Fig. 4

GEN ameliorates the cell injuries caused by H/R in NRVMs. A CCK-8 assay detected cell viability of NRVMs (n = 3). The results are presented as a percentage of the control. Cell viability of NRVMs following different concentrations of GEN exposure was measured by CCK-8 assay. NRVMs were treated with 20 μM and 40 μM with or without H/R insult. B, C Cell supernatants of LDH leakage and the change of cell viability in NRVMs. The cells were treated with the indicated treatments. D, E Expression of p-AMPK, AMPK, p-ACC, ACC, TXNIP, NLRP3, ASC, cleaved caspase-1, N-GSDMD-, IL-1β proteins detected by western blotting (n = 3). F, H Densitometry data of p-AMPK, p-ACC, ACC, TXNIP, NLRP3, ASC, cleaved caspase-1, N-GSDMD-, IL-1β in myocardium of each group of mice. I, J IL-1β and IL-18 in cell supernatants were detected by ELISA assays. K, L Intracellular red and green fluorescence of JC-1 was determined using a confocal microscope. The aggregates and monomers were assessed (n = 3); scale bar = 50 μm. M SOD2 enzymatic activity of NRVMs assayed with a commercial kit. N Representative images of immunofluorescence staining of p-AMPK in cardiomyocytes after different doses of GEN treated and the calculated intensity of p-AMPK (n = 3); scale bar = 20 μm. O Representative images of immunofluorescence staining of NLRP3 after H/R insult with 40 μM GEN treatment and the calculated intensity of NLRP3 (n = 3); scale bar = 20 μm. The data for each group were shown as the mean ± SD; ^#p < 0.05, ^##p < 0.01 vs. Control group; *p < 0.05, **p < 0.01 vs. H/R + Vehicle group