Apoptosis of Renal Cell Carcinoma Cells by Expression of FADD (Fas-Associated Death Domain).
- Author:
In Gab JEONG
1
;
Cheol KWAK
;
Hyeon JEONG
;
Sang Eun LEE
Author Information
1. Departments of Urology, Seoul National University College of Medicine, Korea. urology@snu.ac.kr
- Publication Type:In Vitro ; Original Article
- Keywords:
FADD;
DED;
Carcinoma;
renal cell;
Apoptosis
- MeSH:
Apoptosis*;
Blotting, Western;
Carcinoma, Renal Cell*;
Caspase 3;
Cell Line;
Clone Cells;
Cytoplasm;
DNA Fragmentation;
DNA, Complementary;
Humans;
Plasmids
- From:Korean Journal of Urology
2003;44(5):436-445
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
PURPOSE: The Fas-associated death domain (FADD) constitutes a novel protein that specifically associates with the cytoplasmic death domain of Fas, and induces apoptosis. FADD is composed of a death effector domain (DED) and a death domain. In this study, we evaluated the in vivo antitumor effect of the FADD, or FADD-DED, gene in renal cell carcinoma cells, using a plasmid vector expressing the human FADD and FADD-DED genes. MATERIALS AND METHODS: The cDNA of the human FADD and FADD-DED genes were amplified by RT-PCR, and cloned to the pCR(R) 3.1. The expressions of the cloned FADD and FADD-DED (pCR(R)3.1-FADD and pCR3.1-FADD-DED) were observed by Western blot analysis. The efficacy of the growth inhibition by the cloned FADD and FADD-DED genes was tested, in vitro, on A498 and Caki-1 human renal cell carcinoma cell lines using the MTT assay. To evaluate the apoptosis, DNA fragmentation and caspase-3 assays were performed. RESULTS: Expressions of the FADD protein, and the FADD-DED, of the transfected A498 and Caki-1 cells had increased by 48 and 24 hours, respectively, compared with the control cell lines. The cytotoxicity of the pCR3.1-FADD and pCR3.1-FADD-DED on the A498 and Caki-1 cells significantly increased compared to the empty vector. The increased cytotoxicity of the FADD- or FADD-DED-transfected cell lines was associated with enhanced apoptosis, as assessed by DNA fragmentation and caspase-3 assays. CONCLUSIONS: Our results showed that the cloned FADD or FADD-DED expression plasmid vector efficiently inhibited the growth of A498 and Caki-1 human renal cell carcinoma cell lines. These data suggest that the exogenous FADD or FADD-DED expressions may have therapeutic applications in renal cell carcinomas.
