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Adult mammalian cardiomyocyte regeneration after injury is thought to be minimal. Mononuclear diploid cardiomyocytes (MNDCMs), a relatively small subpopulation in the adult heart, may account for the observed degree of regeneration, but this has not been tested. We surveyed 120 inbred mouse strains and found that the frequency of adult mononuclear cardiomyocytes was surprisingly variable (>7-fold). Cardiomyocyte proliferation and heart functional recovery after coronary artery ligation both correlated with pre-injury MNDCM content. Using genome-wide association, we identified Tnni3k as one gene that influences variation in this composition and demonstrated that Tnni3k knockout resulted in elevated MNDCM content and increased cardiomyocyte proliferation after injury. Reciprocally, overexpression of Tnni3k in zebrafish promoted cardiomyocyte polyploidization and compromised heart regeneration. Our results corroborate the relevance of MNDCMs in heart regeneration. Moreover, they imply that intrinsic heart regeneration is not limited nor uniform in all individuals, but rather is a variable trait influenced by multiple genes.

Mammalian cardiomyocytes have limited regenerative ability. Cardiac disease, such as congenital heart disease and myocardial infarction, causes an initial loss of cardiomyocytes through regulated cell death (RCD). Understanding the mechanisms that govern RCD in the injured myocardium is crucial for developing therapeutics to promote heart regeneration. We previously reported that ferroptosis, a non-apoptotic and iron-dependent form of RCD, is the main contributor to cardiomyocyte death in the injured heart. To investigate the mechanisms underlying the preference for ferroptosis in cardiomyocytes, we examined the effects of anti-ferroptotic reagents in infarcted mouse hearts. The results revealed that the anti-ferroptotic reagent did not improve neonatal heart regeneration, and further compromised the cardiac function of juvenile hearts. On the other hand, ferroptotic cardiomyocytes played a supportive role during wound healing by releasing pro-angiogenic factors. The inhibition of ferroptosis in the regenerating mouse heart altered the immune and angiogenic responses. Our study provides insights into the preference for ferroptosis over other types of RCD in stressed cardiomyocytes, and guidance for designing anti-cell-death therapies for treating heart disease. [ABSTRACT FROM AUTHOR]

During development, sympathetic neurons extend axons along a myriad of distinct trajectories, often consisting of arteries, to innervate one of a large variety of distinct final target tissues. Whether or [...]

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2007.05.037 Byline: Yi-Hui Chen, Mamoru Ishii, Jingjing Sun, Henry M. Sucov, Robert E. Maxson Keywords: Msx1, Msx2; Aorticopulmonary septation; Apoptosis; Cardiac neural crest; Conotruncus; Myocardium; Outflow tract malalignment; Proliferation; Secondary heart field Abstract: Msx1 and Msx2 are highly conserved, Nk-related homeodomain transcription factors that are essential for a variety of tissue-tissue interactions during vertebrate organogenesis. Here we show that combined deficiencies of Msx1 and Msx2 cause conotruncal anomalies associated with malalignment of the cardiac outflow tract (OFT). Msx1 and Msx2 play dual roles in outflow tract morphogenesis by both protecting secondary heart field (SHF) precursors against apoptosis and inhibiting excessive proliferation of cardiac neural crest, endothelial and myocardial cells in the conotruncal cushions. During incorporation of SHF precursors into the OFT myocardium, ectopic apoptosis in the Msx1-/-; Msx2-/- mutant SHF is associated with reduced expression of Hand1 and Hand2, which from work on Hand1 and Hand2 mutants may be functionally important in the inhibition of apoptosis in Msx1/2 mutants. Later during aorticopulmonary septation, excessive proliferation in the OFT cushion mesenchyme and myocardium of Msx1-/-; Msx2-/- mutants is associated with premature down-regulation of p27.sup.KIP1, an inhibitor of cyclin-dependent kinases. Diminished accretion of SHF precursors to the elongating OFT myocardium and excessive accumulation of mesenchymal cells in the conotruncal cushions may work together to perturb the rotation of the truncus arteriosus, leading to OFT malalignment defects including double-outlet right ventricle, overriding aorta and pulmonary stenosis. Author Affiliation: Department of Biochemistry and Molecular Biology, Norris Comprehensive Cancer Center and Hospital, University of Southern California Keck School of Medicine, 1441 Eastlake Avenue, Los Angeles, CA 90033, USA Article History: Received 20 December 2006; Revised 21 May 2007; Accepted 29 May 2007

