PRM1在结肠癌组织中的表达及意义
摘要
结肠癌是我国高发的恶性肿瘤，占胃肠道恶性肿瘤的第三位，其发病率逐年增高，且有年轻化趋势。结肠癌诊断主要依赖肠镜及腹部CT等手段，故大多数患者明确诊断时已处于癌症晚期失去了手术治疗的机会，因此寻找特异的早期诊断标记及分子治疗靶点十分重要。鱼精蛋白（protamines，PRM）被认为与男性不育有关，最新研究表明，PRM1可能是一个新的结肠癌相关的肿瘤/睾丸（Cancer/Testis,CT）抗原基因。本研究的目的在于通过检测结肠癌组织中PRM1-mRNA和蛋白水平的变化，阐述PRM1在临床诊断结肠癌中的意义及其作为基因治疗靶点的可能性。具体研究如下：
1实验路线
1.1收集结肠癌患者外周血，分离血清；收集结肠癌组织、正常结肠组织。
1.2实时定量PCR比较结肠癌组织与正常结肠组织PRM1mRNA表达水平差异。
1.3PRM1mRNA检测用于结肠癌诊断敏感性分析。
1.4免疫组织化学染色观察PRM1在结肠癌组织中的表达。
1.4.1结肠癌组织PRM1表达水平与正常结肠组织PRM1表达水平差异比较。
1.4.2高、中、低分化结肠腺癌组织PRM1表达水平差异比较。
1.5western blot检测比较结肠腺癌与正常结肠组织PRM1表达水平差异。
1.6ELISA试验检测结肠癌患者血清PRM1水平。
1.6.1结肠癌患者血清PRM1水平与健康人差异比较。
1.6.2不同分期结肠癌患者血清PRM1水平差异比较。
1.6.3高、中、低分化结肠腺癌患者血清PRM1水平差异比较。
1.6.4PRM1血清检测用于结肠癌诊断的阳性率。
1.7ELISA试验检测结肠癌患者血清CEA水平。
1.8血清PRM1、CEA检测用于结肠癌诊断敏感性比较。
2结果
2.1结肠癌PRM1mRNA表达水平高于正常结肠组织
53例结肠癌组织平均PRM1-mRNA含量显著高于正常结肠组织，P=0.01116。将定量PCR反应产物进行琼脂糖凝胶电泳，通过辉度扫描比较结肠癌与正常结肠组织PRM1mRNA表达差异，结果与定量PCR结果一致。
2.2组织PRM1mRNA检测用于结肠癌诊断敏感性分析
以结肠癌组织中PRM1-mRNA含量大于等于正常结肠组织PRM1-mRNA含量的2倍以上记为PRM1-mRNA表达阳性。统计结果显示，53例结肠癌病例PRM1mRNA检测总体阳性率为96.2%（51/53）。
2.3免疫组织化学染色观察PRM1在结肠癌组织中的表达
实验采取免疫组织化学染色的方法检测结肠癌组织表达PRM1表达及确定组织分布特点。结果显示，结肠癌细胞胞浆内及癌细胞围成的腺腔内可见PRM1阳性染色颗粒，正常结肠组织细胞内无明显PRM1阳性染色。低分化腺癌细胞内PRM1染色强度明显高于高分化腺癌细胞。
2.4western blot检测PRM1在结肠癌组织中的表达
实验采取western blot的方法检测结肠癌组织与正常结肠组织表达PRM1水平差异。结果显示，结肠癌组织PRM1水平远远高于正常结肠组织，与免疫组化实验一同证明，结肠癌组织整体PRM1表达水平高于健康结肠组织。
2.5ELISA试验检测结肠癌患者血清PRM1水平
2.5.1结肠癌患者血清PRM1水平与健康人差异比较
ELISA试验检测结肠癌患者与健康成年人血清PRM1水平，结果显示，结肠癌患者血清PRM1水平显著高于健康成年人，P<0.05。
2.5.2PRM1血清检测用于结肠癌诊断的阳性率
根据健康成年人PRM1血清水平的95%可信区间判断结肠癌患者血清水平阳性率，结果显示，结肠癌患者血清PRM1检测阳性率为80%（24/30）。
2.5.3不同分期结肠癌患者血清PRM1水平差异比较
比较T_1N_XM_X、T_2N_XM_X、T_3N_XM_X结肠癌患者血清PRM1水平，结果显示，各组患者血清PRM1水平无明显差异，提示结肠癌患者血清PRM1水平升高程度与肿瘤T分期无关。
2.5.4高、中、低分化结肠腺癌患者血清PRM1水平差异比较
按结肠腺癌细胞病理分化程度比较患者血清PRM1水平差异，结果显示，低分化结肠癌患者血清PRM1水平与中分化结肠癌患者相比，差异无显著性，P>0.05。
2.6ELISA试验检测结肠癌患者血清CEA水平
ELISA试验检测结肠癌患者与健康成年人血清CEA水平，结果显示，结肠癌患者血清CEA平均水平显著高于健康成年人平均水平，P<0.05。
2.7血清PRM1、CEA检测用于结肠癌诊断敏感性比较
临床上常用CEA作为结肠癌等肿瘤的血清学诊断标记，故将PRM1与CEA用于结肠癌诊断的敏感性进行比较，结果显示，血清PRM1用于结肠癌诊断的阳性率为80%(24/30)，血清CEA用于结肠癌诊断的阳性率为30%（9/30）。经过统计学检验，p<0.05。证实PRM1用于结肠癌诊断的敏感性高于CEA。
3结论
3.1PRM1mRNA在结肠癌中含量显著高于正常结肠组织。
3.2结肠腺癌细胞及癌细胞围成的腺腔内可见PRM1蛋白阳性染色，且PRM1表达水平与结肠腺癌细胞分化度有关。提示PRM1可能通过自分泌/旁分泌的方式作用于肿瘤细胞。
3.3Western blot实验进一步证明，结肠腺癌组织比正常结肠组织PRM1表达水平显著高于正常肠组织。
3.4结肠癌患者外周血中PRM1水平高于健康成年人，提示PRM1可能通过内分泌的方式作用于远处组织及细胞，具体作用及作用方式有待于进一步研究。
3.5血清PRM1水平用于结肠癌诊断的阳性率高于CEA。
Colon cancer is one high incidence of malignancy in China, accounting for thethird in total malignant tumors of the gastrointestinal. Patients with colon cancerincreased year by year, and are getting younger and younger. Diagnosis of coloncancer mainly depends on the means of colonoscopy and abdominal CT, so mostpatients lost opportunity of surgical treatment when they get clearly diagnosis, solook for specific early diagnostic markers and molecular therapeutic targets is veryimportant. Protamine (PRM) is considered to be related with male infertility, thelatest research shows that, PRM1may be a new colon cancer/testis (Cancer/Testis,CT) antigen gene. The purpose of this study is to elaborated the significance ofPRM1in the clinical diagnosis of colon cancer and the possibility of PRM1as genetherapy targets of colon cancer through the detection of PRM1-mRNA and proteinlevels change in colon cancer tissue. Specific studies are as follows:
Experimental Routes:
1.1Collecte peripheral blood of patients with colon cancer, the serum wasseparated; collected colon cancer tissue and normal colon tissues.
