HIF-1α is a negative regulator of plasmacytoid DC development in vitro and in vivo

Abstract

IntroductionMammalian hematopoiesis in the BM is regulated among others by the oxygen (O 2 ) availability. O 2 concentrations in the BM range from anoxia to 6% opposed to 4%-14% in well-oxygenated tissues, including the blood. 1,2 Recent data indicate that O 2 gradients within the BM participate in keeping hematopoietic stem cells (HSCs) in a low-replicating pluripotent state. HSCs are located in an extremely hypoxic niche as demonstrated by dye-perfusion and engraftment studies. 3,4 Hypoxia-inducible factors 1-3 (HIF-1-HIF-3) are stabilized by a low pO 2 to induce adaptive gene expression. They are heterodimers consisting of distinct O 2 -sensitive ␣-subunits and a stable common ␤-subunit, also known as aryl hydrocarbon receptor nuclear translocator (ARNT). 5 HIF-1 and HIF-2 were recently connected to HSC biology. In their hypoxic niche, HIF-1 maintains HSC quiescence, 6 whereas HIF-2 maintains their self-renewing capacity. 7 HIF-1 also affects embryonic hematopoiesis as demonstrated by defective myeloid and erythroid progenitor formation in HIF-1␣ Ϫ/Ϫ as well as ARNT Ϫ/Ϫ embryos. 8 In adult hematopoiesis, HIF-1 is essential for B-cell progenitor proliferation and mature B-cell subclass differentiation. 9 However, its involvement in mononuclear phagocyte development is unknown. Previous findings indicated defective development of human plasmacytoid dendritic cells (pDCs) under hypoxia in vitro. 10 Therefore, we asked whether HIF-1 regulates DC lineage differentiation in mice. Methods AnimalsHIF-1␣ fl/fl or HIF-2␣ fl/fl mice 11,12 were bred with LysM-Cre transgenic mice 13 in the C57BL/6 background. Age-matched C57BL/6 wild-type (WT) mice were controls. ID2 Ϫ/Ϫ mice and their respective WT control were in the NMRI background. 14 The guidelines of the Hessian animal care and use committee were followed. DC generation from BMFor DC generation in vitro, 2 ϫ 10 6 total BM cells/mL in RPMI 1640 with 10% FCS and 200 ng/mL recombinant murine fms-related tyrosine kinase 3-ligand (Flt3-L, PeproTech) were cultured in 6-well Ultra-Low attachment plates (Corning) for up to 9 days 15 at various O 2 levels as indicated, using a InVivo 2 400 hypoxia workstation (Ruskinn Technologies). Alternatively, cells were cultured with 100M dimethyloxallyl glycine (DMOG, from Biomol). Sorted monocyte/DC progenitors/common DC progenitors (MDPs/ CDPs; 10 4 cells/well) were cultured with 200 ng/mL Flt3-L in 24-well Ultra-Low attachment plates. Flow cytometry and cell sortingBM cells, spleen, or whole blood cells were stained with fluorochromeconjugated antibodies and analyzed on a LSRII/Fortessa flow cytometer (BD Biosciences). MDP/CDP were sorted from lineage Ϫ cell-enriched BM (lineage cell depletion kit and AutoMACS cell separator from Miltenyi Biotec) using a FACSAria III cell sorter (BD Biosciences). For details (antibodies, surface markers, intracellular transcription factor staining procedures), see supplemental Methods (available on the Blood Web site; see the Supplemental Materials link at the top of the online article). pDCs were i...

IRF-5-Mediated Inflammation Limits CD8+ T Cell Expansion by Inducing HIF-1α and Impairing Dendritic Cell Functions during Leishmania Infection

“…Nevertheless, this transcription factor impairs the upregulation of CCR7, a chemokine involved in homing of mature DCs to secondary lymphoid organs [ 53 , 85 ]. Moreover, HIF-1α acts as a negative regulator of plasmacytoid dendritic cell development [ 86 ]. Our results reveal a detrimental role for HIF-1α in DC function during experimental VL.…”

The multifaceted biology of plasmacytoid dendritic cells

“…Hypoxia-inducible factors influence hematopoiesis and maintain HSC function. Recently, it was shown that low oxygen content upregulates ID2 and suppresses FLT3L-induced pDC development, in a manner dependent on HIF1α 227 . GFI1 controls the development and functions of pDCs and cDCs in a STAT3-dependent manner 228 and represses Rag transcription in pDCs 229 .…”

HIF1A and NFAT5 coordinate Na+-boosted antibacterial defense via enhanced autophagy and autolysosomal targeting

“…BMDM from C57BL/6 WT mice and various knock out and transgene mice were generated in Teflon® bags, as described earlier [80]. In addition to C57BL/6 WT mice (Charles River Breeding Laboratories, Sulzfeld, Germany), we used GFP-MAP1LC3 mice [81], cybb -/mice [82] and their littermate controls, atg7 cKO (Atg7 F/F : Lyz2-Cre) mice [83,84] and their littermate controls, and hif1a cKO (Hif1a F/F : Lyz2-Cre) mice [85] and their respective littermate controls as described earlier [86,87]. For infection experiments, we used E. coli HB101 or, for immunofluorescence studies, E. coli harboring pWRG167 [88] and pWRG29 for constitutive expression of superfolder GFP (sfGFP) or super cyan fluorescent protein 3A (sCFP3A), respectively.…”