p27、cyclin D1和cyclin E在膀胱移行细胞癌中的表达及其意义
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摘要
目的 ：研究P27、cyclin D1和cyclin E蛋白在膀胱移行细胞癌中的表达及意义。
方法： 采用免疫组织化学方法检测65例膀胱移行细胞癌的石蜡标本、10例膀胱粘膜非典型增生、8例正常膀胱组织中P27、cyclin D1和cyclin E的表达。
结果： 正常膀胱组织p27阳性表达率为100%，非典型增生标本中p27阳性表达率为60%，低于正常膀胱组织(P<0.05)；膀胱癌组织中p27阳性表达率仅为51%，明显低于正常膀胱组织(P<0.01)。p27阳性表达率在G1、G2、G3膀胱癌中分别为75%，53.8%，26.3%，三组间相比较，差别均有显著性意义(P<0.05)。G1组与G3组比较有显著性意义。Ta-T1、T2-T4两组中，p27的阳性表达率分别为72.9%、25%，两组比较有显著性意义(P<0.01)。 cyclin E在非典型增生组织中阳性表达率为60%，明显高于正常膀胱组织的25%，但统计学上无显著性意义。在膀胱癌G1、G2、G3三组中cyclin E的阳性表达率分别为80%、42.3%、31.6%，三组比较有显著性意义(X2=10.38 P<0.01)，其中G1组cyclin E的阳性表达率明显高于G2、G3组(P<0.01)。在Ta-T1、T2-T4两组中，cyclin E的阳性表达率分别为
62.2%、35.7%，两组比较有显著性意义(P<0.05)。cyclin D1在正常膀胱组织中没有表达，在非典型增生组织中阳性表达率为40%，在肿瘤组织中cyclin D1阳性表达率为38.5%(P<0.05)，两者明显高于正常组织。在G1、G2、G3组膀胱癌中cyclin D1阳性表达率分别为65%、34.6%、18.6%，三组间相比较有显著性意义(P<0.01)。G1组cyclin D1的阳性表达率明显高于G2、G3组(P<0.01)。 在Ta-T1、T2-T4两组中，cyclin D1的阳性表达率分54.1%、17.5%，两组比较有显著性意义(P<0.01)。P27、cyclin D1和cyclin E两两相关性分析显示呈正相关。
结论：本试验表明：P27蛋白表达下降或缺失及cyclin D1和cyclin E过表达可能加速细胞周期转化，参与膀胱癌的发生。cyclin D1过表达可能是膀胱癌形成过程中较早的分子事件。随着膀胱移行细胞癌的病理分级、临床分期增加，P27、cyclin D1和cyclin E而呈下降趋势。检测P27、cyclin D1和cyclin E在膀胱癌组织中的表达，有助于估计肿瘤的恶性程度和判断患者的预后。
Objective To study the expression and it's significance of P27,cyclin D1 and cyclin E protein in transitional cell carcinoma of bladder(TCCB).
Methods Expression of P27 cyclin D1 and cyclin E protein were investigated by immunohistochemistry technique (SP) with 65 cases of transitional cell carcinoma of the bladder, 10 atypical hyperplasia cases and 8 normal bladder cases.
Results The postive immunostaining rate of P27 in normal bladder (100%) was significantly higher than in the atypical hyperplasia and significantly higher than in the TCCB (51%). The postive immunostaining rate of P27 was 75% of G1, 53.8% of G2, 26.3% of G3,72.9% of Ta-T1 and 25% of T2-T4.There was significant difference either among G1,G2,G3(P<0.05) or between Ta-T1 and T2-T4(P<0.01). cyclin E in the atypical hyperplasia(60%) was insignificantly higher than in the normal bladder tissue(25%). The expression rate of cyclin E was 80% of G1,42.3% of G2,31.6% of G3,62.2% of Ta-T1 and 35.7% of T2-T4. There was significant difference either among G1, G2, G3 (P<0.05) or between Ta-T1 and T2-T4 (P<0.01). The expression rate of
G1 was significantly higher than G2 and G3, The expression of cyclin D1 in the normal bladder tissue was negative. The expression rate of cyclin D1 in TCCB (38.5%) and in the atypical hyperplasia(40%)was significantly higher than that in the normal bladder tissue. The expression rate of cyclin D1 was 65% of G1, 34.6% of G2, 18.6% of G3,54.1% of Ta-T1 and 17.5% of T2-T4. There was significant difference either among G1, G2, G3 (P<0.01) or between Ta-T1 and T2-T4 (P<0.01). The expression rate of cyclin D1 of G1 was significantly higher than that of G2 and G3. P27, cyclin D1 and cyclin E were correlated with each other.
