Liver Research Center, Brown Alpert Medical School and Rhode Island Hospital, Providence, RI 02903, USA.
J Viral Hepat. 2010 Oct;17(10):705-13. doi: 10.1111/j.1365-2893.2009.01228.x.
Chronic hepatitis C (HCV) infection is a substantial medical problem that leads to progressive liver disease, cirrhosis, and hepatocellular carcinoma (HCC). The aim of this study was to achieve sustained cellular immune responses in vivo to a HCV nonstructural protein using dendritic cell (DC)-based immunization approach. We targeted the HCV NS5 protein to DCs in vivo by injecting microparticles loaded with this antigen. The DC population was expanded in BALB/C mice (H-2(d) ) by hydrodynamic injection of a plasmid pUMVC3-hFLex expressing the secreted portion of the human Fms-like tyrosine kinase receptor-3 ligand (hFlt3). Mice were subsequently injected with microparticles coated with HCV NS5 protein via the tail vein. Cellular immune responses were determined with respect to secretion of INFγ and IL2 by CD4(+) cells and cytotoxic T-lymphocyte (CTL) assays in vitro; inhibition of tumour cell growth was employed for the assessment of CD8(+) generated activity in vivo. We found that Flt3L treatment expanded the DC population in the spleen to 43%, and such cells displayed a striking upregulation of CD86 as well as CD80 and CD40 co-stimulating molecules. Viral antigen-specific T(H) 1 cytokine secretion by splenocytes was generated, and CTL activity against syngeneic NS5 expressing myeloma target cells was observed. In addition, these cells inhibited tumour growth indicating that NS5-specific robust CTL activity was operative in vivo. Thus, the capability of activating DCs in vivo using the methods described is valuable as a therapeutic vaccine strategy for chronic HCV infection.
慢性丙型肝炎（HCV）感染是一个严重的医学问题，可导致进行性肝疾病、肝硬化和肝细胞癌（HCC）。本研究的目的是通过树突状细胞（DC）免疫接种方法在体内实现对 HCV 非结构蛋白的持续细胞免疫应答。我们通过注射负载这种抗原的微粒将 HCV NS5 蛋白靶向体内的 DC。通过在 BALB/C 小鼠（H-2(d)）中注射表达人 Fms 样酪氨酸激酶受体-3 配体（hFlt3）分泌部分的质粒 pUMVC3-hFLex，扩展 DC 群体。随后，通过尾静脉向小鼠注射涂有 HCV NS5 蛋白的微粒。通过体外测定 CD4(+)细胞分泌 INFγ和 IL2，以及细胞毒性 T 淋巴细胞（CTL）测定，确定细胞免疫应答；通过评估体内 CD8(+)产生的活性，采用抑制肿瘤细胞生长来评估。我们发现 Flt3L 处理将脾中的 DC 群体扩展到 43%，并且这些细胞表现出明显的 CD86 以及 CD80 和 CD40 共刺激分子的上调。脾细胞产生针对病毒抗原的 T(H)1 细胞因子分泌，并且观察到针对表达同种型 NS5 的骨髓瘤靶细胞的 CTL 活性。此外，这些细胞抑制肿瘤生长，表明 NS5 特异性强大的 CTL 活性在体内起作用。因此，使用所描述的方法在体内激活 DC 的能力是治疗慢性 HCV 感染的有价值的治疗疫苗策略。