1.2Compare the differences of PRM1mRNA expression level of colon cancertissue and normal colon tissue via Real-time quantitative PCR.
1.3Analysis sensitivity of PRM1mRNA detection for the diagnosis of coloncancer.
1.4Immunohistochemical staining observe PRM1expression in colon cancertissue
1.4.1Compare PRM1expression level in colon cancer tissue and normalcolon tissue PRM1expression levels.
1.4.2Compare difference of PRM1expression in high differentiated adenocarcinoma, medium differentiated colon adenocarcinoma andpoorly differentiated colon adenocarcinoma.
1.5Compare the differences of PRM1expression level between colonadenocarcinoma and normal colon tissue by western blot.
1.6Detect serum PRM1levels of patients with colon adenocarcinoma.
1.6.1Compare the differences serum levels between patients with colonadenocarcinoma and health controls.
1.6.2Compare the differences serum levels between patients who withadenocarcinoma in different stages.
1.6.3Compare the differences serum levels between well, moderate andpoorly differentiated colon adenocarcinoma.
1.6.4The positive rate of serum PRM1for colon cancer diagnosis.
1.7ELISA assay detected serum CEA level in colon cancer.
1.8Compare the sensitivity of serum PRM1, CEA for diagnosis colonadenocarcinoma.
2. Result
2.1The PRM1mRNA levels in the colon cancer is higher than that in normalcolon tissues
The average PRM1-mRNA content of53cases of colon cancer tissue wassignificantly higher than that in normal colon tissue, P=0.01116. The same resultsconsistent with the results of the quantitative PCR was got through the luminancescanning when quantitative PCR reaction product was subjected to agarose gelelectrophoresis and compared PRM1mRNA expression difference between coloncarcinoma and normal colon tissue.
2.2Sensitivity analysis of PRM1mRNA detection for colon cancer diagnosis
PRM1-mRNA expression was defined positive when PRM1-mRNAcontent in colon cancer tissue was higher or equal to2times than that in normalcolon tissue. Statistics results showed that the PRM1mRNA positive rate was96.2% (51/53)
2.3Immunohistochemical staining assay detected PRM1expressions incolon cancer tissue
PRM1expression and tissue distribution characteristics were detected viaimmunohistochemistry. The results showed that PRM1positive staining particleswere found in the cytoplasm of colon cancer cells and glandular cavity surroundedby cancer cells. PRM1staining intensity in poorly differentiated adenocarcinomacells was significantly higher than the well-differentiated adenocarcinoma cells.
2.4Detected PRM1expression in colon cancer tissue via western blot
The experiments used western blot to detect PRM1expression levels of coloncancer tissue and normal colon tissue. The results showed that the PRM1level incolon cancer tissue were far higher than that in the normal colon tissue. Western blotexperiment together with the immunohistochemistry experiment proved the PRM1expression level was higher in colon cancer tissue than that in normal colon tissue.
2.5ELISA test detected serum PRM1level of patients with colon cancer
2.5.1Compare serum PRM1level with colon cancer with that in healthypeople.
Detected serum PRM1levels in patient with colone cancer and healthy adultsby ELISA assay, the results showed that the serum PRM1level of patients withcolon cancer were significantly higher than that in healthy adults, P <0.05.
2.5.2The positive rate of serum PRM1used for colon cancer diagnosis
Using the confidence interval for the group of healthy subjects as the cut-offvalue, the sensitivity of PRM1as markers for colon adenocarcinoma were calculatedto investigate its diagnostic efficacies.Ap-value≤0.05was considered to bestatistically significant.The results showed that the serum positive rate of patientswith colon cancer was80%(24/30)
2.5.3Compared PRM1levels of patients with different stages colon cancer
Compare serum PRM1level of patients with T_1N_XM_X、T_2N_XM_X、T_3N_XM_X colon cancer, the results showed that there was no significant differences in serumPRM1level of patients who in different colon stages.
2.5.4Compared PRM1levels of patients with different differentiation
Compared serum PRM1level according to Pathological differentiation of colonadenocarcinoma cells. The results showed that there was no significant differencesin serum PRM1level of patients with different differentiation, P>0.05.
2.6Detected serum CEA level of patients with colon cancer via ELISA test
Using ELISA test to detect serum CEA level of patients with colon cancer andhealthy adult, the results showed that the average serum CEA level of patients withcolon cancer was significantly higher than that of healthy adults (P<0.05)
2.7serum PRM1, CEA detection for colon cancer diagnostic sensitivity
Serum CEA was commonly as a serological diagnostic marker for colon cancerand other tumors, so we compared the diagnostic sensitivity of serum PRM1withCEA as a colon cancer diagnostic marker, the results showed that the diagnosispositive rate of PRM1was80%(24/30) and positive rate of CEA was30%(9/30).
3. Conclusions
3.1PRM1mRNA levels in colon cancer were significantly higher than that innormal colon tissue.
3.2PRM1positive staining particles were found in the cytoplasm of coloncancer cells and glandular cavity surrounded by cancer cells. PRM1performs bothautocrine and paracrine function.
3.3PRM1level in colon cancer tissue was far higher than that in the normalcolon tissue with western blot.
3.4The serum PRM1level of patients with colon cancer is significantly higherthan that of healthy adults. PRM1performs endocrine function.
3.5Serum PRM1level is more sensitive than serum CEA.
