饲粮脂肪酸ω-6/ω-3对鹅脂肪代谢影响及其分子机制的研究
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摘要
随着生活水平的提高,人们越来越关注衰老、肥胖等健康问题。食物的脂肪含量及其脂肪酸组成与健康密切相关。其中-6系列和ω-3系列多不饱和脂肪酸(PUFA)的平衡在稳定细胞膜结构、防止肥胖、抗氧化、提高免疫力等方面具有至关重要的作用。世界卫生组织(WHO)和联合国粮食与农业组织(FAO)提出膳食中脂肪酸-6/ω-3的比例为(5～10)：1,但迄今为止,其适宜的比例尚不明了。
本试验以扬州鹅为试验动物,研究饲粮脂肪酸ω-6/ω-3比例对其产肉性能、脂肪沉积及脂肪酸组成、抗氧化以及免疫功能的影响；同时试验利用荧光定量PCR和基因原位杂交等方法检测脂肪代谢相关基因的表达水平,旨在阐明ω-6/ω-3影响脂肪代谢的机制,从而为生产中扬州鹅的科学饲养和油脂饲料的合理使用提供一些科学指导,也为理想脂肪酸比例模式的深入研究提供一些基础资料。试验分为四个部分进行。
试验一研究了饲粮脂肪酸ω-6/ω-3不同比例对扬州鹅脂肪沉积、血脂代谢和肌肉脂肪酸组成的影响。结果表明：ω-6/ω-3为9：1组时胸、腿肌率均显著高于其他比例,6：1的腹脂率在生长快速期是显著低于其他比例的；ω-6/ω-3对扬州鹅的血脂代谢有一定的影响,其比例为6：1时有较好的降低血脂的效应；鹅肌肉中ω-6/ω-3随饲粮中该比例的下降而下降,并以3：1组最低。以上研究结果表明：适宜比例的ω-6/ω-3(6：1、3：1)可以降低扬州鹅早期腹脂水平,对血脂和血糖水平有一定的改善作用,能够促进胸肌率和腿肌率的提高,从而提高扬州鹅的产肉性能。
试验二研究了饲粮脂肪酸ω-6/ω-3不同比例对扬州鹅脂肪代谢相关酶活性和基因表达的影响。结果表明：ω-6/ω-3为3：1和6：1的低比例饲粮处理组的脂肪酸合成相关的乙酰辅酶A羧化酶(ACC)、苹果酸酶(ME)和脂肪酸合成酶(FAS)的酶活性较低,与脂肪周转代谢相关的肝酯酶(HL)、脂蛋白酯酶(LPL)的酶活性较高；高ω-6/ω-3饲粮处理组的脂肪合成相关基因FAS、ME、ACC表达相对较高。另外,低ω-6/ω-3饲粮处理组的高密度脂蛋白受体(HDL-R)基因表达强度高于高ω-6/ω-3饲粮处理组；而高ω-6/ω-3饲粮处理组的低密度脂蛋白受体(LDL-R)基因表达强度高于低ω-6/ω-3比例饲粮处理组。本试验表明低ω-6/ω-3比例饲粮能够在一定程度上抑制脂肪合成代谢；
试验三研究了饲粮脂肪酸ω-6/ω-3不同比例对扬州鹅抗氧化性能的影响。试验通过4个梯度ω-6/ω-3饲粮的饲养试验,研究ω-6/ω-3对肝脏过氧化脂质、抗氧化酶和肝细胞超微结构的影响。研究结果表明：饲粮ω-6/ω-3对扬州鹅肝脏中的自由基和过氧化脂质有一定的影响,其比例为3：1时对扬州鹅肝脏组织中的丙二醛(MDA)、一氧化氮(NO)、过氧化氢(H2O2)和游离脂肪酸(NEFA)活性有较好的抑制作用；饲粮ω-6/ω-3对扬州鹅肝脏抗氧化酶活性有一定的影响,其比例为3：1时对扬州鹅肝脏总超氧化物歧化酶(T-SOD),谷胱甘肽过氧化物酶(GSH-Px)和过氧化氢酶(CAT)的活性有较好的提高效应；饲粮ω-6/ω-3对鹅肝亚细胞显微结构有一定影响,且较高ω-6/ω-3饲粮组的扬州鹅肝细胞核仁、线粒体膜系结构变化较大。
试验四研究了饲粮脂肪酸ω-6/ω-3不同比例对扬州鹅免疫机能的影响。研究结果表明：不同ω-3/ω-6饲粮对扬州鹅免疫器官的的生长发育影响有所不同,并且其影响还与扬州鹅的日龄有关,其中ω-6/ω-3在6：1时可促进42日龄前扬州鹅免疫器官的发育,而3：1的比例对42日龄后免疫器官的影响较为明显；在56和70日龄时,含低ω-6/ω-3的饲粮能够增加扬州鹅血清中白蛋白/球蛋白比例和免疫球蛋白(包括IgA、IgG和IgM)水平；42-70日龄生长期扬州鹅随日龄增长,其外周血异嗜性粒细胞/淋巴细胞呈下降趋势,单核细胞呈上升趋势,而嗜酸、碱性粒细胞则没有明显的变化规律；饲粮ω-6/ω-3影响42-70日龄扬州鹅外周血白细胞亚群组成。较低的比例能够降低异嗜性细胞：淋巴细胞(H：L)比值；降低嗜酸、碱性粒细胞在白细胞中的百分比,提高单核细胞百分比。
With the improvement of living standards, people are increasingly concerned about senescence, obesity and other health problems. The food's fat content and fatty acid composition are closely related to health. The balances between ω-6series and ω-3series polyunsaturated fatty acids (PUFA) paly a crucial role in stabilizing the cell membranes structure, preventing obesity,antioxidant and improving immunity. WHO(World Health Organization) and FAO(Food and Agriculture Organization) suggest that the proportion of the ω-6/ω-3fatty acids in diet is (5～10):1.While so far, the appropriate ratio is not clear.
