Quantitative analysis of lymphangiogenic markers for predicting metastasis of human gastric carcinoma to lymph nodes

The spread of tumor cells to regional lymph nodes is an early event of gastric cancer metastasis. In our study, we assessed the expression of lymphangiogenic factors and lymphatic endothelial markers in gastric carcinoma tissues and compared expression levels with the status of lymph node metastasis. We also examined the correlation between lymphatic vessel density (LVD) in primary tumors and lymph node metastasis. Paired biopsy samples (tumor and corresponding normal mucosa) of gastric tissue were obtained from 39 patients with gastric carcinoma. The expression of VEGF-C, VEGF-D, VEGFR-3 and podoplanin mRNAs was assessed by real-time quantitative PCR. The expression of VEGF-C (but not of VEGF-D) was significantly greater in patients with lymph node metastasis than in those without metastasis. The expression of lymphatic endothelial markers VEGFR-3 and podoplanin was also significantly greater in the node-positive group. LVD, as assessed by immunohistochemistry for podoplanin, was correlated with lymph node metastasis. These results indicate that quantitative analysis of lymphangiogenic markers in gastric biopsy specimens may be useful in predicting metastasis of gastric cancer to regional lymph nodes.     

Expression of vascular endothelial growth factor receptor-3 by lymphatic endothelial cells is associated with lymph node metastasis in prostate cancer.

PURPOSE: The molecular mechanisms underlying lymph node metastasis are poorly understood, despite the well-established clinical importance of lymph node status in many human cancers. Recently, vascular endothelial growth factor (VEGF)-C and VEGF-D have been implicated in the regulation of tumor lymphangiogenesis and enhancement of lymphatic invasion via activation of VEGF receptor-3. The purpose of this study was to determine the expression pattern of the VEGF-C/VEGF-D/VEGF receptor-3 axis in prostate cancer and its relationship with lymph node metastasis. EXPERIMENTAL DESIGN: The expression pattern of VEGF-C, VEGF-D, and VEGF receptor-3 in localized prostate cancer specimens (n = 37) was determined using immunohistochemistry. RESULTS: Widespread, heterogeneous staining for VEGF-C and VEGF-D was observed in all cancer specimens. Intensity of VEGF-C staining was lower in benign prostate epithelium than in adjacent carcinoma, whereas no difference between benign epithelium and carcinoma was observed for VEGF-D staining. VEGF receptor-3 immunostaining was detected in endothelial cells of lymphatic vessels in 18 of 37 tissue samples. The presence of VEGF receptor-3-positive vessels was associated with lymph node metastasis (P = 0.0002), Gleason grade (P < 0.0001), extracapsular extension (P = 0.0382), and surgical margin status (P = 0.0069). In addition, VEGF receptor-3 staining highlighted lymphatic invasion by VEGF-C-positive/VEGF-D-positive carcinoma cells. CONCLUSIONS: Together, these results suggest that paracrine activation of lymphatic endothelial cell VEGF receptor-3 by VEGF-C and/or VEGF-D may be involved in lymphatic metastasis. Thus the VEGF-C/VEGF-D/VEGF receptor-3 signaling pathway may provide a target for antilymphangiogenic therapy in prostate cancer.     

Vascular endothelial growth factor-D is an independent prognostic factor in epithelial ovarian carcinoma

We assessed the presence of vascular endothelial growth factor (VEGF)-C, VEGF-D and their receptor VEGFR-3 by immunohistochemistry in 59 epithelial ovarian carcinomas, 11 borderline tumours and 20 benign cystadenomas. VEGF-C and VEGF-D were generally expressed in tumour cells and also in endothelia adjacent to tumour nests which showed a strong staining for them. VEGFR-3 was expressed in lymphatic and vascular endothelial cells adjacent to tumour nests. Immunoreactivity was significantly more frequent as lesions progressed from a benign tumour to advanced carcinoma. A strong correlation was found between VEGF-C and VEGF-D detected in carcinoma and VEGFR-3 detected in neighbouring endothelial cells. Increased expression of VEGF-C, VEGF-D and VEGFR-3 was significantly associated with lymph node metastasis and peritoneal metastasis outside the pelvis. There was a significant correlation between the high levels of VEGF-C and VEGF-D proteins, and poor survival. The presence of VEGF-D was an independent prognostic indicator by multivariate analysis. We conclude that VEGF-C, VEGF-D and VEGFR-3 play an important role in lymphatic spread and intraperitoneal tumour development in ovarian carcinoma. Since VEGF-D was found to be an independent predictor of poor outcome, its measurement, together with other prognostic markers may improve prospective identification of patients with a poor prognosis.     

