Effect Of Melaleuca Alternifolia Oil On Small Intestinal Mucosal Morphology And Cellular Immune Function In Broilers

With the continuous development of animal husbandry in China,the illegal addition or even excessive use of antibiotics in feed was very common,which resulted in a series of potential threats,including the production of drug-resistant strains and veterinary drug residues.In the context of the urgent need for high-quality livestock and poultry products,the demand for safe and effective antibiotic substitutes has became a hot research topic for scientists.Melaleuca alternifolia oil is the essential oil steam distilled from the natural plants,melaleuca alterniflolia.It not only has many functions such as antibacterial,anti-inflammatory and anti-virus,but also does not easily lead to bacterial resistance.At present,there are many researches on the use of melaleuca alternifolia oil in the pharmaceutical industry,but less for the livestock and poultry industry.In this study,the effects of melaleuca alternifolia oil on small intestine mucosal morphology,cellular immune function and anti-inflammatory activity were investigated in vivo and in vitro,with the purpose to provide a theoretical basis for its application in broiler breeding.Test one,effects of melaleuca alternifolia oil on small intestinal mucosal morphology and lymphocyte transformation in broilers.The ninety-nine lingnan yellow chickens were divided into three groups.The control group was fed conventionally,while the two experimental groups were fed with 0.012 mL/L and 0.02 mL/L melaleuca alternifolia oil in drinking water.On 15 d,30 d and 45 d,the blood samples were collected and lymphocytes were separated for lymphocyte transformation experiments.On day 45,duodenum,jejunum and ileum samples were collected and paraffin sections were obtained to observe the small intestinal mucosal morphology.The results showed that broilers of two experimental groups had increased peripheral blood lymphocyte transformation rates compared with control group,as well as increased villus height,thinner wall of the intestine,shallower depth of crypts of each small intestine.The ratio of intestinal villi height/crypt depth,the number of intestinal intraepithelial goblet cells and lymphocytes in the two experimental groups was higher than that of the control group.It was concluded that melaleuca alternifolia oil could promote the growth of intestinal villi,thereby improving the morphology of the small intestine,and could enhance the intestinal mucosal immune function and peripheral blood cell immune function.Test two,anti-inflammatory and anti-oxidative effects of melaleuca alternifolia oil in broiler monocytes.Broiler monocytes were isolated and cultured in vitro.The lipopolysaccharide(LPS)was added to the culture medium to stimulate the cells to produce inflammatory reactions and oxidative damage.The effects co-culture of gradient concentration of melaleuca alternifolia oil together with LPS on inflammatory factors and oxidative damage indicators were observed.The mRNA expression of TNF-?,IL-1?,IL-6 and IL-18 were measured by fluorescence quantitative assay.The contents of TNF-?,IL-1?,IL-6 and IL-18 in the cell supernatant were measured by ELISA.The changes of reactive oxygen species(ROS)were measured by flow cytometry.The changes of total antioxidant capacity(T-AOC),malondialdehyde(MDA),catalase(CAT)and superoxide dismutase(SOD)were measured by using antioxidant kit.The results showed that when LPS was added,the mRNA relative expression of TNF-?,IL-1?,IL-6 and IL-18 in the cell and the protein content of TNF-?,IL-1?,IL-6,and IL-18 in the cell supernatant were increased.After the addition of melaleuca alternifolia oil,the mRNA relative expression levels of TNF-?,IL-1?,IL-6 and IL-18 decreased compared with the LPS group,as well as the protein content of TNF-?,IL-1?,IL-6 and IL-18 in the cell supernatant.After stimulating by LPS,the intracellular ROS increased rapidly,while the co-culture of melaleuca alternifolia oil could significantly reduce the increasing of cellular ROS caused by LPS.The antioxidant index of the cells also showed that LPS decreased T-AOC,CAT and SOD activity,and increased MDA concentration.After action of different concentrations of melaleuca alternifolia oil,T-AOC,CAT and SOD activity increased while MDA concentration decreased.The results showed that melaleuca alternifolia oil has anti-inflammatory function by inhibiting the expression of inflammatory factors,and also has anti-oxidation function on monocytes.The results show that melaleuca alternifolia oil could promote the development of small intestine,increase the transformation rate of lymphocytes,and have anti-inflammatory and anti-oxidant function.This study provides a theoretical basis for melaleuca alternifolia oil as feed additive in broiler production.The research on the anti-inflammatory function of melaleuca alternifolia oil on broilers is a blank field,so the results of this study also laid the foundation for further research in the later period.