Comparison of the effects of aconitine on failing cardiomyocytes before and after combined with glycyrrhetinic acid
Objective To compare the different effects of aconitine on failing cardiomyocytes before and after combined with glycyrrhetinic acid.Methods H9c2 cells were randomized into nine groups,i.e.,blank group,model group,glycyrrhetinic acid group,30,120,and 480 μmol·L-1 aconitine groups,30 μmol·L-1 combination group(30 μmol·L-1 aconitine+30 μmol·L-1 glycyrrhetinic acid),120μmol·L-1 combination group(120 μmol·L-1 aconitine+30 μmol·L-1 glycyrrhetinic acid),and 480 μmol·L-1 combination group(480 μmol·L-1 aconitine+30 μmol·L-1 glycyrrhetinic acid).The model of failing H9c2 cardiomyocytes was established by an 24-hour intervention of 2 μmol·L-1 adriamycin in all groups except the blank group.After successful modeling,the blank and model groups were treated with DMEM,and the remaining groups with the corresponding drugs for 24 h.Cardiomyocyte morphology and mitochondrial microstructure were observed by microscopy,and cell viability was determined by CCK-8 assay.Enzyme immunoassay was used to examine the content of lactate dehydrogenase(LDH),Na+-K+-ATPase,Ca2+-ATPase,Ca2+-Mg2+-ATPase,superoxide dismutase(SOD),malondialdehyde(MDA),catalase(CAT),glutathione(GSH),and ATP.Fluorescent probe assay was employed to check the reactive oxygen species content(ROS),mitochondrial membrane potential,and intracellular and mitochondrial Ca2+concentration.Proteins of AMPK pathway and CaMKⅡ pathway were examined by Western blot.Results Compared with model group,120 and 480 μmol·L-1 aconitine decreased cell viability,increased MDA content(P＜0.05 or P＜0.01),480 μmol·L-1 aconitine disrupted the mitochondrial structure,30,120,and 480 μmol·L-1 aconitine increased ROS content,decreased SOD,CAT,and GSH enzyme activities,decreased mitochondrial membrane potential and ATP production,and increased intracellular and mitochondrial Ca2+concentration,decreased Na+-K+-ATPase,Ca2+-ATPase,and Ca2+-Mg2+-ATPase activities(P＜0.05 or P＜0.01),and 480 μmol·L-1 aconitine inhibited AMPK phosphorylation,decreased SERCA2a and PGC-1α protein expression,and promoted RyR2 protein expression(P＜0.05 or P＜0.01).Compared with aconitine group,the combination of glycyrrhizic acid improved mitochondrial structural damage,reduced ROS content,elevated the activities of SOD,CAT,Na+-K+-ATPase and Ca2+-Mg2+-ATPase,and reduced the intracellular and mitochondrial calcium ion concentration(P＜0.05 or P＜0.01),and 30,120 μmol·L-1 combination groups elevated the mitochondrial membrane potential(P＜0.05 or P＜0.01),the 120,480 μmol·L-1 combination groups decreased MDA content,increased GSH and Ca2+-ATPase activities,and elevated ATP content(P＜0.05 or P＜0.01),and the 480 μmol·L-1 combination group inhibited CaMK Ⅱ phosphorylation and elevated PGC-1α protein expression(P＜0.05 or P＜0.01).Conclusion Glycyrrhetinic acid antagonizes the mitochondrial toxicity of aconitine on the failing cardiomyocytes,which may be related to alleviating oxidative stress and calcium overload,as well as increasing mitochondrial energy production.
万方数据
维普
