Combined C5 and CD14 inhibition balances immunological miRNA responses after multiple trauma in pigs.

Abstract

Combined inhibition of complement-component C5 and Toll-Like Receptor CD14 is a promising therapeutic strategy to attenuate post-traumatic inflammatory-complications. The effect of this inhibition on miRNA expression at the fracture site and in systemic circulation was assessed in a porcine multiple-trauma model. Expression of the mRNA targets of deregulated miRNAs was determined at the fracture site and in lung and liver. Two experimental groups, intramedullary nailing (control; n = 8) and intramedullary nailing with combined C5/CD14 inhibition (n = 4), were compared using an established porcine multiple-trauma model. Fracture hematoma, fractured bone, unfractured control bone, lung, and liver samples were collected. Extracellular-vesicles (EVs) were isolated from plasma at 1.5, 2.5, 24, and 72 h post-trauma. MiRNA qPCR array analyses were performed on pooled fxH, fX, CB, and EV samples. In silico mRNA target prediction and mRNA qPCR analyses were conducted on all samples. The fracture site miRNA signature in the C5/CD14 inhibition group was anti-inflammatory compared to intramedullary nailing alone, based on miRNA and mRNA qPCR analyses, with downregulation of anti-osteogenic miRNAs. Furthermore, C5/CD14 inhibition reduced pro-inflammatory and pro-fibrotic EV-carried miRNA expression in systemic circulation and their downstream mRNA targets in lung and liver. Anti-inflammatory miRNA and mRNA expression were upregulated at the injury site in the C5/CD14 inhibition group, while pro-osteogenic pathways were preserved by downregulating anti-osteogenic miRNAs. Systemically, C5/CD14 inhibition reduced EV-carried inflammatory miRNA expression, consistent with mRNA target analyses in lung and liver. Thus, C5/CD14 inhibition may be a novel therapeutic to modulate post-traumatic immune responses and decrease systemic effects of surgical trauma.