Effects of WT1 gene downregulation on apoptosis in porcine fetal fibroblasts

• Published in: In vitro cellular & developmental biology. Animal 2014-06, Vol.50 (6), p.555-561
• Main Authors: An, Peipei, Ding, Yu, Wang, Aibing, Tan, Wentao, Gao, Fei, Fan, Anran, Tang, Bo, Zhang, Xueming, Li, Ziyi
• Format: Article
• Language: English
• Subjects: Animal Genetics and Genomics; Animals; Annexins; Apoptosis; Apoptosis - genetics; bcl-2-Associated X Protein - biosynthesis; Biomedical and Life Sciences; Cell Biology; Cell Culture; CELL GROWTH/DIFFERENTIATION/APOPTOSIS; Cell lines; cell proliferation; Cell Proliferation - genetics; Cells, Cultured; chromosomes; Developmental Biology; Down regulation; Epithelial cells; Fetal Proteins - biosynthesis; Fibroblasts; Fibroblasts - metabolism; gene expression regulation; genes; Kidneys; Life Sciences; Messenger RNA; mutation; plasmids; Proto-Oncogene Proteins c-bcl-2 - biosynthesis; reverse transcriptase polymerase chain reaction; RNA; RNA Interference; RNA, Small Interfering; Stem Cells; Swine; testes; Tumors; Up-Regulation; Western blotting; WT1 Proteins - biosynthesis; WT1 Proteins - genetics; zinc finger motif
• ISSN: 1071-2690; 1543-706X
• DOI: 10.1007/s11626-014-9738-1

Wilms’ tumor gene 1 (WT1) is located on chromosome 11p13. Besides a role in the development of Wilms’ tumor, specific mutations in the Zn finger region are found in Denys-Drash syndrome and Frasier syndrome, both characterized by urogenital abnormalities, sometimes in combination with Wilms’ tumor. Our past study shows that WT1 is expressed in porcine kidney fibroblasts (PKFs) and swine testis cells (ST cells) and is essential for the maintenance of the development and survival of PKFs and ST cells. But we do not know whether WT1 gene was expressed in porcine fetal fibroblasts or not. To further explore whether WT1 was expressed in porcine fetal fibroblasts (PFFs) and its contribution to cell apoptosis, RT-PCR, immunocytochemical staining, and Western blot were used to detect the expression of WT1, the recombinant plasmids of pLV3-WT1 short hairpin ribonucleic acid (shRNA) were used to downregulate the WT1 gene in porcine fetal fibroblasts, and the role of WT1 in cell proliferation was examined by apoptosis analysis also. Our results indicated that WT1 was expressed in PFFs, the pLV3-WT1 shRNA dramatically reduced the expression of WT1, and downregulation of WT1 directly led to early cell apoptosis by downregulating the expression of antiapoptotic gene Bcl-2 and upregulating the expression of proapoptotic gene Bax in PFFs. Our results demonstrate that WT1 is also essential for the maintenance of the survival of PFFs.