Silencing of Bcl-XL expression in human MGC-803 gastric cancer cells by siRNA.
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.
Lei
Xiao-Yong
XY
Institute of Pharmacy and Pharmacology, Nanhua University, Hengyang 421001, China. Lei_xiaoyong@hotmail.com
Zhong
Miao
M
Feng
Lan-Fang
LF
Yan
Chun-Yan
CY
Zhu
Bing-Yang
BY
Tang
Sheng-Song
SS
Liao
Duan-Fang
DF
D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D017209:Apoptosis / Q000235:genetics; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D018390:Gene Targeting / Q000379:methods; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D034741:RNA, Small Interfering / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013274:Stomach Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D014162:Transfection / Q000379:methods; D051020:bcl-X Protein / Q000235:genetics* / Q000378:metabolism*
C494811:BCL2L1 protein, human; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D051020:bcl-X Protein