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

Background: 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.
Methods: 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 hours 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.
Results: 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.
Conclusions: Anti-inflammatory miRNA and mRNA expression were upregulated at the injury site in the C5/CD14 inhibition group, while osteogenesis was 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.