Abstract
Monocytes and their derived macrophages are found at the site of remodeling tissue, such as fracture hematoma, that is exposed to mechanical forces and have been previously implicated in the reparative response. However, the mechanoresponsive of monocytes and macrophages to skeletal tissue-associated mechanical forces and their subsequent contribution to skeletal repair remains unclear. The aim of this study was to investigate the potential of skeletal tissue-associated loading conditions to modulate human monocyte activation and phenotype. Primary human monocytes or the human monocyte reporter cell line, THP1-Blue, were encapsulated in agarose and exposed to a combination of shear and compressive loading for 1 h a day for 3 consecutive days. Exposure of monocytes to mechanical loading conditions increased their pro-inflammatory gene and protein expression. Exposure of undifferentiated monocytes to mechanical loading conditions significantly upregulated gene expression levels of interleukin(IL)-6 and IL-8 compared to free swelling controls. Additionally, multiaxial loading of unstimulated monocytes resulted in increased protein secretion of TNF-α (17.1 ± 8.9 vs. 8 ± 7.4 pg/ml) and MIP-1α (636.8 ± 471.1 vs. 124.1 ± 40.1 pg/ml), as well as IL-13 (42.1 ± 19.8 vs. 21.7 ± 13.6) compared monocytes cultured under free-swelling conditions. This modulatory effect was observed irrespective of previous activation with the M1/pro-inflammatory differentiation stimuli lipopolysaccharide and interferon-γ or the M2/anti-inflammatory differentiation factor interleukin-4. Furthermore, mechanical shear and compression were found to differentially regulate nitric oxide synthase 2 (NOS2) and IL-12B gene expression as well as inflammatory protein production by THP1-Blue monocytes. The findings of this study indicate that human monocytes are responsive to mechanical stimuli, with a modulatory effect of shear and compressive loading observed toward pro-inflammatory mediator production. This may play a role in healing pathways that are mechanically regulated. An in depth understanding of the impact of skeletal tissue-associated mechanical loading on monocyte behavior may identify novel targets to maximize inflammation-mediated repair mechanisms.
Highlights
The repair process following traumatic injury to the musculoskeletal system is known to be influenced by the mechanical environment
Monocytes stimulated with LPS and IFN-γ had significantly higher gene expression levels of the pro-inflammatory genes IL8 and C motif chemokine ligand 2 (CCL2) under free-swelling conditions compared to IL-4 stimulated monocytes (10.3- and 28.3-fold increases, respectively), confirming their pro-inflammatory phenotype (Figure 1A)
Monocytes are found at the site of skeletal tissue injuries resulting from either traumatic bone fractures, or microdrilling of the subchondral bone to facilitate microfracturemediated cartilage repair [23]
Summary
The repair process following traumatic injury to the musculoskeletal system is known to be influenced by the mechanical environment. A wound healing response is initiated during the process of bone fracture repair, as well as marrow stimulation techniques applied in cartilage repair strategies where the subchondral bone is penetrated. This involves inflammatory cell exudation or infiltration to the site of injury, followed by coagulation activation and fibrin clot formation, which is known to regulate monocyte chemotaxis and proliferation [3, 4]. Pro-inflammatory macrophages are associated with high production of pro-inflammatory cytokines and increased microbicidal activity, whereas anti-inflammatory macrophages are associated with wound healing and immunoregulatory functions [3, 6]
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