动物营养学报    2021, Vol. 33 Issue (7): 3949-3958    PDF    
包膜丁酸钠对断奶羔羊免疫和抗氧化指标的影响
陈想1 , 左丽君1 , 王可鑫1 , 贾斌2 , 潘春媛1 , 姜宁1 , 张爱忠1     
1. 黑龙江八一农垦大学动物科技学院, 黑龙江省寒区饲料资源高效利用与营养调控重点实验室, 大庆 163319;
2. 黑龙江省农业科学院畜牧兽医分院, 齐齐哈尔 161000
摘要: 本试验旨在研究包膜丁酸钠(CSB)对断奶羔羊免疫和抗氧化指标的影响。选择42日龄的断奶羔羊24只,随机分成4组,分别为空白对照组(CON组)、抗生素对照组(A CON组)和包膜丁酸钠A组(CSB A组)和包膜丁酸钠B组(CSB B组),每组6个重复,每个重复1只。CON组饲喂不含抗生素的基础饲粮,A CON组在基础饲粮中添加250 mg/kg的黄霉素,CSB A组和CSB B组在基础饲粮中分别添加2和3 g/kg包膜丁酸钠。预试期7 d,正试期28 d。各组分别在试验第14天和第28天选取3只羔羊屠宰取样。结果表明:在试验第14天和第28天,与CON组相比,饲粮中添加包膜丁酸钠对羔羊的肝脏、脾脏、胸腺指数均有所提高(P>0.05)。在试验第14天,与CON组相比,CSB A组与CSB B组血清中肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)的含量显著下降(P < 0.05),A CON组血清中白细胞介素-2(IL-2)、丙二醛(MDA)的含量显著下降(P < 0.05);与A CON组相比,CSB A组与CSB B组血清中IL-6的含量显著下降(P < 0.05)。在试验第14天,与CON组相比,CSB B组肝脏中TNF-α、白细胞介素-1(IL-1)和IL-6的含量显著下降(P < 0.05),CSB A组与CSB B组肝脏中IL-2的含量显著下降(P < 0.05),CSB A组肝脏中总抗氧化能力(T-AOC)和总超氧化物歧化酶(T-SOD)的活性显著上升(P < 0.05)。在试验第28天,与CON组相比,CSB A组血清中IL-6的含量显著下降(P < 0.05),CSB A组与CSB B组血清中TNF-α的含量显著下降(P < 0.05),而血清中T-SOD的活性显著上升(P < 0.05);与A CON组相比,CSB B组血清中TNF-α的含量显著下降(P < 0.05),CSB A组与CSB B组血清中T-SOD的活性显著升高(P < 0.05)。在试验第28天,与CON组相比,CSB A组与CSB B组肝脏中TNF-α、IL-1和IL-2的含量显著下降(P < 0.05),而肝脏中T-AOC和T-SOD的活性显著上升(P < 0.05)。由此可见,饲粮中添加包膜丁酸钠可以抑制断奶羔羊血清和肝脏中促炎因子含量的升高并增强其抗氧化能力,从而在一定程度上保护羔羊免受断奶应激带来的负面影响,其中2 g/kg的添加量较为适宜。
关键词: 包膜丁酸钠    断奶羔羊    免疫器官指数    免疫指标    抗氧化指标    
Effects of Coated Sodium Butyrate on Immune and Antioxidant Indexes of Weaned Lambs
CHEN Xiang1 , ZUO Lijun1 , WANG Kexin1 , JIA Bin2 , PAN Chunyuan1 , JIANG Ning1 , ZHANG Aizhong1     
1. Key Laboratory of Efficient Utilization of Feed Resources and Nutrition Control in Cold Regions of Heilongjiang Province, College of Animal Science and Technology, Heilongjiang Bayi Land Reclamation University, Daqing 163319, China;
2. Animal Husbandry and Veterinary Branch of Heilongjiang Academy of Agricultural Sciences, Qiqihar 161000, China
Abstract: This experiment was conducted to study the effects of coated sodium butyrate on the immune and antioxidant indexes of weaned lambs. Twenty-four 42-day-old weaned lambs were randomly divided into 4 groups: blank control group (CON group), antibiotic control group (A CON group), coated sodium butyrate A group (CSB A group) and coated sodium butyrate B group (CSB B group), with 6 replicates in each group. Lambs in the CON group were fed a basal diet without antibiotics, and those in the A CON group were fed the basal diet supplemented with 250 mg/kg flavomycin, while those in the CSB A group and CSB B group were fed the basal diet supplemented with 2 and 3 g/kg coated sodium butyrate, respectively. The pre-trial period lasted for 7 days, and the experiment lasted for 28 days. Three lambs of each group were slaughtered and sampled on the 14th and 28th day of the experiment. The result showed as follows: On the 14th and 28th day of the experiment, compared with the CON group, the liver, spleen and thymus indexes of lambs were increased when diet supplemented with coated sodium butyrate (P>0.05). On the 14th day of the experiment, compared with the CON group, the contents of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in serum of CSB A group and CSB B group were significantly decreased (P < 0.05), and the contents of interleukin-2 (IL-2) and malondialdehyde (MDA) in serum of A CON group were significantly decreased (P < 0.05); compared with the A CON group, the IL-6 content in CSB A group and CSB B group was significantly decreased (P < 0.05). On the 14th and 28th day of the experiment, compared with the CON group, the contents of TNF-α, interleukin-1 (IL-1) and IL-6 in liver were significantly decreased (P < 0.05), and the IL-2 content in liver of CSB A group and CSB B group was significantly decreased (P < 0.05), while the total antioxidant capacity (T-AOC) and total superoxide dismutase (T-SOD) activity in liver of CSB A group were significantly increased (P < 0.05). On the 28th day of the experiment, compared with the CON group, the content of IL-6 in serum of CSB A group was significantly decreased (P < 0.05), and the TNF-α content in serum of CSB A group and CSB B group was significantly decreased (P < 0.05), while the T-SOD activity in serum of CSB A group and CSB B group was significantly increased (P < 0.05); compared with the A CON group, the TNF-α content in serum of CSB B group was significantly decreased (P < 0.05), while the T-SOD activity in serum of CSB A group and CSB B group was significantly increased (P < 0.05). On the 28th day of the experiment, compared with the CON group, the contents of TNF-α, IL-1 and IL-2 in liver of CSB A group and CSB B group were significantly decreased (P < 0.05), which the T-AOC and T-SOD activity in liver of CSB A group and CSB B group were significantly increased (P < 0.05). Therefore, the addition of coated sodium butyrate in the diet can inhibit the increase of inflammatory cytokines in serum and liver and enhance the antioxidant capacity of weaned lambs, thus protecting lambs from the negative effects of weaning stress to a certain extent, and 2 g/kg of coated sodium butyrate is more reasonable.
Key words: sodium butyrate    weaned lamb    immune organ index    immune index    antioxidant index    

