蛇床子素通过上调p53、下调增殖细胞核抗原和Ki67的表达抗野百合碱所致肺小动脉重构

Osthole reverses pulmonary artery remodeling induced by monocrotaline through increasing expression of p53 and reducing the expression of proliferating cell nuclear antigen and Ki67

目的观察蛇床子素(Ost)对野百合碱(MCT)所致大鼠肺小动脉重构的干预作用,并探讨其机制。方法雄性SD大鼠45只,随机分为正常对照组(n=10)、模型组(n=15)、Ost低剂量组和Ost高剂量组(均n=10)。模型组和Ost低、高剂量组大鼠经颈背部一次性皮下注射MCT 50 mg·kg-1建立肺动脉高压模型,Ost低、高剂量组于造模后第1日开始灌胃给予Ost 10、20 mg·kg-1,每日1次,模型组和正常对照组给予等量双蒸水。给药28 d后,测量大鼠体重、肺重,计算肺重指数;通过HE染色观察肺小动脉病理学变化,采用IPP6.0图像分析软件统计小动脉血管厚度和血管外径与内径比值;real time RTPCR检测肺组织抑癌基因p53、增殖细胞核抗原(PCNA)和Ki67 m RNA的表达,Western blotting检测肺组织PCNA蛋白的表达。结果与正常对照组相比,模型组大鼠体重减轻、肺重和肺重指数明显升高(P<0.01);肺小动脉壁明显增厚、管腔狭窄,肺小动脉血管厚度及血管外径与内径比值明显增大(P<0.01);肺组织p53 m RNA的表达明显下调(P<0.01),Ki67m RNA、PCNA m RNA和蛋白的表达均明显上调(P<0.01)。与模型组相比,Ost高剂量组肺重明显降低(P<0.01);Ost低、高剂量组大鼠体重增加、肺重指数明显降低(P<0.05或P<0.01);肺小动脉壁变薄,肺小动脉血管厚度及血管外径与内径比值明显减小(P<0.05或P<0.01);肺组织p53 m RNA的表达上调(P<0.05或P<0.01),Ki67 m RNA、PCNA m RNA和蛋白的表达均明显下调(P<0.01)。与Ost低剂量组相比,Ost高剂量组大鼠肺小动脉血管厚度减小(P<0.05),p53 m RNA的表达上调(P<0.05)。结论 Ost能抗MCT诱导的肺小动脉重构,其机制可能与上调p53、下调PCNA和Ki67的表达有关。

AIM To investigate the effects of osthole（Ost） on pulmonary artery remodeling induced by monocrotaline（MCT） in rats and to explore the mechanism. METHODS Forty- five male Sprague- Dawley rats were randomly divided into normal control group（n = 10）, model group（n = 15）, low dose of Ost treatment group（n = 10） and high dose of Ost treatment group（n = 10）. All rats were given a single dose of MCT 50 mg·kg-1subcutaneously to establish pulmonary arterial hypertension model except the normal control group. After modeling, the rats in the low dose and higt dose of Ost treatment group were gavaged Ost 10 and 20 mg·kg-1once daily for 28 days. The other rats in the model group and normal control group were given the same amount of double distilled water. After 28 days of administration, the lung and body was weighed and the relative weight ratio of lung to body weight was calculated. The morphological change of the pulmonary artery was observed by HE staining. The vascular thickness and the outer and inner diameter ratio of pulmonary artery were tested by IPP 6.0 image analysis software. The m RNA expression of tumor suppressor gene p53, proliferating cell nuclear antigen（PCNA） and Ki67 were examined by real time RT- PCR. The protein expression of PCNA was detected by Western blotting. RESULTS Compared with those in the normal control group, the weight were lost and the lung weight index was significantly increased, the thickening of pulmonary artery was observed, and the vascular thickness and the outer and inner diameter ratio of pulmonary artery were increased in the model group（P〈0.01）. The m RNA expression of p53 in lung tissues was down- regulated in the model group（P〈0.01）.The m RNA expression of Ki67, the m RNA and the protein expression of PCNA in lung tissues were up-regulated in the model group（ P〈0.01）. Compared with those in the model group, the lung weight was decreased significantly in the high dose of Ost treatment group（P〈0.01）. The weight lost and the value of lung weight index（ P〈0.05 or P〈0.01）, the thickening of pulmonary artery were improved, and the vascular thickness and the outer and inner diameter ratio of pulmonary artery were decreased significantly（P〈0.05 or P〈0.01） in the low and high dose of Ost treatment group. The m RNA expression of p53 in lung tissues was upregulated（P〈0.05 or P〈0.01）, while the m RNA expression of Ki67, the m RNA and the protein expression of PCNA in lung tissues were down- regulated（ P〈0.01） in the low and high dose of Ost treatment group.Compared with those in the low dose of Ost treatment group, the vascular thickness were decreased（P〈0.05）,and the m RNA expression of p53 in lung tissues was up- regulated（P〈0.05） in the high dose of Ost treatment group. CONCLUSION Ost can resist the development of pulmonary artery remodeling induced by MCT in rats,and the mechanism might be related to increase the expression of p53 and reduction the expression of PCNA and Ki67.