Animal Nutrition Institute, Sichuan Agricultural University, Chengdu, 611130, China; Fish Nutrition and Safety Production University Key Laboratory of Sichuan Province, Sichuan Agricultural University, Chengdu, 611130, China; Key Laboratory for Animal Disease-Resistance Nutrition of China Ministry of Education, Sichuan Agricultural University, Chengdu, 611130, China.
Animal Nutrition Institute, Sichuan Agricultural University, Chengdu, 611130, China.
Fish Shellfish Immunol. 2019 Apr;87:546-558. doi: 10.1016/j.fsi.2019.01.056. Epub 2019 Feb 1.
The present study was the first to investigate the effects of dietary vitamin A (VA) on the intestinal physical barrier function associated with oxidation, antioxidant system, apoptosis and cell-cellular tight junction (TJ) in the proximal (PI), mid (MI) and distal (DI) intestines of young grass carp (Ctenopharyngodon idella). Fish were fed graded levels of dietary VA for 10 weeks, and then a challenge test using an injection of Aeromonas hydrophila was conducted for 14 days. Results indicated that dietary VA deficiency caused oxidative damage to fish intestine partly by the reduced non-enzymatic antioxidant components glutathione (GSH) and VA contents as well as reduced antioxidant enzyme activities [not including manganese superoxide dismutase (MnSOD)]. Further results observed that the decreased antioxidant enzyme activities by VA deficiency were partly related to the down-regulation of their corresponding mRNA levels which were regulated by the down-regulation of NF-E2-related factor 2 (Nrf2) mRNA levels and up-regulation of kelch-like-ECH-associated protein (Keap1a) (rather than Keap1b) mRNA levels in three intestinal segments of fish. Meanwhile, VA deficiency up-regulated the mRNA levels of the apoptosis signalling [caspase-3, caspase-8, caspase-9 (rather than caspase-7)] associated with the inhibition of the target of rapamycin (TOR) signalling pathway in three intestinal segments of fish. Additionally, VA deficiency decreased the mRNA levels of TJ complexes [claudin-b, claudin-c, claudin-3, claudin-12, claudin-15a, occludin and zonula occludens-1 (ZO-1) in the PI, MI and DI, as well as claudin-7 and claudin-11a in the MI and DI] linked to the up-regulation of myosin light chain kinase (MLCK) signalling. These results suggested that VA deficiency impaired structural integrity in three intestinal segments of fish. Meanwhile, excessive VA also showed similar negative effects on these indexes. Taken together, the current study firstly demonstrated that VA deficiency impaired physical barrier functions associated with impaired antioxidant capacity, aggravated cell apoptosis and disrupted TJ complexes in the PI, MI and DI, but different segments performed different actions in fish. Based on protecting fish against protein oxidation, the optimal VA levels for grass carp were estimated to be 2622 IU/kg diet.
本研究首次探讨了饲粮维生素 A（VA）对草鱼近端（PI）、中段（MI）和远端（DI）肠道氧化相关的肠道物理屏障功能、抗氧化系统、细胞凋亡和细胞间紧密连接（TJ）的影响。鱼用含有不同水平 VA 的饲料喂养 10 周，然后进行为期 14 天的嗜水气单胞菌注射攻毒试验。结果表明，饲粮 VA 缺乏导致鱼体肠道氧化损伤，部分原因是非酶抗氧化成分谷胱甘肽（GSH）和 VA 含量以及抗氧化酶活性降低（不包括锰超氧化物歧化酶（MnSOD））。进一步的研究结果表明，VA 缺乏导致抗氧化酶活性降低，部分原因是与 NF-E2 相关因子 2（Nrf2）mRNA 水平下调和Kelch 样 ECH 相关蛋白（Keap1a）（而非 Keap1b）mRNA 水平上调相关，这在鱼体三个肠道段中均下调了相应的 mRNA 水平。同时，VA 缺乏上调了与雷帕霉素（TOR）信号通路抑制相关的细胞凋亡信号（caspase-3、caspase-8、caspase-9（而非 caspase-7））的 mRNA 水平。此外，VA 缺乏降低了 TJ 复合物[PI、MI 和 DI 中的 claudin-b、claudin-c、claudin-3、claudin-12、claudin-15a、occludin 和 zonula occludens-1（ZO-1），以及 MI 和 DI 中的 claudin-7 和 claudin-11a]的 mRNA 水平，同时上调了肌球蛋白轻链激酶（MLCK）信号。这些结果表明，VA 缺乏破坏了鱼体三个肠道段的结构完整性。同时，VA 过量对这些指标也有类似的负面影响。综上所述，本研究首次表明，VA 缺乏破坏了 PI、MI 和 DI 中与抗氧化能力下降、细胞凋亡加剧和 TJ 复合物破坏相关的物理屏障功能，但不同肠道段的作用方式不同。基于保护鱼类免受蛋白质氧化的影响，估计草鱼的最佳 VA 水平为 2622 IU/kg 饲料。
