HIF-1α Deletion Modulates the Ratio of Treg/Th17 cells in CD4 T Cells and Improves Vascularized Composite Allotransplantation Survival under Costimulatory Blockade
Cheng-Hung Lin1, Madonna R Anggelia1,2, Huang-Yu Yang3, Yi-Ching Ko3, Nicholas Do1, Gerald Brandacher4, WP Andrew Lee4.
1Center for Vascularized Composite Allotransplantation, Department of Plastic and Reconstructive Surgery, Chang Gung Memorial Hospital, Chang Gung Medical College and Chang Gung University, Taoyuan, Taiwan, 2Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan, 3Department of Nephrology, Chang Gung Memorial Hospital, Taoyuan, Taiwan, 4Department of Plastic and Reconstructive Surgery, Vascularized Composite Allotransplantation (VCA) Laboratory, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
PURPOSE: Hypoxia-inducible factor (HIF-1α) regulates the balance between effector T cells (Teff) and regulatory T cells (Tregs) by enhancing Th17 development (helper T cell subtype) and attenuating Tregs development via degradation of FoxP3, whose expression defines Tregs. The balance between Tregs and Th17 cells impacts allograft survival. This study investigates the consequence of HIF-1α deficiency on T cell differentiation and vascularized composite allograft (VCA) survival using knockout mice.
To assess the effect of HIF-1α deficiency upon T cell subtypes differentiation, na´ve CD4+ T cells derived from wild type (WT) and HIF-1α deficient (HIF-1αfl/flCD4CreC57BL/6) mice were incubated in 2 types helper T cell subtypes (Th1 and Th17) and Tregs-skewing conditions. Degrees of IFN-γ, IL-17A and FoxP3 expressions were then quantified. Mixed lymphocyte reaction (MLR) was then used to assess effect of HIF-1α deficiency on CD4+ T cell proliferation. To evaluate effect on VCA survival, 13 WT and 15 HIF-1αfl/flCD4CreC57BL/6 mice received osteomyocutaneous allografts from Balb/c mice. Immunosuppressive regimen consisted only of co-stimulatory blockade (1 mg anti-CD154 at POD 0, 0.5 mg CTLA4Ig at POD 2). Allograft survival, ratios of Tregs/Teff subpopulations in the periphery and within the allograft were assessed. RESULTS: Unlike from WT mice, the na´ve CD4+ T cells from HIF-1αfl/flCD4CreC57BL/6 mice not expressed IFN-γ and IL-17A after incubated in Th1 and Th17-skewing conditions. However, those cells still expressed FoxP3 in Tregs-skewing conditions although the degree of expression was lower than WT mice (p<0.01). HIF-1α deficient CD4+ T cells did not result in T cells proliferation during MLR analysis. In the transplanted mice, a higher ratio of Tregs/Th1 cells and Tregs/Th17 cells in the periphery was observed in tolerant animals and lower ratio of both in graft rejecting animals (p<0.05). With co-stimulatory blockade, improved allograft median survival time (MST) occurred with HIF-1αfl/flCD4Cremice compared to WT mice (MST = 100 vs 36.5 days, p<0.01). Infusion/adoptive transfer of 5x106 Th17 cells at POD 0 in HIF-1αfl/flCD4Crerecipients disrupted allograft survival (MST = 60 days) (Figure 1).
CONCLUSION: HIF-1α deficiency affects the differentiation of na´ve CD4+ T cells, ratio of Tregs/Teff, and improves allograft survival in the setting of co-stimulatory blockade. Targeting potential mechanisms involved in CD4+ T cell differentiation, such as HIF-1α, may be a viable treatment approach in improving allograft tolerance.
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