In order to clarify the mechanism of fat metabolism by the ω-6/ω-3, provide some scientific guidance for the scientific breeding of Yangzhou geese and the rational using of oil feed in the production and offer some basic information to the thorough study of the ideal fatty acid ratio model.In this thesis, the fat deposition,oxidation resistance, immune function were compared among different ratios of ω-6/ω-3fatty acid of diet in Yangzhou goose. And the fat metabolism-related gene mRNA expression was detected by RT-qPCR and in situ_hybridization, its results as follows:
Firstly, the fat deposition, lipid metabolism and muscle fatty acid composition were analyzed among different ratios of ω-6/ω-3fatty acid of diet in Yangzhou goose. The results showed that the percentage of breast and leg muscle was significantly higher than that of others when the ω-6/ω-3was9:1in diet, and the percentage of abdominal fat serum lipid was significantly lower when the ω-6/ω-3was6:1. Besides, the ratio of ω-6/co-3in muscle changed with that in diet. In summary, appropriate proportions of co-6/co-3(6:1,3:1) can reduce the early level of abdominal fat of Yangzhou goose, do a certain improvement in the lipids and blood glucose levels and contribute to increase the rate of breast muscle and leg muscle in Yangzhou goose.
Secondly, the fat metabolism-related gene mRNA expression was detected by RT-qPCR and in situ hybridization. The results showed that the enzyme activity related to fatty acid synthesis such as Acetyl-coA Carboxylase(ACC)、Malic Enzyme(ME)and Fatty Acid Synthase(FAS) activity was lower when the ratio of ω-6/ω-3, while the enzyme activity related to fat metabolism turnover such as Liver Esterase(HL) and Lipoprotein Lipase(LPL) activity was higher. The genes expression related to synthcticfat such as FAS,ME,ACC was higher when the ratio of ω-6/ω-3was high in diet. In addition, the gene expression intensity of High Density Lipoprotein Receptor(HDL-R) in the treatment with a lower proportion of dietary ω-6/ω-3was higher than with a higher, while the gene expression intensity of Ligh Density Lipoprotein Receptor(LDL-R) in the treatment with a higher proportion of dietary ω-6/ω-3was higher than with a lower. In summary, low proportion of ω-6/ω-3in diet,to a certain extent, inhibited fat synthesis and metabolism.
Thirdly, antioxidant enzymes and liver cell ultrastructure was studied among different ratio of dietary ω-6/ω-3in diet. The results showed that Malondialdehyde(MDA), Nitrogen(NO),Hydrogen peroxide(H2O2) and Nonesterified fatty acid(NEFA) activities were inhibited in diet (ω-6/ω-3=3:1), while Total Superoxide Dismutase(T-SOD), Glutathiome Peroxidase(GSH-Px) and Catalase(CAT) activities enhanced in diet (ω-6/ω-3=3:1). The sub-cellular microstructure was detected also among different ratio of dietary ω-6/ω-3in diet. The results showed that nucleolar, mitochondrial membrane system of hepatocyte was different injured, and minimal injury in lower ω-6/ω-3diet, the bigger the change of nucleolus, mitochondrial membrane system in Yangzhou goose.
Finally, the immune function and immune organ development was studied among different ratio of dietary ω-6/ω-3in diet. It resulted that the immune organ development was promoted in diet (ω-6/ω-3=6:1) before42days old in Yangzhou goose. The serum albumin/globulin ratio, and immunoglobulin content (including IgA, IgG and IgM) was increased at the age of56and70days. With the grown of age, the ratio of peripheral blood different tropism of granulocyte/lymphocyte declined, monocytes had an upward trend, while there is no significant variation in acidophilus and alkaline granulocyte. The constitution of peripheral blood leukocyte subsets in42to70days old Yangzhou goose was effected by the dietary ratios of ω-6/ω-3. A lower proportion may reduce the ratio of H/L and the percentage of acidophilic granulocyte and basophilic granulocyte in the white blood cells, while increase the percentage of monocytes.
