Oxidative ageing: from proximate to ultimate causes

Oxidative stress seems really important in age-related decline and disease—but what causes it? Here I’ve tried to express a broadening perspective, by exploring its core, context and ultimate causes; and largely anchored in human studies where possible.

We all die—what matters is how. While human life expectancy has increased, non-communicable diseases are now the major cause of disability and death globally (WHO and OWID). These are mostly age-related diseases (e.g. CVD, cancer, COPD, dementia, etc.), which develop slowly over time, and coexist as multimorbidity (e.g. most people >65 in US/UK ^1,2); resulting in functional decline/frailty and socioeconomic burden (i.e. ↓ productivity, ↑ sick care). This situation is growing globally, as populations are ageing, and diseases occur earlier—so we may live longer but sicker ¹. Moreover, this invisible epidemic underlies susceptibility to (communicable) infectious diseases, such as COVID-19 ³, elevating chronic disease to acute threat.

Rather than treat each age-related malady as it arises, the other approach is to consider ageing itself ⁴. Ageing (i.e. growing old) involves a gradual systemic decline of tissue function and stability, with increasing susceptibility to disease and eventually death. However, variation between individuals and species highlights disconnect between biological and chronological ageing, and opportunities to identify modulators of healthspan. In particular, the accelerating effect of (natural) radiation is evident; and similarly, since the 1950s ageing was suggested to involve the deleterious side effects of free radicals (i.e. Harman’s seminal free radical theory) ^5,6, with subsequent incorporation into other theories (e.g. antagonistic pleiotropy ⁷, mitohormesis ⁸ and redox ⁹).

The accrued observational evidence supports that human ageing is accompanied by increasing oxidative stress, alongside decreasing antioxidant/adaptive responses ^9–12 (esp. with frailty and disease ^9,12,13, but less so in centenarians ¹⁴); while higher intakes and blood levels of various antioxidants are (dose-dependently) associated with reduced total and cause-specific mortality ^15,16. On the other hand, many antioxidant trials (i.e. mainly vit-A/C/E, β-carotene and/or selenium; primary and secondary prevention; published ~1990–10) and meta-analyses thereof show general failure to lower mortality, and even potential harm ¹⁷ (e.g. previous post). This may be due to overly crude interventions based on early paradigms ^8,9. As such, this article explores a broader perspective.

Redox theory

Free radicals and antioxidants really represent oxidation and reduction (redox) reactions, respectively, within a more complete concept of redox biology, which recognises all such reactions and their place in redox networks connected to metabolism ^18,19. For instance, antioxidants (e.g. vit-C and E) have specific chemistry and cell distribution (e.g. cytoplasm and membranes, respectively), but function together within regenerative networks which draw electrons from intermediary metabolism, and ultimately food (Fig. 1).

Fundamentally, redox reactions couple the biology of life, to the chemistry of our planet and physics of the universe. From a bio-thermodynamic perspective, organisms are open systems, where internal energy/order derives from the external environment. This biochemistry requires manipulation of chemical bonds and electron transfer in thermodynamically favourable redox reactions (i.e. a redox tower). In particular, rising atmospheric oxygen coincided with the evolution of aerobic life, which exploits the electronegativity of oxygen as a terminal electron acceptor in energy metabolism, thereby maximising energy yield; while also serving in biosynthesis and production of reactive oxygen species (ROS—incl. free radicals) ²⁰. This may have been accompanied by concurrent evolution of adaptive antioxidant responses (e.g. Nrf2 ²¹), and elaboration of redox sensing/signalling with increasing metazoan complexity (e.g. NOX ²⁰ and redox proteome ^22,23).

