The Regulatory Effect Of Total Saponins From Physalis Alkekengi L. Var. Francheti (Mast.) Makino On Macrophage Inflammatory Response

Physalis alkekengi L. var. francheti (Mast.) Makino (PA)(Solanaceae), known as "Jin Deng Long" in Chinese, is a perennial herb that is widely distributed in China. The fruit of this herb is edible. This article focused on the bioactivity of total saponins from Physalis alkekengi L. var. francheti (Mast.) Makino (TSP) in anti-inflammation by the methods of ELISA, RT-PCR and Western blot. The main results are as follows:1. The cytotoxic effect of the TSP on RAW264.7cells was tested with the MTT assay. The TSP had no effect on cell viability at the concentration of0-100μg/mL. The IC50of TSP was223.42μg/mL.2. The production of NO, IL-1β, IL-6, TNF-a, MCP-1, and PGE2secreted by LPS-stimulated macrophages were determined by ELISA assay. The results showed that the TSP with the concentration of10μg/mL could significantly (*P<0.05,**P<0.01) reduce the release of NO, IL-6, IL-1β, and MCP-1in the LPS-stimulated macrophages, and reduced20.38%,5.67%,29.80%,31.71%, respectively; TSP with the concentration of50μg/mL could significantly (*P<0.05,**P<0.01) reduce the production of NO, IL-6, IL-1β, MCP-1PGE2and TNF-a in the LPS-stimulated macrophages, and reduced31.40%,40.60%,70.47%,64.52%,8.11%and8.06%, respectively; TSP with the concentration of100μg/mL could significantly (*P<0.05) reduce the secrection of NO, IL-6, IL-1β, MCP-1PGE2and TNF-a in the LPS-stimulated macrophages, and reduced63.71%,72.68%,90.04%,82.57%,18.11%and17.31%, respectively. From the above, the TSP (10,50,100μg/mL) could significantly (*P<0.05,**P<0.01) reduce the release of NO, IL-6, IL-1β, and MCP-1in the LPS-stimulated macrophages in a dose-dependent manner. Compared with the LPS induced groups, groups treated with TSP at50and100μg/mL reduced the production of PEG2and TNF-a, significantly (*P<0.05,**P<0.01).3. The qRT-PCR assay were performed to test the expressions of IL-1β, IL-6, TNF-a, COX-2, iNOS mRNA, which are inflammation related genes. The results showed that the TSP with the concentration of10μ/mL could significantly (*P<0.05) down regulated the expressions of iNOS and IL-1β mRNA in the LPS-stimulated macrophages, and reduced56.54%and25.54%, respectively; TSP with the concentration of50μg/mL could significantly (*P<0.05,**P<0.01) down regulated the expressions of iNOS, TNF-a, IL-6and IL-1β mRNA in the LPS-stimulated macrophages, and reduced61.49%,12.44%,38.94%and 72.49%, respectively; TSP with the concentration of100μg/mL could significantly (*P<0.05,**P<0.01) down regulated the expressions of iNOS, COX-2, TNF-a, IL-6and IL-1β mRNA in the LPS-stimulated macrophages, and reduced79.12%,17.39%,12.48%,72.38%and89.40%, respectively. From the above, the TSP (10,50,100μg/mL) could significantly (*P<0.05,**P<0.01) down regulated the expressions of iNOS, IL-1β mRNA in a concentration-dependent manner. With the compare of the groups induced by LPS, groups treated with TSP with higher concentrations, which could also suppressed the expressions of PEG-2(100μg/mL) mRNA and TNF-a (50,100μg/mL) mRNA, significantly (*P<0.05,**P<0.01).4. The Western blot assays were carried out to examine the expression of IL-1β protein. We found that treatment with10,50and100μg/mL TSP remarkably reduced the IL-1β protein expression in a concentration-dependent manner, and decreased4.68%,89.25%and93.84%, respectively.5. In conclusion, TSP significantly inhibited the release of NO, IL-6, and IL-1β in a dose-dependent manner, and also down-regulated the expressions of related aforementioned genes and protein. In addition, TSP treatment suppressed the elevated COX-2and TNF-α gene expression in lipopolysaccharide-induced RAW264.7. Moreover, the secretions of MCP-1in stimulated macrophages were also restrained in a concentration-dependent way. Our data showed that TSP exhibited anti-inflammatory effect by suppressing the transcription of some inflammatory cytokine genes, such as iNOS, IL-1β, and IL-6, suggesting that the anti-inflammatory properties of TSP may be helpful to mitigate inflammation, and could be regarded as a potential source of natural anti-inflammatory agents.