Development of bioartificial renal tubule devices with lifespan-extended XXXX renal proximal tubular epithelial cells.

@entity1 @entity23032 @entity986 @entity5 @entity1372 @entity177 @entity876 @entity6393 @entity413
BACKGROUND: The bioartificial renal tubule device is a cell therapy system for @entity177 . The major obstacle in the development of the bioartificial renal tubule device is the obtainment of a large number of viable renal tubule cells to seed on the inner surface of hollow fibers. Although our previous studies had used a transformed cell line, they may be dangerous for clinical uses. Therefore, different approaches to amplify renal proximal tubular epithelial cells (RPTEC) in culture without oncogenes, vectors and carcinogens have been required. METHODS: The limitation of the replicative lifespan of @entity1 RPTEC, which is    12 population doublings (PDs), was extended by invalidating messenger RNA of cell cycle-related genes with antisense oligonucleotide or small interfering RNA (siRNA). RESULTS: Periodic transfection of siRNA to a @entity5 suppressor @entity876 or a cyclin-dependent kinase inhibitor @entity986 ( @entity986 ) extended the lifespan by 33 and 63 PDs, respectively, in 3 months of culture. The siRNA-mediated lifespan extension was controllable because cell division ceased within 2 weeks after the transfection was discontinued. Expressions of y-glutamyltransferase 1 and @entity413 transporter 1 were recovered in siRNA-transfected RPTEC cultured on porous membranes. Bioartificial renal tubule devices (0.8 m(2)) constructed with these cells showed reabsorption of water (122.3    4.2 mL/30 min), @entity1372 (18.1    0.7 mEq/30 min) and @entity413 (121.7    4.4 mg/30 min) after 1 week of circulation. Furthermore, @entity6393 and @entity23032 were metabolized by RPTEC in mini-devices (65 cm(2)) within 48 h of circulation. CONCLUSIONS: These approaches enabled us to yield a high enough number of RPTEC for construction of bioartificial renal tubule devices repeatedly. Lifespan-extended RPTEC could recover their specific characteristics by culturing on porous membranes, and bioartificial renal tubule devices constructed with these cells showed good performances of reabsorption and metabolism. SUMMARY: A large number of @entity1 renal tubular cells required for construction of the bioartificial renal tubule device were prepared by extending the lifespan of the primary cells by invalidating mRNA of cell cycle-related genes. Constructed bioartificial renal tubule devices with lifespan-extended cells showed good performances of in vitro examination of reabsorption and metabolism. Requiring no oncogenes, vectors or cell cloning, the RNAi-mediated lifespan extension can help advance tissue-replacement therapy as well as basic research.