血管内皮生长因子-D在胃癌中表达的意义

Objective： To investigate the expression of vascular endothelial growth factor D （VEGF-D） in gastric cancer and its relationship with lymph node metastasis. Methods： 100 cases of gastric carcinoma tissues （50 cases with lymph node metastasis, 50 cases negative） and 30 cases of normal gastric tissues were gathered to detect the expressions of VEGF-C and D proteins by immunohistochemistry. Results： VEGF-C and D were revealed in cytoplasm of gastric carcinoma tissues and normal gastric tissues. The positive rates of VEGF-C and D expressions were significantly higher in gastric cancer tissues than those in the normal ones （51%, 60% vs 10%, 20% respectively; both P 〈 0.05）. There were significant correlations between the positive expression of VEGF-D and lymph node metastasis, lymphatic invasion, positive expression of VEG F-C, but not with tumour size, tissue differentiation, and venous invasion. Conclusion： The expression of VEGF-D is closely related to lymph node metastasis in gastric carcinoma.     

Expression of vascular endothelial growth factor (VEGF)-C and VEGF-D in early gastric carcinoma: correlation with clinicopathological parameters

In this study, we investigated whether expression of vascular endothelial growth factor (VEGF)-C and/or VEGF-D correlates with clinicopathological features of human gastric carcinoma. We immunohistochemically examined the expression of VEGF-C and VEGF-D in 140 archival surgical specimens of submucosally invasive gastric carcinoma. Of these specimens, 32 (22.9%) and 12 (8.6%) showed intense VEGF-C and VEGF-D immunoreactivity in cancer cells, respectively. VEGF-C immunoreactivity was associated with histological type, lymphatic invasion, lymph node metastasis, and microvessel density. No association was identified between VEGF-D immunoreactivity and clinicopathological variables. These results suggest that VEGF-C is a dominant regulator of lymphangiogenesis in early-stage human gastric carcinoma.     

Expression of vascular endothelial growth factor C and D (VEGF-C and -D) is an important risk factor for lymphatic metastasis in undifferentiated early gastric carcinoma

BACKGROUND: Vascular endothelial growth factor C (VEGF-C) and D (VEGF-D) are considered to be potentially lymphangiogenic and can selectively induce hyperplasia of the lymphatic vasculature. In this study, we aimed to clarify the relation between expression of VEGF-C and -D and lymphatic metastasis in early gastric cancers. METHODS: Using the specific antibodies, we classified 105 cases which were treated as gastrectomy with standard lymphadenectomy at the First Department of Surgery, Tokyo University Hospital, between 1994 and 2001, into three groups (diffuse type, focal type and negative type) for VEGF-C and two groups (positive and negative) for VEGF-D. RESULTS: There was a significant correlation between the expression of VEGF-C and -D and lymphatic invasion but not with venous invasion. All of the 22 cases that were negative for VEGF-C and -D were histologically classified as adenocarcinoma of undifferentiated type and showed negative lymph node metastasis and also negative lymphatic invasion. VEGF-C was positive in all tumors of differentiated type, while its expression varied in tumors of undifferentiated type. The VEGF-D positive rate is much lower than that of VEGF-C. In undifferentiated tumors in particular, VEGF-D was positive only in 4/64 (6%) and three of these four had nodal metastasis. Therefore, in tumors of differentiated type, expression of VEGF-C and -D had no clinical relevance. In tumors of undifferentiated type, the negative expression of VEGF-C suggests lack of nodal metastasis, while the positive expression of VEGF-D suggests nodal metastasis. The lymph node metastasis was significantly related to the expression of VEGF-C and -D in adenocarcinomas of undifferentiated type but not in those of differentiated tumors. CONCLUSIONS: In early gastric cancers of histologically undifferentiated type with negative expression of VEGF-C and -D, limited surgery might be safely applied because the possibility of nodal metastasis is very low. These observations are based only on retrospective analysis of a small case series and further evaluation with a larger number of cases is necessary.     

