重组PACAP13神经肽对小鼠胸腺功能的影响  

Effect of the recombinant neuropeptide PACAP13 on thymus function of mice model

作　　者：吴倩妍 梁阳 刘贞银 张华华 Wu Qianyan;LiangYang;Liu Zhenyin;Zhang Huahua(School of Basic Medicin,Guangdong Medical University,Dongguan 523808;Guangdong Maoming Health Vocational College,Maoming 525400;Scientific Research Platform Service Management Center,Guangdong Medical University,Dongguan 523808)

机构地区：[1]广东医科大学基础医学院,东莞523808 [2]广东茂名健康职业学院,茂名525400 [3]广东医科大学科研平台服务管理中心,东莞523808

出　　处：《安徽医科大学学报》2022年第6期877-884,共8页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:31200679);广东省自然科学基金(编号:2018A0303130291)。

摘　　要：目的 在细胞、器官及个体水平探究垂体腺苷酸环化酶激活肽重组13肽(PACAP13肽)对胸腺功能的影响及其机制。方法 通过体外原代培养胸腺上皮细胞(TEC)及骨髓间充质干细胞(MSC),设置淋巴细胞组、淋巴细胞+TEC组、淋巴细胞+MSC组、淋巴细胞+TEC+MSC组,检测PACAP13肽处理后各组淋巴细胞的增殖情况,检测培养液中细胞因子白细胞介素(IL)-1、IL-2、IL-6和转化生长因子(TGF)的含量;上述各组细胞加入地塞米松(Dex)诱导细胞凋亡,AnnexinⅤ-FITC/PI流式双染检测PACAP13肽对各组淋巴细胞凋亡的情况。设置胸腺组、胸腺+TEC组、胸腺+MSC组、胸腺+TEC+MSC组,并添加PACAP13肽,实时定量PCR(qPCR)检测胸腺信号结合T细胞受体删除环(sjTREC)的含量。注射降植烷诱导狼疮模型小鼠,随机分为模型组和PACAP13组,模型组注射0.9%生理盐水,PACAP13组注射PACAP13肽,qPCR检测胸腺、脾脏sjTREC含量;ELISA检测血清狼疮相关自身抗体水平及逆转录PCR(qRT-PCR)检测胸腺相关mRNA表达水平。结果 与淋巴细胞组相比,淋巴细胞+TEC组、淋巴细胞+MSC组、淋巴细胞+TEC+MSC组胸腺淋巴细胞增殖率增加,凋亡率下降(P<0.05);与对照组相比,PACAP13组淋巴细胞增殖率增加,凋亡率下降,细胞培养液中细胞因子IL-1、IL-2、IL-6和TGF增加(P<0.05)。与胸腺组相比,胸腺+TEC组、胸腺+MSC组、胸腺+TEC+MSC组胸腺sjTREC含量增加;与对照组相比,PACAP13肽组胸腺sjTREC含量增加。与模型组相比,PACAP13组狼疮小鼠胸腺和脾脏中sjTREC的含量增加(P<0.05),胸腺中Aire、Fezf2、Foxp3、TGF-βmRNA表达水平增加(P<0.01),胸腺CXCL13mRNA表达水平下降(P<0.01),血清自身抗体抗dsDNA抗体、抗RNP/sm抗体、ANA水平下降(P<0.05)。结论 PACAP13肽能促进胸腺淋巴细胞增殖和抗凋亡,在体外及体内均具有增强胸腺淋巴细胞的生成和输出能力以及降低狼疮小鼠自身抗体水平的免疫调节作用,其作用机制与调节胸腺细Objective To investigate the effect and mechanism of pituitary adenylate cyclase activated peptide recombinant 13 peptide(PACAP13)on thymus at cell,organ and individual levels.Methods Thymic epithelial cells(TEC)and bone marrow mesenchymal stem cells(MSC)were cultured in vitro,lymphocyte group,lymphocyte+TEC group,lymphocyte+MSC group and lymphocyte+TEC+MSC group were set up to detect the proliferation of lymphocytes in each group treated by PACAP13 peptide,and contents of cytokines interleukin(IL)-1,IL-2,IL-6 and transforming growth factor(TGF)in culture medium were detected.Dexamethasone(Dex)was added to induce cell apoptosis,and Annexin V-FITC/PI flow cytometry double staining was used to detect the effect of PACAP13 peptide on cell apoptosis.Thymus group,thymus+TEC group,thymus+MSC group,thymus+TEC+MSC group were set up,and PACAP13 peptide was added,the content of signal joint T cell receptor rearrangement excision cricles(sjTREC)in thymus was detected by real-time quantitative PCR(qRCR).The pristane-induced lupus model mice were randomly divided into model group and PACAP13 group.Model group was injected with 0.9%saline,and PACAP13 group was injected with PACAP13 peptide,thymus and spleen sjTREC levels were detected by qPCR,serum lupus-related autoantibodies were detected by ELISA and thymus-related mRNA expression levels were detected by reverse transcription quantitative PCR(qRT-RCR).Results Compared with the lymphocyte group,the proliferation rate of thymus lymphocytes in the lymphocyte+TEC group,lymphocyte+MSC group,lymphocyte+TEC+MSC group increased,and the apoptosis rate decreased(P<0.05),compared with the control group,the proliferation rate of lymphocytes in the PACAP13 group increased,the rate of apoptosis decreased,and the cytokines IL-1,IL-2,IL-6 and TGF in the cell culture medium increased(P<0.05).Compared with the thymus group,thymus+TEC group,thymus+MSC group,thymus+TEC+MSC group increased the content of thymus sjTREC,compared with the control group,PACAP13 peptide group thymus sjTREC content i

关 键 词：PACAP13肽 胸腺 胸腺上皮细胞 骨髓间充质干细胞 

分 类 号：R392.11[医药卫生—免疫学]