LPS对鹅等级卵泡基质层TLR家族基因表达的影响  

Time Course Effect of Lipopolysaccharide on Toll-Like Receptors Expression in the Goose Follicular Stroma

作　　者：应诗家[1] 戴子淳 郭佳佳[1] 施振旦[1] 

机构地区：[1]江苏省农业科学院畜牧研究所动物品种改良和繁育重点实验室,南京210014

出　　处：《中国农业科学》2017年第6期1147-1156,共10页Scientia Agricultura Sinica

基　　金：江苏省自然科学基金(BK20130718);国家水禽产业技术体系(CARS-43-16)

摘　　要：【目的】研究鹅卵泡基质层TLR家族基因表达谱及其对LPS不同处理时间后的反应性。【方法】在大群散养条件下,实时观察鹅产蛋过程,分别在产蛋后8和2 h以及产前4、16和28 h注射LPS,并在产蛋后8h屠宰,即LPS作用0、6、12、24和36 h,每个时间点各5只鹅。屠宰时,取卵巢,观察卵泡外观形态,并分离F1-F5级卵泡基质层。试验鹅自由饮水、自由采食、自然光照。LPS处理0、24和36 h的单个F1-F5级卵泡基质层或卵泡用于基因表达分析,而其他LPS处理的每只鹅等级卵泡基质层RNA等质量混合后用于基因表达分析。采用普通PCR方法检测10种鸟类TLR家族基因的在鹅等级卵泡基质层的表达谱,其中以脾脏组织为阳性对照,以不加c DNA模板为阴性对照。根据RT-PCR的表达谱结果,采用Real-time PCR检测不同等级卵泡基质层TLRs表达水平以及不同LPS处理时间对基质层TLRs表达水平的影响。对不同等级卵泡和不同LPS处理时间对基质层TLRs表达的数据,采用单因子方差分析,Duncan法多重比较;对变性卵泡与对照组基质层TLRs表达水平的数据,采用T检验分析。【结果】(1)已公开的10种鸟类TLR家族基因,如TLRs 1A、1B、2A、2B、3、4、5、7、15和21基因均在等级卵泡基质层组织中表达。(2)随等级卵泡生长,TLRs 2A和15表达水平逐渐升高;F1级卵泡基质层TLR2A表达水平显著高于F3、F4和F5级卵泡,TLR15表达水平显著高于F5级卵泡;其他TLR家族基因,如TLRs 1A、1B、2B、3、4、5、7和21在不同等级卵泡基质层中的表达水平差异不显著。(3)在LPS处理0、6和12 h时,等级卵泡外观形态无明显变化,但在LPS处理24 h时,3只鹅的等级卵泡呈深黄色胶冻状变性;在LPS处理36 h时,全部试验鹅等级卵泡呈深黄色胶冻状变性。(4)与对照组(LPS处理0 h)相比,TLRs 2A、4和5表达水平在LPS处理12和24 h时显著升高,TLR2B表达水平在LPS处理24 h时显著升高,TLRs 7和15表达水平在LPS�【Objective】 This study aims to investigate the transcriptional profiling of Toll-like receptors（TLRs） and their responses to lipopolysaccharide with different treatment times in the goose follicular stroma. 【Method】 The laying process was monitored in a flock of Yangzhou geese. The geese were injected intravenously with LPS（1.5mg/kg body weight） 8 and 2 h after oviposition, and 4, 16 and 28 h before oviposition. All experimental geese were slaughtered 8 h after oviposition. Therefore, time course of LPS was achieved, namely at 0, 6, 12, 24 and 36 h after injection of LPS. Five geese at each time point were selected. After slaughtering, the ovaries were collected, the follicle morphology was observed, and the stroma of the first largest follicles（F1）, F2, F3, F4 and F5 was isolated. The animals were provided with feed and water ad libitum, and maintained under natural photoperiod. For the geese 0, 24 and 36 h of after LPS treatment, all samples were used for gene expression analysis. For the other geese, RNA samples of follicular stroma of each goose were mixed with equal concentrations for gene expression analysis. RT-PCR was performed to examine the transcriptional profiling of 10 types of avian TLRs in follicular stroma. The spleen tissue was used as the positive control, and the sample without c DNA sample was used as the negative control. The expression levels of TLRs in follicular stroma among different hierarchical follicles and among different time points were tested using Real-time PCR. For data on the time course of LPS and different hierarchical follicles, the significance of differences was analyzed using the one-way ANOVA followed by Duncan＇s multiple range tests. For data between stroma in control group and DF, statistical analysis was carried out using independent-samples T test. 【Result】 All 10 reported TLRs in poultry, namely TLRs 1A, 1B, 2A, 2B, 3, 4, 5, 7, 15 and 21 were expressed in goose stroma of hierarchical follicles. The expression level of TLR2 A exhibited a tend

关 键 词：种鹅 等级卵泡 内毒素脂多糖(LPS) TLRS 

分 类 号：S835[农业科学—畜牧学]