Differentiated transport functions in primary cultures of rabbit collecting ducts.
Differentiated transport functions in primary cultures of rabbit collecting ducts.
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兔集合管原代培养物中分化的运输功能。
DOI：
10.1152/ajprenal.1987.253.6.f1302
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发表时间：
1987
期刊：
影响因子：
--
通讯作者：
Naray-Fejes-Toth,A
中科院分区：
文献类型：
--
作者：
Fejes-Toth,G;Naray-Fejes-Toth,A
用固相免疫吸附法分离兔皮质集合管（CCT）的片段。从“免疫切割”细胞开始的原代培养物随后在多孔底培养皿中生长至汇合。使用一组细胞特异性单克隆抗体和凝集素，通过流式细胞术和免疫细胞化学验证新鲜分离和培养的细胞的纯度。由于持续的矢量运输，培养的上皮细胞单层产生显着的transepithelial溶质梯度。培养48小时后，腔（L）和基底外侧（B）培养基的组成如下：[Na]，L 119 +/- 1.1，B 145 +/- 3.0; [K]，L 11.9 +/- 0.5，B 2.0 +/- 0.3; [PO 4]，L 2.8 +/- 0.2，B 7.3 +/- 0.3;[Cl]，L 134.1 +/- 2.7 B 117.8 +/- 2.2，[mM]; [Ca]，L 42.1 +/- 1.2，B 812 +/- 26，[microM]; pH，L 5.52 +/- 0.12，B 7.15 +/- 0.08。跨上皮渗透浓度差为14.3 +/- 3.5 mosmol/l。管腔侧的葡萄糖量保持不变，而基底外侧的葡萄糖浓度从21降至7.7 +/- 1.0 mM。乳酸主要在基底外侧蓄积（L 1.0 +/- 0.15，B 23.5 +/- 1.7 mM）。这些数据表明，免疫切割的CCT细胞的原代培养物保留了许多其分化的运输功能，因此应证明有用的研究机制的溶质运输在集合管系统。
Fragments of rabbit cortical collecting tubule (CCT) were isolated by solid-phase immunoadsorption with a monoclonal antibody against an ectoantigen on principal cells. Primary cultures, initiated from the "immunodissected" cells, were then grown to confluency in porous bottom dishes. The purity of the freshly isolated and cultured cells was verified by flow cytometry and immunocytochemistry, using a panel of cell-specific monoclonal antibodies and lectins. As a result of continued vectorial transport, the cultured epithelial monolayers generated pronounced transepithelial solute gradients. The composition of the luminal (L) and basolateral (B) media after 48 h of incubation was as follows: [Na], L 119 +/- 1.1, B 145 +/- 3.0; [K], L 11.9 +/- 0.5, B 2.0 +/- 0.3; [PO4], L 2.8 +/- 0.2, B 7.3 +/- 0.3; [Cl], L 134.1 +/- 2.7 B 117.8 +/- 2.2, [mM]; [Ca], L 42.1 +/- 1.2, B 812 +/- 26, [microM]; pH, L 5.52 +/- 0.12, B 7.15 +/- 0.08. The transepithelial osmotic concentration difference was 14.3 +/- 3.5 mosmol/l. The amount of glucose at the luminal side remained unaltered, whereas glucose concentration at the basolateral side decreased from 21 to 7.7 +/- 1.0 mM. Lactic acid accumulated predominantly at the basolateral side (L 1.0 +/- 0.15, B 23.5 +/- 1.7 mM). These data indicate that primary cultures of immunodissected CCT cells retain many of their differentiated transport functions and thus should prove useful for studying the mechanisms of solute transport in the collecting duct system.