Immunohistochemical Examination on the Distribution of Cells Expressed Lymphatic Endothelial Marker Podoplanin and LYVE-1 in the Mouse Tongue Tissue

The clinical study for lingual disease requires the detailed investigation of the lingual lymphatic network and lymphatic marker-positive cells. Recently, it has been reported that several tissue cells and leukocytes express lymphatic markers, LYVE-1 and podoplanin. This study was aimed to clarify the lingual distribution of cells expressing LYVE-1 and podoplanin. In the mouse tongue, podoplanin is expressed in nerve sheaths, lingual gland myoepithelial cells, and lymphatic vessels. LYVE-1 is expressed in the macrophage marker Mac-1-positive cells as well as lymphatic vessels, while factor-VIII was detected in only blood endothelial cells. α-SMA was detected in vascular smooth muscle and myoepithelial cells. Therefore, identification of lymphatic vessels in lingual glands, the combination of LYVE-1 and factor-VIII, or LYVE-1 and Mac-1 is useful because myoepithelial cells express podoplanin and α-SMA. The immunostaining of factor-VIII on lymphatic vessels was masked by the immunostaining to LYVE-1 or podoplanin because lymphatic vessels express factor-VIII to a far lesser extent than blood vessels. Therefore, except for the salivary glands, the combination of podoplanin and α-SMA, or factor-VIII is useful to identify lymphatic vessels and blood vessels with smooth muscle, or blood capillaries.     

Angiogenesis and lymphangiogenesis and expression of lymphangiogenic factors in the atherosclerotic intima of human coronary arteries

Little information regarding the development of lymphangiogenesis in coronary atherosclerosis is available. We immunohistochemically investigated the correlation among intimal neovascularization (CD34 for angiogenesis and lymphatic vessel endothelial hyaluronan receptor-1 [LYVE-1] and podoplanin for lymphangiogenesis), the expression of lymphangiogenic factors (vascular endothelial growth factor [VEGF]-C and VEGF-D), and the progression of atherosclerosis using 169 sections of human coronary arteries from 23 autopsy cases. The more the atherosclerosis advanced, the more often the neointimas contained newly formed blood vessels ( P < .0001). Vascular endothelial growth factor-C was expressed mostly in foamy macrophages and in some smooth muscle cells, whereas VEGF-D was abundantly expressed in both. The number of VEGF-C-expressing cells, but not that of VEGF-D-expressing cells, was increased as the lesion advanced and the number of intimal blood vessels increased ( P < .01). Lymphatic vessels were rare in the atherosclerotic intima (LYVE-1 vs CD34 = 13 vs 3955 vessels) compared with the number seen in the adventitia (LYVE-1 vs CD34 = 360 vs 6921 vessels). The current study suggests that VEGF-C, but not VEGF-D, may contribute to plaque progression and be a regulator for angiogenesis rather than lymphangiogenesis in coronary atherosclerotic intimas. Imbalance of angiogenesis and lymphangiogenesis may be a factor contributing to sustained inflammatory reaction during human coronary atherogenesis.     

Investigation of intratumoural and peritumoural lymphatics expressed by podoplanin and LYVE-1 in the hybridoma-induced tumours

Tumour-associated lymphatics contribute to a key component of metastatic spread, however, the biological interaction of tumour cells with intratumoural and peritumoural lymphatics (ITLs and PTLs) has remained unclear. To address this important issue, we have focused on the morphological and molecular aspects of newly formed lymphatics (lymphangiogenesis) and pre-existing lymphatics in the intratumoural and peritumoural tissues by using a hybridoma-induced tumour model. In the present study, ITLs with very high vessel density within the tumour mass showed small and flattened contours that varied from non-solid-to-solid tumours, whereas PTLs were relatively disorganized and tortuous, and packed with a cluster of tumour cells at the tumour periphery. Lymphatic endothelial cells (LECs) both in ITLs and PTLs were expressed with LYVE-1 and podoplanin in various tumour tissues, in which initial lymphatics were extremely extended and dilated. The tumour cells were frequently detected adhering to or penetrating lymphatic walls, especially near the open junctions. In the metastatic tissues, lymphangiogenic vasculatures occurred within the tumour matrix, and collecting PTLs represented abnormal twisty valve leaflets. The Western blot and RT-PCR analysis showed local variations of LEC proliferating potentials and lymphatic involvement in metastasis by a distinct profile of the protein and mRNA expression by LYVE-1, podoplanin, Prox-1 and vascular endothelial growth factor-3 (VEGFR-3). These findings indicated that both ITLs and PTLs, including enlarged pre-existing and newly formed lymphatics, may play a crucial role in metastasis with an active tumour cell adhesion, invasion, migration and implantation.     

