Impact Of Transcriptional Factor SP1 In Aortic Dissection Via Regulating Vascular Smooth Muscle Cell Phenotype Switching

Objective : To investigate the relationship between transcriptional factor SP1 and phenotype switch of vascular smooth muscle cell(VSMC)in aorta,by detecting the quantity of SP1 and SM22α,marker of contractile phenotype VSMC,in the tissue from human aortic dissection(AD),Sprague-Dawley(SD)rat model of AD,and VSMC cultured in vitro.To investigate the impact of SP1 in occurrence of aortic dissection(AD)Methods:The research included three main parts: Part 1 14 pieces of human aortic wall tissue were collected including 10 pieces from AD patients and 4 from non AD patients.Expression of SP1 and SM22α were detected in both protein and m RNA level via Western-Blot and RT-PCR.And correlation analysis was performed afterwards.Part 2 In this section,we first built an animal model of aortic dissection in SD rats,by administration of β-amino propionitrile(BAPN)in drinking water.Twenty SD rats(3 weeks,male)were involved and divided randomly into two groups,control group(CG)and treatment group(TG).Each group contained 10 rats.Rats in CG were given normal mineral water,while those in TG were given 0.25% BAPN solution as drinking water.General conditions of both groups were observed daily,and weight of every rat was recorded every 2 to 3 days.Rats were anatomized on death during the experiment or on execution after 6 weeks,and the whole aorta was collected.HE/VB staining and transmission electron microscope(TEM)observation of the samples were performed in order to distinguish the existence of AD histologically and morphologically.Sp1 and Sm22α were measured via Western-Blot and RT-PCR.Immunohistochemical staining were also performed to locate the main site of SP1 in VSMC.Part 3.Human aorta smooth muscle cell(HASMC)was cultured in vitro.Meanwhile,human SP1 overexpression adenovirus vector(Ad.SP1)was constructed.The quantity of Sm22α in HASMC was detected via Western-Blot and RT-PCR,following the overexpression of SP1 by transfection of Ad.SP1.Results: 1.From the human samples,we found that SP1 expression of AD tissue was upregulated than that of normal,and SM22α expression is downregulated.Statistic analysis indicated a negative correlation between SP1 and SM22α.2.The animal model was successfully built with an AD occurrence rate of 100% and AD rupture rate of 100% in TG,and non AD or death occurred in CG.This result was confirmed by the HE/VB staining and TEM.It is noted that every AD occurred in TG was THORACIC aortic dissection(TAD),Indicating this method of building AD animal model is simple yet efficient.The results of Western-Blot and RT-PCR of tissue from animal experiment were similar to those from human tissue in part one.So was the statistical analysis,showing a negative correlation between SP1 and SM22α.From the Immunohistochemical staining of the samples,we located SP1 to be mainly distributed inside the nucleus of the VSMC in aorta.Furthermore,there was more heavy staining near the site of rupture.3.The human SP1 overexpression adenovirus vector(Ad.SP1)can efficiently transfection HASMC cultured in vitro,and successfully overexpress its SP1.In this kind of cell,we detected less quantity of contractile phenotype marker SM22α,indicating the cell switched from contractile to synthetic phenotype..Conclusion: In AD tissue,SP1 is upregulated and VSMC phenotype switch occurs,and they are correlated.Vitro experiment revealed that overexpression of SP1 induced VSMC phenotype switch.All these evidences indicate that Sp1 may participate in the occurrence of AD via inducing phenotype switch of VSMC.