舌鳞癌细胞Tca8113中蛋白激酶C和丝裂原活化蛋白激酶激酶/细胞外调节蛋白激酶通路对诱导型一氧化氮合酶表达的影响  

Effects of protein kinase C and motigen-activated protein kinase kinase/extracellular regulated protein kinases signaling pathway on mRNA level of inducible nitric oxide synthase in Tca8113 cells

作　　者：高雪峰[1] 焦海斌 叶昌成 刘英群[1] Gao Xuefeng, diao Haibin, Ye Changcheng, Liu Yingqun(The Affiliated Stomatological Hospital of Harbin Medical University, Harbin 150001, Chin)

机构地区：[1]哈尔滨医科大学附属口腔医院,哈尔滨150001

出　　处：《华西口腔医学杂志》2018年第2期133-139,共7页West China Journal of Stomatology

摘　　要：目的在舌鳞癌细胞Tca8113中,探索与细胞增殖密切相关的诱导型一氧化氮合酶(NOS-2)的表达调控机制。方法采用RNAi技术沉默Tca8113细胞中NOS-2、蛋白激酶C(PKC)-α、PKC-β和PKC-δ的基因;GriessReagent法检测NOS-2基因沉默后一氧化氮(NO)生成量;CCK8法测定细胞增殖活性;实时定量荧光聚合酶链反应(qPCR)技术检测各种方法处理后NOS-2、PKC-α、PKC-β和PKC-δ的基因表达;Westernblotting技术测定丙二醇甲醚醋酸酯(PMA)作用细胞后的细胞外调节蛋白激酶(ERK)1/2磷酸化程度。结果利用NOS-2的siRNA处理后,Tca8113细胞的增殖能力明显降低(P<0.01);PKC的活性与NOS-2的基因表达成负相关(P<0.05);PKC亚型PKC-α、PKC-β和PKC-δ共同参与NOS-2的基因调控(P<0.01);丝裂原活化蛋白激酶激酶(MEK)/ERK通路负调控NOS-2的基因表达(P<0.05);PKC通过MEK/ERK通路负调控NOS-2的基因表达(P<0.05)。结论在Tca8113细胞中,PKC通过MEK/ERK通路负调控与细胞增殖相关的NOS-2的基因表达。Objective To explore the regulatory mechanism of inducible nitric oxide synthase （NOS-2） expression related to proliferation of Tca8113 cells. Methods RNAi mediated by short hairpin RNAs was utilized to knock down NOS-2, protein kinase C （PKC）-α, PKC-β and PKC-δ. Griess Reagent played a significant role on the detection of NO product after NOS-2 silence. The cell proliferation was determined by CCK8 method. Quantitative real-time polymerase chain reaction （q-PCR） was recruited to check the mRNA level of NOS-2, PKC-α, PKC-β and PKC-δ after treated by a variety of ways. Eventually, the measure of phosphorylation of extracellular regulated protein kinases （ERK）1/2 was performed by Western blotting in PMA-treated Tca8113 cells. Results The cell viability of Tca8113 decreased obviously after transfected with NOS-2 siRNA （P〈0.01）. PKC reduced the expression level of NOS-2 mRNA （P〈0.05）. PKC-α, PKC-β and PKC-δ worked together to regulate the level of NOS-2 mRNA （P〈0.01）. Motigen-activated protein kinase kinase （MEK）/ERK signaling pathway regulated the level of NOS-2 mRNA negatively （P〈0.05）. PKC down regulated the level of NOS-2 mRNA through MEK/ERK signaling pathway （P〈0.05）. Conclusion PKC regulates the mRNA level of NOS-2 related to proliferation through MEK/ERK signaling pathway in Tca8113 cells..

关 键 词：诱导型一氧化氮合酶 蛋白激酶C 丝裂原活化蛋白激酶激酶/细胞外调节蛋白激酶通路 细胞增殖 

分 类 号：Q257[生物学—细胞生物学]