Sida rhombifolia L. Reduces Acute Inflammation and NF-κB/AP-1-Driven SEAP Reporter Activity in TPA- and LPS-Stimulated RAW-Blue™ Cells

Sida rhombifolia L. is traditionally used for the treatment of conditions associated with inflammation. In this study, hexane (SrHex), ethyl acetate (SrAcOEt), and methanolic (SrMeOH) extracts were evaluated in a TPA (12-O-tetradecanoylphorbol-13-acetate)-induced auricular edema assay in male CF-1 mice. SrHex, the most active extract, was fractionated into H-F1–H-F5. Selected fractions were further evaluated by histological analysis and in RAW-Blue™ macrophage-derived cells stimulated with TPA or LPS (lipopolysaccharide). Cell viability and NF-κB/AP-1-driven SEAP reporter activity were determined, and fractions H-F3 and H-F5 were chemically characterized by GC-MS. SrHex showed the greatest anti-inflammatory activity among the extracts, whereas H-F3 was the most active fraction in the TPA-induced edema model. Histological and morphometric analyses showed that SrHex and fractions H-F1–H-F3 attenuated TPA-induced increases in epidermal/subcutaneous tissue thickness and collagen layer thickness. In TPA-stimulated RAW-Blue™ cells, SrHex and H-F1–H-F3 reduced NF-κB/AP-1-driven SEAP reporter activity without a substantial loss of cell viability. In LPS-stimulated cells, SrHex and H-F2–H-F5 produced concentration-dependent reductions in SEAP reporter activity while largely preserving cell viability. H-F5 showed the highest apparent potency in the reporter assay, with an IC50 of 20.63 µg/mL and an Emax of 98.04%. H-F1 also reduced SEAP reporter activity, although the effects at higher concentrations should be interpreted cautiously because of the concomitant reduction in the cell viability signal. A preliminary analysis by GC-MS enabled the identification of dihydroactinidiolide, 2-methylene-5α-cholestan-3β-ol, and hexanedioic acid, mono(2-ethylhexyl) ester in H-F3, whereas in the H-F5 fraction, cyclopropanetetradecanoic acid, 2-octyl-methyl ester, and 3′,8,8′-trimethoxy-3-piperidyl-2,2′-binaphthalene-1,1′,4,4′-tetrone were identified. Overall, S. rhombifolia showed anti-inflammatory activity in the TPA model and modulated NF-κB/AP-1-associated reporter activity in the stimulated RAW-Blue™ cells.