發(fā)布時(shí)間：2018-05-07 16:45

  本文選題：葡萄糖 + 精氨酸��； 參考：《吉林農(nóng)業(yè)大學(xué)》2017年碩士論文

【摘要】：為了探討葡萄糖濃度對(duì)豬小腸上皮細(xì)胞精氨酸吸收、轉(zhuǎn)運(yùn)的影響。本試驗(yàn)以仔豬小腸上皮細(xì)胞(IPEC-J2)系為體外培養(yǎng)模型,選取生長(zhǎng)狀態(tài)良好的(IPEC-J2)細(xì)胞,待細(xì)胞融合度達(dá)到90%時(shí),將細(xì)胞在缺乏氨基酸的EBSS平衡鹽溶液和維生素混合的培養(yǎng)基中饑餓2h。然后向葡萄糖濃度為25mM/L,75mM/L,225mM/L的三種培養(yǎng)基中分別加入0mM/L、0.7mM/L、1.4mM/L、2.1mM/L、4.2mM/L精氨酸,配制成15種葡萄糖和精氨酸混合培養(yǎng)基,分別處理細(xì)胞20,40,80,160min,通過(guò)細(xì)胞指數(shù)、精氨酸吸收轉(zhuǎn)運(yùn)體基因及蛋白質(zhì)表達(dá)量的測(cè)定。試驗(yàn)結(jié)果如下:1精氨酸及葡萄糖對(duì)細(xì)胞增殖的影響。試驗(yàn)結(jié)果表明,添加精氨酸、葡萄糖對(duì)細(xì)胞增殖有明顯的促進(jìn)作用,且二者之間存在著極顯著的互作效應(yīng)(P0.01)。葡萄糖濃度一定時(shí),隨著精氨酸濃度的增加,細(xì)胞指數(shù)呈現(xiàn)了先升高后降低的趨勢(shì),在精氨酸濃度為1.4mM/L、2.1mM/L時(shí)對(duì)細(xì)胞增殖的促進(jìn)作用較顯著(P0.05);精氨酸濃度一定時(shí),隨著葡萄糖濃度的增加,細(xì)胞指數(shù)也呈現(xiàn)了先升高后降低的趨勢(shì),且在葡萄糖濃度在75mM/L時(shí),對(duì)細(xì)胞增殖有顯著的促進(jìn)作用(P0.05)。2精氨酸吸收、轉(zhuǎn)運(yùn)載體mRNA表達(dá)的測(cè)定結(jié)果表明:葡萄糖和精氨酸濃度對(duì)精氨酸吸收、轉(zhuǎn)運(yùn)載體的表達(dá)均有顯著影響((P0.05),而且葡萄糖和精氨酸濃度在精氨酸吸收、轉(zhuǎn)運(yùn)體的mRNA表達(dá)上表現(xiàn)出了極顯著的互作效應(yīng)(P0.01)。細(xì)胞處理40min時(shí),葡萄糖濃度為75mM/L,精氨酸濃度為2.1mM/L,顯著提高了精氨酸吸收載體b0,+AT mRNA相對(duì)表達(dá)量(P0.05);細(xì)胞處理至80min時(shí),葡萄糖濃度為75mM/L,精氨酸濃度為2.1m M/L,顯著提高了精氨酸轉(zhuǎn)運(yùn)載體y+LAT1 mRNA相對(duì)表達(dá)量(P0.05)。3根據(jù)試驗(yàn)2得出的結(jié)果,選取試驗(yàn)2中最適精氨酸吸收、轉(zhuǎn)運(yùn)載體mRNA相對(duì)表達(dá)量的葡萄糖濃度及時(shí)間,檢測(cè)精氨酸轉(zhuǎn)運(yùn)載體的蛋白相對(duì)表達(dá)量。蛋白質(zhì)表達(dá)水平層面的試驗(yàn)表明:精氨酸吸收、轉(zhuǎn)運(yùn)載體的蛋白相對(duì)表達(dá)量與mRNA表達(dá)量的變化規(guī)律基本一致。即處理細(xì)胞40min時(shí),精氨酸濃度為2.1mM/L,顯著提高了精氨酸吸收載體b0,+AT蛋白相對(duì)表達(dá)量(P0.05);細(xì)胞處理至80min時(shí),精氨酸濃度為2.1mM/L,顯著提高了精氨酸轉(zhuǎn)運(yùn)載體y+LAT1蛋白相對(duì)表達(dá)量(P0.05)。綜上所述,葡萄糖、精氨酸濃度對(duì)仔豬小腸上皮細(xì)胞的增殖,精氨酸吸收、轉(zhuǎn)運(yùn)載體基因的表達(dá)均具有顯著影響,而且葡萄糖和精氨酸濃度在細(xì)胞增殖以及精氨酸吸收、轉(zhuǎn)運(yùn)體的mRNA表達(dá)上均表現(xiàn)出了極顯著的互作效應(yīng)。本試驗(yàn)中葡萄糖濃度75mM/L,精氨酸濃度2.1mM/L,能夠顯著促進(jìn)豬小腸上皮細(xì)胞(IPEC-J2)吸收、轉(zhuǎn)運(yùn)體b0,+AT和y+LAT1的基因表達(dá)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.
【學(xué)位授予單位】：吉林農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S828

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