Chinese Journal of Animal Nutrition >

2025 , Vol. 37 >Issue 7: 4532 - 4542

DOI: https://doi.org/10.12418/CJAN2025.371

RESEARCH PAPER

Alleviating Effect of Anthocyanins on Decline in Performance, Oxidative Stress, Immune Damage and Liver Injury of Laying Hens Induced by Fumonisin B1

  • ZHENG Xu , 1 ,
  • HAO Erying 1 ,
  • ZHAO Bin 2 ,
  • JI Yifan 1 ,
  • LIU Zhe 1 ,
  • LI Yanhui 1 ,
  • TIAN Yue 1 ,
  • YUAN Wen 1 ,
  • CHEN Hui , 1, *
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  • 1 College of Animal Science and Technology, Hebei Agricultural University, Baoding 071001, China
  • 2 College of Plant Protection, Hebei Agricultural University, Baoding 071001, China
* professor, E-mail:

Received date: 2024-12-02

  Online published: 2025-07-12

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Abstract

This experiment aimed to study the alleviating effects of anthocyanins on the performance decline, oxidative stress, immune damage and liver injury induced by fusarium B1 (FB1) in laying hens. One hundred and eighty 60-week-old Dawu Jinfeng laying hens were randomly divided into three groups: the control group (fed with a basal diet), the fusarium group (fed with the basal diet supplemented with 22.67 mg/kg FB1), and the remission group (fed with the basal diet supplemented with 22.67 mg/kg FB1 and 150 mg/kg anthocyanin). Each group had six replicates, with 10 chickens per replicate. The pre-test period was 14 days, and the formal test period was 56 days. The results showed as follows: compared with the control group, the average daily feed intake, average egg weight and laying rate of laying hens in the fumonisin group were all reduced, while the feed/egg ratio increased; the activities of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) in plasma, as well as the contents of immunoglobulin G (IgG) and immunoglobulin M (IgM), were significantly decreased (P<0.05), whereas the contents of malondialdehyde (MDA), interleukin-2 (IL-2) and sphingomyelin (Sph), along with the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and gamma-glutamyl transferase (GGT), were significantly increased (P<0.05); there was severe hepatocyte necrosis and lymphocyte swelling; the relative abundance of Bacteroidetes in the cecum was significantly reduced (P<0.05), while the relative abundance of Firmicutes was significantly increased (P<0.05). Compared with the fumonisin group, the average daily feed intake and average egg weight of laying hens in the remission group were significantly increased (P<0.05); plasma GSH-Px and SOD activities, as well as IgG and IgM contents, were significantly increased (P<0.05), while MDA and Sph contents, along with AST, ALT and GGT activities, were significantly decreased (P<0.05); lymphocyte volume decreased, and liver cell damage symptoms were alleviated; the relative abundances of Bacteroidetes and Bacteroides in the cecum were significantly increased (P<0.05). In summary, FB1 has caused a decline in the performance, oxidative stress, weakened immune function and liver damage of laying hens; anthocyanins can effectively alleviate the liver injury caused by FB1 in laying hens by improving the antioxidant ability and anti-inflammatory ability of the body.

Key words： laying hens; anthocyanins; fumonitoxin; liver injury; oxidative stress

Cite this article

ZHENG Xu , HAO Erying , ZHAO Bin , JI Yifan , LIU Zhe , LI Yanhui , TIAN Yue , YUAN Wen , CHEN Hui . Alleviating Effect of Anthocyanins on Decline in Performance, Oxidative Stress, Immune Damage and Liver Injury of Laying Hens Induced by Fumonisin B1[J]. Chinese Journal of Animal Nutrition, 2025 , 37(7) : 4532 -4542 . DOI: 10.12418/CJAN2025.371

