Perturbations of Marrow Stromal Cell function during Acute Inflammation

Abstract

Abstract Mesenchymal stromal cells (MSCs) reside in a complex network of blood cells and can differentiate into mature chondrocytes, osteoblasts, and adipocytes. This trilineage differentiation process is tightly regulated by intrinsic and extrinsic signals, leading to transcriptional programming of balanced lineage output. Inflammatory insults to this stromal homeostatic balance occur through the upregulation of activating cytokines. Pro-inflammatory cytokines’ impact on the hematopoietic lineage is well-defined, but similar impacts to stroma was lacking. To define these changes, we studied two well-characterized forms of inflammation: polyinosinic-polycytidylic acid (Poly(I:C)), that activates IFNα, and Lipopolysaccharide (LPS), that through TLR4 promotes IFNγ. Prior studies of inflammatory conditions show bone loss; therefore, we hypothesized that there will be an increase in the MSC pool to compensate for loss of bone maturation. Using flow cytometry and lineage tracing models for stroma, we phenotypically characterized the non-hematopoietic cells’ changes following perturbation. The lineage-specific Cre models used were stroma (Prx1), adipocytes (AdipoQ), and osteoblasts (OCN). Our data shows an increase in MSCs with decreases in osteoblasts and adipocytes with stimulation. Furthermore, we used reverse-phase protein array (RPPA) and cytokine arrays to map the signaling mechanisms and activated pathways. We found an increase in specifically MAPK pathway with an accompanying increase in cytokines linked to this pathways’ activation. Understanding changes in the bone marrow microenvironment during inflammation will allow for therapeutic intervention in mediating hematopoietic diseases such as leukemia.