Hemophilia macrophages exhibit specific defects relating to wound healing

J. Pilch, D. Lessig, M. Bernard, E. C. Schwarz, H. Eichler, L. M. Knowles (Homburg, Germany)

Bleeding disorders - Basic science
Date: 17.02.2017,
Time: 08:00 - 09:15

Objective: Bleeding events in hemophilia can lead to synovial inflammation, cartilage damage and ultimately, joint destruction. It has also been shown that wound healing is delayed in hemophilia patients. Macrophages are specialized in promoting wound healing and tissue regeneration as they infiltrate in blood clot and phagocytose wound debris. We hypothesize that these macrophage functions are relevant to joint health. The objective is to define macrophage function in hemophilia with respect to polarization, phagocytosis and clot invasion.

Methods: Monocytes isolated from hemophilia patients and healthy individuals were treated with M-CSF or GM-CSF to induce macrophage differentiation. Macrophage differentiation was probed by phenotypic analysis as well as immunocytochemistry for TNFα, CD163 and CD206. Expression of Tie2 was assessed by flow cytometry. Phagocytosis was assessed using fluorescent labeled latex beads as well as red blood cells. To determine cell invasion, monocytes were embedded in a 3-dimensional matrix of clotted plasma, differentiated with M-CSF or GM-CSF and probed for podosome formation.

Results: M-CSF, which supports macrophage traits associated with wound healing and tissue repair, induced a spread and elongated phenotype in donor monocytes. Hemophilia monocytes, on the other hand, did not undergo this typical shape change and failed to express differentiation markers such TNFα or CD163 in response to M-CSF. Inflammatory differentiation in response to GM-CSF, however, was at least partially intact. Functionally, deregulation of hemophilia monocytes was reflected in deficits of M-CSF-dependent actions such as clot invasion and phagocytosis. These deficits could be explained by impaired filopodia formation, which may result from reduced expression of the receptor tyrosine kinase Tie2 on hemophilia monocytes.

Conclusion: Hemophilia macrophages are unable to promote adhesive interactions in response to M-CSF. As a result, M-CSF mediated regenerative macrophage functions such as phagocytosis and clot invasion are diminished while inflammatory macrophage functions in response to GM-CSF appear to be preserved. Together, our results suggest that hemophilia macrophages exhibit specific impairments of functions involved in wound healing, which may have important implications for resolving joint inflammation after bleeding.