Redox reactions are used in a myriad of metabolic and signalling pathways. The initial oxidation of nutrients generates a reductive cache (as NAD(P)H) which fuels electron flow through intermediary metabolism; including the incidental and intentional generation of ROS (e.g. via the ETC and NOX, respectively) and counterpoise antioxidant systems, which together regulate basic cell biology. For instance, the abundant thiol-antioxidants assume specific redox potentials (E_h in mV) in specific subcellular compartments (e.g. cytoplasm, mitochondria, nucleus, etc.), staging redox environments where transient/reversible oxidations can modulate macromolecule structure and function (e.g. metabolic/signalling activity) ^22,23, and sustained oxidations mediate destruction/death, as weaponised in immunity (e.g. oxidative burst ²⁴ and cytotoxicity ²⁵). Ultimately, these redox networks follow thermodynamic hierarchy: bioenergetic systems operate at near-equilibrium with high-flux, generating energy for maintenance of thiol-antioxidants at non-equilibrium, where subcellular redox potentials maintain spatiotemporal organisation and structure ¹⁸.

Overall, redox biology represents a fundamental interface between environment and genome, which couples energy transfer to organisation/adaptation ^18,19,23 (i.e. Gibbs free energy to order). On the other hand, ageing and disease typically involve a general oxidation of redox status (e.g. thiols and macromolecules) ⁹—so what gives and with what consequence?

Glutathione

Glutathione (GSH) is the major small molecule antioxidant (i.e. thiol tripeptide—307Da) synthesised by our cells; serving as a redox buffer and regulator of numerous processes (e.g. signalling/cysteine proteome ²⁶, energy/mitochondria ^27,28, growth–death ^28,29, etc.). The reduced to oxidised glutathione ratio (GSH:GSSG) represents a functional marker of redox status, which varies between cell compartments and conditions. In general, ageing is accompanied by decreasing GSH and/or increasing GSSG ^10,30,31, and thereby oxidation of GSH:GSSG. More specifically, in a US study on healthy human plasma, cysteine oxidation was linear with age, whereas GSH oxidation started at ~45yrs old, with considerable variation (i.e. E_h of GSH:GSSG vs. age, R² = 0.32) ^9,32.

Proximally, GSH decline has been associated with a lower biosynthetic rate ^33,34, enzyme expression (e.g. GCL ^10,31 and GSS ³⁵), enzyme activity (e.g. K_m of GCL ^30,36,37 and GR ³⁸) and precursor levels (i.e. glycine and cysteine) ³³; while precursor supplementation has restored GSH and organ function in old animals ^39–43 and humans ^{33–35,44,45} (reviews ^46–48).

For instance, in 2011, a small study on healthy older adults (n=8, age 60–75), with stable-isotope tracing, revealed low precursor levels (i.e. RBC glycine –55%, cysteine –24%), while just 2 weeks of glycine-cysteine supplementation (~0.1g of each/kg/day) almost completely normalised RBC GSH synthesis rates, GSH:GSSG ratio and plasma oxidative markers (i.e. d-ROMS, F₂-isoprostanes and lipid peroxides) to values of younger controls (n=8, age 30–40; + subgrouping for BMI and glycaemia) ³³; and subsequently also mitochondrial fat oxidation and insulin sensitivity ³⁵. The same protocol in older people with HIV similarly improved GSH and oxidative markers, as well as fed/fasting mitochondrial fuel oxidation, insulin sensitivity, blood lipids, body composition, muscle strength ³⁴ and inflammation ⁴⁴. And subsequent trials report sustained responses at 12 and 24 weeks, which declined after treatment withdrawal ^49,50.

Among other research, in 1997, a relatively large placebo-controlled trial on a cohort of mostly older people with chronic diseases (n=262), with the GSH-precursor NAC for 6 months (2x 600mg/day), lowered flu symptoms and restored cell-mediated immunity ⁴⁵. Accordingly, studies on lymphocytes from older people have linked low GSH and oxidative stress to reversible impairment of cytokine secretion and proliferation ^51–53, while treatment of old animals with GSH-precursors has restored type-1 immunity ⁴³ and resistance to infection ⁴². In humans, plasma aminothiol oxidation has also been associated with cognitive decline, cardiovascular dysfunction and mortality (see refs in ⁵⁴), while in animal models GSH-precursors have restored synaptic plasticity (to that of the adult) ⁴⁰, brain markers (e.g. inflammation and neurodegeneration) ⁴¹ and cardiac function/markers (e.g. inflammation and mitochondria) ³⁹. Therefore GSH redox may regulate systemic organ function during ageing, including ‘inflamm-ageing’.