1实验路线
1.1收集结肠癌患者外周血，分离血清；收集结肠癌组织、正常结肠组织。
1.2实时定量PCR比较结肠癌组织与正常结肠组织PRM1mRNA表达水平差异。
1.3PRM1mRNA检测用于结肠癌诊断敏感性分析。
1.4免疫组织化学染色观察PRM1在结肠癌组织中的表达。
1.4.1结肠癌组织PRM1表达水平与正常结肠组织PRM1表达水平差异比较。
1.4.2高、中、低分化结肠腺癌组织PRM1表达水平差异比较。
1.5western blot检测比较结肠腺癌与正常结肠组织PRM1表达水平差异。
1.6ELISA试验检测结肠癌患者血清PRM1水平。
1.6.1结肠癌患者血清PRM1水平与健康人差异比较。
1.6.2不同分期结肠癌患者血清PRM1水平差异比较。
1.6.3高、中、低分化结肠腺癌患者血清PRM1水平差异比较。
1.6.4PRM1血清检测用于结肠癌诊断的阳性率。
1.7ELISA试验检测结肠癌患者血清CEA水平。
1.8血清PRM1、CEA检测用于结肠癌诊断敏感性比较。
2结果
2.1结肠癌PRM1mRNA表达水平高于正常结肠组织
53例结肠癌组织平均PRM1-mRNA含量显著高于正常结肠组织，P=0.01116。将定量PCR反应产物进行琼脂糖凝胶电泳，通过辉度扫描比较结肠癌与正常结肠组织PRM1mRNA表达差异，结果与定量PCR结果一致。
2.2组织PRM1mRNA检测用于结肠癌诊断敏感性分析
以结肠癌组织中PRM1-mRNA含量大于等于正常结肠组织PRM1-mRNA含量的2倍以上记为PRM1-mRNA表达阳性。统计结果显示，53例结肠癌病例PRM1mRNA检测总体阳性率为96.2%（51/53）。
2.3免疫组织化学染色观察PRM1在结肠癌组织中的表达
实验采取免疫组织化学染色的方法检测结肠癌组织表达PRM1表达及确定组织分布特点。结果显示，结肠癌细胞胞浆内及癌细胞围成的腺腔内可见PRM1阳性染色颗粒，正常结肠组织细胞内无明显PRM1阳性染色。低分化腺癌细胞内PRM1染色强度明显高于高分化腺癌细胞。
2.4western blot检测PRM1在结肠癌组织中的表达
实验采取western blot的方法检测结肠癌组织与正常结肠组织表达PRM1水平差异。结果显示，结肠癌组织PRM1水平远远高于正常结肠组织，与免疫组化实验一同证明，结肠癌组织整体PRM1表达水平高于健康结肠组织。
2.5ELISA试验检测结肠癌患者血清PRM1水平
2.5.1结肠癌患者血清PRM1水平与健康人差异比较
ELISA试验检测结肠癌患者与健康成年人血清PRM1水平，结果显示，结肠癌患者血清PRM1水平显著高于健康成年人，P<0.05。
2.5.2PRM1血清检测用于结肠癌诊断的阳性率
根据健康成年人PRM1血清水平的95%可信区间判断结肠癌患者血清水平阳性率，结果显示，结肠癌患者血清PRM1检测阳性率为80%（24/30）。
2.5.3不同分期结肠癌患者血清PRM1水平差异比较
比较T_1N_XM_X、T_2N_XM_X、T_3N_XM_X结肠癌患者血清PRM1水平，结果显示，各组患者血清PRM1水平无明显差异，提示结肠癌患者血清PRM1水平升高程度与肿瘤T分期无关。
2.5.4高、中、低分化结肠腺癌患者血清PRM1水平差异比较
按结肠腺癌细胞病理分化程度比较患者血清PRM1水平差异，结果显示，低分化结肠癌患者血清PRM1水平与中分化结肠癌患者相比，差异无显著性，P>0.05。
2.6ELISA试验检测结肠癌患者血清CEA水平
ELISA试验检测结肠癌患者与健康成年人血清CEA水平，结果显示，结肠癌患者血清CEA平均水平显著高于健康成年人平均水平，P<0.05。
2.7血清PRM1、CEA检测用于结肠癌诊断敏感性比较
临床上常用CEA作为结肠癌等肿瘤的血清学诊断标记，故将PRM1与CEA用于结肠癌诊断的敏感性进行比较，结果显示，血清PRM1用于结肠癌诊断的阳性率为80%(24/30)，血清CEA用于结肠癌诊断的阳性率为30%（9/30）。经过统计学检验，p<0.05。证实PRM1用于结肠癌诊断的敏感性高于CEA。
3结论
3.1PRM1mRNA在结肠癌中含量显著高于正常结肠组织。
3.2结肠腺癌细胞及癌细胞围成的腺腔内可见PRM1蛋白阳性染色，且PRM1表达水平与结肠腺癌细胞分化度有关。提示PRM1可能通过自分泌/旁分泌的方式作用于肿瘤细胞。
3.3Western blot实验进一步证明，结肠腺癌组织比正常结肠组织PRM1表达水平显著高于正常肠组织。
3.4结肠癌患者外周血中PRM1水平高于健康成年人，提示PRM1可能通过内分泌的方式作用于远处组织及细胞，具体作用及作用方式有待于进一步研究。
3.5血清PRM1水平用于结肠癌诊断的阳性率高于CEA。
Colon cancer is one high incidence of malignancy in China, accounting for thethird in total malignant tumors of the gastrointestinal. Patients with colon cancerincreased year by year, and are getting younger and younger. Diagnosis of coloncancer mainly depends on the means of colonoscopy and abdominal CT, so mostpatients lost opportunity of surgical treatment when they get clearly diagnosis, solook for specific early diagnostic markers and molecular therapeutic targets is veryimportant. Protamine (PRM) is considered to be related with male infertility, thelatest research shows that, PRM1may be a new colon cancer/testis (Cancer/Testis,CT) antigen gene. The purpose of this study is to elaborated the significance ofPRM1in the clinical diagnosis of colon cancer and the possibility of PRM1as genetherapy targets of colon cancer through the detection of PRM1-mRNA and proteinlevels change in colon cancer tissue. Specific studies are as follows:
Experimental Routes:
1.1Collecte peripheral blood of patients with colon cancer, the serum wasseparated; collected colon cancer tissue and normal colon tissues.
1.2Compare the differences of PRM1mRNA expression level of colon cancertissue and normal colon tissue via Real-time quantitative PCR.
1.3Analysis sensitivity of PRM1mRNA detection for the diagnosis of coloncancer.
1.4Immunohistochemical staining observe PRM1expression in colon cancertissue
1.4.1Compare PRM1expression level in colon cancer tissue and normalcolon tissue PRM1expression levels.
1.4.2Compare difference of PRM1expression in high differentiated adenocarcinoma, medium differentiated colon adenocarcinoma andpoorly differentiated colon adenocarcinoma.
1.5Compare the differences of PRM1expression level between colonadenocarcinoma and normal colon tissue by western blot.
1.6Detect serum PRM1levels of patients with colon adenocarcinoma.
1.6.1Compare the differences serum levels between patients with colonadenocarcinoma and health controls.
