Distribution and expression of nephrin,podocin and α-actinin in a conditionally immortalized mouse podocyte cell line

Objective: To analyze the distribution and expression of nephrin, podocin and a-actinin in a conditionally immortalized mouse podocyte cell line (MPC 5), which will be in vitro model for studying the relationship among podocyte molecules. Methodology: MPC 5 were induced to propagate and passage by γ-inferon at 33℃, which were subsequently cultured at 37℃ for differentiation for two weeks. The podocyte morphology was observed by phase-contrast microscope. The distribution of nephrin, podocin and α-actinin was revealed by immunofluorescence (IF) staining. Results: ① In the light microscopy, differentiated cells presented with the polygoneal shape and with the formation of processes. ② IF the staining of nephrin and podcin was mainly distributed on the surface of cellular membrane in a filamentous pattern. The staining of α-actinin was specifically localized to the nuclear. ③ Expressions of nephrin, podocin and α-actinin mRNAs in MPC 5 were detected by RT-PCR. ④ Expression of nephrin, podocin, α-actinin and WT1 in MPC 5 were shown by Western blotting with the size of approximately 180 KDα, 45 kdα, 100 KDα and 52 KDα, respectively. Conclusion: For the first time, we demonstrated the expression of nephrin, podocin and α-actinin both at mRNA level and at protein level in mouse MPC 5.