Cell Populations

PT cells were generated from the Immorto-mouse crossed with the Tgfbr2^fl/fl mice as previously described and characterized.¹⁶ PT cells were grown at 33°C in DMEM/F12 supplemented with 2.5% FBS, hydrocortisone, insulin, transferrin, selenium, triiodothyronine, and penicillin/streptomycin (complete PT media) with IFN-γ.¹⁶ Before experiments, PT cells were moved to 37°C, and IFN-γ was removed to induce differentiation. Deletion of TβRII in PT was achieved by adeno-Cre treatment in vitro and verified by immunoblots.¹⁶

For primary cultures, uninjured renal cortices were dissected, minced, and incubated in a shaker for 45 minutes in complete PT media with collagenase I (1 mg/ml), dispase (1 mg/ml), and DNAse at 37°C. The tissue was passed through a 70-μM filter to remove glomeruli, washed two times with complete PT media, plated, and used for experiments at passage 2.