内皮素-1在缺氧性肺动脉高压新生大鼠肺组织中的表达变化及意义

Changes and significance of endothelin-1 in lung tissue of hypoxic pulmonary hypertension development in neo- natal rats

目的了解内皮素-1（ET-1）在缺氧性肺动脉高压（HPH）新生大鼠肺组织中的表达变化及其意义。方法将120只Wistar新生大鼠按随机数字表法分为缺氧组和对照组，缺氧组新生大鼠建立HPH模型，分别在缺氧3、5、7、10、14、21d检测平均肺动脉压力（mPAP），观察肺小动脉形态学改变并计算肺血管重塑指标：肺小动脉血管壁中层厚度占外径的百分比（MT％）、肺小动脉中层横截面积占总横截面积的百分比（MA％）；通过免疫组织化学和Westernblot法分别进行肺组织ET-1蛋白表达的定性和定量分析。结果1．缺氧3、5、7、10、14、21d，缺氧组mPAP[（8．492±1．548）mmHg、（10．022±1．182）mmHg、（11．470±2．868）mmHg、（16．842±2．154）mmHg、（12．824±2．859）mmHg、（21．036±2．590）ITlmHg；1mmHg=0．133kPa]较对照组[（5．141±1．022）mmHg、（8．137±1．057）mlrlHg、（8．730±0．868）mmHg、（12．125±2．541）mmHg、（8．920±0．744）iTlmHg、（11．156±1．644）inmHg]显著升高，差异均有统计学意义（P均〈0．05）。2．缺氧7d新生大鼠出现肺小动脉重塑改变，缺氧组MT％[（53±11）％]和MA％[（60±9）％]分别高于对照组[（49±11）％、（54±8）％]，差异均有统计学意义（P均〈0．05）。3．免疫组织化学结果：缺氧3、5、7d，缺氧组肺血管壁ET．1表达强度（0．614、0．613、0．651）明显高于对照组（0．433、0．386、0．369），差异均有统计学意义（P均〈0．05）。4．Westernblot结果：缺氧3、5、7、10d缺氧组ET-1表达量（4．885±1．391、4．434±1．726、6．309±0．330、2．353_-t-O．961）明显高于对照组（1．698±0．794、1．454±0．776、2．045±0．668、0．766±0．515），差异均有统计学意义（P均〈0．05）。5．相关分析结果：缺氧3—7d总ET-1表达量与该时间总mPAP呈正相关（r=0．459，P〈0．05），而与MT％、MA％，之间均未发现明显相关性（rMT=-0．041，r=0．322，P均〉0．05）。结论HPH新生大鼠肺组织中ET-1蛋白的表达在缺氧早期升高，缺氧后期不再升高，提示存在缺氧耐受。ET-1是新生大鼠HPH发病过程中的早期变化指标，其作用可能主要与肺动脉压力升高有关，进一步提示在缺氧早期即诵讨有效手殷抑制ET．1的表；大对该病的防治可能更为有效。

Objective To know about the effect and significance of endothelin-1 （ET-1） in pathogenesis of hy- poxic pulmonary hypertension （HPH）in neonatal rats. Methods The 120 newborn Wistar rats were randomly divided into hypoxic group and control group, and the rats of hypoxic group were made into HPH model. Mean pulmonary arte- rial pressure（mPAP） was measured on day 3,5,7,10,14 and 21 after hypoxia. The morphological change of pulmonary vessels were observed, and pulmonary vascular remodeling indexes were also obtained, including the ratio of middle wall thickness to external diameter of small pulmonary arteries （ MT% ） and the ratio of media cross-section area to total cross-sectional area of small pulmonary arteries（ MA% ）. Immunohistochemistry and Western blot tests were done to de- termine the expression of ET-1 in lung tissue. Results 1. After hypoxia for 3,5,7,10,14,21 days, the levels of mPAP inhypoxiagroup[（8.492~l.548） mm Hg,（10.022±1.182） mm Hg,（11.470±2.868） mm Hg,（16.842± 2. 154） mm Hg, （ 12. 824 ± 2. 859） mm Hg, （ 21. 036 ± 2. 590） mm Hg; 1 mm Hg = 0. 133 kPa ] were significantly higher than those in control group[ （5. 141 ± 1. 022） mm Hg, （8. 137 ± 1. 057） mm Hg, （8. 730 ±0. 868） mm Hg, （12. 125 ±2.541） mm Hg,（8.920 ±0.744） mm Hg,（ll. 156 ±1.644） mm Hg] （all P〈0.05）.2. Seven days after hypoxia exposure,small pulmonary arterials remodeling was observed. MT% [ （53 ± 11 ） % ] and MA% [ （60 ± 9） % ] in hypoxia group were also significantly higher than those in control group [ （49 ± 11 ） %, （ 54 ± 8） % ] （ all P 〈 0.05 ）. 3. The results of immunohistochemistry：the expression intensities of ET-1 were significantly higher in hypoxia group （0. 614,0. 613,0. 651） after hypoxia for 3,5,7 days than those in control group（0. 433,0. 386,0. 369） （all P 〈 0.05）. 4. The results of Westem blot： the levels of ET-1 were significantly higher in hypoxia group（4. 885 ± 1. 391, 4. 434 ± 1. 726,6. 309 ± 0. 330,2. 353 ± 0. 961 ） after hypoxia for 3,5,7 and 10 days than those in control group （ 1. 698 ± 0.794,1. 454 ± 0. 776,2. 045 ± 0. 668,0. 766 ± 0.515 ） （ all P 〈 0.05 ）. 5. The results of correlation analy-sis ：the total expression of ET-1 protein was positively correlated with the total level of mPAP on 3 -7 days after hypoxia （r = O. 459 ,P 〈 O. 05 ）. However, the expression of ET-1 had no significant correlation with MT% and MA% （rMT = -- 0. 041, rM^ = 0. 322, all P 〉 0.05 ）. Conchmions The level of ET-1 in lung tissue of HPH newborn rats increased in the early stage after hypoxia exposure and no longer increased during the late stage, which indicated that hypoxia tolerance prevailed. ET-1 acted as an early reactive marker in the process of HPH development in neonatal rats and may be mainly associated with elevated mPAP, which further prompted that through valid means to inhibit the expression of ET-1 in the early stage of hypoxia,in which the prevention and treatment of this disease may be more effective.