Anti-inflammatory Effects of Soluble Eggshell Membrane Powder on LPS-Stimulated Caco-2 Cells: Modulation of Proinflammatory Cytokine Gene Expression

Maintaining intestinal epithelial barrier integrity is essential for mucosal homeostasis and immune defense. Bioactive compounds derived from natural sources have attracted interest for their potential to modulate inflammatory responses in the gut. Eggshell membrane, a proteinaceous layer rich in collagen, glycosaminoglycans (including hyaluronic acid and chondroitin sulfate), and bioactive peptides, has demonstrated anti-inflammatory properties in preclinical and clinical settings, although its effects on intestinal inflammatory markers remain largely unexplored. The aim of this study was to evaluate the anti-inflammatory effects of soluble eggshell membrane powder on lipopolysaccharide (LPS)-induced inflammatory gene expression in human intestinal epithelial Caco-2 cells. Cells were pre-stimulated with LPS (50 µg/mL) for 1 h, then co-treated with the eggshell membrane product at concentrations of 0.01% and 0.1% in the continued presence of LPS for an additional 3 h. Gene expression of TNFα, IL1α, IL1β, IL6, and IL10 was quantified by qRT-PCR (n = 4) using the 2−ΔΔCt method and analyzed by one-way ANOVA with Dunnetts post hoc test. Treatment at 0.01% significantly reduced TNFα expression by 55.1 ± 16.7% (p 0.05), IL1α by 86.5 ± 12.0% (p 0.0001), and IL1β by 64.3 ± 15.2% (p 0.01), relative to LPS-only controls, corresponding to 99.0%, 126.9%, and 129.2% attenuation of the LPS-induced increases, respectively. At 0.1%, IL1α was also significantly reduced (−56.9 ± 11.1%, p 0.01), while an inverse dose–response pattern was observed for TNFα and IL1β. IL6 expression was not reduced by treatment and showed a significant increase at 0.1%. IL10 was not significantly affected by either LPS or treatment. The MTT assay confirmed that the product did not affect cell viability at any concentration tested (0.001–0.1%). These results suggest that soluble eggshell membrane powder exerts selective anti-inflammatory activity targeting early NF-κB/TLR4-dependent proinflammatory mediators, and may have potential applications in intestinal health management.