近年来,霉菌毒素对畜牧业造成的危害愈发严重[1]。霉菌毒素是由某些真菌生物在温湿度适宜的条件下在谷物中产生的次生代谢物,特别是来自镰刀菌属的伏马毒素B1(FB1),主要存在于玉米中[2]。FB1能引起肝脏氧化应激[1],还会直接损伤肠道和肝脏,导致家禽生产性能下降和免疫损伤[3]。
花青素是一类重要的黄酮类天然植物色素[4],具有抗炎、抗菌、抗凋亡、抗氧化以及其保护肝脏的特性,可应用于缓解动物的氧化应激、免疫应激以及肝病[4-7]。据文献报道,花青素有3种主要的抗氧化机制,包括氢原子转移(HAT)、单电子转移(SET)和序向质子损失电子转移(SPLET)[8]。研究表明,花青素可以通过去除氧自由基、抑制氧化酶和螯合金属离子,进而抑制脂质过氧化,恢复抗氧化状态,提高对黄曲霉毒素破坏机体的抵抗能力[9-10]。慢性炎症也可导致活性氧(ROS)失衡,补充花青素等抗氧化剂可对抗过量的ROS。
本课题组前期研究了在饲粮中添加不同水平花青素对蛋鸡的影响,结果显示以添加150 mg/kg花青素对蛋鸡的作用效果最佳[11]。在前期研究的基础上,本研究拟通过建立蛋鸡FB1攻毒模型,从肝脏病理学、肝脏功能、抗氧化功能、炎症因子等角度探究花青素对FB1诱导的蛋鸡肝脏损伤的保护作用机制,为预防FB1诱导的蛋鸡肝脏损伤提供理论依据。

1 材料与方法

1.1 试验材料

花青素提取自蓝莓,有效成分花青素含量为25%;FB1是与河北农业大学植物保护学院研究伏马毒素课题组合作发酵制作而成。

1.2 试验设计

采用单因素试验设计,选取健康、体重相近的60周龄大午金凤蛋鸡180只,随机分成3组,每组6个重复,每个重复10只鸡。预试期14 d,正试期56 d。对照组饲喂基础饲粮,伏马毒素组在基础饲粮的基础上添加22.67 mg/kg FB1,缓解组在基础饲粮的基础上添加22.67 mg/kg FB1和150 mg/kg花青素。花青素添加量的设定参照为本课题组筛选的最佳剂量[11]。FB1添加量的设定参照《饲料卫生标准》(GB 13078—2017)。基础饲粮配制参照《鸡饲养标准》(NY/T 33—2004),其组成及营养水平见表1。
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)%

项目 Items 含量 Content
原料 Ingredients
玉米 Corn 63.00
豆粕 Soybean meal 24.50
石粉 Limestone 9.50
大豆油 Soybean oil 1.00
预混料 Premix1) 2.00
合计 Total 100.00
营养水平 Nutrient levels2)
代谢能 ME/(MJ/kg) 12.35
粗蛋白质 CP 16.56
钙 Ca 3.62
总磷 TP 0.46
蛋氨酸 Met 0.30
赖氨酸 Lys 0.82

1)预混料为每千克饲粮提供 The premix provided the following per kg of the diet:VA 8 000 IU,VD3 3 400 IU,VE 9 mg,VK3 1.5 mg,VB1 1.52 mg,VB2 6 mg,VB6 2.4 mg,VB12 0.01 mg,烟酸 nicotinic acid 18 mg,泛酸 pantothenic acid 7.6 mg,叶酸 folic acid 0.6 mg,D-生物素 D-biotin 0.1 mg,Fe (as ferrous sulfate) 30 mg,Cu (as copper sulfate) 9 mg,Zn (as zinc sulfate) 70 mg,Mn (as manganese sulfate) 70 mg,I (as potassium iodide) 0.44 mg,Se (as sodium selenite) 0.16 mg。
2)粗蛋白质、钙和总磷为测定值,参考标准分别为GB/T 6432—2018、GB/T 6436—2018和GB/T 6437—2018;其余为计算值,依据NY/T 33—2004计算得出。CP, Ca and TP were determined, and the reference standards were GB/T 6432—2018, GB/T 6436—2018 and GB/T 6437—2018, respectively; the rest were calculated values, and they were calculated according to NY/T 33—2004.

