酿酒酵母甘露聚糖对绵羊瘤胃上皮细胞β-防御素-1(S BD-1)表达的影响  

Effects of mannan on the expression of SBD-1 in sheep rumen epithelial cells

作　　者：金鑫[1,2] 张曼[1,2] 王云鹤[1,2] 魏方 温婧怡 杨银凤[1,2] JIN Xin;ZHANG Man;WANG Yun-he;WEI Fang;WEN Jing-yi;YANG Yin-feng(College of veterinary,Inner Mongolia Agricultural University,Hohhot 010018,China;Key Laboratory of Clinical Diagnosis and Treatment Technology in Animal Disease,Ministry of Agriculture,Hob_hot 010018,China)

机构地区：[1]内蒙古农业大学兽医学院,内蒙古呼和浩特010018 [2]农业部动物疾病临床诊疗技术重点实验室,内蒙古呼和浩特010018

出　　处：《中国预防兽医学报》2018年第12期1117-1121,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金项目(31560682);内蒙古自治区研究生教育创新计划资助项目(B20171012921)

摘　　要：为探究酿酒酵母菌细胞壁成分甘露聚糖对绵羊瘤胃上皮细胞(RECs)β-防御素-1(SBD-1)表达的影响,本研究用不同浓度的甘露聚糖刺激RECs 8 h后,利用荧光定量PCR (qPCR)和ELISA方法分别检测RECsSBD-1 mRNA的转录水平和蛋白表达变化,确定甘露聚糖诱导SBD-1表达量最高的刺激浓度。然后用该浓度甘露聚糖对RECs进行不同时间的刺激,同样利用qPCR和ELISA方法对SBD-1 mRNA转录水平和蛋白的表达变化进行检测,确定甘露聚糖诱导SBD-1表达的最佳时间。qPCR和ELISA结果显示:不同浓度甘露聚糖刺激RECs8 h后,SBD-1 mRNA转录水平和蛋白的表达量均有所增加,但随着甘露聚糖浓度的增加,SBD-1表达量呈现先升高后降低的趋势,且当甘露聚糖浓度为50μg/mL时SBD-1表达量达到最大,并与对照组相比差异极显著(p<0.01)。当用50μg/mL的甘露聚糖分别刺激RECs不同时间后,SBD-1 mRNA转录水平和蛋白的表达量先是随着刺激时间的延长而呈上升趋势,刺激时间为4 h时SBD-1表达量达到峰值,且极显著高于对照组(p<0.01),之后呈下降趋势。结果表明甘露聚糖能够提高绵羊RECs SBD-1的表达,且甘露聚糖浓度为50μg/mL刺激RECs 4h时,SBD-1 mRNA转录水平和蛋白表达量达到最高。本研究为甘露聚糖在体内如何发挥先天性免疫提供了一种新解释,也为更好的开发利用甘露聚糖制剂提供实践指导。In order to explore the effect of mannan, a cell wall component of Saccharomyces cerevisiae, on the expression of sheep beta-defensin-1 (SBD-1) in ruminal epithelial cells (RECs) of sheep, the RECs were stimulated with different concentrations of mannan for 8hrs, and the transcript level of SBD-1 mRNA and the expression of SBD-1 protein in RECs was detected by real-time PCR (qPCR) and ELISA to determine the concentration of mannan that induced the highest expression of SBD-1. Then, RECs were stimulated for different periods of time with this concentration of mannan, and the transcript level of SBD-1 mRNA and the expression of SBD-1 protein were also detected by qPCR and ELISA, so as to determine the optimal stimulation time for mannan-induced SBD-1 expression. The results of qPCR and ELISA showed that the transcript level of SBD-1 mRNA and the expression of SBD-1 protein were increased after mannan stimulation for 8hrs, but the expression of SBD-1 were increased and then decreased following the increase of concentrations, the expression of SBD-1 was highest when the cells were stimulated with the concentration of 50μg/mL, and significantly higher than the control group (p<0.01). When RECs were stimulated with mannan at 50μg/mL for different periods of time, the transcript level of SBD-1 mRNA and the expression of SBD-1 protein were increased with the extension of stimulating time, which the expression of SBD-1 was highest at 4hrs, and it was significantly higher than the control group (p<0.01), then downward. Therefore, the results of this study indicated that mannan could improve the SBD-1 expression in RECs of ovine. And the transcript level of SBD-1 mRNA and the expression of SBD-1 protein were highest at the concentration of 50μg/mL and 4hrs stimulating. This study provides a new explanation for how mannan exerts innate immunity in vivo and provides practical guidance for the further development and utilization of mannan preparations.

关 键 词：甘露聚糖 绵羊瘤胃上皮细胞 β-防御素-1 荧光定量PCR ELISA 

分 类 号：S852.61[农业科学—基础兽医学]