Dynamic positional fate map of the primary heart-forming region

Here we show the temporal-spatial orchestration of early heart morphogenesis at cellular level resolution, in vivo, and reconcile conflicting positional fate mapping data regarding the primary heart-forming field(s). We determined the positional fates of precardiac cells using a precision electroporation approach in combination with wide-field time-lapse microscopy in the quail embryo, a warm-blooded vertebrate (HH Stages 4 through 10). Contrary to previous studies, the results demonstrate the existence of a “continuous” circle-shaped heart field that spans the midline, appearing at HH Stage 4, which then expands to form a wide arc of progenitors at HH Stages 5-7. Our time-resolved image data show that a subset of these cardiac progenitor cells do not overlap with the expression of common cardiogenic factors, Nkx-2.5 and Bmp-2, until HH Stage 10, when a tubular heart has formed, calling into question when cardiac fate is specified and by which key factors. Sub-groups and anatomical bands (cohorts) of heart precursor cells dramatically change their relative positions in a process largely driven by endodermal folding and other large-scale tissue deformations. Thus, our novel dynamic positional fate maps resolve the origin of cardiac progenitor cells in amniotes. The data also establish the concept that tissue motion contributes significantly to cellular position fate — i.e., much of the cellular displacement that occurs during assembly of a midline heart tube (HH Stage 9) is NOT due to “migration” (autonomous motility), a commonly held belief. Computational analysis of our time-resolved data lays the foundation for more precise analyses of how cardiac gene regulatory networks correlate with early heart tissue morphogenesis in birds and mammals.     

Patterning of the heart field in the chick

In human development, it is postulated based on histological sections, that the cardiogenic mesoderm rotates 180° with the pericardial cavity. This is also thought to be the case in mouse development where gene expression data suggests that the progenitors of the right ventricle and outflow tract invert their position with respect to the progenitors of the atria and left ventricle. However, the inversion in both cases is inferred and has never been shown directly. We have used 3D reconstructions and cell tracing in chick embryos to show that the cardiogenic mesoderm is organized such that the lateralmost cells are incorporated into the cardiac inflow (atria and left ventricle) while medially placed cells are incorporated into the cardiac outflow (right ventricle and outflow tract). This happens because the cardiogenic mesoderm is inverted. The inversion is concomitant with movement of the anterior intestinal portal which rolls caudally to form the foregut pocket. The bilateral cranial cardiogenic fields fold medially and ventrally and fuse. After heart looping the seam made by ventral fusion will become the greater curvature of the heart loop. The caudal border of the cardiogenic mesoderm which ends up dorsally coincides with the inner curvature. Physical ablation of selected areas of the cardiogenic mesoderm based on this new fate map confirmed these results and, in addition, showed that the right and left atria arise from the right and left heart fields. The inversion and the new fate map account for several unexplained observations and provide a unified concept of heart fields and heart tube formation for avians and mammals.     

心脏的起源和细胞谱系

心脏是器官发生过程中最早形成的结构之一.心脏的原基分布图和心脏细胞谱系是心脏起源和形态发生研究的主要方面.总结了近几年来以鸡和小鼠为模型,在心脏发生和心脏前体细胞定位方面的研究情况.     

Light receptor action is critical for maintaining plant biomass at warm ambient temperatures

The ability to withstand environmental temperature variation is essential for plant survival. Former studies in Arabidopsis revealed that light signalling pathways had a potentially unique role in shielding plant growth and development from seasonal and daily fluctuations in temperature. In this paper we describe the molecular circuitry through which the light receptors cry1 and phyB buffer the impact of warm ambient temperatures. We show that the light signalling component HFR1 acts to minimise the potentially devastating effects of elevated temperature on plant physiology. Light is known to stabilise levels of HFR1 protein by suppressing proteasome-mediated destruction of HFR1. We demonstrate that light-dependent accumulation and activity of HFR1 are highly temperature dependent. The increased potency of HFR1 at warmer temperatures provides an important restraint on PIF4 that drives elongation growth. We show that warm ambient temperatures promote the accumulation of phosphorylated PIF4. However, repression of PIF4 activity by phyB and cry1 (via HFR1) is critical for controlling growth and maintaining physiology as temperatures rise. Loss of this light-mediated restraint has severe consequences for adult plants which have greatly reduced biomass.     