断奶是羔羊生长的关键阶段,在断奶过程中母乳被替代为植物性饲料,饲喂模式的改变和与母羊的分离对于羔羊是巨大的应激,有可能会对其生长发育和健康产生不利影响,严重影响机体的免疫力和抗氧化能力[1],表现出体重下降、发病率上升和死亡率增加等问题[2],如何通过营养调控手段增强畜禽抗应激能力,降低应激造成的养殖损失日益受到关注。丁酸在维持细胞的分化和结肠上皮的完整性、提高动物生产性能方面发挥着重要作用[3],生产中常通过添加丁酸钠盐的形式补充丁酸。然而,由于丁酸钠具有难闻的气味和辛辣的味道,对动物采食量会产生负面影响,且暴露在空气中很容易吸湿。包膜处理可以减少丁酸钠的刺激性味道并稳定其化学性质,包膜材料可以在肠道中逐步释放,使丁酸钠在整个肠道持续释放[4]。张瑞阳等[5]、张卫辉[6]分别在仔猪与肉鸡上发现丁酸钠可以促进生长性能同时增强免疫功能;孟庆爽[7]在犊牛上发现丁酸钠能降低腹泻,有利于机体健康;左丽君[8]的试验发现饲粮中添加2和3 g/kg的包膜丁酸钠均可促进断奶羔羊的生长并提高营养物质表观消化率。为了预防羔羊出现断奶应激,更加全面了解丁酸钠对断奶羔羊的营养调控作用,本试验研究了包膜丁酸钠对断奶羔羊免疫和抗氧化指标的影响,旨在为包膜丁酸钠在断奶羔羊养殖中的科学应用提供理论依据。