1.3 饲养管理

本研究的动物试验经河北农业大学实验动物伦理委员会审核(2023029),符合动物保护、动物福利和伦理原则。试验采用3层阶梯笼养,自由采食和饮水。每日15:30收集鸡蛋并观察鸡群状态。

1.4 样品采集

饲养试验结束后,每个重复随机选取2只试验鸡采用翅下静脉采血法采集血液5 mL于肝素抗凝管中,静置15 min后753.253×g离心15 min,收集血浆分装于1.5 mL EP管中,-20 ℃保存备用;将采血后的试验鸡屠宰,取肝脏样品,冲洗,置于10%中性福尔马林固定液中保存。

1.5 测定指标及方法

1.5.1 生产性能测定

每天记录试验鸡的产蛋数、蛋重和采食量,计算平均蛋重、产蛋率、平均日采食量与料蛋比。

1.5.2 血浆抗氧化指标测定

使用试剂盒测定血浆中谷胱甘肽过氧化物酶(GSH-Px)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性以及丙二醛(MDA)含量、总抗氧化能力(T-AOC)。GSH-Px、SOD、CAT活性及MDA含量检测试剂盒(酶联免疫吸附检测法)购自上海江莱生物科技有限公司,T-AOC检测试剂盒(生化法)购自南京建成生物工程研究所。

1.5.3 血浆免疫指标测定

使用试剂盒(酶联免疫吸附检测法)测定血浆中免疫球蛋白A(IgA)、免疫球蛋白G(IgG)、免疫球蛋白M(IgM)、白细胞介素-1β(IL-1β)、白细胞介素-2(IL-2)、肿瘤坏死因子-α(TNF-α)、干扰素-β(IFN-β)含量,试剂盒购自上海江莱生物科技有限公司。

1.5.4 血浆中肝脏功能指标测定

使用试剂盒测定血浆中鞘氨醇(Sph)、肌酐(CRE)、总胆红素(TBIL)含量和谷草转氨酶(AST)、谷丙转氨酶(ALT)、谷氨酰转移酶(GGT)活性。Sph含量和GGT活性检测试剂盒(酶联免疫吸附检测法)购自上海江莱生物科技有限公司,CRE、TBIL含量与AST、ALT、GGT活性检测试剂盒(生化法)购自南京建成生物工程研究所。

1.5.5 肝脏组织病理学观察

肝脏组织经固定、石蜡包埋、切片、苏木精-伊红(HE)染色和封固后,使用显微摄像系统对切片进行图像采集,观察病变。

1.5.6 盲肠菌群结构分析

试验结束后,从每个重复随机选2只鸡屠宰,取盲肠内容物,采用高通量测序技术对盲肠菌群16S rRNA基因进行PCR扩增并测序分析。

1.6 数据统计分析

采用SPSS 20统计软件对试验数据进行单因素方差分析(one-way ANOVA),并采用Duncan氏法进行多重比较,结果以“平均值±标准差”表示,P<0.05表示差异显著。

2 结果与分析

2.1 花青素对FB1攻毒蛋鸡生产性能的影响

由表2可知,与对照组相比,伏马毒素组的平均日采食量、平均蛋重、料蛋比和产蛋率有虽无显著差异(P>0.05),但在数值上平均日采食量、平均蛋重和产蛋率都有所降低,料蛋比有所升高;与伏马毒素组相比,缓解组的平均日采食量与平均蛋重显著升高(P<0.05),料蛋比与产蛋率无显著差异(P>0.05)。
表2 花青素对FB1攻毒蛋鸡生产性能的影响

Table 2 Effects of anthocyanin on performance of laying hens challenged with FB1

项目
Items		 对照组
Control group		 伏马毒素组
Fumonisin group		 缓解组
Remission group		 P值
P-value
平均日采食量 ADFI/g 108.71±0.52ab 106.27±1.85b 110.11±2.82a 0.027
料蛋比 Feed/egg ratio 2.10±0.18 2.11±0.12 2.08±0.14 0.952
平均蛋重 Average egg weight/g 60.45±0.28ab 59.93±0.86b 61.23±0.72a 0.045
产蛋率 Laying rate/% 86.72±7.03 85.23±5.51 87.10±6.76 0.871

同行数据肩标相同字母或无字母表示差异不显著(P>0.05),不同字母表示差异显著(P<0.05)。下表同。

In the same row, values with the same or no letter superscripts mean no significant difference (P>0.05), while with different letter superscripts mean significant difference (P<0.05). The same as below.

