日粮因素对脂肪酸合成酶(FAS)基因表达的影响

动物体脂的控制是一个复杂过程。不同种类不同年龄的动物体脂水平有很大差异,控制机制是什么,哪些是关键的控制因素便成为关注的焦点。近年对动物FAS的调节控制的研究颇受重视,也取得了一些进展。早在20世纪90年代初期,国外部分学者就在不同种类的动物(大鼠、兔、鸡)上证实了动     

Sunflower-seed oil,rapidly-degradable starch,and adiposity up-regulate leptin gene expression in lactating goats

We conducted experiments to evaluate the effects of lipid supplementation and the nature of starchy concentrate on the regulation of leptin synthesis in lactating goats. Multiparous goats in mid- to late lactation received diets based on different forages and containing plant oil or seeds rich in either 18:1c9, 18:2n-6 or 18:3n-3 corresponding to 3%–7% dry matter (DM) as lipid supplements, or diets based on concentrate as either rapidly or slowly degradable starch. The isoenergetic replacement of a part of the concentrate by either oleic sunflower-seed oil, formaldehyde-treated linseeds, or linseed oil did not modify leptinemia and the leptin mRNA concentration in adipose tissues, suggesting a lack of effect of 18:1c9, 18:3n-3, or their biohydrogenation products. Conversely, leptinemia and the leptin mRNA abundance were increased (by 20% and 140%, respectively, P < 0.05) in goats fed sunflower-seed oil under a grassland hay-based diet but not a maize silage-based diet, at similar energy intakes and adiposity. Thus, 18:2n-6 per se may up-regulate leptin gene expression, but the effect could be blunted by other fatty acids formed during the ruminal digestion of sunflower-seed oil when combined with maize silage. Consumption of rapidly but not slowly degradable starch increased (by 17%, P < 0.05) leptinemia. Moreover, during lactation, plasma leptin was positively correlated (P < 0.05) to adiposity parameters and negatively correlated to fiber intake. The results suggest that leptinemia responds poorly to nutritional factors in lactating goats, thus highlighting the physiological need to sustain hypoleptinemia during lactation.     

Hormonal regulation of leptin receptor expression in primary cultures of porcine hepatocytes

A study was conducted to elucidate hormonal control of leptin receptor gene expression in primary cultures of porcine hepatocytes. Hepatocytes were isolated from pigs (52 kg) and seeded into collagen-coated T-25 flasks. Monolayer cultures were established in medium containing fetal bovine serum for 1 day and switched to a serum-free medium for the remainder of the 3-day culture period. To establish basal conditions hepatocytes were maintained in serum-free William's E medium containing 10 nM dexamethasone and 1 ng/ml insulin. For the final 24 h, insulin (1 or 100 ng/ml) or glucagon (100 ng/ml), were added in the presence or absence of 100 nM triiodothyronine (T3). RNA was extracted and quantitative RT-PCR was performed with primers specific for the long form and total porcine leptin receptors. Leptin receptor expression was calculated relative to co-amplified 18S rRNA. Expression of the long form of the leptin receptor was confirmed under basal conditions. Insulin, glucagon and synthetic human proteins (ghrelin and GLP-1) at 100 ng/ml had no influence on leptin receptor expression; the addition of T3 was associated with a marked increase (P < 0.001) in expression of total and long forms of the leptin receptor by 1.6 and 2.4-fold, respectively. Addition of leptin to cells which were pre-treated with T3 for 24 h (to up-regulate leptin receptor expression), confirmed the lack of a direct effect of leptin on glucagon-induced glycogen turnover and cAMP production. These data suggest that porcine hepatocytes may be insensitive to leptin stimulation even when leptin receptor expression is enhanced by T3.     

