| Ammonia is the primary environmental pollution in husbandry with strong pungent oder.Intensive farming model usually makes ammonia concentration to exceed the national standard in inclosed livestock housing in winter.Excessive ammonia in atmosphere affects the human health and the animal welfare level.Pigs can be regarded as important economic animals in agriculture and ideal animal models in comparative medicine.Although comprehensive researches on the toxicological effects of ammonia have been investigated,there was no study confirmed the mechanisms of“microbiota-gut-brain”axis dysfunction induced by ammonia poisoning.“Microbiota-gut-brain”axis involves in immune regulation,nerve communication and so on.As an essential micronutrient for organisms,organic selenium has mitigation effect on ammonia toxicity.Our study established a porcine ammonia-poisoning model and added L-selenomethionine in basal diets to clarity the toxicologic pathogenesis of ammonia on the“microbiota-gut-brain”axis and the molecular mechanisms of mitigation effect of selenium using histomorphology,molecular biology and multi-omics analysis.Twenty-four 125-day-old fattening pigs were divided into four groups on average,called control group（C）,ammonia group（A）,selenium group（Se）,ammonia+selenium group（A+Se）,respectively.On the 30thday,optical/electron microscope were used to observe the pathological changes in the jejunum and hypothalamus,apoptosis characteristics and intestinal villus morphology;commercial kits were used to measure blood ammonia concentration,antiodidant indexes,brain-gut peptides,cytokines,inflammatory factor and the glutamic acid contents.ROS positive density and apoptosis rate were detected quantitatively based on fluorescence-labeling method,meanwhile,apoptosis/immune-related factors were analyzed by immunohistochemistry.High-throughput transcriptome sequencing and 16S r DNA sequencing were used for the target detection of jejunum-hypothalamus transcription profile and enteric microbiota flora.And apoptosis signaling pathway and mitochondrial dynamics-related factors were detected using q RT-PCR and Western blot.Finally,selenium contents in jejunum and hypothalamus were measured by inductively coupled plasma mass spectrometry.These results corroborated that:（1）Excessive ammonia in piggeries had neurotoxicity to fattening pigs.Chronic ammonia exposure could suppress the secretion of the neurotransmitter and diminish the excitability in the central nervous system,leading to the metabolic perturbation in the“microbiota-gut-brain”axis.（2）Excessive ammonia caused the imbalance among mitochondrial dynamics-related factors in jejunum and hypothalamus to induce the mitochondria injury.In addition,ammonia could mediate mitochondrial apoptosis pathway by inhibiting PI3K/Akt/PPAR-γaxis,ptomoting the cell apoptosis process in jejunum and hypothalamus of fattening pigs.（3）Ammonia poisoning would accelerate oxidative stress in jejunum and hypothalamus to mediate Th1/Th2 imbalance and inflammatory lesion.（4）Excessive ammonia could give rise to the changes in the transcription profiles of jejunum/hypothalamus and the disturbance in enteric microbial community.（5）L-selenomethionine could antagonize the neurotoxicity mediated by excessive ammonia through the“microbiota-gut-brain”axis.In conclusion,excessive ammonia inhalation had neurotoxic effects on fattening pigs,which was manifested as ammonia poisoning could alter porcine transcription profiles in the jejunum/hypothalamus and the distribution of intestinal microbes taxa.These process above could induce abnormal physiological process including oxidative stress,immune imbalance and excessive cell apoptosis to mediate inflammation injury in jejunum/hypothalamus.Meanwhile,it showed that ammonia poisoning could cause metabolic disorder in the“microbiota-gut-brain”axis to block the two-way communication between hypothalamus-jejunum network.Additionally,the supplement of L-selenomethionine had obviously mitigative effects on the ammonia toxicity. |
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