Molecular mechanisms of the combination of retinoid and interferon-gamma for inducing differentiation and increasing apoptosis in human glioblastoma T98G and U87MG cells.
Glioblastoma is the deadliest brain tumor that remains incurable. We examined efficacy of combination of retinoid and interferon-gamma (IFN-gamma) in human glioblastoma T98G and U87MG cells. We conjectured that retinoid could induce differentiation with down regulation of telomerase activity to increase sensitivity to IFN-gamma for apoptosis in glioblastoma cells. Indeed, treatment of cells with 1 muM all-trans retinoic acid (ATRA) or 1 muM 13-cis retinoic acid (13-CRA) for 7 days induced astrocytic differentiation with upregulation of glial fibrillary acidic protein (GFAP) and down regulation of telomerase activity. Wright staining and ApopTag assay showed, respectively, morphological and biochemical features of apoptosis in glioblastoma cells following exposure to 200 units/ml IFN-gamma for 48 h. Induction of differentiation was associated with decreases in levels of nuclear factor kappa B (NFkappaB), inducible nitric oxide synthase (iNOS), and production of nitric oxide (NO) so as to increase sensitivity to IFN-gamma for apoptosis. Notably, IFN-gamma induced signal transducer and activator of transcription-1 (STAT-1) to bind to gamma-activated sequence (GAS) of the target gene. Also, IFN-gamma activated caspase-8 and cleaved Bid to truncated Bid (tBid) for translocation to mitochondria. Fura-2 assay showed increases in intracellular free [Ca2+] and activation of calpain in apoptotic cells. Besides, increases in Bax:Bcl-2 ratio and mitochondrial release of cytochrome c and Smac into the cytosol activated caspase-9 and caspase-3 for apoptosis. Taken together, our results showed that retinoid induced astrocytic differentiation with down regulation of telomerase activity and enhanced sensitivity to IFN-gamma for increasing apoptosis in human glioblastoma cells.
Das
Arabinda
A
Department of Neurosciences, Medical University of South Carolina, Charleston, SC 29425, USA.
Banik
Naren L
NL
Ray
Swapan K
SK
D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins; D051023:BH3 Interacting Domain Death Agonist Protein / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism; D020170:Caspase 1 / Q000096:biosynthesis; D053181:Caspase 8 / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D015536:Down-Regulation; D005904:Glial Fibrillary Acidic Protein / Q000096:biosynthesis; D005909:Glioblastoma / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D007249:Inflammation / Q000503:physiopathology; D007371:Interferon-gamma / Q000494:pharmacology*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D015474:Isotretinoin / Q000494:pharmacology; D008928:Mitochondria / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D012176:Retinoids / Q000494:pharmacology*; D019098:Telomerase / Q000378:metabolism; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
D051017:Apoptosis Regulatory Proteins; D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; D002135:Calcium-Binding Proteins; C412403:DIABLO protein, human; D005904:Glial Fibrillary Acidic Protein; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; D012176:Retinoids; D014212:Tretinoin; C033668:calpastatin; D007371:Interferon-gamma; C496319:NOS2 protein, human; D052247:Nitric Oxide Synthase Type II; D019098:Telomerase; D053181:Caspase 8; D020170:Caspase 1; D015474:Isotretinoin