本试验以扬州鹅为试验动物,研究饲粮脂肪酸ω-6/ω-3比例对其产肉性能、脂肪沉积及脂肪酸组成、抗氧化以及免疫功能的影响；同时试验利用荧光定量PCR和基因原位杂交等方法检测脂肪代谢相关基因的表达水平,旨在阐明ω-6/ω-3影响脂肪代谢的机制,从而为生产中扬州鹅的科学饲养和油脂饲料的合理使用提供一些科学指导,也为理想脂肪酸比例模式的深入研究提供一些基础资料。试验分为四个部分进行。
试验一研究了饲粮脂肪酸ω-6/ω-3不同比例对扬州鹅脂肪沉积、血脂代谢和肌肉脂肪酸组成的影响。结果表明：ω-6/ω-3为9：1组时胸、腿肌率均显著高于其他比例,6：1的腹脂率在生长快速期是显著低于其他比例的；ω-6/ω-3对扬州鹅的血脂代谢有一定的影响,其比例为6：1时有较好的降低血脂的效应；鹅肌肉中ω-6/ω-3随饲粮中该比例的下降而下降,并以3：1组最低。以上研究结果表明：适宜比例的ω-6/ω-3(6：1、3：1)可以降低扬州鹅早期腹脂水平,对血脂和血糖水平有一定的改善作用,能够促进胸肌率和腿肌率的提高,从而提高扬州鹅的产肉性能。
试验二研究了饲粮脂肪酸ω-6/ω-3不同比例对扬州鹅脂肪代谢相关酶活性和基因表达的影响。结果表明：ω-6/ω-3为3：1和6：1的低比例饲粮处理组的脂肪酸合成相关的乙酰辅酶A羧化酶(ACC)、苹果酸酶(ME)和脂肪酸合成酶(FAS)的酶活性较低,与脂肪周转代谢相关的肝酯酶(HL)、脂蛋白酯酶(LPL)的酶活性较高；高ω-6/ω-3饲粮处理组的脂肪合成相关基因FAS、ME、ACC表达相对较高。另外,低ω-6/ω-3饲粮处理组的高密度脂蛋白受体(HDL-R)基因表达强度高于高ω-6/ω-3饲粮处理组；而高ω-6/ω-3饲粮处理组的低密度脂蛋白受体(LDL-R)基因表达强度高于低ω-6/ω-3比例饲粮处理组。本试验表明低ω-6/ω-3比例饲粮能够在一定程度上抑制脂肪合成代谢；
试验三研究了饲粮脂肪酸ω-6/ω-3不同比例对扬州鹅抗氧化性能的影响。试验通过4个梯度ω-6/ω-3饲粮的饲养试验,研究ω-6/ω-3对肝脏过氧化脂质、抗氧化酶和肝细胞超微结构的影响。研究结果表明：饲粮ω-6/ω-3对扬州鹅肝脏中的自由基和过氧化脂质有一定的影响,其比例为3：1时对扬州鹅肝脏组织中的丙二醛(MDA)、一氧化氮(NO)、过氧化氢(H2O2)和游离脂肪酸(NEFA)活性有较好的抑制作用；饲粮ω-6/ω-3对扬州鹅肝脏抗氧化酶活性有一定的影响,其比例为3：1时对扬州鹅肝脏总超氧化物歧化酶(T-SOD),谷胱甘肽过氧化物酶(GSH-Px)和过氧化氢酶(CAT)的活性有较好的提高效应；饲粮ω-6/ω-3对鹅肝亚细胞显微结构有一定影响,且较高ω-6/ω-3饲粮组的扬州鹅肝细胞核仁、线粒体膜系结构变化较大。
试验四研究了饲粮脂肪酸ω-6/ω-3不同比例对扬州鹅免疫机能的影响。研究结果表明：不同ω-3/ω-6饲粮对扬州鹅免疫器官的的生长发育影响有所不同,并且其影响还与扬州鹅的日龄有关,其中ω-6/ω-3在6：1时可促进42日龄前扬州鹅免疫器官的发育,而3：1的比例对42日龄后免疫器官的影响较为明显；在56和70日龄时,含低ω-6/ω-3的饲粮能够增加扬州鹅血清中白蛋白/球蛋白比例和免疫球蛋白(包括IgA、IgG和IgM)水平；42-70日龄生长期扬州鹅随日龄增长,其外周血异嗜性粒细胞/淋巴细胞呈下降趋势,单核细胞呈上升趋势,而嗜酸、碱性粒细胞则没有明显的变化规律；饲粮ω-6/ω-3影响42-70日龄扬州鹅外周血白细胞亚群组成。较低的比例能够降低异嗜性细胞：淋巴细胞(H：L)比值；降低嗜酸、碱性粒细胞在白细胞中的百分比,提高单核细胞百分比。
With the improvement of living standards, people are increasingly concerned about senescence, obesity and other health problems. The food's fat content and fatty acid composition are closely related to health. The balances between ω-6series and ω-3series polyunsaturated fatty acids (PUFA) paly a crucial role in stabilizing the cell membranes structure, preventing obesity,antioxidant and improving immunity. WHO(World Health Organization) and FAO(Food and Agriculture Organization) suggest that the proportion of the ω-6/ω-3fatty acids in diet is (5～10):1.While so far, the appropriate ratio is not clear.
