【作者】 王艳芳；

【导师】 倪劲松；

【作者基本信息】 吉林大学 ， 病理学与病理生理学， 2013， 硕士

【摘要】 大肠癌（结肠癌及直肠癌）是常见的恶性肿瘤之一，近年来，其发病率逐年增加，目前已位列世界第三位，仅次于肺癌及胃癌。以往的研究表明，大肠癌的发生与饮食习惯及多种癌基因和抑癌基因的表达异常有关，新近的研究显示，表观修饰在大肠癌的发生发展过程中所起的作用越来越受到人们的重视。有研究显示组蛋白的乙酰化修饰异常与大肠癌的发生密切相关，而组蛋白乙酰化修饰受组蛋白乙酰化酶（histone acetyl transferases，HATs）和组蛋白去乙酰化酶（histonedeacetylase，HDACs）的共同调控。hMOF是组蛋白乙酰化酶MYST家族成员之一，对组蛋白H4的第16位赖氨酸（H4K16）具有特异性的乙酰化作用。对乳腺癌、肾透明细胞癌组织等的研究显示，hMOF表达量的减少将导致整个基因组内组蛋白H4K16乙酰化水平下降，并使细胞阻滞于S期，DNA损伤得不到及时的修复从而发生恶变。但目前为止对于hMOF及H4K16的乙酰化在大肠癌发生发展中的作用尚未见确切报道。目的：探讨hMOF在直肠正常粘膜组织、管状腺瘤及癌组织中的表达，并分析hMOF与临床指标的关系及相关作用机制；同时观察H4K16Ac在直肠正常粘膜组织、管状腺瘤及癌组织中的表达，并分析hMOF与H4K16Ac之间的相关性。方法：采用免疫组化SP法检测hMOF及H4K16AC在直肠正常粘膜组织、管状腺瘤及直肠癌组织中的表达，并分析其与临床各指标之间的相关性；同时采用Western blot及QPCR方法检测hMOF在正常粘膜、管状腺瘤及癌组织中的表达；采用SPSS13.0统计学软件对hMOF及H4K16Ac在直肠癌组织中表达的进行了相关性分析。结果：免疫组化染色结果显示，hMOF在直肠癌组织中的表达明显低于直肠正常粘膜组织及管状腺瘤(P=0.000＜0.01，P=0.001＜0.01），与肿瘤分期、病理分级及有无淋巴结转移呈负相关（P=0.007＜0.01，P=0.036＜0.05, P=0.004＜0.01）；而正常粘膜组织及管状腺瘤间表达无明显差异（P=0.06＞0.05）；Westernblot及实时定量PCR（QPCR）结果显示，hMOF在直肠癌组织中表达明显低于直肠正常粘膜组织(P=0.008＜0.01，P=0.03＜0.05）；H4K16AC的免疫组化结果表明，其在直肠癌组织中的表达明显低于直肠正常粘膜组织及管状腺瘤组织（P=0.000<0.01，P=0.002<0.01），正常粘膜组织及管状腺瘤间表达无明显差异（P=0.094＞0.05），且其表达与病理分级呈负相关（P=0.028＜0.05）；相关性分析结果显示，直肠癌组织中hMOF与H4K16Ac表达明显正相关（P=0.000＜0.01）；hMOF表达与P53表达呈负相关（Kendall’s tau-b=-.5883，P=0.000＜0.01），与EGFR、Bcl-2、HER-2及Ki67无相关性（P＞0.05）。结论：①hMOF及H4K16AC在直肠癌中的表达均明显降低，二者的表达呈明显正相关；②二者的表达与肿瘤的恶性程度及不良预后具有一定的相关性；③hMOF表达下调可能通过抑制H4K16乙酰化进而促进直肠癌的发生和发展；同时；hMOF可通过对P53的调节来控制细胞凋亡，进而调节肿瘤的发生。更多还原