Expression of vascular endothelial growth factor (VEGF)-C and VEGF-D, and their receptor VEGFR-3, during different stages of cervical carcinogenesis

Cervical carcinogenesis has well-defined stages of disease progression including three grades of pre-invasive lesions--cervical intraepithelial neoplasia grades 1-3 (CIN 1-3)--and invasive cervical cancer. However, the biological properties of CIN lesions prone to develop invasive disease are not well defined. Recent observations suggest that early invasive disease spreads to regional lymph nodes in several tumour types and that growth factors (VEGF-C and VEGF-D) involved in new lymphatic vessel formation may play a crucial role in this process. The present study has assessed the expression of VEGF-C and VEGF-D, and their receptor VEGFR-3, in 152 cervical lesions (33 CIN 1, 33 CIN 2, 37 CIN 3, and 49 squamous cell carcinomas) to determine whether expression of lymphangiogenic factors occurs prior to invasion. The presence of lymphatic vessels was determined using LYVE-1 and podoplanin staining, as well as double immunostaining for LYVE-1/CD34 and podoplanin/CD34. In situ hybridization was performed to determine VEGFR-3 mRNA expression. A significant positive correlation was found between VEGF-C, VEGF-D, and VEGFR-3 expression through the different stages of cervical carcinogenesis. Significant differences in protein expression for VEGF-C, VEGF-D, and VEGFR-3 were found between CIN 1-2 and CIN 3 (p<0.001 for all), but not between CIN 3 and cervical cancer. More than 50% of the CIN 3 lesions showed moderate to strong staining for VEGF-C and VEGF-D, whereas most of the early pre-cancerous lesions (CIN 1 and 2) were negative. In cervical cancer, similar observations to those in CIN 3 were found. VEGFR-3 mRNA expression was found in the cytoplasm of epithelial neoplastic cells and VEGFR3 protein expression was found in more than 50% of CIN 3 lesions and cervical cancers, compared with 15% in CIN 1 and 2. These findings suggest an autocrine growth stimulation pattern via VEGFR-3. Adjacent CIN 3 was present in nine cervical cancers and displayed strong expression for VEGF-C, VEGF-D, and VEGFR-3. These results suggest that in cervical carcinogenesis a switch to the lymphangiogenic phenotype may occur at the stage of CIN 3.     

Infantile hemangioma is a proliferation of LYVE-1-negative blood endothelial cells without lymphatic competence.

Infantile hemangiomas are common benign vascular tumors that exhibit a characteristic history of rapid proliferation in the first year of life and slow spontaneous involution during early childhood. The causative pathogenic event responsible for the abnormal endothelial proliferation remains elusive. The recent discovery of an immature phenotype of proliferating hemangioma endothelial cells due to the exclusive expression of the lymphatic endothelial hyaluronan receptor LYVE-1 led to the proposal that infantile hemangiomas are the result of a primary defect in endothelial cell maturation. To test this hypothesis, we looked for the expression of the lymphatic endothelial cell-specific markers LYVE-1, Prox-1, podoplanin and D2-40 in beta4 integrin-negative proliferating and beta4 integrin-positive involuting infantile hemangiomas. As beta4 integrin proved to be a suitable marker for staging infantile hemangiomas, we used it in combination with clinical and histological criteria to objectively determine the proliferative and involutional phases. In immunohistochemical and immunofluorescent stains, hemangioma vessels were negative for all lymphatic endothelial cell-specific markers tested during both proliferation and involution. LYVE-1 immunoreactivity, however, was found in the dense network of perivascular HLA-DR-positive cells with dendritic cell morphology that are supposed to play a role in hemangiogenesis by releasing pro- and antiangiogenic factors. Notably, this LYVE-1 staining failed to correlate with the growth status of infantile hemangiomas. Our results do not support the notion that LYVE-1 expression was restricted to the proliferative phase and downregulated during involution. Thus, LYVE-1 does not seem to be a reliable marker for proliferating infantile hemangiomas. We conclude that the suggested intrinsic defect in endothelial cell maturation is unlikely the cause for the post-natal rapid growth in infantile hemangiomas. In addition, the lack of lymphatic endothelial cell-specific markers implies that infantile hemangiomas are tumors of blood vessels without lymphatic competence.     

