The effects of the WT1 gene on apoptosis and development‐related gene expression in porcine kidney fibroblasts and swine testis cells

Abstract

The Wilm's tumor 1 gene (WT1) encodes zinc finger proteins that function as tumor suppressors, and play important roles in the development of the genito-urinary system and other organs. Its precise function in the development of porcine tissues and organs, which is an attractive transplantation resource for certain human diseases, is still unclear. Here, we sought to define the role of WT1 in porcine kidney and testis tissues using porcine kidney fibroblasts (PKFs) and swine testis (ST) cells as in vitro models, both of which express WT1. The recombinant plasmids pLV3-WT1 shRNA and pIRES2 -WT1-EGFP were constructed to respectively down- and up-regulate the WT1 gene in porcine cells. The role of WT1 in cell proliferation was investigated by RT-PCR, immunocytochemical staining, apoptosis analysis, and Western blot. The pLV3-WT1 shRNA dramatically reduced WT1 expression at both the transcription and protein levels. The down-regulation of WT1 directly led to early cell apoptosis, and changes in Sf1, Sox9, and Gdnf gene expression in PKFs and ST cells. In contrast, up-regulation of WT1 gave no obvious phenotype in ST cells. Our results demonstrate that WT1 is essential for the survival of PKFs and ST cells because it regulates apoptosis- and development-related genes in the cells; however, no obvious effect was observed when WT1 was over-expressed.