Milatz Susanne, Krug Susanne M, Rosenthal Rita, Günzel Dorothee, Müller Dominik, Schulzke Jörg-Dieter, Amasheh Salah, Fromm Michael
Institute of Clinical Physiology, Charité, Campus Benjamin Franklin, Berlin, Germany.
Biochim Biophys Acta. 2010 Nov;1798(11):2048-57. doi: 10.1016/j.bbamem.2010.07.014. Epub 2010 Jul 22.
The paracellular barrier of epithelia and endothelia is established by several tight junction proteins including claudin-3. Although claudin-3 is present in many epithelia including skin, lung, kidney, and intestine and in endothelia, its function is unresolved as yet. We therefore characterized claudin-3 by stable transfection of MDCK II kidney tubule cells with human claudin-3 cDNA. Two clone systems were analyzed, exhibiting high or low claudin-2 expression, respectively. Expression of other claudins was unchanged. Ultrastructurally, tight junction strands were changed toward uninterrupted and rounded meshwork loops. Functionally, the paracellular resistance of claudin-3-transfected monolayers was strongly elevated, causing an increase in transepithelial resistance compared to vector controls. Permeabilities for mono- and divalent cations and for anions were decreased. In the high-claudin-2 system, claudin-3 reduced claudin-2-induced cation selectivity, while in the low-claudin-2 system no charge preference was observed, the latter thus reflecting the "intrinsic" action of claudin-3. Furthermore, the passage of the paracellular tracers fluorescein (332Da) and FD-4 (4kDa) was decreased, whereas the permeability to water was not affected. We demonstrate that claudin-3 alters the tight junction meshwork and seals the paracellular pathway against the passage of small ions of either charge and uncharged solutes. Thus, in a kidney model epithelium, claudin-3 acts as a general barrier-forming protein.
上皮细胞和内皮细胞的细胞旁屏障是由包括闭合蛋白-3在内的几种紧密连接蛋白建立的。尽管闭合蛋白-3存在于包括皮肤、肺、肾和肠道在内的许多上皮细胞以及内皮细胞中，但其功能尚未明确。因此，我们通过用人类闭合蛋白-3 cDNA稳定转染MDCK II肾小管细胞来对闭合蛋白-3进行表征。分析了两个克隆系统，分别表现出高或低的闭合蛋白-2表达。其他闭合蛋白的表达未发生变化。在超微结构上，紧密连接链向不间断的圆形网状环改变。在功能上，转染了闭合蛋白-3的单层细胞的细胞旁电阻显著升高，与载体对照相比，导致跨上皮电阻增加。单价和二价阳离子以及阴离子的通透性降低。在高闭合蛋白-2系统中，闭合蛋白-3降低了闭合蛋白-2诱导的阳离子选择性，而在低闭合蛋白-2系统中未观察到电荷偏好，后者因此反映了闭合蛋白-3的“内在”作用。此外，细胞旁示踪剂荧光素（332Da）和FD-4（4kDa）的通过减少，而对水的通透性不受影响。我们证明闭合蛋白-3改变紧密连接网络，并封闭细胞旁途径以阻止任何电荷的小离子和不带电荷的溶质通过。因此，在肾脏模型上皮细胞中，闭合蛋白-3作为一种通用的屏障形成蛋白发挥作用。
