Tissue engineering of a bioartificial renal tubule assist device: In vitrotransport and metabolic characteristics

Background. Current renal substitution therapy for acute or chronic renal f ailure with hemodialysis or hemofiltration is life sustaining, but continue s to have unacceptably high morbidity and mortality rates. This therapy is not complete renal replacement therapy because it does not provide active t ransport nor metabolic and endocrinologic functions of the kidney, which ar e located predominantly in the tubular elements of the kidney. Methods. To optimize renal substitution therapy, a bioartificial renal tubu le assist device (RAD) was developed and tested in vitro for a variety of d ifferentiated tubular functions. High-flux hollow-tiber hemofiltration cart ridges with membrane surface areas of 97 cm(2) or 0.4 m(2) were used as tub ular scaffolds. Porcine renal proximal tubule cells were seeded into the in traluminal spaces of the hollow fibers, which were pretreated with a synthe tic extracellular matrix protein. Attached cells were expanded in the cartr idge as a bioreactor system to produce confluent monolayers containing up t o 1.5 x 10(9) cells (3.5 x 10(5) cells/cm(2)). Near confluency was achieved along the entire membrane surface, with recovery rates for perfused inulin exceeding 97 and 95% in the smaller and larger units, respectively, compar ed with less than 60% recovery in noncell units. Results. A single-pass perfusion system was used to assess transport charac teristics of the RADs. Vectorial fluid transport from intraluminal space to antiluminal space was demonstrated and was significantly increased with th e addition of albumin to the antiluminal side and inhibited by the addition of ouabain, a specific inhibitor of Na+,K+-ATPase. Other transport activit ies were also observed in these devices and included active bicarbonate tra nsport, which was decreased with acetazolamide, a carbonic anhydrase inhibi tor, active glucose transport, which was suppressed with phlorizin, a speci fic inhibitor of the sodium-dependent glucose transporters, and para-aminoh ippurate (PAH) secretion, which was diminished with the anion transport inh ibitor probenecid. A variety of differentiated metabolic functions was also demonstrated in the RAD. Intraluminal glutathione breakdown and its consti tuent amino acid uptake were suppressed with he irreversible inhibitor of g amma-glutamyl transpeptidase acivicin; ammonia production was present and i ncremented with declines in perfusion pH. Finally, endocrinological activit y with conversion of 25-hydroxy(OH)vitamin D-3 to 1,25-(OH)(2) vitD(3) was demonstrated in the RAD. This conversion activity was up-regulated with par athyroid hormone and down-regulated with increasing inorganic phosphate lev els, which are well-defined physiological regulators of this process in viv o. Conclusions. These results clearly demonstrate the successful tissue engine ering of a bioartificial RAD that possesses critical differentiated transpo rt, and improves metabolic and endocrinological functions of the kidney. Th is device, when placed in series with conventional hemofiltration therapy, may provide incremental renal replacement support and potentially may decre ase the high morbidity and mortality rates observed in patients with renal failure.