乌司他丁对脂多糖介导的小鼠肾脏固有树突状细胞成熟的抑制作用

Ulinastatin inhibits LPS-induced maturation of renal resident dendritic cells

目的研究乌司他丁对脂多糖介导的小鼠肾脏固有树突状细胞（rDC）成熟的影响。方法取C57BL／6J小鼠肾脏，制备单细胞悬液，使用免疫磁珠法分选CDllc＋的rDC，流式细胞仪鉴定rDC纯度。脂多糖刺激24h，促使rDC成熟，给予不同浓度（50、100和200U／L）乌司他丁处理，酶联免疫吸附试验检测上清液中白细胞介素12（IL-12）的分泌水平。100U／L的乌司他丁处理后，流式细胞仪检测rDC表面主要组织相容性复合物（MHC）lI类分子及CD80和CD86分子，以观察乌司他丁对rDC表型及功能的影响。检测趋化因子受体7（CCR7）的表达，计数受巨噬细胞炎症蛋白3G（MIP-38）趋化进入细胞培养小室下层的CDllC＋细胞的数目，分析乌司他丁对rDC趋化能力的影响。结果脂多糖刺激促使rDC成熟，高表达MHCⅡ、CD80、CD86分子，并分泌大量IL-12，而乌司他丁能够抑制脂多糖介导的rDC表型和功能成熟。经乌司他丁处理，CCR7分子的表达由69．2％下降至37％，并且受趋化的CDllc＋细胞数目下降为脂多糖处理者的45．3％。结论乌司他丁能够抑制脂多糖介导的小鼠rDC的成熟和趋化能力上调。

Objective To investigate the inhibitory effect of ulinastatin （UTI）, an ulinary trypsin inhibitor, on the maturation of renal dendritic cells （rDCs） induced by LPS stimulation. Method The kidneys from C57BL/6J （H-2Kb） mice were harvested to make single cell suspension. rDCs were then selected by Magnetic Cell Sorting （MACS） and assessed for content and purity by flow cytometry. The matured rDCs were generated by stimulating rDCs with LPS for 24 h, then treated with different concentrations of UTI （0-200 U/L）. LPS-stimulated cell supernatants were collected for IL-12 measurement by using ELISA. We used 100U/L of UTI to treat matured rDCs, and assessed the maturation phenotype as well as chemokine receptor expression of I）Cs by using flow cytometry. For chemotaxis assay, LPS-stimulated rDCs treated with UTI （100U/L） were loaded into upper chemotaxis chambers. Cells migrating into the lower chambers were counted by using flow cytometry. Result LPS-stimulated rDCs secreted high levels of IL-12 and expressed high levels of CD80, CD86 and MHC class II, and UTI treatment significantly inhibited the production of ILq2 and expression of CD80, CD86 and MHC class II. UTI treatment further suppressed the expression of CCR7 on rDCs, as well as their migratory responses to MIP-3β upon LPS stimulation. Conclusion UTI treatment inhibits LPS-mediated maturation of rI）Cs and interferes with the up-regulation of rDCs chemotaxis.