Fig. 6.

Pinocembrin alleviates IH-induced mitochondria damage and cell inflammation via BNIP3-dependent mitophagy. a The changes of mitochondrial membrane potential (MMP) in BV2 cells with different treatments were analyzed quantitation of fluorescence intensity through flow cytometry. b Quantitative analysis of MMP was represented as the ratio red to green fluorescence. c The levels of cellular mitochondrial ROS (mtROS) were determined by mitoSOX^TM red mitochondrial superoxide indicator using flow cytometry. The quantitation of fluorescence intensity suggested BNIP3 deficiency markedly increased mtROS production, and induced mitochondria damage against the protective effect from pinocembrin. d sh-NC or sh-BNIP3-transfected BV2 cells were exposed to IH for 24 h, and then cells were double-stained with annexin V/PI. The proportion of apoptotic cells were detected by flow cytometry. e mRNA levels for different inflammatory cytokine genes (TNF-α, iNOS, COX-2, IL-6, and IL-1β) in shNC, shBNIP3, PIN, and/or 3MA groups. f, g Total cell lysates prepared from treated microglia were separated on SDS-PAGE gel and detected by western blot analysis. Data are representative of at least 3 independent experiments and presented as mean ± SEM in all assays. ^†p < 0.05 vs. the NA + sh-NC group; ^#p < 0.05 vs. the IH + sh-NC group; *p < 0.05 vs. the IH + sh-NC + PIN group; **p < 0.01 vs. the IH + sh-NC + PIN group; ***p < 0.005 vs. the IH + sh-NC + PIN group