Investigating the Role of Escin in RAW 264.7 Macrophage Inflammatory Models

Corresponding author: sun guang chen, sungc@glmc.edu.cn

DOI: 10.12201/bmr.202504.00003

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  Abstract: Objective This study aimed toTo investigate the regulatory effects of escin on macrophage polarization factors, inflammatory cytokines, and inflammation-related pathways, and to explore its therapeutic mechanisms in rheumatoid arthritis (RA). Methods RAW 264.7 macrophages were treated with escin at concentrations of 0 , 0.5, 1.0, 2.0, 5.0, 10.0, and 20.0 μM. Cell viability was assessed using the thiazolyl blue tetrazolium bromide (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, MTT) assay. An inflammatory model was established by stimulating RAW 264.7 macrophages with lipopolysaccharide (LPS, 500 ng/mL). The experimental groups were divided as follows: Control group (untreated), Model group (500 ng/mL LPS), Escin treatment groups(500 ng/mL LPS + 0.5 μM escin, 500 ng/mL LPS + 1.0 μM escin, 500 ng/mL LPS + 2.0 μM escin, 500 ng/mL LPS + 4.0 μM escin). Western blot analysis?was performed to analyze: The expression of?mitogen-activated protein kinase (MAPK) pathway proteins, including?extracellular signal-regulated kinase (ERK)?and?c-Jun N-terminal kinase (JNK), in LPS-stimulated macrophages. The expression of?macrophage polarization markers:?macrophage mannose receptor (CD206)?and?hemoglobin scavenger receptor (CD163). The levels of the?pro-inflammatory cytokine tumor necrosis factor-α (TNF-α). Results MTT assays confirmed that?0.5-5 μM escin?exhibited no cytotoxicity toward RAW 264.7 cells. Western blot results demonstrated that: 4 μM escin?significantly?downregulated TNF-α expression.?0.5–4 μM escin?markedly?upregulated CD206 and CD163 expression, indicating enhanced M2 polarization.?0.5–4 μM escin?significantly?suppressed ERK expression, while?4 μM escin?notably?inhibited JNK expression, suggesting modulation of the MAPK pathway. Conclusion Escin exerts therapeutic effects on RA by?inhibiting LPS-induced M1 polarization?and?promoting M2 polarization?in RAW 264.7 macrophages. These effects are mediated through the?suppression of the MAPK signaling pathway?(specifically ERK and JNK), which correlates with reduced TNF-α levels and enhanced anti-inflammatory activity.

  Key words: Escin; anti-inflammation; mechanism of action; RAW264.7 macrophages

  Submit time: 2 April 2025

  Copyright: The copyright holder for this preprint is the author/funder, who has granted biomedRxiv a license to display the preprint in perpetuity.
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