烟酸调控高精料诱导瘤胃上皮细胞凋亡机制的研究

发布时间：2018-12-12 02:30

【摘要】：高精料日粮引起的瘤胃酸中毒已成为影响现代反刍动物生产的重要问题。瘤胃酸中毒导致的瘤胃上皮受损是加重并引起酸中毒全身症状的主要因素。B族维生素烟酸已被证实有缓解高精料应激的作用,但其对瘤胃屏障有何影响尚未见报道。本研究通过体内试验来研究高精料日粮中添加烟酸对绵羊瘤胃发酵、抗氧化能力及瘤胃上皮细胞凋亡的影响,以期从分子水平探讨烟酸保护瘤胃上皮、调控瘤胃酸中毒的机制。本试验选择9头装有永久性瘘管的绵羊(34±3 kg BW),采用单因素完全随机试验设计,试验分为3组,每组3个重复。第Ⅰ组为低精料组(日粮精粗比为2:8),试验期为10 d,包括7 d预试期和3 d正试期。第Ⅱ组为高精料组(日粮精粗比为8:2),第Ⅲ组为高精料+烟酸组(日粮精粗比为8:2,烟酸添加量为800 mg/kg日粮干物质)。结果表明:(1)与低精料组相比,高精料组降低了瘤胃pH值(P0.01),极显著增加了瘤胃总挥发性脂肪酸(TVFA)、乙酸、丙酸、丁酸及乳酸的浓度(P0.01)。高精料日粮还极显著提高了瘤胃内NAD+浓度、NADH浓度、NADH/NAD+的比例(P0.01);高精料+烟酸组极显著提高了瘤胃pH值(P0.01),降低了乙酸、丙酸、丁酸、总挥发性脂肪酸(TVFA)及乳酸的浓度(P0.01);同时还极显著降低了NAD+、NADH的浓度及NADH/NAD+的比例(P0.01)。(2)与低精料组相比,高精料组的过氧化氢酶(CAT)、总超氧化物岐化酶(T-SOD)、谷胱甘肽过氧化物酶(GSH-PX)的活性极显著降低,总抗氧化能力(T-AOC)极显著下降,丙二醛(MDA)的浓度极显著提高(P0.01);高精料条件下添加烟酸极显著提高了CAT、GSH-PX的活性,T-AOC增强,降低了MDA的浓度(P0.01),对T-SOD并无影响(P0.05)。(3)与低精料组相比,高精料组的绵羊血清中促炎因子IL-1β、IL-6、TNF-α和抗炎因子IL-4、IL-10的浓度均极显著升高(P0.01),高精料+烟酸组的绵羊血清中促炎因子IL-1β、IL-6、TNF-α和抗炎因子IL-4、IL-10的浓度均极显著低于高精料组(P0.01)。与低精料组相比,高精料组的瘤胃乳头长度、宽度及角质层厚度极显著降低(P0.01),细胞凋亡指数极显著升高(P0.01);高精料+烟酸组绵羊的瘤胃乳头长度、宽度及角质层高于高精料组(P0.01或P0.05),极显著降低了细胞凋亡指数(P0.01)。(4)与低精料组相比,高精料组绵羊的瘤胃上皮Fas、Bcl-2、Bax、P53上调,Caspase-3、Caspase-8、Caspase-9的mRNA表达水平极显著提高(P0.01),高精料条件下添加烟酸极显著地下调Fas、Bcl-2、Bax、P53及Caspase-3、Caspase-8、Caspase-9的表达(P0.01)。综上所述,高精料应激导致绵羊瘤胃内VFA和乳酸浓度提高,pH值降低,上皮细胞抗氧化性能力下降,激活凋亡相关基因P53、Bcl-2、Bax、Fas表达上调,Caspase-3、Caspase-8、Caspase-9表达提高,导致瘤胃上皮细胞凋亡,瘤胃上皮功能受损。而烟酸一定程度上可以缓解高精料应激,提高瘤胃上皮细胞抗氧化能力,抑制凋亡相关基因的表达,抑制了瘤胃上皮细胞的凋亡,保护了瘤胃上皮结构功能完整。
[Abstract]:Rumen acidosis caused by high concentrate diet has become an important problem affecting the production of modern ruminants. The damage of rumen epithelium caused by rumen acidosis is the main factor that exacerbates and causes the systemic symptoms of acidosis. Group B vitamin nicotinic acid has been proved to have the effect of relieving high concentrate stress, but its effect on rumen barrier has not been reported. In this study, the effects of niacin supplementation on rumen fermentation, antioxidant ability and apoptosis of rumen epithelial cells in sheep were studied in vivo. The aim of this study was to explore the mechanism of niacin in protecting rumen epithelium and regulating rumen acidosis at molecular level. Nine sheep with permanent fistula (34 卤3 kg BW),) were randomly divided into 3 groups with 3 repeats. The trial period was 10 days, which included 7 days pretest period and 3 days positive trial period. The second group was high concentrate group (the ration was 8:2), the third group was high concentrate niacin group (the ration of concentrate and crude ration was 8: 2, niacin was 800 mg/kg dry matter). The results showed that: (1) compared with the low concentrate group, the high concentrate group decreased the rumen pH value (P0.01), and significantly increased the concentration of (TVFA), acetic acid, propionic acid, butyric acid and lactic acid in the rumen total volatile fatty acids (P0.01). High concentrate diet also significantly increased rumen NAD concentration, NADH concentration, NADH/NAD ratio (P0.01); The concentration of acetic