血管内皮生长因子D在胃癌中表达的意义

 目的 研究血管内皮生长因子D（VEGF-D）在胃癌组织内表达的意义。方法 选择手术后的100例胃癌（区域淋巴结转移阳性和阴性病例各50例），30例正常胃黏膜设为对照组。利用免疫组化EnVision法，检测研究对象胃组织内VEGF-D和VEGF-C的表达。分析VEGF-D阳性表达与临床病理参数和VEGF-C阳性表达的关系。结果 VEGF-C，VEGF-D的阳性染色为棕黄色，主要位于肿瘤细胞的胞质，局灶性或弥散性分布。VEGF-C，VEGF-D阳性表达以（+）或（++）为主；胃癌VEGF-C、VEGF-D阳性表达率分别为51 %和60 %；对照组分别为10 %和20 %（P＜0.05）；VEGF-D阳性高表达（++）与区域淋巴结转移、淋巴管受侵和VEGF-C阳性表达相关，与肿瘤最大径、肿瘤位置、肿瘤浸润深度、肿瘤肉眼分型、肿瘤组织分化、肿瘤血管受侵无关。结论 VEGF-D在胃癌淋巴道转移过程中具有较重要的作用。     

Role of vascular endothelial growth factor receptor-3/Flt-4 in early-stage cervical cancer

The present study was designed to investigate the role of vascular endothelial growth factor receptor (VEGFR)-3/Flt-4 in the early stages of cervical cancer. VEGFR-3/Flt-4 expression, vascular endothelial growth factor (VEGF)-C and VEGF-D in the early stages (Ia-IIa) of cervical cancer in 41 patients was examined by immunohistochemical analysis. Additionally, the VEGFR-3/Flt-4-marked vascular density (MVD) was examined and the relationship of these factors with clinicopathological factors was analyzed. VEGFR-3/Flt-4 was found to be expressed in lymphatic endothelial cells and, to a certain extent, in vascular endothelial cells. The VEGFR-3/Flt-4-positive vessels were largely distributed in the stroma surrounding the tumor tissues, and these vessels were morphologically divided into blood and lymphatic vessels, respectively. Cancer cells were found to express VEGF-C, VEGF-D and VEGFR-3/Flt-4, and their positive expression rate was 48.7% (20/41), 58.5% (24/41) and 63.4% (26/41), respectively. VEGFR-3/Flt-4 expression in the cancer cells of the cervical cancer patients in our study was found to be correlated to the clinical stage, lymph node metastasis, lymphatic invasion and expression of VEGF-C and VEGF-D. However, this expression was unrelated to menstrual status, histological grade and histological classification. MVD was correlated to the clinical stage and expression of VEGF-C and VEGF-D, but was unrelated to menstrual status, histological grade, histological classification, lymph node metastasis and lymphatic invasion. In conclusion, VEGFR-3/Flt-4 plays an important role in the early stages of cervical cancer.     

