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STAT3、p27和CyclinD1在皮肤恶性黑色素瘤中的表达
Expression of STAT3、p27and CyclinD1in Cutaneous Malignant Melanoma
【作者】 温立霞;
【导师】 陈旻静;
【作者基本信息】 郑州大学 , 外科学(专业学位), 2013, 硕士
【摘要】 背景及目的皮肤恶性黑色素瘤(cutaneous malignant melanoma,CMM)是起源于皮肤黑色素细胞的常见体表恶性肿瘤之一,是增长最快的皮肤恶性肿瘤。其发生发展同人类其他肿瘤一样,是多种因素、多个步骤及阶段共同作用的极为复杂的过程,主要包含癌基因的激活、抑癌基因活性减弱或消失,最终引起细胞周期的改变,导致癌细胞生长失控。近年来肿瘤的基因治疗为肿瘤的治疗带来了新的挑战和机遇,尽管对癌基因及抑癌基因具体作用机制的认识尚存在许多不足之处,但将癌基因和抑癌基因作为突破口的基因治疗是肿瘤治疗中最根本、最有前途的方法之一。STAT3是信号传导子与转录激活子家族(signal transducers and activators of transcription, STATS)的重要成员,一般驻留在细胞质中,当被上游信号通路激活(如:EGFR, c-Met, IL-6受体激活等)后,STAT3磷酸化形成二聚体或与STATS的其他成员形成异质二聚体。激活的STAT3复合物移位到细胞核中可开始STAT3目的基因(包括CyclinD1[1]、 Bcl-xL[2]、c-myc[1]、and VEGF[3]等)的转录。STAT3通过促进细胞周期素D1(CyclinDl)的表达,促使细胞无限增殖,进而促进组织细胞的癌变。细胞周期的改变与癌细胞的生长失控密不可分。细胞周期素依赖性激酶抑制因子(CDK inhibitor, CDKI)可以抑制细胞周期素(Cyclin)与细胞周期素依赖性激酶(Cyclin dependent kinases,CDK)复合物的形成,CDKI的失活导致细胞无限增殖。作为CDKI家族的常见成员之一,p27可以抑制CyclinDl/CDK4与CyclinE/CDK2复合物的活性,从而使细胞停滞于G1期,干扰细胞的无限增殖,从而抑制组织细胞的癌变。为了解STAT3、p27、CyclinD1在皮肤恶性黑色素瘤发病机制中的作用,本研究检测了54例皮肤恶性黑色素瘤、22例交界痣、20例正常皮肤组织中STAT3、p27、CyclinD1的表达,并分析STAT3、p27、CyclinD1三者的相关性。材料与方法选取郑州大学第一附属医院病理科2008年9月到2012年5月归档的皮肤恶性黑色素瘤蜡块54例及皮肤交界痣22例。患者术前均未进行相关治疗,如冷冻、激光、生物治疗等,且均未合并皮肤病、结缔组织病、免疫系统疾病及其他重要脏器疾病。20例正常皮肤组织(对照组)来自于郑州大学第一附属医院整形外科植皮剩余的全厚皮片。1.采用免疫组化SP法检测STA3、p27、CyclinDl蛋白在54例皮肤恶性黑色素瘤、22例交界痣、20例正常皮肤组织中的表达水平,并分析其三者之间的相互关系。2.所有数据采用SPSS17.0统计软件包行卡方检验及相关性分析,检验水准α=0.05,χ2分割检验标准α’=0.0167。结果1.STAT3蛋白主要表达于CMM黑色素细胞的细胞浆,在CMM中呈高表达,阳性表达率为77.8%(42/54),在交界痣和正常皮肤组织中呈低表达,阳性表达率分别为45.5%(10/22)、35.0%(7/20);STAT3在CMM、交界痣及正常皮肤组织之间的表达差异性具有统计学意义(p<0.0167),在交界痣和正常皮肤组织中的表达差异性无统计学意义p>0.0167).2.p27蛋白主要表达于CMM黑色素细胞的细胞核,在CMM中呈低表达,阳性表达率为16.7%(9/54),在交界痣和正常皮肤组织中呈高表达,阳性表达率分别为86.4%(19/22)、95.0%(19/20);p27在CMM、交界痣及正常皮肤组织之间的表达差异性具有统计学意义(p<0.0167),在交界痣和正常皮肤组织中的表达差异性无统计学意义(p>0.0167).3.CyclinD1蛋白主要表达于CMM黑色素细胞的细胞核,在CMM中呈高表达,阳性表达率为81.5%(44/54),在交界痣和正常皮肤组织中呈低表达,阳性表达率分别为45.0%(10/22)、30.0%(6/20);CyclinD1在CMM、交界痣及正常皮肤组织之间的表达差异性具有统计学意义(p<0.0167),在交界痣和正常皮肤组织中的表达差异性无统计学意义(p>0.0167).4.相关性分析:STAT3蛋白的表达与p27的表达呈负相关(r=-0.478, p<0.05), CyclinD1蛋白的表达与p27的表达呈负相关(r=-0.426,p<0.05); STAT3与CyclinD1蛋白的表达呈正相关(相关系数r=0.663,p<0.05)结论1.STAT3、CyclinDl蛋白在CMM中的表达明显高于皮肤交界痣及正常皮肤,它们的高表达可能促进黑色素细胞的生长,从而引起CMM的发生、发展。提示二者可能在CMM的发生、发展中发挥促进作用。2.p27蛋白在CMM中的表达明显低于其在皮肤交界痣、正常皮肤中的表达,它的低表达可能促进CMM的发生,提示p27在CMM发生中可能起抑癌作用。3.STAT3信号通路的组成性激活最终促进细胞增殖,CyclinDl是其下游的一个重要因子,在CMM中其蛋白表达可促进细胞周期的运转,从而参与CMM发生、发展。4.STAT3、p27、CyclinD1可能作为皮肤恶性黑色素瘤细胞治疗的靶点,为STAT3抑制剂应用于CMM提供理论依据。
【Abstract】 Background and PurposeCutaneous malignant melanoma (CMM),as one of familiar malignant tumors of skin,is derived from melanocytes. It is the fastest growing skin malignant tumors. As well as other human tumors, its growth and development are very complicated process, combined action of multiple factors and multiple steps and stages. Mainly include the activation of oncogenes, reducing or disappear of the activity of tumor-suppressor genes, resulting in the changes of cell cycle, cause cancer cells to grow out of control.Signal transducers and activators of transcription3(STAT3) is an important members of signal transducers and activators (STATS) of transcriptions. Generally, STAT3resides in the cytoplasm, when upstream signaling pathways activated (such as EGFR, c-Met, il-6receptor activation, etc.), STAT3phosphorylated to form dimers or form a heterodimer with other members of the STAT family. Activated STAT3complex translocate to nucleus, the transcription of its target genes(include CyclinDl、Bcl-xL、c-myc、and VEGF,etc.)will begin. STAT3promote cancerous tissue cells by promoting the expression of cell CyclinDl and Unlimited cell proliferation.The change of the cell cycle will lead the cancer cells to grow out of control.CDKI can inhibit the compound formation cyclin and CDK, the inactivation of CDKI leads to cell proliferation.As a CDKI,p27can inhibit the activation of