肝移植围手术期缺血-再灌注胃氧化损伤发生机制

Mechanism of ischemia-reperfusion injury-induced gastric oxidative damage during perioperative period of liver transplantation

目的探讨肝移植围手术期缺血-再灌注胃氧化损伤发生的规律及机制。方法将28只SD大鼠按随机数字表法分为4组,每组7只。其中3组建立自体原位肝移植模型。根据移植肝恢复灌注时间设立4、8、16 h组。另1组设为假手术组(Sham组)。取大鼠胃壁组织HE染色后观察胃黏膜损伤的病理学变化。检测胃组织超氧阴离子(O2-)、丙二醛(MDA)、谷胱甘肽(GSH)、谷胱甘肽过氧化物酶(GSH-PX)水平。采用Western blot法检测胃组织硫氧还蛋白(Trx)-2表达。实验数据比较采用单因素方差分析及LSD-t检验。结果 4 h组胃黏膜深层及黏膜下层充血、水肿、出血,腺体排列紊乱,部分区域出血、坏死,糜烂深达黏膜肌层;8、16 h组胃黏膜损伤减轻,仅黏膜浅层和深层充血、水肿,Sham组表现为多数胃黏膜上皮结构完整。4 h组O2-和MDA分别为(185±26)U/mg和(0.4±0.1)nmol/mg,8 h组相应为(192±59)U/mg和(0.5±0.1)nmol/mg,明显高于Sham组的(102±34)U/mg和(0.2±0.1)nmol/mg(LSD-t=4.99,4.23和6.37,4.52;P<0.05)。4 h组GSH和GSH-PX分别为(17±6)mg/g和(781±174)U/mg,8 h组相应为(15±4)mg/g和(750±160)U/mg,明显低于Sham组的(30±6)mg/g和(1 162±215)U/mg(LSD-t=-3.26,-4.01和-3.78,-4.36;P<0.05)。16 h组大鼠胃组织O2-、MDA、GSH、GSH-PX分别为(169±27)U/mg、(0.3±0.1)nmol/mg、(25±8)mg/g、(1 108±183)U/mg,与8 h组比较差异有统计学意义(LSD-t=-2.85,-3.46,2.66,3.69;P<0.05)。4、8 h组Trx-2蛋白相对表达量分别为52±10、43±8,较Sham组的125±16明显减少(LSD-t=-5.34,-6.23;P<0.05)。16 h组Trx-2蛋白表达量为160±18,较8 h组明显升高(LSD-t=4.75,P<0.05)。结论大鼠肝移植缺血-再灌注损伤早期即引起胃氧化损伤,其发生可能与Trx-2表达下降、活性氧增多和机体抗氧化能力下降有关。

Objective To investigate the occurrence rule and mechanism of ischemia-reperfusion injury （IRI）-induced gastric oxidative damage during perioperative period of liver transplantation. Methods Twenty-eight SD rats were randomized into 4 groups according to the random number table. The model of orthotopic autologous liver transplantation was established in 3 groups, which were 4 h, 8 h, 16 h group according to the reperfusion time of the liver grafts. The other group was set as Sham group. Gastric tissues were stained with HE to observe pathological changes of gastric mucosal injury. Superoxide anion （O2-）, malondialdehyde （MDA）, glutathione （GSH） and glutathione peroxidase （GSH-PX） in gastric tissues were detected. The expression of thioredoxin （Trx）-2 in gastric tissues was detected by Western blot. The comparison on experimental data was conducted using one-way analysis of variance and LSD-t test. Results In 4 h group, congestion, edema and hemorrhage were observed in deep stratum and submucous stratum of stomach, as well as disorganized glands, regional hemorrhage and necrosis, and erosion was observed deep to the muscularis mucosa. In 8 h group and 16 h group, gastric mucosal injury was alleviated, and only congestion and edema in superficial and deep layer were observed. In Sham group, the epithelium structure of most gastric mucosa was intact. The 02- and MDA of 4 h group were respectively （185±26） U/mg and （0.4±0.1） nmol/mg and those of 8 h group were respectively （192±59） U/mg and （0.5±0.1） nmol/mg, which were significantly higher than （102±34） U/rag and （0.2±0.1） nmol/mg of Sham group （LSD-t=4.99, 4.23 and 6.37, 4.52; P〈0.05）. GSH and GSH-PX of 4 h group were respectively （17±6） mg/g and （781±174） U/mg and those of 8 h group were respectively （15±4） mg/g and （750±160） U/mg, which were significantly lower than （30±6） mg/g and （1 162±215） U/mg of Sham group （LSD-t=-3,26, -4.01 and -3.78, -4.36; P〈0.05）. The O2-, MDA, GSH and GSH-PX of 16 h group were respectively （169±27） U/mg, （0.3±0.1） nmol/mg, （25±8） mg/g and （1 108±183） U/mg, and significant difference was observed comparing with 8 h group （LSD-t=-2.85, -3.46, 2.66, 3.69; P〈0.05）. The relative expression of Trx-2 in 4 h group and 8 h group were respectively 52±10 and 43±8, which were significantly lower than 125±16 of Sham group （LSD-t=-5.34, -6.23; P〈0.05）. The expression of Trx-2 in 16 h group was 160±18, which was significantly higher than that of 8 h group （LSD-t-4.75, P〈0.05）. Conclusion IRI causes gastric oxidative damage in the early phase after liver transplantation in rats, which may be associated with the decrease of Trx-2 expression, increase of active oxygen and decrease in organic antioxidative ability.