中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
Chinese Journal of Experimental Surgery
2015年
10期
2423-2426
,共4页
黄萍%陈明锴%刘仕倩%沈世强
黃萍%陳明鍇%劉仕倩%瀋世彊
황평%진명개%류사천%침세강
跨膜蛋白16A%门脉高压症%血管重构%丝裂原活化蛋白激酶-细胞外信号调节激酶1/2通路
跨膜蛋白16A%門脈高壓癥%血管重構%絲裂原活化蛋白激酶-細胞外信號調節激酶1/2通路
과막단백16A%문맥고압증%혈관중구%사렬원활화단백격매-세포외신호조절격매1/2통로
Transmembrane 16A%Portal hypertension%Vascular remodeling%Mitogen-activated protein kinase-extracellular signal-regulated kinase 1/2 signal pathway
目的 探讨跨膜蛋白16A(TMEM16A)对原代培养大鼠门静脉平滑肌细胞增殖的作用及其机制.方法 原代培养大鼠门静脉平滑肌细胞,慢病毒转染上调其TMEM16A表达,分别加T16Ainh-A01和U0126后用Western blot法检测蛋白表达变化,流式细胞术观察细胞周期变化,细胞计数试剂盒(CCK-8)观察细胞增殖.结果 成功培养出大鼠门静脉平滑肌细胞并转染了TMEM16A,转染效率约为80％,流式细胞术结果显示过表达TMEM16A组S期细胞比例为(34.44±3.72)％,高于对照组的(15.56±2.36)％,差异有统计学意义(t=7.423,P＜0.01).Western blot显示各组细胞外信号调节激酶1/2(ERK1/2)表达水平差异无统计学意义(t=2.338,P＞0.05).各组磷酸化ERK1/2(p-ERK1/2)表达差异均有统计学意义(P＜0.05).结论 TMEM16A可能通过影响丝裂原活化蛋白激酶(MAPK)-ERK1/2通路活性来调控门静脉平滑肌细胞增殖,其表达在原代培养的大鼠门静脉平滑肌细胞增殖中起重要作用.
目的 探討跨膜蛋白16A(TMEM16A)對原代培養大鼠門靜脈平滑肌細胞增殖的作用及其機製.方法 原代培養大鼠門靜脈平滑肌細胞,慢病毒轉染上調其TMEM16A錶達,分彆加T16Ainh-A01和U0126後用Western blot法檢測蛋白錶達變化,流式細胞術觀察細胞週期變化,細胞計數試劑盒(CCK-8)觀察細胞增殖.結果 成功培養齣大鼠門靜脈平滑肌細胞併轉染瞭TMEM16A,轉染效率約為80％,流式細胞術結果顯示過錶達TMEM16A組S期細胞比例為(34.44±3.72)％,高于對照組的(15.56±2.36)％,差異有統計學意義(t=7.423,P＜0.01).Western blot顯示各組細胞外信號調節激酶1/2(ERK1/2)錶達水平差異無統計學意義(t=2.338,P＞0.05).各組燐痠化ERK1/2(p-ERK1/2)錶達差異均有統計學意義(P＜0.05).結論 TMEM16A可能通過影響絲裂原活化蛋白激酶(MAPK)-ERK1/2通路活性來調控門靜脈平滑肌細胞增殖,其錶達在原代培養的大鼠門靜脈平滑肌細胞增殖中起重要作用.
목적 탐토과막단백16A(TMEM16A)대원대배양대서문정맥평활기세포증식적작용급기궤제.방법 원대배양대서문정맥평활기세포,만병독전염상조기TMEM16A표체,분별가T16Ainh-A01화U0126후용Western blot법검측단백표체변화,류식세포술관찰세포주기변화,세포계수시제합(CCK-8)관찰세포증식.결과 성공배양출대서문정맥평활기세포병전염료TMEM16A,전염효솔약위80％,류식세포술결과현시과표체TMEM16A조S기세포비례위(34.44±3.72)％,고우대조조적(15.56±2.36)％,차이유통계학의의(t=7.423,P＜0.01).Western blot현시각조세포외신호조절격매1/2(ERK1/2)표체수평차이무통계학의의(t=2.338,P＞0.05).각조린산화ERK1/2(p-ERK1/2)표체차이균유통계학의의(P＜0.05).결론 TMEM16A가능통과영향사렬원활화단백격매(MAPK)-ERK1/2통로활성래조공문정맥평활기세포증식,기표체재원대배양적대서문정맥평활기세포증식중기중요작용.
Objective To investigate the effectof transmembrane protein 16(TMEM16A) on proliferation of primary cultured smooth muscle cellof raportal vein (rPVSMCs) and the mechanism.MethodThe primary rPVSMCwere cultured, and transfected with lentiviruto investigate the effecof TMEM16on proliferation of rPVSMCs.T16Ainh-A01 and U0126 were used in both control group and lentivirutransfection group, respectively.Western blotting analysiwaused to detecthe expression of TMEM16A, phosphorylated extracellulasignal-regulated kinase 1/2 (p-ERK1/2) and extracellulasignal-regulated kinase 1/2 (ERK1/2) in rPVSMCs.Flow cytometry and cell counting kit-8 (CCK-8) were used to measure the cell cycle and proliferation of rPVSMCs.ResultWe successfully cultured the rPVSMCand up-regulated the expression of TMEM16A.Flow cytometry showed thathe cell ratio of phase [(35.44 ±3.72)％] in the over-expressed TMEM16group wasignificantly highethan in the control group [(15.56 ±2.36)％ ,P ＜0.05].Western blotting showed thaERK1/2 waexpressioned stablely in 7 group(P ＞ 0.05).There were significandifferencein p-ERK1/2 among groups.Conclusion TMEM16may regulate the proliferation of rPVSMCby influencing the activity of mitogen-activated protein kinase (MAPK)-ERK1/2 signal pathway, and itexpression may play an importanrole in the proliferation of primary cultured rPVSMCs.
