gms | German Medical Science

Objectives: In recent years evidence has accumulated that the complement system, a crucial part of innate immunity, is involved in bone turnover and the pathogenesis of inflammatory bone disorders. Our group showed that the anaphylatoxin receptor C5aR1 is upregulated during osteoblastogenesis, induces an immune response in osteoblasts and stimulates osteoclast activity. After bone fracture, C5aR1 modulates not only the inflammatory response, but also the formation and remodeling of new bone. Furthermore, we demonstrated an interaction of C5aR1 and toll-like receptor (TLR) signaling in osteoblasts in the regulation of C-X-C motive chemokine 10 (CXCL10), an inflammatory and osteoclastogenic factor, which is secreted by osteoblasts in response to bacterial challenges. Therefore, we hypothesize that the combined actions of C5aR1 and TLRs in osteoblasts modulate the immune response and promote osteoclastogenesis in infected bone fractures. To study this, we used a mouse model of lipopolysaccharide (LPS)-induced impaired bone healing. Healing was investigated in control mice and mice lacking C5aR1 specifically in osteoblast.

Methods: Twelve-week-old male C5aR1^fl/fl and C5aR1^Runx2-Cre mice, which lack the C5aR1 specifically in osteoblasts and hypertrophic chondrocytes underwent a femur osteotomy. Afterwards, the animals were randomly divided in two groups. One group received i.p. injections of 25 µg LPS per day for 7 days and the other group PBS as control. Inflammation and bone repair were assessed on days 1, 7, 14, and 21 post-surgery by serum analysis, FACS, µCT, and histomorphometric analyses as well as biomechanical testing of the fracture callus.

Results and conclusion: In C5aR1^fl/fl mice, LPS treatment induced systemic inflammation 1 d post-fracture, confirmed by increased serum levels of IL-1β, IL-6, TNF- α and M-CSF. Callus formation was compromised in LPS treated controls, as indicated by significantly decreased relative bone and cartilage fractions (p=0.002 and 0.005, respectively) on day 7 post-fracture. On day 14, the percentage of bone was significantly reduced but the cartilage content increased in LPS treated control mice (p=0.04 and 0.007, respectively) indicating delayed endochondral bone formation. This resulted in impaired fracture healing 21 d after fracture, confirmed by a significantly reduced bending stiffness (p=0.04), bridging score (p=0.02), and bone content in the callus (p=0.004) Notably, all LPS-induced negative effects were completely abolished in C5aR1^Runx2-Cre mice.

Our data showed that LPS disrupted bone healing by impaired endochondral ossification, resulting in delayed fracture healing. Notably, C5aR1^Runx2-Cre mice revealed an undisturbed bone healing after LPS treatment, indicating that the combined actions of C5aR1 and TLRs in osteoblasts and hypertrophic chondrocytes may contribute to the pathogenesis of infection-induced disturbed fracture healing.