Establishment and functional characterization of the reversibly immortalized mouse glomerular podocytes(imPODs)

Glomerular podocytes are highly specialized epithelial cells and play an essential role in establishing the selective permeability of the glomerular filtration barrier of kidney.Maintaining the viability and structural integrity of podocytes is critical to the clinical management of glomerular diseases,which requires a thorough understanding of podocyte cell biology.As mature podocytes lose proliferative capacity,a conditionally SV40 mutant tsA58-immortalized mouse podocyte line(designated as tsPC)was established from the Immortomouse over 20 years ago.However,the utility of the tsPC cells is hampered by the practical inconvenience of culturing these cells.In this study,we establish a user-friendly and reversibly-immortalized mouse podocyte line(designated as imPOD),on the basis of the tsPC cells by stably expressing the wildtype SV40 T-antigen,which is flanked with FRT sites.We show the imPOD cells exhibit long-term high proliferative activity,which can be effectively reversed by FLP recombinase.The imPOD cells express most podocyte-related markers,including WT-1,Nephrin,Tubulin and Vinculin,but not differentiation marker Synaptopodin.The imPOD cells do not form tumor-like masses in vivo.We further demonstrate that TGFb1 induces a podocyte injury-like response in the FLP-reverted imPOD cells by suppressing the expression of slit diaphragm-associated proteins P-Cadherin and ZO-1 and upregulating the expression of mesenchymal markers,a-SMA,Vimentin and Nestin,as well as fibrogenic factors CTGF and Col1a1.Collectively,our results strongly demonstrate that the newly engineered im-POD cells should be a valuable tool to study podocyte biology both under normal and under pathological conditions.