The Culture Of Glioblastoma-Derived Stem Cells And The Studies On The Mechanism Of TRAIL Resistance

Glioma is the most common primary brain tumors, in which glioblastoma is the most common and most harmful type. Glioblastoma has the characteristics of high incidence, recurrence rate, mortality and low cure rate,"three high and one low".It is a serious threat to human health. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a member of Tumor necrosis factor superfamily, can be expressed in the normal immune system,which can play a role in immune surveillance on tumor cells.TRAIL-pathway is an important anti-tumor way in vivo,it can induce apoptosis in cancer cells; however, clinical trials of TRAIL apoptotic pathway-targeted therapies have had disappointing results. In examining the molecular basis of TRAIL resistance in glioblastomas, we show here that glioblastoma-derived stem cells (GSCs) are resistant to TRAIL treatment. In this study, we generated neurospheres, GSC091106 and GSC001112 and matched serum-grown cultures, SC091106 and SC091112 from glioblastomas surgically removed from patients. The neurospheres were characterized for having the GSC properties including self-renewal capacity, expression of stem cell marker CD 133 and CD 15 and xenograft formation in SCID mouse brain.The serum-grown cultures but not neurospheres underwent TRAIL-induced apoptosis as evident by cell death and caspase-8,-3, and -7 activities. Lack of the cell surface expression of the death receptors, DR4 and DR5, as shown by flow cytometry and the overexpression of cellular FADDlike interleukin-1 -converting enzyme-inhibitory protein (c-FLIP), as identified by western blot in GSC-enriched neurospheres contributed to the TRAIL resistance. Cisplatin treatment led to the up-regulation of DR5 and down-regulation of c-FLIP and thus restore TRAIL apoptotic pathwayin GSC-enriched neurospheres. This study identifies the molecular basis of TRAIL resistance in GSCs and suggests that the combination of chemotherapy and TRAIL may provide an effective treatment of glioblastomas.The conclusion:(1) GSC is resistant to TRAIL-induced apoptosis due to the lack of the cell surface expression of DR4 and DR5 and the overexpression of intracellular c-FLIP proteins.(2) Chemotherapeutic agent cisplatin can up-regulate DR5 and down-regulate c-FLIP in GSC and eliminate the resistance of GSC to TRAIL treatment.(3) The combination therapy of cisplatin and TRAIL may provide a novel and effective treatment for glioblastomas.The main innovations:(1) The glioblastoma-derived stem cells is more resistant to TRAIL induced apoptosis compared with primary cultured glioblastoma cells(2) DR5 and c-FLIP is directly related to TRAIL induced apoptosis in glioblastoma-derived stem cells.(3)Cisplatin can up-regulate DR5 and down-regulate c-FLIP in glioblastoma-derived stem cells.