Immunological characterization of the early human fracture hematoma

Abstract

The initial inflammatory phase of fracture healing is of great importance for the clinical outcome. We aimed to develop a detailed time-dependent analysis of the initial fracture hematoma. We analyzed the composition of immune cell subpopulations by flow cytometry and the concentration of cytokines and chemokines by bioplex in 42 samples from human fractures of long bones <72 h post-trauma. The early human fracture hematoma is characterized by maturation of granulocytes and migration of monocytes/macrophages and hematopoietic stem cells. Both T helper cells and cytotoxic T cells proliferate within the fracture hematoma and/or migrate to the fracture site. Humoral immunity characteristics comprise high concentration of pro-inflammatory cytokines such as IL-6, IL-8, IFNγ and TNFα, but also elevated concentration of anti-inflammatory cytokines, e.g., IL-1 receptor antagonist and IL-10. Furthermore, we found that cells of the fracture hematoma represent a source for key chemokines. Even under the bioenergetically restricted conditions that exist in the initial fracture hematoma, immune cells are not only present, but also survive, mature, function and migrate. They secrete a cytokine/chemokine cocktail that contributes to the onset of regeneration. We hypothesize that this specific microenvironment of the initial fracture hematoma is among the crucial factors that determine fracture healing.

A Pronounced Inflammatory Activity Characterizes the Early Fracture Healing Phase in Immunologically Restricted Patients

“…The amounts of CD14+ monocytes/macrophages (as percentage of all leucocytes found in the fracture hematoma) and CD34+ hematopoietic stem and progenitor cells (as percentage of all mononuclear cells) were increased significantly within SBM IR (surrounding bone marrow of immunologically restricted patients) when compared to SBM of controls and numerically within the FH IR when compared to FH of controls (Figure 1A,B). It should be noted that the data for controls presented in Figure 1 and elsewhere in this manuscript were taken from our recent publication [12]. The amounts of CD3+CD56+ natural killer T (NKT) cells (as percentage of all lymphocytes) and activated CD45RA−CD25+CD3+CD4+ T helper cells (as percentage of all T helper cells) were increased significantly within SBM IR (SBM of immunologically restricted patients) when compared to SBM of controls and significantly within the FH IR when compared to FH of controls (Figure 1C,D).…”

“…We could confirm the inflammatory nature of this initial phase both on RNA and protein levels showing high concentrations of pro-inflammatory cytokines such as IL-1β, Interferon (IFNγ or TNFα and chemokines like Monocyte chemotactic protein 1 (MCP-1), Interferon gamma-induced protein 10 (IP-10) and Regulated on activation, normal T cell expressed and secreted (RANTES) [12,13]. Furthermore, we demonstrated that immune cells invade and become activated [12]. Very little is known about these initial processes of fracture healing in patients with restricted immune functions.…”

“…This drove us to perform a detailed analysis of immune cell populations and pro-inflammatory and regulatory cytokines, chemokines, and factors regulating angiogenesis within fracture hematomas and the surrounding tissue in patients suffering from disorders concerning impact on immune functions. These results were then compared to those of previously analyzed data on fracture hematomas obtained from control patients (patients lacking co-morbidities associated with delayed fracture healing) [12]. Here, to our knowledge, we give the first detailed immunological characterization of the fracture hematoma from immunologically restricted patients.…”

“…This fracture hematoma represents the site of the initial inflammatory phase [11]. We characterized previously the initial inflammatory phase of control patients from the immunological point of view [12]. We could confirm the inflammatory nature of this initial phase both on RNA and protein levels showing high concentrations of pro-inflammatory cytokines such as IL-1β, Interferon (IFNγ or TNFα and chemokines like Monocyte chemotactic protein 1 (MCP-1), Interferon gamma-induced protein 10 (IP-10) and Regulated on activation, normal T cell expressed and secreted (RANTES) [12,13].…”

Glucocorticoids in rheumatoid arthritis: the picture is shaping up

“…These discussions usually come up, especially among GC ‘supporters’ and ‘opponents’,1 when trying to elaborate recommendations on how to use these drugs best in the treatment of rheumatic diseases such as rheumatoid arthritis (RA),2 polymyalgia rheumatica,3 giant cell arteritis,4 systemic lupus erythematodes,5 6 myositis7 and even systemic sclerosis 7. For example, very divergent opinions were learnt during work on the European League Against Rheumatism (EULAR) recommendations (2013 update) for the management of RA 2.…”