Protective effects of Hirudin against compartment syndrome in rabbits through the activation of Nrf2/HO-1.

Abstract

Compartment syndrome generates an oxidative condition causing the death of skeletal muscle cells. Hirudin has antioxidant and anti-inflammatory properties. However, its correlation with the pathway of Nrf2/HO-1 for the protection of the skeletal muscle is unknown. We aimed to evaluate the protective efficacy of double-doses of hirudin in compartment syndrome and its association with Nrf2/HO-1 expression. Compartment syndrome was induced in rabbits and double-doses of hirudin (0-8 ATU/kg) were locally administered to select an optimal Hirudin concentration that protects skeletal muscle from damage. The tissue structural changes, W/D ratio, lipid peroxidation level by MDA assay and inflammatory factors were determined in the skeletal muscle. To determine the musculoprotective efficacy of H8 at 72 h timepoint after compartment syndrome, and its association with the Nrf2/HO-1 pathway, the following assays were performed by TUNEL assay and immunofluorescence: necrosis (high-mobility group box-1; HMGB1), Nrf2, and HO-1. In addition, the HO-1 mRNA was evaluated by qPCR. Hirudin in a dose of 8 ATU/kg (H8) presented the lowest levels of histological damage, fibrosis, W/D ratio and oxidative stress in the studied groups. Moreover, treatment with H8 markedly downregulated the level of inflammatory factors including TNF-a, IL-1β and IL-6. H8 showed a protective effect at 72 h timepoint after compartment syndrome, as revealed by a decrease in the levels of all damage markers. Nuclear translocation Nrf2 and HO-1 staining in cytoplasm were increased, and the levels of HO-1 mRNA were also increased. In conclusion, double-doses of H8 alleviate the death of muscle cells induced by oxidative stress 72 h after compartment syndrome in rabbits. This protective effect is associated with the nuclear translocation of Nrf2 and an elevated expression of HO-1.