5羟色胺4受体激动剂对糖尿病小鼠结肠黏膜巨噬细胞极化的影响

目的研究5-羟色胺4受体（5-hydroxytryptamine 4 receptor, 5-HT4R）激动剂对糖尿病结肠黏膜巨噬细胞极化的影响。方法使用链脲佐菌素（streptozotocin, STZ）建立小鼠1型糖尿病模型，使用5-HT4R激动剂RS67333进行治疗干预，实验动物随机分为3组，即非糖尿病组、糖尿病对照组和糖尿病给药组。通过免疫荧光方法验证巨噬细胞标记物F4/80和iba1在CX3CR1-GFP+细胞中的表达；通过激光共聚焦显微镜观察CX3CR1-GFP+细胞形态和数目分析结肠黏膜巨噬细胞的活化数量；通过RNAscope技术检测精氨酸酶1（arginase 1, Arg1）和一氧化氮合酶（inducible nitric oxide synthase, iNOS）的表达和分布变化。结果免疫荧光结果显示F4/80和iba1免疫阳性的细胞与CX3CR1-GFP+的细胞基本共定位。与非糖尿病小鼠相比，STZ诱导的1型糖尿病模型小鼠结肠黏膜层活化的巨噬细胞数目增多（P<0.001），且M1型巨噬细胞产物iNOS在黏膜层腔侧表达增加（P<0.001），而M2型巨噬细胞产物Arg1在黏膜固有层表达下降（P<0.001）；与糖尿病对照组相比，给予5-HT4R激动剂可以抑制活化的巨噬细胞数目（P<0.001），结肠黏膜iNOS的表达上调（P<0.01）和Arg1的表达下调（P<0.05）。 结论5-HT4R激动剂可以抑制糖尿病小鼠结肠黏膜巨噬细胞向M1型极化。

Effects of 5-HT4R agonist on macrophage polarization in colonic mucosa of diabetic mice

ObjectiveTo investigate the effects of 5-HT4R agonist on the macrophage polarization in the colonic mucosa of diabetic mice. MethodsType 1 diabetes was induced by intraperitoneal injection of streptozotocin(STZ) in CX3CR1-GFP+ C57BL/6 mice. 5-HT4R agonist RS67333 was used for therapeutic intervention. The mice were randomly divided into control group, diabetic group(group D) and RS67333 intervention group(D-RS group). The expression of macrophage biomarkers F4/80 and ionized calcium binding adaptor molecule 1(iba1) in CX3CR1-GFP+ cells was detected by immuno-fluorescence. The activation of macrophages was assessed by observing the change in morphology and number of CX3CR1-GFP+ cells. The expression and distribution of arginase(Arg1) and inducible nitric oxide synthase(iNOS) in colon was detected by RNAscope. ResultsImmunofluorescent assay revealed that F4/80- or iba1- immunoreactive cells and CX3CR1-GFP+ cells were co-localized in cells. Compared with normal control group, the activated macrophages in diabetic group were significantly increased(P<0.001). The expression of iNOS, production of M1 macrophages was increased in the mucosa close to lumen(P<0.001), while the expression of Arg1, production of M2 macrophages was decreased in the laminae propria in diabetic mice(P<0.001). 5-HT4R agonist inhibited diabetes-induced increase of activated macrophages(P<0.001), upregulation of iNOS(P<0.01) and downregulation of Arg1(P<0.05) in the mucosa of colon. Conclusion5-HT4R agonist can inhibit the shift in macrophage polarization from M2 to M1 in diabetic mice.