The Effects And Mechanism Of TGF-? Signaling Pathway In Vascular Smooth Muscle Cells In Abdominal Aortic Aneurysm

Objectives:Aneurysm refers to the abnormal enlargement of the local part of the blood vessel,usually caused by the thinning of the arterial wall due to vascular injury.The mass is undulating and looks like a tumor-like tissue,so it is called an aneurysm.Aneurysm can occur in any blood vessel.Among the aneurysms with high mortality,there are aneurysms that occur in the Willis ring of the brain,thoracic aortic aneurysms that occur in the thoracic aorta,and abdominal aortic aneurysms that occur in the abdominal aorta.Aneurysms such as atrial septal aneurysms and ventricular aneurysms can also occur in the heart after myocardial infarction.Due to the hemodynamic and anatomical features of the abdominal aorta,the incidence and rupture rate of abdominal aortic aneurysms is highest in all aortic aneurysms.Two major risk factors for the development of elderly and male abdominal aortic aneurysms,as well as smoking and hypertension,are also important risk factors.chronic inflammation of the vessel wall,apoptosis of vascular smooth muscle cells,and destructive remodeling of extracellular matrices are important histopathological features of abdominal aortic aneurysms.Therefore,inhibition of vascular inflammation,apoptosis of vascular smooth muscle cells,and maintenance of homeostasis of the extracellular matrix may inhibit the occurrence of abdominal aortic aneurysm and improve the prognosis of abdominal aortic aneurysm.The transforming growth factor?TGF-?1?signaling pathway was inhibited in human abdominal aortic aneurysm tissue samples,and the use of TGF-?1neutralizing monoclonal antibody significantly aggravated abdominal aortic aneurysm in animal models of abdominal aortic aneurysm.These findings suggest that activation of the TGF-?signaling pathway may play a protective role in abdominal aortic aneurysms.There are also articles directly pointing out that the TGF-?signaling pathway plays a protective role in abdominal aortic aneurysms,but the specific mechanism of its protective effect remains unclear.The main purpose of this scientific research is to find a specific mechanism for the protective effect of TGF-?signaling pathway in abdominal aortic aneurysm.On the other hand,the key genes for the occurrence of abdominal aortic aneurysm are sought to provide therapeutic targets for the treatment of abdominal aortic aneurysms.Cell assay:Administration of human smooth muscle cell recombinant protein TGF-?1?5 ng/mL?at different time points,extract RNA from the cells,the genes associated with abdominal aortic aneurysm in the TGF-?signaling pathway reported in the literature were analyzed.Differential genes were screened by real-time polymerase chain reaction?RT-PCR?.The results showed that TGF-?1?5ng/mL?can significantly increase the expression of potential transforming growth factor binding protein 4 in vascular smooth muscle cells.Further examination of the protein level of LTBP4 revealed that changes in protein levels were consistent with changes in transcription levels.Studies have shown that LTBP4 can participate in the assembly process of elastin,promote the formation of elastic fibers,and increase the elasticity of blood vessels.The fracture of the elastic plate and the degradation of the extracellular matrix are also one of the pathological features of the abdominal aortic aneurysm.Therefore,it is proposed that the scientific hypothesis that TGF-?1 promotes the secretion of LTBP4 by vascular smooth muscle cells to increase the formation of elastic fibers is an important mechanism for its protective effect in abdominal aortic aneurysms.The way TGF-?1 regulates downstream gene expression is mainly through activation of downstream transcription factor SMADs,which promotes its phosphorylation translocation into nuclear regulation of downstream target gene expression.After translocation into the nucleus as a transcriptional coactivator,the YAP protein regulates transcription of the gene by binding to a specific transcription factor in the nucleus.There are many common risk factors for atherosclerosis and abdominal aortic aneurysm.Therefore,in order to explore whether YAP protein is involved in TGF-?1 transcriptional regulation of LTBP4.Human smooth muscle cells were treated with TGF-?1,and the phosphorylation status and nuclear localization of the common three phosphorylation sites controlled by YAP protein were detected by immunoblotting.The results showed that TGF-?1 phosphorylates the YAP tyrosine?Y357?site in a time-and dose-dependent manner,and promotes the translocation of YAP into the nucleus.Further protein immunoprecipitation experiments showed that YAP has direct binding to the transcription factor p-SMAD2/3.These results suggest that YAP protein may be involved in the process of TGF-?1 transcriptional regulation of LTBP4.To further verify whether YAP plays an important role in the transcriptional regulation of LTBP4 by TGF-?1.We performed a series of YAP protein adenovirus overexpression and YAP protein siRNA silencing experiments in human vascular smooth muscle cells.The experimental results show that:1.YAP protein can regulate transcription of LTBP4.TGF-?1 up-regulates the expression of LTBP4 by promoting the translocation of YAP protein and p-SMAD2/3 into the nucleus.Animal experiments:Studies have shown that LTBP4 is involved in the assembly of elastin,which in turn increases the formation of elastic fibers and increases the elasticity of blood vessels.Therefore,an elastase-induced abdominal aortic aneurysm model was preferred in animal models to verify whether YAP protein plays a protective role in the regulation of LTBP4 transcription in the formation of abdominal aortic aneurysms.We constructed a smooth muscle cell-specific overexpressing YAP protein mouse.The expression of LTBP4 in the blood vessels of overexpressed mice was first tested.Consistent with the results of the cell experiments:the expression of LTBP4 in the blood vessels of mice overexpressing YAP protein was significantly increased.SMC-YAP^tg mice?8 weeks old?and SMC-YAP^flox?8 weeks old?were divided into the following four groups:SMC-YAP^tg mice?elastase surgery group,sham operation group?,SMC-YAP^flox mice?elastase Surgical group,sham operation group?.The incidence of abdominal aortic aneurysm,the diameter of the tumor and the fracture of the elastic plate were counted two weeks after surgery.The results showed that smooth overexpression of YAP by smooth muscle cells significantly improved the severity of elastase-induced abdominal aortic aneurysm.Combining current experimental results:This study suggests that one of the possible mechanisms by which TGF-?1 plays a protective role in abdominal aortic aneurysm is to increase the assembly of elastin by regulating the expression of LTBP4,thereby increasing the stability of aneurysms and Inhibits the continued expansion of an aneurysm.In this process,YAP protein plays a key role as a transcriptional coactivator and can be used as a new target for the treatment of AAA.