Overexpression of FADD enhances 5-fluorouracil-induced apoptosis in colorectal adenocarcinoma cells.
To investigate the mechanism of enhancing apoptosis-inducing effects of 5-fluorouracil on human colorectal adenocarcinoma cells by stable transfection of extrinsic Fas-associated death domain protein (FADD) gene, both in vitro and in vivo. FADD gene of stable overexpression was determined by reverse transcription polymerase chain reaction (RT-PCR) assay and Western blotting assay. After treatment with 5-fluorouracil as an apoptotic inducer, in vitro cell growth activities were investigated by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Cell apoptosis and its rates were evaluated by TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling) assay and flow cytometry of annexin V-FITC/PI staining. To examine the combination therapeutic effect of FADD and 5-fluorouracil, tumor xenograft model was prepared for in vivo study. Compared with SW480 and SW480/neo cells, FADD mRNA and protein levels of SW480/FADD cells were higher. Chemosensitivity and apoptosis rates of SW480/FADD cells were remarkably higher than SW480 and SW480/neo cells when treated with 5-fluorouracil. In in vivo study, overexpression of FADD increased the efficacy of 5-fluorouracil-induced inhibition of tumor growth in nude mice. Stable overexpression of extrinsic FADD gene can conspicuously ameliorate apoptosis-inducing effects of 5-fluorouracil on colorectal adenocarcinoma cells, which is a novel strategy to improve chemotherapeutic effects on colorectal cancer.
Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, 430060 Hubei, People's Republic of China.
Anning
A
Yin
Yingan
Y
Jiang
Xianfeng
X
Zhang
Hesheng
H
Luo
D000230:Adenocarcinoma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D053200:Fas-Associated Death Domain Protein / Q000096:biosynthesis* / Q000235:genetics; D005472:Fluorouracil / Q000494:pharmacology* / Q000627:therapeutic use; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D015854:Up-Regulation / Q000187:drug effects* / Q000502:physiology; D023041:Xenograft Model Antitumor Assays / Q000379:methods
D053200:Fas-Associated Death Domain Protein; D005472:Fluorouracil