TGF-β2调控血管平滑肌细胞表型转化在静脉血栓管壁重塑中的作用及其机制

Effect and mechanism of phenotypic transformation of vascular smooth muscle cells regulated by TGF-βon the venous thrombus wall remodeling

目的探讨静脉血栓后管壁重塑过程中转化生长因子β2(TGF-β2)调控血管平滑肌细胞(SMC)表型转化的作用及其机制。方法用0、1、10、20 ng/ml TGF-β2刺激人脐静脉平滑肌细胞(HVSMCs)24 h,以及10 ng/ml TGF-β2刺激HVSMCs 0、2、6、24 h,分别提取RNA,采用RT-PCR检测HVSMCs表型标志物(收缩型:α-SMA和Elastin;合成型:Col1A1和Col1A2)mRNA的表达。建立SD大鼠下腔静脉血栓模型,将大鼠随机分为假手术组(n=5)、血栓组(n=9)和TGF-β2实验组(n=5)。造模后第4天和第7天,采用HE和Masson染色进行验证。造模后第14天,用含TGF-β2(10 ng/只)的水凝胶在血栓局部处理24 h,采用RT-PCR检测各组静脉管壁SMC表型标志物mRNA的表达。结果TGF-β2可上调HVSMC收缩型和合成型标志物mRNA的表达且呈剂量和时间依赖性(P<0.05),其中对收缩型标志物mRNA表达的上调作用更为明显。HE和Masson染色显示急性期(造模后第4天)血栓形成和慢性期(造模后第7天)血栓机化再通,证实下腔静脉血栓造模成功。造模后第14天,与假手术组比较,血栓组静脉管壁SMC收缩型标志物mRNA的表达明显下调,合成型标志物mRNA的表达明显上调(P<0.05)。在血栓局部使用TGF-β2处理后,收缩型标志物mRNA的表达上调更为明显(P<0.05)。结论TGF-β2可诱导HVSMC收缩型和合成型标志物的表达,其中对收缩型标志物的上调作用更为明显。在大鼠下腔静脉血栓模型中,TGF-β2可明显上调SMC收缩型标志物的表达,有利于维持血栓段静脉管壁SMC的收缩表型。

Objective To investigate the role and mechanism of transforming growth factor-β2(TGF-β2)on regulating the phenotype transformation of vascular smooth muscle cells(SMC)during the wall remodeling after venous thrombosis.Methods Human umbilical vein smooth muscle cells(HVSMCs)were treated with different concentration(0,1,10,20 ng/ml)of TGF-β2 for 24 hours,or treated with 10 ng/ml TGF-β2 at different time points(0,2,6,24 h).RT-PCR was performed to detect the mRNA expressions of phenotype markers(contractile type:α-SMA and Elastin;synthetic type:Col1A1 and Col1A2).SD rat models of inferior vena cava thrombosis(IVCT)were established.Rats were randomly divided into sham surgery group(n=5),thrombus group(n=9)and TGF-β2 experimental group(n=5).The successful establishment of rat model was confirmed by HE and Masson staining of vein wall tissues on 4 and 7 days after modelling.At the 14th day after modelling,IVCT rats were locally treated with hydrogel containing TGF-β210 ng per rat for 24 hours.RT-PCR was used to detect the expression of SMC phenotypic marker mRNA in the vein wall of each group.Results TGF-β2 up-regulated mRNA expressions of both contractile and synthetic markers in HVSMCs in a dose-and time-dependent manner(P<0.05).Notely,the mRNA expression of contractile markers was more up-regulated than that of synthetic markers.HE and Masson staining showed the acute thrombosis phase 4 days after modelling and the chronic organized thrombus and recanalization 7 days after modelling,confirming the successful establishment of IVCT models.At the 14th day after modelling,compared with the sham surgery group,the mRNA expression in SMC venous wall contractile markers of thrombus group was down-regulated obviously,and was up-regulated markedly in synthetic markers(P<0.05).When TGF-β2 was used to locally treat the thrombus,the expressions of contractile markers mRNA were more significantly up-regulated(P<0.05).Conclusions TGF-β2 may induce the expression of contractile and synthetic markers in HVSMCs,particularly up-regulating the contractile markers.In IVCT rats model,TGF-β2 can significantly up-regulate the mRNA expressions of SMC contraction markers,which is in favour of maintenance of SMC contractile phenotype in the thrombus vein wall.