肾脏固有树突状细胞在肾缺血-再灌注期间的变化

Changes of renal resident dendritic cells during kidney ischemia-reperfusion injury

目的探讨在肾脏缺血-再灌注损伤(IRI)期间肾脏固有树突状细胞(rDC)的变化。方法采用C57BL/6J小鼠建立双侧肾脏热缺血模型,再灌注24、48 h后取肾组织制备单细胞悬液,流式细胞仪分析CD45^+细胞和CD11c^+rDC的比例变化;采用绿色荧光蛋白和白喉毒素受体标记(CD11c^+GDTR)的小鼠肾组织制作单细胞悬液,流式细胞仪分析CD11c^+rDC的比例及表型;采用CD11c^+GDTR小鼠建立双侧肾脏热缺血模型,再灌注24 h后取肾组织制备单细胞悬液,MACS磁珠富集CD45^+细胞,经流式细胞仪分析rDC表面共刺激分子表达情况。结果再灌注24 h后C57BL/6J小鼠肾脏内CD45^+细胞比例明显增加,48 h后其比例进一步升高,再灌注24 h后CD11c^+rDC数量同样持续升高,但其占CD45^+细胞的比例出现明显下调,48 h后恢复并较Sham组轻度升高;CD11c^+GDTR小鼠正常肾脏CD45^+细胞比例低于1%,其中约40%为CD11c^+的肾脏rDC,主要呈CD11b^(int)F4/80^-MHCⅡ^+;再灌注24 h后CD11c^+F4/80^-亚群rDC表面共刺激分子CD40、CD80、CD86均显著升高。结论热IRI后rDC比例、数量及其表面共刺激分子表达均增加,提示热IRI后肾脏rDC浸润增多且表型成熟。

Objective To investigate the changes of renal resident dendritic cells （rDC）during kidney ischemia-reperfusion injury （IRI）. Methods C57BL/6J mice models with bilateral renal warm ischemia were established. The kidney tissue was prepared for single cell suspension at 24 h and 48 h after reperfusion. The changes in the percentage of CD45 ＋cells and CD1 1 c ＋rDCs were evaluated by flow cytometry. The renal tissues of mice labeled with green fluorescent protein and diphtheria toxin receptor （CD1 1 c ＋GDTR ） were prepared for single cell suspension. The percentage and phenotype of CD1 1 c ＋rDCs were analyzed by flow cytometry. CD1 1 c ＋GDTR mice models with bilateral renal warm ischemia were established. The renal tissue was prepared for single cell suspension at 24 h after reperfusion. CD45 ＋ cells was gathered by magnetic-activated cell separation （MACS ）. The expression levels of co-stimulatory molecules on the rDC surface were analyzed by flow cytometry. Results At 24 h after reperfusion,the percentage of CD45 ＋cells in the kidney of C57BL/6J mice was significantly elevated,and further increased at 48 h after reperfusion. At 24 h after reperfusion,the quantity of CD1 1 c ＋rDCs was equally increased,whereas the percentage of CD1 1 c ＋rDCs in CD45 ＋cells was dramatically declined and restored at 48 h after reperfusion,slightly higher compared with that in the sham group. In healthy CD1 1 c ＋GDTR mice,the percentage of CD45 ＋cells in the kidney was lower than 1%,consisting of approximately 40%of CD1 1 c ＋rDCs,which mainly presented as CD11bintF4/80 -MHCⅡ＋. At 24 h after reperfusion,the percentage of CD11c ＋F4/80 -subset rDC surface co-stimulatory molecules was significantly enhanced,such as CD40,CD80 and CD86. Conclusions Following warm IRI,the percentage and quantity of rDCs,and the expression level of rDC surface co-stimulatory molecule are significantly increased,prompting that renal rDC infiltration is increased and phenotype becomes matured.