【摘要】：本試驗(yàn)將純化后的β-伴大豆球蛋白(7S)和大豆球蛋白(11S)以不同劑量添加到斷奶仔豬的日糧中,建立仔豬致敏模型。采用ELISA方法檢測斷奶仔豬血清中的免疫球蛋白,石蠟切片、免疫組織化學(xué)的方法檢測仔豬小腸黏膜上皮緊密連接蛋白ZO-1的表達(dá)與分布,以及實(shí)時(shí)熒光定量PCR方法研究大豆抗原蛋白對斷奶仔豬小腸黏膜上皮緊密連接蛋白ZO-1 mRNA相對表達(dá)量的影響,探討7S和11S對斷奶仔豬血清IgG亞型及Ig E和腸粘膜緊密連接蛋白ZO-1表達(dá)的影響。選取70頭21日齡“杜×長×大”斷奶仔豬,隨機(jī)分成7組,每組10頭仔豬。對照組(A組)飼喂基礎(chǔ)日糧,β-伴大豆球蛋白(β-conglycinin,7S)B、C、D組和大豆球蛋白(Glycinin,11S)E、F、G組在24 d分別添加500μg·kg-1·W-1、2000μg·kg-1·W-1、5000μg·kg-1·W-1的7S或11S。分別于24、26、28、30日齡采集血液,測定血清中免疫球蛋白含量。試驗(yàn)結(jié)束時(shí),每組選取5頭仔豬放血致死,采集各段小腸,其中空腸分前、中、后三段采取,置于4%多聚甲醛和液氮中保存,分別用于檢測仔豬小腸黏膜上皮緊密連接蛋白ZO-1的表達(dá)與分布和檢測各段小腸ZO-1 mRNA的相對表達(dá)量。試驗(yàn)結(jié)果表明:1、大豆抗原蛋白致敏仔豬血清IgG、IgG1、IgG2、IgG3、IgG4和IgE水平升高,其中7S試驗(yàn)組高于11S試驗(yàn)組。2、大豆抗原蛋白致敏仔豬腸黏膜ZO-1的相對表達(dá)量降低,且與7S和11S在仔豬日糧中的添加劑量呈負(fù)相關(guān);添加劑量相同的7S組ZO-1的相對表達(dá)量低于11S組。3、各試驗(yàn)組仔豬腸黏膜緊密連接蛋白ZO-1 mRNA的相對表達(dá)量均降低,與7S和11S在仔豬日糧中的添加劑量呈負(fù)相關(guān)。添加劑量相同的7S組ZO-1 mRNA的相對表達(dá)量低于11S組。試驗(yàn)證實(shí),β-伴大豆球蛋白和大豆球蛋對斷奶仔豬有明顯的致敏作用,β-伴大豆球蛋白致敏性高于大豆球蛋白;β-伴大豆球蛋白和大豆球蛋顯著提高致敏仔豬血清中的免疫球蛋白IgG、IgG1、IgG2、IgG3、IgG4和IgE水平,降低腸粘膜上皮緊密連接蛋白ZO-1相對表達(dá)量以及ZO-1 mRNA相對表達(dá)量。
[Abstract]:In this experiment, purified 尾 -concomitant soybean globulin (7s) and soybean globulin (11s) were added to weaned piglets' diets in different doses to establish sensitizing model of piglets. The expression and distribution of tight junction protein (ZO-1) in small intestinal mucosal epithelium of weaned piglets were detected by ELISA method, paraffin section and immunohistochemical method. The effect of soybean antigen protein on the expression of tight junction protein (ZO-1 mRNA) in small intestinal mucosa of weaned piglets was studied by real-time fluorescence quantitative PCR. To investigate the effects of 7s and 11s on serum IgG subtypes and the expression of Ig E and intestinal mucosal tight junction protein (ZO-1) in weaned piglets. 70 21 day-old weaned piglets were randomly divided into 7 groups with 10 piglets in each group. The control group (group A) was fed with basal diet, 尾 -concomitant soybean globulin (尾-conglycinin,7S) group D and soybean globulin (Glycinin,11S) estradiol. Group G was treated with 渭 g kg-1 W-1G 2000 渭 g kg-1 W-1N 5000 渭 g kg-1 W-1 7s or 11s on the 24th day, respectively. Blood samples were collected at the age of 2830 days, and the content of immunoglobulin in serum was determined. At the end of the experiment, 5 piglets in each group were selected to bleed to death, and the small intestine was collected, in which the jejunum was divided into three sections: anterior, middle and posterior, and stored in 4% paraformaldehyde and liquid nitrogen. It was used to detect the expression and distribution of tight junction protein (ZO-1) in intestinal mucosal epithelium of piglets and to detect the relative expression of ZO-1 mRNA in small intestine. The results showed that: 1. The levels of serum IgG,IgG1,IgG2,IgG3,IgG4 and IgE increased in soybean antigen protein-sensitized piglets, and the relative expression of ZO-1 in intestinal mucosa of 7s group was higher than that of 11s test group, and the relative expression of ZO-1 in intestinal mucosa of soybean antigen protein-sensitized piglets was lower than that of 11s test group. There was a negative correlation between 7s and 11s in the diet of piglets. The relative expression of ZO-1 in 7s group with the same additive content was lower than that in 11s group. 3. The relative expression of ZO-1 mRNA in intestinal mucosal tight junction protein was decreased in all experimental groups, which was negatively correlated with the additive amount of 7s and 11s in diet of piglets. The relative expression of ZO-1 mRNA in 7s group with the same additive content was lower than that in 11s group. The results showed that 尾 -concomitant soybean globulin and soybean ball egg could sensitize weaned piglets, and 尾 -concomitant soybean globulin was more sensitive than soybean globulin. 尾 -concomitant soybean globulin and soybean ball egg significantly increased the levels of immunoglobulin IgG,IgG1,IgG2,IgG3,IgG4 and IgE in serum of sensitized piglets, and decreased the relative expression of ZO-1 and ZO-1 mRNA in intestinal epithelium.
【學(xué)位授予單位】：安徽農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S828

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