2.2 花青素对FB1攻毒蛋鸡血浆抗氧化指标的影响

由表3可知,与对照组相比,伏马毒素组蛋鸡血浆中GSH-Px、SOD、CAT活性显著降低(P<0.05),MDA含量显著升高(P<0.05),T-AOC差异不显著(P>0.05);与伏马毒素组相比,缓解组蛋鸡血浆中GSH-Px、SOD活性显著升高(P<0.05),MDA含量显著降低(P<0.05),CAT活性和T-AOC差异不显著(P>0.05)。
表3 花青素对FB1攻毒蛋鸡血浆抗氧化指标的影响

Table 3 Effects of anthocyanin on plasma antioxidant indexes of laying hens challenged with FB1

项目
Items		 对照组
Control group		 伏马毒素组
Fumonisin group		 缓解组
Remission group		 P值
P-value
谷胱甘肽过氧化物酶 GSH-Px/(U/mL) 756.26±6.51a 748.14±10.39b 760.11±1.48a 0.004
超氧化物歧化酶 SOD/(U/mL) 284.02±1.84a 280.98±4.90b 285.11±0.73a 0.013
过氧化氢酶 CAT/(U/mL) 20.84±0.60a 19.19±1.26b 19.88±0.57b 0.006
丙二醛 MDA/(nmol/mL) 0.66±0.21b 1.02±0.27a 0.72±0.12b 0.007
总抗氧化能力 T-AOC/(mmol/L) 3.22±0.11 3.16±0.03 3.23±0.08 0.125

2.3 花青素对FB1攻毒蛋鸡血浆免疫指标的影响

由表4可知,与对照组相比,伏马毒素组蛋鸡血浆中IgG、IgM含量显著降低(P<0.05),IL-2含量显著升高(P<0.05),其余指标均差异不显著(P>0.05);与伏马毒素组相比,缓解组蛋鸡血浆中IgG、IgM含量显著升高(P<0.05),其余指标均差异不显著(P>0.05)。
表4 花青素对FB1攻毒蛋鸡血浆免疫指标的影响

Table 4 Effects of anthocyanin on plasma immune indexes of laying hens challenged with FB1

项目
Items		 对照组
Control group		 伏马毒素组
Fumonisin group		 缓解组
Remission group		 P值
P-value
干扰素-β IFN-β/(pg/mL) 708.54±5.19 712.53±1.40 710.86±3.54 0.080
免疫球蛋白A IgA/(μg/mL) 833.29±3.70 835.13±1.61 835.83±1.50 0.087
免疫球蛋白G IgG/(μg/mL) 2 165.68±12.02a 2 140.72±42.07b 2 171.20±6.53a 0.029
免疫球蛋白M IgM/(μg/mL) 283.13±1.47a 279.83±4.41b 283.82±0.57a 0.006
白细胞介素-2 IL-2/(pg/mL) 457.64±3.23b 460.16±1.09a 458.28±1.37ab 0.043
白细胞介素-1β IL-1β/(pg/mL) 161.38±17.53 169.32±30.81 160.09±25.17 0.756
肿瘤坏死因子-α TNF-α/(ng/mL) 2 248.48±19.20 2 229.12±43.02 2 240.65±53.32 0.516

2.4 花青素对FB1攻毒蛋鸡血浆中肝脏功能指标的影响

由表5可知,与对照组相比,伏马毒素组蛋鸡血浆中Sph含量与AST、ALT、GGT活性显著升高(P<0.05),CRE与TBIL含量差异不显著(P>0.05);与伏马毒素组对比,缓解组蛋鸡血浆中Sph含量与AST、ALT、GGT活性显著降低(P<0.05),CRE、TBIL含量差异不显著(P>0.05)。
表5 花青素对FB1攻毒蛋鸡血浆中肝脏功能指标的影响

Table 5 Effects of anthocyanin on liver function indexes in plasma of laying hens challenged with FB1

项目
Items		 对照组
Control group		 伏马毒素组
Fumonisin group		 缓解组
Remission group		 P值
P-value
鞘氨醇 Sph/(ng/mL) 9.27±0.76b 11.68±2.76a 9.43±1.01b 0.013
肌酐 CRE/(μmol/L) 24.55±4.22 26.66±3.95 23.73±4.72 0.257
总胆红素 TBIL/(μmol/L) 8.74±1.25 8.63±1.14 9.01±1.30 0.744
谷草转氨酶 AST/(U/L) 19.00±2.68b 22.13±2.98a 18.34±3.86b 0.015
谷丙转氨酶 ALT/(U/L) 4.68±0.46b 6.38±0.86a 4.61±0.48b <0.001
谷氨酰转移酶 GGT/(ng/mL) 575.80±2.84b 578.33±1.05a 575.59±2.48b 0.021

