摘要
目的探讨IL-6和B细胞活化因子(B cell activating factor,BAFF)的融合蛋白真核表达质粒对干燥综合征动物模型非肥胖糖尿病小鼠(non-obese diabetic mouse,NOD)唾液腺、泪腺组织病理的影响,阐明IL-6/BAFF融合蛋白真核表达质粒对NOD小鼠可能的治疗机制。方法构建IL-6/BAFF融合蛋白真核表达质粒,转染CHO细胞,Western blot检测融合蛋白的表达水平。IL-6/BAFF融合蛋白真核表达质粒免疫BALB/c小鼠,每2周1次,共3次,免疫结束后处死小鼠,ELISA法检测血清中抗IL-6和抗BAFF抗体效价。将21只NOD小鼠随机数字表法分为空白组、阴性对照组、治疗组。治疗组小鼠肌肉注射IL-6/BAFF融合蛋白真核表达质粒,阴性对照组小鼠肌肉注射对照质粒,空白组不做处理,每周1次,6次免疫后处死NOD小鼠,心脏采血取血清,ELISA法检测血清中抗IL-6和抗BAFF抗体及细胞因子IL-6、BAFF、IFN-γ、IL-10、IL-17A的表达水平;取脾组织流式细胞术检测小鼠脾细胞中Th17、Treg、Th1、Th2的比例;HE染色观察小鼠泪腺、唾液腺中灶状淋巴细胞浸润情况和病理改变。结果IL-6/BAFF融合蛋白真核表达质粒使BALB/c小鼠血清中抗IL-6和抗BAFF抗体水平升高(P<0.05)。治疗组NOD小鼠血清中抗IL-6和抗BAFF抗体含量升高(P<0.01);IL-6、BAFF、IFN-γ、IL-10、IL-17A的表达量低于空白组和阴性对照组(P<0.01);脾细胞中Treg、Th2细胞比例增高(P<0.05);泪腺及唾液腺中腺泡大小不一和形态不规则明显改善,导管扩张减轻,灶状淋巴细胞浸润减少。结论IL-6/BAFF融合蛋白真核表达质粒可诱导NOD小鼠体内抗IL-6和抗BAFF抗体产生,进而减轻炎症因子的表达水平,逆转Th17/Treg、Th1/Th2的平衡,同时改善泪腺及唾液腺内不规则的腺泡结构及导管扩张,减少灶状淋巴细胞浸润。本研究结果表明IL-6/BAFF融合蛋白真核表达质粒可以作为靶向T、B细胞的潜在治疗靶点。
Objective To investigate the effects of a eukaryotic expression plasmid for IL-6 and B-cell activating factor(BAFF)fusion protein on the histopathological changes in salivary and lacrimal glands of non-obese diabetic(NOD)mice with Sjögren′s syndrome and to elucidate the possible therapeutic mechanism of IL-6/BAFF fusion protein eukaryotic expression plasmid in NOD mice.Methods The eukaryotic expression plasmid for IL-6/BAFF fusion protein was constructed.After transfecting CHO cells with the plasmid,the expression of IL-6/BAFF fusion protein was detected by Western blot.BALB/c mice were injected with the plasmid every two weeks for three times and the titers of anti-IL-6 and anti-BAFF antibodies were measured by ELISA.Twenty-one NOD mice were randomly divided into three groups(control group,empty vector group and therapy group)by numerical table method.The mice in the therapy group were injected with the IL-6/BAFF fusion protein eukaryotic expression plasmid once a week for six times and the mice in the empty vector group were injected with empty plasmid.The levels of anti-IL-6 and anti-BAFF antibodies as well as cytokines(IL-6,BAFF,INF-γ,IL-10 and IL-17A)in mouse serum samples were detected by ELISA.The proportions of Th17,Treg,Th1 and Th2 cells in mouse splenocytes were measured by flow cytometry.Focal lymphocyte infiltration and pathological changes in the lacrimal and salivary glands of mice were observed under light microscopy after HE staining.Results The eukaryotic expression plasmid for IL-6/BAFF fusion protein increased the levels of anti-IL-6 and anti-BAFF antibodies in the serum of BALB/c mice(P<0.05).The levels of anti-IL-6 and anti-BAFF antibodies in the serum of NOD mice in the therapy group increased(P<0.01),while the expression of IL-6,BAFF,INF-γ,IL-10 and IL-17A in NOD mice in the therapy group was lower than that in the control group and the empty vector group(P<0.05).The percentages of Treg and Th2 cells in the splenocytes of NOD mice increased after treatment(P<0.05).Moreover,the eukaryot
作者
高雅静
庞春艳
尹芳蕊
吕凤凤
王永福
Gao Yajing;Pang Chunyan;Yin Fangrui;Lyu Fengfeng;Wang Yongfu(Department of Rheumatism and Immunology,the First Affiliated Hospital of Baotou Medical College,Inner Monolia University of Secience and Technology,Baotou 014010,China;Inner Mongolia Autoimmunology Key Laboratory,the First Affiliated Hospital of Baotou Medical College,Inner Monolia University of Secience and Technology,Baotou 014010,China)
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2022年第9期714-721,共8页
Chinese Journal of Microbiology and Immunology
基金
国家自然科学基金项目(81760302)。
