摘要
目的：众多研究发现，肿瘤的生长、转移与肿瘤血管生成密切相关,而血管内皮生长因子（vascular endothelial growth factor, VEGF）是目前所证实的最有力、最具特征性的促血管生成因子。研究表明：许多人类恶性肿瘤包括乳腺癌组织中都发现VEGF的过度表达。本课题主要应用酶联免疫吸附实验（ELISA）研究乳腺癌患者手术前后血浆VEGF（P-VEGF）水平的变化情况，并与乳腺良性疾病患者及健康对照组进行比较，希望找到其变化规律，为乳腺癌的监测、尤其是转移灶的早期发现提供依据。本课题还利用免疫组化法比较研究了肿瘤组织中VEGF、ER、PR的表达情况。
方法：选择2002年12月至2004年1月在河北医科大学第四医院外科住院治疗的可手术的乳腺癌患者73例（浸润性导管癌39例，浸润性小叶癌28例，髓样癌5例，粘液腺癌1例），于手术前、手术后七天及三十天左右采用ELISA测量其P-VEGF水平。还选择有远处转移的乳腺癌（以下称转移性乳腺癌）患者9例，其中影像学发现有远处转移者6例，细胞学检查证实有远处淋巴结转移者3例，（9例中原发灶为浸润性小叶癌者3例，分别出现肺转移、肝转移和骨转移；原发灶为浸润性导管癌者1例，出现对侧腋窝淋巴结转移和局部复发；病理类型不详5例，分别出现肺转移，肝转移，肺肝转移，同侧锁骨上淋巴结转移，局部复发并同侧锁骨上淋巴结转移）。选择乳腺良性疾病患者22例（乳腺囊性
增生病7例，乳腺纤维腺瘤6例，乳腺腺病5例，导管内乳头状瘤病2例，乳腺炎伴脓肿形成1例，乳管扩张伴慢性炎症1例），健康对照组18例，用ELISA测量其P-VEGF水平，对所得结果进行比较分析。用免疫组化法检测肿瘤组织中VEGF、ER、PR的表达情况，并进行统计分析。
结果
1转移性乳腺癌患者P-VEGF浓度的范围23.7pg/ml～445pg/ml，中位数94.9pg/ml；可手术的乳腺癌患者P-VEGF浓度的范围1.84pg/ml～229pg/ml，中位数17.00pg/ml；乳腺良性疾病患者P-VEGF浓度的范围3.81pg/ml～30.4pg/ml，中位数8.89pg/ml；健康对照组P-VEGF浓度的范围1.53pg/ml～32.6pg/ml，中位数5.62pg/ml。最高值是一例病理类型不详，有肺肝转移的患者，其P-VEGF浓度为445pg/ml。转移性乳腺癌患者P-VEGF浓度明显高于可手术的乳腺癌、乳腺良性疾病患者和对照组（p值均小于0.05）。可手术的乳腺癌患者P-VEGF浓度高于健康对照组（p<0.05），但与乳腺良性疾病比较无统计学差异（p>.05）。乳腺良性疾病与健康对照组比较P-VEGF水平无统计学差异（p>0.05）。
2浸润性导管癌患者P-VEGF浓度（即手术前的浓度）的范围3.90pg/ml～229pg/ml，中位数22.19pg/ml；浸润性小叶癌患者P-VEGF浓度（即手术前的浓度）的范围是1.84pg/ml～45.9pg/ml，中位数8.79pg/ml。乳腺髓样癌患者P-VEGF浓度（即手术前的浓度）的范围6.74pg/ml～143.4pg/ml，中位数13.7pg/ml。浸润性导管癌患者P-VEGF水平明显高于浸润性小叶癌患者、良性疾病组及健康对照组（p值均小于0.05），与髓样癌比较无统计学差异（p>0.05）。
浸润性小叶癌患者P-VEGF水平与髓样癌、良性疾病组和健康对照组比较均无统计学差异（p值均大于0.05）。髓样癌患者P-VEGF水平高于健康对照组（p<0.05），但与良性疾病比较无统计学差异（p>.05）。转移性乳腺癌患者的P-VEGF浓度高于浸润性导管癌患者(p=0.000)、浸润性小叶癌患者(p=0.000)和髓样癌患者（p=0.028）。而且浸润性小叶癌远处转移的患者P-VEGF水平明显高于可手术的浸润性小叶癌患者的手术前水平(p=0.009)。
3乳腺浸润性导管癌患者手术后七天左右P-VEGF浓度（范围3.96pg/ml～209pg/ml，中位数19.47pg/ml）与手术前及良性疾病组比较差别均无统计学意义（p>0.05）,但高于健康对照组（p<0.05）。手术后三十天左右P-VEGF浓度（范围1.98pg/ml～87pg/ml，中位数5.86pg/ml）低于手术前和手术后七天左右的水平（p值均小于0.05），与良性疾病组和健康对照组比较无统计学差异（p值均大于0.05）。
4乳腺浸润性小叶癌患者手术后七天左右P-VEGF浓度（范围7.81pg/ml～174.5pg/ml，中位数13.2pg/ml）与手术前及良性疾病组比较均无统计学差异（p值均大于0.05）,但要高于健康对照组（p<0.05）。手术后三十天左右P-VEGF浓度（范围10.72pg/ml～16.54pg/ml，中位数14.04pg/ml）与手术前、手术后七天、良性疾病组和健康对照组的P-VEGF比较均无统计学差异（p值均大于0.05）。
5乳腺浸润性导管癌患者与浸润性小叶癌患者手术后七天及三十天左右的P-VEGF水平比较均无差异（p=0.290，p=0.162）。
6可手术乳腺癌患者，无论是浸润性导管癌，还是浸润
性小叶癌，手术前P-VEGF水平与肿瘤组织中VEGF、ER和PR的表达均无相关性（p值均大于0.05）。与临床分期（TNM），淋巴结转移状况，月经状况，肿瘤大小均无相关性（p值均大于0.05）。
7浸润性导管癌组织中VEGF和浸润性小叶癌组织中VEGF的表达均与ER、PR的表达无关联性（p值均大于0.05），与临床分期（TNM），淋巴结转移状况，肿瘤大小，月经情况亦无关联性（p值均大于0.05）。且两者之间比较无差异（χ2=0.210,p=0.647）。
结论
1 转移性乳腺癌患者P-VEGF水平高于可手术的乳腺癌、乳房良性疾病和健康对照?