2.5 花青素对FB1攻毒蛋鸡肝脏组织病理学的影响

由图1所示,与对照组相比,伏马毒素组蛋鸡的肝细胞严重坏死、淋巴细胞肿大,肝细胞损伤严重;与伏马毒素组相比,缓解组蛋鸡的淋巴细胞体积减小,肝细胞损伤症状减轻。
图1 肝脏组织病理学变化(HE染色)

箭头a指示的是变性的肝细胞,箭头b指示的是淋巴细胞。

Fig.1 Histopathological changes of liver (HE staining, 400×)

Arrow a indicates degenerated hepatocytes and arrow b indicates lymphocytes.

2.6 花青素对FB1攻毒蛋鸡盲肠菌群结构的影响

2.6.1 盲肠内容物有效操作分类单元(OTU)数统计

由图2可知,各组间共有的OTU数为1 578个,对照组特有的OTU数为423个,伏马毒素组特有的OTU数为443个,缓解组特有的OTU数为389个。
图2 盲肠内容物OTU维恩图

A为对照组,B为伏马毒素组,C为缓解组。下图同。

Fig.2 OTU Venn diagram of cecal content

A is the control group, B is the fumonisin group, and C is the remission group. The same as below.

2.6.2 盲肠菌群相对丰度

由图3可知,各组盲肠菌群中相对丰度大于1%的菌门有拟杆菌门(Bacteroidota)、厚壁菌门(Firmicutes)、变形菌门(Proteobacteria)、放线菌门(Actinobacteriota)、脱硫菌门(Desulfobacterota)。由表6可知,与对照组相比,伏马毒素组拟杆菌门的相对丰度显著降低(P<0.05),且变形菌门的相对丰度显著升高(P<0.05)。与伏马毒素组相比,缓解组拟杆菌门的相对丰度显著升高(P<0.05),且变形菌门的相对丰度有下降的趋势。
图3 门水平上盲肠菌群组成

Bacteroidota:拟杆菌门;Firmicutes:厚壁菌门;Proteobacteria:变形菌门; Actinobacteriota放线菌门;Desulfobacterota:脱硫菌门;Spirochaetota:螺旋菌门;Patescibacteria:髌骨细菌门;Halobacterota:盐古菌门;Verrucomicrobiota:疣微菌门;Deferribacterota:脱铁杆菌门;Other:其他;Unclassified:未分类。

Fig.3 Composition of cecal microflora at phylum level

表6 花青素FB1攻毒蛋鸡盲肠菌群在门水平相对丰度的影响

Table 6 Effects of anthocyanin on relative abundance of cecal microflora at phylum level of laying hens challenged with FB1%

项目
Items		 对照组
Control group		 伏马毒素组
Fumonisin group		 缓解组
Remission group		 P值
P-value
拟杆菌门 Bacteroidota 49.74±3.95a 44.61±4.27b 48.76±2.64a 0.026
厚壁菌门 Firmicutes 30.75±2.92 41.51±4.13 40.29±2.37 0.750
变形菌门 Proteobacteria 0.78±0.37b 4.96±4.49a 2.63±1.23ab 0.019
放线菌门 Actinobacteriota 2.69±1.19 2.52±0.82 2.01±0.70 0.326
脱硫菌门 Desulfobacterota 1.35±0.50 1.19±0.24 1.34±0.60 0.748
螺旋菌门 Spirochaetota 0.89±0.30 0.83±0.21 1.03±0.41 0.457
髌骨细菌门 Patescibacteria 0.36±0.15 0.69±0.42 0.51±0.40 0.177
盐古菌门 Halobacterota 0.19±0.17 0.43±0.37 0.29±0.26 0.250
疣微菌门 Verrucomicrobiota 0.20±0.20 0.40±0.41 0.09±0.03 0.088
脱铁杆菌门 Deferribacterota 0.22±0.16 0.09±0.06 0.30±0.33 0.171
图4显示的是属水平上盲肠菌群组成,在各组盲肠内容物中共检测出10个共有菌属,其中拟杆菌属(Bacteroides)为优势菌属。由表7可知,与对照组相比,伏马毒素组拟杆菌属的相对丰度有下降的趋势,乳杆菌属的相对丰度有上升的趋势。与伏马毒素组相比,缓解组拟杆菌属的相对丰度显著升高(P<0.05),乳杆菌属的相对丰度有升高的趋势。
图4 属水平上盲肠菌群组成