CyclinDl/CDK4and CyclinE/CDK2,thus made the cell cycle stagnate in G1and interfere with the cells of infinite proliferation, thus inhibiting tissue cells become cancerous. To understand the role of STAT3、p27and CyclinDl in the pathogenesis of CMM, this study tested the expression of STAT3, p27, CyclinD1in54cases CMM,22case mole skin junction,20cases normal skin, and analyze the correlation of the three factors.Materials and methods54case CMM and22case mole skin junction were selected from the department of pathology in the first affiliated hospital of Zhengzhou University which archived from September2008to May2012. All patients had no therapy before,for example, freezing,laser, biotherapy and so on.They had no skin disorder connective tissue disease, immune disorders and other important organs diseases.20cases of normal skin which came from full thick skin grafts of plastic surgery were chosen as control groups.The SP immunohistochemical technique was used to detect the expression of STAT3, p27, CyclinD1in the development of CMM. The statistics data were processed by SPSS17.0, using Chi. square test and correlation analysis. The test standard is0.05and the standard of X2division is0.0167.Results1. STAT3expressed mainly on the cytoplasm of melanoma cells. There was a high expression in CMM, their positive expression is77.8%(42/54). There were low expression in mole skin junction or normal skin, their positive expression were45.5%(10/22、35.0%(7/20). There was a statistical significance between the CMM and mole skin junction or normal skin(P<0.0167).There is no statistical significance between mole skin junction and normal skin(p>0.0167).2. p27expressed mainly on the nucleus of melanoma cells. There was a low expression in CMM, their positive expression is16.7%(9/54). There were low expression in mole skin junction or normal skin, their positive expression were86.4%(19/22)、95.0%(19/20).There was a statistical significance between the CMM and mole skin junction or normal skin (p<0.0167). There is no statistical significance between mole skin junction and normal skin (p>0.0167).3. CyclinDl expressed mainly on the nucleus of melanoma cells. There was a high expression in CMM, their positive expression is81.5%(44/54). There were low expression in mole skin junction or normal skin, their positive expression were45.0%(10/22、30.0%(6/20). There was a statistical significance between the CMM and mole skin junction or normal skin(p<0.0167).There is no statistical significance between mole skin junction and normal skin(p>0.0167).4. The correlation analysis showed that there was a significant negative correlation between STAT3and p27(r=-0.478,p<0.05). There was a significant negative correlation between CyclinDl and p27(r=-0.426,p<0.05). There was a significant positive correlation between STAT3and CyclinDl (r=0.663,p<0.05).Conclusion1. Both the expressions of STAT3protein and CyclinD1protein in CMM are higher than in mole skin junction and normal skin. Their high expression may promote the growth of the melanocytes, thus cause the happening of the CMM. Prompt that the two factors may play a positive role in the occurrence of the CMM.2. The expression of p27in CMM is lower than in mole skin junction and normal skin. Its low expression may promote the proliferation of the melanocytes, thus cause the happening of the CMM. P27might play a tumor suppressor role in CMM for their low expressions.3. The constitutive activation of STAT3signal transduction pathways eventually promote cell proliferation, CyclinD1is its an important downstream factors, and its protein expression in the CMM can promote the operation of cell cycle to participate in the occurrence and development of CMM.4.STAT3、p27、CyclinD1may be new targets for cancer therapy, and provide the theory basis of STAT3inhibitors used in the treatment of CMM.