The Mechanism Of Action Of Telomerase Activity Changes In The Development Of Atherosclerotic Plaques

BackgroundVascular calcification,especially atherosclerotic calcification,easily lead to hypertension,myocardial ischemia,left ventricular hypertrophy and heart failure,which is an important risk factor for the high morbidity and mortality of cardiovascular and cerebrovascular diseases.Vascular calcification,innitaed by various cells,is similar to the bone development,active,highly regulabe,preventable and invertible biological process.The current theories of vascular calcification formation mainly include the bone morphogenetic proteins regulation,lipid metabolic disorder,apoptosis body matrix vesicles and oxidative stress,but the specific molecular mechanism is still unclear.Vascular calcification mainly occurs in vascular medial vascular smooth muscle cells,which can detect telomerase expression.The previous studies reported that the telomere length of atherosclerotic plaque VSMCs were significantly shorter than normal vascular,as well as telomerase activity,but the role of telomerase in vascular calcification is unclear.Several studies have shown that the NF-?B signal pathway play a key role in the process of vascular calcification.The inflammatory factors promote the expression of bone morphogenetic proteins and other related protein by activating the NF-?B pathway and affect the process of vascular calcification.A recent study found that heterogeneous nuclear ribonucleoprotein A1 combined with the I?B? and mediated I?B? degration by protease hydrolysis,and activated the NF-?B signal pathway.Our preliminary data showed that the expression level of hTERT(human telomerase reverse transcriptase)and hnRNP A1 significantly increased during HUASMCs differentiation from contractile to synthetic smooth muscle cells.It has been reported that hnRNP A1 can prolong the telomere length and prevent cells premature senescence caused by excessive telomere loss.Therefore,we aimed to study whether hnRNP A1 can promote the calcification of vascular smooth muscles via telomerase regulation and NF-?B pathway.Materials and Methods1.The human umbilical artery smooth muscle cells(HUASMCs)were separated and primarily cultured,and a eukaryotic expression vector pcDNA3.1-hnRNP A1 was constructed to study the role of hnRNP A1 in the hTERT regulation and HU ASMCs proliferation and apoptosis.2.The HUASMCs calcification model was established to explore the mechanisms of hnRNP A1 in arterial calcification.Results1.The results showed that the expression level of hTERT and hnRNP A1 significantly increased during HUASMCs differentiation from contractile to synthetic smooth muscle cells.Overexpression of hnRNP A1 upregulated telomerase activity 2.7-fold,but has no effects on proliferation and apoptosis of HUASMCs.2.HUASMCs calcification model showed that the overexpression of hnRNP A1 in HUASMCs obviously promoted the cell calcification,but the role seems not to be mediated via the I?B?/NF-?B signal pathway.ConclusionIn our study,hnRNP A1 and hTERT significantly increased during HUASMCs phenotypic switch process,and overexpression of hnRNP A1 upregulated telomerase activity and promoted cell calcification,whereas not mediated by the I?B?/NF-?B pathway.BackgroundAtherosclerosis is an aging associated chronic inflammatory respone.The monocyte/macrophage plays a critical role in the inflammation,which involved in all stages of atherosclerosis.The activation of monocyte/macrophage will exacerbate inflammatory process,vascular smooth muscle cells calcification and lipid accumulation,ultimately lead to plaque rupture,sudden death,acute myocardial infarction,and ischemic stroke.The telomerase activity has been reported to increase during atherosclerosis formation;however,the molecular mechanisms underlying telomerase activation remain unknown.Our previous work showed that human telomerase reverse transcriptase(hTERT)expression was particularly observed in the macrophage-rich region of atherosclerotic lesions.Interestingly,we have identified that a particular microRNA,miR-216a,was remarkably up-regulated in macrophages differentiation in phorbol-12-myristate-13-acetate(PMA)-stimulated THP-1 monocyte cells,paralleling with the increased expression of hTERT.In the present study,we aimed to examine the role of a particular microRNA,miR-216a,in the activation of telomerase during macrophages differentiation.Methods1.To evaluate telomerase activity in the atherosclerotic lesion,we performed immunohistochemical and immunofluorescent test on the atherosclerotic carotid plaques(n=5)and the corresponding near normal carotid arteries from patients with carotid atherosclerosis undergoing carotid endarterectomy.2.THP-1 monocyte cells,as the precursor of macrophages,induced by PMA stimulus for 48 hours,were cultured in vitro to investigate the regulation role of miR-216a on hTERT expression.The atherosclerosis model of ApoE-/-male mice was performed to determine whether miR-216a media the monocyte/macrophage activation and involved in the development of carotid atherosclerotic plaques.Results1.Firstly,we found that hTERT expression was negligible in normal human carotid arteries,but highly presented in atherosclerotic lesions.Particularly,the elevated level of hTERT expression was observed in macrophage-rich region.The co-localization of hTERT with macrophage CD68 immuno-reactivity was further verified in the atherosclerotic plaques by immunofluorescent staining analysis.2.In vitro,the results showed that miR-216a overexpression markedly increased hTERT mRNA expression and telomerase activity.Furthermore,we found miR-216a mediates the activation of telomerase in macrophages differentiation via the SMAD3/NF-?B pathway.3.The ApoE-/-mouse model showed that miR-216a promoted M1 macrophages polarization and inhibited M2 differentiation in carotid atherosclerosis lesions,and also,miR-216a reduced the type ? collagen in plaques,which is associated with the stability of the plaques.ConclusionOur data indicated that miR-216a mediates the activation of telomerase in macrophages differentiation via the SMAD3/NF-?B pathway.MiR-216a promotes M1 macrophages differentiation and reduces type ? collagen in mice atherosclerosis lesions,ultimately accelerates the development of atherosclerotic plaque.