Loss of GSH redox may involve broader changes to sulfur metabolism ³⁰. In the ageing mouse liver, there were increased transsulfuration metabolites, including homocysteine (154%), which was shown to lower the affinity of GCL for its substrates, potentially restraining GSH synthesis ³⁷. The lower methionine further suggests slow homocysteine remethylation by MS (B₁₂/folate-dependant) or BHMT (betaine-dependant) pathways. Similarly, a human autopsy study showed the ageing brain is accompanied by elevated homocysteine and lower B₁₂, methionine and SAM/SAH ratios ⁵⁵. In healthy adults, correlations were also reported between redox markers and NAD(H), with homocysteine, folate and B₁₂, in blood and cerebrospinal fluid ^56,57. Stepping further out, several other amino acids decline during ageing, including serine, a precursor of glycine and cysteine, and source of 1-carbon units (via SHMT) for folate-dependent methylation ⁵⁸. In human skin cells, age-related mitochondrial respiration defects were linked to epigenetic suppression of glycine synthesis (i.e. GCAT and SHMT2 genes) ⁵⁹. Further, in mid-age people with fatty liver (i.e. NAFLD), low serine/glycine was linked to low GSH, which improved with serine supplementation ⁶⁰.

Regeneration of GSH (and other antioxidants) depends on electrons from NADPH (Fig. 1), which mainly come from cytosolic pentose phosphate (PPP) and mitochondria-driven pathways (e.g. 1-carbon, Krebs cycle and NNT) ^61,62. With ageing, NADPH may decline in cells ⁶³, but increase in human plasma within a dysregulated NAD metabolome ⁶⁴. Notably, in a small trial on older adults (n=12, mean age 72), the NAD-precursor nicotinamide riboside acutely (post 2hrs) improved redox status (e.g. RBC NAD(P)H, GSH and F₂-isoprostanes) and exercise performance ³⁸. NAD may decline with age in relation to increased consumption and decreased synthesis ^65,66; the former being associated with oxidative DNA damage and PARP activity in humans ^64,67,68, and senescence/inflammation-driven CD38 activity in animals ⁶⁹. Regardless, NAD use (i.e. via PARP, CD38 or SIRT) specifically removes the ADP-ribose portion, leaving NAM, while resynthesis requires ATP and ribose (as PRPP) ⁶¹. Upstream, glucose metabolism via the PPP generates ribose (as R5P), which also fuels de novo purine synthesis and salvage of AMP (i.e. precursor to ADP/ATP). Notably, purine synthesis requires various cofactors, including glycine and folate ⁷⁰. Moreover, in ageing humans, low plasma NAD was accompanied by increased NAM and methyl-NAM ⁶⁴; while NAM methylation consumes SAM and may be promoted by ATP deficit and inflammation ^71,72. In summary, all these studies suggest potential links from GSH/redox to broader changes in 1-carbon ⁶² and NAD metabolism ⁶³.

Nrf2 networks

Various antioxidant enzymes also decline during ageing ¹⁰, while their manipulation may affect healthspan ⁷³. However, of central importance may be the transcription factor Nrf2 ¹⁰. Nrf2 serves as a master regulator of redox homeostasis and cytoprotection, by controlling the expression of hundreds of genes, including antioxidants/GSH. Nrf2 has constitutive activity and is heavily induced by oxidative/stress conditions. However, basal and/or inducible Nrf2 activity declines during ageing and related disease ^10,11 (e.g. in humans ^74–80); and in relation to altered GCLC promoter binding and low GSH ^81,82. Note, in the studies above, neither glycine-cysteine or GSH supplementation restored Nrf2 expression ^35,42. Moreover, in animals and humans, high-dose antioxidant supplements (e.g. vit-C/E or NAC) can even inhibit exercise-induced Nrf2 signalling and mitohormesis ^8,83,84. This may represent a significant shortfall of many antioxidant approaches, since Nrf2 controls so much of relevance to ageing; including many antioxidant/detox systems, mitochondria and intermediary metabolism (e.g. NADPH, 1-carbon, etc.), iron metabolism, AGE metabolism (i.e. glyoxalase-1), autophagy, DNA repair, etc. ^85–88.

