RÉSUMÉ
Objective To investigate the effect of all-trans retinoic acid (ATRA) on expressions of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in glioma stem cells (GSCs). Methods GSCs were isolated from human glioblastoma cell line U87 and identified by detecting the expressions of CD133 and nestin with immunofluorescence staining. The obtained GSCs were divided into control group,empty vector group (cultured with dimethyl sulfoxide [DMSO]) and ATRA treatment group (cultured with 10 nmool/L ATRA).After 10 d of differentiation; the proliferation of the treated GSCs was evaluated using CCK8 assay; the expressions of glial fibrillary acidic protein (GFAP),β-tubulin Ⅲ and galactoeerebroside (GralC) in the cells were detected by immunofluorescence.VEGF and bFGF levels in cultured supernatant were measured by ELISA; the mRNA expressions of VEGF and bFGF were detected by RT-PCR. Results The target antibodies of neural stem cells (NSCs), CD133 and nestin,positively expressed in the GSCs; differentiated GSCs can differentiate several kinds of homologous daughter cells,which expressed the cell markers of astrocytes,neurons and oligodendrocytes: GFAP, β-tubulin LⅢ and GalC, respectively. The percentage of GFAP-positive differentiated GSC s in the ATRA treatment group was significantly higher as compared with that in the other 2 groups after 10 d of differentiation (P＜0.05); the speed of proliferation of GSCs in ATRA treatment group was obviously slower than that in the other 2 groups 3-7 d after differentiation (P＜0.05).The VEGF and bFGF levels and the mRNA expression levels of VEGF and bFGF in GSCs of the ATRA treatment group 24 h after differentiation were also significantly lower than those in the other 2 groups (P＜0.05). Conclusion ATRA can induce the differentiation of GSCs and inhibit its proliferation.It may exerts its anti-glioblastoma effect through the VEGF and bFGF signaling pathways.
RÉSUMÉ
Objective To study the effect of transforming growth factor-β (TGF-β) signaling pathway blockage on vasculogenic mimicry (VM) in gliomas and explore its possible mechanism.Methods Three-dimentional culture was performed on the glioma cell lines U251 and SHG44; the effects of U251 culture supematant and TGF-β on VM formation of SHG44 cells were observed; the capability of VM formation of U251 and SHG44 cells after being treated with 0 μg/mL (PBS group),15 μg/mL TGF-β neutralizing antibody (Ab15 group) and 30 μg/mL TGF-β neutralizing antibody (Ab30 group) was evaluated.ELISA was used to detect the concentrations of vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF) in the supematant of U251 cells from the blank group,PBS group,Ab 15 group and Ab30 group and the concentrations of VEGF and PDGF in the supernatant of SHG44 cells from the blank group,TGF-β treatment group,PBS group,Ab15 group and Ab30 group.Results VM was formed in the U251 cells while not in the SHG44 cells during the three-dimentional culture; SHG44 cells could only gather into colonies of different sizes.U251 culture supernatant could induce SHG44 cells to form VM,enjoying the most obvious effect at 24-48 h of culture; TGF-β could not induce SHG44 cells to form VM.The number of U251 cells annulation in PBS group,Ab15 group and Ab30 group decreased in sequence with significant difference (P＜0.05).The number of U251 cells armulation in SHG44 cells cultured in U251 culture supematant from the PBS group,Ab15 group and Ab30 group decreased in sequence after being added TGF-β antibody with significant difference (P＜0.05).As compared with that in the blank group and PBS group,significant decrease of VEGF and PDGF concentrations in the U251 cells from Ab15 group and Ab30 group was noted (P＜0.05); as compared with that in the blank group and TGF-β treatment group,significant increase of VEGF and PDGF concentrations in the SHG44 cells from PBS group,Ab15 group and Ab30 group was noted.Conclusion Blockage of TGF-β signaling pathways inhibits VM in glioma,and it maybe probably due to the decrease of VEGF and PDGF expressions..