c-met在良性，癌前和恶性外阴肿瘤中的表达及意义
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摘要
原癌基因C-MET属于具酪氨酸激酶活性的生长因子受体类，编码肝细胞生长因子／离散因子(HGF／SF)受体，是细胞内信息传导的重要组分。目前已证实，C-MET在肿瘤的发展、浸润和转移过程中发挥重要作用，它的过度表达在胃癌、乳腺癌、前列腺癌、胰腺癌、甲状腺癌、肝癌、子宫内膜癌等腺癌相对报道较多，而鳞癌报道较少如食管鳞癌、口腔鳞癌、喉癌、头颈鳞癌、子宫颈癌等。然而，C-MET基因表达与外阴癌变关系的研究国内外尚未见报道。
目的
通过检测c-met基因蛋白在良性外阴尖锐湿疣(VCA)、癌前病变的外阴上皮内瘤变(VIN)、癌旁外阴营养不良(VD)和恶性外阴鳞状细胞癌(VSCC)这一组外阴病变中的表达情况来阐明c-met在外阴癌变中的作用，并进一步分析其表达与外阴鳞癌患者的临床病理特征(年龄、临床分期、组织分化程度及淋巴结状态)之间的关系。
材料和方法
对象：选取中国医科大学第一附属医院妇科1995—2003年间收治病例，外阴鳞癌30例，外阴上皮内瘤变25例，癌旁外阴营养不良20例(包括增生型和硬化苔藓型各10例)，尖锐湿疣15例，5例正常外阴表皮作对照。(所有标本组织均为福尔马林固定，石蜡包埋)。试剂：采用亲和纯化的兔多克隆抗c-met抗体，工作浓度为1：200。应用链酶亲和素—过氧化物酶复合物免疫组化染色方法(S-ABC方法)。采用卡方检验，对c-met蛋白在不同外阴病变中的表达及与外阴鳞癌的临床病理特征的关系进行统计学分析，P＜0.05为差异有显著性。
结果
c-met表达在80％(24／30)的VSCC，阳性细胞百分率为70.3％；20％
(万10)的VINlll，阳性细胞百分率为17.4%;26.7%(4/巧)的vcA中，阳
性细胞百分率为54.3%。无表达于VINI一11和正常外阴表皮。在VSCC
中，癌旁增生型营养不良c一met表达阳性病例百分数为80%(8/10)，阳性
细胞百分率为43 .8%，癌旁硬化苔醉型营养不良无表达。从巩Nl一n经
过VINm到VSCC，。一met表达具有统计学意义上的差异(P<0 .05)。c一
met抗体的免疫反应在VSCC中较强，在viNnl中较弱。。一met更强表达
于高分化型的VsCC中(阳性细胞百分率为68 .7%)，与中/低分化型(阳性
细胞百分率分别为33 .1%和30.3%)相比具有统计学意义(P<0.05)。在
vscC中，c一met表达与年龄、临床分期和淋巴结转移无统计学意义上的相
关性(P>0 .05)。
结论
1.本研究发现在正常外阴表皮，未见c一met的表达，提示C一met没有
参与正常外阴鳞状上皮细胞的生长和分化过程。
2.。一met在外阴鳞癌、癌旁的增生型营养不良、外阴上皮内瘤变viNlll
及外阴尖锐湿庆中均可见阳性表达，提示无论细胞有无异型性，。一met的
表达与细胞的增殖活跃有关。
3.在外阴鳞癌中c一met表达的阳性细胞数及免疫反应强度高于vi-
Mn，在vlNI一n和正常外阴表皮中无表达，提示C一met的表达与外阴鳞
状细胞恶性转变有关，是外阴癌前病变的晚期事件。
4.高分化VSCC中C一met表达明显高于中/低分化，说明C一met表达
与组织分化程度有关，c一met表达与VSCC恶性进展呈负相关。
The c - met oncogene belongs to a member of growth factor families, it encodes the hepatocyte growth factor /scatter factor( HGF/SF) receptor and has the tyrosine - kinase activity. It is one of the important parts in the cellular information transmitting function, c - met has been shown to fufill a number of important roles in tumor progression, invasion and metastasis. Expression of c - met in human adenocarcinoma is well described, such as gastric, breast, prostatic, pancreatic, thyroid, heaptocellular and endomentrial carcinomas. But few studies on its expression in squamous cell carcinomas have been reported, such as esophageal, oral,laryngeal,head and neck,uterine cervix carcinomas,and its expression in vulvar squamous cell carcinoma has not previously been reported.
