低氧胁迫对瘤背石磺Toll样受体4、血细胞和免疫酶活性的影响  

Effects of hypoxia stress on Toll-like receptor 4,blood cells and immune enzyme activities in Onchidium reevesii

作　　者：肖海明 土志涵 饶榕城 陈锡林 钱畅 张小明 沈和定[1,2,3] XIAO Haiming;TU Zhihan;RAO Rongcheng;CHEN Xilin;QIAN Chang;ZHANG Xiaoming;SHEN Heding(Shanghai Collaborative Innovation Center for Cultivating Elite Breeds and Green-culture of Aquaculture Animals,Shanghai Ocean University,Shanghai 201306,China;Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources,Ministry of Education,Shanghai Ocean University,Shanghai 201306,China;National Demonstration Center for Experimental Fisheries Science Education,Shanghai Ocean University,Shanghai 201306,China)

机构地区：[1]上海海洋大学,上海市水产动物良种创制与绿色养殖协同创新中心,上海201306 [2]上海海洋大学,水产种质资源发掘与利用教育部重点实验室,上海201306 [3]上海海洋大学,水产科学国家级实验教学示范中心,上海201306

出　　处：《水产学报》2024年第6期37-49,共13页Journal of Fisheries of China

基　　金：上海市协同创新中心任务(A1-3605-21-000202)。

摘　　要：为探究瘤背石磺面临低氧环境时固有免疫的应激机制,实验以RACE法对瘤背石磺TLR4基因进行全长克隆,并进行生物信息学分析,测定了低氧胁迫下瘤背石磺TLR4基因的表达变化,以及分析了血细胞活力和溶菌酶(LSZ)活性、肝脏中超氧化物歧化酶(SOD)和碱性磷酸酶(ALP)活性。结果显示,瘤背石磺TLR4基因cDNA全长共3605 bp,包括编码956 aa氨基酸的2817 bp开放阅读框。系统进化树表明,瘤背石磺TLR4基因与光滑双脐螺TLR4基因的进化地位较为接近。qRT-PCR结果显示,TLR4基因在瘤背石磺7个组织中均有表达,其中表达量最高的是肝脏。低氧胁迫下,各组织中TLR4基因的表达量皆显著上升,其中神经节在4 h时率先达到峰值。此外,血细胞活力、LSZ活性、ALP活性都是呈先下降后上升的趋势,而SOD活性呈先上升再下降再上升的趋势,波动幅度最为明显。研究结果初步说明了TLR4的生理功能,为探究潮间带生物免疫系统的低氧应激机制提供理论参考。In order to explore the mechanism of innate immunity in Onchidium reevesii under hypoxia stress,the cDNA sequence of Toll-like receptor 4(TLR4)gene was cloned by RACE-PCR,and bioinformatics analysis was conducted.In addtion,the expression changes of TLR4,the activity changes of hemocyte,lysozyme(LSZ),superoxide dismutase(SOD)and alkaline phosphatase(AKP)under hypoxia stress were determined.The results showed that the full length of TLR4 cDNA was 3,605 bp,the length of open reading frame was 2,817 bp,and it encoded a total of 956 amino acids.The phylogenetic tree results suggest that the TLR4 gene was the closest to the TLR4 gene of Biomphalaria glabrata.The results of qRT-PCR indicated that the TLR4 gene was expressed in all tissues,and the relative expression in the hepatopancreas was the highest.The expression of each tissue increased significantly under hypoxia stress and the expression of ganglion reached its peak earlier than the other tissues at 4 h.In addition,hemocyte viability,LSZ activity and AKP activity decreased at first and then increased,while SOD activity increased at first,then decreased and finally increased,which was more sensitive to hypoxia stress than the others.The results provide a data base for illustrating TLR4 biological functions and a theoretical foundation for investigation the mechanism of the immune system in intertidal macrobenthos under hypoxia stress.

关 键 词：瘤背石磺 TLR4 基因克隆 低氧胁迫 固有免疫 

分 类 号：S968.3[农业科学—水产养殖]