Diversification of Cardiomyogenic Cell Lineages in Vitro

The ability of undifferentiated cardiogenic mesoderm to generate diversified myogenic phenotypes was assayed in a minimal culture system. During cardiogenesis in vivo, the anterior and posterior segments of the avian heart have distinct patterns of contractile protein gene expression when they first differentiate. To assess the potential of undifferentiated cardiogenic tissue to diversify into distinct anterior and posterior lineages prior to heart formation, cardiogenic mesoderm and endoderm were removed together from the embryo at Hamburger and Hamilton stages 4-8. Explants from each of these stages differentiated in defined medium as indicated by the expression of muscle-specific genes. However, the ability to express the atrial-specific myosin heavy chain (AMHC1) mRNA was confined to posterior cardiac progenitors. Diversification was not dependent on anterior endoderm, suggesting that inductive interactions between the mesoderm and endoderm are not necessary to maintain diversified cardiac lineages after stage 4. The diversified potential of explanted cardiogenic tissue was altered with retinoic acid treatment, resulting in the activation of AMHC1 gene expression in the anterior progenitors. Anterior cardiogenic cells removed from the embryo at stage 8, when the heart begins to differentiate in vivo, are not susceptible to the alteration of diversified phenotype by retinoic acid treatment. Therefore, the potential to form distinct cardiomyogenic cell lineages is present in the anterior lateral plate mesoderm soon after gastrulation and the maturation of these lineages in a positionally dependent manner is maintained in a simple defined culture system in vitro.     

The convergence of Notch and MAPK signaling specifies the blood progenitor fate in the Drosophila mesoderm

Blood progenitors arise from a pool of pluripotential cells (“hemangioblasts”) within the Drosophila embryonic mesoderm. The fact that the cardiogenic mesoderm consists of only a small number of highly stereotypically patterned cells that can be queried individually regarding their gene expression in normal and mutant embryos is one of the significant advantages that Drosophila offers to dissect the mechanism specifying the fate of these cells. We show in this paper that the expression of the Notch ligand Delta (Dl) reveals segmentally reiterated mesodermal clusters (“cardiogenic clusters”) that constitute the cardiogenic mesoderm. These clusters give rise to cardioblasts, blood progenitors and nephrocytes. Cardioblasts emerging from the cardiogenic clusters accumulate high levels of Dl, which is required to prevent more cells from adopting the cardioblast fate. In embryos lacking Dl function, all cells of the cardiogenic clusters become cardioblasts, and blood progenitors are lacking. Concomitant activation of the Mitogen Activated Protein Kinase (MAPK) pathway by Epidermal Growth Factor Receptor (EGFR) and Fibroblast Growth Factor Receptor (FGFR) is required for the specification and maintenance of the cardiogenic mesoderm; in addition, the spatially restricted localization of some of the FGFR ligands may be instrumental in controlling the spatial restriction of the Dl ligand to presumptive cardioblasts.     

Development of cardiac beat rate in early chick embryos is regulated by regional cues

The mesoderm of each of the paired lateral heart-forming regions (HFRs) in the stage 5-7 chick embryo includes prospective conus (pre-C), ventricle (pre-V), and sinoatrial (pre-SA) cells, arranged in a rostrocaudal sequence (C-V-SA). With microsurgery we divided each HFR into three rostrocaudally arranged segments. After 24 hr of further incubation, each segment differentiated into a spontaneously beating vesicle of heart tissue to form a multiheart embryo. The cardiac vesicles in these embryos expressed left-right and rostrocaudal beat rate gradients: the left caudal pre-SA mesoderm produced tissue with the fastest beat rate of the six while the rostral vesicle formed from right pre-C was the slowest. In another operation, we prevented the HFRs from fusing in the midline by cutting through the anterior intestinal portal at stage 8, to produce cardia bifida (CB) embryos with an independently beating half-heart on each side. In these cases, the left half-heart of 87.2% of CB embryos beat faster than the right, confirming the left-right difference in intrinsic beat rate. To assess whether the future beat rate of each region is already determined in the st 5-7 HFR, we exchanged rectangular fragments of left pre-SA mesoderm and attached endoderm with right pre-C fragments to yield a left HFR with the sequence C-V-C and a right HFR with the sequence SA-V-SA. A CB operation was subsequently performed on these exchange embryos to prevent fusion of the lateral HFRs. Preconus mesoderm, transplanted to the pre-SA region, differentiated into tissue with a rapid beat rate, while pre-SA mesoderm relocated to the preconus region formed heart tissue with a slow spontaneous rate typical of the conus. In 73% of the exchange CB embryos, the left half-heart beat faster than the right, despite the origins of its mesoderm. The exchanged mesoderm developed a rate that was appropriate for its new location rather than the site of origin of the mesodermal fragment. In a third set of operations, we implanted a fragment of st 15 differentiated conus tissue into a site lateral to the left caudal HFR in st 5, 6, and 7 embryos, and subsequently performed CB operations on them. The implant caused the adjacent half-heart to develop with a slower beat rate than in unoperated or sham-operated controls.(ABSTRACT TRUNCATED AT 400 WORDS)     

