神经介素U在体外对猪树突细胞活性及功能的影响

[目的]本文旨在探讨不同浓度神经介素U（NMU）对外周血单核源树突细胞（DC）刺激T淋巴细胞增殖和相关细胞因子分泌的影响。[方法]采集小梅山猪外周血，用集落刺激因子（GM-CSF）和IL-4联合诱导外周血单核细胞形成未成熟树突细胞，再加入脂多糖（LPS）刺激获得成熟的树突细胞，同时观察DC形态。加入不同浓度（0.01、0.1、1、10、100、1 000 nmol？L-1）NMU并分别培养2、4、8和12 h后，收集细胞上清液，用ELISA法测定IL-4、IL-5和IL-13的浓度。加入不同浓度（0.1、1、10、100、1 000 nmol？L-1）NMU培养24 h后，收集细胞，用CCK-8试剂盒和细胞凋亡检测试剂盒分别检测DC刺激混合淋巴细胞时细胞的增殖和DC凋亡率。[结果]NMU（0.1~100 nmol？L-1）能够抑制未成熟树突细胞（iDC）的细胞凋亡（P<0.01），且NMU与NMU+LPS作用效果相一致，10 nmol？L-1NMU组与10 nmol？L-1 NMU+LPS组都能降低iDC凋亡，凋亡率分别为2.383%和2.360%，与LPS组相比，NMU+LPS组抑制iDC细胞凋亡效果极显著（P<0.01），说明NMU与LPS协同发挥抑制iDC细胞凋亡作用；与对照组相比，0.1~100 nmol？L-1NMU能极显著促进mDC分泌IL-5（P<0.01）和抑制IL-4分泌（P<0.01）。NMU对IL-13的影响呈现多样性，低剂量（0.01~0.1 nmol？L-1）NMU在2、4 h抑制mDC细胞分泌IL-13（P<0.05），中剂量（1~10 nmol？L-1）NMU在2 h先抑制mDC分泌IL-13（P<0.05），4 h后又促进其分泌（P<0.05），高剂量（100~1 000 nmol？L-1）NMU则促进mDC分泌IL-13（P<0.05）。经NMU诱导后，iDC和mDC均能促进T淋巴细胞增殖（P<0.01）。[结论]在一定浓度范围内，NMU能够抑制猪树突细胞的凋亡，并提高其细胞活性和促进树突细胞刺激淋巴细胞增殖的功能，并能影响树突细胞细胞因子分泌，提示神经介素U参与了对猪免疫功能的调节。
[Objectives]The paper aims to explore the efficiency of neuromedin U (NMU)with different concentrations treatments on the cytokine secretion and T lymphocyte proliferation by pig dendritic cells (DC). [Methods]In this study,immature dendritic cells (iDC)were originated from pig peripheral blood monocytes cultured with GM-CSF and IL-4,and then triggered DC maturation with LPS. Varied doses of NMU (0.01,0.1,1,10,100,1 000 nmol？L-1)were added into iDC and mature dendritic cells (mDC)at different time points (2,4,8,12 h),then the culture supernatant of mDC was collected to detect IL-4,IL-5 and IL-13 by ELISA kit. Finally,the apoptosis of DC and the capacity of DC to evaluate T cell proliferation were determined by apoptosis detection kit and CCK-8 kit. [Results]NMU (0.1-100 nmol？L-1)can inhibit the apoptosis of iDC (2.383%) (P<0.01),which was consistent with the effect of NMU+LPS (2.360%). Furthermore,compared with LPS,NMU+LPS can significantly inhibit the apoptosis of iDC,which suggested that NMU had a synergistic effect with LPS in inhibiting apoptosis of iDC. Compared with control group,NMU (0.1-100 nmol？L-1)can significantly increase the expression of IL-5 (P<0.01)and reduce the expression of IL-4 (P<0.01). However,the secretion of IL-13 displayed a diversity trend. It turned out that low NMU (0.01-0.1 nmol？L-1)can inhibit the expression of IL-13 in mDC during 2 h to 4 h (P<0.05),and medium dose (1-10 nmol？L-1)NMU can inhibit the IL-13 release within 2 hours while it can increase the IL-13 secretion after 4 hours (P<0.05). High dose (100-1 000 nmol？L-1)NMU increased the expression of IL-13