Accordingly, the Nrf2 pathway has been linked to ageing phenotypes. For instance, Nrf2 signalling is suppressed in human progeria (a genetic disease of premature ageing), where experimental evidence suggests it may drive the condition ⁸⁹. More generally, in a cohort of community-dwelling adults (n=350, age ≥65), lower Nrf2 expression in blood was associated with frailty (independent of comorbidities) ¹³. In preclinical studies, Nrf2 deficiency replicates several major transcriptomic changes occurring in the human brain during ageing and Alzheimer’s ⁹⁰; and regulates many other age-related changes, including sarcopenia ⁹¹, vascular function ⁷⁷, immunity ⁴³, neuroinflammation ⁹², neurogenesis ⁹³, lens defence ⁷⁴, collagen breakdown ⁹⁴, carcinogenesis ⁹⁵, etc.; as well as many cellular hallmarks of ageing ^11,96. Consequently, in animal models, Nrf2 supports stress-resistance, healthspan and lifespan ^{10,11,90,95,97}; as does overexpression of target genes GCL ³⁰ and G6PD ⁹⁸ (produces NADPH in PPP). Also noteworthy, NADPH-dependant mitochondrial reductase TXNRD2 (and GR) has been associated with cross-species lifespan (i.e. birds, rodents and primates) ⁹⁹.

Why does Nrf2 activity decline with age? Nrf2 is normally sequestered in the cytoplasm by Keap1, while being regulated at multiple levels by many other proteins and modifications ¹⁰. Age-related changes include decreased Nrf2 expression, with corresponding changes to various positive/negative regulators (e.g. Keap1 ^75,80, Bach1 and c-Myc ^78,82,100) and epigenetics ^10,11. For instance, in human lens cells, age and metabolic stresses were reported to lower DNA methylation of the Keap1 promoter to suppress Nrf2 activity ^80,101. More broadly, Nrf2 functions within integrated metabolite-sensing networks which also decline, including sirtuins and AMPK ¹⁰² (Fig. 2). For instance, NAD-dependent sirtuins stimulate Nrf2 (e.g. SIRT1 ^103–105, 2 ¹⁰⁶ and 6 ¹⁰⁷), reciprocally ^108–110 (via p53 ¹¹⁰); while Nrf2/GSH may also support NAD levels ^111–113 (e.g. ↑ NQO1 ^112,113, ribose/purines ⁸⁵; ↓ PARP ^64,67, CD38 ^114,115 and SASP ^69,116,117). And the energy (AMP:ATP) sensor AMPK induces NAD synthesis (via NAMPT), SIRT1 ¹⁰² and Nrf2 ^118,119 (via Bach1 ¹¹⁹), also potentially reciprocally ⁸⁷ (e.g. via Trx1 ¹²⁰).

Besides NAD ^65,66, these integrated pathways might decline in relation to various other signals and metabolites. For instance, they are induced by 1-carbon metabolites (e.g. serine ⁵⁸ and folate ^121,122), and several others linked to methionine metabolism (which may decline during ageing), including the methyl donor SAM ^55,123, the polyamine spermidine ^124,125, the circadian hormone melatonin ^126,127 and the gaseous mediator H₂S ^128–131. Meanwhile, as above, human ageing can involve decreased methionine (i.e. precursor to SAM; and thereby polyamines, melatonin, DNA methylation, etc.) and increased homocysteine (i.e. precursor to methionine and cysteine/H₂S ¹³²); in relation to oxidative stress ¹³³ and B₁₂ deficiency ⁵⁵, and ultimately impaired MS activity ¹³⁴. Whereas Nrf2 activity may reciprocally oppose all this by supporting serine ^85,135 and homocysteine metabolism (i.e. ↑ B₁₂/MS ^136,137 and CBS/CSE ¹³⁸).

