D. Storing genetic information in the nucleus
Why: NAD+ (nicotinamide adenine dinucleotide) is a coenzyme that shuttles electrons during redox reactions, including those in glycolysis, the citric acid cycle, and oxidative phosphorylation, making it central to cellular energy production.
Q2. What general trend in NAD+ levels is commonly observed as people age?
- A. NAD+ levels rise steadily throughout adulthood
- B. NAD+ levels remain constant regardless of age
- C. NAD+ levels tend to decline with age in many tissues ✓
- D. NAD+ levels fluctuate randomly with no clear pattern
Why: Research in animal models and human tissue samples has consistently reported that NAD+ levels tend to decrease with advancing age in many tissues, although the magnitude and mechanisms vary across tissues and studies.
Q3. Which enzyme family is a major consumer of NAD+ and is linked to its age-related decline?
- A. Lactate dehydrogenases
- B. Sirtuins and PARP enzymes ✓
- C. Catalase and superoxide dismutase
- D. DNA polymerases
Why: Sirtuins and poly(ADP-ribose) polymerases (PARPs) use NAD+ as a substrate. Increased activity of these enzymes, often triggered by DNA damage and inflammation, can deplete NAD+ pools over time.
Q4. Which of the following is a precursor that the body can use to synthesise NAD+?
- A. Vitamin C
- B. Tryptophan
- C. Nicotinamide riboside ✓
- D. Retinol
Why: Nicotinamide riboside, along with nicotinamide, nicotinic acid, and tryptophan, can serve as NAD+ precursors through salvage or de novo pathways. Nicotinamide riboside is the most studied of these in ageing research.
Q5. Why is understanding NAD+ decline across the lifespan considered important in ageing research?
- A. NAD+ decline is the only known cause of ageing
- B. It may help explain links between declining metabolic function and age-associated diseases ✓
- C. NAD+ decline proves that ageing cannot be influenced
- D. NAD+ is only relevant in childhood development
Why: Falling NAD+ levels may contribute to reduced mitochondrial function, impaired DNA repair, and altered metabolism, which are linked to many age-associated conditions. However, it is one factor among many rather than the sole cause of ageing.
Module 3
Module 3: How NAD+ Fuels Cellular Energy
How NAD+ Fuels Cellular Energy
Knowledge check
Q1. What is the primary role of NAD+ in cellular energy metabolism?
- A. It directly stores energy as glucose in muscle cells
- B. It acts as an electron carrier that accepts electrons and converts to NADH during metabolic reactions ✓
- C. It breaks down proteins into amino acids for immediate energy
- D. It replaces oxygen as the final electron acceptor in the body
Why: NAD+ (nicotinamide adenine dinucleotide) functions as a coenzyme that accepts electrons during reactions such as glycolysis and the citric acid cycle, becoming NADH. The NADH then delivers these electrons to the electron transport chain to help produce ATP.
Q2. In which cellular location does the electron transport chain that uses NADH primarily occur?
- A. Cytoplasm
- B. Nucleus
- C. Inner mitochondrial membrane ✓
- D. Endoplasmic reticulum
Why: The electron transport chain is embedded in the inner mitochondrial membrane. NADH donates electrons to the first complex of this chain, driving the flow of electrons that ultimately powers ATP synthesis.
Q3. Which metabolic pathway produces NADH during the breakdown of glucose in the cytoplasm?
- A. Glycolysis ✓
- B. Urea cycle
- C. Beta-oxidation of fatty acids in the liver only
- D. Photosynthesis
Why: Glycolysis splits glucose into pyruvate in the cytoplasm and generates NADH by reducing NAD+. This is one of the first steps in extracting energy from glucose.
Q4. What is the main product of oxidative phosphorylation that NAD+-derived electrons help create?
- A. Carbon dioxide
- B. Lactic acid
- C. ATP ✓
- D. Glycogen
Why: Electrons from NADH move through the electron transport chain, pumping protons and creating a gradient. ATP synthase then uses this gradient to produce ATP, the main energy currency of the cell.