鸡肝脏型脂肪酸结合蛋白基因启动子活性分析

PCR扩增鸡L-FABP基因5′侧翼区约2kb的DNA片段,进行克隆并测序,构建了鸡L-FABP基因报告基因系列缺失载体,瞬时转染进入人肝癌细胞系,利用双荧光素酶报告基因系统测定了荧光素酶活性。在线分析软件发现鸡L-FABP基因启动子区存在HNF-1、SREBP-1、AP-1、C/EBP、Oct-1、TATA、CCAAT、GATA-1等调控元件,没有发现CpG岛。报告基因结果表明鸡L-FABP基因启动子-2 076bp/-20bp区域具有最强的启动子活性,-522bp/-20bp区域启动子活性最弱;C/EBPα可以显著的抑制鸡L-FABP基因的表达,这些结果为深入研究鸡L-FABP的表达调控机制奠定了基础。     

鸡EX-FABP基因PCR-SSCP分析

以AA白鸡、广西鸡、E1矮小鸡品系为试验材料,采用PCR-SSCP技术对鸡细胞外脂肪酸结合蛋白基因(EX-FABP)的2个片段进行PCR扩增,分析了EX-FABP基因在3个鸡品系的多态性。结果表明:EX-FABP基因在2对引物扩增片段中均存在PCR-SSCP多态性。对于引物1扩增片段,3个鸡品系均检测到BB基因型,AA基因型只出现在AA白鸡中;而且B等位基因频率在3个鸡品系中均明显高于A等位基因频率。对于引物2扩增片段,3个鸡品系均检测到HH和Hh基因型,hh基因型只出现在E1矮小鸡中;H等位基因频率在3个品系中均明显高于h等位基因频率。引物1和引物2的多态性片段测序分析表明,EX-FABP基因第641位点和645位点分别发生了单碱基的转换(G-A和T-C),第3264位点发生了转换(T-C)。     

In ovo leptin administration affects hepatic lipid metabolism and microRNA expression in newly hatched broiler chickens

Background: A leptin-like immunoreactive substance has been found in chicken eggs and has been implicated in serving as a maternal signal to program offspring growth and metabolism. In the present study, we investigated the effects of in ovo leptin administration on hatch weight, serum and hepatic concentrations of metabolites and hormones, as well as on the expression of genes involved in hepatic lipid metabolism and the predicted microRNAs (miRNAs) targeting the affected genes. To this end we injected fertile eggs with either 0.5 μg of recombinant murine leptin or vehicle (PBS) before incubation. Results: Prenatally leptin-exposed chicks showed lower hatch weight, but higher liver weight relative to the body weight, compared to the control group. In ovo leptin treatment increased the hepatic content and serum concentration of leptin in newly hatched chickens. The hepatic contents of triglycerides (TG) and total cholesterol (Tch) were decreased, whereas the serum levels of TG, Tch and apolipoprotein B (ApoB) were increased. The hepatic mRNA expression of sterol regulator element binding protein 1 (SREBP-1c), SREBP-2, hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) and cholesterol 7α-hydroxylase 1 (CYP7A1) was significantly up-regulated, as was the protein content of both SREBP-1c and SREBP-2 in hepatic nuclear extracts of leptin-treated chickens. Moreover, out of 12 miRNAs targeting SREBP-1c and/or HMGCR, five were significantly up-regulated in liver of leptin-treated chicks, including gga-miR-200b and gga-miR-429, which target both SREBP-1c and HMGCR. Conclusions: These results suggest that leptin in ovo decreases hatch weight, and modifies hepatic leptin secretion and lipid metabolism in newly hatched broiler chickens, possibly via microRNA-mediated gene regulation.     

脂肪酸对基因表达的影响及调控

脂肪酸是一类重要的营养物质,是动物体内的能量来源,同时也是生物膜的组成成分,在细胞生化过程中也同样起着重要作用。研究发现,脂肪酸可通过对基因表达的影响,对代谢、生长发育以及细胞分化发挥重要的调控作用。真核生物基因表达的调控大致可分为转录前、转录、转录后、翻译和翻译后等5个阶段的调控。脂肪酸通过细胞膜受体信号途径和转录因子活化途径调节基因表达,而多不饱和脂肪酸主要从基因转录和mRNA的稳定性两个方面调节基因表达。文章综述了脂肪酸,尤其是多不饱和脂肪酸对基因表达的影响及调控机制。     

In ovo leptin administration affects hepatic lipid metabolism and microRNA expression in newly hatched broiler chickens

Background

A leptin-like immunoreactive substance has been found in chicken eggs and has been implicated in serving as a maternal signal to program offspring growth and metabolism. In the present study, we investigated the effects of in ovo leptin administration on hatch weight, serum and hepatic concentrations of metabolites and hormones, as well as on the expression of genes involved in hepatic lipid metabolism and the predicted microRNAs (miRNAs) targeting the affected genes. To this end we injected fertile eggs with either 0.5 μg of recombinant murine leptin or vehicle (PBS) before incubation.