基质细胞衍生因子-1α和白细胞介素-1β诱导淋巴管内皮表型的作用

目的探讨基质细胞衍生因子-1α(SDF-1α)及白细胞介素-1β(IL-1β)诱导内皮细胞表达淋巴管表型的作用。方法 SDF-1α和IL-1β分别诱导内皮细胞株CRL-1730,用Real-time PCR、Western blotting及免疫细胞化学等方法检测其内皮及淋巴管标志物,的表达情况。结果 SDF-1α诱导培养之后,CRL-1730细胞株的内皮细胞标志物血管性血友病因子(vWF)、血管内皮钙黏蛋白(VE-cadherin)、血管内皮生长因子受体(VEGFR)2随其浓度增高而表达降低,淋巴管标志物平足蛋白(podoplanin)、同源异形盒蛋白-1(Prox-1)和淋巴管内皮透明质酸受体-1(LYVE-1)随其浓度增高而表达增高。IL-1β诱导之后,CRL-1730细胞株的vWF、VEGFR2和podoplanin、prox-1、LYVE-1的变化趋势同SDF-1α,而VE-cadherin的表达量基本不变。结论 SDF-1α和IL-1β都能够诱导血管内皮细胞表达淋巴管标志物。     

Lymphatics and bone

There is controversy regarding whether lymphatic vessels are present or absent in bone. Although lymphangiomas have been described in bone, lymphatic vessels have not been identified morphologically with certainty in any other benign or malignant bone tumors or in normal human bone. In this study, we determined by immunohistochemistry, using 2 specific lymphatic endothelial cell markers, LYVE-1 and podoplanin, whether lymphatics are present in normal bone and a wide range of primary and secondary bone neoplasms. In normal bone, LYVE-1+/podoplanin+ lymphatic vessels were not identified in cortical or cancellous bone but were seen in connective tissue overlying the periosteum. With the exception of lymphangioma, Gorham-Stout disease, and hemangioendothelioma, primary benign and malignant bone tumors (as well as secondary carcinomas) that were confined to bone did not contain lymphatic vessels. Primary and secondary bone tumors that had extended through the bone cortex contained LYVE-1+/podoplanin+ lymphatic vessels that seemed to extend for a short distance from surrounding soft tissues into the tumor. Three cases of osteosarcoma that had extended through the bone cortex and had lymph node metastases were all found to contain lymphatic vessels within the tumor. These results indicate that the lymphatic circulation is unlikely to play a role in bone fluid transport in normal bone and that lymphatic vessels are absent from most primary and secondary tumors confined to bone. These findings also suggest that lymphangiogenesis is not involved in the disease progression of most primary bone tumors and that carcinomatous metastasis to bone does not occur via lymphatics.     

Biology of the lymphatic marker LYVE-1 and applications in research into lymphatic trafficking and lymphangiogenesis

The pace of research into the lymphatic system continues to accelerate with the availability of new molecular markers. One such marker, LYVE-1, the lymphatic receptor for the extracellular matrix mucopolysaccharide hyaluronan, has been a key component of many important studies on embryonic and tumour-induced lymphangiogenesis, and continues to be used for the detection and isolation of lymphatic endothelial cells. However, LYVE-1 is interesting in its own right. Being a member of the Link protein family whose only other major hyaluronan receptor is directly involved in leukocyte migration and tumour metastasis, LYVE-1 is already implicated in the trafficking of cells within lymphatic vessels and lymph nodes. The current challenge is to determine the precise roles played by LYVE-1 and other scavenger type receptors in the immune functions of the lymphatics as well as to use LYVE-1 and other markers to investigate the way in which tumours exploit lymphatic vessels for metastasis.     