Q5. Why is maintaining a healthy balance of NAD+ and NADH important for cells?
- A. NADH is toxic and must be fully eliminated
- B. The NAD+/NADH ratio drives metabolic reactions, so imbalances can impair energy production ✓
- C. NAD+ only functions in the nucleus and not in energy pathways
- D. NADH and NAD+ are identical molecules with no functional difference
Why: The ratio of NAD+ to NADH governs the direction and efficiency of many redox reactions. Since NAD+ is needed to keep glycolysis and the citric acid cycle running, a shift in this balance can reduce the cell's capacity to generate energy.
Module 4
Module 4: NAD+ and DNA Repair
NAD+ and DNA Repair
Knowledge check
Q1. Which molecule is a primary substrate consumed by PARP enzymes when they repair damaged DNA?
- A. ATP
- B. NAD+ ✓
- C. FAD
- D. Coenzyme A
Why: PARP (poly ADP-ribose polymerase) enzymes use NAD+ as a substrate, cleaving it to nicotinamide and ADP-ribose and attaching poly(ADP-ribose) chains to proteins at sites of DNA damage to recruit repair machinery.
Q2. What is the main role of PARP1 when a single-strand DNA break occurs?
- A. It permanently silences the damaged gene
- B. It detects the break and signals repair proteins to the site ✓
- C. It replicates the damaged strand before repair
- D. It converts NAD+ directly into ATP
Why: PARP1 binds rapidly to DNA breaks and, once activated, synthesises poly(ADP-ribose) chains that act as a signal and scaffold to recruit base excision repair and other DNA repair factors.
Q3. Why is excessive PARP activation considered a concern in ageing and disease?
- A. It increases the number of mitochondria in cells
- B. It can deplete cellular NAD+ and ATP, potentially triggering cell dysfunction or death ✓
- C. It causes DNA to become more tightly coiled
- D. It prevents any cell division from occurring
Why: Because PARP consumes NAD+ each time it is activated, severe or chronic DNA damage can drain the NAD+ pool, impairing energy metabolism and sirtuin activity and contributing to cellular decline.
Q4. Which NAD+-dependent family of enzymes is closely linked with PARP in the regulation of DNA repair and genomic stability?
- A. Sirtuins ✓
- B. Cytochrome P450 enzymes
- C. Aldehyde dehydrogenases
- D. Kinases of the MAP pathway
Why: Sirtuins (such as SIRT1, SIRT6) are NAD+-dependent deacylases that also participate in DNA repair and chromatin regulation, so PARP and sirtuins compete for the same NAD+ pool, linking DNA damage to metabolic status.
Q5. Which of the following best describes the relationship between NAD+ levels and DNA repair efficiency?
- A. Higher NAD+ availability generally supports more efficient PARP-mediated repair, while NAD+ depletion impairs it ✓
- B. NAD+ levels have no effect on DNA repair
- C. Lower NAD+ levels always accelerate DNA repair
- D. NAD+ only repairs DNA in the nucleus after cell death
Why: Since PARP enzymes require NAD+ as a substrate, adequate NAD+ levels are needed for effective repair; when NAD+ is low, the repair response is compromised, which is one proposed mechanism linking NAD+ decline to ageing.
Module 5
Module 5: Sirtuins and the NAD+ Connection
Sirtuins and the NAD+ Connection
Knowledge check
Q1. What is the primary role of NAD+ in sirtuin activity?
- A. It acts as an enzyme that breaks down sirtuin proteins
- B. It serves as a substrate that sirtuins require to carry out their deacetylation reactions ✓
- C. It binds to DNA to prevent sirtuins from accessing genes
- D. It converts sirtuins into their inactive form
Why: Sirtuins are NAD+-dependent deacetylases. They consume NAD+ as a co-substrate while removing acetyl groups from target proteins, releasing nicotinamide as a byproduct. Without adequate NAD+, sirtuin activity declines.
