Vascular endothelial growth factor receptor-3 in lymphangiogenesis in wound healing

Vascular endothelial growth factor receptor-3 (VEGFR-3) is essential for embryonic cardiovascular development, but thereafter becomes confined to the lymphatic endothelium in adult tissues. We have here studied VEGFR-3 expression in experimental wounds of pigs and chronic inflammatory wounds of humans. In healing incisional and punch biopsy wounds made in the dorsal skin of pigs, angiogenic blood vessels, identified by use of the blood vascular endothelial markers vWF and PAL-E and the basal lamina protein laminin, developed into the granulation tissue stroma from day 4 onward, being most abundant on days 5 and 6 and regressing thereafter. VEGFR-3-positive vessels were observed in the granulation tissue from day 5 onward. These vessels were distinct from the PAL-E/laminin/vWF-positive vessels and fewer in number, and they appeared to sprout from pre-existing VEGFR-3-positive lymphatic vessels at the wound edge. Unlike the blood vessels, very few VEGFR-3-positive lymphatic vessels persisted on day 9 and none on day 14. In chronic wounds such as ulcers and decubitus wounds of the lower extremity of humans, VEGFR-3 was also weakly expressed in the vascular endothelium. Our results suggest that transient lymphangiogenesis occurs in parallel with angiogenesis in healing wounds and that VEGFR-3 becomes up-regulated in blood vessel endothelium in chronic inflammatory wounds.     

Expression of vascular endothelial growth factor-C and its receptor in invasive micropapillary carcinoma of the breast

Invasion to lymphatic vessels and metastasis to lymph nodes are frequent complications in invasive micropapillary carcinoma (IMPC) of human breast cancer. Vascular endothelial growth factor-C (VEGF-C) and its receptor, VEGFR-3 have been implicated as the important factors in the formation of lymphatic vessels and recent experimental evidence strongly suggests that lymphangiogenesis in tumor promotes lymphatic metastasis. To clarify the mechanism of its occurrence, the expression of VEGF-C, VEGFR-3 and lymphatic vessel density (LVD) was examined in 40 cases of IMPC (pure and mixed type) and in 40 cases of pseudo-IMPC. Cytoplasmic expression of VEGF-C and VEGFR-3 were more frequent in tumor cells of IMPC compared to those of pseudo-IMPC. A significant positive correlation was found between the expression of VEGF-C and VEGFR-3 in both IMPC and pseudo-IMPC. The expression of VEGF-C was also significantly associated with higher peritumoral LVD, lymphatic invasion and number of lymph node metastasis in IMPC. These findings suggest that VEGF-C promotes the proliferation of peritumoral lymphatic vessels and that lymphatic invasion and metastasis to lymph nodes are frequently induced in IMPC of breast.     

Transgenic induction of vascular endothelial growth factor-C is strongly angiogenic in mouse embryos but leads to persistent lymphatic hyperplasia in adult tissues

Vascular endothelial growth factor-C (VEGF-C) is the quintessential lymphangiogenic growth factor that is required for the development of the lymphatic system and is capable of stimulating lymphangiogenesis in adults by activating its receptor, VEGFR-3. Although VEGF-C is a major candidate molecule for the development of prolymphangiogenic therapy for defective lymphatic vessels in lymphedema, the stability of lymph vessels generated by exogenous VEGF-C administration is not currently known. We studied VEGF-C-stimulated lymphangiogenesis in inducible transgenic mouse models in which growth factor expression can be spatially and temporally controlled without side effects, such as inflammation. VEGF-C induction in adult mouse skin for 1 to 2 weeks caused robust lymphatic hyperplasia that persisted for at least 6 months. VEGF-C induced lymphangiogenesis in numerous tissues and organs when expressed in the vascular endothelium in either neonates or adult mice. Very few or no effects were observed in either blood vessels or collecting lymph vessels. Additionally, VEGF-C stimulated lymphangiogenesis in embryos after the onset of lymphatic vessel development. Strikingly, a strong angiogenic effect was observed after VEGF-C induction in vascular endothelium at any point before embryonic day 16.5. Our results indicate that blood vessels can undergo VEGF-C-induced angiogenesis even after down-regulation of VEGFR-3 in embryos; however, transient VEGF-C expression in adults can induce long-lasting lymphatic hyperplasia with no obvious side effects on the blood vasculature.     