VEGF、VEGF-C和VEGF-D在甲状腺乳头状癌组织中的表达及意义

背景与目的:研究表明血管内皮生长因子C和D(vascularendothelialgrowthfactor-Cand-D,VEGF-C,VEGF-D)与淋巴管的生成有关。局部淋巴结转移是甲状腺乳头状癌的特征,本研究的目是探讨血管内皮生长因子及其C和D(VEGF、VEGF-C和VEGF-D)蛋白在甲状腺乳头状癌的表达及其意义。方法:应用免疫组织化学SP法检测115例甲状腺乳头状癌和20例结节性甲状腺肿中VEGF、VEGF-C和VEGF-D蛋白的表达。结果:甲状腺乳头状癌VEGF、VEGF-C和VEGF-D的阳性率分别为79.1%、87.0%和72.2%。结节性甲状腺肿组织中VEGF、VEGF-C和VEGF-D的阳性率分别为30.0%、15.0%和20.0%。甲状腺乳头状癌VEGF、VEGF-C和VEGF-D的阳性率较结节性甲状腺肿明显增高。VEGF的表达与甲状腺癌的大小有关。VEGF、VEGF-C和VEGF-D在淋巴结转移组和非转移组的阳性率分别为84.7%、93.2%、83.1%和73.2%、80.4%、60.7%。VEGF-C和VEGF-D的阳性表达与甲状腺乳头状癌的淋巴结转移显著相关(P<0.05;P<0.01)。结论:VEGF与甲状腺乳头状癌的生长有关。VEGF-C和VEGF-D与淋巴结转移密切相关,提示甲状腺乳头状癌标本VEGF-C和VEGF-D的检测可作为预测肿瘤转移的指标之一。     

血管内皮生长因子D及其受体3在乳腺癌淋巴转移中作用的研究进展

乳腺癌是女性最常见的恶性肿瘤。乳腺癌淋巴转移与患者的预后密切相关。最近研究表明淋巴管生成可能会主动促进淋巴转移的发生,而血管内皮生长因子家族的部分成员在这一过程中发挥了重要作用,如血管内皮生长因子C、血管内皮生长因子D及其受体3等。但是,对乳腺癌中血管内皮生长因子D的作用及其预后价值、血管内皮生长因子受体3与乳腺癌淋巴管生成的关系等问题仍有争议。本文对近年国内外有关血管内皮生长因子D及其受体3在乳腺癌淋巴转移中作用的研究进展作一综述。     

The effect of the expression of vascular endothelial growth factor (VEGF)-C and VEGF receptor-3 on the clinical outcome in patients with gastric carcinoma

Aims

We aimed to investigate the relationship among VEGF-C/VEGFR-3 expression, lymphatic metastasis and patient prognosis in gastric carcinoma.

Material and methods

VEGF-C and VEGFR-3 expression in gastric carcinoma tissues obtained from 204 patients who underwent curative gastrectomy (105 cases presented with lymph node metastasis and 99 cases without metastasis) was examined immunohistochemically. There was no significant difference in the other clinicopathologic variables except for postoperative pathological tumor stage (pT) and TNM stage between the two groups. The results were statistically processed.

Results

The results showed that VEGF-C was located mainly in the cytoplasm of tumor cells and VEGFR-3 was found predominantly in the endothelium of lymphatic vessels. VEGF-C and VEGFR-3 expression was more frequent in gastric carcinoma tissues than that in normal gastric tissues, 54.90% and 35.29% respectively, which revealed that the expression of VEGF-C and VEGFR-3 was significantly stronger in patients with lymph node metastasis than in those without metastasis. Patients who had positive staining for VEGF-C showed significantly less favorable survival rates compared with patients who had negative staining for VEGF-C. The survival rates of patients who had positive staining for VEGFR-3 also were significantly lower compared with patients who had negative staining for VEGFR-3. Patients who had positive staining for both VEGF-C and VEGFR-3 exhibited the most unfavorable prognosis. Multivariate analysis demonstrated that the expression of VEGF-C and VEGFR-3 was an independent prognostic determinant. In addition, faint to moderate VEGF-C expression was detected in normal gastric epithelial cells (18/204, 8.9%).

Conclusions

VEGF-C and VEGFR-3 expression could serve as a prognostic biomarker in patients with gastric carcinoma.     

Impact of vascular endothelial growth factor-C and -D expression in human pancreatic cancer: its relationship to lymph node metastasis.

