CXCR4 signaling controls dendritic cell location and activation at steady state and in inflammation

Abstract

Dendritic cells (DCs) encompass several cell subsets that collaborate to initiate and regulate immune responses. Proper DC localization determines their function and requires the tightly controlled action of chemokine receptors. All DC subsets express CXCR4, but the genuine contribution of this receptor to their biology has been overlooked. We addressed this question using natural CXCR4 mutants resistant to CXCL12-induced desensitization and harboring a gain of function that cause the warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) Syndrome (WS), a rare immunodeficiency associated with high susceptibility to the pathogenesis of human papillomavirus (HPV). We report a reduction in the number of circulating plasmacytoid DCs (pDCs) in WHIM patients, whereas that of conventional DCs is preserved. This pattern was reproduced in an original mouse model of WS, enabling us to show that the circulating pDC defect can be corrected upon CXCR4 blockade and that pDC differentiation and function are preserved, despite CXCR4 dysfunction. We further identified proper CXCR4 signaling as a critical checkpoint for Langerhans-cell and DC migration from the skin to lymph nodes, with corollary alterations of their activation state and tissue inflammation in a model of HPV-induced dysplasia. Beyond providing new hypotheses to explain the susceptibility of WHIM patients to HPV pathogenesis, this study shows that proper CXCR4 signaling establishes a migration threshold that controls DC egress from CXCL12-containing environments and highlights the critical and subset-specific contribution of CXCR4 signal termination to DC biology.

Multi-Tissue Characterization of GILZ Expression in Dendritic Cell Subsets at Steady State and in Inflammatory Contexts

“…Histology assessment confirmed marked skin inflammation, with increased epidermis and dermis thickness (Figure 5a,b). DC subsets were identified (Figure 5c) and their analysis showed that dysplasia-associated inflammation was driving MHC II level diminution in LCs and skin DC subsets (Supplemental Figure S4a), as expected [33,40]. As for GILZ expression, we first confirmed that LCs and cDC2 expressed the highest GILZ levels among skin DCs in the FVB/N background (Figure 5d and Supplemental Figure S4b, DC subsets from WT mice), as in the C57BL/6J background (Figure 1).…”

“…For skin cell isolation, tissue pieces were dissected from mouse ear or from depilated (Veet Cream for sensitive skin, used post-mortem) mouse belly and flanks, and processed as previously described [33,36]. Briefly, tissue pieces were incubated for 90 min in dispase (2.5 U/mL, Gibco BRL) at 37 • C. The epidermis and dermis were then separated and cut into small pieces and further incubated for 1 h in a solution of collagenase D (1 mg/mL, Roche) and DNase I (12.5 U/mL, Roche) at 37 • C. Tissue dissociation was then completed using 18-G syringes and samples were further incubated for 15 min at 37 • C before filtration through 70-µm cell strainers.…”

“…To characterize GILZ expression in skin DCs at steady state, skin biopsies were taken from the flanks of C57BL/6J mice and cells were isolated by combining enzymatic digestion with mechanical tissue dissociation [33,36]. We used a gating strategy excluding CD64 + macrophages and allowing for the identification of five skin DC subsets, including LCs, CD103 − cDC1, CD103 + cDC1, cDC2, and CD207 − CD11b − double negative (DN) dermal DCs [4], based on the differential expression of the CD11b, CD103 and CD207 markers (Table 2 and Figure 1a).…”

“…We next sought to evaluate the impact of chronic inflammation on GILZ expression by skin DCs. We took advantage of a transgenic mouse model of HPV-induced dysplasia that is associated with skin inflammation [33,39]. Skin biopsies were harvested from HPV mice and their littermate controls (WT).…”

“…Acute inflammation induced by epicutaneous fluorescein isothiocyanate (FITC) application [31] induced GILZ downregulation in skin LCs and cDC2, as well as in migDC1 recovered from skin-draining (SD)LNs. Such GILZ downregulation in skin DCs was however not detected in a context of chronic skin inflammation associated with human papillomavirus (HPV)induced dysplasia [32,33]. Finally, GILZ was expressed by tumor infiltrating (TI)DCs in a model of HPV-induced tumors, and cancer was associated with opposite modulation of GILZ in cDC1 and cDC2 subsets recovered from tumor-draining (TD)LNs.…”