肉用型和蛋用型鸡胚胎期皮下脂肪组织发育特征及FASN基因表达差异分析

Development characteristics of subcutaneous adipose tissue and differential expression analysis of FASN gene in the embryonic stage of broiler-type and layer-type chickens

为了探究爱拔益加(AA)肉鸡和海兰灰蛋鸡胚胎期皮下脂肪组织发育特征和差异,试验选取AA肉鸡和海兰灰蛋鸡种蛋各100枚进行孵化,将孵化24 h记为1胚龄(E1),从能分离到皮下脂肪组织时的胚龄开始至出壳当天(D0)选取6个时间点,每个时间点每个品种选取8枚发育正常的鸡胚,称量胚胎重(E20包含卵黄囊重量)/体重及皮下脂肪组织(主要包括颈部、胸部及腿部的皮下脂肪)重量,计算相对皮下脂肪组织含量;称量结束后随机从8枚鸡胚中选取3枚鸡胚,取其左侧后肢皮下脂肪组织制作石蜡切片进行H.E.染色,观察皮下脂肪组织细胞发育状态和测定皮下脂肪组织细胞平均面积,同时提取皮下脂肪组织RNA,检测不同时间点脂肪酸合成酶(FASN)基因的mRNA相对表达量。结果表明:AA肉鸡和海兰灰蛋鸡胚胎重/体重均随孵化时间增加而总体呈增加趋势,均在E20后增长趋势趋于稳定,且E20和D0间差异不显著(P>0.05),但均极显著高于E12、E14、E16、E18(P<0.01);不同时间点AA肉鸡的胚胎重/体重均高于海兰灰蛋鸡,其中在E12时差异不显著(P>0.05),E14时差异显著(P<0.05),E16、E18、E20和D0时差异极显著(P<0.01)。在落盘前(E12~E18),AA肉鸡和海兰灰蛋鸡皮下脂肪组织重量、相对皮下脂肪含量和皮下脂肪细胞平均面积均随胚龄增加而增加,且同一品种不同时间点间均差异极显著(P<0.01)。在落盘后(E18后),AA肉鸡和海兰灰蛋鸡不同时间点的皮下脂肪组织重量均差异不显著(P>0.05),但AA肉鸡皮下脂肪重量均极显著高于海兰灰肉鸡(P<0.01);AA肉鸡和海兰灰蛋鸡相对皮下脂肪含量均呈先降低后升高趋势;在E18时,海兰灰蛋鸡相对皮下脂肪含量极显著高于AA肉鸡(P<0.01);AA肉鸡和海兰灰蛋鸡皮下脂肪细胞平均面积变化逐渐趋于稳定,不同时间点间均差异不显著(P>0.05)。在E12~D0时,海兰灰蛋鸡皮下脂肪细胞平均面积均大于AA肉鸡,且除在E12时两品种间差异不显著(P>0.05)外,在E14~D0时两品种间均差异极显著(P<0.01)。FASN基因在AA肉鸡和海兰灰蛋鸡皮下脂肪组织中有着不同的表达模式,即在AA肉鸡中,不同时间点皮下脂肪组织中FASN基因mRNA表达量均差异不显著(P>0.05);而在海兰灰蛋鸡中,FASN基因在E12~E18时呈上升趋势,在E18时达到最高,之后呈下降趋势,其中E18时FASN基因mRNA表达量与E12、E14和E20时差异显著(P<0.05),与D0时差异极显著(P<0.01),与E16时差异不显著(P>0.05)。在E12~E20时,海兰灰蛋鸡皮下脂肪组织中FASN基因mRNA表达量均显著或极显著高于AA肉鸡(P<0.05或P<0.01);而在D0时,AA肉鸡FASN基因mRNA表达量高于海兰灰蛋鸡,但差异不显著(P>0.05)。说明鸡胚胎期皮下脂肪组织是晚期发育性状,且胚胎期海兰灰蛋鸡的皮脂沉积速度快于AA肉鸡。

