Total Glycosides of Ranunculus Japonius Prevent Hypertrophy in Cardiomyocytes via Alleviating Chronic Ca2+ Overload

ObjectiveTo evaluate the in vitro anti-hypertrophic effect of total Glycosides of Ranunculus Japonius (TGRJ).MethodsNeonatal rat cardiomyocytes were cultured and hypertrophy was induced by administrating isoproterenol (ISO, 10 μmol/L) or angiotensin II (Ang II, 1 μmol/L) for 48 hours. In the treatment groups, cells were pretreated with TGRJ (0.3 g/L) for 30 minutes prior to hypertrophic stimuli. The anti-hypertrophic effects of TGRJ were examined by measuring cell size, total protein content, and protein synthesis. Intracellular free Ca2+ concentration ([Ca2+]i) was evaluated using fluorescence dye Fura-2/AM. Sacroplasmic/endoplasmic reticulum Ca2+ ATPase 2a (SERCA2a), atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP), and beta-myosin heavy chain (β-MHC) protein expression levels were measured by Western blotting. SERCA2a activity was assayed by p-nitrophenal phosphate disodium salt hexahydrate method.ResultsIncreased cell size, total protein content, and protein synthesis following ISO or Ang II stimulation were significantly inhibited by pretreatment with TGRJ (all P<0.05). This anti-hypertrophic effect of TGRJ was confirmed by its suppressing effect on elevated expression of the three hypertrophic related genetic markers, ANP, BNP, and β-MHC. In addition, TGRJ inhibited ISO or Ang II induced up-regulation of [Ca2+] under chronic but not acute conditions. And ISO or Ang II induced down-regulation of SERCA2a expression and activity was also effectively rectified by TGRJ pretreatment.ConclusionsThe results of present study suggested that TGRJ could prevent ISO or Ang II induced cardiac hypertrophy through improving chronic [Ca2+] disorder, might via normalizing SERCA2a expression and activity.