Absence of lymphangiogenesis and intratumoural lymph vessels in human metastatic breast cancer

Early metastasis to lymph nodes is a frequent complication in human breast cancer. However, the extent to which this depends on lymphangiogenesis or on invasion of existing lymph vessels remains controversial. Although proliferating intratumoural lymphatics that promote nodal metastasis have been demonstrated in experimental breast tumours overexpressing VEGF-C, it has yet to be determined whether the same phenomena occur in spontaneous human breast cancers. To address this important issue, the present study investigated the lymphatics in primary human breast carcinoma (75 cases of invasive ductal and lobular breast cancer) by quantitative immunohistochemical staining for the lymphatic endothelial hyaluronan receptor LYVE-1, the blood vascular marker CD34, and the nuclear proliferation marker pKi67. None of the breast carcinomas was found to contain dividing lymph vessels, even in areas of active haemangiogenesis. Furthermore, the majority of non-dividing lymph vessels were confined to the tumour periphery where their incidence was low and unrelated to tumour size, grade or nodal status; rather, their density was inversely correlated with tumour aggressiveness as assessed by macrophage density (p = 0.009), and blood microvessel density (p = 0.05, Spearman Rank), as well as with distance from the tumour edge. Finally, a proportion of the peritumoural lymphatics contained tumour emboli associated with hyaluronan, indicating a possible role for LYVE-1/hyaluronan interactions in lymphatic invasion or metastasis. These results suggest that naturally occurring breast carcinomas invade and destroy lymph vessels rather than promoting their proliferation; that breast tumour lymphangiogenesis may not always occur at physiological VEGF-C levels; and that nodal metastasis can proceed via pre-existing lymphatics.     

Immunohistochemical detection of human small lymphatic vessels under normal and pathological conditions using the LYVE-1 antibody

The spread of tumor cells via lymphatic vessels to the lymph nodes is an important indicator of malignancy. However, previous markers used to identify lymphatic endothelium gave ambiguous results in immunohistochemical analyses with paraffin-embedded tissues. In this study, we attempted to prepare a polyclonal antibody against human lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) for detecting lymphatic vessels using immunohistochemistry. The antibody was raised against a region near the transmembrane anchor of LYVE-1 in New Zealand white rabbits. Immunostainings with anti-LYVE-1 and von Willebrand factor antibodies were performed in various normal and pathological tissues. LYVE-1 expression was confined to the endothelial surface of lymphatic vessels but was not found in the endothelium of blood vessels, which were positive for von Willebrand factor. Our LYVE-1 polyclonal antibody was useful for the identification of small lymphatic vessels in normal human tissues. In addition, the immunostaining enabled us to distinguish lymphatic invasion by malignant tumor cells from blood vessel invasion using paraffin-embedded sections. In conclusion, our polyclonal antibody against the transmembrane anchor of the peptide can be used to detect human lymphatic vessels under various conditions.     

LYVE-1 immunohistochemical assessment of lymphangiogenesis in endometrial and lung cancer

AIMS/METHODS: Normal and malignant pulmonary and endometrial tissues were analysed for lymphatic vessels to assess the process of lymphangiogenesis and its role at these sites, using specific immunostaining for LYVE-1 and the panendothelial marker CD31. RESULTS: Lymphatics were clearly demonstrated in some normal tissues (myometrium, bronchial submucosa, and intestinal submucosa), but not in others (endometrium and alveolar tissue). LYVE-1 positive lymphatic vessels were detected at the tumour periphery of endometrial and lung carcinomas, but not within the main tumour mass. Double staining for LYVE-1 and the MIB1 proliferation marker revealed a higher proliferation index in lymphatic endothelial cells at the invading front of endometrial carcinomas, compared with myometrial areas distal to the tumour. Lung and endometrial carcinomas did not have an intratumorous lymphatic network. CONCLUSIONS: Although lymphangiogenesis may occur at the invading tumour front, incorporated lymphatics do not survive. Therefore, the dissemination of cancer cells through the lymphatics may occur by invasion of peripheral cancer cells into the adjacent normal lymphatics, or through shunts eventually produced at the invading tumour front as a consequence of active angiogenesis and lymphangiogenesis.     

Lymphatic development in mouse small intestine.

Lymphatic vessels in the small intestine serve as essential conduits for the absorption and transport of lipids from the intestine to the thoracic duct. Although the morphology and function of the intestinal lymphatic vasculature are well known, little is known about the embryonic development of these vessels. In this study, we examined development of lymphatic and blood vasculatures in the intestinal tube during mouse embryonic development by immunostaining with recently discovered molecular markers for lymphatic endothelial cells: LYVE-1, VEGFR3, Prox-1, and podoplanin. Immature lymphatics became detectable in mesentery, but not in intestinal tube, around E13.5-E14.5, while organized lymphatic vessel plexuses and capillaries were observed in intestinal tube and villi around E17.5. These lymphatic plexuses and capillaries in the intestinal tube appeared to be formed through an active branching process associated with activation of VEGFR3 and involvement of LYVE-1+ macrophages. Our data also reveal that the lymphatic vessels in the intestinal tube, unlike the blood vessels, have not originated from the mesoderm of intestine. All lymphatic vessels in the intestinal tube originated by extension of mesenteric lymphatic vessels through an active branching process. Although the formation of lymphatic vessels follows the formation of blood vessels in the intestine, a mature lymphatic vasculature is formed before birth. Together, our study reveals the temporal and spatial windows of intestinal lymphatic development during embryonic development in mouse.     