PURPOSE: The aim of this study was to evaluate the expression of vascular endothelial growth factor (VEGF)-C and -D in pancreatic cancer and to reveal its relation to lymph node metastasis. EXPERIMENTAL DESIGN: Formalin-fixed, paraffin-embedded blocks were obtained from 58 patients with pancreatic head cancer. All of the patients underwent a curative resection. The total number of resected lymph nodes was 1,058. The expressions of VEGF-C and -D were evaluated by immunohistochemical staining. To evaluate the relation to lymph node metastasis, the expressions of VEGF-C and -D between the marginal and central portions in the tumor were compared. When >25% of the tumor cells showed distinct staining, the portion was judged as high expression. RESULTS: The two groups with high expression of VEGF-C (P = 0.015) and VEGF-D (P = 0.020) in the marginal portion had a significantly higher incidence of lymph node metastasis compared with the groups with low expression, respectively. Furthermore, the group with high expression of both VEGF-C and -D in the marginal portion had a higher incidence of lymph node metastasis compared with the group with low expression (P = 0.007). The 5-year survival rate of patients with high expression of both VEGF-C and -D in the marginal portion was significantly lower than that of patients with low expression of both VEGF-C and -D (P = 0.017). CONCLUSIONS: VEGF-C and -D expression in tumor cells in the marginal portion of the tumor significantly associated with lymphatic metastasis and prognosis in patients with pancreatic head cancer.     

Clinical significance of VEGF-A, -C and -D expression in esophageal malignancies

Vascular endothelial growth factors (VEGF)-A, -C and -D are members of the proangiogenic VEGF family of glycoproteins. VEGF-A is known to be the most important angiogenic factor under physiological and pathological conditions, while VEGF-C and VEGF-D are implicated in the development and sprouting of lymphatic vessels, so called lymphangiogenesis. Local tumor progression, lymph node metastases and hematogenous tumor spread are important prognostic factors for esophageal carcinoma (EC), one of the most lethal malignancies throughout the world. We found solid evidence in the literature that VEGF expression contributes to tumor angiogenesis, tumor progression and lymph node metastasis in esophageal squamous cell carcinoma (SCC), and many authors could show a prognostic value for VEGF-assessment. In adenocarcinoma (AC) of the esophagus angiogenic properties are acquired in early stages, particularly in precancerous lesions like Barrett's dysplasia. However, VEGF expression fails to give prognostic information in AC of the esophagus. VEGF-C and -D were detected in SCC and dysplastic lesions, but not in normal mucosa of the esophagus. VEGF-C expression might be associated with lymphatic tumor invasion, lymph node metastases and advanced disease in esophageal SCC and AC. Therapeutic interference with VEGF signaling may prove to be a promising way of anti-angiogenic co-treatment in esophageal carcinoma. However, concrete clinical data are still pending.     

VEGF-C和VEGFR-3对乳腺癌淋巴管生成和淋巴结转移的影响

目的:探讨VEGF-C及VEGFR-3对乳腺癌淋巴结转移的影响。方法:采用免疫组化SP法检测乳腺病、乳腺纤维腺瘤和乳腺癌中VEGF-C及VEGFR-3的表达状态。结果:10例乳腺病及纤维腺瘤VEGF-C均阴性,也无VEGFR-3阳性淋巴管;48例乳腺癌组织VEGF-C阳性率为70.83%(34/48);淋巴结转移者VEGF-C阳性率为92.59%(25/27),显著高于无转移者(42.86%,9/21)(P<0.01)。VEGFR-3阳性淋巴管条数随VEGF-C强度增强而增多。在淋巴结转移者中,VEGFR-3阳性淋巴管条数(6.03)多于无淋巴结转移者(2.01)(P<0.01)。结论:乳腺癌组织中VEGF-C的表达状态与VEGFR-3阳性淋巴管数量及淋巴结转移关系密切。     

The expression of vascular endothelial growth factor C and its receptors in non-small cell lung cancer.