The gut microbiome

The gut microbiome shifts during ageing and can be used to predict chronological age ¹³⁹. This may be of consequence to systemic metabolism and redox. For instance, the prevalence of small intestinal bacterial overgrowth (SIBO) may increase with age ^139–141, which could increase nutrient competition with host, resulting in malabsorption (e.g. B₁₂ ¹⁴⁰). Similarly, the gut microbiome also regulates systemic GSH metabolism, by limiting serine/glycine availability, which might be of importance in age-related metabolic disease ^60,142. On the other hand, the colonic microbiome is emerging as a source of host folate ^143,144 (also an auxotrophic nutrient for beneficial butyrate-producing bacteria ¹⁴⁵­), the NAD-precursor NAM ¹⁴⁶ and polyamines ¹²⁴.

The gut microbiome also profoundly regulates immunity; and since 1907, Metchnikoff already suggested gut barrier disruption may drive chronic inflammation and senescence ¹⁴⁷. Accordingly, recent studies in older humans have shown that microbial markers (i.e. LPS, LBP and sCD14) are elevated in plasma ^148,149 and correlate inflammation, glucose control (i.e. HbA1c) and poor physical function ^148–150; whereas centenarians had lower LPS than even young adults ¹⁵¹. In animal models, the gut microbiota drives inflamm-ageing ¹⁴⁷, in association with gut dysbiosis (e.g. ↓ Akkermansia; ↑ TM7 and proteobacteria ¹⁵²), gut inflammation and microbial translocation ^147,152. Microbial components can also trigger production of antimicrobial ROS/RNS, which may burden systemic redox homeostasis. Accordingly, blood LPS correlated NOX2 and oxidative stress in elderly people with neurodegenerative diseases ¹⁵³.

Age-related oxidative stress and organ dysfunction was also recently linked to the meta-organismal TMAO pathway ^154–158. Here, dietary TMA-precursors (e.g. carnitine and choline) fuel microbial production of TMA, which is converted by liver FMO3 to TMAO, which circulates blood and is excreted by the kidneys. In animals and humans, blood TMAO increases with age ^154–158, in relation to oxidative stress and vascular dysfunction ^155,157,158; and neuroinflammation, cognitive decline ¹⁵⁶ and dementia ¹⁵⁹. In animals this occurs alongside gut dysbiosis, including overgrowth of proteobacteria and Desulfovibrio (a sulfur-reducing and TMA-producing genus), and increased FMO3 expression ¹⁵⁸, while suppression of microbial TMA formation normalises vascular dysfunction ^155,157. Ultimately, TMAO may suppress SIRT1/3 activity to promote oxidative stress, inflammation ¹⁶⁰ and senescence ¹⁵⁴.

On the other hand, there are likely reciprocal interactions. For instance, gastric acid secretion declines with age, while gastritis increases, which may promote maldigestion and B₁₂ deficiency ¹⁶¹. Of potential relevance, acid-secreting parietal cells of the stomach are extremely densely populated with mitochondria, and their function is sensitive to redox and energy status ^162,163. Gut oxidative stress may also slow intestinal motility to promote SIBO ^164,165, and directly affect microbiome balance ¹⁶⁶ and diversity ¹⁶⁷. Note, SOD1-deficient mice also have reduced gut Akkermansia and NAM metabolism ¹⁴⁶. Also, age-related gut microbiota change was dependent on inflammatory signalling and reversible with anti-TNF therapy ¹⁴⁷.

The environment

Currently, there seem only minor associations between genetic variation and ageing, although a recent meta-analysis of genomic studies implicated those relating to CVD and heme metabolism; and thereby iron-based oxidative stress ¹⁶⁸. Many environmental factors are associated with age-related disease and mortality. Accordingly, the reciprocal metabolic and microbial networks discussed above suggest vicious cycles which may be receptive to many environmental cues (e.g. diet, toxins, microbes, exercise, sleep, psych/stress, etc.). Our exposure to these factors throughout life constitute the exposome, which may accumulate as memories/scars in multiple bio-systems/structures to erode quality and induce hallmarks of ageing (e.g. redox ⁹, information ¹⁶⁹ and extracellular matrix (ECM) theories ¹⁷⁰); some of which may be readily reversible, others less so. Indeed, cells are equipped with mechanisms to repair and recycle damaged components, while altering the extracellular environment (e.g. ECM ¹⁷⁰, blood plasma ¹⁷¹ or Cys redox ¹⁷²) appears sufficient to rejuvenate old cells/tissues—so what about the external environment?