In order to clarify the mechanism of fat metabolism by the ω-6/ω-3, provide some scientific guidance for the scientific breeding of Yangzhou geese and the rational using of oil feed in the production and offer some basic information to the thorough study of the ideal fatty acid ratio model.In this thesis, the fat deposition,oxidation resistance, immune function were compared among different ratios of ω-6/ω-3fatty acid of diet in Yangzhou goose. And the fat metabolism-related gene mRNA expression was detected by RT-qPCR and in situ_hybridization, its results as follows:
Firstly, the fat deposition, lipid metabolism and muscle fatty acid composition were analyzed among different ratios of ω-6/ω-3fatty acid of diet in Yangzhou goose. The results showed that the percentage of breast and leg muscle was significantly higher than that of others when the ω-6/ω-3was9:1in diet, and the percentage of abdominal fat serum lipid was significantly lower when the ω-6/ω-3was6:1. Besides, the ratio of ω-6/co-3in muscle changed with that in diet. In summary, appropriate proportions of co-6/co-3(6:1,3:1) can reduce the early level of abdominal fat of Yangzhou goose, do a certain improvement in the lipids and blood glucose levels and contribute to increase the rate of breast muscle and leg muscle in Yangzhou goose.
Secondly, the fat metabolism-related gene mRNA expression was detected by RT-qPCR and in situ hybridization. The results showed that the enzyme activity related to fatty acid synthesis such as Acetyl-coA Carboxylase(ACC)、Malic Enzyme(ME)and Fatty Acid Synthase(FAS) activity was lower when the ratio of ω-6/ω-3, while the enzyme activity related to fat metabolism turnover such as Liver Esterase(HL) and Lipoprotein Lipase(LPL) activity was higher. The genes expression related to synthcticfat such as FAS,ME,ACC was higher when the ratio of ω-6/ω-3was high in diet. In addition, the gene expression intensity of High Density Lipoprotein Receptor(HDL-R) in the treatment with a lower proportion of dietary ω-6/ω-3was higher than with a higher, while the gene expression intensity of Ligh Density Lipoprotein Receptor(LDL-R) in the treatment with a higher proportion of dietary ω-6/ω-3was higher than with a lower. In summary, low proportion of ω-6/ω-3in diet,to a certain extent, inhibited fat synthesis and metabolism.
Thirdly, antioxidant enzymes and liver cell ultrastructure was studied among different ratio of dietary ω-6/ω-3in diet. The results showed that Malondialdehyde(MDA), Nitrogen(NO),Hydrogen peroxide(H2O2) and Nonesterified fatty acid(NEFA) activities were inhibited in diet (ω-6/ω-3=3:1), while Total Superoxide Dismutase(T-SOD), Glutathiome Peroxidase(GSH-Px) and Catalase(CAT) activities enhanced in diet (ω-6/ω-3=3:1). The sub-cellular microstructure was detected also among different ratio of dietary ω-6/ω-3in diet. The results showed that nucleolar, mitochondrial membrane system of hepatocyte was different injured, and minimal injury in lower ω-6/ω-3diet, the bigger the change of nucleolus, mitochondrial membrane system in Yangzhou goose.
Finally, the immune function and immune organ development was studied among different ratio of dietary ω-6/ω-3in diet. It resulted that the immune organ development was promoted in diet (ω-6/ω-3=6:1) before42days old in Yangzhou goose. The serum albumin/globulin ratio, and immunoglobulin content (including IgA, IgG and IgM) was increased at the age of56and70days. With the grown of age, the ratio of peripheral blood different tropism of granulocyte/lymphocyte declined, monocytes had an upward trend, while there is no significant variation in acidophilus and alkaline granulocyte. The constitution of peripheral blood leukocyte subsets in42to70days old Yangzhou goose was effected by the dietary ratios of ω-6/ω-3. A lower proportion may reduce the ratio of H/L and the percentage of acidophilic granulocyte and basophilic granulocyte in the white blood cells, while increase the percentage of monocytes.
引文
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