1.6.2Compare the differences serum levels between patients who withadenocarcinoma in different stages.
1.6.3Compare the differences serum levels between well, moderate andpoorly differentiated colon adenocarcinoma.
1.6.4The positive rate of serum PRM1for colon cancer diagnosis.
1.7ELISA assay detected serum CEA level in colon cancer.
1.8Compare the sensitivity of serum PRM1, CEA for diagnosis colonadenocarcinoma.
2. Result
2.1The PRM1mRNA levels in the colon cancer is higher than that in normalcolon tissues
The average PRM1-mRNA content of53cases of colon cancer tissue wassignificantly higher than that in normal colon tissue, P=0.01116. The same resultsconsistent with the results of the quantitative PCR was got through the luminancescanning when quantitative PCR reaction product was subjected to agarose gelelectrophoresis and compared PRM1mRNA expression difference between coloncarcinoma and normal colon tissue.
2.2Sensitivity analysis of PRM1mRNA detection for colon cancer diagnosis
PRM1-mRNA expression was defined positive when PRM1-mRNAcontent in colon cancer tissue was higher or equal to2times than that in normalcolon tissue. Statistics results showed that the PRM1mRNA positive rate was96.2% (51/53)
2.3Immunohistochemical staining assay detected PRM1expressions incolon cancer tissue
PRM1expression and tissue distribution characteristics were detected viaimmunohistochemistry. The results showed that PRM1positive staining particleswere found in the cytoplasm of colon cancer cells and glandular cavity surroundedby cancer cells. PRM1staining intensity in poorly differentiated adenocarcinomacells was significantly higher than the well-differentiated adenocarcinoma cells.
2.4Detected PRM1expression in colon cancer tissue via western blot
The experiments used western blot to detect PRM1expression levels of coloncancer tissue and normal colon tissue. The results showed that the PRM1level incolon cancer tissue were far higher than that in the normal colon tissue. Western blotexperiment together with the immunohistochemistry experiment proved the PRM1expression level was higher in colon cancer tissue than that in normal colon tissue.
2.5ELISA test detected serum PRM1level of patients with colon cancer
2.5.1Compare serum PRM1level with colon cancer with that in healthypeople.
Detected serum PRM1levels in patient with colone cancer and healthy adultsby ELISA assay, the results showed that the serum PRM1level of patients withcolon cancer were significantly higher than that in healthy adults, P <0.05.
2.5.2The positive rate of serum PRM1used for colon cancer diagnosis
Using the confidence interval for the group of healthy subjects as the cut-offvalue, the sensitivity of PRM1as markers for colon adenocarcinoma were calculatedto investigate its diagnostic efficacies.Ap-value≤0.05was considered to bestatistically significant.The results showed that the serum positive rate of patientswith colon cancer was80%(24/30)
2.5.3Compared PRM1levels of patients with different stages colon cancer
Compare serum PRM1level of patients with T_1N_XM_X、T_2N_XM_X、T_3N_XM_X colon cancer, the results showed that there was no significant differences in serumPRM1level of patients who in different colon stages.
2.5.4Compared PRM1levels of patients with different differentiation
Compared serum PRM1level according to Pathological differentiation of colonadenocarcinoma cells. The results showed that there was no significant differencesin serum PRM1level of patients with different differentiation, P>0.05.
2.6Detected serum CEA level of patients with colon cancer via ELISA test
Using ELISA test to detect serum CEA level of patients with colon cancer andhealthy adult, the results showed that the average serum CEA level of patients withcolon cancer was significantly higher than that of healthy adults (P<0.05)
2.7serum PRM1, CEA detection for colon cancer diagnostic sensitivity
Serum CEA was commonly as a serological diagnostic marker for colon cancerand other tumors, so we compared the diagnostic sensitivity of serum PRM1withCEA as a colon cancer diagnostic marker, the results showed that the diagnosispositive rate of PRM1was80%(24/30) and positive rate of CEA was30%(9/30).
3. Conclusions
3.1PRM1mRNA levels in colon cancer were significantly higher than that innormal colon tissue.
3.2PRM1positive staining particles were found in the cytoplasm of coloncancer cells and glandular cavity surrounded by cancer cells. PRM1performs bothautocrine and paracrine function.
3.3PRM1level in colon cancer tissue was far higher than that in the normalcolon tissue with western blot.
3.4The serum PRM1level of patients with colon cancer is significantly higherthan that of healthy adults. PRM1performs endocrine function.
3.5Serum PRM1level is more sensitive than serum CEA.
引文
[1]廖真防.E-cadherin,VEGF,CD44V6在结肠癌中的表达及其临床意义[D].重庆医科大学硕士论文,2007.5:6
[2] Frattini M, Balestra D, Suardi S, et al. Different genetic featuresassociated withcolon and rec-tal carcinogenesis［J］. Clin CancerRes,2004,10(12):4015-4021.
[3]徐建明,正确认识结肠癌和直肠癌诊治理念的差异[J].中国实用外科杂志,2012,32(9):703-705
[4] Bernstein LR, Liotta LA. Molecular mediator of interactions withextracelluarmatrix components in metastasis and angiogenesis. Curr Opin Oncol,1994,6(1):106-113.
[5] Skinner SA，Frydman GM, Obrif PE, et al. Microvascular structure of benign andmalignan tumor of the colon in humans. Dig Dis Sci,1995,40(2):373-384.
[6] Krishnan J, Kirkin V, Steffen A, et al. Differential in vitro expression of vascularendothelial growth factor(VEGF)-C and VEGF-D in tumors and its relationshipto lymphatic metastasis in immunocompetents rats.Cancer Ras,2003,63:713-722.
