• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>World Journal of Gastroenterology >Generation of cytotoxic T cell against HBcAg using retrovirally transduced dendritic cells
【24h】

Generation of cytotoxic T cell against HBcAg using retrovirally transduced dendritic cells

机译：使用逆转录病毒转导的树突状细胞产生针对HBcAg的细胞毒性T细胞

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

AIM: Cytotoxic T lymphocytes (CTLs) play an important role in resolving HBV infection. In the present study, we attempted to evaluate the efficiency of bone marrow-derived dendritic cells (DCs) transduced with recombinant retroviral vector bearing hepatitis B virus (HBV) core gene and the capability of generating CTLs against HBcAg by genetically modified DCs in vivo. METHODS: A retroviral vector containing HBV core gene was constructed. Replicating DC progenitor of C57BL/6 mice was transduced by retroviral vector and continually cultured in the presence of recombinant mouse granulocyte-macrophage colony-stimulating factor (rmGM-CSF) and interleukin-4(IL-4) for 6 days. LPS was added and cultured for additional two days. The efficiency of gene transfer was determined by PCR, Western blot and FACS. Transduced DCs immunized C57BL/6 mice subcutaneously 2 times at an one-week interval. Intracellular IFN-γ and IL-4 of immunized mice lymphocytes were analyzed. Generation of CTLs in lymphocytes stimulated with mitomycin C-treated EL4-C cell which stably expresses HBcAg was determined by LDH release assays. RESULTS: Recombinant retroviral expression vector (pLCSN) was positively detected by PCR as well as enzyme digestion with EcoRI and BamH I. Retroviruses were generated by pLCSN transfection packing cell and the virus titer was 3 x 10~5 CFU/ml. Indirect immunofluorescence and FACS showed that HBV core gene was expressed in murine fibroblasts. Transduced bone marrow cells had capability of differentiating into DCs in vitro in the presence of rmGM-CSF and rmIL-4. The result of PCR showed that HBV core gene was integrated into the genome of transduced DCs. Western blot analysis showed that HBV core gene was expressed in DCs. The transduction rate was 28 % determined by FACS. Retroviral transduction had no influence on DCs expressions of CD80 and MHC class Ⅱ. HBcAg specific CTLs and Th1 type immune responses could be generated in the mice by using transduced DCs as antigen presenting cells (APCs). progenitors expresses efficiently HBcAg, and genetically modified DCs evoke a higher CTLs response than HBcAg in vivo.
机译：目的：细胞毒性T淋巴细胞（CTL）在解决HBV感染中起重要作用。在本研究中，我们尝试评估用携带乙型肝炎病毒（HBV）核心基因的重组逆转录病毒载体转导的骨髓来源树突状细胞（DC）的效率，以及在体内通过基因修饰的DC产生针对HBcAg的CTL的能力。方法：构建含有HBV核心基因的逆转录病毒载体。通过逆转录病毒载体转导C57BL / 6小鼠的复制DC祖细胞，并在重组小鼠粒细胞-巨噬细胞集落刺激因子（rmGM-CSF）和白介素-4（IL-4）存在下连续培养6天。加入LPS，再培养两天。通过PCR，蛋白质印迹和FACS确定基因转移的效率。转导的DC以一周间隔两次皮下免疫C57BL / 6小鼠。分析了免疫小鼠淋巴细胞的细胞内IFN-γ和IL-4。通过LDH释放测定法确定了用丝裂霉素C处理的稳定表达HBcAg的EL4-C细胞刺激的淋巴细胞中CTL的产生。结果：通过PCR以及EcoRI和BamH I酶切检测，重组逆转录病毒表达载体（pLCSN）均被阳性检测。pLCSN转染包装细胞可产生逆转录病毒，病毒滴度为3 x 10〜5 CFU / ml。间接免疫荧光和流式细胞仪检测表明，HBV核心基因在鼠成纤维细胞中表达。在存在rmGM-CSF和rmIL-4的情况下，转导的骨髓细胞具有体外分化为DC的能力。 PCR结果表明，HBV核心基因已整合到转导DC的基因组中。蛋白质印迹分析表明，HBV核心基因在DC中表达。通过FACS确定的转导率为28％。逆转录病毒转导对CD80和Ⅱ类MHC的DC表达没有影响。通过将转导的DC用作抗原呈递细胞（APC），可在小鼠中产生HBcAg特异性CTL和Th1型免疫应答。祖细胞能有效表达HBcAg，而转基因的DC在体内引起的CTL响应要高于HBcAg。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号