The use of LYVE-1 antibody for detecting lymphatic involvement in patients with malignant melanoma of known sentinel node status

BACKGROUND: Sentinel node (SN) status is the most important prognostic indicator in patients with cutaneous melanoma without clinically evident metastatic spread, but the procedure is associated with considerable morbidity. The LYVE-1 lymphatic marker offers the possibility of studying lymphangiogenesis and tumour metastasis within the primary excision. AIMS: To establish whether lymphatic vessel numbers/distribution within the primary tumour correlated with SN status. To assess whether tumour cells were easily demonstrable within lymphatics and could be used as a surrogate for SN status. METHODS: Double immunostaining for LYVE-1 and S100 in cutaneous biopsies from 18 SN+ patients with no lymphatic/vascular involvement on routine histology and 18 SN- patients matched for tumour thickness and ulceration.RESULTS: Lymphatic vessels were detected in all cases. Vessels within the tumour mass were suggestive of active lymphangiogenesis; those outside were mainly mature vessels with well defined walls. Tumour cells within lymphatics were detected in one of 18 SN- and five of 18 SN+ patients. Lymphatics containing tumour cells were all outside the tumour mass in well formed vessels, suggesting melanoma cell invasion into preformed lymphatics. There was no significant difference in lymphatic counts between SN+ and SN- patients. Although peritumorous lymphatic counts were higher in ulcerated than non-ulcerated melanomas, they did not vary with Breslow thickness. CONCLUSION: LYVE-1 staining can reliably demonstrate lymphatic vessel distribution, but lymphatic counts cannot predict melanoma metastatic potential and cannot substitute for SN biopsy. LYVE-1 immunostaining can detect melanoma cells within lymphatics, but is unreliable in predicting melanoma metastasis, failing to detect metastatic spread in more than two thirds of patients with regional node metastasis.     

Kaposi's sarcoma-associated herpesvirus infection of blood endothelial cells induces lymphatic differentiation

Kaposi's sarcoma-associated herpesvirus (KSHV) is necessary for KS, a highly vascularized tumor predominated by endothelial-derived spindle cells that express markers of lymphatic endothelium. Following KSHV infection of TIME cells, an immortalized human dermal microvascular endothelial cell (DMVEC) line, expression of many genes specific to lymphatic endothelium, including VEGFR3, podoplanin, LYVE-1, and Prox-1, is significantly increased. Increases in VEGFR3 and podoplanin protein are also demonstrated following latent infection. Examination of cytokine secretion showed that KSHV infection significantly induces hIL-6 while strongly inhibiting secretion of IL-8, a gene product that is decreased by differentiation of blood to lymphatic endothelial cells. These studies support the hypotheses that latent KSHV infection of blood endothelial cells drives their differentiation to lymphatic endothelial cells.     

Endostatin inhibits tumour lymphangiogenesis and lymphatic metastasis via cell surface nucleolin on lymphangiogenic endothelial cells

Endostatin has potent anti‐endothelial and anti‐angiogenic functions. Endostatin was reported to reduce lymphangiogenesis by down‐regulating the level of VEGF‐C in tumour tissues. However, there is little evidence for the direct function of endostatin on lymphangiogenic endothelial cells and lymphangiogenic vessels. Here, we report that cell surface nucleolin, which was reported as an endostatin receptor mediating its anti‐angiogenic and anti‐tumour functions, is also selectively expressed on the cell surface of lymphangiogenic endothelial cells both in vitro and in vivo. Treatment of primary mouse lymphatic endothelial cells (mLECs) by endostatin inhibits mLEC migration, tubule formation, and activation of the Erk pathway in mLECs, while neutralization of cell surface nucleolin or nucleolin knockdown results in loss of the anti‐lymphatic endothelial activities of endostatin. Also, anti‐nucleolin antibody or lentivirus delivered nucleolin siRNA abolishes the anti‐lymphangiogenic function of endostatin in the Matrigel plug assay. Endostatin remarkably inhibits tumour‐associated lymphangiogenesis, leading to reduced lymphatic metastasis. Systemic blockade of nucleolin notably abolishes the anti‐lymphangiogenic and anti‐lymphatic metastatic functions of endostatin. Importantly, endostatin does not affect quiescent lymphatics in normal organs, which is consistent with the lack of expression of cell surface nucleolin in quiescent lymphatics. Taken together, our results demonstrate that endostatin directly acts on lymphangiogenic endothelial cells via cell surface nucleolin, which provides a novel mechanism for the inhibition of tumour lymphangiogenesis and lymphatic metastasis by endostatin. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Angiosarcomas express mixed endothelial phenotypes of blood and lymphatic capillaries: podoplanin as a specific marker for lymphatic endothelium

