Krishnamurti U, Chen Y, Michael A, Kim Y, Fan W W, Wieslander J, Brunmark C, Rondeau E, Sraer J D, Delarue F, Tsilibary E C
Department of Laboratory Medicine and Pathology, University of Minnesota, Minneapolis, USA.
Lab Invest. 1996 Mar;74(3):650-7.
The use of human glomerular epithelial cells (HGEC) in research has been severely restricted by several obstacles, which have been circumvented by the generation of T-SV40 immortalized human visceral glomerular epithelial cells (Delarue et al, 1991). In this work, we compared the primary and immortalized HGEC for expression of integrin and some nonintegrin surface receptors. We also studied the adhesion of both types of HGEC to glomerular basement membrane (GBM), type IV collagen (tIV), and its major noncollagenous NC1 domain. The integrins mediating adhesion of HGEC to tIV were also examined. Expression of integrin and some nonintegrin cell surface receptors was analyzed by flow cytometry. Adhesion to GBM, tIV, and its major noncollagenous NC1 domain was studied by direct solid phase cell adhesion assays. Identification of integrins mediating adhesion of HGEC to tIV was achieved by inhibition of cell adhesion using monoclonal antibodies to integrin subunits. The primary and immortalized HGEC share phenotypic characteristics, and alpha3beta1 appeared to be the major integrin present on both HGEC types. The kinetics of binding to GBM, tIV, and its noncollagenous NCI domain were similar in both the primary and immortalized HGEC, although the latter displayed a somewhat weaker binding. Both the primary and immortalized HGEC displayed significantly better adhesion to NC1-alpha3 compared with NC1-alpha1, alpha3beta1 appears to be the major integrin mediating the adhesion of HGEC to tIV. Our studies suggest that alpha3beta1 is the major integrin present on HGEC. This has been confirmed by flow cytometric analysis. In addition, we demonstrated a functional role for this integrin in mediating attachment of HGEC to tIV. Our data also demonstrate a preference in binding of HGEC to alpha3 chains of NC1 compared with alpha1 chains of NC1. These findings were seen in both the primary and immortalized HGEC. The T-SV40 immortalized HGEC can therefore serve as a very useful tool to study glomerular visceral cell biology.
人类肾小球上皮细胞（HGEC）在研究中的应用受到了诸多障碍的严重限制，而T-SV40永生化人类内脏肾小球上皮细胞的产生（Delarue等人，1991年）克服了这些障碍。在这项工作中，我们比较了原代和永生化HGEC中整合素和一些非整合素表面受体的表达。我们还研究了这两种类型的HGEC对肾小球基底膜（GBM）、IV型胶原蛋白（tIV）及其主要非胶原NC1结构域的黏附。我们还检测了介导HGEC与tIV黏附的整合素。通过流式细胞术分析整合素和一些非整合素细胞表面受体的表达。通过直接固相细胞黏附试验研究对GBM、tIV及其主要非胶原NC1结构域的黏附。使用针对整合素亚基的单克隆抗体抑制细胞黏附来鉴定介导HGEC与tIV黏附的整合素。原代和永生化HGEC具有共同的表型特征，α3β1似乎是两种类型HGEC上存在的主要整合素。原代和永生化HGEC与GBM、tIV及其非胶原NCI结构域结合的动力学相似，尽管后者的结合力稍弱。与NC1-α1相比，原代和永生化HGEC对NC1-α3的黏附均显著更好，α3β1似乎是介导HGEC与tIV黏附的主要整合素。我们的研究表明，α3β1是HGEC上存在的主要整合素。这已通过流式细胞术分析得到证实。此外，我们证明了这种整合素在介导HGEC与tIV附着中的功能作用。我们的数据还表明，与NC1的α1链相比，HGEC对NC1的α3链的结合具有偏好性。在原代和永生化HGEC中均观察到了这些发现。因此，T-SV40永生化HGEC可作为研究肾小球内脏细胞生物学的非常有用的工具。