OBJECTIVE
To clarify the role of c - met in vulvar carcinogenesis through examination of its expression in a spectrum of vulvar lesions ranging from benign vulvar con-dyloma acuminate (VCA) through premalignant vulvar intraepithelial neoplasia (VIN) and vulvar dystrophy{ VD) near the carcinoma to malignant vulvar squamous cell carcinomas (VSCC) conditions and to analyze the relationship between c - met expression and clinicalpathologic features of vulvar squamous carcinomas including age,clinical stage,histopathologic tumor grade and lymph node status.
MATERIALS AND METHODS
The subjects were Thirty vulvar squamous cell carcinomas( VSCC ) , Twenty - five vulvar intraepithelial neoplasia ( VIN), Twenty vulvar dystrophy ( VD)
near the carcinoma including ten hyperplastic dystrophy ( HD) and ten Lichen sclerosis(LS) ,Fifteen vulvar condyloma acuminata( VCA)from the department of gynecological pathology at the first affiliated hospital of China Medical University between 1995 and 2003. For the controls,5 specimens of normal vulvar skin were used. ( all the tissues were formalin fixed and paraffin - embeded). All the specimens were detected by the technique of the immunohistochemical S - ABC with the employed affinity - purified rabbit polyclonal anti - c - met antibody (1 ;200). For statistical analysis,the chi - square test were used, c - met expression was compared with clinicalpathologic features of VSCC. P value less than 0.05 were regarded as significant.
RESULTS
c -met was expressed in 80% (24/30) of VSCC,The positive cells rate were 70. 3% ;20% (2/10) of VINIII ,The posive ceUs rate were 17. 4% ;26. 7% (4/15) of VCA,The positive cells rate were 54. 3% , and was undetectable in VINI - II and normal epithelium. In VSCC, c - met was expressed in 80% ( 8/ 10) of hyperplastic dystrophy (HD) near the carcinoma, The positive cells rate were 43. 8% , and was undetectable in lichen sclerosis( LS). c - met expression were statistically significant differences from VINI - II through VINIII to VSCC ( p < 0.05 ). The immunoreactivity to c - met antibody was strong in VSCC, and weak in VIN Iff. c - met was more intensely expressed in well differentiated VSCC (The positive cell rate were 68.7% ) than moderately and poorly differentiated VSCC (The positive cell rate were 33. 1% and 30. 3% ) ( p <0. 05). There was no statistically significant correlation between c - met expression and age,clinical stage and lymph node metastasis in VSCC(P >0.05).
CONCLUSIONS
1. We found in normal vulvar epithelium, c - met expression was absent which suggested that c - met is not involved in growth and differentiation of normal vulvar squamous epithelial cells.
2. c - met was expressed in VSCC , VD, VIN III and VCA,From which we can see c - met expression is associated with active proliferation of cells no mat-ter whether they have different characters.
3. The positive cells rate and immunoreactivity degree to c - met antibody were higher in VSCC than in VIN Iff and undetectable in VINI - II and normal epithelium which suggested c - met gene expression may be related to vulvar squamous cell malignant transformation and may be a late event in vulvar prema-lignant lesion.
