人肺癌细胞中蛋白激酶C亚类与cAMP-PKA系统相关性的研究

STUDIES ON THE RELATIONSHIP BETWEEN PROTEIN KINASE C α ISOFORM AND cAMP PROTEIN KINASE A SYSTEM IN HUMAN LUNG CANCER LTEPa 2 CELLS

为了探讨蛋白激酶Ｃα（ＰＫＣα）及其与环腺苷酸－蛋白激酶Ａ（ｃＡＭＰ－ＰＫＡ）信号系统的相关性，用六甲撑双乙酰胺（ｈｅｘａｍｅｔｈｙｌｅｎｅｂｉｓａｃｅｔａｍｉｄｅ，ＨＭＢＡ）和ＰＫＣ抑制剂ＣａｌｐｈｏｓｔｉｎＣ分别处理人肺癌细胞ＬＴＥＰａ－２，统计细胞生长百分数，测定ｃＡＭＰ水平。用斑点杂交方法检测ＰＫＣα基因表达，并测定表达反义ＰＫＣα的人肺癌ＬＴＥＰａ－２细胞中ＰＫＣ与ＰＫＡ激酶活性。结果显示：５μｍｏｌ／ＬＨＭＢＡ处理人肺癌ＬＴＥＰａ－２细胞３ｄ后，细胞增殖明显被阻抑，ＰＫＣα基因表达下降，ｃＡＭＰ水平升高。０．１５ｍｇ／ＬＰＫＣ抑制剂ＣａｌｐｈｏｓｔｉｎＣ导致人肺癌细胞增殖速度减慢时，ｃＡＭＰ水平也升高。在表达反义ＰＫＣα的人肺癌ＬＴＥＰα－２细胞中，当ＰＫＣα基因和蛋白表达被阻抑、ＰＫＣ活性明显降低的同时，ＰＫＡ活性则显著升高。结果提示：在人肺癌细胞中ＰＫＣα亚类和ｃＡＭＰ－ＰＫＡ通路显示出拮抗关系。ＰＫＣα和ＰＫＡ两套信号系统相互作用。

To study the relationship between protein kinase Cα isoform and cAMP protein kinase A system. Treating human lung cancer cells LTEPa 2 with HMBA(hexamethylene bisacetamide) and PKC inhibitor Calphostin C respectively, counting growth rate and detecting the level of cAMP, analysing the expression of PKCα gene using Dot blot in treated human lung cancer LTEPa 2 cells and assying the activity of PKC and PKA in LTEPa 2 cells expressing antisense PKCα.The results demonstrated that human lung cancer LTEPa 2 cells showed reduced growth rate and decreased expression of PKCα gene after treatment with 5μmol/L HMBA,on the contrary,cAMP level increased.When PKA activity was inhibited in LTEPa 2 cells treated with 0 15mg/L Calphostin C,a specific inhibitor of PKC,cAMP level increased in LTEPa 2 cells with slow growth rate.The PKA activity increased obviously in LTEPa 2 cells expressing antisense PKCα mRNA,which displayed markedly decreased PKCα level(RNA,protein content and enzymatic activiey).The results indicated that an antagonistical relationship between PKCα and cAMP PKA pathway existed in LTEPa 2 cells.These two signaling systems regulate proliferation and transformation of human lung cancer cells cooperatively.