【Abstract】 Colorectal cancer (Colorectal cancer, CRC) is one of the most common malignant tumor inthe world, and it is effected in health seriously. CRC is a consequence of many factors, normalintestinal epithelium to complete deterioration would take many years or even decades. Includingmore and more molecular biology event occurs, such as DNA methylation, MYH (mutyh)mutation, deletion of IGF-II mark, serrated polyps and so on. The activation of oncogenes andthe inactivation of tumor suppressor genes also play an important role in the process of CRCoccurrence. Recent studies showed that epigenetics change was also one of the pathogenesis ofCRC. Epigenetics includes DNA methylation, histone modification, micRNA interference,however, the histone modification plays an important part in the regulation of cell cycle,chromatin remodeling, DNA repair and proteinexpression; Human MOF (human males absent onthe first, hMOF) is one of the members of histone acetyltransferase family, it has the function ofspecifically recognise acetylated H4K16and P53. Some research confirms that hMOF canparticipated in the functions include gene transcription, change of the conformation in chromatin,cell cycle regulation, repair of DNA damage, apoptosis and maintaining the nuclear structuredirect or indirect by the effects of acetylation. The study showed that, the expression of hMOF inrenal cell carcinoma, glioma, and other types of malignancies was significantly lower than that innormal tissue. With the increasing of grade malignancy, loss of hMOF expression is more obvious,it showed that loss of hMOF expression plays an important role in the process ofcellcarcinogenesis. At the same time the study also showed that the role of hMOF is inhibit theacetylation of histone H4K16. But at present the acetylation of hMOF and H4K16in thedevelopment of rectal cancer have not been reported.Our research aims to combined detect the expression hMOF and H4K16Ac in colorectalnormal mucosa, tubular adenoma tissue and colorectal carcinoma to analysis their correlation, andexplore the significance and the mechanism of their expression in rectal cancer.1. Experimental method(1)In this study, we used immunohistochemical staining SP two-step method detected theexpression level of hMOF and H4K16Ac protein in66cases of rectal cancer tissues and15casestubular adenoma to analyze the correlation with clinical index.(2)Western blot and QPCR were used to analyse the expression of hMOF in44cases of freshspecimens of rectal cancer and its normal mucosa tissue.(3)Statistical methods to analyse the correlation between the expression of hMOF andH4K16Ac in rectal cancer tissues. 2. Experimental results(1) The results of immunohistochemistry showed that hMOF was positively expressed withbrown, the staining was located in the nucleus. The rate of positive expression of hMOF innormal mucosa tissue, rectal tubular adenoma tissue and rectal cancer tissue were100%,93.3%,and34.9%. The expression of hMOF in normal mucosa tissue and rectal tubular adenoma tissueshowed no significant difference (P=0.06＞0.01). The expression of hMOF was significantlylower than the former (P=0.000＜0.01，P=0.001＜0.01). The expression of hMOF hassignificantly positive correlation with tumors stage, pathological differentiation and lymph nodemetastasis (P=0.007＜0.01，P=0.036＜0.01，P=0.004＜0.01).(2) The results of Western blot and QPCR showed that the expression of hMOF andH4K16Ac in rectal cancer tissues was significantly lower than that in fresh normal mucosa.(P=0.008＜0.01,P=0.03＜0.05).(3) The rate of positive expression of H4K16Ac in normal mucosa tissue, rectal tubularadenoma tissue and rectal cancer tissue was100%,80%,42.4%. The expression of H4K16Ac innormal mucosa tissue and rectal tubular adenoma tissue showed no significant difference(P=0.094＞0.01). The expression of H4K16Ac was significantly lower than the former(P=0.000＜0.01，P=0.004＜0.01). The expression of hMOF has significantly positive correlationwith pathological differentiation (P=0.000＜0.01).(4) The statistical results showed that the expression of hMOF and H4K16Ac havesignificantly positive correlation in rectal cancer tissues (P=0.000＜0.01).3. conclusion(1) The expression of hMOF protein in colorectal carcinoma was lower and the expressionwas decrease with the increase of pathological differentiation and tumors stage, and it wasnegatively correlated with lymph node metastasis;(2) The expression of H4K16Ac protein in colorectal carcinoma was lower; and theexpression was decrease with the increase of pathological differentiation;(3) hMOF protein was positively correlated with H4K16Ac protein in the tissue of colorectalcarcinoma.更多还原