Mechanisms of embryonic vascular mural cell differentiation and the onset of blood flow regulation

点击翻译按钮获取中文摘要

Blood vessels respond to increased or decreased metabolic needs by regulating blood flow, adjusting to ensure tissues and organs receive adequate blood supply. The multi-layered vascular wall has two key cell types, the inner endothelial lining and the external mural' layer. Pericytes are single mural cells that associate with smaller vessels on the vascular tree, while vascular Smooth Muscle Cells (vSMCs) form layers on larger vessels. In embryonic development, once the endothelial lining of the vessel assembles, it immediately is subjected to fluid flow forces. Naked' vessels can only passively carry flow, but as mural cells adhere and differentiate, they begin to contract and relax to ensure steady blood supply.***In preliminary data we have determined an early vSMC transcriptome and discovered mechanisms of paracrine signalling leading to vSMC differentiation. We have developed Computational Fluid Dynamics analysis of blood flow. We have developed methods to quantitate vessel diameter changes in vivo in response to pharmacological agents and we have determined the timing of both marker expression and of functional contraction and relaxation responses. These preliminary data and novel techniques now poise us to ask questions at the junction of developmental biology and vascular function.***In our NSERC program we hypothesize that blood flow and vascular mural cells each influence each other during development through multiple mechanisms.******Aim 1: Probe how mural cell attachment to the developing endothelium controls blood flow***Here we study the interacting effects of vSMC and pericyte differentiation on blood flow. We integrate computational fluid dynamics analysis of vessel architecture with live imaging data of mural cells over different developmental stages from the period before mural cells associate with endothelium through the stages where they become contractile. We genetically ablate mural cells to understand differences in vessel architecture. We design constructs for optogenetic activation of contraction in vSMCs.***Aim 2: Understand the role of flow-sensitive, mechanosensory pathways in vSMC differentiation***Here we test the relative roles of mechanosensation and signalling pathways reliant on blood flow on mural cell differentiation probing the function of 3 mechanosensory proteins and a circulating BMP on smooth muscle differentiation.***Aim 3: Determine how Ca2+ dynamics change with mural cell differentiation and how spatial control of Ca2+ affects the vascular network.***This aim will investigate the molecular mechanisms of functional vSMC differentiation by understanding how vascular tone develops through the regulation of Ca2+ dynamics using the live reporter GCamp5G expressed in pericytes or vSMCs to determine how Ca2+ handling changes over the period when mural cells gain contractile function.**