VEGF-D in association with VEGFR-3 promotes nodal metastasis in human invasive lobular breast cancer

We assessed the expression of vascular endothelial growth factors (VEGF-C and VEGF-D) in breast cancer cells and the density of lymph vessels and VEGF receptor-3 (VEGFR-3)-positive vessels in and around the tumor in invasive lobular breast cancer. We found significant correlation between peritumoral lymph vessel density and presence of lymph node metastases (P=.001) and the number of metastatic lymph nodes (P<.001). A significant correlation was detected between tumor cell VEGF-D expression and lymph node status (P=.001) and density of lymphatic vessel endothelial receptor (LYVE)-1-positive vessels (P=.035). VEGFR-3+/VEGF-D+ and VEGFR-3+/VEGF-C+ tumors had a significantly higher number of metastatic lymph nodes than tumors with other staining patterns (P<.001). Tumors positive for neither VEGF-D nor VEGFR-3 had a lower density of LYVE-1+ vessels than tumors with other staining patterns (P=.033). Our results indicate that peritumoral lymph vessel density is associated with lymph node metastases in invasive lobular breast cancer and that invasive lobular cancer producing VEGF-D, surrounded by VEGFR-3+ vessels, has a significantly higher peritumoral lymph vessel density and a higher number of metastatic lymph nodes.     

血管内皮生长因子C及其受体在MNNG诱发的大鼠大肠癌细胞及淋巴管的表达

目的 观察大鼠大肠癌组织内血管内皮生长因子-C(VEGF-C)及其受体3(VEGFR-3)的表达情况,探讨VEGF-C及其受体VEGFR-3在肿瘤淋巴转移中的作用.方法 采用甲基硝基亚硝基胍(MNNG)诱发的大鼠大肠癌模型,应用免疫组织化学(SABC法)技术检测29例大鼠原发性大肠癌组织中VEGF-C及VEGFR-3蛋白,观察VEGF-C及VEGFR-3在大肠癌组织内的表达.结果 正常大肠组织内未见VEGF-C阳性表达,但可见淋巴管内皮细胞VEGFR-3阳性表达.在大肠癌组织内,VEGF-C蛋白表达于癌细胞,早期和中晚期癌的阳性表达率分别是75%和100%,(P＜0.05).VEGFR-3主要表达于淋巴管内皮细胞,早期和中晚期癌组织内淋巴管的阳性表达率分别是58.33%和94.12%(P＜0.05).结论 大鼠大肠癌VEGF-C的表达与肿瘤进展有关,推测VEGF-C通过受体VEGFR-3诱导淋巴管生成:VEGFR-3在淋巴管的阳性表达均随肿瘤进展增高,可能与大肠癌淋巴转移有关.     

血管内皮生长因子C及其受体3在人恶性黑色素瘤组织内的表达及意义

目的 观察人恶性黑色素瘤组织内血管内皮生长因子C(VEGF-C)及其受体3(VEGFR-3)的表达,探讨VEGF-C和VEGFR-3在恶性黑色素瘤淋巴管生成及淋巴道转移中的作用.方法 取人恶性黑色素瘤组织48例(石蜡标本30例,术后新鲜组织18例),应用免疫组织化学和RT-PCR技术,观察VEGF-C和VEGFR-3蛋白及mRNA在恶性黑色素瘤组织内的表达情况.以淋巴管内皮透明质酸受体(LYVE-1)标记淋巴管,计数恶性黑色素瘤组织淋巴管数密度.结果 VEGF-C和VEGFR-3蛋白主要表达于恶性黑色素瘤细胞胞浆内,在肿瘤周围的血管和淋巴管内皮上也可见VEGFR-3蛋白表达,VEGF-C和VEGFR-3蛋白在淋巴结转移组恶性黑色素瘤组织内的表达水平明显高于无淋巴结转移组(P<0.05).在18例新鲜恶性黑色素瘤中,淋巴结转移组VEGF-C和VEGFR-3mRNA的表达明显高于无淋巴结转移组(P<0.01).LYVE-1表达于肿瘤间质内的淋巴管内皮细胞,淋巴结转移组恶性黑色素瘤组织中的淋巴管数密度(LMVD)为9.845±2.454,无淋巴结转移组恶性黑色素瘤组织中的淋巴管数密度为6.534±2.193,淋巴结转移组恶性黑色素瘤组织内的淋巴管数密度明显高于无淋巴结转移组(P<0.01).结论恶性黑色素瘤组织内VEGF-C表达明显增高,并通过上调其受体VEGFR-3的表达促进恶性黑色素瘤组织内淋巴管的生成,从而促进恶性黑色素瘤的淋巴道转移.     

