谷氨酰胺对大鼠肠缺血再灌注损伤的保护作用及相关机制

Protective effects of glutamine on intestinal ischemia reperfusion injury in rats and its mechanism

目的研究谷氨酰胺对肠缺血再灌注损伤(IR)大鼠的作用及相关机制。方法按照体重将大鼠随机分为5组:对照组、实验-A组(术前静脉给谷氨酰胺)、实验-B组(术前灌胃给谷氨酰胺)、实验-C组(术中静脉给谷氨酰胺)和实验-D组(术中肠腔给谷氨酰胺),每组5只。造模前,实验-A组和实验-B组经尾静脉注射和灌胃给予谷氨酰胺1. 5 mg·kg^(-1),于开腹后,实验-C组和实验-D组分别经肠系膜上静脉和肠腔给予谷氨酰胺1. 5 mg·kg^(-1),对照组经尾静脉注射给予0. 9%NaCl 1 mL,1次/天,连续4 d。用血管夹夹闭肠系膜上血管制备大鼠肠IR模型。缺血50 min,再灌注1,12 h后,取肠标本和血标本。用酶联免疫吸附试验检测血中的肿瘤坏死因子-α(TNF-α)含量;用反转录聚合酶链反应法检测肠标本组织中的核因子-κB(NF-κB)和过氧化物酶体增殖物激活受体γ(PPAR-γ) mRNA表达水平;用免疫印迹法检测NF-κB和PPAR-γ蛋白表达水平。结果肠IR 12 h后,对照组、实验-A组、实验-B组、实验-C组和实验-D组的血清中TNF-α含量分别是(419. 04±10. 55),(311. 77±9. 81),(224. 53±3. 36),(318. 77±15. 64)和(436. 20±22. 50)ng·L^(-1),实验-A组和实验-B组与对照组比较,差异均有统计学意义(均P <0. 05);实验-A组和实验-B组与与实验-D组比较,差异均有统计学意义(均P <0. 05)。这5组的NF-κB mRNA含量分别是1,0. 51±0. 03,0. 61±0. 07,0. 62±0. 09和0. 92±0. 05;这5组的PPAR-γmRNA含量分别是1,2. 91±0. 22,2. 52±0. 24,1. 48±0. 12和1. 39±0. 13,实验-A组与对照组和实验-D组比较,NF-κB和PPAR-γmRNA表达量的差异均有统计学意义(均P <0. 05)。NF-κB和PPAR-γ蛋白含量趋势与mRNA一致。IR 1 h的上述指标结果与12 h一致。结论谷氨酰胺通过降低NF-κB活性和增加PPAR-γ活性,对肠IR大鼠起到保护作用。

Objective To investigate the effects of glutamine on intestinal ischemia reperfusion injury(IR)in rats and the related mechanisms.Methods According to the weight,rats were divided into five groups:control group,experimental-A group,experimental-B group,experimental-C group and experimental-D group.The A group and B group were preoperatively given glutamine(1.5 mg·kg-1)through tail vein injection and intragastric administration,respectively.The control group was preoperatively administered 1 mL 0.9%sodium chloride was given for the rat each day by tail vein injection.The experimental-C group and experimental-D group were given glutamine(1.5 mg·kg-1)through injection of superior mesenteric vessel and intestinal cavity,respectively,after open abdomen,once a day for 4 days.Meanwhile,the model of intestinal IR was established by clamping the superior mesenteric vessel.Intestinal tissues and blood samples were taken after ischemia for 50 minutes,reperfusion for 1 h and 12 h.The changes of tumor necrosis factor-alpha(TNF-α)content in blood samples were detected by enzyme-linked immunosorbent assay.RT-PCR were used to detect nuclear factor kappa-B(NF-κB)and peroxisome proliferator-activated receptor gamma(PPAR-γ)mRNA in intestinal specimens of rats with intestinal IR.Western-blot were used to detect NF-κB and PPAR-γprotein expression.Results After intestinal reperfusion for 12 h,serum TNF-αlevels in control,experimental-A,experimental-B,experimental-C and experimental-D groups were respectively(419.04±10.55),(311.77±9.81),(224.53±3.36),(318.77±15.64)and(436.20±22.50)ng·L-1;comparison between experimental-A group and experimental-B group with control group,the difference was statistically significant(all P<0.05);comparison between experimental-A group and experimental-B group with experimental-D group,the difference was statistically significant(all P<0.05).The expression levels of NF-κB mRNA in the five groups were 1,0.51±0.03,0.61±0.07,0.62±0.09 and 0.92±0.05,respectively;meanwhile,the expression levels of PPAR-γmRNA in the five groups were 1,2.91±0.22,2.52±0.24,1.48±0.12 and1.39±0.13,respectively;comparison between experimental-A group with control group and experimental-D group,the difference was statistically significant(all P<0.05).The trend of protein results is consistent with that of gene.The trend of 1 h results is consistent with that of 12 h.Conclusion Glutamine can play a protective role to intestinal IR rats via a mechanism of reducing the activity of NF-κB and increasing the activity of PPAR-γ.