钩藤生物碱对自发性高血压大鼠胸主动脉成纤维细胞凋亡/增殖及FN、LN的影响

Effects of Rhynchophylla Alkaloids on Vascular Adventitial Fibroblast Apoptosis and Proliferation in the Thoracic Aorta of Spontaneously Hypertensive Rats

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DOI：

中文关键词: 钩藤碱异钩藤碱钩藤总生物碱自发性高血压主动脉外膜成纤维细胞凋亡/增殖

英文关键词:rhynchophylline isorhynchophylline rhynchophylla alkaloid spontaneous hypertension vascular adventitial fibroblasts apoptosis/proliferation

基金项目:国家自然科学基金资助项目(No.30300466);教育部新世纪优秀人才支持计划资助项目(No.NCET-07-0522);山东省自然科学基金资助项目(No.Y2006C99)

作者	单位
戴国华	山东中医药大学附属医院心内科
孙敬昌	山东中医药大学
齐冬梅	山东中医药大学
周洪雷	山东中医药大学
李运伦	山东中医药大学附属医院心内科

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中文摘要:

目的研究钩藤碱、异钩藤碱和钩藤总生物碱对自发性高血压大鼠(spontaneously hypertensive rat,SHR)胸主动脉血管外膜成纤维细胞(vascular adventitial fibroblasts,VAF)凋亡、增殖及Bcl-2、Bax、c-Fos、c-Myc、层黏连蛋白(laminin,LN)、纤维层黏连蛋白(fibronectin,FN)的影响。方法选择8周龄雄性SHR40只,随机分为模型组、卡托普利组(17.5mg/kg)、异钩藤碱组(5.0mg/kg)、钩藤碱组(5.0mg/kg)和钩藤总生物碱组(50.0mg/kg),每组8只。另选取8周龄雄性Wistar大鼠8只作为正常组。模型组和正常组灌胃给予等容量生理盐水。各组给药容积为每次10mL/kg,每周给药6天,连续8周,随体重变化调整给药量。采用AnnexinV-FITC结合PI染色、流式细胞术分析胸主动脉VAF凋亡率;免疫组化法检测胸主动脉VAFBcl-2、Bax、c-Myc、c-Fos、FN及LN蛋白表达;原位杂交法检测胸主动脉VAF转化生长因子-β1(transforming growth factor,TGF-β1)mRNA表达。结果与模型组比较,各用药组干预4、8周大鼠尾动脉收缩压均降低,胸主动脉VAF凋亡率、Bax蛋白均升高,Bcl-2、c-Fos、FN、LN蛋白及TGF-β1mRNA均降低,钩藤碱组、异钩藤碱组及钩藤总生物碱组c-Myc蛋白表达均降低,差异均有统计学意义(P<0.05,P<0.01)。与卡托普利组比较,钩藤碱组、异钩藤碱组及钩藤总生物碱组大鼠尾动脉收缩压、c-Fos蛋白表达及TGF-β1mRNA差异无统计学意义(P>0.05);胸主动脉VAF凋亡率升高,Bcl-2、c-Myc及LN蛋白表达均降低,差异均有统计学意义(P<0.05,P<0.01);钩藤碱组、异钩藤碱组Bax蛋白表达均升高,差异均有统计学意义(P<0.05,P<0.01);异钩藤碱组FN蛋白表达降低,差异有统计学意义(P<0.05)。钩藤碱组、异钩藤碱组及钩藤总生物碱组间比较,差异无统计学意义(P>0.05)。结论钩藤碱、异钩藤碱和钩藤总生物碱可能通过调节原癌基因Bcl-2和Bax蛋白表达,促进SHR胸主动脉VAF凋亡,通过下调c-Fos、c-Myc蛋白和TGF-β1mRNA表达途径抑制SHR胸主动脉VAF增殖,并通过下调FN和LN蛋白表达而减轻细胞外基质的沉积,从而改善胸主动脉外膜重塑,降低尾动脉收缩压。

英文摘要:

Objective To study the effects of rhynchophylline,isorhynchophylline,and rhynchophylla alkaloids on the vascular adventitial fibroblasts(VAF) apoptosis and proliferation in thoracic aorta of spontaneously hypertensive rats(SHR),and on the Bcl-2,Bax,c-Fos,c-Myc,laminin(LN),and fibronectin(FN).Methods Forty 8-week old male SHR were randomly divided into five groups,i.e.,the model group,the captopril group(17.5 mg/kg),the isorhynchophylline group(5.0 mg/kg),the rhynchophylline group(5.0 mg/kg),and the rhynchophylla alkaloids group(50.0 mg/kg),8 in each group.In addition,eight 8-week old male Wistar rats were selected as the normal group.Equal volume of normal saline was given to rats in the normal group and the model group by gastrogavage.Rats in the rest groups were perfused with isovolumic medication solution(10 mL/kg),six days per week for eight successive weeks.The dosage of drugs was adjusted according to the change of body weight.The VAF apoptosis rate of the thoracic aorta was measured by Annexin V-FITC combined with PI dyeing and flow cytometry.The protein expressions of thoracic aortic Bcl-2,Bax,c-Myc,c-Fos,FN,and LN were detected by immunohistochemical assay.The adventitial transforming growth factor β1(TGF-β1) mRNA expression in the thoracic aorta was detected by in situ hybridization method.Results Compared with the model group,the tail arterial systolic pressure decreased,the VAF apoptosis and the protein expression of Bax increased,Bcl-2,c-Fos,FN,LN,and TGF-β1 mRNA all decreased in the thoracic aorta of SHR in each treatment group after 4-and 8-week of intervention.Rhynchophylline,isorhynchophylline,and rhynchophylla alkaloids could inhibit the protein expression of c-Myc with statistical difference(P<0.05,P<0.01).Compared with the captopril group,there was no statistical difference in decreasing the tail arterial systolic pressure,the protein expression of c-Fos and the mRNA expression of TGF-β1 among the rhynchophylline group,the isorhynchophylline group,and the rhynchophylla alkaloids group(P>0.05)。There was statistical difference in increased VAF apoptosis and decreased protein expressions of Bcl-2,c-Myc,and LN(P<0.05,P<0.01).There was statistical difference in increased protein expression of Bax between the rhynchophylline group and the isorhynchophylline group(P<0.05,P<0.01).There was statistical difference in decreased protein expression of FN in the isorhynchophylline group(P<0.05).There was no significant difference among the rhynchophylline group,the isorhynchophylline group,or the rhynchophylla alkaloids group(P>0.05).Conclusions Rhynchophylline,isorhynchophylline,and rhynchophylla alkaloids might promote the VAF apoptosis in the thoracic aorta of SHR by regulating the protein expressions of Bcl-2 and Bax.They might inhibit the VAF proliferation by restraining protein expressions of c-Fos,c-Myc,and TGF-β1 mRNA.They also might improve the thoracic aorta wall reconstruction and decrease the tail arterial systolic pressure by down-regulating the protein expressions of FN and LN,and attenuating the deposition of extracellular matrix.

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