人喉癌组织中VEGF-C、VEGF-D及其受体3的表达及意义

目的 探讨喉癌组织中VEGF—C、D和受体VEGFR-3的表达及其在喉癌进展中的作用。方法 取人喉癌标本12例,正常及良性病变喉组织10例,免疫组化法观察VEGF—C、VEGF—D、VEGFR-3以及LYVE-1的表达。结果 VEGF—C和VEGF—D主要表达于喉癌细胞胞浆内,喉癌组织中VEGF—C和VEGF—D表达的阳性率明显高于正常及良性病变喉组织（P〈0．05）;VEGFR-3主要表达于基底层的癌细胞,在喉癌组织中VEGFR-3表达的阳性率明显高于正常和良性病变组织中（P〈0．01）,并且VEGFR-3的表达与VEGF—C、VEGF—D的表达显著正相关（P〈0．01）。LYVE—1仅见表达于淋巴管内皮细胞。结论 喉癌组织中VEGF—C、VEGP-D的表达明显增高,推测可能通过与VEGFR-3的结合促进喉癌组织中淋巴管的生成;LYVE-1是淋巴管内皮细胞较特异的标记物。     

VEGFR-3 in adult angiogenesis.

Vascular endothelial growth factor receptor 3 (VEGFR-3, Flt-4), the receptor for vascular endothelial growth factors (VEGFs) C and D, is expressed on lymphatic endothelium and may play a role in lymphangiogenesis. In embryonic life, VEGFR-3 is essential for blood vessel development. The purpose of this study was to investigate whether VEGFR-3 is also involved in blood vessel angiogenesis in the adult. This was studied in human tissues showing angiogenesis and in a model of VEGF-A-induced iris neovascularization in the monkey eye, by the use of immunohistochemistry at the light and electron microscopic level. VEGFR-3 was expressed on endothelium of proliferating blood vessels in tumours. In granulation tissue, staining was observed in the proliferative superficial zone in plump blood vessel sprouts, in the intermediate zone in blood vessels and long lymphatic sprouts, and in the deeper fibrous zone in large lymphatics, in a pattern demonstrating that lymphangiogenesis follows behind blood vessel angiogenesis in granulation tissue formation. At the ultrastructural level, VEGFR-3 was localized in the cytoplasm and on the cell membrane of endothelial cells of sprouting blood vessels and sprouting lymphatics. In monkey eyes injected with VEGF-A, blood vessel sprouts on the anterior iris surface and pre-existing blood vessels in the iris expressed VEGFR-3. In conclusion, these results support a role for VEGFR-3 and its ligands VEGF-C and/or VEGF-D in cell-to-cell signalling in adult blood vessel angiogenesis. The expression of VEGFR-3 in VEGF-A-induced iris neovascularization and in pre-existing blood vessels exposed to VEGF-A suggests that this receptor and possibly its ligands are recruited in VEGF-A-driven angiogenesis.     

直肠腺癌VEGF-C及VEGFR-3的表达与淋巴转移的关系

目的为探讨癌细胞淋巴管转移机理,观察血管内皮生长因子-C(VEGF-C)和血管内皮生长因子受体-3(VEGFR-3)在直肠腺癌组织及淋巴管的表达。方法取人直肠腺癌手术材料30例,用免疫组化方法检测癌区和癌周正常区VEGF-C和VEGFR-3的表达情况。结果VEGF-C主要表达在直肠腺癌的癌细胞胞浆,VEGFR-3主要在淋巴管内皮细胞有阳性表达,两者在癌区的表达率均高于正常区直肠组织;癌区淋巴管的平均面密度(33.81±5.67)高于正常区平均面密度(20.13±3.27)。结论VEGF-C和VEGFR-3在人直肠腺癌中的过表达,可能与淋巴管增生和扩张,促进癌细胞的淋巴转移有关。     