Angiosarcomas apparently derive from blood vessel endothelial cells; however, occasionally their histological features suggest mixed origin from blood and lymphatic endothelia. In the absence of specific positive markers for lymphatic endothelia the precise distinction between these components has not been possible. Here we provide evidence by light and electron microscopic immunohistochemistry that podoplanin, a approximately 38-kd membrane glycoprotein of podocytes, is specifically expressed in the endothelium of lymphatic capillaries, but not in the blood vasculature. In normal skin and kidney, podoplanin colocalized with vascular endothelial growth factor receptor-3, the only other lymphatic marker presently available. Complementary immunostaining of blood vessels was obtained with established endothelial markers (CD31, CD34, factor VIII-related antigen, and Ulex europaeus I lectin) as well as podocalyxin, another podocytic protein that is also localized in endothelia of blood vessels. Podoplanin specifically immunolabeled endothelia of benign tumorous lesions of undisputed lymphatic origin (lymphangiomas, hygromas) and was detected there as a 38-kd protein by immunoblotting. As paradigms of malignant vascular tumors, poorly differentiated (G3) common angiosarcomas (n = 8), epitheloid angiosarcomas (n = 3), and intestinal Kaposi's sarcomas (n = 5) were examined for their podoplanin content in relation to conventional endothelial markers. The relative number of tumor cells expressing podoplanin was estimated and, although the number of cases in this preliminary study was limited to 16, an apparent spectrum of podoplanin expression emerged that can be divided into a low-expression group in which 0-10% of tumor cells contained podoplanin, a moderate-expression group with 30-60% and a high-expression group with 70-100%. Ten of eleven angiosarcomas and all Kaposi's sarcomas showed mixed expression of both lymphatic and blood vascular endothelial phenotypes. By double labeling, most podoplanin-positive tumor cells coexpressed endothelial markers of blood vessels, whereas few tumor cells were positive for individual markers only. From these results we conclude that (1) podoplanin is a selective marker of lymphatic endothelium; (2) G3 angiosarcomas display a quantitative spectrum of podoplanin-expressing tumor cells; (3) in most angiosarcomas, a varying subset of tumor cells coexpresses podoplanin and endothelial markers of blood vessels; and (4) all endothelial cells of Kaposi's sarcomas expressed the lymphatic marker podoplanin.     

人胰腺癌淋巴管的分布及形态观察

目的观察人胰腺癌淋巴管的分布及形态结构,探讨胰腺癌淋巴道转移机制。方法取手术后人胰腺癌标本21例,应用免疫组化染色法LYVE-1标记淋巴管进行淋巴管计数,半薄切片光镜观察和超薄切片透射电镜观察胰腺癌组织淋巴管的形态及分布特点。结果胰腺癌组织中LYVE-1染色阳性的脉管具有淋巴管的形态学特征,可见癌周组织的微淋巴管数量较癌旁"正常区"有所增加(P<0.01);半薄切片光镜下可见癌周边区和"正常区"淋巴管存在,癌中心区未见有淋巴管;电镜下癌周边区淋巴管内皮细胞连接开放,部分内皮细胞破裂溶解,管壁不完整。淋巴管内皮细胞的线粒体、高尔基体等细胞器改变。结论胰腺癌组织淋巴管主要位于癌周围浸润区的纤维结缔组织中,且淋巴管数量较癌旁"正常区"增多,淋巴管内皮超微结构改变。胰腺癌淋巴管转移可能通过增多的淋巴管的内皮连接开放和对内皮细胞的破坏溶解作用进入淋巴管管壁。     

