Open AccessSilencing of Bcl‐XL Expression in Human MGC‐803 Gastric Cancer Cells by siRNA
Author(s) -
LEI XiaoYong,
ZHONG Miao,
FENG LanFang,
YAN ChunYan,
ZHU BingYang,
TANG ShengSong,
LIAO DuanFang
Publication year - 2005
Publication title -
acta biochimica et biophysica sinica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.771
H-Index - 57
eISSN - 1745-7270
pISSN - 1672-9145
DOI - 10.1111/j.1745-7270.2005.00077.x
Subject(s) - transfection , small interfering rna , green fluorescent protein , microbiology and biotechnology , gene silencing , bcl xl , flow cytometry , apoptosis , biology , acridine orange , expression vector , chemistry , cell culture , gene , programmed cell death , biochemistry , recombinant dna , genetics
To investigate the inhibitory effect of the Bcl‐XL small interfering RNA (siRNA) on Bcl‐XL gene expression in the human gastric cancer cell line MGC‐803, green fluorescent protein (GFP) siRNA was constructed and transfected into MGC‐803 cells, together with GFP expression vector pTrace SV40. GFP expression levels were observed using fluorescence microscopy. Bcl‐XL siRNA and negative siRNA were then constructed and stably transfected into MGC‐803 cells. RT‐PCR and immunofluorescence were used to detect the expression of Bcl‐XL. Spontaneous apoptosis was detected by acridine orange (AO) and flow cytometry. Results were as follows: (1) 48 h after GFP expression vector and GFP siRNA co‐transfection, the expression level of GFP in the GFP siRNA group was much lower than the negative siRNA group, according to fluorescence microscopy results. The mRNA and protein levels of Bcl‐XL in Bcl‐XL siRNA stable transfectants were reduced to almost background level compared with negative siRNA transfectants or untreated cells. (2) Changes in nucleus morphology was observed by AO staining nucleic and flow cytometry analysis, which showed that stable Bcl‐XL siRNA transfectants have an increased spontaneous apoptosis (21.17%±1.26% vs. 1.19%±0.18% and 1.56%±0.15% respectively, P <0.05 vs. negative siRNA or untreated control). siRNA targeting GFP or Bcl‐XL genes can specifically suppress GFP or Bcl‐XL expression in MGC‐803 cells, and Bcl‐XL siRNA can increase spontaneous apoptosis. Bcl‐XL siRNA may be a beneficial agent against human gastric adenocarcinoma. Edited by
You‐Xin JIN