Effects Of IFN-γ And TNF-α On The Apoptosis Of Insulinoma Cells Line Min6

Objective: To investigate the effects of IFN-γ and TNF-α on the apoptosis of isletcells and the insulinoma cell line MIN6.Methods:1. NOD/Ltj mouse islets cell viability was analyzed by MTT assay aftertreatment of IFN-γ and TNF-α.2. MIN6cell viability was analyzed by MTT assayafter treatment of IFN-γ and TNF-α. Morphological change of MIN6cells wasobserved under the inverted microscope. Nucleus morphological change of MIN6cells was observed under the laser scanning confocal microscope. DNA ladder wasanalyzed by agarose electrophoresis. Effect of MIN6was detected after treatment ofIFN-γ and TNF-α by FACS. Early expression of Caspase-3in apoptosis of MIN6cellswas detected by western blotting.Results:1. By MTT it shows that it has cytotoxicity on islet by treatment of IFN-γalone, and it significantly decreased cell viability when treatment of IFN-γ and TNF-αtogether.2. By MTT it showes that it has cytotoxicity to MIN6by treatment of IFN-γ orTNF-α alone, and it has synergism cytotoxicity to MIN6cells when treatment ofIFN-γ and TNF-α together in a time-dependent way (P<0.05). MIN6cells grow wellin a tile laminated shape. MIN6cells change into spherical shape and begin to floatafter treatment with IFN-γ or TNF-α for12,24or48hours. It has little effect onchange of nucleus shape when treatment of IFN-γ or TNF-α alone for48hours. Whentreatment of IFN-γ and TNF-α together, it showes classical apoptotic morphologysuch as cell nucleus shrinkage, chromatin condensation, and so on. Agaroseelectrophoresis displays that MIN6DNA shows obvious DNA apopotic fragmentsstimulated by IFN-γ and TNF-α. It has significant DNA apoptotic fragments whentreatment with IFN-γ and TNF-α together. In contrast to IFN-γ or TNF-α alone, IFN-γand TNF-α synergism had an obvious inhibition in the viability of MIN6cells andcaused significant apoptosis in MIN6cells. Casepase-3was down regulated aftertreatment of IFN-γ and TNF-α together.Conclusions:1. IFN-γ has toxic effect to islet cells from NOD/Ltj mice, and it has a significant apoptosis in islet cells when treatment IFN-γ and TNF-α together.2. IFN-γ or TNF-α has cytotoxicity to MIN6cells, and it has synergismcytotoxicity when treatment IFN-γ and TNF-α together in a time-dependent way. Itcaused a significant early apoptosis in MIN6cells not necrosis.3. Through down regulating caspase-3, IFN-γ and TNF-α induces MIN6cellapoptosis.