title: Knockdown of secreted protein acidic and rich in cysteine (SPARC) expression diminishes radiosensitivity of glioma cells.
abstract: Secreted protein acidic and rich in cysteine (SPARC) has been shown to play an important role in the promotion of glioma. In this study, we investigated the effects of downregulated SPARC expression on the radiosensitivity of human glioma U-87MG cells and its possible mechanism. With a small-interfering RNA (siRNA) expression plasmid vector targeting SPARC, we obtained the stably transfected cells in which the expression of SPARC was successfully downregulated. Then, the cells were irradiated with 60Co-γ-rays and analyzed by several methods, such as clonogenic assay, flow cytometry, comet assay, and western blotting. Clonogenic assay showed that downregulation of SPARC expression enhanced cell survival after radiation. Flow cytometry analysis indicated that SPARCsiRNA decreased cell apoptosis responding to irradiation. Analysis of signaling molecules with western blotting showed that the level of Akt phosphorylation was increased in irradiated U-87MG/SPARCsiRNA cells. Further, cell-cycle analysis by flow cytometry showed enhanced G2 accumulation in U-87MG/SPARCsiRNA cells after irradiation. Comet assay revealed that SPARCsiRNA promoted the repair of radiation-induced DNA damage. Our results suggest that inhibition of SPARC expression may diminish the radiosensitivity of human glioma U-87MG cells. One of the mechanisms for this effect may be associated with the reduced cell apoptosis responding to radiation, which may be contributed by the phosphoinositide 3-kinase/AKT pathway activation. Moreover, enhanced G2 accumulation and increased DNA repair may also account for the decreased radiosensitivity.
authors.affiliation: School of Radiation Medicine and Protection, Medical College of Soochow University, Suzhou, Jiangsu Province, China.
authors.forename: Haiyan
authors.initials: H
authors.lastname: Liu
authors.forename: Haowen
authors.initials: H
authors.lastname: Zhang
authors.forename: Xin
authors.initials: X
authors.lastname: Jiang
authors.forename: Yan
authors.initials: Y
authors.lastname: Ma
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Xu
authors.forename: Shuang
authors.initials: S
authors.lastname: Feng
authors.forename: Fenju
authors.initials: F
authors.lastname: Liu
mesh_terms: D017209:Apoptosis / Q000235:genetics; D001932:Brain Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000532:radiotherapy*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000235:genetics; D004249:DNA Damage / Q000235:genetics; D004260:DNA Repair / Q000235:genetics; D015536:Down-Regulation; D016195:G2 Phase / Q000235:genetics; D055785:Gene Knockdown Techniques / Q000379:methods; D005910:Glioma / Q000235:genetics / Q000378:metabolism* / Q000532:radiotherapy*; D006801:Humans; D015676:Osteonectin / Q000096:biosynthesis / Q000172:deficiency* / Q000235:genetics*; D010766:Phosphorylation / Q000235:genetics; D046988:Proteasome Endopeptidase Complex; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D011836:Radiation Tolerance; D014162:Transfection / Q000379:methods
chemical_list: D015676:Osteonectin; C121623:PSMF1 protein, human; D011506:Proteins; D034741:RNA, Small Interfering; D051057:Proto-Oncogene Proteins c-akt; D046988:Proteasome Endopeptidase Complex