抗STAT3shRNA对树突状细胞成熟影响研究及其靶向防龋DNA疫苗构建

doi:10.13701/j.cnki.kqyxyj.2016.03.006

口腔医学研究 ›› 2016, Vol. 32 ›› Issue (3): 234-238.DOI: 10.13701/j.cnki.kqyxyj.2016.03.006

抗STAT3shRNA对树突状细胞成熟影响研究及其靶向防龋DNA疫苗构建

刘苗苗¹, 张艳¹, 张偲¹, 樊明文^{1, 2*}, 郭继华^{1, 2*}

1. 武汉大学口腔医学院牙体牙髓科，湖北省口腔基础医学重点实验室-省部共建国家重点实验室培育基地口腔生物医学教育部重点实验室（武汉大学）湖北武汉 430079；
2. 武汉大学口腔医院牙体牙髓科湖北武汉 430079

收稿日期:2015-12-21 出版日期:2016-03-28 发布日期:2016-03-29
通讯作者: 樊明文,电话:027-87647443
郭继华,电话:027-87686208
作者简介:刘苗苗（1989~ ），女，江苏人，硕士，主要从事牙体牙髓病学相关基础研究和临床工作。
基金资助:
国家自然科学基金资助项目(编号:81170955)
中央高校基本科研业务费资助(编号:2042014kf0270)

Effect of Anti-STAT3 shRNA on Dendritic Cell Maturation and Corresponding Targeted Anti-caries DNA Vaccine Construction.

LIU Miao-miao¹, ZHANG Yan¹, ZHANG Si¹, FAN Ming-wen^1,2*, GUO Ji-hua^1,2*

1. Hubei-MOST KLOS & KLOBME, School & Hospital of Stomatology, Wuhan University, Wuhan 430079, China;
2. Department of Endodontics, School & Hospital of Stomatology, Wuhan University, Wuhan 430079, China

Received:2015-12-21 Online:2016-03-28 Published:2016-03-29

摘要/Abstract

摘要： 目的: 研究以慢病毒为载体的抗小鼠STAT3 shRNA对树突状细胞(DC)成熟的影响,构建抗STAT3 shRNA树突状细胞靶向防龋DNA 疫苗。方法: 制备抗小鼠STAT3 shRNA慢病毒,体外转染树突状细胞系DC2.4,用Western杂交检测抑制STAT3表达的效果;用流式细胞术检测CD40、CD80和CD86表达;在抑制DC2.4的STAT3表达后,用LPS刺激,流式细胞术检测其对DCs成熟的影响;将针对STAT3的特异性shRNA整合入DC靶向防龋DNA疫苗pGJA-P/VAX,转染HEK-293细胞检测抑制STAT3表达效果。结果: 抗STAT3 shRNA慢病毒转染DC2.4后,可明显抑制STAT3表达,并增加DC表面标记物CD40、CD80和CD86的表达;用LPS刺激后,CD40、CD80和CD86表达进一步增强;抗STAT3 shRNA DC靶向防龋DNA疫苗转染HEK-293细胞后可以抑制STAT3表达。结论: 小鼠抗STAT3 shRNA慢病毒可有效抑制DC2.4 STAT3的表达,STAT3沉默后可促进DC2.4成熟。成功构建抗STAT3 shRNA DC靶向防龋DNA疫苗。

关键词: STAT3, 慢病毒, 树突状细胞, shRNA, DNA疫苗

Abstract: Objective: To investigate the effect of anti-STAT3 shRNA on dendritic cell maturation and to construct an anti-careis DNA vaccine carrying anti-STAT3 shRNA to enhance the efficacy of DNA vaccination. Methods: Dendritic cell line DC2.4 was infected by lentivirus containing shRNA targeting mouse STAT3. Total cellular protein was collected to detect expression level of STAT3 by western blot. CD40, CD80 and CD86 of DC2.4 were analyzed by FACS. LPS was used after infection to stimulate DC maturation. Anti-careis DNA vaccine carrying anti-STAT3 shRNA was constructed by cloning shRNA fragment into DC target anti-caries DNA vaccine pGJA-P/VAX. The efficiency of its inhibition on STAT3 expression was confirmed in HEK-293 cells. Results: Expression of STAT3 decreased in DC2.4 infected by lentivirus containing shRNA targeting STAT3. The expression levels of CD40, CD80 and CD86 of DC2.4 increased upon STAT3 knockdown, which were further up-regulated after the stimulation of LPS. Anti-caries DNA vaccine with anti-STAT3 shRNA could inhibit STAT3 expression. Conclusion: Lentiviral mediated shRNA interference targeting STAT3 could inhibit the STAT3 expression level of DC2.4 and increase the expression of CD40, CD80 and CD86 molecules and the maturation of dendritic cells. Anti-caries DNA vaccine against STAT3 was constructed successfully.

Key words: STAT3 Lentivirus, Dendritic cell, shRNA, DNA vaccine

中图分类号:

R780.1

刘苗苗, 张艳, 张偲, 樊明文, 郭继华. 抗STAT3shRNA对树突状细胞成熟影响研究及其靶向防龋DNA疫苗构建[J]. 口腔医学研究, 2016, 32(3): 234-238.

LIU Miao-miao, ZHANG Yan, ZHANG Si, FAN Ming-wen, GUO Ji-hua. Effect of Anti-STAT3 shRNA on Dendritic Cell Maturation and Corresponding Targeted Anti-caries DNA Vaccine Construction.[J]. Journal of Oral Science Research, 2016, 32(3): 234-238.

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抗STAT3shRNA对树突状细胞成熟影响研究及其靶向防龋DNA疫苗构建

Effect of Anti-STAT3 shRNA on Dendritic Cell Maturation and Corresponding Targeted Anti-caries DNA Vaccine Construction.

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