Conclusions These findings suggest that the down or negative-expression of P27 and overexpression of cyclin D1 and cyclin E might accelerate the progression of the cell cycle and promote the carcinogenesis of the TCCB.The overexpression of cyclin D1 might be an relatively earlier event in the initition of the TCCB. With malignance of tumor increasing, the expression rate of them was decreasing. Analysis of P27、cyclin D1 and cyclin E expression in TCCB was useful for us to estimate the malignance and prognosis.
方法： 采用免疫组织化学方法检测65例膀胱移行细胞癌的石蜡标本、10例膀胱粘膜非典型增生、8例正常膀胱组织中P27、cyclin D1和cyclin E的表达。
结果： 正常膀胱组织p27阳性表达率为100%，非典型增生标本中p27阳性表达率为60%，低于正常膀胱组织(P<0.05)；膀胱癌组织中p27阳性表达率仅为51%，明显低于正常膀胱组织(P<0.01)。p27阳性表达率在G1、G2、G3膀胱癌中分别为75%，53.8%，26.3%，三组间相比较，差别均有显著性意义(P<0.05)。G1组与G3组比较有显著性意义。Ta-T1、T2-T4两组中，p27的阳性表达率分别为72.9%、25%，两组比较有显著性意义(P<0.01)。 cyclin E在非典型增生组织中阳性表达率为60%，明显高于正常膀胱组织的25%，但统计学上无显著性意义。在膀胱癌G1、G2、G3三组中cyclin E的阳性表达率分别为80%、42.3%、31.6%，三组比较有显著性意义(X2=10.38 P<0.01)，其中G1组cyclin E的阳性表达率明显高于G2、G3组(P<0.01)。在Ta-T1、T2-T4两组中，cyclin E的阳性表达率分别为
62.2%、35.7%，两组比较有显著性意义(P<0.05)。cyclin D1在正常膀胱组织中没有表达，在非典型增生组织中阳性表达率为40%，在肿瘤组织中cyclin D1阳性表达率为38.5%(P<0.05)，两者明显高于正常组织。在G1、G2、G3组膀胱癌中cyclin D1阳性表达率分别为65%、34.6%、18.6%，三组间相比较有显著性意义(P<0.01)。G1组cyclin D1的阳性表达率明显高于G2、G3组(P<0.01)。 在Ta-T1、T2-T4两组中，cyclin D1的阳性表达率分54.1%、17.5%，两组比较有显著性意义(P<0.01)。P27、cyclin D1和cyclin E两两相关性分析显示呈正相关。
结论：本试验表明：P27蛋白表达下降或缺失及cyclin D1和cyclin E过表达可能加速细胞周期转化，参与膀胱癌的发生。cyclin D1过表达可能是膀胱癌形成过程中较早的分子事件。随着膀胱移行细胞癌的病理分级、临床分期增加，P27、cyclin D1和cyclin E而呈下降趋势。检测P27、cyclin D1和cyclin E在膀胱癌组织中的表达，有助于估计肿瘤的恶性程度和判断患者的预后。
Objective To study the expression and it's significance of P27,cyclin D1 and cyclin E protein in transitional cell carcinoma of bladder(TCCB).