Expression of vascular endothelial growth factor (VEGF)-C and that of its receptors were assessed in non-small cell lung cancer. Immunohistochemistry revealed positive VEGF-C expression in 38.7% (24/62) of the patients studied. A significant positive correlation was found between VEGF-C in cancer cells and VEGF receptor-3 (VEGFR-3) in vascular endothelial cells, but not between VEGF-C in cancer cells and VEGFR-2 in endothelial cells. In this cohort of lung cancer patients, VEGF-C expression was significantly associated with lymph node metastasis, lymphatic vessel invasion, and worse outcomes after the operation. Although the independent prognostic impact of VEGF-C and VEGFR-3 was not clear, VEGFR-2 expression in endothelial cells retained the independency as the prognostic indicator. In light of these findings, we conclude that VEGF-C plays an important role in lymphatic invasion/metastasis and tumour progression in non-small cell lung cancer.     

血管内皮生长因子C、D在鼻咽癌组织中的表达及其临床意义

背景与目的:血管内皮生长因子C(vascular endothelial growth factor-C,VEGF-C)和D(vascular endothelial growth factor-D,VEGF-D)是目前已鉴定出的淋巴管生长因子,有研究表明肿瘤组织中VECF-C或VEGF-D过表达与淋巴转移有关.本研究旨在探讨鼻咽癌组织中VEGF-C和VEGF-D的表达情况及其临床意义.方法:采用免疫组化SP法检测66例鼻咽癌组织中VEGF-C和VEGF-D的表达情况,同时检测血管内皮生长因子受体3(vascular endotheliaI growth factor receptor-3,VEGFR-3)和CD34染色情况并计数微淋巴管密度(lymphatic microvessel density,LMVD)和微血管密度(microvessel density,MVD).结果:VEGF-C高表达率在鼻咽癌组织中(54.5%)较鼻咽非肿瘤组织中(26.3%)高(P＜0.05);鼻咽癌伴区域淋巴结转移或T分期晚者,VEGF-C高表达率增高,单因素及多因素Logistic回归分析均表明区域淋巴结转移与VEGF-C高表达相关(P＜0.05),但VEGF-C高表达与性别、年龄、5年生存率、LMVD、MVD等因素无关(P＞0.05).VEGF-D阳性表达率在鼻咽癌组织中(69.7%)较鼻咽非肿瘤组织中(42.1%)高(P＜0.05);鼻咽癌中VEGF-D阳性表达与性别、年龄、T分期、区域淋巴结转移、LMVD、MVD等因素无关(P＞0.05),但与VEGF-C高表达显著正相关(P＜0.01),VEGF-D阳性表达者5年生存率(50.0%)显著低于VEGF-D不表达者(85.0%)(P＜0.01).结论:鼻咽癌中VEGF-C高表达与区域淋巴结转移密切相关;VEGF-D阳性表达与区域淋巴结转移无关,但与VEGF-C高表达正相关,且与5年生存率密切相关.     

Expression analysis of vascular endothelial growth factors and their relationships to lymph node metastasis in human colorectal cancer

This study was undertaken to determine whether expressions of the vascular endothelial growth factor (VEGF) family (VEGF-A, VEGF-B, VEGF-C, and VEGF-D) are correlated with clinicopathological parameters, with particular reference to lymph node metastasis in colorectal cancer. Total RNA was isolated from 82 surgical specimens of colorectal cancer and matched to normal mucosa with (n = 41) or without (n = 41) lymph node metastasis. The mRNA expression of each VEGF family member was quantified by real-time quantitative (RTQ) RT-PCR assay. VEGF-B and VEGF-C mRNA were significantly higher both in the tumors with lymph node metastasis (p = 0.027 and p = 0.024, respectively) and in tumors with lymphatic invasion (p = 0.042 and p = 0.005, respectively). In contrast, VEGF-D mRNA was down-regulated in tumors with lymphatic involvement (p = 0.047). Among the other clinicopathological factors, we noted that VEGF-A mRNA was higher in tumors with liver metastasis than in those without (p = 0.018) and was higher in tumors with venous invasion than in those without (p = 0.007). The results of this study demonstrate that high levels of VEGF-B, C and low levels of VEGF-D mRNA expression are associated with lymph node metastasis and lymphatic involvement. These results suggest that a balance among VEGF-B, VEGF-C, and VEGF-D might contribute to the lymphangiogenic process and metastasis in colorectal cancer.