4. c - met was more intensely expressed in well differentiated VSCC than
目的
通过检测c-met基因蛋白在良性外阴尖锐湿疣(VCA)、癌前病变的外阴上皮内瘤变(VIN)、癌旁外阴营养不良(VD)和恶性外阴鳞状细胞癌(VSCC)这一组外阴病变中的表达情况来阐明c-met在外阴癌变中的作用，并进一步分析其表达与外阴鳞癌患者的临床病理特征(年龄、临床分期、组织分化程度及淋巴结状态)之间的关系。
材料和方法
对象：选取中国医科大学第一附属医院妇科1995—2003年间收治病例，外阴鳞癌30例，外阴上皮内瘤变25例，癌旁外阴营养不良20例(包括增生型和硬化苔藓型各10例)，尖锐湿疣15例，5例正常外阴表皮作对照。(所有标本组织均为福尔马林固定，石蜡包埋)。试剂：采用亲和纯化的兔多克隆抗c-met抗体，工作浓度为1：200。应用链酶亲和素—过氧化物酶复合物免疫组化染色方法(S-ABC方法)。采用卡方检验，对c-met蛋白在不同外阴病变中的表达及与外阴鳞癌的临床病理特征的关系进行统计学分析，P＜0.05为差异有显著性。
结果
c-met表达在80％(24／30)的VSCC，阳性细胞百分率为70.3％；20％
(万10)的VINlll，阳性细胞百分率为17.4%;26.7%(4/巧)的vcA中，阳
性细胞百分率为54.3%。无表达于VINI一11和正常外阴表皮。在VSCC
中，癌旁增生型营养不良c一met表达阳性病例百分数为80%(8/10)，阳性
细胞百分率为43 .8%，癌旁硬化苔醉型营养不良无表达。从巩Nl一n经
过VINm到VSCC，。一met表达具有统计学意义上的差异(P<0 .05)。c一
met抗体的免疫反应在VSCC中较强，在viNnl中较弱。。一met更强表达
于高分化型的VsCC中(阳性细胞百分率为68 .7%)，与中/低分化型(阳性
细胞百分率分别为33 .1%和30.3%)相比具有统计学意义(P<0.05)。在
vscC中，c一met表达与年龄、临床分期和淋巴结转移无统计学意义上的相
关性(P>0 .05)。
结论
1.本研究发现在正常外阴表皮，未见c一met的表达，提示C一met没有
参与正常外阴鳞状上皮细胞的生长和分化过程。
2.。一met在外阴鳞癌、癌旁的增生型营养不良、外阴上皮内瘤变viNlll
及外阴尖锐湿庆中均可见阳性表达，提示无论细胞有无异型性，。一met的
表达与细胞的增殖活跃有关。
3.在外阴鳞癌中c一met表达的阳性细胞数及免疫反应强度高于vi-
Mn，在vlNI一n和正常外阴表皮中无表达，提示C一met的表达与外阴鳞
状细胞恶性转变有关，是外阴癌前病变的晚期事件。
4.高分化VSCC中C一met表达明显高于中/低分化，说明C一met表达
与组织分化程度有关，c一met表达与VSCC恶性进展呈负相关。
The c - met oncogene belongs to a member of growth factor families, it encodes the hepatocyte growth factor /scatter factor( HGF/SF) receptor and has the tyrosine - kinase activity. It is one of the important parts in the cellular information transmitting function, c - met has been shown to fufill a number of important roles in tumor progression, invasion and metastasis. Expression of c - met in human adenocarcinoma is well described, such as gastric, breast, prostatic, pancreatic, thyroid, heaptocellular and endomentrial carcinomas. But few studies on its expression in squamous cell carcinomas have been reported, such as esophageal, oral,laryngeal,head and neck,uterine cervix carcinomas,and its expression in vulvar squamous cell carcinoma has not previously been reported.
OBJECTIVE
To clarify the role of c - met in vulvar carcinogenesis through examination of its expression in a spectrum of vulvar lesions ranging from benign vulvar con-dyloma acuminate (VCA) through premalignant vulvar intraepithelial neoplasia (VIN) and vulvar dystrophy{ VD) near the carcinoma to malignant vulvar squamous cell carcinomas (VSCC) conditions and to analyze the relationship between c - met expression and clinicalpathologic features of vulvar squamous carcinomas including age,clinical stage,histopathologic tumor grade and lymph node status.
