葡萄糖浓度对猪小肠上皮细胞精氨酸吸收、转运的影响

发布时间：2018-05-07 16:45

本文选题：葡萄糖 + 精氨酸　；参考：《吉林农业大学》2017年硕士论文

【摘要】：为了探讨葡萄糖浓度对猪小肠上皮细胞精氨酸吸收、转运的影响。本试验以仔猪小肠上皮细胞(IPEC-J2)系为体外培养模型,选取生长状态良好的(IPEC-J2)细胞,待细胞融合度达到90%时,将细胞在缺乏氨基酸的EBSS平衡盐溶液和维生素混合的培养基中饥饿2h。然后向葡萄糖浓度为25mM/L,75mM/L,225mM/L的三种培养基中分别加入0mM/L、0.7mM/L、1.4mM/L、2.1mM/L、4.2mM/L精氨酸,配制成15种葡萄糖和精氨酸混合培养基,分别处理细胞20,40,80,160min,通过细胞指数、精氨酸吸收转运体基因及蛋白质表达量的测定。试验结果如下:1精氨酸及葡萄糖对细胞增殖的影响。试验结果表明,添加精氨酸、葡萄糖对细胞增殖有明显的促进作用,且二者之间存在着极显著的互作效应(P0.01)。葡萄糖浓度一定时,随着精氨酸浓度的增加,细胞指数呈现了先升高后降低的趋势,在精氨酸浓度为1.4mM/L、2.1mM/L时对细胞增殖的促进作用较显著(P0.05);精氨酸浓度一定时,随着葡萄糖浓度的增加,细胞指数也呈现了先升高后降低的趋势,且在葡萄糖浓度在75mM/L时,对细胞增殖有显著的促进作用(P0.05)。2精氨酸吸收、转运载体mRNA表达的测定结果表明:葡萄糖和精氨酸浓度对精氨酸吸收、转运载体的表达均有显著影响((P0.05),而且葡萄糖和精氨酸浓度在精氨酸吸收、转运体的mRNA表达上表现出了极显著的互作效应(P0.01)。细胞处理40min时,葡萄糖浓度为75mM/L,精氨酸浓度为2.1mM/L,显著提高了精氨酸吸收载体b0,+AT mRNA相对表达量(P0.05);细胞处理至80min时,葡萄糖浓度为75mM/L,精氨酸浓度为2.1m M/L,显著提高了精氨酸转运载体y+LAT1 mRNA相对表达量(P0.05)。3根据试验2得出的结果,选取试验2中最适精氨酸吸收、转运载体mRNA相对表达量的葡萄糖浓度及时间,检测精氨酸转运载体的蛋白相对表达量。蛋白质表达水平层面的试验表明:精氨酸吸收、转运载体的蛋白相对表达量与mRNA表达量的变化规律基本一致。即处理细胞40min时,精氨酸浓度为2.1mM/L,显著提高了精氨酸吸收载体b0,+AT蛋白相对表达量(P0.05);细胞处理至80min时,精氨酸浓度为2.1mM/L,显著提高了精氨酸转运载体y+LAT1蛋白相对表达量(P0.05)。综上所述,葡萄糖、精氨酸浓度对仔猪小肠上皮细胞的增殖,精氨酸吸收、转运载体基因的表达均具有显著影响,而且葡萄糖和精氨酸浓度在细胞增殖以及精氨酸吸收、转运体的mRNA表达上均表现出了极显著的互作效应。本试验中葡萄糖浓度75mM/L,精氨酸浓度2.1mM/L,能够显著促进猪小肠上皮细胞(IPEC-J2)吸收、转运体b0,+AT和y+LAT1的基因表达及蛋白合成。
[Abstract]:To investigate the effect of glucose concentration on arginine uptake and transport in porcine intestinal epithelial cells. In this experiment, the porcine intestinal epithelial cell line IPEC-J22 was used as the culture model in vitro. When the cell fusion degree reached 90%, the cells were starved for 2 h in the medium containing EBSS balanced salt solution and vitamins lacking amino acids. And then to the three kinds of media with glucose concentration of 25 mm / L, 75 mm / L, 225 mm / L, respectively, adding 0 mm / L 0. 7 mm / L 1. 4 mm / L L ~ (2. 1) mm / L L ~ (2) 2 mm / L arginine, and then preparing a mixed medium of 15 kinds of glucose and arginine, and treating the cells for 20, 40, 80, 160 mins, respectively, through cell index, Determination of arginine absorption transporter gene and protein expression. The results were as follows: the effects of arginine and glucose on cell proliferation. The results showed that the addition of arginine could significantly promote the proliferation of the cells, and there was a significant interaction effect between the two. When glucose concentration was constant, the cell index increased first and then decreased with the increase of arginine concentration. When the concentration of arginine was 1.4 mm 路L ~ (-1) L ~ (-1) mg / L, the cell proliferation was significantly promoted (P _ (0.05), and when the concentration of arginine was constant, With the increase of glucose concentration, the cell index increased first and then decreased. When glucose concentration was at 75mM/L, the cell proliferation was significantly promoted by P0.050.2. arginine absorption. The results of mRNA expression of transport vector showed that the concentration of glucose and arginine had a significant effect on the absorption of arginine, and the expression of transport vector had a significant effect on the absorption of arginine, and the concentration of glucose and arginine was absorbed in arginine. The mRNA expression of the transporter showed a significant interaction effect (P0.01). When cells treated with 40min, glucose concentration was 75 mm / L, arginine concentration was 2.1 mm / L, which significantly increased the relative expression of arginine absorption vector b0, AT mRNA, P0.05N, and increased the relative expression of AT mRNA to 80min. Glucose concentration was 75 mm / L, arginine concentration was 2.1 m / L, which significantly increased the relative expression of arginine transport vector y LAT1 mRNA. According to the results of experiment 2, the optimum arginine absorption was selected. The glucose concentration and time of the relative expression of mRNA and the relative protein expression of arginine transport vector were detected. The results of protein expression level showed that the relative expression of arginine and transport vector was consistent with that of mRNA. When treated with 40min, arginine concentration was 2.1 mm 路L ~ (-1), which significantly increased the relative expression of arginine absorption vector b0 and AT protein (P0.05), and increased the relative expression of arginine transport vector y LAT1 protein (P0.05) when treated with 80min (2.1 mm 路L ~ (-1). In conclusion, glucose and arginine concentrations have a significant effect on the proliferation, arginine absorption and expression of transport vector genes of small intestinal epithelial cells of piglets, and glucose and arginine concentrations have significant effects on cell proliferation and arginine absorption. The mRNA expression of transporter showed significant interaction effect. In this study, glucose concentration of 75mm / L and arginine concentration of 2.1 mm / L significantly promoted the absorption of IPEC-J22, gene expression and protein synthesis of transporters b0, AT and y LAT1 in porcine intestinal epithelial cells.
【学位授予单位】：吉林农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S828

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