枸杞多糖对LPS诱导的人视网膜色素上皮细胞炎性反应的影响及机制

Effect and relative mechanism of Lycium Barbarum Polysaccharide on LPS-induced inflammatory response within human retinal pigment epithelial cells

摘要:

目的：探索枸杞多糖(LBP)对细菌脂多糖(LPS)诱导的人视网膜色素上皮(ARPE-19)细胞的炎性反应的影响及其可能介导的信号通路。

方法：通过LPS刺激体外培养的ARPE-19细胞构建炎性反应模型。将细胞随机分为5组，空白组使用完全培养基培养，LPS组给予含10μg/mL LPS的完全培养基刺激24h，低、中、高浓度LBP组分别给予0.1、0.5、1mg/mL LBP完全培养基孵育24h后给予含10μg/mL LPS的完全培养基刺激24h。应用CCK-8观察细胞存活率、实时荧光定量PCR检测相关炎性因子的mRNA表达量及Western-blot检测NF-κB/MAPK信号通路相关磷酸化蛋白表达量的变化。

结果：与正常细胞相比，LPS刺激后，ARPE-19的存活率下降。同时，随着外源性LBP浓度增加，ARPE-19的细胞存活率升高，且细胞内炎性因子IL-1β、IL-6和MCP-1的表达量降低，并伴随NF-κB/MAPK通路的相关磷酸化蛋白(p-p65、p-IκBα、p-JNK、p-ERK及p-p38)的表达下降。

结论：枸杞多糖通过抑制胞内炎性因子及NF-κB/MAPK通路相关因子的磷酸化，从而阻止LPS诱导人视网膜色素上皮细胞产生的炎性反应。

Abstract:

AIM: To explore the effect of lycium barbarum polysaccharides(LBP)on inflammatory response of human retinal pigment epithelial cells(ARPE-19)induced by lipopolysaccharide(LPS)and its possible signal pathway.

METHODS: ARPE-19 cells were stimulated by LPS in vitro to construct the inflammatory injury cell model. Primarily, the cells were divided into five groups randomly. The blank group was cultured in complete medium, and the LPS group was stimulated with complete medium containing 10μg/mL LPS for 24h. The low, medium and high concentration LBP groups were incubated with complete medium importing 0.1, 0.5 and 1mg/mL LBP for 24h separately, and then stimulated with complete medium containing 10μg/mL LPS for 24h. We used the CCK-8 method to observe the cell survival rate, real-time fluorescent quantitative PCR to detect the mRNA expression of inflammatory factors and Western blot to test the changes of phosphorylated protein within the signaling pathway of NF-κB/MAPK.

RESULTS: Compared with normal cells, the survival rate of ARPE-19 cells was decreased after the LPS stimulation. With the increase of exogenous LBP concentration, the survival rate of ARPE-19 cell was gradually increased, while the inflammatory factors expression of cytokines IL-1β, IL-6 and MCP-1 were reduced accompany with the phosphorylated proteins(p-p65, P-IκBα, p-JNK, p-ERK and p-p38)of NF-κB/MAPK signaling pathway were decreased.

CONCLUSION: LBP prevents LPS-induced inflammatory response of ARPE-19 by inhibiting the intracellular inflammatory factors and the phosphorylation of the related protein within NF-κB/MAPK signaling pathway.