A bioartificial kidney device with polarized secretion of immune modulators

The accumulation of protein-bound toxins in dialyzed patients is strongly associated with their high morbidity and mortality. The bioartificial kidney device (BAK), containing proximal tubule epithelial cells (PTEC) seeded on functionalized synthetic hollow fiber membranes (HFM), may be a powerful solution for the active removal of those metabolites. In an earlier study, we developed an upscaled BAK containing conditionally immortalized human PTEC (ciPTEC) with functional organic cationic transporter 2 (OCT2). Here, we first extended this development to a BAK device having cells with the organic anionic transporter 1 (OAT1), capable of removing anionic uremic wastes. We confirmed the quality of the ciPTEC monolayer by confocal microscopy and paracellular inulin-FITC leakage, as well as, by the active transport of anionic toxin, indoxyl sulfate (IS). Furthermore, we assessed the immune-safety of our system by measuring the production of relevant cytokines by the cells after lipopolysaccharide (LPS) stimulation. Upon LPS treatment, we observed a polarized secretion of pro-inflammatory cytokines by the cells: 10-fold higher in the extraluminal space, corresponding to the urine compartment, as compared to the intraluminal space, corresponding to the blood compartment. To the best of our knowledge, our work is the first to show this favorable cell polarization in a BAK upscaled device

A bioartificial kidney device with polarized secretion of immune modulators

The accumulation of protein-bound toxins in dialyzed patients is strongly associated with their high morbidity and mortality. The bioartificial kidney device (BAK), containing proximal tubule epithelial cells (PTEC) seeded on functionalized synthetic hollow fiber membranes (HFM), may be a powerful solution for the active removal of those metabolites. In an earlier study, we developed an upscaled BAK containing conditionally immortalized human PTEC (ciPTEC) with functional organic cationic transporter 2 (OCT2). Here, we first extended this development to a BAK device having cells with the organic anionic transporter 1 (OAT1), capable of removing anionic uremic wastes. We confirmed the quality of the ciPTEC monolayer by confocal microscopy and paracellular inulin-FITC leakage, as well as, by the active transport of anionic toxin, indoxyl sulfate (IS). Furthermore, we assessed the immune-safety of our system by measuring the production of relevant cytokines by the cells after lipopolysaccharide (LPS) stimulation. Upon LPS treatment, we observed a polarized secretion of pro-inflammatory cytokines by the cells: 10-fold higher in the extraluminal space, corresponding to the urine compartment, as compared to the intraluminal space, corresponding to the blood compartment. To the best of our knowledge, our work is the first to show this favorable cell polarization in a BAK upscaled device

Human proximal tubule epithelial cells cultured on hollow fibers : living membranes that actively transport organic cations

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

Human proximal tubule epithelial cells cultured on hollow fibers : living membranes that actively transport organic cations

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