肿瘤坏死因子对谷氨酰胺促大鼠小肠蛋白质合成的影响及途径

Inhibitory effect of TNF-α on glutamine-enhanced protein synthesis in the rat small intestine and its pathway

目的谷氨酰胺(glutamine,Gln)需经过载体转运至细胞内才能被利用,感染状态下Gln疗效不佳。肿瘤坏死因子(tumor necrosis factor,TNF-α)是感染中重要的调节因子,故研究TNF-α与Gln促蛋白质合成及其与Gln载体之间的关系对揭示感染状态下Gln代谢途径、提高疗效具有重要意义。文章研究TNF-α对Gln促大鼠小肠蛋白质合成及其对细胞膜Gln载体ASCT2mRNA、蛋白表达的影响,揭示TNF-α影响Gln促蛋白质合成的途径。方法 30只雄性SD大鼠按单纯随机法分3组:对照组、Gln组和TNF-α组。所有大鼠均行3 d全肠外静脉营养。对照组仅给予普通全肠外营养;Gln组给予添加丙氨酰谷氨酰胺二肽的肠外营养,Gln剂量为0.3 g/(kg·d),计72 h;TNF-α组给予添加丙氨酰谷氨酰胺二肽的肠外营养,并在第3天持续24h静脉滴注TNF-α,速度5μg/(kg·h)。所有大鼠均在取材前0.5h一次性静脉注射[L-15N]亮氨酸,剂量1.0 mmol/kg。实验结束时分别测定血浆TNF-α、Gln浓度、小肠Gln浓度、蛋白质合成率、ASCT2载体mRNA及蛋白表达水平。结果 TNF-α组血浆TNF-α浓度显著高于其他2组;TNF-α组血浆及小肠中Gln浓度最高;Gln组和TNF-α组小肠中蛋白质合成率均高于对照组,且Gln组最高;以上各组之间差异有统计学意义(P<0.01)。对照组ASCT2的mRNA及蛋白表达均显著低于其他2组(P<0.01)。Gln组ASCT2的蛋白显著高于其他2组(P<0.01)。结论 TNF-α能够升高大鼠血浆及小肠中Gln的浓度,抑制Gln的促蛋白质合成作用,其途径是TNF-α抑制氨基酸载体的转运功能,Gln不能顺利进入细胞,导致组织蛋白质合成下降,这种抑制发生在翻译及以后水平。

Objective The functions of glutamine （Gln） are based on its transport into target cells via transporters in the cytomembrane. Previous studies showed that Gin was not effective in nutritional regimens for patients with sepsis. Since TNF-α is one of the major mediators in sepsis, studies on the relations among TNF-α, Gln and its transporters are of vital importance for an insight into the pathway of Gln metabolism and Gin-enhanced therapeutic effect in sepsis. Our aim was to determine in vivo whether and how TNF-α affects Gin-enhanced protein synthesis, the mRNA and protein expressions of ASCT2, and to disclose the pathway via which TNF-α affects Gin-enhanced protein synthesis. Methods Thirty male Sprague-Dawley rats were randomly assigned to a control （total parenteral nutrition, TPN ）, a Gin and a TNF-α group. All the rats received isonitrogenous and isoenergetic TPN solutions for 3 days. In addition, the rats in the Gin group were given Gin at 0.3 g per kg of body weight per d for 72 hours, and those in the TNF-α group administered Gin at the same dose followed by intravenous infusion of TNF-α in the last 24 hours. At 30 minutes before sample collection, all the rats were mainlined with [ L-15N] -leucine at 1.0 mmol per kg of body weight. Then we measured the concentrations of TNF-α and Gin in the plasma and small intestine, and determined the rate of fractional synthesis and the mRNA and protein expressions of ASCT2. Results The concentration of TNF-α in the plasma was significantly higher in the TNF-α group than in the other two （P 〈 0.01 ）, and so was that of Gin in the plasma and small intestine （P 〈 0.01 ）. The rate of fractional synthesis was dramatically increased in the TNF-α and Gln groups as compared with the control, and it was the highest in the Gin group （P 〈0.01 ）. The mRNA and protein expressions of ASCT2 were remarkably lower in the control group than in the other two （ P 〈 0.01 ）, while its protein expression was significantly higher in the Gln than in the TNF-α and control groups （ P 〈 0.01 ）. Conclusion TNF-α can increase the concentration of Gin in the plasma and small intestine and attenuate Gin-enhanced protein synthesis in the small intestine of rats. Its pathway may involve the inhibitory effect of TNF-α on the function of the Gin transporter to uptake the Gln into target cells for protein synthesis and this inhibition can be found at and after the level of protein translation.