Laboratory of Pathology and Immunology of Aquatic Animals, Ocean University of China, No. 5, Yushan Road, Qingdao 266003, PR China.
Fish Shellfish Immunol. 2013 Aug;35(2):539-45. doi: 10.1016/j.fsi.2013.05.004. Epub 2013 May 24.
Our previous study has demonstrated that an integrin β subunit of Chinese shrimp (Fenneropenaeus chinensis) (FcβInt) involved in WSSV infection. In order to further elucidate the potential role of the FcβInt in the WSSV infection, expression response of FcβInt to WSSV infection in shrimp hemocytes was investigated after intra-muscular injection with the virus. Following time-course hemocytes sampling, the expression variation of FcβInt in hemocytes was examined by flow cytometric immunofluorescence assay (FCIFA) and enzyme linked immunosorbent assay (ELISA) using the monoclonal antibody (Mab) 2C5 against FcβInt, which was successfully produced with recombinant partial FcβInt and exhibited binding to a 120 kDa hemocyte protein. Meanwhile, the dynamic state of FcβInt mRNA level and WSSV copies in hemocytes were determined by quantitative real-time PCR. The result of FCIFA showed that FcβInt was mainly expressed on the semi-granular and granular cells, which was down-regulated at 6 h post infection (p.i.), and significantly increased to the peak level at 12 h p.i., then decreased to the control level by 24 h. However, FcβInt on the hyaline cells was lowly expressed and didn't show active response to the viral infection. The variation of FcβInt concentrations in total hemocytes determined by ELISA was roughly in accordance with the changing tendency of FcβInt expressed on the semi-granular and granular cells. FcβInt mRNA level in total hemocytes was significantly up-regulated to the peak level at 12 h p.i. Moreover, the number of WSSV copies in hemocytes began to exhibit a significant increase at 24 h p.i. The present study demonstrated that WSSV infection would induce a differential regulation of FcβInt expression in different type hemocytes, which provided valuable evidences for the close correlation between FcβInt and WSSV infection.
我们之前的研究表明，中国对虾（Fenneropenaeus chinensis）的整合素β亚基（FcβInt）参与了 WSSV 感染。为了进一步阐明 FcβInt 在 WSSV 感染中的潜在作用，我们在肌肉内注射病毒后，研究了虾血细胞中 FcβInt 对 WSSV 感染的表达反应。通过时间过程血细胞采样，使用针对 FcβInt 的单克隆抗体（Mab）2C5 通过流式细胞术免疫荧光分析（FCIFA）和酶联免疫吸附试验（ELISA）检测了 FcβInt 在血细胞中的表达变化，该单克隆抗体是用重组部分 FcβInt 成功制备的，并且与 120 kDa 的血细胞蛋白结合。同时，通过定量实时 PCR 测定了 FcβInt mRNA 水平和血细胞中 WSSV 拷贝数的动态状态。FCIFA 的结果表明，FcβInt 主要表达在半颗粒细胞和颗粒细胞上，感染后 6 小时（pi）下调，12 小时达到峰值，然后在 24 小时下降到对照水平。然而，透明细胞上的 FcβInt 表达水平较低，对病毒感染没有表现出活跃的反应。ELISA 测定的总血细胞中 FcβInt 浓度的变化与半颗粒细胞和颗粒细胞上表达的 FcβInt 的变化趋势大致相符。总血细胞中 FcβInt mRNA 水平在感染后 12 小时显著上调至峰值。此外，24 小时后血细胞中的 WSSV 拷贝数开始显著增加。本研究表明，WSSV 感染会诱导不同类型血细胞中 FcβInt 表达的差异调节，为 FcβInt 与 WSSV 感染之间的密切相关性提供了有价值的证据。