Objective：Many studies discover the growth and metastasis of cancer are closely associated with the formation of tumors blood vessels. Vascular endothelial growth factor(VEGF) is the strongest growth factor to stimulate angiogenesis. There is overexpression of VEGF in many human malignant tumors, including carcinoma of the breast. In this study, enzyme-linked immunosorbent assay（ELISA） was used to examine the levels of preoperative and postoperative plasma VEGF（P-VEGF） in breast cancer. They were also compared with benign breast disease and healthy controls. Hoping to find the changing regulation of P-VEGF and to provide strong evidence to the early finding of replase and metastasis in breast cancer. The expression of VEGF, ER, PR in tumor tissue were also examined by immunohistochemistry.
Methods：The study included 95 patients with 22 benign breast diseases(7 cystic hyperplasia, 6 fibroadenoma, 5 benign mastopathy, 2 intraductal papilloma, 1 mastitis, 1 duct ectasia), 73 localized breast cancer(39 invasive ductal carcinoma, 28 invasive lobular carcinoma, 5 medullary carcinoma,1 mncinous carcinoma), 9 distant metastasis breast cancer( 3 metastatic lobular carcinoma, 1 metastatic ductal carcinoma, 5 unknown),
together with 18 health controls. Those plasma samples were analyzed for VEGF by using ELISA.We also monitored the P-VEGF levels in localized breast cancer, whose blood samples were collected before surgery and on about seventh or thirtieth postoperative days. P-VEGF distributions between different groups were compared. Immunohistochemistry staining for VEGF, ER, PR was performed. All statistical analyses were performed by using SPSS software 10.0.
Results: 1 Significantly higher P-VEGF concentrations were found in metastatic breast cancer (range 23.7 pg/ml～445pg/ml, median 94.9 pg/ml) compared with those in localized breast cancer (range 1.84 pg/ml～229pg/ml, median 17.00 pg/ml), those in benign breast diseases(range 3.81 pg/ml～30.4 pg/ml, median 8.89 pg/ml), and those in healthy controls (range 1.53 pg/ml～32.6 pg/ml, median 5.616 pg/ml) (p<0.05 for all those comparisons). The highest concentration (445 pg/ml) was seen in a patient with metastatic neoplasms in lung and liver.The P-VEGF concentrations of locoregional breast cancer were higher than those of healthy controls (p<0.05), but had no significant difference with benign breast diseases (p>0.05). And the difference between benign breast diseases and healthy controls was not significant (p>0.05).
2 Patients with invasive ductal carcinoma (range 3.90 pg/ml～ 229pg/ml, median 22.19pg/ml) had higher P-VEGF levels than those with lobular carcinoma (range 1.84 pg/ml ～45.9 pg/ml, median 8.79pg/ml; p<0.05), benign diseases (p<0.05) and
healthy controls (p<0.05),but not higher than those with medullary carcinoma (range 6.47 pg/ml ～143.3 pg/ml, median 13.7pg/ml; p>0.05).The P-VEGF levels of the patients with locoregional lobular carcinoma were not higher than those of the patients with medullary carcinoma,benign diseases and healthy controls (p>0.05 for all).The P-VEGF levels in patients with medullary carcinoma were higher than those in healthy controls (p<0.05) ,but not higher than those in benign diseases (p>.05).The patients of invasive lobular cancer with distant metastases had higher P-VEGF levels than those without distant metastases.
3 The preoperative P-VEGF levels in invasive ductal cancer ranged from3.90 pg/ml to 229 pg/ml.(median,22.19pg/ml).The levels measured on about seventh postoperative day (range 3.96 pg/ml～209pg/ml, median 19.47 pg/ml) showed no significant difference with the preoperative levels (p>0.05) and those in benign breast diseases (p>0.05), but were higher than those in healthy controls (p<0.05).The levels found on about thirtieth postoperative day (range 1.98 pg/ml ～ 87 pg/ml, median 5.86 pg/ml)were significantly lower than the preoperative and the seventh-postoperative levels(p<0.05). And there were no significant difference between them and healthy controls (p>0.05) or benign breast diseases (p>
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