Study Of Smooth Muscle Cell And Endothelia Cell In Vascular Injury And Molecular Mechanisms

Chapter 1 SM22-alpha VSMC-lineage tracking during vascular remodeling in a femoral artery injury modelObjectives:Vascular smooth muscle cell(VSMC) expansion and Endothelia cell damage are the main feature of vascular wall thickening after an injury during the vascular remodeling.However,SMC depletion at the early stages warrants investigation into the sources of SMCs during vascular remodeling.To determine the fate of vascular smooth muscle cells after vascular injury.Methods:we generated a transgenic mouse line,SM22Cre mice expressing a tamoxifen-inducible Cre recombinase,which is controlled by the smooth muscle-specific SM22 promoter.SM22Cre mice were crossed with mice carrying the yellow fluorescent protein(YFP) controlled by the ubiquitous R26R promoter but with a floxed stop codon in between;in effect,tamoxifen injections resulted in YFP expression exclusively in vascular SMCs.We established a femoral artery wire injury model for Sm22aCreYFP mice,and harvested tissues at the following time points:hour 0,day 1,day 7,day 14 and day 21.Results:In Sm22aCreYFP mice-wire injury model,we found that approximately 50%of vascular SMCs express YFP in the media with a single-layer of endothelial cells in the intima.At day 21,the YFP-expressing cells in the media decreased to around 10%.In contrast, YFP-expressing cells composed approximately 50%of the total neointimal cells.Conclusion:These findings demonstrate that the medial SMCs contribute to the formation of the neointimal SMCs in vascular remodeling. Chapter 2 Tie2-endothelial lineage tracking in a murine vein graft modelObjectives:It is very important to study proliferate endothelial cell and smooth muscle cell and mutual relation and morphology during a vascular remodling,for those are essential vascular cells.To determine the fate of endothelial cells after vein graft dring the vascular remodeling.Methods:we generated a transgenic mouse line,Tire2Cre mice, expressing a tamoxifen-inducible Cre recombinase,which is controlled by the endothelia cell cell-specific Tie2 promoter.Tire2Cre mice were crossed with mice carrying the yellow fluorescent protein(YFP) controlled by the ubiquitous R26R promoter but with a floxed stop codon in between;in effect,tamoxifen injections resulted in YFP expression exclusively in vascular endothelial cells.We established a Jugular vein graft to femoral artery model for Tie2creYFP mice,and harvested tissues at the following time points:hour 0,day3 day,day 7,day 14 and day 35.Results:In Tie2CreYFP mice-Jugular vein graft model,we found that Endothelial-lineage cells lose endothelial markers and gain SMC characteristics as they proliferate in the neointima in vivo.The endothelial cells(eoma cells) which were treated with TGFβ1 for 4 days gain SMC characteristics in vitro.Conclusion:These findings demonstrate that Endothelial-lineage cells contribute to the formation of the neointimal SMCs in vascular remodeling. Chapter 3 cyclin dependent kinase inhibitors play a important role in vascular injuryObjectives:p21^Cip1,one of the cyclin dependent kinase inhibitors,is implicated in physiologic cell turnover during normal regenerative cycles, as well as being active players in vascular remodeling.To determine lesion formation in mice with homozygous deletion of p21^Cip1 resulted in accelerated proliferation of vascular smooth muscle cells.Methods:We established a wire injury model for p21-/- mice and wild type mice,and harvested tissues at the following time points:day7 and day 14.Results:The p21-/- mice increase neointima formation compare to WT mice indicates that p21^Cip1 plays a important role in restraining excessive proliferation during vascular wound repair in local vascular cells.The deletion of p21^Cip1 increased apoptosis in vascular cells,which suggests that p21^Cip1 plays a role in the control of both proliferation and cell death.Conclusion:These findings demonstrate that p21^Cip1 activity in local vascular cells is essential for regulation of both cell proliferation and death in response to vascular injury,and it is a key mediator of vascular proliferation in response to injury.