MATERIALS AND METHODS
The subjects were Thirty vulvar squamous cell carcinomas( VSCC ) , Twenty - five vulvar intraepithelial neoplasia ( VIN), Twenty vulvar dystrophy ( VD)
near the carcinoma including ten hyperplastic dystrophy ( HD) and ten Lichen sclerosis(LS) ,Fifteen vulvar condyloma acuminata( VCA)from the department of gynecological pathology at the first affiliated hospital of China Medical University between 1995 and 2003. For the controls,5 specimens of normal vulvar skin were used. ( all the tissues were formalin fixed and paraffin - embeded). All the specimens were detected by the technique of the immunohistochemical S - ABC with the employed affinity - purified rabbit polyclonal anti - c - met antibody (1 ;200). For statistical analysis,the chi - square test were used, c - met expression was compared with clinicalpathologic features of VSCC. P value less than 0.05 were regarded as significant.
RESULTS
c -met was expressed in 80% (24/30) of VSCC,The positive cells rate were 70. 3% ;20% (2/10) of VINIII ,The posive ceUs rate were 17. 4% ;26. 7% (4/15) of VCA,The positive cells rate were 54. 3% , and was undetectable in VINI - II and normal epithelium. In VSCC, c - met was expressed in 80% ( 8/ 10) of hyperplastic dystrophy (HD) near the carcinoma, The positive cells rate were 43. 8% , and was undetectable in lichen sclerosis( LS). c - met expression were statistically significant differences from VINI - II through VINIII to VSCC ( p < 0.05 ). The immunoreactivity to c - met antibody was strong in VSCC, and weak in VIN Iff. c - met was more intensely expressed in well differentiated VSCC (The positive cell rate were 68.7% ) than moderately and poorly differentiated VSCC (The positive cell rate were 33. 1% and 30. 3% ) ( p <0. 05). There was no statistically significant correlation between c - met expression and age,clinical stage and lymph node metastasis in VSCC(P >0.05).
CONCLUSIONS
1. We found in normal vulvar epithelium, c - met expression was absent which suggested that c - met is not involved in growth and differentiation of normal vulvar squamous epithelial cells.
2. c - met was expressed in VSCC , VD, VIN III and VCA,From which we can see c - met expression is associated with active proliferation of cells no mat-ter whether they have different characters.
3. The positive cells rate and immunoreactivity degree to c - met antibody were higher in VSCC than in VIN Iff and undetectable in VINI - II and normal epithelium which suggested c - met gene expression may be related to vulvar squamous cell malignant transformation and may be a late event in vulvar prema-lignant lesion.
4. c - met was more intensely expressed in well differentiated VSCC than
引文
1. Liu Y, et al. The human hepatocyte growth factor receptor gene, complete structure lorganization and promoter characterization. Gene, 1998,215 (1): 159-169
2. Donald L, Vignae, Narsimhan R, et al. Hepatocyte growth factor (HGF) stimulates the tyrosine kinase activity of the reeepter encoded by the protoneogene C-met[J]. Oneogene 1991;6:501-504.
3. Cooper C S, park M, et al. Molecular cloning of a new transforming gene from a chemically transformed human cell line. Nature 1984;311:29-33.
4. Weidoer KM, Dicesare S, Sachs M, et al. Interaction between GabI and the cmet receptor tyrosine Kinase is responsible for epithelial morphogenesis; Nature, 1996,384(6605):173-176
5. Jinl, Fchs A, Sehnitt SJ, et al. Expression of scatter factor and c-met receptor in benign and malignant breast tissue, cancer 1997;79:749-760.
6. Kuniyasu H, Yasui W, Kitadai Y, et al. Frequent amplification of the C-met gene in scirrhous type stomach cancer. Biochem Biophys Res Commun. 1992; 189:227-232.
