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B. Liu, Z. Li, S. P. Mahesh, F. S. Hwang, S. Pantanelli, W. O. Siu, R. B. Nussenblatt; GITR Promotes Activation Induced Apoptosis of Human NK Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):5181.
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To investigate glucocorticoid-induced tumor necrosis factor receptor (GITR)’s role in activation-induced cell death (AICD) of NK cells and possible mechanisms.
Human PBMCs from normal donors were isolated from their buffy coat using Ficoll gradient centrifugation. Highly purified NK cells were obtained from isolated PBMCs by magnetic sorting technique using a negative NK isolation kit. Combination of IL2 and IL15 was used to stimulate NK cells. Apoptotic cells were detected by staining of cells with the combination of annexin-V-FITC and Via-probeTM (7-Amino-actinomycin D, 7-AAD) followed by flow cytometry. SDS-PAGE and western blot analysis were used to detect apoptosis related signaling molecules.
About 45% of the NK population from human PBMCs constitutively expresses GITR. Upon activation by cytokines, NK cells greatly up-regulated GITR expression. TLR4 agonist LPS also up-regulated GITR expression in purified NK cells. A stimulating GITR monoclonal antibody reduced proliferation and enhanced apoptosis of cytokine-activated NK cells. The mitochondrial pathway, but not the Fas-FasL pathway was involved in GITR-enhanced apoptosis. Finally, we found that IL2/IL15 stimulation elevated cytoplasmic expression of p21cip1/waf1 in NK cells while GITR activation decreased IL2/IL15-induced cytoplasmic expression of p21cip1/waf1.
The results support the notion that GITR functions as a costimulator in cytokine-induced AICD of NK cells and reveal a novel function of GITR in the regulation of innate immune response.
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