【作者】 姆巴东；

【导师】 程邦昌； 赫捷；

【作者基本信息】 武汉大学 ， 胸心外科， 2004， 博士

【摘要】 抑癌基因(常是隐性基因)的异常表达或突变是致瘤作用机制之一。人类基因的很多区域，发生改变时可致细胞的无控性生长。肺癌中最引人注目的是位于3号染色体短臂区域的抑癌基因。杂合缺失研究显示3P缺失与多种肿瘤有关，尤其是3p21.3，在肺癌和卵巢癌中有高度杂合缺失，表明在该区域可能存在潜在的抑癌基因。小细胞肺癌中该位点杂合缺失高达90％，在其它肿瘤，如乳腺癌、胃、卵巢、睾丸及肾肿瘤中也发现较高的杂合缺失。已在该区域发现19个候选抑癌基因。Semaphorin家族的SEMA3B(Semaphorin 3B)及SEMA3F(Semaphorin 3F)是其中之一。零星关于SEMA3B及SEMA3F mRNA表达方面及其抑癌作用机制的研究集中于肺癌细胞系中。但该基因在原发性肺癌组织中的转录表达与临床病理关系的研究尚无报道。已有研究报道食管癌频繁的3p区域杂合缺失，但尚未见SEMA3B及SEMA3F mRNA表达与食管癌的临床病理关系研究报道。 本研究首先从SEMA3B及SEMA3F基因的转录表达水平，采用RT-PCR方法检测52例中国人群肺癌患者远癌正常肺组织和肺癌组织中SEMA3B及SEMA3F mRNA的表达情况。结果显示肺癌组织中SEMA3B基因产物的表达水平为51.9％(27／52)，明显低于远癌正常肺组织100％(52／52)(p＜0.001)。肺癌组织中SEMA3B基因产物的表达水平与肺癌TNM分期及淋巴结转移有密切关系(p＜0.05)，与肺癌类型，原发肿瘤大小、部位，患者的年龄、性别及吸烟与否均无明显关系(p＞0.05)。研究又发现SEMA3B表达异常与病理类型、分化程度、病理分期无显著关系。SEMA3F表达异常与病理类型、分化程度、病理分期、淋巴结转移无显著关系。SEMA3F基因在肺癌和远癌正常肺组织表达率分别为47.8％(22／46)、100％(46／46)，有非常显著性统计和差异(p＜0.001)。在肺癌中SEMA3F的表达缺失与病理形态、分化程度、病理分期、淋巴结转移无显著关系(p＞0.05)。SEMA3B和SEMA3F基因表达缺失率分别为48.1％(25／52)和44.2％(23／52)两种基因的表达水平差异无显著性，但表达性质不全相同。二者累计表达缺失率为69.231％(36／52)，其中13例两个基因都表达缺失，前者和后者的单独缺失分别为12例和10例，差异无统计学意义(p＞0.05)。 然后，本研究采用逆转录聚合酶链反应(RT-PCR)方法检测46例食管癌组织及远癌正常食管组织SEMA3B和SEMA3F基因的表达水平。结果显示：SEMA3B在食管癌组织中表达率为50%(23/46)，明显低于相应远癌正常食管组织100%(46/46)(P<0.ool)。sEMA3B在食管腺癌、小细胞癌、鳞状上皮细胞癌中的表达率分别为80%(4/5)、0%(o/3)、50%(19/38)，三组表达差异有显著性切<0.05);SEMA3B在表达异常与病理形态、分化程度、病理分期、淋巴结转移无显著相关。SEMA3F基因在食管癌和远癌正常食管组织表达率分别为47.8%(22/46)、100%(46/46)、(p<0刀01)。在食管癌中SEMA3F的表达缺失与病理形态、分化程度、病理分期、淋巴结转移无显著关系(P>0.05)。SEMA3B和SEMA3F基因表达缺失率分别为50%(23/46)和47.8%(22/46)，两种基因的表达水平上无显著差异，但表达性质不全相同。二者累计表达缺失率为73.91%(34/46)，其中13例两个基因都表达缺失，前者和后者的单独缺失分别为10例和11例，差异无统计学意义 (P>0.OS)。 本实验首次检验了中国人群肺癌患者远癌组织和肿瘤组织中SEMA3B、SEMA3F基因的表达情况。发现肺癌存在SEMA3B、SEMA3F基因的表达缺失，与正常肺组织相比有显著差异，提示SEMA3B和SEMA3F基因的表达缺失与原发性肺癌的发生发展密切相关;首次发现SEMA3B基因的表达缺失与肺癌淋巴结转移密切相关，提示SEMA3B基因的表达缺失与原发性肺癌的预后相关。本研究首次在人体食管癌标本中检测SEMA3B和SEMA3F基因转录本的表达情况，发现二者转录本在中国食管癌患者癌组织中的高度表达缺失，充分表明该基因在中国食管癌的发生、发展中起了极为重要的作用，发挥了抑癌基因的功能。联合检测两种基因可提高诊断水平。SEMA3B和SEMA3F的表达异常是肺癌、食管癌发生、发展的的早期事件，为肺癌、食管癌的早期诊断和基因治疗提供了理论依据。更多还原

