LEPR-expressing cells are key regulators for a sustained hematopoietic response during chronic inflammation

Abstract

The blood system is established and maintained by hematopoietic stem and progenitor cells (HSPCs) that are located in the bone marrow (BM) where they are surrounded by different types of supportive cells. Accumulating evidence suggests that important extrinsic regulatory mechanisms have evolved to maintain stemness of HSCs and guide HSPC differentiation into more mature blood cell lineages. We set out to study how the BM microenvironment supports the hematopoietic system in adaptation to inflammation. Therefore wild-type (WT) mice were stimulated with LPS or polyIC to mimic gram-negative bacterial or viral infection, respectively. Based on expression of CD45 and Ter119 to exclude hematopoietic cells in conjuction with Sca1, CD31 and CD140b, we are able to flow-cytometrically define several non-hematopoietic cell types in mouse BM: so-called PαS cells which were shown to have mesenchymal stem cell characteristics, a stromal cell population enriched for CXCL12-abundant reticular cells (CARs), and endothelial cells (ECs). These different cell types were isolated and their gene expression profiles were determined and compared to steady-state WT mice. We found that Il6 is significantly and specifically up-regulated during LPS stimulation by the cell population enriched for CARs. This could be validated using qPCR and by injecting BM chimeric mice (Wt>Wt, Wt>Il6-/-, Il6-/->Wt, Il6-/->Il6-/-) with PBS and LPS. We could show that IL6 protein is almost exclusively produced by non-hematopoietic cells. We then generated cell type-specific Il6-deficient mice by crossing LepR-Cre mice with IL6fl/fl mice, and could confirm that IL6 is mainly produced by non-hematopoietic cells that are enriched for CARs. To functionally study the regulatory role of IL6 on hematopoiesis during inflammation, WT and Il6-/- mice were injected with PBS or LPS over 3 weeks every other day. We analyzed phenotypically defined hematopoietic stem and progenitor cells as well as more mature myeloid cell types, and observed a significant difference in absolute cell numbers of these cell types in LPS-treated WT mice compared to LPS-treated Il6-/- mice. We could observe the same significant difference in absolute cell numbers when we repeated the experiment with LepR-Cre; IL6fl/fl mice and their LepR-Cre negative littermates. Our preliminary data suggests that CAR cell produced IL6 is essential for a sustained myelopoietic response during chronic LPS treatment. IL6 may act as a proliferative stimulus for the early HSPC compartment during infection to provide a reservoir of highly proliferative myeloid progenitors in order to counterbalance inflammation-driven cell losses.