摘要
目的:探讨沉默信息调节因子6(silent information regulator 6,SIRT6)对四氯化碳(carbon tetrachloride,CCl_(4))诱导的小鼠肝纤维化作用和机制,以及肝星状细胞在SIRT6沉默后其下游通路的表达变化。方法:将30只雄性C57BL/6J小鼠随机分为正常对照组(n=6)和模型组(造模2、4、8、12周,n=24),采用腹腔注射CCl_(4)制备肝纤维化小鼠模型,每周2次,持续12周。检测各组小鼠血清丙氨酸氨基转移酶(alanine aminotransferase,ALT)、天门冬氨酸氨基转移酶(aspartate aminotransferase,AST)含量评估肝损伤;HE、Masson染色观察小鼠肝脏病理学改变;免疫组织化学染色检测小鼠肝脏α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)的表达;Western blot检测肝脏α-SMA、SIRT6、第9位赖氨酸乙酰化的组蛋白H3(acetyl histone H3 at Lys9,H3K9ac)、第56位赖氨酸乙酰化的组蛋白H3(acetyl histone H3 at Lys56,H3K56ac)、白细胞介素1β(interleukin-1β,IL-1β)和IL-18蛋白的表达。将大鼠肝星状细胞(hepatic stellate cells-T6,HSC-T6)进行SIRT6基因沉默,分为NC siRNA组和SIRT6 siRNA组。Western blot检测SIRT6、H3K9ac和H3K56ac蛋白的表达。结果:与正常对照组比较,模型组小鼠血清中ALT和AST含量均显著升高(P<0.05);HE染色和Masson结果显示,模型组肝脏病理变化逐渐加重,胶原纤维沉积逐渐增多;免疫组织化学染色与Western blot均显示,在8和12周模型组中肝脏α-SMA表达显著增加(P<0.05);Western blot检测结果显示,CCl_(4)造模后各组小鼠肝脏中SIRT6蛋白表达均低于正常对照组(P<0.05),并随肝纤维化的加重逐渐降低;同时模型组小鼠肝脏H3K9ac、H3K56ac、IL-1β和IL-18表达水平在8、12周时明显升高(P<0.05);SIRT6沉默后,与NC siRNA组比较,SIRT6 siRNA组中的H3K9ac和H3K56ac显著增加(P<0.05)。结论:SIRT6缺乏通过H3K9ac和H3K56ac的异常增多使得IL-1β和IL-18的表达升高,加剧炎症反应而促进肝纤维化进程,提示SIRT6的表达异常参与了肝纤维化进
AIM:This study aims to investigate the effects and mechanisms of silent information regulator 6(SIRT6)on carbon tetrachloride(CCl_(4))-induced liver fibrosis in mice,as well as the expression changes in the downstream pathways of hepatic stellate cells after SIRT6 silencing.METHODS:Thirty male C57BL/6J mice were randomly divided into a normal control group(n=6)and a model group(modeling at 2,4,8,12 weeks,n=24).A liver fibrosis model in mice was prepared by intraperitoneal injection of CCl_(4) twice a week for 12 weeks.Serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)levels were measured to assess liver injury.Hematoxylin-eosin(HE)and Masson staining were used to observe the pathological changes in mouse liver tissues.Immunohistochemical staining was conducted to detect the expression ofα-smooth muscle actin(α-SMA)in the liver,Western blot analysis was used to measure the expression of liverα-SMA,SIRT6,acetyl histone H3 at Lys9(H3K9ac),acetyl histone H3 at Lys56(H3K56ac),interleukin-1β(IL-1β),and IL-18 proteins.Hepatic stellate cells-T6(HSC-T6)underwent SIRT6 gene silencing,divided into NC siRNA group and SIRT6 siRNA group,with Western blot performed to detect the expression of SIRT6,H3K9ac,and H3K56ac proteins.RESULTS:Compared with the normal control group,the serum ALT and AST levels in the model group were significantly increased(P<0.05);HE and Masson staining results showed that the pathological changes in the liver of the model group worsened over time,with an increase in collagen fiber deposition.Both immunohistochemistry and Western blot showed that the expression of liverα-SMA significantly increased at 8 and 12 weeks in the model group(P<0.05).Western blot results showed that the expression of SIRT6 protein in the liver of all model group mice was lower than that in the normal control group(P<0.05),and decreased gradually with the progression of liver fibrosis;also,the expression levels of H3K9ac,H3K56ac,IL-1β,and IL-18 in the liver of the model group mice were significantly el
作者
林夏萍
黄仙玉
孙健
李映莹
徐云燕
黄波
杨媛
吴芹
LIN Xiaping;HUANG Xianyu;SUN Jian;LI Yingying;XU Yunyan;HUANG Bo;YANG Yuan;WU Qin(Key Laboratory of the Basic Pharmacology of Ministry of Education and Joint International Research Laboratory of Ethno-medicine of Ministry of Education,Zunyi Medical University,Zunyi 563006,China;Department of Gastroenterology,Af-filiated Hospital of Zunyi Medical University,Zunyi 563003,China)
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2024年第9期1652-1659,共8页
Chinese Journal of Pathophysiology
基金
国家自然科学基金资助项目(No.82060750)。
关键词
肝纤维化
四氯化碳
肝星状细胞
沉默信息调节因子6
炎症因子
liver fibrosis
carbon tetrachloride
hepatic stellate cells
silent information regulator 6
inflammatory factors
