摘要
目的研究降钙素基因相关肽(calcitonin gene related peptide,CGRP)对人外周血中Th17(IL-17producing T cell)和Treg(regulatory T cell)细胞分化的调节作用。方法采用免疫磁珠法(magnetic activated cell sorting,MACS)分离人外周血中CD4^+CD45RA^+T细胞,即初始CD4^+T细胞,在一定条件下分别体外诱导其向Th17细胞和Treg细胞分化,之后用流式细胞术测定Th17和Treg细胞所占比例;用实时定量RT-PCR方法检测Th17和Treg细胞分化的特异性转录因子维甲酸相关核孤儿受体γt(orphan nuclear receptorγt,RORγt)和转录因子FoxP3(foxhead box protein 3)mRNA的表达水平;用酶联免疫吸附测定法(enzyme-linked immunosorbent assay,ELISA)检测Th17细胞上清中IL-17A的含量,进而从多个方面探讨CGRP对Th17和Treg细胞分化的影响。结果 CGRP可剂量依赖性增加Th17细胞所占的比例和RORγt mRNA的表达水平(P<0.05);另外,CGRP也促进了IL-17A的分泌(P<0.05)。同时,CGRP能剂量依赖性抑制Treg细胞所占的比例和FoxP3mRNA的表达(P<0.05)。结论 CGRP可促进人外周血中初始CD4^+T细胞向Th17细胞分化,而抑制其向Treg细胞分化。
Objective To dissect the role of caleitonin gene related peptide(CGRP) on IL-17 producing T cell(Th17) and regulatory T cell(Treg) differentiation. Methods Naive CD4+T cells were isolated from human Peripheral blood by magnetic activated cell sorting (MACS) selection, and cultured under Th17-polarizing and Treg-polarizing condition, The proportion of Th17 cells and Treg cells were detected by flow cytometry, the relative expression levels of retionid-related orphan nuclear receptor Tt(ROR'/t) mRNA and foxhead box protein 3(FoxP3) mRNA were measured by real-time RT-PCR. In addition, we detected the production of IL-17A by enzyme-linked immunosorbent assay, we explored the effects of CGRP on the differentiation of Th17 and Treg cells form Several aspects. Results CGRP increased the proportion of Th17 cells and the expression of RORyt mRNA(P〈0. 05). Additionally, CGRP also increased the production of IL-17A (P〈 0.05), these effects of CGRP were in dose-dependent manner. Meanwhile, CGRP inhibited the percentage of Treg cells and the expression of FoxP3 mRNA also in dose-dependent way(P〈0.05), peaking at 10-6 M. Conclusion These results demonstrate that CGRP facilitates naive CD4+T cells from human peripheral blood differentiate to Th17 cells, but suppresses them differentiate to Treg cells.
出处
《河北医科大学学报》
CAS
2017年第3期326-330,共5页
Journal of Hebei Medical University
基金
河北省高等学校科学研究计划(QN2015006)
河北省医学科学研究重点课题(20150626
20150631)