Results

Prenatally leptin-exposed chicks showed lower hatch weight, but higher liver weight relative to the body weight, compared to the control group. In ovo leptin treatment increased the hepatic content and serum concentration of leptin in newly hatched chickens. The hepatic contents of triglycerides (TG) and total cholesterol (Tch) were decreased, whereas the serum levels of TG, Tch and apolipoprotein B (ApoB) were increased. The hepatic mRNA expression of sterol regulator element binding protein 1 (SREBP-1c), SREBP-2, hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) and cholesterol 7α-hydroxylase 1 (CYP7A1) was significantly up-regulated, as was the protein content of both SREBP-1c and SREBP-2 in hepatic nuclear extracts of leptin-treated chickens. Moreover, out of 12 miRNAs targeting SREBP-1c and/or HMGCR, five were significantly up-regulated in liver of leptin-treated chicks, including gga-miR-200b and gga-miR-429, which target both SREBP-1c and HMGCR.

Conclusions

These results suggest that leptin in ovo decreases hatch weight, and modifies hepatic leptin secretion and lipid metabolism in newly hatched broiler chickens, possibly via microRNA-mediated gene regulation.     

Sexual maturation in hens is not associated with increases in serum leptin and the expression of leptin receptor mRNA in hypothalamus

Background

In mammals, leptin is an attractive candidate for mediating the metabolic signal and the reproductive function via the specific receptor in hypothalamus. However, till now, the role of leptin on reproduction in birds is less well established. This experiment was conducted to elucidate the role of leptin on the onset of reproduction in bird, as a first step, to detect the changes of peripheral leptin and leptin receptor mRNA expression in hypothalamus between mature and immature hens at the same age. 120 ISA brown pullets at D60 were allocated randomly into two groups, long light (LL) group being raised under artificial light regimes with incrementally increased light phase (from 8 L:16D to 14 L:12D) and short light (SL) group raised on consistent light (8 L:16D) for 12 wk.

Results

The results showed that pullets in LL group reached sexual maturation 15 d earlier than those in SL group. Serum E₂ showed a significant increase with age, but no difference was observed between two groups. Serum leptin concentration decreased significantly from D112 to D136 in LL, and was markedly higher in LL group than that in SL at D112, while there was no significant difference between two groups at D136. Leptin receptor and GnRH-I mRNA expression in hypothalamus were significantly increased with age, yet there was no significant difference between SL and LL chickens at the same age. The expression of FSH-β and LH-β mRNA in pituitary was increased with age but did not show significant difference between LL and SL group. GnRH-I mRNA expression was very rich in pineal gland, and decreased from D112 to D136 in LL but not in SL group, and there was no difference between two groups at the same age.

Conclusions

These results indicate that the earlier sexual maturation in hens induced by long-light regime is not accompanied with an increase in serum leptin or leptin receptor gene expression in hypothalamus, or genes expression in HPG axis.     

Sexual maturation in hens is not associated with increases in serum leptin and the expression of leptin receptor mRNA in hjpothalamus

