高压氧下调HIF-1α表达抑制动静脉内瘘内膜增生的研究

Hyperbaric oxygen downregulates hypoxia-inducible factor-1α and inhibits venous neointimal hyperplasia of arteriovenous fistula：an experimental study

摘要:

目的：研究高压氧疗法（hyperbaric oxygen，HBO） 对于动静脉内瘘（arteriovenous fistula，AVF）的静脉内膜增生（venous neointimal hyperplasia，VNH）的影响。方法：36只健康新西兰兔随机分为3组（每组12只），分别为肾衰假手术组（假手术组）、肾衰动静脉内瘘高压氧干预组（实验组）、肾衰动静脉内瘘组（对照组）。各组均以腺嘌呤诱导的方式建立慢性肾功能衰竭（chronic renal failure，CRF）模型，实验组、对照组新西兰兔建立AVF，并对实验组进行高压氧治疗。分别于开始HBO干预后第7、14、28天各组截取4只新西兰兔AVF静脉段血管，用HE染色评估各组内膜厚度，免疫组织化学技术和Western blot 检测缺氧诱导因子-1α（hypoxia-inducible factor-1α，HIF-1α）、血管内皮生长因子（vascular endothelial growth factor，VEGF）、基质金属蛋白酶2（matrix metalloproteinase-2，MMP-2）的表达。结果：①28 d时，假手术组未观察到AVF静脉端静脉内膜增生，实验组、对照组平均静脉内膜增生厚度分别为（0.200 5±0.034 0）和（0.351 4±0.062 0） mm，3组静脉内膜增生厚度差异具有统计学意义（均P＜0.05）。②28 d假手术组、对照组、实验组HIF-1α平均光密度（mean optical density，MOD）分别为0.003 1±0.000 8、0.201 6±0.046 1、0.099 1±0.027 6，3组HIF-1α MOD差异具有统计学意义（均P＜0.05）。③28 d假手术组、对照组、实验组VEGF分别为0.008 3±0.001 9、0.214 3±0.052 6、0.105 1±0.040 6。3组VEGF MOD差异具有统计学意义（均P＜0.05）。④28 d假手术组、对照组、实验组MMP-2 MOD分别为0.005 1±0.014 6、0.238 4±0.090 3、0.109 0±0.041 5，3组MMP-2 MOD差异具有统计学意义（均P＜0.05）。⑤28 d假手术组、对照组、实验组HIF-1α表达水平分别为0.114 9±0.010 2、0.193 7±0.016 6、0.151 0±0.018 5，3组HIF-1α表达水平差异具有统计学意义（均P＜0.05）。⑥28 d假手术组、对照组、实验组VEGF表达水平分别为0.093 7±0.011 6、0.214 3±0.052 6、0.238 6±0.008 3。3组VEGF表达水平差异具有统计学意义（均P＜0.05）。⑦28 d假手术组、对照组、实验组MMP-2表达水平分别为0.121 6±0.010 4、0.229 5±0.026 0、0.175 5±0.011 8，3组MMP-2 MOD差异具有统计学意义（均P＜0.05）。结论：HBO治疗对于AVF组织内的HIF-1α、VEGF、MMP-2表达及血管内膜增生有明显的抑制作用。

Abstract:

Objective：To investigate the effect of hyperbaric oxygen（HBO） on venous neointimal hyperplasia（VNH） of arteriovenous fistula（AVF）. Methods：A total of 36 healthy New Zealand rabbits were randomly divided into sham-operation+renal failure group （sham-operation group），renal failure+AVF+HBO intervention group（experimental group），and renal failure+AVF group（control group），with 12 rabbits in each group. A model of chronic renal failure was established by adenine induction；the rabbits in the experi－mental group and the control group were used to establish a model of AVF，and those in the experimental group were given HBO therapy. Four rabbits were selected from each group on days 7，14，and 28 of HBO intervention，and the vessels of the vein with AVF were collected. HE staining was performed to evaluate intimal thickness，and immunohistochemistry and Western blot were used to measure the expression of hypoxia-inducible factor-1α（HIF-1α），vascular endothelial growth factor（VEGF），and matrix metallopro－teinase-2（MMP-2）. Results：On day 28，VNH was not observed in the sham-operation group，and the mean thickness of VNH was 0.200 5±0.034 0 mm in the experimental group and 0.351 4±0.062 0 mm in the control group；there was a significant difference in the thickness of VNH between the three groups. On day 28，there was a significant difference in HIF-1α MOD between the sham-op－eration group，the control group，and the experimental group（0.003 1±0.000 8 vs. 0.201 6±0.046 1 vs. 0.099 1±0.027 6，P<0.05）. On day 28，VEGF MOD was 0.008 3±0.001 9 in the sham-operation group，0.214 3±0.052 6 in the control group，and 0.105 1±0.040 6 in the experimental group，and there was a significant difference in VEGF MOD between the three groups（P<0.05）. On day 28，MMP-2 MOD was 0.005 1±0.014 6 in the sham-operation group，0.238 4±0.090 3 in the control group，and 0.109 0±0.041 5 in the experimental group，and there was a sig－nificant difference in MMP-2 MOD between the three groups（P<0.05）. On day 28，the expression level of HIF-1α was 0.114 9±0.010 2 in the sham-operation group，0.193 7±0.016 6 in the control group，and 0.151 0±0.018 5 in the experimental group，and there was a significant difference in the expression level of HIF-1α between the three groups（P<0.05）. On day 28，the expression level of VEGF was 0.093 7±0.011 6 in the sham-operation group，0.214 3±0.052 6 in the control group，and 0.238 6±0.008 3 in the experimental group，and there was a significant difference in the expression level of VEGF between the three groups（P<0.05）. On day 28，the expression level of MMP-2 was 0.121 6±0.010 4 in the sham-operation group，0.229 5±0.026 0 in the control group，and 0.175 5±0.011 8 in the experimental group，and there was a significant difference in MMP-2 MOD between the three groups（P<0.05）. Conclusion：HBO therapy can downregulate the expression of HIF-1α，VEGF，and MMP-2 in AVF tissue and significantly inhibit VNH.