Treatment Of Aneurysm By Transplantation Endothelial Progenitor Cells Transfected With Ang-1 Gene And The Differentiation Of EPCs In Aneurysm

Backgroup:The treatments of intracranial aneurysm(ICA)include endovascular coiling and surgical clipping approach.Compared with the surgical clipping approach,endovascular coiling has a higher risk for retreatment.This recurrence may be due to aneurysm sac growth,coil compaction and incomplete endothelialization of the aneurysm neck.Among them,the incomplete endothelialization of aneurysm neck plays a crucial role in the recurrence.Improving endothelialization of the aneurysm neck may therefore prevent blood from entering the aneurysm,thereby alleviating the risk of recurrence.Studies involving human clinical studies have indicated that the impaired function and decreased number of circulating Endothelial progenitor cells(EPCs)in aneurysm patients may contribute to aneurysm formation.EPCs can differentiate to endothelial cells(ECs)and contribute to vascular repair.Angiogenin-1(Ang-1)is a member of secreted glycoprotein family.Ang-1 is a key regulator for EPCs and angiogenesis events.Ang-1 signals primarily through the transmembrane receptor tyrosine kinase(Tie2),which is expressed in vascular endothelium and EPCs.Angl-Tie2 signal is a key regulator of angiogenic process including EPCs migration,differentiation,and tube formation.High serum Ang-1 levels mean a good functional outcome for aneurysmal subarachnoid hemorrhage patients.Therefore,overexpression of Ang-1 may enhance the function of EPCs,the Ang-1-EPCs transplantation may accelerate the endothelialization of aneurysm neck.In a previous research,we found that combined treatment with SDF-la-coated coils and EPCs transplantation could accelerate clot organization and endothelial cell proliferation.Immunofluorescence analysis suggested that ?-smooth muscle actin-positive cells were in organized tissues in aneurysm sacs,but it was not clear whether smooth muscle cells come from EPCs or not.In summary,the aim of the present study is to enhance EPCs functions by overexpressing Ang-1.And then we evaluate the effect of overexpressing Ang-1 in EPCs on the endothelialization in aneurysm sacs and necks.Moreover,we explore the differentiation of the transplanted EPCs in aneurysm.Upregulation of Ang-1 gene expression in EPCs through gene transfer may be a novel therapeutic strategy for aneurysm neck endothelial repair.Part ? Overexpression of the Ang-1 gene improves function and endothelialization capacity of EPCs in aneurysm modelsObjective:Proliferation and migration of endothelial progenitor cells(EPCs)play a critical role in vascular repairing processes.The aim of our study is to explore the effects of overexpressing Ang-1 in EPCs on aneurysm repairing and remodeling in a rat model of aneurysm.Materials and methods:EPCs were transfected with an adenovirus serotype 5 vector expressing Ang-1 or NC.The effects of overexpressing Ang-1 on viability and functioning of EPCs were measured via tube formation,wound healing assay and MTT assays.The endothelial repair process in the coiled aneurysms was evaluated via hematoxylin eosin staining,Masson trichrome staining and immunofluorescence analysis on days 14 after coil implantation.Results:Ang-1 significantly promoted the migration,tube formation and proliferation ability of EPCs in vitro.Histological analysis showed that the aneurysm neck endothelium was more integrated in the Ang-1-EPCs treated rats.Further study demonstrated that smooth muscle cells were in organized tissue in aneurysm sacs.Conclusions:The present study demonstrated that Ang-1 gene overexpression enhanced the tube formation,migration and proliferation ability of EPCs in vitro.Upregulation of Ang-1 gene expression accelerated focal fibrous tissue remodeling and the endothelialization of aneurysm neck orifice.Part ? The effect of smooth muscle cells on the differentiation of EPCs and differentiation of EPCs in aneurysmObjective:To explore the effect of smooth muscle cells(SMCs)on the differentiation of EPCs and whether EPCs can differentiate into SMCs in aneurysm.Materials and methods:EPCs were indirectly co-cultured with SMCs.Then real-time PCR were used to examine expressions of SMCs marker,i.e.,a-SMA,SM22a,CD31 and vWF.After the coil implantation,GFP-labeled EPCs(GFP-EPCs)were infused into vein immediately.The differentiation of EPCs in aneurysm was measured via immunofluorescence on day 14 after coil implantation.Results:Our results showed that co-culture of EPCs with SMCs increaseed the expression of a-SMA and SM22a(P<0.05),CD31(P>0.05),and decreased the expression of vWF(P<0.05).Immunofluorescence showed GFP and a-SMA double-positive cells in organized tissue in aneurysm sacs.Conclusions:SMCs played an important role in the differentiation of EPCs and EPCs could differentiate into SMCs and contribute to aneurysm repairing.