Methods Expression of P27 cyclin D1 and cyclin E protein were investigated by immunohistochemistry technique (SP) with 65 cases of transitional cell carcinoma of the bladder, 10 atypical hyperplasia cases and 8 normal bladder cases.
Results The postive immunostaining rate of P27 in normal bladder (100%) was significantly higher than in the atypical hyperplasia and significantly higher than in the TCCB (51%). The postive immunostaining rate of P27 was 75% of G1, 53.8% of G2, 26.3% of G3,72.9% of Ta-T1 and 25% of T2-T4.There was significant difference either among G1,G2,G3(P<0.05) or between Ta-T1 and T2-T4(P<0.01). cyclin E in the atypical hyperplasia(60%) was insignificantly higher than in the normal bladder tissue(25%). The expression rate of cyclin E was 80% of G1,42.3% of G2,31.6% of G3,62.2% of Ta-T1 and 35.7% of T2-T4. There was significant difference either among G1, G2, G3 (P<0.05) or between Ta-T1 and T2-T4 (P<0.01). The expression rate of
G1 was significantly higher than G2 and G3, The expression of cyclin D1 in the normal bladder tissue was negative. The expression rate of cyclin D1 in TCCB (38.5%) and in the atypical hyperplasia(40%)was significantly higher than that in the normal bladder tissue. The expression rate of cyclin D1 was 65% of G1, 34.6% of G2, 18.6% of G3,54.1% of Ta-T1 and 17.5% of T2-T4. There was significant difference either among G1, G2, G3 (P<0.01) or between Ta-T1 and T2-T4 (P<0.01). The expression rate of cyclin D1 of G1 was significantly higher than that of G2 and G3. P27, cyclin D1 and cyclin E were correlated with each other.
Conclusions These findings suggest that the down or negative-expression of P27 and overexpression of cyclin D1 and cyclin E might accelerate the progression of the cell cycle and promote the carcinogenesis of the TCCB.The overexpression of cyclin D1 might be an relatively earlier event in the initition of the TCCB. With malignance of tumor increasing, the expression rate of them was decreasing. Analysis of P27、cyclin D1 and cyclin E expression in TCCB was useful for us to estimate the malignance and prognosis.
引文
[1] Walsh P,Reitik A,Vaughan D,et al.Campbell's Urology,7th Edition Harcourt Publishers Limited,1998:2360-4
[2] Hortwell LH,Kastan MB. Cell cycle control and cancer.Science, 1994,266(5192):1821-1828.
[3] Haas K,Staller P,Geisencetal K. Mutual reguirement of CDK4 and Myc in malignant
transformation:evidence for cyclinD1/CDK4 and p16INK4A as upstream regulators of Myc.Oncogene,1997,15(21):2615-2623.
[4] Polyak K,Lee H,Erdjument-Bromage A,etal.Cloning of P27Kipl,acyclin-deplendent kinase inhibitor and a potential mediator of extracellular antimitogenic signals.Cell,1994,78(1):59-66.
[5] Chen J,Williagham T,Shuford M,etal.Tunor suppression and inhibition of aneuploid cell accumulation in human brain tumor cells by ectopic overexpression of the cyclin-dependent kinase inhibitor P27KIPI.J Clin invest,1996,97(8):1983-1988.
[6] Kamai T. Takagi K, 4 sami H et al. Prognostic significance of P27kipl and ki67 expression in carcinoma of the yenal pelvis and wreter. (3) V int, 2000, 86:14-19.
[7] Del Pizzo JJ. Borkowski A. Jacobs SC. et al loss of cell cycle regulators P27kipl and cyclinE in transitional cell carcinoma of the bladder correlates with tumor grade and patient survival. Am J Pathol, 1999, 155:1129- 1136
[8] Frierson HF,Gaffey MJ,Zukerberg LR,.et al.Immunohistochemical detection and gene amplification of cyclinD1 in mammary infiltrating ductal carcinoma.Mod Pathol 1996;9:723-30.
