Lv Sunjian, Lu Binjie, Xu Jiehao, Xu Haisheng, Zhao Jing, Li Song, Li Yiqun, Chen Yuyin
College of Animal Sciences, Zhejiang University, 866 Yuhangtang Road, Hangzhou, Zhejiang Province 310058, China.
College of Animal Sciences, Zhejiang University, 866 Yuhangtang Road, Hangzhou, Zhejiang Province 310058, China.
Dev Comp Immunol. 2015 Jul;51(1):56-64. doi: 10.1016/j.dci.2015.02.016. Epub 2015 Mar 2.
Peroxinectin possesses the features of both peroxidase activity and adhesive property and plays important roles in innate immune system of crustaceans. In this study, the sequence of peroxinectin of Eriocheir sinensis (EsPX) was analyzed and its expression in response to exterior stimulation was detected in both in vivo and in vitro examination. We showed that the full-length cDNA sequence was composed of 2701 bp and owned a molecular mass of 85.2 kDa and a theoretical pI (isoelectric point) of 6.91. Real-time PCR revealed that the EsPX was mainly distributed in the muscle, hemocytes and stomach. Furthermore, the EsPX was verified to be located in hyalinocytes, semigranulocytes and granulocytes, and was distributed throughout the cytoplasm and nucleus, especial in cytoplasm. After injected with beads, lipopolysaccharide (LPS) and Aeromonas hydrophila, the EsPX mRNA expression was significantly up-regulated and peaked up at 4, 2 and 16 h respectively (P <0.05). In the in vitro experiment, the stimulation of LPS and beads also induced a prominent boost of EsPX protein in primary cultured hemocytes. The expression of EsPX was peaked up at 4 and 8 h for LPS and beads challenged groups respectively, followed by remarkable release of the incremental EsPX into the extracellular matrix. These findings suggested that the expression of EsPX was susceptible to exterior stimulation, and that the highly expressional EsPX would be released into extracellular matrix by the exterior stimulus.
过氧化物酶激活蛋白兼具过氧化物酶活性和黏附特性，在甲壳动物的先天免疫系统中发挥重要作用。本研究对中华绒螯蟹过氧化物酶激活蛋白（EsPX）的序列进行了分析，并通过体内和体外实验检测了其对外界刺激的响应表达。结果表明，其全长cDNA序列由2701 bp组成，分子量为85.2 kDa，理论等电点为6.91。实时荧光定量PCR显示，EsPX主要分布于肌肉、血细胞和胃中。此外，EsPX被证实定位于透明细胞、半颗粒细胞和颗粒细胞中，且分布于整个细胞质和细胞核，尤其在细胞质中。注射珠子、脂多糖（LPS）和嗜水气单胞菌后，EsPX mRNA表达显著上调，分别在4、2和16 h达到峰值（P<0.05）。在体外实验中，LPS和珠子刺激也诱导原代培养血细胞中EsPX蛋白显著增加。LPS和珠子刺激组的EsPX表达分别在4和8 h达到峰值，随后增加的EsPX显著释放到细胞外基质中。这些结果表明，EsPX的表达易受外界刺激影响，且高表达的EsPX会因外界刺激而释放到细胞外基质中。
