Effects of Glycine tomentella Hayata extract on cytokines and clearance of apoptotic cells

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Objective : I-Tiao-Gung, Glycine tomentella Hayata(GTH), has been used as a traditional herbal medicine to treat rheumatic diseases in Kinmen and Taiwan, indicative of its anti-inflammatory effect. However, the possible mechanism of the pharmacological effect has not been completely delineated. The aim of this research is to investigate the effects of GTH on proinflammatory cytokines production in mouse macrophage cell line RAW264.7 and human monocyte cell line U937. Methods : Mouse macrophage cell line RAW264.7 and human monocyte cell line U937 were cultured with lipopolysaccharide(LPS) in the presence or absence of ethanol extract of I-Tiao-Gung. The gene expression of proinflammatory cytokines: IL-1β, IL-6,TNF-α, iNOS and transglutaminase 2 (TG2) were assayed by reverse transcriptase-polymerase chain reaction. The production of IL-1β, IL-6, TNF-α and metalloproteinase-9 (MMP-9) were assayed by ELISA and gelatin zymography.The phagocytosis activiti of RAW264.7 was analyzed by ingesting apoptotic cells and latex-beads and then detected at 4h by FACS. Results : Our results showed that the ethanol extract of GTH 165μg/ml inhibited the gene expression as well as the production of proinflammatory cytokines: IL-1β(55%), IL-6(56%) in LPS-activated macrophages, but only IL-6 (79%)in LPS-activated human monocyte. Furthermore, the gene expression of TNF-α and iNOS had no significant difference compared with the LPS-activated cell. In addition, the ethanol extract of GTH also inhibited the production of proinflammatory mediators, transglutaminase 2 (TG2)(47%) and metalloproteinase-9 (MMP-9) (57%) in LPS-activated macrophages. Morever, TG2 mRNA expression was enhanced 58% than control in the treatment wth GTH alone.In the experiment of phagocytosis, it was found that RAW264.7 had higher ability to ingest apoptotic cells (156% enhanced) and no significant difference in latex-beads. Conclusion : In our study, we found the ethanol extract of GTH suppressed the LPS-induced production of IL-1β, IL-6, transglutaminase 2 (TG2) and metalloproteinase-9 (MMP-9)in mouse macrophage cell line RAW264.7. The inhibition of IL-6 also exerted in human monocyte cell line U937. The clearance of apoptotic cells was also enhanced. This would indicate the anti-inflammatory potencies of this traditional herbal remedy.