D-苧烯对人胰腺癌细胞法尼基蛋白转移酶、p21~(ras)膜结合的抑制作用的体外试验

目的探讨ｄ－艹宁烯在化学诱发实验性肿瘤及人实体瘤的防治作用及其机制。方法采用Ｗｅｓｔｅｒｎｂｌｏｔｔｉｎｇ方法观察了ｄ－艹宁烯对胰腺癌细胞中与膜结合的ｐ２１ｒａｓ蛋白表达和法尼基蛋白转移酶活性的抑制作用。同时，采用免疫组化法观察了ｄ－艹宁烯对ｐ２１ｒａｓ在细胞膜上定位的影响。并以Ｎｏｒｔｈｅｒｎｂｌｏｔｉｎｇ方法检测ｄ－艹宁烯对Ｈ－ｒａｓ癌基因在人胰腺癌细胞系中的表达。结果ｄ－艹宁烯对人胰腺癌细胞呈现的抑制作用，其机制可能与其抑制法尼基蛋白转移酶活性，从而降低ｐ２１Ｈ－ｒａｓ蛋白的异戊二烯化修饰有关。ｄ－艹宁烯可降低ｐ２１ｒａｓ蛋白的膜结合，增加胞浆ｐ２１ｒａｓ蛋白的积蓄。在经ｄ－艹宁烯处理过的胰腺癌细胞加入抗ｐ２１ｒａｓ抗体，采用免疫组化法也可观察到上述现象。结论法尼基蛋白转移酶的抑制和ｐ２１ｒａｓ蛋白膜结合与ｐ２１ｒａｓ定位的改变密切相关。ｐ２１ｒａｓ法尼基化的抑制作用改变了它在细胞内的定位，从而影响其生物学活性，但尚未发现与Ｈ－ｒａｓ癌基因表达的密切相关。

Inhibition of farnesyl protein transferase and p21 ras memebrane association by d limonene in human pancreas tumor cells in vitro

Objective Monoterpene d limonene inhibits the plasma membrane associated p21 ras expression and the posttranslational isoprenylation of p21 ras ,a mechanism that may contribute to its efficacy in the chemoprevention and therapy of chemically induced rodent cancers and some human solid tumor cells.Methods In the present study,the relative abilities of d limonene to inhibit membrane associated p21 ras expression in pancreas tumor cell(PaCa) was carried out with Western blotting,and the inhi bition of farnesyl protein transferase (FPTase) activity during the ras protein isoprenylation and cell proliferation were determi ned.Concomitantly,the effects of d limonene on p21 ras localization by immunohistochemistry and H ras oncogene expression in PaCa tumor cell line by Northern blotting were observed.The results showed that d limonene inhibited FPTase activity,thus to reduce p21 H ras isoprenylation.D limonene could decrease p21 ras membrane association and increase cytosolic accumulation of p21 ras .This phenomenon was also noted when d limonene treated PaCa cells were stained immunohistochemically with anti p21 ras antibody.Conclusion It is suggested that the inhibition of FPTase activity was closely related with the inhibition of p21 ras membrane association and the alteration of p21 ras localization.Inhibition of farnesylation of p21 ras altered their intracellular localization and,hence,disrupted their biological activity,but no relationship with H ras oncogene expression was found.