Genetic immunization against hepatitis B virus with calcium phosphate nanoparticles in vitro and in vivo
Genetic immunization against hepatitis B virus with calcium phosphate nanoparticles in vitro and in vivo
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磷酸钙纳米颗粒体外和体内针对乙型肝炎病毒的基因免疫
DOI：
10.1016/j.actbio.2020.04.021
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发表时间：
2020
影响因子：
9.7
通讯作者：
Epple Matthias
中科院分区：
文献类型：
--
作者：
Rojas-Sanchez Leonardo;Zhang Ejuan;Sokolova Viktoriya;Zhong Maohua;Yan Hu;Lu Mengji;Li Qian;Yan Huimin;Epple Matthias
磷酸钙纳米颗粒负载有质粒DNA和toll样受体配体（TLR），即CpG或鞭毛蛋白，以激活抗原呈递细胞（APC）如树突状细胞（DC）。在体外对功能化纳米粒子在HeLa、C2 C12和BHK-21细胞系中进行了研究，重点是两种特异性蛋白的表达。以编码增强型绿色荧光蛋白（EGFP）的EGFP-DNA为模型质粒，通过荧光显微镜和流式细胞术优化转染效率。通过体外和体内免疫实验评价了负载TLR配体和编码B病毒表面抗原（pHBsAg）的质粒DNA的磷酸钙纳米颗粒，以鉴定预防性B病毒（HBV）疫苗的可能候选物。纳米颗粒在三种细胞系中诱导了HBsAg的强烈表达。在脾细胞中，共刺激分子CD 80和CD 86的表达增强。在小鼠肌肉注射后，纳米颗粒诱导HBsAg表达、抗原特异性T细胞应答和抗原特异性抗体应答（IgG 1）。重要性声明B型肝炎是全球最常见的病毒感染之一。对于预防性免疫，可以使用携带佐剂（刺激分子）和编码病毒抗原的DNA的纳米颗粒。在将这种纳米颗粒给予细胞后，它们被细胞吸收，其中DNA被转录成病毒抗原（蛋白质）。该病毒抗原诱导病毒特异性免疫应答。这一点在体外细胞培养和小鼠体内的伸展静脉研究中都得到了证实。
Calcium phosphate nanoparticles were loaded with plasmid DNA and toll-like receptor ligands (TLR),i.e.CpG or flagellin, to activate antigen-presenting cells (APCs) like dendritic cells (DCs). The functionalized nanoparticles were studiedin vitroon HeLa, C2C12 and BHK-21 cell lines, focusing on the expression of two specific proteins. EGFP-DNA, encoding for enhanced green fluorescent protein (EGFP), was used as a model plasmid to optimize the transfection efficiencyin vitroby fluorescence microscopy and flow cytometry. Calcium phosphate nanoparticles loaded with TLR ligands and plasmid DNA encoding for the hepatitis B virus surface antigen (pHBsAg) were evaluated byin vitroandin vivoimmunization experiments to identify a possible candidate for a prophylactic hepatitis B virus (HBV) vaccine. The nanoparticles induced a strong expression of HBsAg in the three cell lines. In splenocytes, the expression of the co-stimulatory molecules CD80 and CD86 was enhanced. After intramuscular injection in mice, the nanoparticles induced the expression of HBsAg, the antigen-specific T cell response, and the antigen-specific antibody response (IgG1).Statement of SignificanceHepatitis B is one of the most frequent viral infections worldwide. For preventive immunization, nanoparticles can be used which carry both an adjuvant (a stimulatory molecule) and DNA encoding for a viral antigen. After administration of such nanoparticles to cells, they are taken up by cells where the DNA is transcribed into the viral antigen (a protein). This viral antigen is inducing a virus-specific immune response. This was shown both byin vitrocell culture as well as by an extensivein vivostudy in mice.