CD40 ligation induces tissue factor expression in human vascular smooth muscle cells

Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are important regulators of blood and lymphatic vessel growth and vascular permeability. Both blood and lymphatic vessels of the upper respiratory tract play important roles in pathological conditions, such as infections and tumors. Here we have studied the expression of VEGF-C and its receptor VEGFR-3 in the upper respiratory system by Northern blot analysis and immunohistochemistry of human tissues, and in situ mRNA hybridization of developing mouse embryos and β-galactosidase staining of mouse embryos having a LacZ marker gene in the VEGFR-3 gene locus. The results demonstrate expression of VEGF-C and VEGFR-3 in the developing and adult nasal respiratory epithelium and in the nasal vascular plexus, respectively. Unlike in most other tissues, in the nasal mucosa VEGFR-3 is expressed in both blood and lymphatic vessels. Expression of VEGF-C was also detected in nasal and nasopharyngeal tumor islands, which were surrounded by VEGFR-3-positive angiogenic blood vessels. These results suggest that VEGF-C and VEGFR-3 have a role in the development of the nasal submucosal vascular plexus and in its normal function and that they are associated with angiogenesis in nasal and nasopharyngeal tumors.     

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.     

Expression of the Vascular Endothelial Growth Factor C Receptor VEGFR-3 in Lymphatic Endothelium of the Skin and in Vascular Tumors

It is difficult to identify lymph vessels in tissue sections by histochemical staining, and thus a specific marker for lymphatic endothelial cells would be more practical in histopathological diagnostics. Here we have applied a specific antigenic marker for lymphatic endothelial cells in the human skin, the vascular endothelial growth factor receptor-3 (VEGFR-3), and show that it identifies a distinct vessel population both in fetal and adult skin, which has properties of lymphatic vessels. The expression of VEGFR-3 was studied in normal human skin by in situ hybridization, iodinated ligand binding, and immunohistochemistry. A subset of developing vessels expressed the VEGFR-3 mRNA in fetal skin as shown by in situ hybridization and radioiodinated vascular endothelial growth factor (VEGF)-C bound selectively to a subset of vessels in adult skin that had morphological characteristics of lymphatic vessels. Monoclonal antibodies against the extracellular domain of VEGFR-3 stained specifically endothelial cells of dermal lymph vessels, in contrast to PAL-E antibodies, which stained only blood vessel endothelia. In addition, staining for VEGFR-3 was strongly positive in the endothelium of cutaneous lymphangiomatosis, but staining of endothelial cells in cutaneous hemangiomas was weaker. These results establish the utility of anti-VEGFR-3 antibodies in the identification of lymphovascular channels in the skin and in the differential diagnosis of skin lesions involving lymphatic or blood vascular endothelium.     

食管癌血管内皮生长因子和受体的表达及在癌转移中的作用

目的观察人食管癌组织和淋巴管中血管内皮生长因子(VEGF-C)及其受体-3(VEGFR-3)的表达,探讨食管癌的淋巴道转移机制。方法取临床手术切除的食管癌组织,用免疫组化方法检测人食管癌早期和进展期癌细胞或淋巴管对VEGF-C及其VEGFR-3的表达情况。结果在食管癌的癌细胞中可见VEGF-C阳性表达,阳性颗粒主要定位于肿瘤细胞胞浆内。淋巴管内皮细胞仅见VEGFR-3阳性表达,VEGFR-3在血管和癌细胞中也存在少量阳性表达。进展期食管癌VEGF-C和VEGFR-3的表达率和表达强度均强于早期。结论食管癌癌细胞VEGF-C的表达和淋巴管内皮细胞上VEGFR-3的表达均与肿瘤进展呈正相关,推测VEGF-C通过受体VEGFR-3促进食管癌组织淋巴管生成,从而引起癌淋巴道转移。     

Localization of vascular endothelial growth factor-D in malignant melanoma suggests a role in tumour angiogenesis

