Conditionally Immortal Slc4a11-/- Mouse Corneal Endothelial Cell Line Recapitulates Disrupted Glutaminolysis Seen in Slc4a11-/- Mouse Model.

摘要: Purpose To establish conditionally immortal mouse corneal endothelial cell lines with genetically matched Slc4a11+/+ and Slc4a11-/- mice as a model for investigating pathology therapies SLC4A11 associated congenital hereditary dystrophy (CHED) Fuchs' dystrophy. Methods We intercrossed H-2Kb-tsA58 (Immortomouse) expressing an IFN-γ dependent temperature-sensitive mutant of the SV40 large T antigen (tsTAg) C57BL/6 mice. The growth characteristics was assessed by doubling time. Ion transport activities (Na+/H+ exchange, bicarbonate, lactate, Slc4a11 ammonia transport) were analyzed intracellular pH measurement. metabolic status analyzing TCA cycle intermediates via gas chromatography mass spectrometry (GC-MS). Results immortalized cells (MCECs) remained proliferative through passage 49 maintained similar active ion activity. As expected, proliferation temperature sensitive dependent. MCECs exhibited decreased capacity, reduced NH3:H+ transport, altered expression glutaminolysis enzymes to mouse, proportion derived from glutamine compensatory increases in glucose flux compared MCECs. Conclusions This is first report immortalization remains active, except transporter activity Furthermore, recapitulate defects observed endothelium, providing excellent tool study pathogenesis mutations dystrophies screen potential therapeutic agents.