1 材料与方法 1.1 试验材料

包膜丁酸钠为市购,丁酸钠含量为30%;羔羊(德国美利奴羊×杜泊羊)购于黑龙江中升牧业有限公司。

1.2 试验设计与饲粮

选择42日龄的断奶羔羊24只,随机分成4组,分别为空白对照组(CON组)、抗生素对照组(A CON组)和包膜丁酸钠A组(CSB A组)和包膜丁酸钠B组(CSB B组),每组6个重复,每个重复1只羊,单笼饲养。根据断奶羔羊体重和营养需要特点,参照我国《肉羊饲养标准》(NY/T 816—2004)配制基础饲粮,其组成及营养水平见表 1。CON组饲喂不含抗生素的基础饲粮,A CON组在基础饲粮中添加250 mg/kg的黄霉素,CSB A组和CSB B组在基础饲粮中分别添加2和3 g/kg包膜丁酸钠。各组羔羊常规饲养,自由采食、饮水。

表 1 基础饲粮组成及营养水平(风干基础) Table 1 Composition and nutrient levels of the basal diet (air-dry basis)  
1.3 饲养管理

试验前对羊舍进行冲洗消毒处理。羔羊采用全舍饲饲养管理,试验期间,试验羊每日饲喂2次,饲喂时间为07:00和17:00,精料和粗料分开饲喂,先精后粗,预试期7 d,正试期28 d。预试期内观察每只羊的采食量,并将每日饲喂的包膜丁酸钠早晚分2次拌入羔羊饲喂的精料中。供给充足清洁饮水,每天清粪、消毒,观察断奶羔羊健康状况。

1.4 样品采集

在正试期第14天和第28天,分别从每组中随机选3只羔羊,于早晨空腹称重后颈静脉采血10 mL,室温静置1 h,使用离心机4 000×g离心15 min,制备血清,将血清分装入1 mL离心管中,置于-20 ℃冰柜中冷冻保存备用。

羔羊静脉采血后立即屠宰,采集羔羊的免疫器官(肝脏、脾脏、胸腺),并去除结缔组织,用滤纸擦干血液,称取湿重,计算其器官指数。取肝脏,用预冷的生理盐水漂洗血渍、滤纸拭干,称取0.5 g,加生理盐水4.5 mL,于冰浴中充分匀浆,制成10%组织匀浆,4 ℃、3 500 r/min离心10 min,取上清液于-20 ℃保存待测。

1.5 测定指标及方法 1.5.1 免疫器官指数

免疫器官指数(g/kg)=免疫器官湿重/活体重。

1.5.2 血清、肝脏免疫指标

免疫球蛋白G(IgG)、白细胞介素-1(IL-1)、白细胞介素-2(IL-2)、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)含量均采用酶联免疫吸附测定试剂盒测定,试剂盒购于苏州卡尔文工程有限公司,按照试剂盒内说明书的步骤操作。

1.5.3 血清、肝脏抗氧化指标

谷胱甘肽过氧化物酶(GSH-Px)活性采用二硫代二硝基苯甲酸法测定,总超氧化物歧化酶(T-SOD)活性采用经胺法测定,总抗氧化能力(T-AOC)采用比色法测定,丙二醛(MDA)含量采用硫代巴比妥酸法测定。上述指标测定所有试剂盒均由南京建成生物工程研究所提供,测定仪器为全功能酶标仪,所有操作均按说明书步骤进行。

1.6 数据处理与分析

采用Excel 2007对试验数据进行初步统计,然后使用SPSS 19.0统计软件进行单因素方差分析,并采用Tukey’s多重比较分析组间差异显著性,P < 0.05表示差异显著。

2 结果与分析 2.1 包膜丁酸钠对断奶羔羊免疫器官指数的影响

图 1图 2图 3所示,在试验第14天与第28天,与CON组相比,饲粮中添加抗生素和包膜丁酸钠后羔羊的肝脏、脾脏和胸腺指数均有所提高,但各组间并无显著差异(P>0.05)。

