Flo8基因缺失白念珠菌甘露聚糖通过IL-10调控溃疡性结肠炎的机制研究

Mechanism of mannan from flo8-deficient Candida albicans in regulating ulcerative colitis via interleukin-10

摘要:

目的探讨flo8基因缺失白念珠菌来源的甘露聚糖(flo8-deficient Candida albicans, flo8Δ-M)在小鼠溃疡性结肠炎(ulcerative colitis, UC)模型中的调控作用及机制。 方法通过热碱法提取白念珠菌甘露聚糖。构建野生型(wild type, WT)、Clec4n基因敲除小鼠UC模型，在相应时间点对小鼠进行甘露聚糖灌胃操作，检测小鼠体质量、小鼠结肠长度，评估小鼠疾病活动指数，ELISA检测结肠IL-6、IL-10水平。flo8Δ-M刺激小鼠骨髓来源树突状细胞(bone marrow-derived dendritic cells, BMDCs)，ELISA检测上清液的IL-10水平，Western印迹法检测BMDCs中脾脏酪氨酸激酶(spleen tyrosine kinase, Syk)的磷酸化水平，qPCR检测IL-10表达水平。 结果体内实验表明，flo8Δ-M灌胃处理组UC小鼠体质量、结肠长度、IL-10水平显著增加，疾病活动指数、IL-6水平显著降低(P＜0.05)。体外实验显示，flo8Δ-M通过Dectin-2-Syk信号通路显著诱导BMDCs分泌IL-10(P＜0.05)，最后在Clec4n基因敲除UC小鼠模型中验证了Dectin-2的关键调控作用。 结论flo8基因缺失白念珠菌来源的甘露聚糖通过Dectin-2-Syk信号通路介导抗炎细胞因子IL-10产生，在小鼠溃疡性结肠炎模型中起保护作用。

Abstract:

ObjectiveTo investigate the role and mechanism of mannan derived from flo8-deficient Candida albicans(flo8Δ-M) in regulating ulcerative colitis(UC) in a mouse model. MethodsMannan from C. albicans was extracted using a hot alkali method. The UC models were established in both wild-type(WT) and Clec4n knockout mice(Clec4n-/-). The mice were administered mannan via gavage at specified time points, and the weight changes, colon length and disease activity index(DAI) were monitored. The colonic levels of interleukin(IL)-6 and IL-10 were measured by enzyme-linked immunosorbent assay(ELISA). The bone marrow-derived dendritic cells(BMDCs) in the mice were stimulated with flo8Δ-M, and the IL-10 levels in the supernatant were quantified by ELISA. The phosphorylation of spleen tyrosine kinase(Syk) in BMDCs was assessed by Western blotting, and the IL-10 mRNA expression was determined by quantitative real-time polymerase chain reaction(qPCR). ResultsIn vivo experiments demonstrated that flo8Δ-M gavage significantly increased the body weight, colon length, and IL-10 levels, while reduced the DAI and IL-6 levels in UC mice(all P<0.05). In vitro experiments indicated that the flo8Δ-M significantly induced IL-10 secretion in BMDCs via the Dectin-2-Syk signaling pathway(P<0.05). The critical regulatory role of Dectin-2 was further validated in Clec4n knockout UC mice. ConclusionMannan derived from flo8-deficient C. albicans exerts protective effects in UC mouse model by promoting anti-inflammatory cytokine IL-10 production through the Dectin-2-Syk signaling pathway.