acid, propionic acid, butyric acid, total volatile fatty acid (TVFA) and lactic acid decreased significantly (P0.01) in high concentrate niacin group. At the same time, the concentration of NAD, NADH and the ratio of NADH/NAD (P0.01). (2) were significantly decreased in the high concentrate group compared with the low concentrate group, the catalase (CAT), total superoxide dismutase (T-SOD) in the high concentrate group was significantly lower than that in the low concentrate group. Glutathione peroxidase (GSH-PX) activity, total antioxidant capacity (T-AOC) and malondialdehyde (MDA) concentration were significantly decreased (P0.01). Under the condition of high concentration of niacin, the activity of CAT,GSH-PX was significantly increased, T-AOC was enhanced, the concentration of MDA was decreased (P0.01), and the concentration of T-SOD was not affected (P0.05). (3) compared with the low concentrate group. The concentrations of proinflammatory factor IL-1 尾, IL-6,TNF- 伪 and anti inflammatory factor IL-4,IL-10 in sheep serum of high concentrate group were significantly higher than those of high concentrate group (P0.01), and IL-1 尾, IL-6, in serum of high concentrate group were significantly higher than those of high concentrate group (P0.01). The concentrations of TNF- 伪 and anti inflammatory factor IL-4,IL-10 were significantly lower than those of high concentrate group (P0.01). Compared with the low concentrate group, the rumen papilla length, width and cuticle thickness in the high concentrate group were significantly decreased (P0.01), and the apoptosis index was significantly increased (P0.01). The length, width and cuticle of rumen nipple in high concentrate niacin group were higher than those in high concentrate group (P0.01 or P0.05), and the apoptosis index (P0.01). (4) was significantly lower than that in low concentrate group. The rumen epithelium Fas,Bcl-2,Bax,P53 was up-regulated and the mRNA expression of Caspase-3,Caspase-8,Caspase-9 was significantly increased in high concentrate group (P0.01), and niacin significantly down-regulated Fas,Bcl-2,Bax, in high concentrate group (P0.01). P53 and Caspase-3,Caspase-8,Caspase-9 expression (P0.01). In conclusion, high concentrate stress increased the concentration of VFA and lactic acid in rumen of sheep, decreased the pH value, decreased the antioxidant ability of epithelial cells, up-regulated the expression of apoptosis-related gene P53, Bcl-2, and increased the expression of Caspase-3,Caspase-8, in sheep rumen. The increased expression of Caspase-9 resulted in apoptosis of rumen epithelial cells and impaired function of rumen epithelium. To some extent, niacin can relieve high concentrate stress, improve the antioxidant ability of rumen epithelial cells, inhibit the expression of apoptosis-related genes, inhibit the apoptosis of rumen epithelial cells, and protect the structure and function of rumen epithelium.
【学位授予单位】：江西农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S826.5

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