Expression of angiogenic and lymphangiogenic factors by tumours may influence the route of metastatic spread. Vascular endothelial growth factor (VEGF) is a regulator of tumour angiogenesis, but studies of the inhibition of solid tumour growth by neutralizing anti-VEGF antibodies indicated that other angiogenic factors may be involved. VEGF-D may be an alternative regulator because like VEGF it is angiogenic and it activates VEGF receptor-2 (VEGFR-2), an endothelial cell receptor which is a key signalling molecule in tumour angiogenesis. This study reports the generation of monoclonal antibodies to the receptor-binding domain of VEGF-D and the use of these antibodies to localize VEGF-D in malignant melanoma. VEGF-D was detected in tumour cells and in vessels adjacent to immunopositive tumour cells, but not in vessels distant from the tumours. These findings are consistent with a model in which VEGF-D, secreted by tumour cells, activates endothelial cell receptors and thereby contributes to the regulation of tumour angiogenesis and possibly lymphangiogenesis. In addition, VEGF-D was detected in the vascular smooth muscle, but not the endothelium, of vessels in adult colon. The endothelium of these vessels was negative for VEGFR-2 and VEGFR-3. As VEGF receptors can be up-regulated on endothelium in response to vessel damage and ischaemia, these findings of a specific localization of VEGF-D in smooth muscle of the blood vessels suggest that VEGF-D produced by vascular smooth muscle could play a role in vascular repair by stimulating the proliferation of endothelial cells.     

NF-κBp65、VEGF-C及受体VEGFR-3在乳腺癌组织中的表达

目的检测乳腺癌组织内NF-κBp65、VEGF-C及受体VEGFR-3的表达水平,并观察它们与临床病理特征的关系,为乳腺癌的早期诊断、治疗提供依据。方法采用免疫组化染色法检测50例乳腺癌组织及10癌旁组织内NF-κBp65、VEGF-C及受体VEGFR-3的表达,并分析与临床病理因素的关系。结果乳腺癌组织内NF-κBp65蛋白表达的阳性率为76.0%,明显高于癌旁对照组织30.0%,二者相比差异显著(P<0.05);乳腺癌组织内VEGF-C蛋白表达的阳性率为84.0%,明显高于癌旁对照组织20.0%,二者相比差异显著(P<0.05);乳腺癌组织内VEGFR-3蛋白表达的阳性率为88.0%,明显高于癌旁对照组VEGFR-3阳性表达率20.0%,两者相比差异显著(P<0.05);乳腺癌组织内NF-κBp65、VEGF-C及受体VEGFR-3三者的表达存在显著相关性(P<0.05),均与肿瘤淋巴转移密切相关(P<0.05),而与病人年龄、肿瘤大小无关(P>0.05)。结论乳腺癌中NF-κBp65的表达可能上调VEGF-C的表达,进而导致肿瘤周围淋巴管增生、扩张,促进肿瘤细胞向区域淋巴结转移。     

VEGF-C、VEGFR-3和iNOS在人乳腺癌淋巴管的表达

目的检测乳腺癌组织血管内皮生长因子-C(VEGF-C)及其受体(VEGFR-3)、iNOS基因表达的相关性及这三个基因mRNA表达水平与淋巴管密度(LVD)的相关性,为阐明乳腺癌淋巴管生成的分子机理提供实验依据。方法RT-PCR检测乳腺癌组织及正常乳腺组织VEGF-C、VEGFR-3、iNOSmRNA的表达水平;免疫组织化学染色法检测淋巴管内皮细胞上VEGFR-3的表达,测定淋巴管密度。结果乳腺癌LVD为20.35±4.23,显著高于正常对照组(P<0.05);乳腺癌组织VEGF-C、VEGFR-3、iNOSmRNA三者的表达率分别为74.0%、84.0%、82.0%,显著高于正常对照组(P<0.05);VEGF-C、VEGFR-3、iNOS阳性组中LVD分别为21.34±3.45、18.54±4.68、17.43±4.76,显著高于对照组(P<0.05)。结论VEGF-C、VEGFR-3、iNOSmRNA表达与淋巴管密度之间呈正相关,并且3者之间的表达亦具有相关性,这些基因的表达增高可能在乳腺癌淋巴管生成过程中具有重要作用。     

Ultrastructural development of Kaposi's sarcoma in relation to the dermal microvasculature

