Abstract:

Ｏｂｊｅｃｔｉｖｅ：To explore effects of siRNA knockdown podocalyxin on the structure and function of podocytes.
Ｍｅｔｈｏｄｏｌｏｇｙ：Podocytes were cultured in RPMI 1640 medium at 33 ℃,and switch to 37 ℃ for differentiation and maturation.Cells were then divided into 5 groups:Vehicle,Mock,Shscramble control(Shnc),siRNAPODXL (Si),and ezrin inhibition group(NSC305787).Fluorescence microscopy was used to evaluate the changes in cell structure and actin cytoskeleton after phalloidin staining of Factin.The wound healing experiment and Transwell test were used to assess cell migration,and flow cytometry was used to detect apoptosis of podocyte.Western blot and RTqPCR were used to quantify  change of PODXL and pezrin gene and protein.
Ｒｅｓｕｌｔｓ：(1) Cytoskeleton of podocyte in Vehicle group was normal,while in the cells with PODXL knockdown or with ezrin inhibited,Factin stress fibers were rearranged and lost (P<005).There was no significant change of Factin stress fibers in the Mock and Shnc groups compared with Vehicle group.(2) The wound healing and Transwell experiments both showed that  migration of podocyte in the Si group,ezrin inhibition group were slower than that of the other three groups.(3) Flow cytometry showed that apoptosis increased in cells with PODXL knockdown or ezrin inhibition.(4) Western blotting and RTPCR analysis showed that  PODXL was successfully knocked down in  Si group,accompanied by downregulation of pezrin but not total ezrin compared with  cells in the Vehicle group; (5) Compared with Vehicle group,the activity of RhoA decreased significantly in Si group and ezrin inhibition group,but there was no significant change of RhoA activity in  Mock and Shnc groups compared with vehicle group.
Ｃｏｎｃｌｕｓｉｏｎ：
PODXL plays a key role on  podocyte cytoskeletons and survival,and  maintains the level of podocyte pezrin which may be crucial for podocyte cytoskeletons and survival

Key words: podocytes, cytoskeleton, apoptosis