Differential effects of flunixin meglumine and meloxicam on TNF- α production in LPS-stimulated equine neutrophils in vitro

Abstract

Systemic inflammatory response syndrome (SIRS) induced by endotoxemia is usually secondary to colitis and is a major cause of high morbidity and mortality in horses. Non-steroidal anti-inflammatory drugs (NSAIDs), such as flunixin meglumine (FM) and meloxicam (MX), are used to improve clinical outcomes in SIRS/endotoxemia. These NSAIDs suppress tumor necrosis factor-alpha (TNF-α) levels; however, the underlying mechanisms remain unclear. The aim of this study was to investigate the inhibitory effects of FM and MX on TNF-α in lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMCs) and neutrophils in vitro. Blood samples were collected from three healthy thoroughbred horses, and PBMCs and neutrophils were isolated using density gradient centrifugation. Cells were cultured with LPS (0.3 ng/mL) and FM or MX (5, 10, or 50 μM). TNF-α gene expression was analyzed using quantitative real-time PCR, and protein levels were measured using ELISA. No significant inhibitory effects of FM or MX on TNF-α gene or protein expression were observed in LPS-stimulated PBMCs. However, FM significantly inhibited the increase in TNF-α protein levels in LPS-stimulated neutrophils in a concentration-dependent manner (p < 0.05). MX showed a similar tendency in LPS-stimulated neutrophils, but the differences were not significant. The regulation of neutrophil-derived TNF-α by FM administration could be a promising therapeutic strategy for equine SIRS/endotoxemia, providing mechanistic insight for optimizing anti-inflammatory therapy.