Abstract
Inflammation plays a central role in the onset and progression of numerous diseases, and macrophages are key regulators of this process through their polarization toward pro- or anti-inflammatory phenotypes. Natural bioactive compounds have emerged as promising candidates to modulate inflammatory responses. Amongst these, genistein, a soybean-derived isoflavone, was selected due to its reported anti-inflammatory and antioxidant profile across different cell types. In this study, we evaluated the effects of genistein in a lipopolysaccharide (LPS)-induced macrophage model to investigate its cellular and molecular mechanisms of action. Genistein treatment at subcytotoxic concentration preserved cell viability, did not induce apoptosis or cell cycle arrest, and reduced oxidative stress. In LPS-stimulated macrophages, treatment with 5 µg/mL genistein significantly decreased nitric oxide production and downregulated pro-inflammatory markers while upregulating an anti-inflammatory mediator, thus reverting macrophage polarization to an anti-inflammatory phenotype. Protein analyses confirmed reduced expression of iNOS and cytokines such as IL-6 and TNF-α. Furthermore, genistein restored cell morphology and inhibited NF-κB p65 nuclear translocation, indicating its capacity to counteract LPS-driven pro-inflammatory signalling. These findings support the role of genistein in modulating macrophage polarization and inflammatory responses, highlighting its therapeutic potential for the management of acute and chronic inflammation-related disorders. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1038/s41598-026-42357-7.