CTP-HBcAg18-27-Tapasin联合佐剂CpG ODN免疫转基因小鼠的免疫应答研究

doi:10.12092/j.issn.1009-2501.2018.03.005

中国临床药理学与治疗学 ›› 2018, Vol. 23 ›› Issue (3): 271-276.doi: 10.12092/j.issn.1009-2501.2018.03.005

CTP-HBcAg_18-27-Tapasin联合佐剂CpG ODN免疫转基因小鼠的免疫应答研究

吴姗姗，唐余燕，陈小华，张毅，王洁玲，汤正好，臧国庆，余永胜

上海交通大学附属第六人民医院感染病科，上海 200233

收稿日期:2017-08-30 修回日期:2018-02-20 出版日期:2018-03-26 发布日期:2018-03-28
通讯作者: 余永胜，男，博士，主任医师，硕士生导师，研究方向：病毒性肝炎免疫治疗。 Tel: 021-64369181-8673 E-mail:yuyongsheng@medmail.com.cn
作者简介:吴姗姗，女，在读硕士，研究方向：病毒性肝炎免疫发病机制及免疫治疗。
基金资助:
上海教育委员会科研创新项目（15ZZ013）；上海市卫生和计划生育委员会（201640283）

Immune response elicited in HBV-transgenic mice by cytoplasmic transduction peptide-HBcAg18-27-Tapasin adjuvanted with CpG ODN

WU Shanshan, TANG Yuyan, CHEN Xiaohua, ZHANG Yi, WANG Jieling, TANG Zhenghao, ZANG Guoqing, YU Yongsheng

Department of Infectious Diseases,Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai 200233, China

Received:2017-08-30 Revised:2018-02-20 Online:2018-03-26 Published:2018-03-28

摘要/Abstract

摘要：

目的: 观察以非甲基化寡聚脱氧核苷酸基序（CpG oligodeoxynucleotides,CpG ODN）为佐剂联合融合蛋白胞质转导肽（CTP）-HBcAg18-27-Tapasin免疫乙型病毒性肝炎(hepatitis B virus,HBV)转基因小鼠的免疫反应。方法:HBV转基因小鼠随机分为5组，实验组：CTP-HBcAg18-27-Tapasin+CpG ODN 组，对照组：CTP-HBcAg18-27-Tapasin，干扰素（IFN-α）组，CpG ODN组，空白组为生理盐水组。融合蛋白及CpG ODN经肌肉注射免疫小鼠，流式细胞仪检测病毒特异性T细胞反应（HBV-specific cytotoxic T cell,CTL）变化，全自动免疫分析仪检测血清乙肝表面抗原(hepatitis B virus surface antigen,HBsAg)水平，ELISA检测T淋巴细胞培养上清白介素（IL）-2、γ-干扰素（IFN-γ）水平，实时荧光定量PCR(real time quantitative polymerase chain reaction,qRT-PCR)检测HBV DNA水平，免疫组化观察HBV转基因小鼠肝脏中HBsAg水平。结果:CTP-HBcAg18-27-Tapasin+CpG ODN组中IFN-γ+CD8+双阳性T细胞百分比增高，细胞培养上清IL-2、IFN-γ水平升高，肝脏病理显示炎性细胞增多，免疫组化显示HBsAg表达水平明显下降，同时对血清HBsAg及HBV DNA水平进行比较，CTP-HBcAg18-27-Tapasin有明显抑制作用。结论:CpG-ODN作为佐剂可以增强融合蛋白CTP-HBcAg18-27-Tapasin诱导的HBV转基因小鼠抗病毒免疫应答。

关键词: 胞质转导肽, 抗病毒反应, 佐剂

Abstract:

AIM: To investigate the effects of fusion protein of cytoplasmic transduction peptide (CTP) -HBcAg18-27-Tapasin adjuvanted with CpG ODN on immune response and anti-viral response in HBV transgenic mice. METHODS: HBV-transgenic mice were randomly divided into 5 groups: experimental group: CTP-HBcAg18-27-Tapasin+CpG ODN, control group: CpG ODN, IFN-α and CTP-HBcAg18-27-Tapasin, vehicle group: normal saline. Serum samples were collected at various times before and after immunization. Intracellular cytokine in T lymphocytes was analyzed by flow cytometry, and the levels of IL-2 and IFN-γ secreted by T lymphocytes were detected by ELISA. The expression of HBsAg in liver sections was detected by immunohistochemistry. Sera levels of HBV DNA and HBsAg were examined by qRT-PCR and automatic enzyme-linked immunosorbent assay, respectively. RESULTS:An increased percentage of IFN-γ+CD8+ T cells was detected in CTP-HBcAg18-27-Tapasin+CpG ODN group and predominantly Th1 cellular immune response. In addition, the expression levels of serum HBsAg and HBV DNA were significantly inhibited, along with evident down-regulation of HBsAg in liver samples than CTP-HBcAg18-27-Tapasin alone group. CONCLUSION: CpG ODN acts as adjuvant could enhance anti-virus immune responses in HBV transgenic mice immunized with fusion protein CTP-HBcAg18-27-Tapasin.

Key words: CTP-HBcAg18-27-Tapasin, antiviral immune response, adjuvant

中图分类号:

R965.2

吴姗姗，唐余燕，陈小华，张毅，王洁玲，汤正好，臧国庆，余永胜. CTP-HBcAg_18-27-Tapasin联合佐剂CpG ODN免疫转基因小鼠的免疫应答研究[J]. 中国临床药理学与治疗学, 2018, 23(3): 271-276.

WU Shanshan, TANG Yuyan, CHEN Xiaohua, ZHANG Yi, WANG Jieling, TANG Zhenghao, ZANG Guoqing, YU Yongsheng . Immune response elicited in HBV-transgenic mice by cytoplasmic transduction peptide-HBcAg18-27-Tapasin adjuvanted with CpG ODN[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2018, 23(3): 271-276.

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CTP-HBcAg_18-27-Tapasin联合佐剂CpG ODN免疫转基因小鼠的免疫应答研究

Immune response elicited in HBV-transgenic mice by cytoplasmic transduction peptide-HBcAg18-27-Tapasin adjuvanted with CpG ODN

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