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Ethnopharmacological relevance: Smilax glabra, a rhizome of the Liliaceae plant, is a widely used traditional Chinese medicine for anti-cardiac hypertrophy treatment.We have previously found that Smilax glabra flavonoids (SGF) exerted such anti-cardiac hypertrophy activity.However, the mechanism of this activity of SGF has not been clarified yet.Materials and Methods: This study was to investigate the inhibitory role of SGF on intracellular Ca2+ release in rat cardiomyoblast cells (H9C2).Intracellular Ca2+ release was determined by Ca2+ indicator fluorescence (fluo 4-AM) in H9C2 cell line.Results: SGF at concentrations of 0.25, 0.5, 1.0 mg/ml significantly inhibited the phenylephrine or angiotensin II induced intracellular Ca2+ release in a dose-dependent manner.Furthermore, SGF could also inhibit RyR agonist caffeine induced Ca2+ release and PE-induced Ca2+ release under the condition in which IP3 receptors were blocked with 2-APB.Nevertheless, SGF had no impact on PE-induced Ca2+ release under the condition in which RyRs were blocked with tetracaine.Conclusions: Our results suggest that the protective effects of SGF are mediated via targeting inhibition of RyR mediated intracellular Ca2+ release.