Lentivirus-mediated RNA interference suppresses APRIL expression and enhances chemosensitivity in colorectal cancer cells.
- Author:
Jing GUAN
1
;
Ai-min SUN
;
Li-hui WANG
;
Mei-rong HE
Author Information
- Publication Type:Journal Article
- MeSH: Apoptosis; drug effects; Cell Cycle; drug effects; Cell Line, Tumor; Cell Proliferation; drug effects; Colorectal Neoplasms; drug therapy; metabolism; pathology; Drug Resistance, Neoplasm; Fluorouracil; pharmacology; Humans; Lentivirus; genetics; RNA Interference; Tumor Necrosis Factor Ligand Superfamily Member 13; genetics; metabolism
- From: Journal of Southern Medical University 2011;31(9):1600-1604
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo investigate the effects of lentivirus-mediated RNA interference (RNAi) targeting a proliferation-inducing ligand (APRIL) on the chemosensitivity to 5-FU of colorectal cancer cell line LoVo.
METHODSThe lentiviral vector siRNA-APRIL was constructed and verified by PCR and DNA sequencing. LoVo cells were transfected with siRNA-APRIL plasmid, non-targeting siRNA plasmid, or empty plasmid. Forty-eight hours after the transfection, the cells were examined for APRIL expression using Western blot. Seventy-two hours after treatment with 10 µg/ml 5-FU, flow cytometry was used to detect the cell apoptosis and cell cycle changes. The cell growth inhibition rate following 5-FU exposure was detected by MTT assay.
RESULTSPCR analysis and DNA sequencing demonstrated that the RNAi sequence targeting APRIL gene was successfully inserted into the lentiviral vector. siRNA-APRIL transfection resulted in obviously reduced expression of APRIL in LoVo cells. After 5-FU exposure, the apoptosis rate of siRNA-APRIL-transfected cells were increased to (21.12∓3.35)%, significantly higher than that in cells transfected with the non-targeting plasmid or the empty plasmid [(13.06∓1.92)% and (12.28∓1.79)%, respectively, P<0.01]; the cell number in G0/G1 phase increased while that in G2/M phase decreased in siRNA-APRIL-transfected cells. The growth inhibition rate in siRNA-APRIL group was (59.67∓5.03)%, significantly higher than that in the other two groups [(42.33∓4.16)% and (39.67∓4.73)%, respectively, P<0.01].
CONCLUSIONLentivirus-mediated RNAi targeting APRIL can effectively suppress the expression of APRIL in LoVo cells and enhance the chemosensitivity of the cells to 5-FU.