葡萄籽原花青素对饲喂高精料绵羊结肠上皮屏障的影响及作用机制研究

doi:10.16431/j.cnki.1671-7236.2024.10.008

摘要/Abstract

摘要： 【目的】探究葡萄籽原花青素(GSPs)对高精料绵羊结肠上皮屏障的影响及通过Nrf2/ARE通路保护结肠上皮损伤的作用机制。【方法】选取48只120日龄、体重(22.75 kg±1.20 kg)相近的杜寒F1代杂交公羔羊，随机分为4组，每组12只羊。各组试验羔羊基础饲粮中GSPs的添加量分别为0(对照组，CON)、10(10GSPs组)、20(20GSPs组)、40(40GSPs组)mg/kg BW。试验期60 d，其中预试期15 d，正试期45 d。正试期结束次日早晨每组屠宰6只羊，采集结肠组织进行形态学和炎性细胞浸润分析，检测结肠组织白细胞介素2(IL-2)、IL-4、前列腺素E₂(PGE₂)等炎症因子水平及核因子E2相关因子2(Nrf2)、血红素氧合酶1(HO-1)、谷胱甘肽过氧化物酶4(GPx4)等相关基因和蛋白表达量。【结果】 ①对照组羔羊结肠黏膜上皮细胞严重缺失，固有层裸露，肠腺排列疏松，间隙增宽，紧密连接蛋白缺失、受损，而20GSPs和40GSPs组羔羊结肠形态结构和紧密连接蛋白分布表达正常；②随着GSPs添加量的升高，羔羊结肠组织髓过氧化物酶(MPO)、诱导型一氧化氮合酶(iNOS)酶活性及一氧化氮(NO)、PGE2、IL-2水平线性降低(P＜0.05)；③与对照组相比，20GSPs和40GSPs组羔羊结肠组织中Nrf2、OH-1基因和蛋白表达量均显著升高(P＜0.05)，40GSPs组结肠组织中GPx4基因表达量升高(P＜0.05)；④与对照组相比，20GSPs和40GSPs组羔羊结肠组织中Toll-样受体4(TLR4)基因和蛋白表达量均显著降低(P＜0.05)，40GSPs组结肠组织中核因子κB(NF-κB)P65基因和蛋白表达量均显著降低(P＜0.05)；20GSPs和40GSPs组羔羊结肠组织中肿瘤坏死因子α(TNF-α)基因和蛋白及IL-1β蛋白表达量均显著降低(P＜0.05)。【结论】 GSPs可以通过激活Nrf2通路，抑制NF-κB炎症信号通路活化，降低结肠组织局部炎症，缓解饲喂高精料绵羊结肠上皮组织损伤。

关键词: 葡萄籽原花青素; 高精料; 绵羊; 结肠上皮; 炎症损伤; 信号通路

Abstract: 【Objective】 This study was conducted to investigate the effects of grape seed proanthocyanidins (GSPs) on colon epithelial barrier of sheep fed a high-concentrate diet and the mechanism of alleviating colon epithelial injury through Nrf2/ARE pathway.【Method】 A total of forty-eight 4-month-old 1/2 Dorper × 1/2 thin-tailed Han crossed ram lambs with similar body weight (22.75 kg±1.20 kg) were randomly assigned into four experimental treatment groups，with 12 lambs in each group. Lambs in each group were supplemented with the basal diets of 0 (CON group)，10(10GSPs group)，20(20GSPs group) and 40 mg/kg BW GSPs (40GSPs group)，respectively.The feeding experiment lasted for 60 days，including 15 days adaptation period and 45 days experimental period.On the morning after the trial period ended，6 sheep in each group were slaughtered and colon tissues were collected for morphological and inflammatory cell infiltration analysis.The levels of interleukin-2 (IL-2),IL-4,prostaglandin E₂ (PGE₂) and other inflammatory factors，and the expression levels of nuclear factor-like-2 factor (Nrf2)，heme oxygenase-1 (HO-1)，glutathione peroxidase 4 (GPx4) and other related genes and proteins were detected in colon tissues.【Result】 ①In CON group，colon mucosal epithelial cells were seriously missing，lamina propria was exposed，intestinal gland arrangement was loose，spaces were widened，tight junction protein was missing and damaged. Whereas，the colonic morphological structure and tight junction protein distribution was normal in 20GSPs and 40GSPs groups.②The enzyme activities of myeloperoxidase (MPO) and inducible nitric oxide synthase (iNOS)，and the concentrations of nitric oxide (NO)，IL-2 in colon tissues were linearly decreased with the increase of GSPs supplementation (P＜0.05).③Compared with CON group，the gene and protein expression of Nrf2 and HO-1 in colon tissues in 20GSPs and 40GSPs groups were significantly increased (P＜0.05)，and the expression of GPx4 gene in colon tissues in 40GSPs group was significantly increased (P＜0.05).④Compared with CON group，the gene and protein expression of Toll-like receptor 4 (TLR4) in colon tissues of lambs in 20GSPs and 40GSPs groups were significantly decreased，and the gene and nuclear protein expression of nuclear factor kappa B (NF-κB) p65 in colon tissues of lambs in 40GSPs group were significantly decreased (P＜0.05). The gene and protein expression of tumor necrosis factor α (TNF-α) and protein expression levels of IL-1β in colon tissues of lambs in 20GSPs and 40GSPs groups were also significantly lower than those in CON group (P＜0.05).【Conclusion】 GSPs could reduce local inflammation of colon tissue and alleviate colon epithelial tissue damage by activating Nrf2 pathway and inhibiting the activation of NF-κB inflammatory signaling pathway in sheep fed high-concentrate diet.

Key words: grape seed procyanidins; high-concentrate diet; sheep; colonic epithelium; inflammatory injury; signaling pathway

中图分类号:

S816.7

牟春堂, 魏焕, 谢蕴哲, 张建新, 郝小燕. 葡萄籽原花青素对饲喂高精料绵羊结肠上皮屏障的影响及作用机制研究[J]. 中国畜牧兽医, 2024, 51(10): 4270-4281.

MU Chuntang, WEI Huan, XIE Yunzhe, ZHANG Jianxin, HAO Xiaoyan. Effect of Grape Seed Proanthocyanidins on Colon Epithelial Barrier of Sheep Fed a High-Concentrate Diet and Its Mechanism[J]. China Animal Husbandry and Veterinary Medicine, 2024, 51(10): 4270-4281.

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