[7]曾益新.肿瘤学[M].北京:人民卫生出版社,第二版,2003:309
[8] Ferrar N，Henzel WJ．Pituitary follicular cell secrete anovel heparin bindinggrowth factors pecific for vascular endothelial cells[J]．Biochem Biophys ResCommun，1989，161(2)：851
[9] Iljin K,Karkkanen MJ, Lawrence EC, et al. VEGFR3gene structure, regulatoryregion, and sequence ploy morphisms [J]. FASEB J.2001,15(6):1028-1036
[10] Yancopoulos GD,Davis S, Gale NW, et al. Vascular specific growth factors andblood vessel formation [J]. Nature,2000,407:242-248
[11] Ferrara N, Gerber HP, LeCouter J,et al. The biology of VEGF and itsreceptors.Nat Med,2003,9:669–676
[12] Werther K, Christensen IJ, Brunner N,Nielsen HJ,et al, Soluble vascularendothelial growth factor levels in patients with primary colorectalcarcinoma.The Danish RANX05Colorectal Cancer Study Group. Eur J SurgOncol,2000,26:657–662
[13] Akbulut H, Altuntas F, Akbulut KG,Ozturk G, Cindoruk M, Unal E, Icli F,etal.Prognostic role of serum vascular endothelial growth factor, basic fibroblastgrowth factor and nitric oxidein patients with colorectalcarcinoma.Cytokine,2002,20:184–190
[14] Takcacki K, Yamaguchi A, Urano T. et al. Expression of CD44variant exon8-10in colorectal cancer and its relationship to metastasis [J]. Jpn J CancerRes,1995,86(3):292-297
[15]Bendardaf R,Elzagheid A, E-cadherin, CD44s and CD44v6correlate with tumourdiffer-entiation in colorectal cancer.Oncol Rep,2005,May;13(5):831-835
[16] Wheelock MJ．Johnson KR， Cadherins as modulators of cellular phenotype．AnnuRev， Cell Dev Biol,2003,19:207-235
[17] Eidelman S,Caroline H.Expression of the cell-cell adhesion glycoproteincell-CAM120/80in normal human tissue and tumors[J].Am JPathol,1989,135:101-110
[18] Chalmerl J，Hofler H，Atkinson MJ．Mapping of a Cadherin gene claster to aregin of chromosome5subject to frequent allelic loss in carcinoma[J].Genomics,1999,57(1):160-163
[19] Berx G，Van Roy F，The E-cadherin／catenin complex：an important gatekeeperin breast cancer tuniorigenesis and malignant progression Breast CancerRes,2001,3(5)：289-293
[20] Bondi J,Bukholm G, An increase in the number of adhesion proteins with alteredexpression is associated with an increased risk of cancer death for coloncarcinoma patients.[J].Int J Colorectal Dis,2006,21(3):231-237
[21] Zamzani N，Marchetti P，Castedo M，et al.Reduction in mtochondrial potentialconstitutes an early irreversible step of programmed lymphacyte death in vivo.JExp Med，1995，181:1661-1672
[22] Petit PX，LeCouer H，Zorn E，et al.Alterations of mitochondrials structure andfunction are early events of dexamethasone-induced thymocyte apoptosis.J CellBiol.1995，130:156-167
[23] Baggeto LG.ROle of mitochondria in carcinogenesis.EurJCancer，1993;29A:156-159
[24] Okochi O，Hibi K，Uemura T，et al.Detection of mitochondrial DNA alterationsin the serum of hepatocellular carcinoma patients.Clin CancerRes.2002，8:2875-2878
[25] Shay JW，Werbin H.New evidence for insertion of mitochondrial DNAsequencesinto the human genome:significance for cancer and aging.Mutation·26·Res.1992，275:227-235
[26] Uiang BC.Evidence for association of mitochondrial DNA sequenceamplificantion and nuclear localization in human low-grade gliomas.MutationRes，1996，354:27-33
[27] KirchesE，Mawrin C，Schneider-Stoek R，et al.Mitochondrial DNA as a clonaltumor cell marker:gliomatosis cerebri.J Neurooncol.2003，61(1):1-5
[28] Tong BC,Ha PK,Dhir K,et al.Mitochondrial DNAalterations in thyroid cancer.JSurg oncol.2003,82:170-173
[29] Blanc H，Adams CW，Wallace DC，et al.Different nucleotide changes in the largerRNA gene of the mitochondrial DNA confer chloramphenicol resistance on twohuman cell lines.Nucleic Acids Res，1981，9:5785-5795
[30] Tate PH，Bird AP.Effects of DNA methylation on DNA-binding proteins andgene expression.Curr Opin Genet Dev.1993:3:226-231.
[31] Robertson KD，Uzvolgyi E，Liang G Talmadge C，Sumegi J，Gonzales FA，Jones PA.Thehuman DNA methyltransferases(DNMTs)l，3a and3b:coordinatemRNA expression in normal tissues and overexpression in tumors.Nucleic AcidsRes.1999:27:2291-2298
[32] Smith HA, McNeel DG. The SSX family of cancer-testis antigens as targetproteins for tumor therapy [J]. Clin Dev Immunol,2010,150591.
[33] De Smet C，Lurquin C，Lethe B，et a1．DNA methylation is the primary silencingmechanism for a set of germ lineand tumor-specific genes with fl CpG-richpromoter[J]．Mol Cell Bi0l，1999，19:7327-7335
[34] GUREA O,WEI1Jm OLD LJ et a1.The SSX gene family:characterization of9complete genes[J].bit J Cancer,2002,l0l(5)：448453．
[35] Mintz B，ilmensee K．Normal genetically mosaic mice produced from malignantteratocarcinorna cels[J]．Proc NatlAcad Sci USA。1975，72：3585-3589
[36] Scanlan MJ,Gure AO,Jungbluth AA,et a1.Cancer/testis antigens:an expandingfamily of targets for cancer im-munotherapy[J].Immunol Rev,2002,188:22-32
[37]刘亦肖,张传军.肿瘤睾丸抗原CTA的研究进展[J].陕西师范大学学报,2006,34:128-132
[38] Ono T,Kurashige T,Harada N,et a1.Identification of proscrosin binding proteinsp32precursor as a human cancer/testis antigen[J].Proc Natl Acad Sci U S A,2001,98:3282-3287．
[39] Louknov DI,PugachevaE,VatdinS,eta1.BORIS,a novd male germ-linespecificprotein associ-ated with epigenetic reprogramming events,shares the same11-zinc finger domain with CTCF.the insulator protein involved in readingimprinting marks in the soma[J].Proc Natl Acad Sci USA,2002,99:6806-6811．
[40] Shan J,Yuan L,Xiao Q,et a1.TSP50,a possible protease in human testes,isactivated in breast CAMleer epithelial cels[J].Cancer Res,2002,62:290-294.
[41]蔡胜利,冷希圣,杜如墨.肿瘤特异性抗原MAGE家族的研究进展[J].中华外科杂志,2001.39(8).
[42] Aoki Carrell DT．Human protamines and the developing spermatid：theirstructure，function，expression and relationship with male infertility，Asian JAndrol2003；5(4)：315-324
[43] Choudhary SK，Wykes．A haploid expressed gene cluster exists as a singlechromatin domain in human sperm[J].J Biol Chem．1995．270(15)：8755-8762．
[44] Balhorn R, Reed S, Tanphaichit r N. Aberrant protamine1/protamine2ratios insperm of infertile human males [J].Experientia,1988,44(1):52-55.