In order to investigate the development characteristics and differences of subcutaneous adipose tissue of Arbor Acres(AA) broilers and Hy-Line grey laying hens in embryonic stage,100 eggs were selected from AA broilers and Hy-Lline grey laying hens for incubation,respectively.Incubation up to 24 h was denoted as 1 embryo age(E1).Six time points were selected from the beginning of embryo age when subcutaneous adipose tissue could be isolated to the day when the chicks were hatched(D0).Eight normal developing chicken embryos were randomly selected in each time points of each variety to weigh the embryo weight(yolk sac weight included at E20) or body weight and subcutaneous adipose tissue weight(mainly including subcutaneous fat in neck,chest and legs),which were then used to calculate the relative subcutaneous adipose tissue content.At the end of weighing,three embryos were randomly selected from the eight embryos,and the subcutaneous adipose tissue of the left hind limb was collected to make paraffin section for H.E.staining.The development status of subcutaneous adipose tissue cells was observed and the average area of subcutaneous adipose tissue cells was measured.Meanwhile,RNA from subcutaneous adipose tissue was extracted to detect FASN mRNA expression at different time points.The results showed that the embryo weight/body weight of AA broilers and Hy-Line grey laying hens increased with the increase of incubation time,and the increase trend gradually tended to be stable after E20.There was no significant difference in the increase trend between E20 and D0(P>0.05),but they were extremely significantly higher than that of E12,E14,E16 and E18(P<0.01).The embryo weight/body weight of AA broilers was higher than that of Hy-Line grey laying hens at different time points,but the difference was not significant at E12(P>0.05);the difference was significant at E14(P<0.05),and the difference was extremely significant at E16,E18,E20 and D0(P<0.01).The subcutaneous adipose tissue weight,relative content of subcutaneous fat and average area of subcutaneous adipocytes of AA broilers and Hy-Line grey laying hens increased with the increase of embryo age before setting(E12-E18),and the differences were significant among different time points of the same breed(P<0.05).There was no significant difference in the subcutaneous adipose tissue weight among different time points of the same breed for AA broilers and Hy-Line grey laying hens(P>0.05),but the subcutaneous adipose tissue weight of AA broilers was extremely significantly higher than that of Hy-Line grey laying hens after E18(P<0.01).The relative subcutaneous fat content of AA broilers and Hy-Line grey laying hens decreased first and then increased.At E18,the relative subcutaneous fat tissue content of Hy-Line grey laying hens was extremely significantly higher than that of AA broilers(P<0.01).The change of average area of subcutaneous adipocytes gradually stabilized for AA broilers and Hy-Line grey laying hens,and there was no significant difference among different time points of the same breed(P>0.05).At E12~D0,the average area of subcutaneous adipocytes of Hy-Line grey laying hens was larger than that of AA broilers,and the difference between the two breeds was extremely significant from E14 to D0(P<0.01),except at E12(P>0.05).FASN gene has different expression patterns in subcutaneous adipose tissue of AA broilers and Hy-line grey laying hens.There was no significant difference in the mRNA expression of FASN gene in the subcutaneous adipose tissue of AA broilers among different time points(P>0.05).In Hy-Line grey laying hens,FASN gene increased at E12-E18,reaching the highest at E18,and then decreased.The mRNA expression level of FASN gene at E18 was significantly different from that of E12,E14 and E20(P<0.05),and extremely significantly different from that of D0(P<0.01),but not significantly different from E16(P>0.05).The mRNA expression of FASN gene in the subcutaneous adipose tissue of Hy-Line grey laying hens was significantly or extremely significantly higher than that of AA broilers at E12-E20(P<0.05 or P<0.01).At D0,the mRNA expression of FASN gene in AA broilers was higher than that in Hy-Line grey laying hens,but the difference was not significant(P>0.05).The results indicated that the subcutaneous adipose tissue was a late-stage developmental trait in the embryonic stage of the chicken,and the sebum deposition rate of Hy-Line gray laying hens was faster than that of AA broilers.