Phytochemical Characterization and Anti-inflammatory Effects of Azadirachta indica Extract on LPS-Induced Intestinal Epithelial Cells
Keywords:
Azadirachta indica, Intestinal inflammation, LPS, Oxidative stress, Epithelial barrier, PhytochemicalsAbstract
Background: Inflammatory activation of intestinal epithelial cells is a key mechanism underlying digestive disorders such as inflammatory bowel disease and endotoxin-mediated mucosal injury. Azadirachta indica (neem) (A. indica) is rich in bioactive compounds with reported anti-inflammatory and antioxidant properties, yet its molecular effects on LPS-induced intestinal epithelial inflammation remain insufficiently characterized.
Methods: Methanolic extracts of A. indica leaves were subjected to phytochemical quantification and LC-MS/HPLC profiling to identify major bioactive compounds. Human intestinal epithelial cells (Caco-2) were pretreated with neem extract (25-100 µg/mL) and challenged with LPS (1 µg/mL). Cellular viability (MTT assay), pro-inflammatory cytokine production (IL-6, TNF-α, IL-1β, ELISA), oxidative stress markers (ROS, MDA, SOD), gene expression (NF-κB, COX-2, iNOS, tight junction proteins), and apoptosis (flow cytometry) were assessed.
Results: Phytochemical analysis confirmed high levels of phenolics, flavonoids, tannins, saponins, and limonoids (azadirachtin, nimbin), consistent with potential bioactivity. The extract exhibited no cytotoxicity up to 100 µg/mL and dose-dependently suppressed LPS-induced IL-6, TNF-α, and IL-1β production. Treatment also reduced ROS accumulation and lipid peroxidation while restoring SOD activity. Gene expression analysis demonstrated downregulation of NF-κB, COX-2, and iNOS, alongside recovery of tight junction proteins (Occludin, ZO-1). Apoptosis induced by LPS was significantly attenuated.
Conclusions: A. indica extract exerts potent anti-inflammatory, antioxidant, and barrier-protective effects in LPS-stimulated intestinal epithelial cells. The data suggest that neem phytochemicals act via multi-target modulation of oxidative stress and NF-κB signaling, highlighting their therapeutic potential for inflammatory digestive disorders. Further in vivo studies are warranted to confirm efficacy and optimize dosage for translational applications.
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