[9] Toyoshima H, Hunter T. P27 a novel inhibitor of G1 cyclin-cdk protein kinase activity, is related to P21.Cell.1994.78: 67-74.
[10] Luo Y, Hurwitz J. Cell-cycle inhibition by indepent CDK and PCAN-binding domains in p21/cipl. Nature, 1995, 375:159-161.
[11] Pietenpol JA.Assignment of the human P27kip gene to 12P13 and its analysis in leukemias.Cancer Res.1995.55:1206-1210 .
[12] Bullrich F, Maclachlan TK,Sang N,et al.Chromosomal mapping of members of the cdc2 family of protein kinases. Cdk3. cdk6. POSSLRE and PJTALER.and a cdk inhibitor p27kip. To regions involved in human cancer. Cancer Res. 1995,55:1 199.
[13] Millard,-S-S, Yan,-J-S, Nguyen,-H, et al. Enhanced ribosomal association of p27(Kip1) mRNA is a mechanism contributing to accumulation during growth
arrest. J-Biol-Chem. 1997 Mar 14; 272(11): 7093-8.
[14] Aprelikova,-O, Xiong,-Y, Liu,-E-T, et al. Both p16 and p21 families of cyclin-dependent kinase (CDK) inhibitors block the phosphorylation of cyclin-dependent kinases by the CDK-activating kinase. J-Biol-Chem. 1995 Aug 4; 270(31): 18195-7.
[15] Xu,-X, Nakano,-T, Wick,-S, et al. Mechanism of Cdk2/Cyclin E inhibition by p27 and p27 phosphorylation. Biochemistry. 1999 Jul 6; 38(27): 8713-22.
[16] Deng c, Zhang P ,Harper JW,et al .Mice lacking P21 undergo nomal development but are defctive in G1 checkpiont control.Cell 1995, 82;675-684 .
[17] Tsukamoto S,Sakada T,ushijima C,e al.Reduced expression of cell cycle regulator P27(kip1) correlates with a shortened survival in non-small cell lung cancer.Lung Cancer,2001,34(1):83-90.
[18] Nohara T,Ryo T,Iwamoto S,et al.expression of cell-cycle regulator P27 is correlated to the prognosis and ER expression in breast carcinoma patients.Oncology,2001,60(1):94-100.
[19] Haitel A,wiener HG,neudert B,et al .Expression of the cell cycle proteins p21, p27, and pRb in clear cell renal cell carcinoma and their prognostic significance.Urology. 2001 Sep;58(3):477-81
[20] Cheung TH,Lo KW,Yu MM,et al. Aberrant expression of p21(WAF1/CIP1) and p27(KIP1) in cervical carcinoma.Cancer Lett. 2001 Oct 22;172(1):93-8.
[21] Nakavama K,Kineman R,Shiane M,et al. Mice lacking p27(Kip1) display increased body size, multiple organ hyperplasia, retinal dysplasia, and pituitary tumors.Cell. 1996 May 31;85(5):707-20.
[22] Pagano M,Tam SW,Theodoras AM,et al. Role of the ubiquitin pathway in regulating abundance of the cyclin-dependent kinase inhibitor. Science,1995,269(5224):682-685.
[23] Motokura T,Keyomarsi K,Kronenberg HM, et al.Cloning and characterization
of human cyclin D3,a cDNA closely related in sequence to the PRADI/cyclin D1 proto-oncogene.J Biol Chem 1992；267：20412-20415.
[24] Matsushime H,Even ME,Strom DK, et al.Identification and properties of an atypical catalytic subunit(p34PSK-J3/cdk4)for mammaliman D type G1 cyclins.Cell 1992；71：323-334
[25] Quelle DE,Ashmun RA,Shurtleff J, et al.Overexpression of mouse D-type cyclins accelerates G1 phase in rodent fobroblasts. Genes Dve 1993;7：1559-1571.
