谷氨酰胺联合脐血间充质干细胞移植在大鼠肠缺血再灌注损伤中作用的研究
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Effects of Glutamine Combined with umbilical cord blood mesenchymal stem cell transplantation on intestinal ischemia reperfusion injury in rats
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    摘要:

    【摘要】目的:探讨谷氨酰胺联合脐血间充质干细胞(MSCs)移植在大鼠肠缺血再灌注损伤中作用的研究。方法:体外复苏并培养脐血间充质干细胞移植前备用,观察CM-DiI荧光标记后脐血间充质干细胞的去向。80只SD大鼠随机分为正常对照组,缺血再灌注损伤组,谷氨酰胺组,MSCs移植组及联合组每组各15只。对照组采用生理盐水灌肠,损伤组采用TNB (S乙醇稀释)灌肠,在TNBS建模后1h,谷氨酰胺组于尾静脉输入谷氨酰胺0.45 g/kg、MSCs移植组于尾静脉输入1?010 L-1 脐血间充质干细胞悬液,联合组尾静脉输入谷氨酰胺0 .45 g/kg 脐血间充质干细胞悬液1?010 L-1 。通过ELISA法检测各组大鼠血清中肠脂肪酸结合蛋白(IFABP)、白介素-6(IL-6)、超氧化物歧化酶(SOD)的含量;各组于再灌注1h、3h后检测肠组织含水率;通过RT-PCR、Western blot观察大鼠肠黏膜上皮细胞Caspase-3、NF-kB、Bcl-2在谷氨酰胺联合MSCs移植后的mRNA和蛋白的表达情况。结果:通过荧光示踪法观察到移植的MSCs细胞分布于肠粘膜淋巴组织内和腺上皮细胞间,表明MSCs可能参与了肠缺血再灌注损伤的修复过程。各组大鼠血清中SOD、IFABP、IL-6的含量变化比较,损伤组血清中IFABP、IL-6的含量较对照组显著增加,而谷氨酰胺组,MSCs移植组及联合组与之比较,则显著减少,联合组减少更为明显,损伤组血清中SOD的含量较对照组显著减少,而谷氨酰胺组,MSCs移植组及联合组与之比较,则显著增高,联合组增高更为明显( P<0.05)。再灌注1h和3h,损伤组肠组织含水率均明显高于对照组;与损伤组相比,谷氨酰胺组、MSCs移植组及联合组肠组织含水率值均显著降低,联合组降低更为明显,而谷氨酰胺组、MSCs移植组差异无统计学意义( P>0.05)。与对照组比较,损伤组肠黏膜上皮细胞Caspase-3、NF-kB的mRNA和蛋白表达明显上调,Bcl-2的mRNA和蛋白表达明显下调( P<0.05),而谷氨酰胺组、MSCs移植组及联合组与之比较,Caspase-3、NF-kB的mRNA和蛋白表达明显下调,Bcl-2的mRNA和蛋白表达明显上调( P<0.05),谷氨酰胺组及MSCs移植组之间无统计学差异( P>0.05),但两组与联合组比较,差异明显( P<0.05)。结论:谷氨酰胺组及MSCs移植后,明显减轻了大鼠肠缺血再灌注损伤程度,其可能通过抑制Caspase-3、NF-kB表达和促进Bcl-2表达减轻肠黏膜缺血再灌注损伤。?

    Abstract:

    [Abstract] objective: To investigate the effect of Glutamine Combined with umbilical cord blood mesenchymal stem cells (MSCs) transplantation on intestinal ischemia-reperfusion injury in rats. Methods:Isolation of umbilical cord blood mesenchymal stem cells in vitro, and observe its fate after CM-DiI fluorescent labeling. 80 SD rats were randomly divided into normal control group, ischemia reperfusion injury group, glutamine group, MSCs transplantation group and combined group with 15 rats in each group. Control group using saline enema, injury group using TNBS (ethanol dilution) enema, and after TNBS modeling 1h, at the end of intravenous glutamine 0.45 g/kg in the glutamine group, MSCs transplantation group in the tail vein of 1 ?010/L umbilical cord blood mesenchymal stem cell suspension, and combined group intravenous glutamine 0.45 g/kg and 1?010/L umbilical cord blood mesenchymal stem cell suspension. ELISA were used to detect midgut fatty acid binding protein (iFABP), interleukin 6 (IL-6), superoxide dismutase (SOD) content in rat serum. The water content of intestinal tissue were detected by 1h and 3h after reperfusion in each group. Expression of NF-kB, Bcl-2 and Caspase-3 mRNA after glutamine combined with MSCs in rat intestinal epithelial cells were observed by RT-PCR and Western blot. Results:The fluorescent tracer method to observe the transplanted MSCs cells distributed in the intestinal mucosal lymphoid tissues and epithelial cells gland, it indicated that MSCs might be involved in the repair process of intestinal ischemia-reperfusion injury. Comparison of contents of SOD, IFABP and IL-6 in serum of rats in each group, the content of IFABP and IL-6 in the serum of injured group was significantly higher than that in the control group, in the glutamine group, the MSCs transplantation group and the combined group were significantly reduced, and the combined group was more obvious. The content of SOD in the injured group was significantly lower than that in the control group, and glutamine group, MSCs transplantation group and the combined group significantly increased, the combined group increased significantly (P<0.05). The water content of intestinal tissue in the injured group was significantly higher than that in the control group after reperfusion 1h and 3h,.Compared with the injury group, the water content of the glutamine group, MSCs transplantation group and the combined group were significantly lower, and the combination group decreased more significantly, and there was no significant difference in glutamine group and MSCs transplantation group (P>0.05). Compared with the control group, injury group intestinal epithelial cells Caspase-3,NF-kB mRNA and protein expression were significantly increased, and expression of Bcl-2 mRNA and protein were significantly lower (P <0.05), the expression of Caspase-3 and NF-kB mRNA and protein were significantly reduced in the glutamine group, MSCs transplantation group and combined group, expression of Bcl-2 mRNA and protein were significantly increased (P <0.05), between glutamine group and MSCs transplantation group were no significant difference (P> 0.05), but the two groups compared with the combined group, the difference were significant (P<0.05). Conclusions: After glutamine and MSCs transplantation, the degree of intestinal ischemia reperfusion injury was obviously reduced in rats, it may reduce intestinal mucosal ischemia-reperfusion injury by inhibiting the expression of Caspase-3 and NF-kB and promoting the expression of Bcl-2.

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王炳杰.谷氨酰胺联合脐血间充质干细胞移植在大鼠肠缺血再灌注损伤中作用的研究[J].中国比较医学杂志,,().

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  • 收稿日期:2016-03-18
  • 最后修改日期:2016-04-18
  • 录用日期:2016-06-15
  • 在线发布日期: 2016-11-28
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