维生素D_(3)对黄颡鱼头肾极化分型的巨噬细胞影响

VITAMIN D_(3) ON POLARIZATION PHENOTYPE OF HEAD KIDNEY MACROPHAGES IN PELTEOBAGRUS FULVIDRACO

研究以黄颡鱼(Pelteobagrus fulvidraco)头肾巨噬细胞为研究对象,通过细菌脂多糖(LPS)和环磷酸腺苷(cAMP)分别诱导M1型和M2型极化,200 pmol/L维生素D_(3)孵育后对其形态学特征、生物学功能及极化相关基因的表达进行分析鉴定来确定维生素D_(3)在巨噬细胞极化中的调节作用。结果表明,维生素D_(3)能降低诱导后M1型和M2型巨噬细胞的死亡率,并增强巨噬细胞的吞噬活性。在M1型巨噬细胞中维生素D_(3)能够抑制活性氧(ROS)和炎症介质一氧化氮(NO)的产生,降低超氧阴离子自由基的活力,白介素1β(IL-1β)和肿瘤坏死因子-α(TNF-α)的表达水平显著降低(P<0.05);在M2型细胞中能够增加精氨酸酶的活性,显著增加白介素10(IL-10)和转化生长因子(TGF-β)的表达水平(P<0.05),最终抑制巨噬细胞向M1表型极化,促进巨噬细胞向M2表型极化,发挥抗炎作用;黄颡鱼头肾巨噬细胞中Nos-2和Arg-2分别是M1和M2巨噬细胞的生物标记基因。研究结果为进一步研究鱼类巨噬细胞极化及维生素D_(3)对极化调节的影响提供了重要依据。

As a key class of immune cells,macrophages are not only the first line of defense against pathogen invasion,but also play different functions through polarization.Macrophages can secrete pro-inflammatory factors and cause excessive inflammatory response,resulting in a variety of inflammatory diseases when polarizing into M1 type.Macrophages can secrete anti-inflammatory factors and play an anti-inflammatory role when polarizing into M2 type.Macrophages are characterized by phenotypic heterogeneity and functional diversity,and it is important to target macrophagetype polarization through nutritional regulation.Vitamin D_(3)has been proved to play important role in fish anti-inflammatory response.In this study,the effects of 200 mol/L vitamin D_(3)on polarization phenotype of head kidney macrophages of Pelteobagrus fulvidraco were studied.The primary macrophage cells of Pelteobagrus fulvidraco head kidney were isolated and cultured.After being challenged by LPS and cAMP to induce macrophage polarization,the cell morphological changes were observed by inverted light microscopy,and the functional changes were studied by measuring survival rate,phagocytosis,reactive oxygen species and nitric oxide production,superoxide anion radical and arginase activity,as well as the related gene expression charactering in different macrophages polarization states.The results showed that vitamin D_(3)reduced the mortality of M1 and M2 macrophages and enhanced the phagocytic activity of macrophages.In M1 macrophages,vitamin D_(3)inhibited the production of reactive oxygen species(ROS)and inflammatory mediator nitric oxide(NO),reduced the activity of superoxide anion free radical,and decreased the expression of interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)(P<0.05).The activity of arginase as well as the expression of interleukin-10(IL-10)and transforming growth factor(TGF-β)in M2 cells was up-regulated by vitamin D_(3)(P<0.05).In conclusion,vitamin D_(3)inhibited the polarization of macrophages to M1 phenotype,promoted the polarization of macrophages to M2 phenotype,and played an anti-inflammatory role.In the current study,Nos-2 and arg-2 were found to be biomarker genes of M1 and M2 macrophages,respectively.These results preliminarily reveal the mechanism of vitamin D_(3)on the polarization of head kidney macrophages of Pelteobagrus fulvidraco,and favor the regulation of macrophage phenotypes in the treatment of inflammation,which provides useful information for further study on the polarization of fish macrophages and the effect of vitamin D_(3)on polarization regulation.