[26] Wang TC, Cardiff RD,Zurerberg.L,et al. Mammary hyperplasia and carcinoma in MMTV-cyclin D1 transgenic mice(J).nature,1995,369(6482):669-671
[27] Ishikawa T,Furihata M.Ohtsuki Y,et al. Cyclin D1 overexpression related to retinoblastoma protein expression as a prognositic marker in human oesophageal squamous cell carcionoma Br J Cancer 1998;77;92-97.
[28] Maed K,Chung Y.Kang S,et al.Cyclin D1 overexpression and prognosis in colorectal adenocarcinoma,Oncology 1998;55;145-151.
[29] Caputi M. Prognostic role of cyclin D1 in non small cell lung cancer, an imunohistochemical analysis.Eur J Histochem .1997;41;133-138.
[30] Ogawa K, Kimoto N, Asamoto M,et al. Aberrant expression of p27(Kip1) is associated with malignant transformation of the rat urinary bladder epithelium. Carcinoma 2000 Jan;21(1):117-121.
[31] Lee CC, Yamamoto S, Morimura K,et al.Cancer Significance of cyclin D1 overexpression in transitional cell carcinomas of the urinary bladder and its correlation with histopathologic features. 1997 Feb 15;79(4):780-789
[32] Geng Y, Eaton EN, Picon M,et al. Regulation of cyclin E transcription by E2Fs and retinoblastoma protein. Oncogene 1996 Mar 21; 12 (6): 1173-1180.
[33] Koff A,Cross F,Fisher A,Schumacher J et al,Human cyclins E,a new cyclin that intereacts with two members of CDC2 gene family. Cell,1991;257：1217-1228.
[34] Gong J . Threshold expression of cyclin E but not D type cyclins characterizes
normal and tumour cells entering S phase Cell.Prolif, 1995 .28; 337-346.
[35] Akama Y. Frequent amplification of the cyclin E gene in human gastric carcinomas .JPn J.Cancer Res , 1995;86：617-662.
[36] Ohtsubo,M. Human cyclin E, a nuclear protein essential for the G1-to-S phase transition. Mol Cell Biol, 1995. 15.2612-2624.
[37] Lundberg AS,Weinberg RA.Functional inactivation of the retinoblastoma protein requires sequential modification by a least two distinct cyclin-cdk complexes. J .Mol Cell Biol.1998；18：753-756.
[38] Leng X, Connell-Crowley L, Goodrich D,et al. S-Phase entry upon ectopic expression of G1 cyclin-dependent kinases in the absence of retinoblastoma protein phosphorylation. Curr Biol 1997 Sep 1; 7(9):709-712.
[39] Santoni-Rugiu E, Falck J, Mailand N,et al.Involvement of Myc activity in a G(1)/S-promoting mechanism parallel to the pRb/E2F pathway. Mol Cell Biol 2000 May;20(10):3497-3509.
[40] Sakaguchi T, Watanabe A, Sawada H,et al.Prognostic value of cyclin E and p53 expression in gastric carcinoma. Cancer 1998 Apr 1;82 (7):1238-1243.
[41] Sutter T, Doi S, Carnevale KA, et al.Expression of cyclins D1 and E in human colon adenocarcin J Med 1997;28(5-6):285-309.
[42] Scott KA,Walker RA.Lack of cyclin E immunoreactivity in non-malignant breast and association with proliferation in breast cancer. Br J Cancer 1997;76 (10):1288-1292.
[43] Sui L, Dong Y, Ohno M, et al. Implication of malignancy and prognosis of p27 (kip1), Cyclin E, and Cdk2 expression in epithelial ovarian tumors. Gynecol Oncol 2001 Oct; 83 (1): 56-63.
[44] Del Pizzo JJ. Borkowski A. Jacobs sc. et al loss of cell cycle regulators P27kipl and cyclinE in transitional cell carcinoma of the bladder correlates with tumor grade and patient survival. Am J Pathol, 1999, 155:1129-1136.