Summary Ultrastructural subtypes of endothelial cells in Kaposi's sarcoma were compared with lymphatics and the normal dermal microcirculation in different stages of lesional development. In the earliest patch stage, lymphatic channels, recognised by their dissecting growth pattern and a lack of a basal lamina and pericytes, were prominent. Venous endothelium was recognised by virtue of its multilaminated basal lamina and often showed markedly irregular luminal and abluminal cytoplasmic projections. As the histological stage progressed toward spindle cells, venous endothelium showed a tetrad of changes: dissolution of the basal lamina; fragmentation and disappearance of the pericyte sheath, decreased and often rudimentary intercellular junctions and markedly reduced numbers of Weibel-Palade bodies. These were also features of spindle cells, which were rarely seen to emerge from narrow vascular channels of indeterminate type. Spindle cells showed sparse intercellular junctions and minimal basal lamina but no Weibel-Palade bodies. These progressive venous alterations thus resulted in a mixed intermediate subtype of endothelium with the morphological traits resembling lymphatics as well as venous blood vessels. The mixed subtype included endothelial tubes surrounded by a complete basal lamina but lacking pericytes, and much more commonly, tubes with pericytes but a scanty basal lamina. Both forms had remarkably few or no Weibel-Palade bodies. In the spindle cell stage, normal vessels were largely replaced by the mixed subtype and an indeterminate type of frequently disrupted endothelial tube which lacked a basal lamina as well as a pericytic investment. Dissecting lymphatic channels could not be confidently distinguished from the latter vessels. Direct anatomical connections between morphologically recognizable lymphatics and venules, blood capillaries or endothelial tubes of mixed subtype were difficult to demonstrate. Lymphaticovenous shunts were represented by gaps in all endothelial subtypes, the gaps sometimes due to cell degeneration.     

The ultrastructure of lymphatic valves in rabbits and mice

The ultrastructure of lymphatic valves was studied in rabbits and mice. The lymphatic valves usually consist of two cusps but three or four are sometimes present. The cusps are covered by endothelium. Along the free edge of the cusps are endothelial cells which can differentiate morphologically from other endothelial cells. They are named “tip-cells”; they have pseudopod-like projections and abundant cytoplasmic filaments 60–90 Å in diameter. Vesicles occur in endothelial cells of both lymphatic vessels and their valves; they are on the luminal or connective tissue side and are never provided with a diaphragm like that frequently observed in blood vessels. Joining endothelial cells are zonulae occludentes but desmosomes are not observed. No open intercellular junctions are encountered along the valvular endothelium. A basement membrane (basal lamina) is more frequently observed in valves than in walls of lymphatic vessels. Connective tissue in the cusps consists of collagenous fibrils, fine filaments and fibroblasts.     

人脐带血淋巴管内皮祖细胞的分化及其生物学特征

目的研究脐带血中CD34^＋／CD133^＋／VEGFR-3^＋淋巴管内皮祖细胞经VEGF—C诱导向内皮细胞分化过程中生物学特征的变化,并探讨其分化的机制。方法取脐带血,用PercoU密度梯度离心法分离单形核细胞,再用流式细胞仪分选CD34^＋／CD133^＋／VEGFR-3^＋细胞,然后用VEGF—C诱导分化。在扫描电镜和透射电镜下观察细胞表面形态和细胞内结构的变化,并在激光扫描共焦显微镜下观察特征性标志物的表达变化。结果脐带血中的淋巴管内皮祖细胞表达CD34、CD133和VEGFR-3。CD34^＋／CD133^＋／VEGFR-3^＋细胞经VEGF-C诱导后7d,呈长梭形,细胞伸出板状伪足和丝状伪足,出现较多短的微绒毛。表面可见细胞小凹,细胞质中含有丰富的线粒体和粗面内质网。诱导后14d,细胞已具有内皮细胞的特征,表达淋巴管内皮特异性标志物LYVE-1和5-核苷酸酶,CD133表达消失,细胞质中可见Weibel-Palade小体。结论脐带血中存在CD34^＋／CD133^＋／VEGFR-3^＋淋巴管内皮祖细胞,这些细胞在VEGF—C诱导作用下可能通过VEGF—C／VEGFR-3信号途径分化为淋巴管内皮细胞。