Ameliorative Effects of Royal Jelly Against Ibuprofen-Induced Hepatic Toxicity: Molecular Regulation via Oxidative Stress, Cytokine Signaling, Cell Death Mechanisms, and Endoplasmic Reticulum Stress

Abstract Ibuprofen (IBU), a widely used nonsteroidal anti-inflammatory drug (NSAID), is associated with hepatotoxic effects, particularly at high doses or with prolonged use. These effects are mainly mediated through oxidative stress, inflammation, and apoptosis. This study investigates the hepatoprotective potential of royal jelly (RJ), a bee-derived product rich in bioactive compounds, against IBU-induced liver injury in rats. Male Sprague Dawley rats were divided into five groups: i) control, ii) RJ (300 mg/kg), iii) IBU (400 mg/kg), iv) IBU + RJ (150 mg/kg), and v) IBU + RJ (300 mg/kg). All treatments were administered orally for five days. Biochemical, molecular, and histopathological analyses were performed to asssess liver enzyme activities (ALT; alanine aminotransferase, AST; aspartate aminotransferase, ALP; alkaline phosphatase), as well as oxidative stress markers including malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione (GSH). The expression of inflammation-related genes (NF-κB; nuclear factor kappa B, TNF-α; tumor necrosis factor alpha, IL-1β; interleukin-1 Beta, TLR4; toll-like receptor 4) was evaluated. Expressions of autophagy markers ( Beclin-1, LC3A; light chain 3A, LC3B; light chain 3B, Atg5; A-autophagy related 5, Atg7; autophagy related 7 ) , apoptotic markers (p53; tumor protein P53, Bax; BCL2 associated X, Bcl-2; B-cell lymphoma 2, Caspase-3, Caspase-9), and Endoplasmic Reticulum (ER) stress indicators (ATF6; activating transcription factor 6, PERK; protein kinase RNA-like endoplasmic reticulum kinase, IRE1; inositol-requiring enzyme 1, CHOP; C/EBP homologous protein, GRP78; glucose-regulated protein 78) were also measured. Additionally, the PI3K/Akt/mTOR ( phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin ) pathway and MMP2/MMP9 (matrix metalloproteinases) were evaluated. IBU significantly increased liver enzymes, lipid peroxidation, and pro-inflammatory and pro-apoptotic signaling while decreasing antioxidant defense and PI3K/Akt/mTOR activity. RJ co-administration markedly ameliorated these alterations, restoring antioxidant levels, reducing inflammation and apoptosis, and improving liver histology. These findings suggest that RJ exhibits significant hepatoprotective properties against NSAID-induced liver injury.