[2] Hortwell LH,Kastan MB. Cell cycle control and cancer.Science, 1994,266(5192):1821-1828.
[3] Haas K,Staller P,Geisencetal K. Mutual reguirement of CDK4 and Myc in malignant
transformation:evidence for cyclinD1/CDK4 and p16INK4A as upstream regulators of Myc.Oncogene,1997,15(21):2615-2623.
[4] Polyak K,Lee H,Erdjument-Bromage A,etal.Cloning of P27Kipl,acyclin-deplendent kinase inhibitor and a potential mediator of extracellular antimitogenic signals.Cell,1994,78(1):59-66.
[5] Chen J,Williagham T,Shuford M,etal.Tunor suppression and inhibition of aneuploid cell accumulation in human brain tumor cells by ectopic overexpression of the cyclin-dependent kinase inhibitor P27KIPI.J Clin invest,1996,97(8):1983-1988.
[6] Kamai T. Takagi K, 4 sami H et al. Prognostic significance of P27kipl and ki67 expression in carcinoma of the yenal pelvis and wreter. (3) V int, 2000, 86:14-19.
[7] Del Pizzo JJ. Borkowski A. Jacobs SC. et al loss of cell cycle regulators P27kipl and cyclinE in transitional cell carcinoma of the bladder correlates with tumor grade and patient survival. Am J Pathol, 1999, 155:1129- 1136
[8] Frierson HF,Gaffey MJ,Zukerberg LR,.et al.Immunohistochemical detection and gene amplification of cyclinD1 in mammary infiltrating ductal carcinoma.Mod Pathol 1996;9:723-30.
[9] Toyoshima H, Hunter T. P27 a novel inhibitor of G1 cyclin-cdk protein kinase activity, is related to P21.Cell.1994.78: 67-74.
[10] Luo Y, Hurwitz J. Cell-cycle inhibition by indepent CDK and PCAN-binding domains in p21/cipl. Nature, 1995, 375:159-161.
[11] Pietenpol JA.Assignment of the human P27kip gene to 12P13 and its analysis in leukemias.Cancer Res.1995.55:1206-1210 .
[12] Bullrich F, Maclachlan TK,Sang N,et al.Chromosomal mapping of members of the cdc2 family of protein kinases. Cdk3. cdk6. POSSLRE and PJTALER.and a cdk inhibitor p27kip. To regions involved in human cancer. Cancer Res. 1995,55:1 199.
[13] Millard,-S-S, Yan,-J-S, Nguyen,-H, et al. Enhanced ribosomal association of p27(Kip1) mRNA is a mechanism contributing to accumulation during growth
arrest. J-Biol-Chem. 1997 Mar 14; 272(11): 7093-8.
[14] Aprelikova,-O, Xiong,-Y, Liu,-E-T, et al. Both p16 and p21 families of cyclin-dependent kinase (CDK) inhibitors block the phosphorylation of cyclin-dependent kinases by the CDK-activating kinase. J-Biol-Chem. 1995 Aug 4; 270(31): 18195-7.
[15] Xu,-X, Nakano,-T, Wick,-S, et al. Mechanism of Cdk2/Cyclin E inhibition by p27 and p27 phosphorylation. Biochemistry. 1999 Jul 6; 38(27): 8713-22.
[16] Deng c, Zhang P ,Harper JW,et al .Mice lacking P21 undergo nomal development but are defctive in G1 checkpiont control.Cell 1995, 82;675-684 .
[17] Tsukamoto S,Sakada T,ushijima C,e al.Reduced expression of cell cycle regulator P27(kip1) correlates with a shortened survival in non-small cell lung cancer.Lung Cancer,2001,34(1):83-90.
[18] Nohara T,Ryo T,Iwamoto S,et al.expression of cell-cycle regulator P27 is correlated to the prognosis and ER expression in breast carcinoma patients.Oncology,2001,60(1):94-100.