Diet is the natural source of metabolites for microbiome, metabolism and redox biology ¹⁹, with potential to influence age-related changes. Accordingly, in a cohort of Irish elderly (n=178, mean age 78), health status was related to gut microbiota, which was related to diet, where the most microbial and dietary diversity related to a low fat/high fibre pattern ¹⁷³. Similarly, in a larger trial, adherence to a Mediterranean-style diet (i.e. NU-AGE diet) was associated with favourable modulation of the gut microbiome in relation to other health markers ¹⁷⁴; and in another large study, higher Med-diet scores (and lower BMI) associated with less oxidised thiol redox in mid-age people ¹⁷⁵. Such plant-rich diets might favourably modulate pathways discussed above (e.g. ↑ carbs/fibre, antiox, folate, polyamines; ↓ Met:Gly, carnitine, heme-iron, etc.). On the other hand, better nutritional status was associated with SIRT1 expression in older adults, independent of Med-diet scores ¹⁷⁶. Also, high protein intake, and even more so plasma urea, was associated with markedly lower NAD levels in older adults (independent of age, gender, eGFR and calories), implicating amino acid catabolism in NAD decline ⁶⁸; while in mice high protein intake upregulated liver mitochondrial protein content (incl. NAD transporter) and oxidative stress ¹⁷⁷.

Whole plant foods (e.g. fruit, veg and grains) are dose-dependently associated with healthspan and longevity ^178,179, and represent the foundation of many nutritional guidelines. Furthermore, dietary antioxidant capacity is also dose-dependently related to lower total, CVD and cancer mortality ^15,16. Whole plant foods are especially rich in many antioxidants ¹⁸⁰ and related phytochemicals ¹⁸¹ which can promote Nrf2 activity in cells (e.g. polyphenols, glucosinolates, etc.) ¹⁸²; e.g. as redox cofactors ¹⁸³, ROS-stimulants ^184,185 or via other pathways (e.g. epigenetics ¹⁸⁶, AMPK/SIRT1 ^105,109, etc.). While non-nutrient phytochemicals typically have limited absorption and rapid metabolism/clearance, brief Nrf2 stimulation has lasting effects on protein expression, which accumulates with repeat stimulation ¹⁸⁷. Moreover, limited polyphenol absorption leads to accumulation in the gut, where microbial metabolism generates smaller secondary metabolites, which are more bioavailable and persist in circulation ^188,189.

Modulation of the Nrf2 pathway can occur within hours after a single meal in young healthy humans: decreasing after a processed high-fat meal (at 5hrs 75%), but stimulated by addition of polyphenols (at 3hrs 250%) ¹⁹⁰. Importantly, some longer trials with polyphenol-rich foods/extracts have improved redox status in older people (e.g. mean age 50–60 ^191–193 or 60–76 ^194–197; w/disease ^{192–194,197} and AMPK/SIRT/Nrf2 ^191,194,197), alongside various organ markers/functions (Table), suggesting the body remains responsive during ageing and disease. Similarly, in ageing animals, Nrf2/redox and organ function is improved by phenols (e.g. resveratrol ¹⁰⁹, green tea catechins ¹⁹⁸, cocoa epicatechin ¹⁹⁹, curcumin ²⁰⁰, amla ²⁰¹ and sesamol ²⁰²) and glucosinolates (e.g. sulforaphane ^43,74,92); as well as by probiotics ^203,204 and butyrate ²⁰⁵. Perhaps noteworthy, unlike GSH-precursors ³⁹, polyphenols also improved the ECM (e.g. arteries ²⁰⁰, muscle ²⁰⁶ and skin ²⁰⁷).