[45] Word WS, Coffey DS. DNA packaging and orgnization in mammalianspermatozoa: comparision with somatic cells [J].Biol Reprod,1991,44(4):569-571.
[46] Bianchi F, Pula G. P2protamine from human sperm are Zincfinger proteins withone Cys2/His2motif [J]. BiochemBiophys Res Commum,1992,182(1):540-543.
[47] Nonchev S, Tsanev R. Protamine histone replacement and DNA replication inthe male mouse pronucleus [J]. Mol Reprod Dev,1990,25(1):72-76.
[48] Perreault SD, Barbee RR, Ecstein KH, et al. Interspecies differences in thestability of mammalian sperm nuclei assessed in vivo by sperm microinjectionand in vitro by flow cytometry [J]. Bio Reprod,1998,39(1):157-167.
[49] Hecht NB．Reguletion of haploid expressed genes in male germ cells ReprodFert·1990，88I679
[50]赵亚力,张思仲.精蛋白基因转录的组织和阶段发育特异性初步研究[J].中华医学遗传学杂志,1995,12(3):150-153
[51] Grimes SR,Meistrich ML,Platz RD,et a1. Nuclear protein transitions in rat testisspermatids.Exp Cell Res,1977.110：31-39
[52] Mascotti DP．LohmanTM．Thermodynamics of oligoarginines binding to RNAand DNA．Biochemistry．1997．36：7272-7279
[53] Dadoune JP:The nuclear status of human sperm cells.Micron,1995,26:323.
[54] DAI Wen-ping,LIU Ping.HAN Yu-hai·CHEN Xi-ao-mei．FEI Ren-ren&XUE
[55] FEI Ren-ren.CHEN Hui.XIONG Hui-fang.wAN Qi-zhi.FENG Jian-fang，ZHANG Jing-bo＆.Xue She-pu.The studies on human sperm nucleus-analysisof sperm nuclear basic proteins in100infertile men.Acta Anatomica Sinica(inChinese).1996,27(3):256-258．
[56] LI Jian-guo.ZHU Jing-song.wU Ming-zhang· WANG Yi-fei,FEI Ren-ren＆.CHEN Hui.Elec-trophoretic analysis of nuclear proteins of ejaculated spermsfrom spouses of women with unexplained spon-taneous abortion．Reproductionand Contraception in (Chinese),1996,l6(1):59-64．
[57] Balhorn R.Reed S＆.Tanphaichitr N.Aberrant prota-nine1/protamine2ratios insperm of infertile human males.Experientia,1988,44:52.
[58] Olo C,Jung-Ha H,WilF~WD,eta1.Protanfme2deficiency leads to sperm DNAdamage and embryo death in mice. Biol Reprod2003,69:211-217
[59] ChoC，WillisWD，GouldingEH，etaLHaploinsuficiency of protamine-1or-2causes infertility in mice．Nat Genet2001；28：82-86
[60]吴小芳,陈晖,陈松,等.人与大小鼠精子中精蛋白mRNA的分析[J].生殖与避孕2001,21(4):200-205
[61] Tanaka H, Miyagawa Y, Tsujimura A, Matsumiya K, Okuyama A andNishimune Y (2003) Single nucleotide polymorphisms in the protamine-1and-2genes of fertile and infertile human male populations. Mol Hum Reprod9:69–73.
[62] Schlicker M, Schnulle V, Schneppel L, Vorob’ev VI and Engel W (1994)Disturbances of nuclear condensation in human spermatozoa: search for mutations inthe genes for protamine1, protamine2and transition protein1. Hum Reprod9:2313–2317.
[63] Iguchi N, Yang S, Lamb DJ and Hecht NB (2005) A protamine SNP: one geneticcause of male infertility? J Medical Genet [Advanced online publication30September2005].
[64] Miyagawa Y, Nishimura H, Tsujimura A, Matsuoka Y, Matsumiya K, OkuyamaA, Nishimune Y and Tanaka H (2005) Single-nucleotide polymorphisms andmutation analyses of the TNP1and TNP2genes of fertile and infertile humanmale populations.[J] Androl26:779–786.
[65] Shirley CR, Hayashi S, Mounsey S, Yanagimachi R and Meistrich ML (2004)Abnormalities and reduced reproductive potential of sperm from Tnp1-andTnp2-null double mutant mice. Biol Reprod71:1220–1229.
[66] Suganuma R, Yanagimachi R and Meistrich ML (2005) Decline in fertility ofmouse sperm with abnormal chromatin during epididymal passage as revealed byICSI. Hum Reprod20:3101–3108.
[67] Sliverst roni L. Histone instead of protamine interminal germ cells of infertileoligospermic men [J]. Fertil Steril,1976,27(3):1428-1430.
[68]陈宇,费仁仁,吴小芳.大鼠精蛋白基因在MEL细胞中的表达[J].中国医学科学院学报2002,24(6):601-605
[69] Gupta G,Sharma R,Chattopadhyay TK,et a1.Clinical sig-nificance of spermprotein17expression and immunogenicity in esophageal cancer[J].Int J Cancer,2007,120(8):1739-1748
[70] Chen YT,Scanlan MJ.A testicular antigen aberrant-ly expressed in humancancers detected by autolo-gous screening[J].Proc Natl Acad Sci USA,1997,94(5):1914-1918
[71]滕小梅,韩泽广,黄健. PRM1，一个新的候选结肠癌相关CT抗原基因的识别及其在结肠癌组织中的表达分析[J].现代检验医学杂志,2008,23(3):1-4
[72] Phillips RK, Quill DS, Dudley HA.Experimental colon neoplasia enhanced byextract of sperm (protamine)1[J]. J R Soc Med..77(10):830-2
[2] Frattini M, Balestra D, Suardi S, et al. Different genetic featuresassociated withcolon and rec-tal carcinogenesis［J］. Clin CancerRes,2004,10(12):4015-4021.
[3]徐建明,正确认识结肠癌和直肠癌诊治理念的差异[J].中国实用外科杂志,2012,32(9):703-705
[4] Bernstein LR, Liotta LA. Molecular mediator of interactions withextracelluarmatrix components in metastasis and angiogenesis. Curr Opin Oncol,1994,6(1):106-113.
[5] Skinner SA，Frydman GM, Obrif PE, et al. Microvascular structure of benign andmalignan tumor of the colon in humans. Dig Dis Sci,1995,40(2):373-384.
[6] Krishnan J, Kirkin V, Steffen A, et al. Differential in vitro expression of vascularendothelial growth factor(VEGF)-C and VEGF-D in tumors and its relationshipto lymphatic metastasis in immunocompetents rats.Cancer Ras,2003,63:713-722.