[19] Haitel A,wiener HG,neudert B,et al .Expression of the cell cycle proteins p21, p27, and pRb in clear cell renal cell carcinoma and their prognostic significance.Urology. 2001 Sep;58(3):477-81
[20] Cheung TH,Lo KW,Yu MM,et al. Aberrant expression of p21(WAF1/CIP1) and p27(KIP1) in cervical carcinoma.Cancer Lett. 2001 Oct 22;172(1):93-8.
[21] Nakavama K,Kineman R,Shiane M,et al. Mice lacking p27(Kip1) display increased body size, multiple organ hyperplasia, retinal dysplasia, and pituitary tumors.Cell. 1996 May 31;85(5):707-20.
[22] Pagano M,Tam SW,Theodoras AM,et al. Role of the ubiquitin pathway in regulating abundance of the cyclin-dependent kinase inhibitor. Science,1995,269(5224):682-685.
[23] Motokura T,Keyomarsi K,Kronenberg HM, et al.Cloning and characterization
of human cyclin D3,a cDNA closely related in sequence to the PRADI/cyclin D1 proto-oncogene.J Biol Chem 1992；267：20412-20415.
[24] Matsushime H,Even ME,Strom DK, et al.Identification and properties of an atypical catalytic subunit(p34PSK-J3/cdk4)for mammaliman D type G1 cyclins.Cell 1992；71：323-334
[25] Quelle DE,Ashmun RA,Shurtleff J, et al.Overexpression of mouse D-type cyclins accelerates G1 phase in rodent fobroblasts. Genes Dve 1993;7：1559-1571.
[26] Wang TC, Cardiff RD,Zurerberg.L,et al. Mammary hyperplasia and carcinoma in MMTV-cyclin D1 transgenic mice(J).nature,1995,369(6482):669-671
[27] Ishikawa T,Furihata M.Ohtsuki Y,et al. Cyclin D1 overexpression related to retinoblastoma protein expression as a prognositic marker in human oesophageal squamous cell carcionoma Br J Cancer 1998;77;92-97.
[28] Maed K,Chung Y.Kang S,et al.Cyclin D1 overexpression and prognosis in colorectal adenocarcinoma,Oncology 1998;55;145-151.
[29] Caputi M. Prognostic role of cyclin D1 in non small cell lung cancer, an imunohistochemical analysis.Eur J Histochem .1997;41;133-138.
[30] Ogawa K, Kimoto N, Asamoto M,et al. Aberrant expression of p27(Kip1) is associated with malignant transformation of the rat urinary bladder epithelium. Carcinoma 2000 Jan;21(1):117-121.
[31] Lee CC, Yamamoto S, Morimura K,et al.Cancer Significance of cyclin D1 overexpression in transitional cell carcinomas of the urinary bladder and its correlation with histopathologic features. 1997 Feb 15;79(4):780-789
[32] Geng Y, Eaton EN, Picon M,et al. Regulation of cyclin E transcription by E2Fs and retinoblastoma protein. Oncogene 1996 Mar 21; 12 (6): 1173-1180.
[33] Koff A,Cross F,Fisher A,Schumacher J et al,Human cyclins E,a new cyclin that intereacts with two members of CDC2 gene family. Cell,1991;257：1217-1228.
[34] Gong J . Threshold expression of cyclin E but not D type cyclins characterizes
normal and tumour cells entering S phase Cell.Prolif, 1995 .28; 337-346.
[35] Akama Y. Frequent amplification of the cyclin E gene in human gastric carcinomas .JPn J.Cancer Res , 1995;86：617-662.
[36] Ohtsubo,M. Human cyclin E, a nuclear protein essential for the G1-to-S phase transition. Mol Cell Biol, 1995. 15.2612-2624.
[37] Lundberg AS,Weinberg RA.Functional inactivation of the retinoblastoma protein requires sequential modification by a least two distinct cyclin-cdk complexes. J .Mol Cell Biol.1998；18：753-756.
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