【Abstract】 Tumorigenesis results from the abnormal regulation of genes involved in cellular homeostasis. One mechanism involve the alteration in gene expression or mutation of tumor suppressor genes, which are recessive genes where mutation results of loss of function. There are several regions within the human genome that when altered, lead to uncontrolled cell proliferation. The short arm of chromosome 3 has been shown to exhibit high loss of heterozygosity (LOH) in several types of cancer including ovarian, kidney, lung, and testicular cancers. In particular, overlapping homozygous deletions in lung cancers have been identified in region 3p21.3 where already 19genes were found. In particular the 3p21.3 region exhibits a high LOH in lung, breast, stomach and ovarian carcinomas suggesting the existence of a putative tumor suppressor gene within this region. In small cell lung cancer, >90% of the tumors exhibits LOH at this site. Two Class III Semaphorin genes SEMA3B and SEMA3F are ones of them. A few articles converge about mRNA expression and suppressor mechanism on tumor cell line, but up to now, there is no publication about SEMA3B and SEMA3F mRNA expression and their tumor suppressor function mechanism in lung and esophageal carcinoma tissue. But the association between the expression of this two SEMA and their clinicopathologic characters was still unexplored in primary lung cancer. The 3p LOH in esophageal carcinoma has been established but there is no publication yet about SEMA3B and SEMA3F in esophageal carcinoma.In this study, we firstly used the RT-PCR method to evaluate the transcription expression of SEMA3B and SEMA3F in 52 cases of Chinese lung cancer tissues and adjacent normal tissues. The result showed that, the SEMA3B and SEMA3F transcription were identified in all non-cancer tissues but was not found in 53.2%(27/52) and 55.2(29/52) carcinoma tissues (p<0.001). Loss of SEMA3B expression was significantly high in patients with positive lymph node metastasis 64.3% (18/28) compared with those without lymph node metastasis 29.2 %( 7/24) (p<0.05). The rate of loss of SEMA3B mRNA expression was correlated with TNM stage (p<0.05), which was more frequently observed in those with advanced tumor stage. There is no significant association of abnormal expression with histological type, differentiation grade stage of tumors or age, sex, and smoking index. SEMA3B and SEMA3F expression loss respectively were 48.1% (25/52) and 44.2%(23/52) with no significant difference in rate but with different pattern: the cumulative incidence of expression losswas69.231% (36/46) . SEMA3B and SEMA3F were differently lost in 10 and 11 cases respectively, both the two genes expression were lost in 13 cases.We secondly used the RT-PCR method to evaluate the expression level of SEMA3B and SEMA3F genes in 46 cases of esophageal cancer tissues and adjacent normal tissues. The result showed that, the SEMA3B and SEMA3F transcripts were identified in all non-cancer tissues but was not found respectively in 50%(23/46) and 47.8%(22/46) carcinoma tissues respectively (pO.OOl) with distinct expression patterns. No significant association of abnormal SEMA3B and SEMA3F mRNA expression with histological type, differentiation grade of tumors or patients age and sex was observed (pX).05).Our trial was the first one which studies the relationship between SEMA3B, SEMA3F and clinicopathological characters of lung and esophageal cancer in Chinese population and the first one establishing correlation between lymph node metastasis and lung cancer and their correlation with esophageal carcinoma. We respectively discovered SEMA3B and SEMA3F transcription significantly inactivating in lung and esophageal cancer, suggesting that SEMA3B and SEMA3F genes could play a role of TSG in both lung and esophageal carcinoma. SEMA3B mRNA inactivation has been found associated with lung cancer lymph node metastasis suggest that SEMA3B gene transcription inactivation correlate with lung cancer prognosis. Simultaneous check up of the two genes can improve the diagno更多还原