Bacteroides:拟杆菌属;Rikenellaceae_RC9_gut_group:理研菌科RC9肠道群;Faecalibacterium:粪杆菌属;Lactobacillus:乳杆菌属;Olsenella:欧陆森氏菌属;Ruminococcus_torques_group:瘤胃球菌属扭链群;Phascolarctobacterium:考拉杆菌属;Prevotellaceae_UCG-001:普雷沃氏菌科UCG-001;Erysipelatoclostridium:红斑丹毒丝菌属;Desulfovibrio:脱硫弧菌属;Other:其他;Unclassified:未分类。

Fig.4 Composition of cecal microflora at genus level

表7 花青素对FB1攻毒蛋鸡盲肠菌群在属水平相对丰度的影响

Table 7 Effects of anthocyanin on relative abundance of cecal microflora at genus level of laying hens challenged with FB1%

项目
Items		 对照组
Control group		 伏马毒素组
Fumonisin group		 缓解组
Remission group		 P值
P-value
拟杆菌属 Bacteroides 19.64±3.48ab 17.18±2.86b 21.25±2.69a 0.044
理研菌科RC9肠道群
Rikenellaceae_RC9_gut_group		 9.49±0.96 7.86±1.30 8.64±1.97 0.109
粪杆菌属 Faecalibacterium 3.22±0.56 3.13±0.90 3.19±0.90 0.976
乳杆菌属 Lactobacillus 2.28±1.25 2.32±1.67 4.54±2.96 0.068
欧陆森氏菌属 Olsenella 2.42±1.19 2.06±0.75 1.65±0.69 0.251
瘤胃球菌属扭链群 Ruminococcus_torques_group 1.67±0.19 1.58±0.50 1.86±0.63 0.481
考拉杆菌属 Phascolarctobacterium 2.04±1.22 1.41±0.55 1.35±0.45 0.195
普雷沃氏菌科UCG-001 Prevotellaceae_UCG-001 1.74±0.52 1.62±0.60 1.42±0.23 0.401
红斑丹毒丝菌属 Erysipelatoclostridium 1.35±0.42 1.23±0.66 1.35±0.49 0.880
脱硫弧菌属 Desulfovibrio 1.34±0.50 1.17±0.23 1.32±0.60 0.747

3 讨论

3.1 花青素对FB1攻毒蛋鸡生产性能的影响

饲料发霉后会产生一定量的霉菌毒素,过量的霉菌毒素尤其是FB1会影响家禽的生产性能,如饲料转化率、平均日采食量等。Ruan等[12]研究发现,与对照组相比,饲粮中添加酶菌毒素显著降低了蛋鸡1~8周和9~12周的蛋重、产蛋率,并且增加了料蛋比。本试验结果显示,与对照组相比,伏马毒素组蛋鸡的平均日采食量与平均蛋重有下降的趋势,与前者一致,但料蛋比差异不显著;与伏马毒素组相比,缓解组蛋鸡的平均日采食量与平均蛋重显著升高,推测花青素具有缓解FB1所造成的蛋鸡生产性能下降的作用。

