Objective: To investigate whether ginsenoside Rb1 (Rb1) can protect human umbilical vein endothelial cells (HUVECs) against high glucose-induced apoptosis and examine the underlying mechanism.
Methods: HUVECs were divided into 5 groups: control group (5.5 mmol/L glucose), high glucose (HG, 40 mmol/L) treatment group, Rb1 (50 µ mol/L) treatment group, Rb1 plus HG treatment group, and Rb1 and 3-(1H-1,2,3-triazol-4-yl) pyridine (3-TYP, 16 µ mol/L) plus HG treatment group. Cell viability was evaluated by cell counting kit-8 assay. Mitochondrial and intracellular reactive oxygen species were detected by MitoSox Red mitochondrial superoxide indicator and dichloro-dihydro-fluorescein diacetate assay, respectively. Annexin V/propidium iodide staining and fluorescent dye staining were used to measure the apoptosis and the mitochondrial membrane potential of HUVECs, respectively. The protein expressions of apoptosis-related proteins [Bcl-2, Bax, cleaved caspase-3 and cytochrome c (Cyt-c)], mitochondrial biogenesis-related proteins [proliferator-activated receptor gamma coactivator 1-alpha, nuclear respiratory factor-1 and mitochondrial transcription factor A)], acetylation levels of forkhead box O3a and SOD2, and sirtuin-3 (SIRT3) signalling pathway were measured by immunoblotting and immunoprecipitation.
Results: Rb1 ameliorated survival in cells in which apoptosis was induced by high glucose (P<0.05 or P<0.01). Upon the addition of Rb1, mitochondrial and intracellular reactive oxygen species generation and malondialdehyde levels were decreased (P<0.01), while the activities of antioxidant enzymes were increased (P<0.05 or P<0.01). Rb1 preserved the mitochondrial membrane potential and reduced the release of Cyt-c from the mitochondria into the cytosol (P<0.01). In addition, Rb1 upregulated mitochondrial biogenesis-associated proteins (P<0.01). Notably, the cytoprotective effects of Rb1 were correlated with SIRT3 signalling pathway activation (P<0.01). The effect of Rb1 against high glucose-induced mitochondria-related apoptosis was restrained by 3-TYP (P<0.05 or P<0.01).
Conclusion: Rb1 could protect HUVECs from high glucose-induced apoptosis by promoting mitochondrial function and suppressing oxidative stress through the SIRT3 signalling pathway.
Keywords: SIRT3 signalling pathway; apoptosis; ginsenoside Rb1; high glucose; human umbilical vein endothelial cells; mitochondria.