[7]曾益新.肿瘤学[M].北京:人民卫生出版社,第二版,2003:309
[8] Ferrar N，Henzel WJ．Pituitary follicular cell secrete anovel heparin bindinggrowth factors pecific for vascular endothelial cells[J]．Biochem Biophys ResCommun，1989，161(2)：851
[9] Iljin K,Karkkanen MJ, Lawrence EC, et al. VEGFR3gene structure, regulatoryregion, and sequence ploy morphisms [J]. FASEB J.2001,15(6):1028-1036
[10] Yancopoulos GD,Davis S, Gale NW, et al. Vascular specific growth factors andblood vessel formation [J]. Nature,2000,407:242-248
[11] Ferrara N, Gerber HP, LeCouter J,et al. The biology of VEGF and itsreceptors.Nat Med,2003,9:669–676
[12] Werther K, Christensen IJ, Brunner N,Nielsen HJ,et al, Soluble vascularendothelial growth factor levels in patients with primary colorectalcarcinoma.The Danish RANX05Colorectal Cancer Study Group. Eur J SurgOncol,2000,26:657–662
[13] Akbulut H, Altuntas F, Akbulut KG,Ozturk G, Cindoruk M, Unal E, Icli F,etal.Prognostic role of serum vascular endothelial growth factor, basic fibroblastgrowth factor and nitric oxidein patients with colorectalcarcinoma.Cytokine,2002,20:184–190
[14] Takcacki K, Yamaguchi A, Urano T. et al. Expression of CD44variant exon8-10in colorectal cancer and its relationship to metastasis [J]. Jpn J CancerRes,1995,86(3):292-297
[15]Bendardaf R,Elzagheid A, E-cadherin, CD44s and CD44v6correlate with tumourdiffer-entiation in colorectal cancer.Oncol Rep,2005,May;13(5):831-835
[16] Wheelock MJ．Johnson KR， Cadherins as modulators of cellular phenotype．AnnuRev， Cell Dev Biol,2003,19:207-235
[17] Eidelman S,Caroline H.Expression of the cell-cell adhesion glycoproteincell-CAM120/80in normal human tissue and tumors[J].Am JPathol,1989,135:101-110
[18] Chalmerl J，Hofler H，Atkinson MJ．Mapping of a Cadherin gene claster to aregin of chromosome5subject to frequent allelic loss in carcinoma[J].Genomics,1999,57(1):160-163
[19] Berx G，Van Roy F，The E-cadherin／catenin complex：an important gatekeeperin breast cancer tuniorigenesis and malignant progression Breast CancerRes,2001,3(5)：289-293
[20] Bondi J,Bukholm G, An increase in the number of adhesion proteins with alteredexpression is associated with an increased risk of cancer death for coloncarcinoma patients.[J].Int J Colorectal Dis,2006,21(3):231-237
[21] Zamzani N，Marchetti P，Castedo M，et al.Reduction in mtochondrial potentialconstitutes an early irreversible step of programmed lymphacyte death in vivo.JExp Med，1995，181:1661-1672
[22] Petit PX，LeCouer H，Zorn E，et al.Alterations of mitochondrials structure andfunction are early events of dexamethasone-induced thymocyte apoptosis.J CellBiol.1995，130:156-167
[23] Baggeto LG.ROle of mitochondria in carcinogenesis.EurJCancer，1993;29A:156-159
[24] Okochi O，Hibi K，Uemura T，et al.Detection of mitochondrial DNA alterationsin the serum of hepatocellular carcinoma patients.Clin CancerRes.2002，8:2875-2878
[25] Shay JW，Werbin H.New evidence for insertion of mitochondrial DNAsequencesinto the human genome:significance for cancer and aging.Mutation·26·Res.1992，275:227-235
[26] Uiang BC.Evidence for association of mitochondrial DNA sequenceamplificantion and nuclear localization in human low-grade gliomas.MutationRes，1996，354:27-33
[27] KirchesE，Mawrin C，Schneider-Stoek R，et al.Mitochondrial DNA as a clonaltumor cell marker:gliomatosis cerebri.J Neurooncol.2003，61(1):1-5
[28] Tong BC,Ha PK,Dhir K,et al.Mitochondrial DNAalterations in thyroid cancer.JSurg oncol.2003,82:170-173
[29] Blanc H，Adams CW，Wallace DC，et al.Different nucleotide changes in the largerRNA gene of the mitochondrial DNA confer chloramphenicol resistance on twohuman cell lines.Nucleic Acids Res，1981，9:5785-5795
[30] Tate PH，Bird AP.Effects of DNA methylation on DNA-binding proteins andgene expression.Curr Opin Genet Dev.1993:3:226-231.
[31] Robertson KD，Uzvolgyi E，Liang G Talmadge C，Sumegi J，Gonzales FA，Jones PA.Thehuman DNA methyltransferases(DNMTs)l，3a and3b:coordinatemRNA expression in normal tissues and overexpression in tumors.Nucleic AcidsRes.1999:27:2291-2298
[32] Smith HA, McNeel DG. The SSX family of cancer-testis antigens as targetproteins for tumor therapy [J]. Clin Dev Immunol,2010,150591.
[33] De Smet C，Lurquin C，Lethe B，et a1．DNA methylation is the primary silencingmechanism for a set of germ lineand tumor-specific genes with fl CpG-richpromoter[J]．Mol Cell Bi0l，1999，19:7327-7335
[34] GUREA O,WEI1Jm OLD LJ et a1.The SSX gene family:characterization of9complete genes[J].bit J Cancer,2002,l0l(5)：448453．
[35] Mintz B，ilmensee K．Normal genetically mosaic mice produced from malignantteratocarcinorna cels[J]．Proc NatlAcad Sci USA。1975，72：3585-3589
[36] Scanlan MJ,Gure AO,Jungbluth AA,et a1.Cancer/testis antigens:an expandingfamily of targets for cancer im-munotherapy[J].Immunol Rev,2002,188:22-32
[37]刘亦肖,张传军.肿瘤睾丸抗原CTA的研究进展[J].陕西师范大学学报,2006,34:128-132
[38] Ono T,Kurashige T,Harada N,et a1.Identification of proscrosin binding proteinsp32precursor as a human cancer/testis antigen[J].Proc Natl Acad Sci U S A,2001,98:3282-3287．
[39] Louknov DI,PugachevaE,VatdinS,eta1.BORIS,a novd male germ-linespecificprotein associ-ated with epigenetic reprogramming events,shares the same11-zinc finger domain with CTCF.the insulator protein involved in readingimprinting marks in the soma[J].Proc Natl Acad Sci USA,2002,99:6806-6811．
[40] Shan J,Yuan L,Xiao Q,et a1.TSP50,a possible protease in human testes,isactivated in breast CAMleer epithelial cels[J].Cancer Res,2002,62:290-294.