7. Nakashiro k, Hayashi Y, et al. Immunhistochemical expression of hepatocyte growth factor and c-met/HGF receper in benign and malignanthumanprostatetissue. OnolRep. 2003, Sep-Oct; 10 (5):1149-53.
8. Furukawa T, Duguid WP, Kobari M, et al. Hepatocyte growth factor and met receptor expression in human pancreatic carcinogenesis. AM J pathol, 1995; 147:889-95.
9. Iaggiora P, Lorenzato A, et al. The RON and MET Oncogene are Co -expressed in human ovarian carcinomas and cooperate in activating invasiveness. Exp Cell Res. 2003 Aug 15:288(2):382-9.
10. Di Renzo M. F, Olivero M, Ferros, et al. Overexpression of the c-met/HGF receptor gene in human thyroid carcinomas. Oneogene. 1992; 7:2549-2553.
11. Luic, Park M, Tsaom-s, et al. Overexpression of c-met proto on cogene but not epidermal growth factor receptor or c-met B -2 in primary human color-
ectal carcinomas. Oncogene. 1992;7:181-185.
12．宁宏，张劲松等．C-MET原癌基因MRNA在睑板腺癌中的表达．中华眼科杂志；2001：5．37(3)．
13. WagatsumaS, Konnor, Satos, et al. Tumor engiogenesis, hepatocyte growth factor and c-met expression in endomentrial carcinoma, cancer, 1998;82:520-530.
14. ProteH, TribouletP, et al. Over expression of stronelysin-3,Bm-40/sparc, and Met genes in human esophageal carcinoma. Clin cancer Res 1998 Jun;4 (6):1375-82.
15. HuYc, et al. Profiling of differentially expressed cancer-related genes in esophageal squamous cell carcinoma(ESCC) using human cancer cdna arrays. Overexpression of oncogene met correlates with tumor differentiation in ESCC. Clin cancer Res 2001,Nov;7(11):519-25.
16. Cortesina G, et al. Staging of head and neck squamous cell carcinoma using the met oncogene product as marker of tumor cell in lymph node metastasis. Int J cancer. 2000, May 20;89 (3):286-92.
17. Uchida D, Kawamata H, et al. Role of HGF/c-met system in invasion and metastasis of oral squamous cell carcinoma cells in vitro and its clinical significance. Int J Cancer 2001 Aug 15;93(4):489-96.
18. Sawatsubashi M, Sasatomi E, et al. Expression of c-met in laryngeal carcinoma. Virchows Arch 1998 Apr;432(4):331.
19. Baykal C ,Ayhan A, et al. Overexpression of the c-met/HGF receptor and its prognostic significance in uterine cervix carcinomas. Gynecol Oncol. 2003, Feb;88(2):123-9.
20. Shimabukuro K,Ichinose S,et al. Hepatocyte growth factor/scatter factor is implicated in the mode of stromal invasion of uterine sqamous cervical cancer. Gynecol Oncol. 2001,Nov:83(2):205-15.
21. Morivama T, Kataoka H, et al. Concomitant expression of hepatocyte growth factor( HGF), HGF activator and c-met genes in human gliomas in vitro. [J]. FEBS,lect, 1995;372;78-82。
22. Chao-nan Qian, xiang guo, et al. Met Protein Expression Level Correlates With Survival in Patients with Late-Stage Nasopharyngeal Carcinoma. Canc-
er Res. 62,589-596,Jan15,2002.
2. Donald L, Vignae, Narsimhan R, et al. Hepatocyte growth factor (HGF) stimulates the tyrosine kinase activity of the reeepter encoded by the protoneogene C-met[J]. Oneogene 1991;6:501-504.
3. Cooper C S, park M, et al. Molecular cloning of a new transforming gene from a chemically transformed human cell line. Nature 1984;311:29-33.