Q2. Sirtuins are best described as which type of enzyme?
- A. Kinases that add phosphate groups to proteins
- B. Proteases that cut proteins into fragments
- C. NAD+-dependent deacetylases ✓
- D. Polymerases that synthesise new DNA strands
Why: Sirtuins belong to a family of class III histone deacetylases that require NAD+ to remove acetyl groups from lysine residues on histones and other proteins. This deacetylation regulates gene expression, metabolism, and stress responses.
Q3. How many sirtuin proteins (SIRT1 to SIRT7) are found in mammals?
- A. Three
- B. Five
- C. Seven ✓
- D. Twelve
Why: Mammals have seven sirtuins, SIRT1 through SIRT7, which are located in different cellular compartments. SIRT1 is mainly nuclear, SIRT3 is mitochondrial, and SIRT6 is involved in DNA repair and chromatin maintenance.
Q4. Which sirtuin is most closely associated with regulating mitochondrial function and energy metabolism?
- A. SIRT1
- B. SIRT2
- C. SIRT3 ✓
- D. SIRT7
Why: SIRT3 is the major mitochondrial deacetylase. It regulates enzymes involved in the citric acid cycle, fatty acid oxidation, and oxidative stress responses, making it central to cellular energy metabolism.
Q5. Why is the link between NAD+ levels and sirtuin activity important in ageing research?
- A. NAD+ levels rise sharply with age, which accelerates sirtuin activity
- B. NAD+ levels tend to decline with age, which may reduce sirtuin activity and contribute to age-related decline ✓
- C. Sirtuins produce NAD+, so ageing has no effect on their relationship
- D. NAD+ only affects sirtuins in young organisms
Why: NAD+ levels generally fall as organisms age, which may limit the activity of NAD+-dependent sirtuins. Researchers study this connection because restoring NAD+ is hypothesised to support sirtuin function and healthier ageing, though many questions remain unresolved.
Module 6
Module 6: Lifestyle Factors That Influence NAD+ Levels
Lifestyle Factors That Influence NAD+ Levels
Knowledge check
Q1. Which of the following lifestyle factors is most consistently linked to higher NAD+ levels in the body?
- A. Chronic sleep deprivation
- B. Regular physical exercise ✓
- C. Prolonged daily sitting without breaks
- D. A diet high in refined sugar
Why: Regular physical exercise has been shown in studies to support NAD+ levels, particularly in muscle tissue, by influencing the enzymes involved in NAD+ synthesis and metabolism. Sleep deprivation, prolonged sitting, and high refined sugar intake are generally associated with poorer metabolic health and are not linked to higher NAD+ levels.
Q2. Alcohol consumption can affect NAD+ levels because:
- A. Alcohol metabolism uses up large amounts of NAD+ in the liver ✓
- B. Alcohol directly produces NAD+ in the bloodstream
- C. Alcohol increases the activity of the sirtuin enzymes
- D. Alcohol has no measurable effect on NAD+ metabolism
Why: When the liver breaks down alcohol, enzymes such as alcohol dehydrogenase and aldehyde dehydrogenase rely on NAD+ as a cofactor. Heavy drinking can therefore draw down the body's NAD+ pool, which is one reason it is considered a factor that may lower NAD+ levels.
Q3. Which dietary pattern is generally considered supportive of healthy NAD+ metabolism?
- A. A diet rich in whole foods, vegetables, and adequate protein ✓
- B. A diet consisting mainly of ultra-processed snacks
- C. A diet that eliminates all B vitamins
- D. A diet with very high levels of added sugar
Why: NAD+ is synthesised from precursors such as tryptophan and B3 vitamins (niacin), which are found in whole foods, protein sources, and vegetables. A balanced whole-food diet supplies these building blocks, whereas ultra-processed or sugar-heavy diets provide few of them and are linked to metabolic stress.
Q4. Why is sleep considered relevant to NAD+ levels?
