Adenovirus-mediated transfer of siRNA against basic fibroblast growth factor mRNA enhances the sensitivity of glioblastoma cells to chemotherapy.
Basic fibroblast growth factor (bFGF) is an important growth factor for glioma cell proliferation and invasion. BFGF is overexpressed in malignant gliomas and its level is associated with malignant grades and clinical outcome of patients with gliomas. Small interfering RNAs (siRNA) are synthetic forms of microRNA made of short double stranded RNA, and they effectively catalyze the degradation of their target mRNA. The use of siRNA has become a key method in the suppression of gene expression and the development of therapeutic agents. In this study, we used an adenovirus(Ad)-mediated transfer of siRNA against bFGF mRNA (Ad-bFGF-siRNA) to study the effect of down-regulating bFGF expression on the sensitivity of glioma cells to chemotherapeutics. An optimal siRNA sequence specific for bFGF mRNA was cloned into an adenoviral vector and transfected into three glioma cell lines: U251, A172, and LN229. Methyl thiazolyl tetrazolium (MTT) assays were used to examine changes in cell proliferation, and changes in bFGF mRNA and protein levels in U251 cells were detected using quantitative RT-PCR and Western blot, respectively. Apoptosis of U251 cells was detected using Hoechst staining and flow cytometry, with expression of apoptosis-related proteins evaluated by Western blot. Following the transfection of a bFGF-specific siRNA, mRNA and protein levels of bFGF decreased significantly. Lower rates of proliferation and increased levels of apoptosis also were associated with the Ad-bFGF-siRNA-transfected group compared to control group. Decreased expression of Bcl-2, Bcl-xL, Jak-1, and STAT-3 and increased expression of Bax also were detected in the Ad-bFGF-siRNA-transfected group. For cells treated with both Ad-bFGF-siRNA and chemotherapeutics, a significant reduction in cell survival was observed compared to treatment with Ad-bFGF-siRNA or chemotherapeutics alone. Overall, we found that targeting bFGF mRNA with a siRNA resulted in lower rates of proliferation, increased apoptosis, and enhanced sensitivity of glioma cells to chemotherapy drugs. This suggests that specific targeting of bFGF mRNA may provide a novel approach for the treatment of glioblastoma multiforme (GBM).
Feng
Xuequan
X
Department of Neurosurgery, Tianjin First Center Hospital, Fukang Road No 24, 300192 Tianjin, China.
Zhang
Biao
B
Wang
Jinhuan
J
Xu
Xinnv
X
Lin
Na
N
Liu
Hongsheng
H
D000256:Adenoviridae / Q000235:genetics*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D001932:Brain Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000473:pathology; D002330:Carmustine / Q000494:pharmacology; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D016222:Fibroblast Growth Factor 2 / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D005909:Glioblastoma / Q000188:drug therapy* / Q000235:genetics* / Q000473:pathology; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013713:Teniposide / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016222:Fibroblast Growth Factor 2; D013713:Teniposide; D002330:Carmustine