3.2 花青素对FB1攻毒蛋鸡抗氧化能力的影响

机体在遭受各种有害刺激时会诱导细胞发生氧化应激[13]。氧化应激会产生大量ROS,而ROS过多的积累会导致抗氧化系统失衡,造成机体细胞内脂质过氧化,出现氧化损伤。MDA是脂质过氧化的终产物,作为反映氧化应激的指标之一,其含量升高标志着氧化损伤的发生[14-15]。本研究结果显示,伏马毒素组蛋鸡血浆中MDA含量显著升高,说明FB1可以导致蛋鸡机体氧化应激。SOD活性的增强与自由基清除能力的提升呈正相关;CAT可催化超氧自由基生成过氧化氢来降低ROS水平,而且具备解毒和修复细胞损伤的能力;GSH-Px可以消除细胞外的过氧化氢,参与谷胱甘肽的运输,催化脂质过氧化物的分解并生成相应的醇,从而减轻过氧化物对机体的损害[16-17]。在抗氧化防御系统中,SOD将游离的氧复原为过氧化氢,再经CAT将过氧化氢转化为水和氧气,此过程中GSH-Px可与SOD协同作用,将其转化为无毒的羟基化合物,防止游离的氧沉积与脂质过氧化物MDA的形成[18]。本研究结果显示,与对照组相比,FB1显著降低了蛋鸡血浆中GSH-Px、SOD和CAT活性,严重破坏了蛋鸡的抗氧化能力,从而导致机体内的MDA累积,血浆中MDA含量显著升高。姚中磊等[19]研究表明,在饲粮中添加100 mg/kg花青素可显著提高肉鸡血清中SOD、CAT活性与T-AOC,并且显著降低MDA含量。在体外的研究显示,黑米花青素通过抑制ROS的积累和增加SOD和CAT的活性,减轻过氧化氢诱导的PC12细胞损伤[20]。牟春堂等[21]研究发现,饲粮中添加葡萄籽原花青素可显著提升育肥绵羊肝脏中GSH-Px活性并且显著降低MDA含量。本试验结果表明,与伏马毒素组对比,缓解组蛋鸡血浆中GSH-Px、SOD和CAT活性显著提高,且MDA含量显著降低,与前人研究结果一致,表明花青素能够显著改善FB1对蛋鸡造成的氧化应激损伤,推测花青素与其他天然抗氧化剂有类似功效。

3.3 花青素对FB1攻毒蛋鸡免疫功能的影响

免疫球蛋白在体内起防御作用,IgA、IgM和IgG是禽类重要的免疫因子[22]。家禽先天免疫系统中的淋巴细胞能够产生多种抗体,如IgM、IgG和IgA,作为免疫的第1道防线[23]。IgA参与黏膜免疫应答,IgM参与急性感染过程,IgG被认为是初级免疫应答中最持久、最重要的抗体,可提供既往感染的信息[24]。IL-2能够促进TNF-α释放,是评价免疫功能强弱的关键指标。本试验结果显示,伏马毒素组蛋鸡血浆中IgG、IgM含量显著下降,IL-2含量显著升高,表明蛋鸡机体的免疫系统遭受破坏,免疫细胞遭受损伤。董磊等[25]研究发现,饲粮中添加原花青素使母猪血液中IgA、IgG、IgM含量显著升高,并使促炎因子白细胞介素-1(IL-1)、IL-2、干扰素-γ(IFN-γ)含量显著下降。本试验结果显示,与伏马毒素组相比,缓解组蛋鸡血浆中IgG、IgM含量显著升高,IL-2含量有降低的趋势,这与前人研究结果基本一致,表明花青素能够改善FB1给蛋鸡所带来的免疫损伤,证明花青素有潜在的药用作用。

3.4 花青素对FB1攻毒蛋鸡肝脏功能的影响

Sph在肝脏中具有多种生物学功能,其代谢紊乱与肝脏疾病的发生密切相关,通过研究Sph的代谢途径和调控机制,可以为肝脏疾病的治疗提供新的靶点和策略[26]。Qin等[27]研究发现,Sph激酶会增加FB1诱导鞘脂的积累,从而导致Sph含量增加。ALT、AST和GGT是检测肝脏损伤和评估肝脏功能的关键血清生物标志物[28]。ALT和AST主要分布在肝细胞中,是氨基酸与酮酸两者相互关联的关键酶。当肝细胞受损时,其通透性增强,肝细胞中的ALT和AST释放到血液中,导致血液中ALT和AST活性急剧上升[29]。本试验结果显示,试验鸡饲喂含FB1饲粮后,Sph、AST、ALT与GGT含量显著升高,表明FB1对蛋鸡的肝脏造成了一定的损伤。Xu等[30]研究发现,花青素可以下调Sph的含量,进而调节鞘脂代谢。本试验结果表明,与伏马毒素组相比,缓解组蛋鸡血浆中Sph含量显著降低,推测花青素对鞘脂代谢有一定的保护作用,也间接证明了花青素对FB1导致的肝脏损伤具有缓解作用。Cai等[31]研究发现,100 mg/(kg BW·d)的紫薯花青素可使小鼠血清中ALT活性降低,基本恢复到正常水平。Du等[32]研究发现,100 mg/kg蒲公英黄酮可使脂多糖(LPS)攻毒组的乌鳢血清中的ALT与AST活性显著降低。本试验结果显示,缓解组蛋鸡血浆中ALT、AST、GGT活性可降低至对照组的水平,推测花青素可以缓解FB1所导致的蛋鸡肝脏损伤。