Decreased macrophage number and activation lead to reduced lymphatic vessel formation and contribute to impaired diabetic wound healing

Impaired wound healing is a common complication of diabetes. Although it is well known that both macrophages and blood vessels are critical to wound repair, the role of wound-associated lymphatic vessels has not been well investigated. We report that both the presence of activated macrophages and the formation of lymphatic vessels are rate-limiting to the healing of diabetic wounds. We have previously shown that macrophages contribute to the lymphatic vessels that form during the acute phase of corneal wound healing. We now demonstrate that this is a general phenomenon; cells that co-stain for the macrophage marker F4/80 and the lymphatic markers LYVE-1 (lymphatic vascular endothelium hyaluronate receptor) and podoplanin contribute to lymphatic vessels in full-thickness wounds. LYVE-1-positive lymphatic vessels and CD31-positive blood vessels were significantly reduced in corneal wound healing in diabetic mice (db/db) (P < 0.02) compared with control (db/+) mice. Glucose treatment of control macrophages led to the down-regulation of the lymphatic-specific receptor VEGFR3 and its ligands, vascular endothelial growth factor-C and -D (VEGF-C, -D). Interleukin-1beta stimulation rescued diabetic macrophage function; application of interleukin-1beta-treated db/db-derived macrophages to wounds in db/db mice induced lymphatic vessel formation and accelerated wound healing. These observations suggest a potential therapeutic approach for healing wounds in diabetic patients.     

Expression of vascular endothelial growth factor receptor 3 in blood and lymphatic vessels of lung adenocarcinoma

Vascular endothelial growth factor receptor 3 (VEGFR-3) has been proposed as a marker for lymphatic endothelial cells. This study investigated the expression of VEGFR-3 in the tumour vessels of lung adenocarcinoma and evaluated whether VEGFR-3 staining was useful for identifying lymphatic vessels within the tumour stroma. It also explored whether active growth of lymphatic vessels occurred in lung adenocarcinoma. Formalin-fixed, paraffin-embedded specimens obtained from 60 cases of lung adenocarcinoma, including five cases of pure bronchiolo-alveolar carcinoma (BAC) without stromal, vascular, and pleural invasion, were examined. No VEGFR-3-positive vessels were observed in pure BAC, but varying numbers of VEGFR-3-positive vessels were found in 39 of 55 (70.9%) invasive adenocarcinomas. A comparison of serial sections stained for VEGFR-3, CD31, and laminin-1 showed that most of the VEGFR-3-positive vessels appeared to be blood vessels (CD31-positive, laminin-1-positive), but some had the characteristics of lymphatic vessels (variable staining for CD31, little or no staining for laminin-1). VEGFR-3 staining highlighted lymphatic invasion by cancer cells; this invasion could not be detected by CD31 or haematoxylin and eosin (H&E) staining. Active growth of lymphatic vessels (as indicated by nuclear Ki-67 labelling of the endothelium) was observed in five tumours, four of which showed a high level of lymphatic invasion by cancer cells. It was concluded that VEGFR-3 immunostaining did not discriminate clearly between vascular and lymphatic endothelial cells, since expression of VEGFR-3 can be up-regulated in tumour blood vessels. However, VEGFR-3 staining combined with laminin-1 and CD31 staining would be useful for identifying lymphatic vessels and their invasion by tumour cells in a more objective way. Finally, proliferation of lymphatic endothelial cells may occur in association with lymphatic invasion by cancer cells.     

肺癌组织及淋巴管E-cadherin、β-catenin和LYVE-1的表达

目的观察肺癌组织E-cadherin和β-catenin的表达,LYVE-1特异性标记淋巴管;探讨钙黏蛋白及其受体在癌细胞淋巴道转移中的作用。方法取肺癌手术材料30例,通过免疫组化法,观察E-cadherin、β-catenin和LYVE-1在癌细胞及淋巴管的表达。结果癌细胞对E-cadherin、β-catenin呈阳性表达,低分化组、有淋巴结转移组E-cadherin表达减弱。淋巴管对LYVE-1阳性表达,E-cadherin阴性表达,β-catenin弱阳性表达。结论LYVE-1在淋巴管特异性表达;E-cadherin表达与肿瘤分化程度和有无淋巴结转移呈负相关;E-cadherin/β-catenin复合物对肿瘤细胞与淋巴管内皮细胞的黏附不起主要作用。