[41]蔡胜利,冷希圣,杜如墨.肿瘤特异性抗原MAGE家族的研究进展[J].中华外科杂志,2001.39(8).
[42] Aoki Carrell DT．Human protamines and the developing spermatid：theirstructure，function，expression and relationship with male infertility，Asian JAndrol2003；5(4)：315-324
[43] Choudhary SK，Wykes．A haploid expressed gene cluster exists as a singlechromatin domain in human sperm[J].J Biol Chem．1995．270(15)：8755-8762．
[44] Balhorn R, Reed S, Tanphaichit r N. Aberrant protamine1/protamine2ratios insperm of infertile human males [J].Experientia,1988,44(1):52-55.
[45] Word WS, Coffey DS. DNA packaging and orgnization in mammalianspermatozoa: comparision with somatic cells [J].Biol Reprod,1991,44(4):569-571.
[46] Bianchi F, Pula G. P2protamine from human sperm are Zincfinger proteins withone Cys2/His2motif [J]. BiochemBiophys Res Commum,1992,182(1):540-543.
[47] Nonchev S, Tsanev R. Protamine histone replacement and DNA replication inthe male mouse pronucleus [J]. Mol Reprod Dev,1990,25(1):72-76.
[48] Perreault SD, Barbee RR, Ecstein KH, et al. Interspecies differences in thestability of mammalian sperm nuclei assessed in vivo by sperm microinjectionand in vitro by flow cytometry [J]. Bio Reprod,1998,39(1):157-167.
[49] Hecht NB．Reguletion of haploid expressed genes in male germ cells ReprodFert·1990，88I679
[50]赵亚力,张思仲.精蛋白基因转录的组织和阶段发育特异性初步研究[J].中华医学遗传学杂志,1995,12(3):150-153
[51] Grimes SR,Meistrich ML,Platz RD,et a1. Nuclear protein transitions in rat testisspermatids.Exp Cell Res,1977.110：31-39
[52] Mascotti DP．LohmanTM．Thermodynamics of oligoarginines binding to RNAand DNA．Biochemistry．1997．36：7272-7279
[53] Dadoune JP:The nuclear status of human sperm cells.Micron,1995,26:323.
[54] DAI Wen-ping,LIU Ping.HAN Yu-hai·CHEN Xi-ao-mei．FEI Ren-ren&XUE
[55] FEI Ren-ren.CHEN Hui.XIONG Hui-fang.wAN Qi-zhi.FENG Jian-fang，ZHANG Jing-bo＆.Xue She-pu.The studies on human sperm nucleus-analysisof sperm nuclear basic proteins in100infertile men.Acta Anatomica Sinica(inChinese).1996,27(3):256-258．
[56] LI Jian-guo.ZHU Jing-song.wU Ming-zhang· WANG Yi-fei,FEI Ren-ren＆.CHEN Hui.Elec-trophoretic analysis of nuclear proteins of ejaculated spermsfrom spouses of women with unexplained spon-taneous abortion．Reproductionand Contraception in (Chinese),1996,l6(1):59-64．
[57] Balhorn R.Reed S＆.Tanphaichitr N.Aberrant prota-nine1/protamine2ratios insperm of infertile human males.Experientia,1988,44:52.
[58] Olo C,Jung-Ha H,WilF~WD,eta1.Protanfme2deficiency leads to sperm DNAdamage and embryo death in mice. Biol Reprod2003,69:211-217
[59] ChoC，WillisWD，GouldingEH，etaLHaploinsuficiency of protamine-1or-2causes infertility in mice．Nat Genet2001；28：82-86
[60]吴小芳,陈晖,陈松,等.人与大小鼠精子中精蛋白mRNA的分析[J].生殖与避孕2001,21(4):200-205
[61] Tanaka H, Miyagawa Y, Tsujimura A, Matsumiya K, Okuyama A andNishimune Y (2003) Single nucleotide polymorphisms in the protamine-1and-2genes of fertile and infertile human male populations. Mol Hum Reprod9:69–73.
[62] Schlicker M, Schnulle V, Schneppel L, Vorob’ev VI and Engel W (1994)Disturbances of nuclear condensation in human spermatozoa: search for mutations inthe genes for protamine1, protamine2and transition protein1. Hum Reprod9:2313–2317.
[63] Iguchi N, Yang S, Lamb DJ and Hecht NB (2005) A protamine SNP: one geneticcause of male infertility? J Medical Genet [Advanced online publication30September2005].
[64] Miyagawa Y, Nishimura H, Tsujimura A, Matsuoka Y, Matsumiya K, OkuyamaA, Nishimune Y and Tanaka H (2005) Single-nucleotide polymorphisms andmutation analyses of the TNP1and TNP2genes of fertile and infertile humanmale populations.[J] Androl26:779–786.
[65] Shirley CR, Hayashi S, Mounsey S, Yanagimachi R and Meistrich ML (2004)Abnormalities and reduced reproductive potential of sperm from Tnp1-andTnp2-null double mutant mice. Biol Reprod71:1220–1229.
[66] Suganuma R, Yanagimachi R and Meistrich ML (2005) Decline in fertility ofmouse sperm with abnormal chromatin during epididymal passage as revealed byICSI. Hum Reprod20:3101–3108.
[67] Sliverst roni L. Histone instead of protamine interminal germ cells of infertileoligospermic men [J]. Fertil Steril,1976,27(3):1428-1430.
[68]陈宇,费仁仁,吴小芳.大鼠精蛋白基因在MEL细胞中的表达[J].中国医学科学院学报2002,24(6):601-605
[69] Gupta G,Sharma R,Chattopadhyay TK,et a1.Clinical sig-nificance of spermprotein17expression and immunogenicity in esophageal cancer[J].Int J Cancer,2007,120(8):1739-1748
[70] Chen YT,Scanlan MJ.A testicular antigen aberrant-ly expressed in humancancers detected by autolo-gous screening[J].Proc Natl Acad Sci USA,1997,94(5):1914-1918
[71]滕小梅,韩泽广,黄健. PRM1，一个新的候选结肠癌相关CT抗原基因的识别及其在结肠癌组织中的表达分析[J].现代检验医学杂志,2008,23(3):1-4
[72] Phillips RK, Quill DS, Dudley HA.Experimental colon neoplasia enhanced byextract of sperm (protamine)1[J]. J R Soc Med..77(10):830-2