Isolation and characterization of a novel mouse lymphatic endothelial cell line: SV-LEC
Isolation and characterization of a novel mouse lymphatic endothelial cell line: SV-LEC
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DOI：
10.1089/lrb.2005.3.105
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发表时间：
2005-01-01
影响因子：
1.4
通讯作者：
Alexander, J. S.
中科院分区：
文献类型：
--
作者：
Ando, T.;Jordan, P.;Alexander, J. S.
背景资料：淋巴系统调节间质液和蛋白质平衡，并通过调节白细胞和抗原运输到淋巴结来调节免疫应答。本文描述了一种稳定的小鼠肠系膜外膜组织淋巴管内皮细胞系（SV-LEC），该细胞系不同于血液主动脉（AEC）和静脉（VEC）内皮细胞，其基于几种淋巴标志物（例如，Prox-1、LYVE-1、Flt-4）。SV-LEC还响应于TNF-α表达MAdCAM-1，这是在VEC中观察到的效应，但在AEC中没有。从表达温度敏感性SV40大T细胞的小鼠肠系膜外膜分离淋巴管内皮细胞（SV-LEC），（"永生小鼠"，H-2K（B）-tsA58）在低氧气氛中，在补充有VEGF-C的D-缬氨酸取代的MEM中用低氧培养物选择（0%0 - 2、5%C02和95%N-2）与5 mM巯基乙酸盐。采用RT-PCR、免疫印迹和荧光显微镜检测细胞特异性标志物（Flt-4、LYVE-1、Prox-1）和紧密连接蛋白（ZO-1）的表达。MAdCAM-1（一种高内皮微静脉标记物）的表达也在对TNF-α、IL-1 β和IFN-γ的反应中进行了检测。LYVE-1和Prox-1的免疫印迹显示SV-LEC和VEC上有强表达，但AEC上没有。结论：SV-LEC表达多种淋巴管内皮细胞标志物，其中部分标志物与VEC共有，而与AEC无关，SV-LEC可作为体外淋巴管功能模型的一个有用系统。
Background: The lymphatic system regulates interstitial fluid and protein balance and modulates immune responses by regulating leukocyte and antigen traffic to lymph nodes. The present article describes a stable mouse lymphatic endothelial cell line from mesenteric adventitial tissue (SV-LEC) which is distinct from blood aortic (AEC) and venous (VEC) endothelial cells, based on expression of several lymphatic markers (e.g., Prox-1, LYVE-1, Flt-4). SV-LEC also expresses MAdCAM-1 in response to TNF-alpha, an effect seen in VEC, but not AEC.Methods and Results: Lymphatic endothelial cells (SV-LEC) were isolated from mesenteric adventitia from mice expressing temperature-sensitive SV40 large T ('Immortomouse', H-2K(b)-tsA58) selected with hypoxia culture in D-valine-substituted MEM supplemented with VEGF-C in a low oxygen atmosphere (0% O-2, 5% CO2, and 95% N-2) with 5 mM thioglycolate. Expression of lymphatic-specific markers (Flt-4, LYVE-1, Prox-1) and the tight junction proteins (ZO-1) were examined by RT-PCR, immunoblotting, and fluorescent microscopy. MAdCAM-1 (a high endothelial venular marker) expression was also examined in response to TNF-alpha, IL-1 beta and IFN-gamma.Results: Message for Flt-4 and LYVE-1 was detected on SV-LEC. Immunoblotting for LYVE-1 and Prox-1 showed strong expression on SV-LEC and VEC, but not AEC. Occludin expression was seen in all cell types, junctional ZO-1 was detected at SV-LEC and VEC junctions, not AEC.Conclusion: SV-LEC expresses several lymphatic endothelial markers, some of which are shared with VEC, but not AEC, and may represent a useful system for modeling lymphatic function in vitro.