A strategy to discover new organizers identifies a putative heart organizer

Organizers are regions of the embryo that can both induce new fates and impart pattern on other regions. So far, surprisingly few organizers have been discovered, considering the number of patterned tissue types generated during development. This may be because their discovery has relied on transplantation and ablation experiments. Here we describe a new approach, using chick embryos, to discover organizers based on a common gene expression signature, and use it to uncover the anterior intestinal portal (AIP) endoderm as a putative heart organizer. We show that the AIP can induce cardiac identity from non-cardiac mesoderm and that it can pattern this by specifying ventricular and suppressing atrial regional identity. We also uncover some of the signals responsible. The method holds promise as a tool to discover other novel organizers acting during development

Cardiac progenitor migration and specification: The dual function of Wnts

The heart is the first organ to function during vertebrate development and cardiac progenitors, are among the first cell lineages to be established from mesoderm cells emerging from the primitive streak during gastrulation. Cardiac progenitors have been mapped in the epiblast of pre-streak embryos. In the early chick gastrula they are located in the mid-primitive streak, from which they enter the mesoderm bilaterally. However, migration routes of cardiac progenitors have never been directly observed within the embryo and the factor(s) controlling their movement are not known. Furthermore, it is not understood how signals controlling cell movement are integrated with those that determine cell fate. Long-term video microscopy combined with GFP labelling and image processing enabled us to observe the movement patterns of prospective cardiac cells in whole embryos in real time. Embryo manipulations and the analysis of explants suggest that Wnt3a plays a crucial role in guiding these cells through a RhoA dependent mechanism involving negative chemotaxis. Wnt3a is expressed at high levels in the amniote primitive streak and ectopic signalling activity caused wider movement trajectories resulting in cardia bifida, which was rescued by dominant-negative Wnt3a. Our studies revealed Wnt3a-RhoA mediated chemo-repulsion as a novel mechanism guiding cardiac progenitors. This activity can act at long-range and does not interfere with cardiac cell fate specification

Differentiation of Embryonic Stem Cells into Cardiomyocytes in a Microfluidic System

The differentiation process of murine embryonic stem cells into cardiomyocytes was investigated with a compliant microfluidic platform which allows for versatile cell seeding arrangements, optical observation access, long-term cell viability, and programmable uniaxial cyclic stretch. Specifically, two environmental cues were examined with this platform—culture dimensions and uniaxial cyclic stretch. First, the cardiomyogenic differentiation process, assessed by a GFP reporter driven by the α-MHC promoter, was enhanced in microfluidic devices (µFDs) compared with conventional well-plates. The addition of BMP-2 neutralizing antibody reduced the enhancement observed in the µFDs and the addition of exogenous BMP-2 augmented the cardiomyogenic differentiation in well plates. Second, 24 h of uniaxial cyclic stretch at 1 Hz and 10% strain on day 9 of differentiation was found to have a negative impact on cardiomyogenic differentiation. This microfluidic platform builds upon an existing design and extends its capability to test cellular responses to mechanical strain. It provides capabilities not found in other systems for studying differentiation, such as seeding embryoid bodies in 2D or 3D in combination with cyclic strain. This study demonstrates that the microfluidic system contributes to enhanced cardiomyogenic differentiation and may be a superior platform compared with conventional well plates. In addition to studying the effect of cyclic stretch on cardiomyogenic differentiation, this compliant platform can also be applied to investigate other biological mechanisms.Singapore-MIT Alliance for Research and TechnologyAmerican Heart AssociationNational Science Foundation (U.S.) (Science and Technology Center (EBICS): Emergent Behaviors of Integrated Cellular Systems, Grant CBET-0939511)International Research & Development Program (Grant number 2009-00631

Examining the cardiac nk-2 genes in early heart development

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