3.5 花青素对FB1攻毒蛋鸡肝脏组织病理学的影响

真菌毒素的代谢和积累主要发生在肝脏,其在清除毒素、产生蛋白质和维持代谢稳态等方面起着核心作用,在机体的氧化应激与免疫应激方面也是不可或缺的重要器官[33-34]。真菌毒素会改变肝脏形态,诱导炎症,增加细胞凋亡和氧化应激[35]。本试验研究表明,与对照组相比,伏马毒素组蛋鸡的肝细胞变性严重,并伴有淋巴细胞的增加,表明FB1对肝细胞造成了一定的破坏。有研究表明,黄酮类化合物可以减轻肝脏的炎症症状[36]。本研究结果表明,蛋鸡饲粮中添加150 mg/kg花青素可以缓解FB1所带来的肝脏坏死的情况,说明花青素具有保护肝脏的潜在药用价值。

3.6 花青素对FB1攻毒蛋鸡盲肠菌群结构的影响

肠道菌群对机体的影响极其复杂,免疫系统、神经系统和肝脏疾病与肠道菌群息息相关,可以通过添加特定的药物或者添加剂来改善机体肠道健康[37-38]。研究表明,黄酮类物质可以调节畜禽的肠道菌群结构,抑制有害菌生长,改善肠道健康[39-41]。Zhao等[40]研究发现,黄酮类物质可以改善厚壁菌门和拟杆菌门等微生物的比例,从而改善肠道健康。拟杆菌属是拟杆菌门的一个关键属,其有助消化、产能、维持倡导稳态等功能。本试验结果显示,与伏马毒素组相比,饲粮中添加150 mg/kg花青素可以显著提高盲肠中拟杆菌门与拟杆菌属的相对丰度,这说明花青素提高了蛋鸡肠道中有益菌的占比,改善了肠道健康。在本试验中,变形菌门为蛋鸡盲肠中第三优势菌门,其含有较多的致病菌,运用一定的手段降低变形菌门的相对丰度有利于肠道健康[42]。Wang等[43]研究发现,蔓越莓花青素能够降低小鼠肠道中变形菌门的相对丰度。在本试验中,与伏马毒素组相比,饲粮中添加150 mg/kg花青素有降低蛋鸡盲肠中变形菌门相对丰度的趋势,说明花青素抑制了蛋鸡肠道中有害菌的繁殖。乳杆菌属是厚壁菌门中的一种革兰氏阳性菌,其大多数为益生菌,可利用与消耗多糖[44]。乳杆菌属作为机体重要的菌群,其菌株可以相互黏附或与其他菌株黏附形成聚集,这与生物膜的形成、黏附、定植和宿主生理功能密切相关[45]。Wang等[43]的研究表明,饲粮中添加蔓越莓花青素可以增加小鼠肠道中乳杆菌属的相对丰度。本试验结果显示,与伏马毒素组相比,饲粮中添加150 mg/kg花青素可以显著提高蛋鸡盲肠中乳杆菌属的相对丰度,说明花青素可以增加蛋鸡肠道菌群有益菌的占比,改善肠道健康。

4 结论

饲粮中添加150 mg/kg的花青素提高了FB1攻毒蛋鸡的平均日采食量与平均蛋重,增加了肠道中有益菌占比,减少了有害菌占比,增强了血浆中抗氧化酶活性以及免疫球蛋白含量,使肝细胞与血液中功能酶活性趋于稳定,有效缓解了FB1所导致的蛋鸡氧化应激、免疫损伤以及肝脏损伤。

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