数据柱形用平均值和标准误表示,标注不同小写字母表示差异显著(P < 0.05),无字母或相同字母表示差异不显著(P>0.05)。下图同。 Data column was represented by mean and standard deviation, and marked with different small letters showed significant difference (P < 0.05), while with no letter or the same letters mean no significant difference (P>0.05). The same as below. 图 1 包膜丁酸钠对断奶羔羊肝脏指数的影响 Fig. 1 Effects of coated sodium butyrate on liver index of weaned lambs
图 2 包膜丁酸钠对断奶羔羊脾脏指数的影响 Fig. 2 Effects of coated sodium butyrate on spleen index of weaned lambs
图 3 包膜丁酸钠对断奶羔羊胸腺指数的影响 Fig. 3 Effects of coated sodium butyrate on thymus index of weaned lambs
2.2 包膜丁酸钠对羔羊血清免疫指标的影响

表 2可知,在试验第14天,各组间血清中IgG、IL-1的含量差异不显著(P>0.05);与CON组相比,A CON组血清中TNF-α、IL-2和IL-6的含量显著下降(P < 0.05),CSB A组与CSB B组血清中TNF-α和IL-6的含量显著下降(P < 0.05);与A CON组相比,CSB A组与CSB B组血清中TNF-α的含量无显著差异(P>0.05),CSB A组血清中IL-2的含量显著升高(P < 0.05),CSB A组与CSB B组血清中IL-6的含量显著下降(P < 0.05)。

表 2 包膜丁酸钠对断奶羔羊血清免疫指标的影响 Table 2 Effects of coated sodium butyrate on serum immune indexes of weaned lambs  

在试验第28天,各组间血清中IgG、IL-1和IL-2的含量差异不显著(P>0.05);与CON组相比,A CON组血清中TNF-α和IL-6的含量显著下降(P < 0.05),CSB A组与CSB B组血清中TNF-α的含量显著下降(P < 0.05),CSB A组血清中IL-6的含量显著下降(P < 0.05);与A CON组相比,CSB B组血清中TNF-α的含量显著下降(P < 0.05),血清中IL-6的含量显著提高(P < 0.05)。

2.3 包膜丁酸钠对断奶羔羊肝脏免疫指标的影响

表 3可知,在试验第14天,与CON组相比,A CON组肝脏中IL-1和IL-2的含量显著下降(P < 0.05),CSB A组肝脏中IL-2和IL-6的含量显著下降(P < 0.05),CSB B组肝脏中TNF-α、IL-1和IL-2的含量显著下降(P < 0.05);与A CON组相比,CSB A组肝脏中IL-2和IL-6的含量显著下降(P < 0.05),CSB B组肝脏中TNF-α的含量显著下降(P < 0.05),CSB A组与CSB B组肝脏中IL-1的含量显著提高(P < 0.05)。

表 3 包膜丁酸钠对断奶羔羊肝脏免疫指标的影响 Table 3 Effects of coated sodium butyrate on liver immune indexes of weaned lambs  

在试验第28天,与CON组相比,A CON组、CSB A组和CSB B组肝脏中TNF-α、IL-1和IL-2的含量显著下降(P < 0.05);与A CON组相比,CSB A组肝脏中TNF-α及CSB B组肝脏中IL-1的含量显著下降(P < 0.05),CSB A组肝脏中IL-1及CSB B组肝脏中IL-2的含量显著提高(P < 0.05)。

2.4 包膜丁酸钠对断奶羔羊血清抗氧化指标的影响

表 4可知,在试验第14天,各组间血清中T-AOC及T-SOD、GSH-Px的活性没有显著差异(P>0.05);A CON组血清中MDA的含量显著低于CON组(P < 0.05)。

表 4 包膜丁酸钠对断奶羔羊血清抗氧化指标的影响 Table 4 Effects of coated sodium butyrate on serum antioxidant indexes of weaned lambs

