Analysis of the expression of cell cycle regulating genes in gastrointestinal tumors

应用免疫组织化学和Western blotting方法检测细胞周期调控基因G1、CDK2、4、CDK抑制物(CDK4I/pI6)、Rb在正常粘膜、增生性腺瘤、腺瘤和晚期结直肠癌中的表达。在正常粘膜中，每种蛋白仅在腺体底部的少数细胞中检测到。在增生性粘膜中，Cyclin D和E在腺体底部的上部细胞中大量表达。在腺瘤中，细胞周期蛋白D：55/66(83%)、细胞周期蛋白E：42/50(84%)高表达。在腺瘤中，CDK2、p16和Rb以及细胞周期蛋白D、E均有高表达，如细胞周期蛋白D：8/8(100%)、细胞周期蛋白E：13/13(100%)、细胞周期蛋白2：10/12(83%)、p16：10/14(71%)、Rb：10/12(83%)。最后，在晚期癌症中，所有的蛋白都过表达，例如，细胞周期蛋白D：2…More 6/45(58%)，Cyclin E：41/45(91%)，CDK2：34/45(76%)，CDK4：39/45(87%)，p16：59/60(98%)，Rb：34/45(76%)。Western blotting的定量分析进一步证实，这些蛋白在结肠癌中的表达比正常粘膜高几倍。此外，Rb过度磷酸化(失活)的比率是正常粘膜的1.46倍。相邻切片的免疫组织化学对比分析表明，高表达Rb的癌细胞也高表达Cyclin E/CDK2，高表达CDK4的癌细胞也高表达p16。综上所述，在以腺瘤-癌序列为基础的多步结肠癌发生过程中，细胞周期蛋白D和E在增生期或腺瘤期过表达，CDK2和CDK4I/p16在腺瘤期过表达，CDK4在晚期癌期过表达。因此，细胞周期调节因子G1、CDKs和CDK抑制物的表达与结肠癌的发生密切相关。细胞周期蛋白E/CDK2的过度表达可能参与了Rb蛋白的过度磷酸化。较少

The expression of cell cycle regurating genes, eg., positive regulators : G1 cyclins (cyclin D,E), cyclin dependent kinases (CDK2,4), negative regulators : CDK inhibitor (CDK4I/pI6), RB,in normal mucosa, hyperplasia, adenoma, carcinoma in adenoma and advanced carcinoma of colorectum was analyzed by immunohistochemistry and Western blottings. In normal mucosa, each protein was detected only in a few cells of bottom parts of the gland. In hyperplastic mucosa, the expression of cyclin D and E was detected in large number of cells of upper parts from the bottom of the gland. In adenomas, the overexpression of these proteins are frequently detected, eg., cyclin D : 55/66 (83%), cyclin E : 42/50 (84%). In carcinoma in adenoma, the overexpression of CDK2, p16 and RB,as well as cyclin D,E,was noted, eg., cyclin D : 8/8 (100%), cyclin E : 13/13 (100%), CDK2 : 10/12 (83%), p16 : 10/14 (71%), RB : 10/12 (83%). Finally, in advanced carcinomas, all the proteins were overexpressed, eg., cyclin D : 2 … More 6/45 (58%), cyclin E : 41/45 (91%), CDK2 : 34/45 (76%), CDK4 : 39/45 (87%), p16 : 59/60 (98%), RB : 34/45 (76%). The quantitative analysis by Western blottings further confirmed a few times higher expression of these proteins in colon carcinomas compared to normal mucosa. Also, the rate of hyperphosphorylated (inactivated) form of RB was shown to be 1.46 times higher compared to normal mucosa. The comparative analysis using immunohistochemistry on the adjacent sections revealed that the cancer cells overexpressing RB also overexpressed cyclin E/CDK2 and those overexpressing CDK4 also expressed p16. In summary, during the multistep colon carcinogenesis based on adenoma-carcinoma sequence, cyclin D and E are overexpressed from the stage of hyperplasia or adenoma, CDK2 and CDK4I/p16 are overexpressed from the stage of carcinoma in adenoma, and CDK4 is overexpressed at the stage of advanced carcinoma. Thus, it was suggested that the expression of cell cycle regulators such as G1 cyclins, CDKs and CDK inhibitors are deeply involved in colon carcinogenesis. And, the overexpression of cyclin E/CDK2 was thought to be involved in hyperphosphorylation of RB protein. Less