肿胀激活状态下蛋白激酶C同工酶亚型在胃癌耐药细胞系的亚细胞分布变化

目的　探讨肿胀激活状态下，传统型蛋白激酶C(cPKC)同工酶亚型在胃癌SGC7901及其耐药细胞系SGC7901/VCR的表达、亚细胞分布变化及其意义。方法　采用免疫荧光及免疫印迹方法，观察正常状态及持续低渗灌流不同时间cPKC同工酶亚型的亚细胞分布变化。利用免疫荧光双标记及共聚焦显微镜技术观察PKCα与P-糖蛋白（Pgp）的共表达。结果　在正常状态下，cPKC的4种同工酶亚型在胃癌SGC7901及其耐药细胞SGC7901/VCR细胞质及细胞膜均有表达PKCα在耐药细胞表达呈强阳性，在药敏细胞表达呈阳性，PKCβ

Alteration of subcellular distribution of protein kinase C isoforms in swelling-activated multi-drug-resistant gastric cancer cells and its significance

Objective　To study the alteration of expression and subcellular distribution of classical protein kinase (cPKC) isoforms in gastric cancer cells SGC7901 and their multidrug-resistant cell line SGC7901/VCR under condition of swelling activation and to study the significance of such alterations. Methods　Immuno-fluorescence technique and Wstern Blotting were used to determine the expression and subcellular distribution of cPKC isoforms in gastric cancer cells SGC7901 and its multidrug-resistant cell line SGC7901/VCR under normal condition and during continuous hypotonic perfusion. The co-expression of p-glycoprotein (Pgp) and protein kinase Cα(PKCα) was visualized by immunofluorescence double labeling and laser confocus microscopy. Results　Under the normal condition, the four isoenzymes of cPKC were expressed in both the cell membrane and the nuclei of the gastric cancer cells SGC7901 and their multidrug-resistant cell line SGC7901/VCR; PKCαwas strongly positively expressed in the multidrug-resistant cells and positively expressed in the drug-sensitive cells; PKCβⅠ and PKCβⅡ were positively expressed in both drug-resistant and drug-sensitive cells, and PKCγwas strongly positively expressed in both cells. In drug-resistant cell line, cell-swelling and translocation of PKCαand PKCγwere observed ten minutes after continuous hypotonic perfusion. Thirty minutes after the perfusion, almost all the PKCα was translocated into the cell membrane of SGC7901/VCR, part of the PKCγ was translocated into the nucleus, and the cell volume increased to two to three times as much as before. Sixty minutes later, the subcellular distribution of PKCαand PKCγ and the cell volume returned to normal. In drug-sensitive cells, 10～20 minutes after the continuous hypotonic perfusion nearly all the PKCαwas translocated into the cell membrane, part of PKCγwas translocated into the cell membrane, and the cell volume increased to two to three times as much as before. Forty minutes later, PKCα and PKCγ had basically returned to normal. The subcellular distribution of PKCβⅠ and PKCβⅡremained unchanged in both SGC7901 and SGC7901/VCR. Co-expression of Pgy and PKCα was observed in both drug-sensitive and drug-resistant cells, especially in the former. Conclusion　 Only PKCα and PKCγ isoforms play an important role in the regulation of signal transduction of Pgy and cell volume under continuous perfusion and are related to the expression of PKCα and Pgy. PKCβⅠ and PKCβⅡ may have nothing to do with such processes.