【作者】 王二玲；

【导师】 周光宏；

【作者基本信息】 南京农业大学 ， 动物营养与饲料科学， 2009， 硕士

【摘要】 本文旨在研究视黄酸对β-胡萝卜素的反馈调控。首先研究了β-胡萝卜素15,15’-加单氧酶(βCMOOX)在肉仔鸡体内的组织特异性,在此基础上,系统研究了全反式视黄酸(ATRA)和13-顺式视黄酸(13-cis RA)对肉仔鸡不同组织内βCMOOX活性及其mRNA表达的影响,为进一步研究β-胡萝卜素代谢机制提供理论基础。肉仔鸡体内βCMOOX的组织特异性研究使用半定量RT-PCR的方法研究βCMOOX在三黄鸡不同组织内的表达,并用高效液相色谱法测定各组织中βCMOOX的活性。结果表明βCMOOX mRNA在正常鸡的心脏、肝脏、脾脏、肾脏、肠道、睾丸和骨骼肌组织中均有表达,且在空肠内表达水平最高(P<0.01),在肺中几乎没有表达；βCMOOX在各组织内(除肺脏外)均有活性,且空肠和十二指肠内活性最高(P<0.05)。这就显示βCMOOX在鸡体内的分布具有组织特异性。视黄酸对不同组织内βCMOOX活性及其mRNA表达的影响研究了ATRA和13-cis RA对肉仔鸡十二指肠、空肠、肝脏和肾脏内βCMOOX活性和βCMOOX mRNA表达的影响.试验将48只18日龄肉仔鸡随机分为6组：其中1组为正常日粮组,其它五组均饲喂低维生素A日粮。在56-59日龄,正常日粮组每只鸡每天灌服1ml大豆油,其余各组每只鸡每天分别灌服1ml大豆油、0.25μmol·L-1ATRA、1μmol·L-1ATRA、4μmol·L-1ATRA和4μmol·L-113-cis RA,视黄酸均溶于大豆油。结果表明,ATRA浓度为4μmol·L-1时,显著降低十二指肠(P<0.01)、空肠(P<0.01)和肾脏(P<0.05)内βCMOOX活性,抑制十二指肠(P<0.01)、空肠(P<0.01)和肾脏(P<0.01)内βCMOOX mRNA的表达,而对肝脏内βCMOOX活性和βCMOOX mRNA的表达无明显作用；相同浓度的13-cis RA作用不明显(P>0.05).与低维生素A对照组相比,0.25μmol·L-1、1μmol·L-1和4μmol·L-1 ATRA对肉仔鸡十二指肠、空肠和肾脏内βCMOOX活性和βCMOOX mRNA的表达均有显著抑制作用,且随着浓度的增高,抑制作用增强。这就说明βCMOOX可能在反转录时受到ATRA的调控,并且这种调控机制存在组织特异性。更多还原

【Abstract】 The aim of this study was to investigate the feedback regulation of metabolism ofβ-carotene by retinoic acid in different issues of broilers. The Issue-specific of P-carotene-15,15’-monooxygenase (PCMOOX) was firstly studied in broilers. Then, the effect ofβCMOOX activity and the expression of its mRNA by all-trans retinoic acid (ATRA) and 13-cis retinoic acid(13-cis RA) in different issue of broilers, to supply rational for the further study of the metabolic mechanism ofβ-carotene.In the first experiment, the expression of PCMOOX mRNA andβCMOOX activity in different tissues of chickens were studied, used the methods of Semi-quantitative RT-PCR and HPLC. The results indicated thatβCMOOX mRNA was expressed in tissues including heart, liver, spleen, lung, kidney, duodenum, jejunum, ileum, muscle and testis, not expressed in lung; Furthermore, it was expressed in jejunum with the highest level. The PCMOOX activity was found in every issue, except lung, and the highest level in duodenum and jejunum. This showed that the distribution ofβCMOOX in chicken has tissue specificity.The second experiment was conducted to study the regulation ofβCMOOX activity and the expression of its mRNA by ATRA and 13-cis RA in different issues of broilers. Forty eight 18-day-old broilers were divided into six groups, one group was feed normal diet, and the others were feed the diet contained low vitamin A. On 56-59 day old, the normal-diet group was drenched 1 mL soybean oil per broiler per day, the other groups were differently drenched 1 mL soybean oil,0.25μmol·L-1 ATRA,1μmol·L-1 ATRA,4μmol·L-1 ATRA and 4μmol·L-1 13-cis RA,ATRA and 13-cis RA were dissolved in soybean oil. The results indicated that 4μmol·L-1 ATRA significantly decreased theβCMOOX activity of duodenum (P<0.01),jejunum (P<0.01)and kidney (P<0.05), and suppressed the expression of PCMOOX mRNA of duodenum (P<0.01),jejunum (P<0.01)and kidney (P<0.01),but not affected that of liver; 4μmol·L-1 13-cis retinoic acid did not affect theβCMOOX activity and the expression ofβCMOOX mRNA of duodenum, jejunum, liver and kidney. Compared to the control group,0.25μmol·L-1,1μmol·L-1 and 4μmol·L-1 ATRA all significantly decreased PCMOOX activity and suppressed rthe expression ofβCMOOX mRNA of the duodenum, jejunum and kidney of broilers, the inhibition was enhanced with the concentration raised up, as well. This showed that the regulation ofβCMOOX might take place at the transcriptional level, moreover the regulation ofβCMOOX gene expression by RA showed differences between several tissues.更多还原