在试验第28天,与CON组相比,CSB A组和CSB B组血清中T-AOC和GSH-Px的活性呈现上升状态,MDA的含量呈现与之相反的趋势,但组间差异并不显著(P>0.05),CSB A组和CSB B组血清中T-SOD的活性显著升高(P < 0.05);与A CON组相比,CSB A组和CSB B组血清中T-SOD的活性显著升高(P < 0.05)。

2.5 包膜丁酸钠对断奶羔羊肝脏抗氧化指标的影响

表 5可知,在试验第14天,与CON组相比,CSB A组肝脏中T-AOC和T-SOD的活性显著升高(P < 0.05),各组间肝脏中GSH-Px的活性和MDA的含量无显著差异(P>0.05)。

表 5 包膜丁酸钠对断奶羔羊肝脏抗氧化指标的影响 Table 5 Effects of coated sodium butyrate on liver antioxidant indexes of weaned lambs

在试验第28天,与CON组相比,A CON组、CSB A组和CSB B组肝脏中T-AOC和T-SOD的活性显著升高(P < 0.05);与A CON组相比,CSB A组肝脏中T-SOD的活性显著降低(P < 0.05),各组间肝脏中GSH-Px的活性和MDA的含量无显著差异(P>0.05)。

3 讨论 3.1 丁酸钠对断奶羔羊免疫器官指数的影响

器官指数是评价动物机体机能强弱的常用生物学指标,其大小在一定程度上反映了该器官的机能状态和发育情况[9]。随着能量摄入量的增加,动物的机体与器官发育也会随之加快[10]。在正常的状态下,动物可以调节自身器官发育与生长发育保持一致。本试验研究发现,添加包膜丁酸钠对断奶羔羊肝脏、脾脏、胸腺指数有提高的趋势,但差异不显著,其原因可能是内脏与动物体重的增长达到了相适应的比例,这种适宜的比例能较好地承担起动物机体的新陈代谢。由此可以推断,添加包膜丁酸钠可以促进羔羊对各营养成分的摄入,来供给羔羊机体的免疫器官发育。

3.2 包膜丁酸钠对断奶羔羊血清免疫指标的影响

在一定范围内血清免疫球蛋白含量越高则说明免疫效果越好。IgG是发挥主要免疫作用的免疫球蛋白,占动物血清免疫球蛋白总量的75%~80%,此外免疫球蛋白在肠道中发挥的作用同样也是巨大的[11]。赵会利等[12]研究发现,丁酸钠有提高断奶犊牛血清IgG含量的趋势,从而提高机体免疫功能和抗病力。本研究也发现,包膜丁酸钠可以提高断奶羔羊血清中IgG的含量。包被丁酸钠可提高仔猪的免疫功能,其原因可能与其有效成分丁酸的酸化性能有关,丁酸可通过调节肠道pH改善肠道菌群平衡,进而提高机体免疫力[6]。白细胞介素具有参与炎症反应、激活调节免疫细胞和传递信息等多种生物学功能[13],因此阻断白细胞介素类和TNF-α均可减轻炎症程度[14]。短链脂肪酸已被发现与炎症调节是密切相关的[15],已有研究能充分证实丁酸可以减少促炎因子的产生[15-18]。本研究中,在试验第14天,与CON组相比,CSB B组断奶羔羊血清中TNF-α和IL-6的含量显著降低;在试验第28天,与CON组相比,CSB A组和CSB B组断奶羔羊血清中TNF-α的含量显著降低,同时CSB A组血清中IL-6的含量显著降低。上述结果表明,饲粮中添加包膜丁酸钠可减少TNF-α和白细胞介素类促炎因子的产生。

