佛波酯对SMMC-7721人肝癌细胞粘附及整合蛋白基因表达的调控作用

Regulation of PMA on SMMC 7721 Human Hepatocarcinoma Cell Adhesion and Integrin Gene Expression

用促癌剂佛波酯（ＰＭＡ）作用于ＳＭＭＣ－７７２１人肝癌细胞，研究细胞表面的主要粘附分子α５β１整合蛋白基因表达及相应细胞粘附行为的改变．用１００ｎｍｏｌ／ＬＰＭＡ作用ＳＭＭＣ－７７２１人肝癌细胞，发现其作用因时间的长短而异，作用３０、６０、１２０ｍｉｎ分别增加细胞与纤连蛋白（Ｆｎ）粘附１８．８％、３８．７％和５６．６％，作用６、１２ｈ分别降低４４．０％、３７．４％，而不影响与多聚赖氨酸的粘附．使用足量的抗α５和／或抗β１单抗预先封闭细胞与Ｆｎ的结合点，再将细胞与Ｆｎ粘附，发现α５单抗单独使用可将ＳＭＭＣ－７７２１细胞与Ｆｎ的粘附抑制２０％左右，β１单抗则抑制１４％，两者联合使用时可封闭４０％左右的粘附，提示该细胞表面存在除α５β１外的其它整合蛋白在介导着细胞与Ｆｎ的粘附．进一步应用Ｎｏｒｔｈｅｒｎｂｌｏｔ方法，分析整合蛋白基因表达，发现１００ｎｍｏｌ／ＬＰＭＡ抑制α５亚基转录，以３０ｍｉｎ最明显，抑制达８３．１％，作用６、１２ｈ抑制率仍为４６．６％、４３．６％．还就ＰＭＡ影响细胞粘附和整合蛋白基因表达的可能机理作了讨论．

It was studied that the gene expression of integrin α5β1 and the ability of cell adhesion of SMMC 7721 hepatocarcinoma cell were changed in the presence of 100 nmol/L phorbol 12 myristate 13 acetate(PMA).The ability of cell adhesion to extracellular matrix fibronectin(Fn) was increased by 18 8%,38 7% and 56 6% of the control for 30, 60 and 120 min treatment with 100 nmol/L PMA,respectively.But after 6 and 12 hours,it was decreased to 57 4% and 61 1% of the untreated cells,while the adhesion to polylysine was remained the same.Using anti α5 and anti β1 mAb with sufficient amount to pre block the adhesion point,it was found that the adhesion to Fn was inhitited about 40%.It seems that there were other integrins mediating the cell adhesion to Fn besides α5β1.The results obtained from the Northern blot with α5 cDNA probe showed that PMA could downregulate the transcription of integrin α5β1 gene between reaction time of 10 min to 12 hours.The transcription was lowered to 16 9% of the control after 30 min.The inhibition rate was still 43 6% after 12 hours treatment.The possible mechanism by which PMA influences the cell adhesion and integrin gene expression has been discussed.