Excess endothelial production of reactive oxygen species (ROS) has been found to play a major role in the pathogenesis of many cardiovascular diseases. Ageing is a primary risk factor of cardiovascular diseases, however the role of oxidative stress in the development of vascular dysfunction in the elderly remains unknown. In this study we used wild-type C57BL/6 J mice, at young (3–4 m) and old (20–24 m) age to investigate the potential role of ROS in age-related metabolic disorders and vascular dysfunction. The body weight, heart weight and fasting serum glucose were recorded. There was no significant difference in the heart/body weight ratio, and the fasting glucose between the two groups. However, there was a significant increase in blood pressure in ageing mice (141 mm Hg) compared to young mice (126 mm Hg) measured by volume pressure recording tail plethysmography. Vascular tone was examined using aortic rings in an organ bath. There was no significant difference in vessel relaxation to a NO donor (sodium nitroprusside) between the young and ageing mice. However, there was a significant decrease in endothelium- dependent vessel relaxation to acetylcholine in ageing mice compared to young mice. There was a significant increase in the levels of superoxide production by the ageing vessels compared to the young controls as measured using lucigenin-chemiluminescence. In conclusion, there was a significant increase in the levels of ROS production in the vasculature of ageing mice, which might contribute to the age-related endothelial dysfunction and high blood pressure.

Hyperglycaemia is a hallmark of type 2 diabetes and plays a prominent role in oxidative stress-induced endothelial cell dysfunction and the development of cardiovascular diseases. Recently, a diet enriched with short-chain fatty acids (SCFAs) i.e. acetate, propionate and butyrate, has been found to improve insulin sensitivity and reduce blood pressure in clinical studies; however, the underlying mechanism remains elusive. In this study we investigated the effect of SCFAs on high glucose-induced endothelial dysfunction in vitro. Human pulmonary microvascular endothelial cells (HPMECs) were pre-incubated with acetate (5mM), propionate (4mM) or butyrate (4mM) for 1 hour before adding high glucose (30mM) and cultured for 24 h in the presence of SCFAs. Compared to cells cultured in the normal glucose medium (5mM), high glucose induced a significant increase (48.5±12.6%; p<0.05) in endothelial reactive oxygen species (ROS) production as measured by both NADPH-dependant lucigenin-chemiluminescence and DHE fluorescence. Increased ROS production was accompanied with a significant increase in cell death (30.66±8.3%; p<0.05) as measured by both MTT and trypan blue exclusion assays, and capillary structure breakdown of HPMEC cultured on the metrigels. We then examined the effects of SCFAs on endothelial ROS production, and found that SCFAs slightly increased the levels of ROS production in HPMEC cultured in 5mM glucose medium. However, in the presence of high glucose, SCFAs significantly inhibited high glucose-induced ROS production, reduced cell death and preserved capillary structure on the metrigels. In conclusion, SCFAs inhibit high glucose-mediated endothelial oxidative stress and improve endothelial cell function. SCFAs may represent an effective therapeutic strategy as a food supplement for cardiovascular complications linked to hyperglycaemia and diabetes.

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Ageing has been recognised to be a major risk factor for the development of cardiovascular disease and growing evidence suggests a role for oxidative stress. NADPH oxidase 2 has been reported to be a major source of reactive oxygen species (ROS) generation in the cardiovascular system, however, the role of this enzyme in age-related metabolic disorders and vascular diseases remains unclear. In this study we used age-matched wild-type (WT) and Nox2-deficient (Nox2 −/− ) mice on a C57BL/6 background at young (3-4 month) and ageing (20-24 month) to investigate the role of Nox2 in age-related oxidative stress, metabolic disorders and vascular dysfunction. There was an age-related increase in blood pressure in WT mice (126 mmHg for young and 148 mmHg for ageing) (P<0.05); however the blood pressure remained at lower levels in Nox2 −/− ageing mice. Compared with young WT mice, WT ageing mice had significantly high levels of fasting serum insulin and this was accompanied with delayed clearance of glucose (P<0.05) indicating insulin resistance. However, no indication of insulin resistance was found in Nox2 −/− ageing mice. The endothelial function was examined using aortic rings in an organ bath. Compared to the young controls, there was a significant decrease in the endothelium-dependent relaxation to acetylcholine in WT ageing aortas (Emax 72% for young and 64% for ageing, P<0.05). However, endothelial function was well preserved in Nox2 −/− ageing aortas (Emax 83% for young and 80% for ageing. The ROS production was then measured in aortic sections by DHE fluorescence and a significant increase was found in ROS production in WT ageing aortas as compared to WT young controls (P<0.05). However, there was a significantly lower level of ROS production (P<0.05) in both age-matched Nox2 −/− aortas. In conclusion, Nox2-derived oxidative stress plays an important role in ageing-associated metabolic disorders and vascular dysfunction, and targeting Nox2 represents a valuable therapeutic strategy to treat these aging-related diseases.