目的 观察早期和晚期二氯乙酸盐（dichloroacetate, DCA）干预对左肺切除(pneumonectomy,PE)联合野百合碱（monocrotaline,MCT）皮下注射诱导大鼠肺动脉高压（pulmonary arterial hypertension, PAH）的治疗作用以及对生存素(survivin)表达的影响。方法 雄性SD大鼠(体重200 g～250 g) 采用随机数字表法随机分为5组,每组8只：正常对照组（N）,DCA对照组(D),肺高压模型组（PM）,早期DC预防组（PMD1）,晚期DCA治疗组（PMD2）。除N组和D组外,其余组别大鼠均行左肺切除,并于14 d后项背部皮下注射MCT(剂量为60 mg/kg）。同时PMD1组和PMD2组分别在左肺切除14 d和 35 d后开始每天给予DCA（剂量为80 mg•kg-1•d-1）灌胃,而PM组在左肺切除14 d后给予相同容积的生理盐水灌胃,D组同N组大鼠,只是左肺切除14 d后同样给予相同剂量的DCA灌胃。PMD1组和PMD2组大鼠分别在左肺切除35 d和56 d行开胸进行肺动脉压力（PAP）的测量。测量后处死动物,取肺组织进行常规病理染色,采用荧光定量PCR法检测survivin mRNA的表达,采用western blot 法检测大鼠survivin蛋白的表达。结果 PM组大鼠平均PAP为(37.8±1.3) mm Hg (1 mm Hg=0.133 kPa),肺小血管中膜厚度占外径的百分比 (WT%)和右心室与左心室加室间隔之比 (RV /LV + S)分别为（40±8)%和（44.8±2.2）,survivin mRNA和Western blot蛋白表达分别为（4.03±0.27）和（0.684±0.055）,与N组、D组比较均明显升高( P＜0.05)。早期DCA预防后PAP明显下降为（26.2±1.50）mm Hg,WT%和RV /LV + S分别降为（24±3) %和（29.5±1.9 ),而survivin mRNA和Western blot蛋白表达也分别降为（2.00±0.12)和（0.415±0.027),与PM组比较均明显下降 (P＜0.05),而晚期干预后PAP为(36.0±1.60) mm Hg,WT%和RV /LV + S分别为(38±3) %和(42.0±1.3), survivin mRNA和Western blot蛋白表达也分别(3.77±0.45) 和(0.732±0.101) ,与PM组比较没有统计学差异（P＞0.05）。结论 Survivin在PAH的发生及肺血管重构中起重要作用,而DCA早期预防能缓解PAP升高和肺血管中膜增厚以及降低survivin的表达量,然而晚期干预后和PM组比较差异没有统计学意义。
Objective To observe the effect of dichloroacetate (DCA） on pulmonary arterial pressure and the expression of survivin in rat model of severe pulmonary arterial hypertention. Methods 40 male Sprague-Dawley rats (8-wk-old,weight 200 g-250 g) were randomly divided into 5goups, each of which had eight rats: normal group(N), Dichloroacetate control group(D), left pneumonectom（PE）+MCT group (PM),early DCA treated group(PMD1), and later DCA treated group(PMD2). Except for group N and D, other groups were induced by pneumonectomy combined with monocrotaline 60 mg/kg subcutaneous injection after pneumonectomy(D14). Group PMD1 and PMD2 were respectively received dichloroacetate 80 mg•kg-1•d-1 after pneumonectomy(D14) and D35 by intragastric gavage once a day for 21 consecutive days. At the same time, the equivalent volume of sterile saline solution was substituted for DCA in group PM. Rats in group D were treated as same as in group N except for receiving DCA gavage treatment as described in group PMD. At D35 (PMD1)and D56(PMD2), rats were anesthetized with ketamine (150 mg/kg, i.p.) and atropine surphate (50 μg, i.p.). Pulmonary arterial pressure was measured by direct puncture of pulmonary artery trunk with i.v. Catheter (22 gauge) connected to pressure transducer. Rats were euthanized by intravenous injection with potassium chloride,and inferior lobe of right lung were dissected, snap-frozen in the liquid nitrogen and kept at –80 ℃ until analyzed. The residual lobes were fixed in neutral formaldehyde for further HE stain and morphometric analysis. The mRNA expression of survivin was analyzed with Realtime-PCR. The protein expression of survivin was detected with Western blot. Results The mean PAP of group PM is (37.80±1.3) mm Hg (1 mm Hg=0.133 kPa), the percentage of medial thickening (WT%) and right ventricular hypertrophy index(RV /LV + S) are （40±8)%and（44.8±2.2）, respectively.The expression of survivin mRNA and protein level are（4.03±0.27）and（0.684±0.055）, respectively. compared with group N and group D , they were markedly increased in group PM (P<0.05). Early treatment with DCA significantly reduced the mean PAP, the percentage of medial thickening, and right ventricular hypertrophy index, they are（26.2±1.50）mm Hg，（24±3) %，and（29.5±1.9 ) , respectively. Meanwhile, the expression of survivin mRNA and protein level were markedly reduced to （2.00±0.12)and（0.415±0.027), compared with group PM, they were significantly reduced in group PMD1（P<0.05）.Whereas later treatment with DCA has no significant effect（P＞0.05）, the mean PAP, the percentage of medial thickening ,and right ventricular hypertrophy index are(36.0±1.60) mm Hg, (38±3) %,and(42.0±1.3), respectively. the expression of survivin mRNA and protein level were (3.77±0.45)and(0.732±0.101), respectively. Conclusions Survivin plays an important role in the development of pulmonary hypertention and the remodeding of pulmonary vascular. DCA alleviates the deterioration of pulmonary arterial hypertension, medial thickening of pulmonary arterioles and protein expression of survivin in pneumonectomized rats injected with MCT, whereas there were no significant difference was found between PM and PMD2.
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