Another important component of a Mediterranean diet is fish, which itself generally has positive associations with healthspan, in relation to omega-3s ^208–210; and while supplement trials seem less consistent, this may involve several confounding factors obscuring benefit ^211,212. Further, a meta-analysis of RCTs suggests omega-3s can also improve general blood redox markers ²¹³; and a recent trial with EPA/DHA (2700mg/day) showed robust induction of Nrf2 in PBMCs in T2D ²¹⁴. Accordingly, while polyunsaturated fatty acids are highly susceptible to oxidation, especially omega-3s, they generate specific products (e.g. J₃-isoprostanes ²¹⁵, 4-HHE ²¹⁶ and oxygenase-derived resolvin D1 ^217,218) which can induce Nrf2 antioxidant responses. In mice, fish oil increased DHA/4-HHE and HO-1 in multiple organs ²¹⁶; and a genetically increased omega-3/6 ratio induced Nrf2/HO-1 and lowered UVB-induced skin oxidative stress, inflammation and carcinogenesis ^219,220. Therefore omega-3 status (and omega-6/3 ratios) may influence redox homeostasis and age-related disease; with a notable abundance of DHA in brain and retina ²²¹.

On the other hand, toxins can disrupt redox homeostasis, as with TMAO above. Note, deep-sea fish also contain particularly high levels of preformed TMAO ²²² (e.g. cod vs. salmon ²²³). Moreover, our oceans are now polluted and the positive health associations with marine omega-3s may be offset by concomitant toxin/PCB intake ^209,210. Another important factor may be lifelong accumulation of redox-active metals ⁹. For instance, in a representative Mediterranean population (n=815), plasma oxidative stress correlated age, heme-iron intake (from meat and fish) and transferrin saturation (i.e. iron status) ²²⁴. Further, several toxic metals (e.g. Pb, Cd and Hg) have also been found to accumulate with age in specific organs, where they may promote oxidative stress and disease ^225–228. And regarding air pollution, chronic or high exposure to urban particulate matter can inhibit Nrf2 activity ²²⁹; as can cigarette smoke via oxidation products ²³⁰ and post-translational modifications (prevented by resveratrol) ²³¹.

Advanced glycation end-products (AGEs, aka. glycotoxins) also accumulate with age and disease in relation to inflammation and oxidative stress ²³². AGEs can form endogenously and exogenously in food. In particular, heat-processed animal and high-fat foods are rich in AGEs ²³³, which have partial absorption (~10–30%) and affect host and gut microbiome ²³⁴. In healthy US adults, there are correlations between diet and blood AGEs, inflammatory and oxidative markers ^235–237; all of which were lowered simply by switching to wet cooking methods ^236,237. Further, in a 1-year RCT on older people with MetS (n=138, mean age 61), a low-AGE diet also greatly normalised insulin sensitivity ²³⁸. Moreover, the ability of calorie restriction to extend animal health and lifespan has also been linked to lower dietary AGEs, with favourable modulation of GSH redox and oxidative stress ²³⁹. Note, AGEs may especially accumulate on long-lived macromolecules (e.g. ECM, esp. cartilage and bone) ²³², where they can mediate various deleterious effects ¹⁷⁰, including inhibition of SIRT1/Nrf2 activity ^{101,237,238,240} (prevented by curcumin ²⁴¹). In animal models, at least arterial AGE accumulation can be quickly reversed (e.g. curcumin ²⁰⁰ and spermidine ²⁴²).

Exercise is also associated with healthspan, which may involve hormesis and Nrf2 activity ^83,84. Accordingly, an active lifestyle was associated with preserved Nrf2 activity in skeletal muscle of older people ⁷⁵. Further trials have shown that acute exercise-induced Nrf2 signalling is impaired in PBMCs ⁷⁶, while aerobic training improved resistance to oxidative stress in forearm ²⁴³. Similarly, in old animals, moderate exercise training improved SIRT1/redox in skeletal muscle ²⁴⁴ and Nrf2/redox in heart ²⁴⁵. Note however, exercise and weight loss did not improve leaky gut markers in older people ¹⁴⁸, or those with cardiometabolic disease ¹⁵⁰, while polyphenols did in MetS ²⁴⁶ and T2D ¹⁹⁷. Also, while there is suggestion that (compared to antioxidant vitamins) phytochemicals may not interfere with exercise adaptations ²⁴⁷, or even support them ⁸³, they still can in pharmacologic dose/form in young/aged humans (e.g. epicatechin ²⁴⁸ and resveratrol ²⁴⁹).

