A Novel Flow Bioreactor for In Vitro Microvascularization

Although the importance of fluid flow for proper vascular development and function in vivo is well recognized, microvascular formation in response to flow has not been well evaluated in a three-dimensional (3D) environment in vitro. In this study, we developed a novel 3D in vitro perfusion system that allows direct investigation of the effects of shear stress on the development of microvasculature in vitro. This system utilizes a 3D collagen gel for suspension of vascular cells and mesenchymal stem cells, through which flow is directly perfused. We characterized the flow conditions and demonstrate the impact of flow on the development of microvasculature using a coculture of endothelial cells and mesenchymal stem cells. With the unique ability to apply bulk flow through the collagen gels, and to estimate shear stress within the constructs, this perfusion system provides a flexible platform for developing a controllable biomimetic environment that can be adapted for a variety of investigations of microvascularization

Quiescence and activation of stem and precursor cell populations in the subependymal zone of the mammalian brain are associated with distinct cellular and extracellular matrix signals

The subependymal zone (SEZ) of the lateral ventricles is one of the areas of the adult brain where new neurons are continuously generated from neural stem cells (NSCs), via rapidly dividing precursors. This neurogenic niche is a complex cellular and extracellular microenvironment, highly vascularized compared to non-neurogenic periventricular areas, within which NSCs and precursors exhibit distinct behavior. Here, we investigate the possible mechanisms by which extracellular matrix molecules and their receptors might regulate this differential behavior. We show that NSCs and precursors proceed through mitosis in the same domains within the SEZ of adult male mice—albeit with NSCs nearer ependymal cells—and that distance from the ventricle is a stronger limiting factor for neurogenic activity than distance from blood vessels. Furthermore, we show that NSCs and precursors are embedded in a laminin-rich extracellular matrix, to which they can both contribute. Importantly, they express differential levels of extracellular matrix receptors, with NSCs expressing low levels of α6β1 integrin, syndecan-1, and lutheran, and in vivo blocking of β1 integrin selectively induced the proliferation and ectopic migration of precursors. Finally, when NSCs are activated to reconstitute the niche after depletion of precursors, expression of laminin receptors is upregulated. These results indicate that the distinct behavior of adult NSCs and precursors is not necessarily regulated via exposure to differential extracellular signals, but rather via intrinsic regulation of their interaction with their microenvironment

Prdm6 controls heart development by regulating neural crest cell differentiation and migration.

The molecular mechanisms that drive the acquisition of distinct neural crest cell (NCC) fates is still poorly understood. Here, we identify Prdm6 as an epigenetic modifier that temporally and spatially regulates the expression of NCC specifiers and determines the fate of a subset of migrating Cardiac NCCs (CNCCs). Using transcriptomic analysis, genetic and fate mapping approaches in transgenic mice, we show that disruption of Prdm6 is associated with impaired CNCC differentiation, delamination, and migration, and leads to patent ductus arteriosus (DA)and ventricular noncompaction. Bulk and single-cell RNA-seq analyses of DA and CNCC identify Prdm6 as a regulator of a network of CNCC specification genes including Wnt1, Tfap2b, and Sox9. Loss of Prdm6 in CNCCs diminishes its expression in pre-EMT cluster, resulting in the retention of NCC in the dorsal neural tube. This defect is associated with diminished H4K20 mono-methylation and G1-S progression and augmented Wnt1 transcript levels in pre-EMT and neural tube clusters, which we show is the major driver of the impaired CNCC migration. Altogether, these findings reveal Prdm6 as a key regulator of CNCC differentiation and migration and identify Prdm6 and its regulated network as potential targets for the treatment of congenital heart diseases