- A. Sleep converts NAD+ directly into energy for the brain
- B. Poor or insufficient sleep disrupts circadian rhythms that regulate NAD+ synthesis and metabolism ✓
- C. NAD+ is only produced during sleep and never during waking hours
- D. Sleep has no connection to cellular energy pathways
Why: Many NAD+-related processes follow circadian rhythms, and the enzymes that synthesise and recycle NAD+ show daily variation. Chronic poor sleep can disrupt these rhythms, which is why sleep quality is considered one of the lifestyle factors relevant to NAD+ balance.
Q5. Which statement about smoking and NAD+ is most accurate?
- A. Smoking increases NAD+ because it stimulates metabolism
- B. Smoking has no effect on cellular energy systems
- C. Smoking contributes to oxidative stress and DNA damage, which increases demand for NAD+-dependent repair enzymes and can lower NAD+ availability ✓
- D. Smoking only affects the lungs and not cellular metabolism
Why: Smoking produces oxidative stress and DNA damage throughout the body. Repair enzymes such as PARPs consume NAD+ when responding to DNA damage, so higher damage loads can deplete the NAD+ pool. This makes smoking a factor associated with lower NAD+ availability.
Module 7
Module 7: Current Research, Evidence, and Open Questions
Current Research, Evidence, and Open Questions
Knowledge check
Q1. Which molecule is the central subject of NAD+ research in ageing?
- A. Nicotinamide adenine dinucleotide, a coenzyme found in all living cells ✓
- B. Adenosine triphosphate, the main energy currency of the cell
- C. Collagen, the structural protein of connective tissue
- D. Melatonin, a hormone that regulates sleep cycles
Why: NAD+ (nicotinamide adenine dinucleotide) is a coenzyme present in every living cell. It takes part in redox reactions in metabolism and serves as a substrate for sirtuins and PARPs, which is why it is studied in relation to ageing.
Q2. What is the main reason NAD+ levels are thought to decline with age?
- A. Increased consumption by enzymes such as CD38 and PARPs, alongside reduced synthesis ✓
- B. Complete loss of the nicotinamide precursor from the diet
- C. Permanent deactivation of the mitochondria after puberty
- D. Accumulation of NAD+ in the bloodstream that blocks cell uptake
Why: Research suggests NAD+ falls with age because enzymes that consume it, such as CD38 and PARP, become more active, while the biosynthetic pathways that regenerate it can become less efficient. The exact balance and its causes are still being studied.
Q3. Which of the following is a precursor that can be used to raise NAD+ levels?
- A. Nicotinamide riboside ✓
- B. Retinol acetate
- C. Ascorbic acid
- D. Biotin
Why: Nicotinamide riboside (NR), along with nicotinamide mononucleotide (NMN) and niacin, is a precursor that the body can convert into NAD+. NR and NMN have been the most studied in human and animal research on ageing.
Q4. What is a key open question in current NAD+ and ageing research?
- A. Whether raising NAD+ levels in humans reliably extends healthy lifespan, since many results come from animal studies ✓
- B. Whether NAD+ has any biological role at all in human cells
- C. Whether NAD+ supplements are already proven to reverse all age-related disease
- D. Whether NAD+ can be absorbed through the skin only
Why: Much of the evidence comes from mice, yeast and worms, and human trials have so far been small and short. Whether NAD+-boosting interventions extend healthy lifespan in people, and at what dose, remains an open question.
Q5. Why should a beginner be cautious when interpreting headlines about NAD+ supplements?
- A. Animal and early human findings often do not translate directly to proven benefits in people ✓
- B. NAD+ supplements are always banned in every country
- C. NAD+ is toxic at any measurable concentration in the body
- D. Ageing research is only conducted on plants
Why: Headlines often report results from mice or cell studies. Early human trials may show that NAD+ levels rise safely, but that does not yet prove improved health outcomes or longevity, so claims should be weighed against the strength of the evidence.