All-trans Retinoic Acid Inhibit Medulloblastoma Cells Growth Mechanism

Backround:Medulloblastoma is a kind of cerebellum neuroepithelial tumor,it is the most common central nervous system tumors in children.The survival times reach 5 years of patients was 70%,however tumor metastasis in a part of high risk patients, their 5-years survival rate is only 25%.All-trans retinoic acid(ATRA) is a comparative matured and adopted retinoid medicine in the clinical cancer chemical therapy and prevention.It has been widely used for its abilities in anti-proliferation, pro-differentiation,and the inhibition of cells growth in many tumors.Objective:To study the anti-proliferation effects of ATRA on MB39 cells and the approach of its action.Methods:Medulloblastoma cell line MB39 cells were treated with ATRA at different concentrations.The proliferation inhibition was examined by CCK-8 assay.Flow cytometry(FITC/PI double staining flow cell method and caspase-3 staining flow cell method) was used to detect the apoptosis.Western blotting was used to detect the expression of IRS-1,PI3K,Akt,p-Akt,NF-κB,IKKα,IκBαand the expression of apoptosis-related protein Bax,Bcl-2.We establish a culture system to isolate and passage tumor stem cells from MB39 cell line.Futhermore,we investigate the effection of ATRA on MB39 stem/progenitor cells.And identify the MB39 stem/progenitor cells with immunofluorescence cell stain technology. Results:Treatment of ATRA could result in the growth inhibition of MB39 cells,and the cell density significantly decreased(P＜0.01).The MB39 cells display a normal fibroblast-like morphology under the microscope before the induction,and the ATRA treated cells displayed the typically apoptosis morphlogy.The ATRA-treated MB39 cells display obvious apoptosis by flowcytometry(P＜0.01 ),and it is caspase-3 dependent apoptosis pathway.Western blotting investigate the changes of MB39 protein with the treatment of ATRA for 24 hours,the results show the expression of IRS-1, PI3K,Akt,p-Akt,NF-κB,Bcl-2 were significantly declined,and the expression of IKKα,IκBα,Bax were enhanced.After inducing cultrue of MB39 cells,we obtain a small part cells which possess stem cell characteristics;ATRA can inhibit MB39 cells forming sphere and growth suspended.