The mechanisms of the formation of cardiogenic afferent effects after the development of local immune damage to the heart]

Sensitivity of cardiac receptors to several substances after local immune heart damage and the nature of cardiogenic influences on the circulation were studied in acute experiments o anesthetized dogs. The depressor reflexes from the heart were shown to disappear during 30 min. after immune heart damage, and vagal afferent impulse activity decreased. After immune heart damage, cardiac sympathetic afferent fibres were more sensitive to endogenous biological substances than to vagal ones. The sympathetic cardiac afferent system is found to be more sensitive to chemical agents, which is a decisive factor in formation of cardiogenic influences on the circulation during pathological processes in the heart.     

Evidence for a role of Smad6 in chick cardiac development.

Bone morphogenetic proteins (BMPs), members of the transforming growth factor-beta (TGF-beta) superfamily, are obligatory growth factors for early embryogenesis and heart formation. SMAD proteins transduce signals of the TGF-beta superfamily. We isolated chicken Smad6 (cSmad6), a member of inhibitory SMADs, and found its expression to be remarkably restricted to the developing heart, eyes, and limbs. cSmad6 expression was detected in the cardiogenic region of stage 5 embryos and overlapped Nkx2-5 and bmp-2, -4, and -7 expression. Throughout development, cSmad6 was expressed strongly in the heart, primarily in the myocardium, endocardium, and endocardial cushion tissue. Myocardial expression of cSmad6 was stronger in the forming septum, where highly localized expression of bmp-2 and -4 was also observed. Ectopically applied BMP-2 protein induced the expression of cSmad6, a putative negative regulator of BMP-signaling pathway, in anterior medial mesoendoderm of stage 4-5 embryos. In addition, blocking of BMP signaling using Noggin downregulated cSmad6 in cardiogenic tissue. cSmad1, one of the positive mediators of BMP signaling, was also expressed in cardiogenic region, but was not BMP-2 inducible. Our data suggest that cSmad6 has a role in orchestrating BMP-mediated cardiac development. We propose the possible mechanism of action of cSmad6 as modulating BMP signal by keeping a balance between constitutively expressed pathway-specific cSmad1 and ligand-induced inhibitory cSmad6 in the developing heart.     

Reassessment of Isl1 and Nkx2-5 cardiac fate maps using a Gata4-based reporter of Cre activity

Isl1 and Nkx2-5-expressing cardiovascular progenitors play pivotal roles in cardiogenesis. Previously reported Cre-based fate-mapping studies showed that Isl1 progenitors contribute predominantly to the derivatives of the second heart field, and Nkx2-5 progenitors contributed mainly to the cardiomyocyte lineage. However, partial recombination of Cre reporter genes can complicate interpretation of Cre fate-mapping experiments. We found that a Gata4-based Cre-activated reporter was recombined by Isl1^Cre and Nkx2-5^Cre in a substantially broader domain than previously reported using standard Cre-activated reporters. The expanded Isl1 and Nkx2-5 cardiac fate maps were remarkably similar, and included extensive contributions to cardiomyocyte, endocardial, and smooth muscle lineages in all four cardiac chambers. These data indicate that Isl1 is expressed in progenitors of both primary and secondary heart fields, and that Nkx2-5 is expressed in progenitors of cardiac endothelium and smooth muscle, in addition to cardiomyocytes. These results have important implications for our understanding of cardiac lineage diversification in vivo, and for the interpretation of Cre-based fate maps.     

Contribution of cardiogenic oscillations to gas exchange in constant-flow ventilation

The contribution of cardiogenic oscillations to gas exchange during constant-flow ventilation was examined in 11 dogs. With the use of two variations of cardiopulmonary bypass to maintain the systemic and pulmonary circulation, the influence of cardiogenic oscillations was removed by arresting the heart. Cardiac arrest by ventricular fibrillation was associated with a mean decrease in alveolar ventilation of 43% in five dogs on right and left heart bypass. However, successful defibrillation and return of the prearrest level of alveolar ventilation could not be achieved; thus we studied six dogs on left heart bypass. Alveolar ventilation decreased an average of 37% with cardiac arrest, and defibrillation resulted in a return of alveolar ventilation to 81% of the prearrest value. These results are consistent with previous predictions that cardiogenic oscillations are an important mechanism of gas transport during constant-flow ventilation.