Jansen J, De Napoli I E, Fedecostante M, Schophuizen C M S, Chevtchik N V, Wilmer M J, van Asbeck A H, Croes H J, Pertijs J C, Wetzels J F M, Hilbrands L B, van den Heuvel L P, Hoenderop J G, Stamatialis D, Masereeuw R
Department of Pharmacology and Toxicology, Radboud Institute for Molecular Life Sciences, Nijmegen, The Netherlands.
Department of Physiology, Radboud Institute for Molecular Life Sciences, Nijmegen, The Netherlands.
Sci Rep. 2015 Nov 16;5:16702. doi: 10.1038/srep16702.
The bioartificial kidney (BAK) aims at improving dialysis by developing 'living membranes' for cells-aided removal of uremic metabolites. Here, unique human conditionally immortalized proximal tubule epithelial cell (ciPTEC) monolayers were cultured on biofunctionalized MicroPES (polyethersulfone) hollow fiber membranes (HFM) and functionally tested using microfluidics. Tight monolayer formation was demonstrated by abundant zonula occludens-1 (ZO-1) protein expression along the tight junctions of matured ciPTEC on HFM. A clear barrier function of the monolayer was confirmed by limited diffusion of FITC-inulin. The activity of the organic cation transporter 2 (OCT2) in ciPTEC was evaluated in real-time using a perfusion system by confocal microscopy using 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide (ASP(+)) as a fluorescent substrate. Initial ASP(+) uptake was inhibited by a cationic uremic metabolites mixture and by the histamine H2-receptor antagonist, cimetidine. In conclusion, a 'living membrane' of renal epithelial cells on MicroPES HFM with demonstrated active organic cation transport was successfully established as a first step in BAK engineering.
生物人工肾（BAK）旨在通过开发用于细胞辅助清除尿毒症代谢产物的“活性膜”来改善透析。在此，将独特的人类条件永生化近端肾小管上皮细胞（ciPTEC）单层培养在生物功能化的聚醚砜（MicroPES）中空纤维膜（HFM）上，并使用微流控技术进行功能测试。通过沿着HFM上成熟ciPTEC紧密连接处大量表达的闭合蛋白-1（ZO-1）蛋白，证明了紧密单层的形成。通过FITC-菊粉的有限扩散证实了单层的明确屏障功能。使用灌注系统通过共聚焦显微镜，以4-（4-（二甲基氨基）苯乙烯基）-N-甲基碘化吡啶鎓（ASP(+)）作为荧光底物，实时评估ciPTEC中有机阳离子转运体2（OCT2）的活性。初始ASP(+)摄取受到阳离子尿毒症代谢产物混合物和组胺H2受体拮抗剂西咪替丁的抑制。总之，在BAK工程的第一步中，成功建立了具有活性有机阳离子转运功能的MicroPES HFM上的肾上皮细胞“活性膜”。
