辛伐他汀对肺纤维化大鼠肺组织血管新生及VEGF和PF4基因表达的影响

发布时间：2019-02-12 14:47

【摘要】：目的：观察辛伐他汀对博莱霉素（BLM）诱导肺纤维化大鼠肺组织血管新生及血管内皮生长因子（VEGF）和血小板第4因子（PF4）基因表达的影响，探讨辛伐他汀治疗肺纤维化的分子生物学机制，为辛伐他汀的临床应用提供理论依据。方法：6周龄健康SD雄性大鼠96只，采用随机数字表法分成4组，分别为正常对照组(A组)、博来霉素组（B组）、醋酸泼尼松治疗组(C组)、辛伐他汀治疗组(D组)，每组24只。B、C、D组使用博来霉素溶液以5mg/kg气管内一次性给药建立肺纤维化的动物模型，A组则以同样的方法气管内一次性注入等体积生理盐水。C组、D组在博来霉素造模后第1天开始分别以泼尼松混悬液5mg/kg·d、辛伐他汀混悬液10mg/kg·d灌胃，而A组和B组分别以等体积的生理盐水（10ml/kg）每日灌胃。分别于造模后第7、14、28天取材，各组每次随机处死大鼠8只，共32只。肺组织HE染色观察肺泡炎及肺纤维化改变；采用免疫组化法（SP法）检测各组大鼠肺组织血管新生情况及VEGF、PF4蛋白的表达，采用RT-PCR法检测各组大鼠肺组织VEGF及PF4mRNA的表达。结果：①肺组织病理学结果显示A组大鼠肺组织结构清晰完整，无充血、水肿、渗出，无纤维化病变。B组第7d表现为轻度肺泡炎，肺泡间隔增宽，较多炎细胞浸润，可见成纤维细胞增生；第14d肺泡炎达到高峰，，肺泡间隔明显增宽，炎细胞浸润加重，成纤维细胞明显增殖；第28d肺纤维化明显，肺泡腔塌陷或消失，成纤维细胞大量增生，可见胶原大量沉积。激素及辛伐他汀治疗组肺泡炎和纤维化程度较模型组各相应时间点均有不同程度的减轻。②各组之间相同时间点比较：B、C组大鼠肺组织微血管密度（MVD）高于A组，差异有统计学意义；B、C两组之间比较，差异无统计学意义；D组微血管密度明显低于B组，差异有统计学意义。③免疫组化和RT-PCR结果：各组之间相同时间点比较， B组、C组肺组织VEGF表达明显高于A组，差异有统计学意义；B、C两组间比较差异无统计学意义；D组肺组织VEGF表达明显低于B组，差异有统计学意义。B组、C组肺组织PF4表达明显低于A组，差异有统计学意义；B、C两组间比较差异无统计学意义；D组肺组织PF4表达明显高于B组，差异有统计学意义。结论：①辛伐他汀能够抑制肺纤维化大鼠肺组织中的微血管新生，减轻肺泡炎和肺纤维化的程度。②辛伐他汀抗肺纤维化的机制可能与其抑制肺组织内VEGF的表达，相对上调PF4的表达，抑制病理性血管新生有关。③糖皮质激素不能减轻肺纤维化过程中肺组织的血管新生。
[Abstract]:Aim: to observe the effects of simvastatin on angiogenesis, vascular endothelial growth factor (VEGF) and platelet factor 4 (PF4) gene expression in lung tissue of bleomycin (BLM) induced pulmonary fibrosis rats. To explore the molecular biological mechanism of simvastatin in the treatment of pulmonary fibrosis and to provide theoretical basis for the clinical application of simvastatin. Methods: 96 healthy SD male rats aged 6 weeks were randomly divided into 4 groups: normal control group (group A), bleomycin group (group B), prednisone acetate treatment group (group C) and simvastatin treatment group (group D). 24 rats in each group were treated with bleomycin solution and intratracheal administration of bleomycin solution to establish pulmonary fibrosis model. Group A was given the same amount of saline in the trachea, and group C was injected with the same volume of saline in the trachea at one time, and the rats in group C were given intratracheal administration of bleomycin in one dose. Group D was treated with prednisone suspension (5mg/kg d) and simvastatin suspension (10mg/kg d) on the first day after bleomycin modeling, while group A and group B were intragastrically perfused with normal saline (10ml/kg) of the same volume. The rats were randomly killed in each group on the 7th day 1428 day after the model, altogether 32 rats were killed. The changes of pulmonary alveolitis and pulmonary fibrosis were observed by HE staining, the expression of VEGF,PF4 protein and angiogenesis in lung tissue were detected by immunohistochemical method (SP), and the expression of VEGF and PF4mRNA in lung tissue were detected by RT-PCR method. Results: 1 the lung histopathological results showed that the lung tissue structure of group A was clear and complete, without hyperemia, edema, exudation and fibrosis. Fibroblast proliferation was observed. On the 14th day, the alveolar inflammation reached the peak, the alveolar septum widened obviously, the inflammatory cell infiltration increased, the fibroblast proliferated obviously, and on the 28th day, the pulmonary fibrosis was obvious, the alveolar cavity collapsed or disappeared, the fibroblasts proliferated and a large amount of collagen was deposited. The pulmonary alveolitis and fibrosis degree of steroid and simvastatin treatment group were alleviated in different degrees compared with the corresponding time points in model group. 2 comparison of the same time points among groups: the (MVD) of lung tissue in group B C was higher than that in group A. The difference was statistically significant. There was no significant difference between the two groups. The microvessel density in group D was significantly lower than that in group B. the difference was statistically significant. 3 Immunohistochemical and RT-PCR results showed that the expression of VEGF in group B and C was significantly higher than that in group A at the same time. The expression of VEGF in group D was significantly lower than that in group B. the expression of PF4 in group B and C was significantly lower than that in group A (P < 0.05). The expression of PF4 in group D was significantly higher than that in group B (P < 0.05). Conclusion: 1 Simvastatin can inhibit the angiogenesis of pulmonary tissue and alleviate the degree of alveolitis and pulmonary fibrosis in rats with pulmonary fibrosis. 2 the mechanism of simvastatin against pulmonary fibrosis may be related to its inhibition of VEGF expression in lung tissue. Relative up-regulation of PF4 expression and inhibition of pathological angiogenesis. 3 glucocorticoid can not reduce pulmonary angiogenesis during pulmonary fibrosis.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R563

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