JMJD3对小鼠肝脏缺血再灌注损伤的影响研究

Study on the effect of JMJD3 on liver ischemia-reperfusion injury in mice

目的探讨含十字形结构域蛋白3(JMJD3)在小鼠肝脏缺血再灌注损伤(IRI)时的作用,并探究抑制JMJD3能否减轻小鼠肝脏IRI。方法建立小鼠IRI模型,Western blot、免疫组织化学检测JMJD3的表达及定位;建立AML-12肝细胞的缺氧复氧损伤模型,Western blot检测JMJD3的蛋白水平变化情况;用ShRNA沉默AML-12肝细胞中的JMJD3,CCK-8检测沉默JMJD3对AML-12细胞活力的影响,同时抑制C-jun氨基末端激酶(JNK)的活性进一步探究JNK-p53信号通路是否调控JMJD3;在小鼠IRI模型中验证GSK-J4预处理抑制JMJD3对肝脏IRI的作用。结果小鼠肝脏IRI能导致JMJD3的表达增加,且JMJD3主要表达于肝细胞中,缺氧复氧损伤也能导致AML-12蛋白水平增加。沉默JMJD3能减轻缺氧复氧损伤导致的AML-12的凋亡,且JMJD3受JNK-p53信号通路调控。在小鼠IRI模型中,GSK-J4预处理能明显减轻IRI 6、24 h后血清中丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)水平,苏木素-伊红(HE)和TUNEL染色发现抑制JMJD3能缓解IRI 6 h病理改变和降低肝细胞凋亡。结论抑制JMJD3能减轻小鼠肝脏IRI,JMJD3受JNK-p53信号通路调控。

Objective To investigate the role of Jumonji domain-containing protein 3(JMJD3)in the mouse liver ischemia reperfusion injury(IRI),and whether inhibition of JMJD3 could alleviate liver IRI.Methods Established the mice IRI model,Western blot and immunohistochemistry were used to detect the expression and location of JMJD3.Established a hypoxia-reoxygenation injury model of AML-12 hepatocytes,Western blot was used to detect changes in the protein level of JMJD3;Silenced JMJD3 in AML-12 cell with ShRNA,CCK-8 was used to detect the effect of silencing JMJD3 on the cell viability of AML-12,while inhibiting the activity of C-jun N-terminal kinase(JNK)to further explore whether the JNK-p53 signaling pathway regulating JMJD3.Verified that GSK-J4 pretreatment inhibited the effect of JMJD3 on liver IRI in the mice IRI model.Results Mice liver IRI increased the expression of JMJD3,and JMJD3 was mainly expressed in liver cells.Hypoxia and reoxygenation injury could also lead to an increase in AML-12 protein levels.Silencing JMJD3 reduced the apoptosis of AML-12 caused by hypoxia and reoxygenation injury,and JMJD3 was regulated by the JNK-p53 signaling pathway.In the mice IRI model,GSK-J4 pretreatment significantly reduced the levels of alanine aminotransferase(ALT)and aspartate aminotransferase(AST)after 6 and 24 hours of IRI,and hematoxylin-eosin(HE)staining and TUNEL staining found that inhibiting JMJD3 could alleviate the pathological changes of IRI 6 h and reduce hepatocyte apoptosis.Conclusion Inhibition of JMJD3 could alleviate mouse liver IRI,JMJD3 is regulated by JNK-p53 pathway.