Fig 2: Compound C decreases insulin signalling and reduced NaHS-induced glucose uptake in IR cells. PA-treated C2C12 cells incubated with compound C (20 µM, 30 min) were cultured with DMEM containing 50 µM NaHS for 30 min. (a) NaHS-induced phosphorylation of IRS1, PI3K, AKT and AMPK was blocked by compound C. (b) NaHS-induced enhancement of glucose uptake was inhibited by compound C (20 µM, 30 min) in IR cell models. *p < 0.05, **p < 0.01, ***p < 0.001. Data are shown as the mean ± sem.

Fig 3: STO-609 inhibition of the effect of NaHS to stimulate the insulin signalling pathway and AMPK to improve glucose uptake in the IR cell model. (a) C2C12 myoblasts were treated with the CaMKKß inhibitor, STO-609 (25 µM, 30 min), and stimulated with or without insulin (10 nM, 10 min). STO-609 blocked the phosphorylation of IRS1, PI3K, AKT and AMPK induced by NaHS. (b) Additional increases of the NaHS-induced 2-NBDG uptake was blocked by STO-609 (25 µM, 30 min) in PA-induced IR C2C12 cells. **p < 0.01, ***p < 0.001. Data are shown as the mean ± sem.

Fig 4: Schematic illustration of the mechanism for the preventive effect of H2S on the aging process. Oxidative stress is induced by reduced antioxidants, imbalanced protein homeostasis and dysfunctional mitochondria. Additionally, oxidative stress induces mitochondrial mutations and dysfunction. Mitochondrial mutations are also closely related with the mitochondrial repair function. Mitochondrial dysfunction contributed to the activation of apoptosis and the loss of irreplaceable cells that, ultimately, led to the aging of tissues or organs and age-related disease. Exogenous H2S attenuates oxidative stress-induced aging by mechanisms that enhance antioxidants, improve mtDNA repair capability and strengthen protein homeostasis (proteostasis). The protective mechanisms were implemented by activation of the PI3K/AKT and CaMKKβ/AMPK pathways.

Fig 5: Western blot analysis of the effects of metformin on the (A and B) ERK1/2/AMPK and the (C and D) UPR in PC12 cells. *P<0.05, **P<0.01, ***P<0.001. The data are presented as the means ± SD; n=6 for each subgroup. D-gal, D-galactose; Met, metformin; U, U0126; CC, compound C.