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Abstract
Background Hepatic ischemia-reperfusion injury (HIRI) is a major complication in clinical situations such as liver transplant, hypovolemic shock, and severe trauma, critically affecting patient prognosis. Therapeutic interventions targeting HIRI are limited. We hypothesized that inhibition of Bromodomain containing protein-4 (BRD4) could attenuate IRI-mediated inflammation and hepatocellular death. The present study aimed to investigate the effect of BRD4 inhibition using nitroxoline (NTX) against HIRI.
Methods Identification of potential drug and disease targets, followed by protein-protein interactions was done using network pharmacology (IDDF2024-ABS-0206 Figure 1. Network pharmacology analysis). Highly interactive targets were validated through molecular docking. To further validate, HIRI was induced in male SD rats (n=6) via occlusion of the portal triad for 1 hour, followed by 24 hours of reperfusion. Rats in the treatment group (n=6) were dosed with NTX (25, 50, and 75mg/kg, orally) for 7 days before induction of HIRI. Blood samples were taken, animals were sacrificed, and livers were harvested for further analysis (IDDF2024-ABS-0206 Figure 2A. Representative liver photographs from different study groups). Liver function tests (LFTs), oxidative stress parameters, molecular markers (BRD4, NFκB, HIF1A, NLRP3, ATF4, CHOP, GRP78, XBP1), inflammatory markers (IL-1β, TNF-α, IL-6), and histological analysis were performed.
Results Gene ontology and KEGG pathway enrichment analysis showed that potential targets enriched cellular homeostasis, I-kappa B/NFκB complex, NFκB binding, HIF-1 signaling pathway. Six highly interactive proteins identified using Cytoscape showed a good binding affinity for NTX. NTX treatment (50 and 75mg/kg) markedly improved LFTs, MDA, nitrite, reduced GSH levels, and SOD, catalase, and myeloperoxidase activity compared to the disease control group. A significant decrease in the levels of BRD4, ER stress markers, and inflammatory markers was observed in NTX-treated groups compared to disease control. Furthermore, histopathological analysis revealed a notable reduction in sinusoidal congestion, nuclear pyknosis, vacuolization, and necrosis in the NTX-treated group compared to the disease group (IDDF2024-ABS-0206 Figure 2B. Representative images of H&E stained liver sections).
Network pharmacology analysis
Representative liver photographs from different study groups
Representative images of H&E stained liver sections
Mechanism of nitroxoline in amelioration of hepatic ischemia-reperfusion injury
Conclusions These data through network pharmacology, biochemical, molecular, and histological analyses demonstrated that administration of NTX was effective in reducing HIRI through alleviation of BRD4-mediated inflammation and ER stress (IDDF2024-ABS-0206 Figure 3. Mechanism of nitroxoline in amelioration of hepatic ischemia-reperfusion injury).