3.3 包膜丁酸钠对断奶羔羊肝脏免疫指标的影响

肝脏是机体毒素降解和代谢的主要器官,同时也是内毒素侵害时最易受损的器官[19]。TNF-α作为促炎因子的代表,处于炎性反应的调控地位,也是肝细胞死亡和损害的主要调控因子。Cermanova等[20]研究发现,丁酸钠能明显抑制肝脏中TNF-α mRNA的表达。丁酸钠能显著改善亚急性瘤胃酸中毒(SARA)奶山羊肝组织的炎症反应,能显著抑制肝脏中炎症基因TNF-α与白细胞介素-1β(IL-1β)的表达[21]。丁酸可进一步降低非酒精性脂肪性肝病大鼠血清和肝脏内毒素水平,可使肝脏组织中TNF-αIL-1、IL-2、IL-6、γ-干扰素(IFN-γ)等促炎因子的表达明显下调[22]。丁酸钠对自身免疫性肝炎模型小鼠肝脏损伤的保护作用可能与其抑制Toll样受体4(TLR4)信号通路,下调炎性因子IL-6和TNF-α的表达有关[23]。吴锦露等[24]研究发现,丁酸钠可直接调节辅助性T细胞17(Th17)的免疫平衡,抑制肝脏TLR4炎症通路,下调肝脏促炎因子的表达。TLR4广泛分布于肝脏组织细胞表面[25],高迁移率族蛋白B1(HMGB1)是早期介导肝脏损伤的主要内源性配体[26],HMGB1作为TLR4内源性配体在肝脏损伤中起重要作用[27],而丁酸钠能有效抑制HMGB1的产生[28]。在本研究中,试验第14天和第28天,饲粮中添加包膜丁酸钠均降低了肝脏中TNF-α、白细胞介素类促炎因子的含量,由此推断,丁酸钠对断奶羔羊的肝脏免疫起到了促进作用。

3.4 包膜丁酸钠对断奶羔羊血清抗氧化指标的影响

机体受到外界刺激,体内会产生大量的自由基,过量的自由基会使细胞代谢紊乱,从而威胁养殖动物的健康。T-AOC由酶促和非酶促抗氧化防御体系共同组成,反映了机体总抗氧化、清除自由基的能力;T-SOD、GSH-Px是抗氧化系统中最主要的酶,能够有效清除自由基;MDA是体内脂质过氧化物,过量产生会破坏细胞膜完整性、使细胞发生突变、对抗氧化防御系统造成损伤[29]。岳敏等[30]研究发现丁酸钠可通过提高母猪血清中超氧化物歧化酶(SOD)的活性,降低血清中MDA的含量,来提高机体的抗氧化能力。犊牛断奶前后在酸化奶中加入丁酸钠可提高血清中T-AOC和T-SOD的活性,并且能显著降低犊牛血清中MDA的含量[31]。张卫辉[6]发现,在饲粮中添加丁酸钠可以显著提高21日龄肉鸡血清中SOD的活性,并且显著降低血清中MDA的含量,但对血清中GSH-Px的活性无显著影响。本试验结果显示,饲粮中添加丁酸钠可提高断奶羔羊血清中T-AOC及T-SOD、GSH-Px的活性,并且降低血清中MDA的含量,这与上述前人报道所得的结论是一致的,说明丁酸钠能够提高机体的抗氧化能力。

3.5 包膜丁酸钠对断奶羔羊肝脏抗氧化指标的影响

肝脏是动物重要的代谢与免疫器官。丁酸钠可改变动物组织的抗氧化指数,表明丁酸钠能改善氧化应激状态[32]。Lan等[33]研究表明,以1 200 mg/kg的剂量添加丁酸钠缓解了由较高的环境温度引起的胸肌和肝脏氧化应激,从而改善肉鸡的肉品质和肝功能。与饲喂高谷物饲粮的山羊相比,饲喂添加丁酸钠饲粮的山羊血清中T-SOD的活性以及T-AOC增加,肝脏和血浆中MDA的含量均下降[34]。本试验结果表示,在试验第14天,与CON组相比,CSB A组羔羊肝脏中T-AOC和T-SOD的活性显著升高;在试验第28天,CSB A组和CSB B组羔羊肝脏中T-AOC和T-SOD的活性显著升高。上述结果说明,饲粮中添加包膜丁酸钠可提高羔羊肝脏的抗氧化能力。

4 结论

饲粮中添加包膜丁酸钠可以抑制断奶羔羊血清和肝脏中促炎因子含量的升高,并且还可增加断奶羔羊机体和肝脏的抗氧化能力,从而在一定程度上保护其免受断奶应激带来的负面影响;2和3 g/kg的包膜丁酸钠的作用效果基本一致,从经济效益方面考虑,选择2 g/kg的包膜丁酸钠较为合理。

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