Effect Of Supplementation Of Vitamin D₃ In Diet On The Expression Of Antimicrobial Peptides Genes In Chicken

Antimicrobial peptide,a kind of polypeptide substance which has antibiotic activity, is a significant effector molecule of the animal immune system's function execution,and takes part in the animals' innate immunity and adaptive immunity.A further understanding on the chick's defense mechanism of different organization in different physiological situation at different growth stage could be obtained by studying the expression time sequence and tissue specificity of antimicrobial peptide.Enhancing the expression ability of the organ's entogenous antimicrobial peptide by means of allogenic material,so as to increase the defense ability,and provide new method for the healthy cultivation of animal husbandry.The test detected the expression distribution of chick's AvBD-1 and fowlicidin-1,2,3 gene in spleen,uropygial gland,liver,kidney and bursa of fabricius by RT-PCR(Reverse Transcript-Polymerase Chain Reaction).The result showed that there had expression of spleen,uropygial gland,liver,kidney,thymus gland and bursa of fabricius of the Sichuan mountains Silky fouls,and the expression amount of AvBD-1(Avian beta-defensin-1),fowlicidin-3 gene in the spleen,bursa of fabricius,thymus gland,uropygial gland is higher than that in liver and spleen.Fowlicidin-1,2 had most expression amount in spleen, uropygial gland,liver,kidney,thymus gland and bursa of fabricius.Different does of Vitamin D₃(VD₃) added into silky fouls' daily,and then detected the expression amount of antibiotic peptide AvBD-1and fowlicidin-1 mRNA by real-time quantitation RT-PCR.The result showed that compared to group A(not added VD₃),added 800 IU/kg VD₃ to 6400 IU/kg VD₃ could enhance the expression of AvBD-1,fowlicidin-1 gene in bursa of Fabricius(P＜0.01),and between 800 to 3200 IU/kg addition,the expression ability of AvBD-1,fowlicidin-1 in bursa of Fabricius increased when rising of addition amount.In thymus gland,compared to group A,which had not added VD₃,the addition of 3200～6400 IU/kg could enhance the expression amount of AvBD-1 gene(P＜0.01),and the expression amount was most.However,the addition of 800,1600 IU/kg VD₃ had no notable difference as compared to control group on the expression of AvBD-1 gene in thymus gland.As to fowlicidin-1 gene,the highest expression ability in thymus gland is the addition of 1600 IU/kg VD3,which had remarkably difference when compared to other groups(P＜0.01).Compared to the control group A,the addition of 3200 IU/kg,6400 IU/kg vitamin D₃ could increase(P＜0.01);there was no remarkable difference between 0 IU/kg and 800 IU/kg dose group,When compared to 3200 IU/kg and 6400 IU/kg dose group.Thus,adding proper dose of VD₃ could enhance the expression of antibacterial peptide in bursa of Fabricius and thymus gland of silky foul in mountainous region in Sichuan province.To learn much more whether the expression regulation of VD₃ of chick's antibacterial peptide was related to VDR,The test detected the influence of different dose of VD₃ on the expression ability of VDR in chick's fabricius gland by real-time RT-PCR.The result showed that compared to other groups(0,800,3200,6400 IU/kg),the addition of 1600 IU/kg vitamin D₃ could enhance the expression ability of VDR gene in fabricius gland(P＜0.01),while there was no notable difference of VDR gene expression in other VD₃ dose groups(P＞0.05).There was no remarkable difference between the expression ability of VDR mRNA and that of AvBD-1,fowlicidin-1 gene mRNA(P＞0.05).Furthermore,the analysis on special sequence of promoter sequence of antibacterial peptide fowlicidin-1 gene of silky foul in mountainous region in Sichuan province indicated that there was no transcription factor binding site of VDR/RXR.Therefore,it still couldn't prove that the addition of VD₃ in the diet,stimulating the expression of foul's endogenous antibacterial peptide,was mediated by VDR and regulated by the VDRE in the promoter sequence of antibacterial peptide gene.The analysis on the special sequence of promoter sequence indicated that there existed not only several transcription factor binding sites(NF-κβ,Ap-1,NF-IL6,Sp1 etc.) related to immunological reaction and inflammatory reaction,but also transcription factor binding sites,such as SREBP-1(sterol regulatory element-binding protein 1,which is the significant regulation and control factor),ER(estrogen hormone receptor),HSF(heat shock factor) and so like.