Institute of Clinical Physiology, Charité, Campus Benjamin Franklin, Freie Universität and Humboldt-Universität, 12200 Berlin, Germany.
J Cell Sci. 2010 Jun 1;123(Pt 11):1913-21. doi: 10.1242/jcs.060665. Epub 2010 May 11.
Whether or not significant amounts of water pass the tight junction (TJ) of leaky epithelia is still unresolved, because it is difficult to separate transcellular water flux from TJ-controlled paracellular water flux. Using an approach without differentiating technically between the transcellular and paracellular route, we measured transepithelial water flux with and without selective molecular perturbation of the TJ to unequivocally attribute changes to the paracellular pathway. To this end, MDCK C7 cells were stably transfected with either claudin-2 or claudin-10b, two paracellular cation-channel-forming TJ proteins that are not endogenously expressed in this cell line. Claudin-2 is typical of leaky, water-transporting epithelia, such as the kidney proximal tubule, whereas claudin-10b is present in numerous epithelia, including water-impermeable segments of the loop of Henle. Neither transfection altered the expression of endogenous claudins or aquaporins. Water flux was induced by an osmotic gradient, a Na(+) gradient or both. Under all conditions, water flux in claudin-2-transfected cells was elevated compared with vector controls, indicating claudin-2-mediated paracellular water permeability. Na(+)-driven water transport in the absence of an osmotic gradient indicates a single-file mechanism. By contrast, claudin-10b transfection did not alter water flux. We conclude that claudin-2, but not claudin-10b, forms a paracellular water channel and thus mediates paracellular water transport in leaky epithelia.
水是否大量通过渗漏上皮的紧密连接（TJ）仍未解决，因为很难将细胞旁水通量与 TJ 控制的细胞旁水通量分开。我们使用一种不区分细胞旁和细胞内途径的方法，测量了 TJ 选择性分子扰动前后的跨上皮水通量，以明确将变化归因于细胞旁途径。为此，我们用 claudin-2 或 claudin-10b 稳定转染 MDCK C7 细胞，这两种 TJ 蛋白都是细胞旁阳离子通道形成蛋白，在该细胞系中不表达。 Claudin-2 是渗漏性、水转运上皮的典型代表，如肾脏近端小管，而 claudin-10b 存在于许多上皮中，包括 Henle 环的水不可渗透部分。转染均未改变内源性 Claudin 或水通道蛋白的表达。水通量由渗透梯度、Na+梯度或两者共同诱导。在所有条件下，与载体对照相比，claudin-2 转染细胞的水通量升高，表明 Claudin-2 介导的细胞旁水通透性。在没有渗透梯度的情况下，Na+驱动的水转运表明是单分子机制。相比之下，claudin-10b 转染并未改变水通量。我们得出结论， Claudin-2 而不是 claudin-10b 形成细胞旁水通道，因此介导渗漏性上皮的细胞旁水转运。