4. Weidoer KM, Dicesare S, Sachs M, et al. Interaction between GabI and the cmet receptor tyrosine Kinase is responsible for epithelial morphogenesis; Nature, 1996,384(6605):173-176
5. Jinl, Fchs A, Sehnitt SJ, et al. Expression of scatter factor and c-met receptor in benign and malignant breast tissue, cancer 1997;79:749-760.
6. Kuniyasu H, Yasui W, Kitadai Y, et al. Frequent amplification of the C-met gene in scirrhous type stomach cancer. Biochem Biophys Res Commun. 1992; 189:227-232.
7. Nakashiro k, Hayashi Y, et al. Immunhistochemical expression of hepatocyte growth factor and c-met/HGF receper in benign and malignanthumanprostatetissue. OnolRep. 2003, Sep-Oct; 10 (5):1149-53.
8. Furukawa T, Duguid WP, Kobari M, et al. Hepatocyte growth factor and met receptor expression in human pancreatic carcinogenesis. AM J pathol, 1995; 147:889-95.
9. Iaggiora P, Lorenzato A, et al. The RON and MET Oncogene are Co -expressed in human ovarian carcinomas and cooperate in activating invasiveness. Exp Cell Res. 2003 Aug 15:288(2):382-9.
10. Di Renzo M. F, Olivero M, Ferros, et al. Overexpression of the c-met/HGF receptor gene in human thyroid carcinomas. Oneogene. 1992; 7:2549-2553.
11. Luic, Park M, Tsaom-s, et al. Overexpression of c-met proto on cogene but not epidermal growth factor receptor or c-met B -2 in primary human color-
ectal carcinomas. Oncogene. 1992;7:181-185.
12．宁宏，张劲松等．C-MET原癌基因MRNA在睑板腺癌中的表达．中华眼科杂志；2001：5．37(3)．
13. WagatsumaS, Konnor, Satos, et al. Tumor engiogenesis, hepatocyte growth factor and c-met expression in endomentrial carcinoma, cancer, 1998;82:520-530.
14. ProteH, TribouletP, et al. Over expression of stronelysin-3,Bm-40/sparc, and Met genes in human esophageal carcinoma. Clin cancer Res 1998 Jun;4 (6):1375-82.
15. HuYc, et al. Profiling of differentially expressed cancer-related genes in esophageal squamous cell carcinoma(ESCC) using human cancer cdna arrays. Overexpression of oncogene met correlates with tumor differentiation in ESCC. Clin cancer Res 2001,Nov;7(11):519-25.
16. Cortesina G, et al. Staging of head and neck squamous cell carcinoma using the met oncogene product as marker of tumor cell in lymph node metastasis. Int J cancer. 2000, May 20;89 (3):286-92.
17. Uchida D, Kawamata H, et al. Role of HGF/c-met system in invasion and metastasis of oral squamous cell carcinoma cells in vitro and its clinical significance. Int J Cancer 2001 Aug 15;93(4):489-96.
18. Sawatsubashi M, Sasatomi E, et al. Expression of c-met in laryngeal carcinoma. Virchows Arch 1998 Apr;432(4):331.
19. Baykal C ,Ayhan A, et al. Overexpression of the c-met/HGF receptor and its prognostic significance in uterine cervix carcinomas. Gynecol Oncol. 2003, Feb;88(2):123-9.
20. Shimabukuro K,Ichinose S,et al. Hepatocyte growth factor/scatter factor is implicated in the mode of stromal invasion of uterine sqamous cervical cancer. Gynecol Oncol. 2001,Nov:83(2):205-15.
21. Morivama T, Kataoka H, et al. Concomitant expression of hepatocyte growth factor( HGF), HGF activator and c-met genes in human gliomas in vitro. [J]. FEBS,lect, 1995;372;78-82。
22. Chao-nan Qian, xiang guo, et al. Met Protein Expression Level Correlates With Survival in Patients with Late-Stage Nasopharyngeal Carcinoma. Canc-
er Res. 62,589-596,Jan15,2002.