Evolution vs. entropy

The biochemistry of life ultimately follows the physics of the universe. In particular, the 2^nd law of thermodynamics describes how all isolated systems spontaneously evolve toward equilibrium, with dissipation of all energy—i.e. a state of maximum entropy (S, as joules per kelvin). As open systems, organisms continually take in food/energy, while expelling entropic waste, enabling maintenance of their internal order (i.e. non-equilibrium) indefinitely ^250,251. More specifically, aerobic life harnesses non-equilibrium electrochemistry to drive bioenergetics, while coupling the Gibbs free energy to self-organisation (e.g. non-equilibrium biochemistry, biosynthesis and barriers). Order is further protected by homeostatic/defence pathways which commit resources toward clearance of molecular damage, dysfunctional cells and exogenous threats (e.g. toxins and infections).

However, this gradually fails with ageing, which is considered a state of increasing disorder analogous to entropy ^251–256 (i.e. energy and information entropy ^169,256); characterised by accumulation of stochastic modifications to macromolecules (e.g. via glycation ^170,253, DNA methylation ^169,256, etc.) and loss of non-equilibrium (e.g. via chemical potentials/gradients and barriers ⁹), with increasing cell/tissue heterogeneity and dysfunction, and susceptibility to disease and death (i.e. ‘…a rush toward equilibrium’ ²⁵¹). In essence, ageing may involve a gradual loss of molecular fidelity/clarity due to loss of thermodynamic efficacy (i.e. energy–order) ²⁵⁴. Notably, ageing rate varies widely between species, and in some may be negligible, suggesting the extent of self-maintenance may be determined by ecological niche and evolutionary pressures ^250,252,255. As such, in most animals, through reproductive maturation (and parenthood) there may be decreasing selective pressure (i.e. evolutionary value) on such homeostatic/repair systems, thus constraining their function.

On the other hand, while evolution (past environment) shapes the genes which ultimately determine capacity for longevity ²⁵², the present environment still constantly modulates physiology. Accordingly, various redox modulators have rejuvenated age and disease-related organ dysfunction in older humans, animals and cells (Table). Therefore, our current environment may be limiting, and even late optimisation, via removal of disruptive/toxic stressors and re-engagement of homeostatic pathways, may enable some restoration of redox biology, thermodynamic efficacy and physiological function. This may improve/square healthspan and disease-limited lifespan, to more fully express genomic potential; beyond which we may be particularly limited in our ability to clear accumulative extracellular changes and ultimately to maintain aesthetics and function ^170,232,253.

Perspective

In summary, much age-related physiological decline seems tied to oxidation of redox status (i.e. oxidative ageing, or ‘oxid-ageing’), in relation to various proximate metabolic and signalling changes, which link to microbial and environmental factors (e.g. diet/lifestyle), and ultimate evolutionary constraints. Importantly, many biological changes seem to remain amenable throughout ageing and disease, where various redox modulators have rejuvenated age-related organ dysfunction. Therefore redox status may be a continuous determinate of physiological function, and regulator of ageing rate and reversibility, which links past and present environment to thermodynamic efficacy and healthspan.

The flexibility of ageing might have some provocative social implications. For instance, to what extent does our notion of ‘normal ageing’ encompass preventable and even reversible functional decline and disease, and therefore suboptimal, unhealthy ageing? Is age-related functional decline an early manifestation of subclinical disease and prodromal to age-related disease? Are the top causes of morbidity and mortality an inevitable consequence of living longer, or environmental mismatch—i.e. our success or failure? Is unhealthy ageing coupled to an unhealthy planet? Moreover, recently the fundamental biological features of spaceflight were revealed, with many notable similarities to ageing, including chronic oxidative stress ²⁵⁷. Therefore optimising our redox biology may be important not just for health on this planet, but for enabling travel to others.

In memory of JM

Resources

• Diet: Harvard, EAT-Lancet
• Databases: polyphenols, polyamines, AGEs

References

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