Effective fraction of Gualou-Xiebai-Banxia decoction inhibits the apoptosis of myocardial cells induced by ox-LDL via FGF21/FGFR1/?Klotho-FRS2 signal pathway

Ethnopharmacological relevance: The Gualou-Xiebai-Banxia decoction (GXBD), a classical traditional Chinese medicine (TCM) formula, has beneficial effects in turbid phlegm obstruction syndrome, a type of coronary heart disease (CHD). However, the underlying mechanism and effective constituents of GXBD remain elusive. Our previous studies have shown that the effective constituents of GXBD may be enriched in the n-butanol fraction (GXB-N) and water fraction (GXB-W), the targets of which remain unknown. Aim of the study: To investigate whether GXB-N and GXB-W protect myocardial cells (MCs) via fibroblast growth factor 21 (FGF21) signaling and, if so, to elucidate the underlying mechanisms. Furthermore, to investigate the targets of GXB-N and GXB-W as potential therapeutic targets for cardiovascular disease (CVD). Materials and methods: Cell viability and apoptosis were assayed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl2H-tetrazolium bromide (MTT) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays, respectively. The content of FGF21 in the medium was measured using enzyme-linked immunosorbent assay (ELISA). Protein expression was detected using immunofluorescence and western blotting. Results: Apoptosis increased markedly in MCs exposed to oxidized low density lipoprotein (ox-LDL) 100 mu g/mL, with increased expression of FGF21, FGFR1 and ss Klotho, phosphorylation of fibroblast receptor substrate 2 alpha (FRS2 alpha) was suppressed. Following incubation with GXB-N and GXB-W 200 mu g/mL, the expression of FGF21, FGFR1, and ss Klotho and the phosphorylation of FRS2 alpha were increased. Conclusion: Ox-LDL may inhibit the phosphorylation of FRS2 alpha, inducing considerable FGF21 resistance and resulting in MC apoptosis. GXB-N and GXB-W restored and enhanced FGF21 sensitivity in MCs, consequently rescuing cells from ox-LDL-induced apoptosis. The FGF21-FRS2 alpha signal pathway may be part action targets of these two effective fractions of GXBD.