Lee C C, Ichihara T, Yamamoto S, Wanibuchi H, Sugimura K, Wada S, Kishimoto T, Fukushima S
First Department of Pathology and Department of Urology, Osaka City University Medical School, 1-4-3 Asahi-machi, Abeno-ku, Osaka 545-8585, Japan.
Carcinogenesis. 1999 Sep;20(9):1697-708. doi: 10.1093/carcin/20.9.1697.
The cyclin-dependent kinase (CDK) inhibitor p27(KIP1) exerts its growth suppressive effects by targeting the cyclin-CDK complexes. Reduced protein levels of p27(KIP1) have been reported in numerous human cancers and this has been attributed to increased degradation. However, few reports have addressed the significance of p27(KIP1) expression in chemical carcinogenesis of rodents. In a rat two-stage urinary bladder carcinogenesis model, with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) initiation followed by promotion with sodium L-ascorbate (Na-AsA), we evaluated the expression of p27(KIP1) protein using immunohistochemistry during various stages of urinary bladder carcinogenesis. In addition, we evaluated the mRNA expression profiles for p27(KIP1), p21(WAF1/Cip1) and p53 in tumors. Fisher 344 rats were initiated with 0.05% BBN in the drinking water for 4 weeks and then administered 5% Na-AsA in the diet. Immunohistochemical examination revealed p27(KIP1) protein to be constitutively expressed in normal urothelium, simple hyperplasia and in most papillary and nodular (PN) hyperplasias and small papillomas, but diminished or absent in large papillomas and in transitional cell carcinomas. An inverse correlation between expression of p27(KIP1) and cell proliferation was generally observed. Quantitation of mRNA by multiplex reverse transcription-PCR showed a significant downregulaton of p27(KIP1), p21(WAF1/Cip1) and p53 mRNA in tumors. More than 50% reduction in p27(KIP1) mRNA expression was observed in 42 and 47% of tumors at weeks 18 and 24, respectively; similar reduction in p21(WAF1/Cip1) mRNA expression was observed in 58 and 73% of tumors at weeks 18 and 24, and in p53 mRNA expression in 50 and 73% of tumors at weeks 18 and 24, respectively. None of the 25 tumors we examined by PCR-single-strand conformational polymorphism analysis had p53 mutations. These data imply that abnormal down-regulation of p27(KIP1), p21(WAF1/Cip1) and/or p53 in tumor cells may contribute to the malignant progression of tumors during rat two-stage bladder carcinogenesis.
细胞周期蛋白依赖性激酶（CDK）抑制剂p27（KIP1）通过作用于细胞周期蛋白 - CDK复合物发挥其生长抑制作用。在许多人类癌症中都报道了p27（KIP1）蛋白水平降低，这被认为是由于降解增加所致。然而，很少有报道探讨p27（KIP1）表达在啮齿动物化学致癌过程中的意义。在大鼠膀胱两步致癌模型中，先用N - 丁基 - N -（4 - 羟基丁基）亚硝胺（BBN）启动，然后用L - 抗坏血酸钠（Na - AsA）促进，我们在膀胱致癌的各个阶段使用免疫组织化学评估p27（KIP1）蛋白的表达。此外，我们评估了肿瘤中p27（KIP1）、p21（WAF1 / Cip1）和p53的mRNA表达谱。将Fisher 344大鼠的饮用水中加入0.05%的BBN持续4周，然后在饮食中给予5%的Na - AsA。免疫组织化学检查显示，p27（KIP1）蛋白在正常尿路上皮、单纯增生以及大多数乳头状和结节状（PN）增生及小乳头状瘤中持续表达，但在大乳头状瘤和移行细胞癌中减少或缺失。通常观察到p27（KIP1）表达与细胞增殖之间呈负相关。通过多重逆转录 - PCR对mRNA进行定量分析显示，肿瘤中p27（KIP1）、p21（WAF1 / Cip1）和p53 mRNA显著下调。在第18周和第24周时，分别有42%和47%的肿瘤中p27（KIP1）mRNA表达降低超过50%；在第18周和第24周时，分别有58%和73%的肿瘤中p21（WAF1 / Cip1）mRNA表达有类似降低，以及在第18周和第24周时，分别有50%和73%的肿瘤中p53 mRNA表达降低。我们通过PCR - 单链构象多态性分析检测的25个肿瘤中均未发现p53突变。这些数据表明，肿瘤细胞中p27（KIP1）、p21（WAF1 / Cip1）和/或p53的异常下调可能在大鼠膀胱两步致癌过程中促进肿瘤的恶性进展。
