摘要
目的探讨腺病毒介导热休克蛋白70(HSP70)对缺氧性肺动脉高压(HPH)新生大鼠肺的保护作用。方法选取7~10日龄健康、清洁级Wistar新生大鼠128只,按随机数字表法随机分为HPH组和对照组。HPH组根据转染液不同分为盐水组、空病毒组、HSP70组,转染后置于80 mL/L氮氧混合气体的低氧舱内建立HPH模型。造模3、7、10、14 d测定各组新生大鼠平均肺动脉压力(mPAP)。应用反转录PCR和Western blot检测各组新生大鼠肺组织中HSP70、缺氧诱导因子-1α(HIF-1α)、内皮素-1(ET-1)、诱导型一氧化氮合酶(iNOS) mRNA及蛋白表达。结果1.缺氧3、7、10、14 d盐水组mPAP水平(M,Q:12.00,2.50;15.00,2.00;18.00,1.75;20.00,2.25)与对照组(M,Q:9.50,4.75;10.50,1.00;13.00,1.00;15.50,3.25)比较显著增高,差异均有统计学意义(z=-3.28、-3.40、-3.34、-3.06,均P〈0.01);空病毒组(M,Q:13.50,2.00;15.50,1.75;18.00,1.00;22.00,4.25)与对照组比较显著增高,差异均有统计学意义(z=-2.83、-3.40、-3.42、-2.97,均P〈0.01);HSP70组缺氧3、7、10 d的mPAP水平(M,Q:8.50,4.00;10.50,1.00;13.00,1.00)与对照组比较差异均无统计学意义(z=-0.43、-0.00、-3.06,均P〉0.05)。2.HSP70组缺氧3、7、10 d各时间点间HIF-1α、ET-1、iNOS mRNA比较差异均有统计学意义(F=6.321、9.669、6.333,均P〈0.01),HSP70蛋白比较差异均有统计学意义(F=16.463、3.637、17.749,均P〈0.01)。3.缺氧3、7、10 d盐水组HIF-1α mRNA显著高于对照组,差异均有统计学意义(q=4.312、9.106、6.151,均P〈0.01);空病毒组显著高于对照组,差异均有统计学意义(q=3.982、9.235、5.352,均P〈0.01);缺氧3、7 d HSP70组低于空病毒组,差异均有统计学意义(q=6.083、11.031,均P〈0.05)。缺氧3、7、10 d盐水组ET-1 mRNA显著高于对照组,差异均有统计学意义(q=5.112、10.086、6.264,均P〈0.01);空病毒组显著高于对照组,差异均有统计学意义(q=4.182、12.238、5.864,均P〈0.01);缺氧3、7、10 d HSP70组低于空病毒组,差异均有统计学意义(q=6.912、10.235、7.021,均P〈0.05)。缺氧3、7、10 d盐水组iNOS mRNA显著高于对照组,差异均有统计学意义(q=4.998、8.056、5.369,均P〈0.01);空病毒组显著高于对照组,差异均有统计学意义(q=4.778、10.138、5.154,均P〈0.01),缺氧3、7、10 d HSP70组低于空病毒组,差异均有统计学意义(q=7.819、9.838、6.156,均P〈0.05)。缺氧3、7、10 d盐水组HIF-1α蛋白显著高于对照组,差异均有统计学意义(q=3.146、3.012、4.106,均P〈0.05);缺氧10 d空病毒组显著高于对照组,差异有统计学意义(q=3.468,P〈0.05);缺氧3、7、10 d HSP70组显著低于空病毒组,差异均有统计学意义(q=3.876、4.108、4.021,均P〈0.05)。缺氧3、7、10 d HSP70组ET-1蛋白显著低于盐水组,差异均有统计学意义(q=3.367、2.983、3.246,均P〈0.05);且显著低于空病毒组,差异均有统计学意义(q=3.268、2.678、3.567,均P〈0.05)。缺氧3、7、10 d盐水组iNOS蛋白显著高于对照组,差异均有统计学意义(q=3.360、3.567、3.567,均P〈0.05),HSP70组低于空病毒组,差异均有统计学意义(q=3.126、3.908、3.087,均P〈0.05)。结论腺病毒介导HSP70可以提高HPH新生大鼠肺组织HSP70表达,下调HIF-1α、ET-1、iNOS表达,降低肺动脉压力。
ObjectiveTo investigate the protective effect of adenovirus mediated heat shock protein 70 (HSP70) on lungs in neonatal rats with hypoxic pulmonary hypertension(HPH).MethodsOne hundred and twenty-eight 7-10 d healthy Wistar neonatal rats were randomly divided into HPH model group and control group.HPH group was divided into saline group, empty virus group, and HSP70 group according to the transfection solution.HPH model was established in the hypoxia cabin of 80 mL/L nitrogen oxygen mixed gas after transfection.The mean pulmonary artery pressure(mPAP) was measured after 3, 7, 10 and 14 days of hypoxia in each group.The mRNA and protein expression of HSP70, hypoxia inducible factor-1 alpha(HIF-1α), endothelin-1(ET-1) and inducible nitric oxide synthase(iNOS) in the lung tissues of neonatal rats were detected by using reverse transcription-PCR and Western blot respectively.Results(1)The mPAP level was significantly higher in saline group(M, Q: 12.00, 2.50; 15.00, 2.00; 18.00, 1.75; 20.00, 2.25) than that in control group(M, Q: 9.50, 4.75; 10.50, 1.00; 13.00, 1.00; 15.50, 3.25), and the differences were significant(z=-3.28, -3.40, -3.34, -3.06, all P〈0.01); and the diffe-rences were also significant between empty virus group (M, Q: 13.50, 2.00; 15.50, 1.75; 18.00, 1.00; 22.00, 4.25)and control group (z=-2.83, -3.42, -3.40, -2.97, all P〈0.01) in 3, 7, 10, and 14 days; but there was no significant difference between HSP70 group (M, Q: 8.50, 4.00; 10.50, 1.00; 13.00, 1.00)and the control group in 3, 7, and 10 days (z=-0.43 -0.00, -3.06, all P〉0.05). (2)The expressions of HSP70 mRNA among the groups were statistically significant(F=6.321, 9.669, 6.333, all P〈0.01), and the expressions of HSP70 protein also had significant difference(F=16.463, 3.637, 17.749, all P〈0.01). (3)The level of HIF-1α mRNA in saline group was significantly higher than that of the control group, and the differences were statistically significant (q=4.312, 9.106, 6.151, all P〈0.01); and the level of HIF-1α mRNA in empty virus group was also significantly higher than that in the control group, and the differences were statistically significant ( q=3.982, 9.235, 5.352, all P〈0.01) in 3, 7, and 10 days; hypoxia in HSP70 group was lower than that of the empty virus group in 3, 7 days, and the differences were statistically significant (q=6.083, 11.031, all P〈0.05). The level of ET-1 mRNA in saline group was significantly higher than that in the control group(q=5.112, 10.086, 6.264, all P〈0.01), in empty virus group was significantly higher than that in the control group, and the differences were statistically significant (q=4.182, 12.238, 5.864, all P〈0.01) in 3, 7, and 10 days, but in HSP70 group it was lower than that in the empty virus group in 3, 7, and 10 days, and the differences were statistically significant (q=6.912, 10.235, 7.021, all P〈0.05). The level of iNOS mRNA in saline group was significantly higher than that of the control group, and the differences were statistically signi-ficant (q=4.998, 8.056, 5.369, all P〈0.01), in empty virus group was significantly higher than that in the control group, and the differences were statistically significant (q=4.778, 10.138, 5.154, all P〈0.01) in 3, 7, and 10 days, but in HSP70 group it was lower than that in the empty virus group in 3, 7, and 10 days, and the differences were statistically significant (q=7.819, 9.838, 6.156, all P〈0.05). The level of HIF-1α protein in saline group was significantly higher than that of the control group in 3, 7, and 10 days, and the differences were statistically significant (q=3.146, 3.012, 4.106, all P〈0.05), in empty virus group was significantly higher than that of the control group in 10 days, and the difference was statistically significant (q=3.468, P〈0.05); but in HSP70 group it was lower than that in the empty virus group in 3, 7, and 10 days, and the differences were statistically significant (q=3.876, 4.108, 4.021, all P〈0.05). The level of ET-1 protein of HSP70 group was lower than that of the saline group, the differences were statistically significant(q=3.367, 2.983, 3.246, all P〈0.05), in HSP70 group was lower than that of the empty virus, and the differences were statistically significant (q=3.268, 2.678, 3.567, all P〈0.05). The level of iNOS protein in saline group was significantly higher than that in the control group in 3, 7, and 10 days, and the diffe-rences were statistically significant (q=3.360, 3.567, 3.567, all P〈0.05), but in HSP70 group it was lower than that in the empty virus group, and the differences were statistically significant (q=3.126, 3.908, 3.087, all P〈0.05).ConclusionAdenovirus mediated HSP70 can improve the HSP70 expression in HPH, down-regulate the expression of HIF-1α, ET-1, iNOS, and reduce pulmonary arterial pressure.
出处
《中华实用儿科临床杂志》
CSCD
北大核心
2017年第6期451-456,共6页
Chinese Journal of Applied Clinical Pediatrics
基金
国家自然科学基金(81360104)