Background： In mammals, leptin is an attractive candidate for mediating the metabolic signal and the reproductive function via the specific receptor in hypothalamus. However, till now, the role of leptin on reproduction in birds is less well established. This experiment was conducted to elucidate the role of leptin on the onset of reproduction in bird, as a first step, to detect the changes of peripheral leptin and leptin receptor mRNA expression in hypothalamus between mature and immature hens at the same age. 120 ISA brown pullets at D60 were allocated randomly into two groups, long light （LL） group being raised under artificial light regimes with incrementally increased light phase （from 8 L：]6D to 14 L：]2D） and short light （SL） group raised on consistent light （8 L：16D） for 12 wk. Results： The results showed that pullets in LL group reached sexual maturation 15 d earlier than those in SL group. Serum E2 showed a significant increase with age, but no difference was observed between two groups. Serum leptin concentration decreased significantly from D112 to D136 in LL, and was markedly higher in LL group than that in SL at D112, while there was no significant difference between two groups at D136. Leptin receptor and GnRH-I mRNA expression in hypothalamus were significantly increased with age, yet there was no significant difference between SL and LL chickens at the same age. The expression of FSH-13 and LH-13 mRNA in pituitary was increased with age but did not show significant difference between LL and gland, and decreased from D112 to D136 in LL but not groups at the same age. SL group. GnfiH-I mRNA expression was very rich in pinea n SL group, and there was no difference between two Conclusions： These results indicate that the earlier sexual maturation in hens induced by long-light regime is not accompanied with an increase in serum leptin or leptin receptor gene expression in hypothalamus, or genes expression in HPG axis.     

Identification of leptin gene polymorphisms associated with carcass traits and fatty acid composition in Japanese Black cattle

Previous studies have indicated that some leptin gene polymorphisms were associated with economically important traits in cattle breeds. However, polymorphisms in the leptin gene have not been reported thus far in Japanese Black cattle. Here, we aimed to identify the leptin gene polymorphisms which are associated with carcass traits and fatty acid composition in Japanese Black cattle. We sequenced the full‐length coding sequence of leptin gene for eight Japanese Black cattle. Sequence comparison revealed eight single nucleotide polymorphisms (SNPs). Three of these were predicted to cause amino acid substitutions: Y7F, R25C and A80V. Then, we genotyped these SNPs in two populations (JB1 with 560 animals and JB2 with 450 animals) and investigated the effects on the traits. Y7F in JB1 and A80V in JB2 were excluded from statistical analysis because the minor allele frequencies were low (< 0.1). Association analysis revealed that Y7F had a significant effect on the dressed carcass weight in JB2; R25C had a significant effect on C18:0 and C14:1 in JB1 and JB2, respectively; and A80V had a significant effect on C16:0, C16:1, C18:1, monounsaturated fatty acid and saturated fatty acid in JB1. The results suggested that these SNPs could be used as an effective marker for the improvement of Japanese Black cattle.     

多不饱和脂肪酸对脂肪代谢酶基因表达的影响

多不饱和脂肪酸不仅可以抑制脂肪合成酶基因的表达,还能对脂质氧化相关酶基因的表达起促进作用,从而调节机体脂质代谢过程。本文主要就多不饱和脂肪酸的分类和来源、对脂肪合成及氧化相关酶基因表达的影响作一综述。     

柴胡舒肝汤对脂肪肝模型动物的影响

采用高脂日粮饲喂海兰褐蛋公雏,制作脂肪肝动物模型,再以柴胡舒肝汤进行干预。结果显示,试验组鸡只逐渐出现类似脂肪肝综合征的临床症状,实验室检验血清呈黄色稠膏凝脂状,血脂含量显著升高,剖检肝脏肿大呈土黄色泽,肝脏脂肪含量明显升高,表明以高脂日粮加丙基硫氧嘧啶人工造模成功。中药预防组在其饮水中加入4.5mL柴胡舒肝汤,结果较好地对抗了高脂日粮与丙基硫氧嘧啶的作用,试验鸡没有出现相应临床症状与实验室检验指标,表明柴胡舒肝汤对高脂日粮加丙基硫氧嘧啶所致脂肪肝有预防作用。对已发生脂肪肝病变的鸡以柴胡舒肝汤进行治疗,使相关临床症状逐步消失,肝脏病变与其他检验指标得到恢复,表明柴胡舒肝汤对高脂日粮加丙基硫氧嘧啶所致脂肪肝有治疗作用,可将在其饮水中加入4.5mL柴胡舒肝汤作为推荐剂量。试验中没有发现动物对柴胡舒肝汤呈现不良反应。