蜕皮甾酮通过抑制MEK/ERK通路减轻小鼠肝脏缺血再灌注损伤OA

AIM:To investigate the protective effect of ecdysterone on mice with hepatic ischemia-reperfusion injury(HIRI)through regulation of mitogen-activated protein kinase kinase(MEK)/extracellular signal-regulated kinase(ERK)signaling pathway,and to elucidate its underlying mechanism in inhibiting hepatocyte apoptosis.METHODS:(1)A mouse liver organoid hypoxia/reoxygenation(H/R)model was established.Organoids were divided into control,model,low-and high-dose ecdysterone,U0126,and high-dose ecdysterone combined with U0126 groups.Hematoxylin-eosin(HE)staining was used to observe morphological changes,and cell viability was assessed using a cell viability assay kit.Immunofluorescence was performed to detect cleaved caspase-3 expression to evaluate apoptosis.(2)A 70%hepatic ischemia-reperfusion model was established in mice.Male C57BL/6J mice were randomly divided into control,model,low-and high-dose ecdysterone,U0126,and high-dose ecdysterone combined with U0126 groups(n=6).After 24 h of re-perfusion,serum alanine aminotransferase(ALT),aspartate aminotransferase(AST),tumor necrosis factor-α(TNF-α),and interleukin-6(IL-6)levels were measured.Liver injury was evaluated by HE staining combined with Suzuki scoring.Apoptosis was detected using TUNEL assay.The expression of apoptosis-related proteins and MEK/ERK pathway compo-nents was analyzed by Western blot.RESULTS:The H/R treatment caused structural damage to organoids,cell viability reduction,and cleaved caspase-3 expression increase.Compared with model group,ecdysterone treatment improved or-ganoid morphology,increased cell viability,and reduced cleaved caspase-3 expression.In vivo,compared with control group,serum levels of liver function indicator(ALT and AST)and inflammatory factor(TNF-α and IL-6)in model group were significantly increased(P<0.01),Suzuki scores were significantly elevated(P<0.01),and the proportion of TUNEL-positive cells increased.The levels of phosphorylated MEK,ERK and c-JUN proteins significantly increased(P<0.01),whereas that of B-cell leukemia/lymphoma-2(BCL-2)and BCL-extra-large(BCL-XL)significantly decreased(P<0.01).Compared with model group,ecdysterone at different doses in combination with U0126 significantly reduced serum levels of liver function indicators and inflammatory factors(P<0.01),decreased Suzuki scores(P<0.01),and reduced the pro-portion of TUNEL-positive cells.Meanwhile,BCL-2-associated X protein(BAX)expression was down-regulated,where-as BCL-2 and BCL-XL expression was up-regulated(P<0.01),and the phosphorylation levels of MEK,ERK and c-JUN were reduced(P<0.01).The high-dose ecdysterone and combination treatment groups showed stronger effects.CON-CLUSION:In mouse HIRI and liver organoid H/R models,ecdysterone alleviates liver injury and inflammatory responses and inhibits hepatocyte apoptosis by suppressing abnormal activation of the MEK/ERK signaling pathway,thereby exerting hepatoprotective effects.