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2005.11.008 Byline: Bibha Choudhary (a), Yoshihiro Ito (b), Takako Makita (a), Tomoyo Sasaki (b), Yang Chai (b), Henry M. Sucov (a) Keywords: Neural crest; TGF[beta]; Type II receptor; Persistent truncus arteriosus; Interrupted aortic arch; DiGeorge syndrome Abstract: Previous studies have demonstrated that TGF[beta] induces a smooth muscle fate in primary neural crest cells in culture. By crossing a conditional allele of the type II TGF[beta] receptor with the neural crest-specific Wnt1cre transgene, we have addressed the in vivo requirement for TGF[beta] signaling in smooth muscle specification and differentiation. We find that elimination of the TGF[beta] receptor does not alter neural crest cell specification to a smooth muscle fate in the cranial or cardiac domains, and that a smooth muscle fate is not realized by trunk neural crest cells in either control or mutant embryos. Instead, mutant embryos exhibit with complete penetrance two very specific and mechanistically distinct cardiovascular malformations -- persistent truncus arteriosus (PTA) and interrupted aortic arch (IAA-B). Pharyngeal organ defects such as those seen in models of DiGeorge syndrome were not observed, arguing against an early perturbation of the cardiac neural crest cell lineage. We infer that TGF[beta] is an essential morphogenic signal for the neural crest cell lineage in specific aspects of cardiovascular development, although one that is not required for smooth muscle differentiation. Author Affiliation: (a) Institute for Genetic Medicine, Keck School of Medicine, School of Dentistry, University of Southern California, 2250 Alcazar St., IGM240, Los Angeles, CA 90033, USA (b) Center for Craniofacial Molecular Biology, School of Dentistry, University of Southern California, Los Angeles, CA 90033, USA Article History: Received 14 July 2005; Revised 26 October 2005; Accepted 8 November 2005

The role of transforming growth factor [beta] in formation of neural crest cells in some aspects of cardiovascular development in mouse embryos is studied.

The cytokine erythropoietin (Epo) is an essential factor promoting the survival, proliferation, and differentiation of erythroid progenitor cells. Epo expression and the initial phase of definitive erythropoietic differentiation in the fetal liver (E9-E12) are compromised in mouse embryos lacking the retinoic acid receptor RXR[alpha]. Our previous work demonstrated that the Epo gene is a direct target of retinoic acid action, via a retinoic acid receptor binding site in the Epo gene enhancer. However, Epo expression and erythropoietic differentiation become normalized in RXR[alpha] mutants from E12. In this study, we have investigated the molecular mechanisms underlying the transition in Epo gene regulation from RXR[alpha]-dependence to RXR[alpha]-independence. We find that three independent regulatory components are required for high level Epo expression in the early fetal liver: ligand-activated retinoic acid receptors, the hypoxia-regulated factor HIF1, and GATA factors. By E11.5, the fetal liver is no longer hypoxic, and retinoic acid signaling is no longer active; Epo expression from E11.5 onward is enhancer-independent, and is driven instead by basal promoter elements that provide a sufficient level of expression to support further erythropoietic differentiation. Keywords: Erythropoiesis; Erythropoietin; Fetal liver; Retinoic acid; Hypoxia; HNF4

Erythropoietin (EPO) is an essential growth factor that regulates erythrocyte production in mammals. In this study, we demonstrate a novel role of EPO in regulating angingenesis in vivo. Epo and Epo receptor (EpoR) are expressed in the vasculature during embryogenesis. Deletion of Epo or EpoR leads to angiogenic defects starting at E 10.5, 2 days before ventricular hypoplasia and 3 days before the onset of the embryonic lethal phenotype. Overall, angioganesis was severely affected in the mutant embryos: vascular anomalies included decreased complexity of the vessel networks. However, de novo vasculogenesis remained intact, consistent with the differential expression of Epo and EpoR during the early stages of embryonic development. The aforementioned angiogenesis defect can be partially rescued by expressing human EPO during embryogenesis. Moreover, Ang-1 expression is regulated by EPO/EPOR under normoxic conditions. Taken together, oar results suggest important roles of EPO and EPOR in angiogenesis. Keywords: Erythropoietin; Angiogenesis; Gene targeting; VEGF; Angiopoietin

Neural crest cells are multipotential progenitors that contribute to various cell and tissue types during embryogenesis. Here, we have investigated the molecular and cellular mechanism by which the fate of neural crest cell is regulated during tooth development. Using a two-component genetic system for indelibly marking the progeny of neural crest cells, we provide in vivo evidence of a deficiency of CNC-derived dental mesenchyme in Msx1 null mutant mouse embryos. The deficiency of the CNC results from an elevated CDK inhibitor p[19.sup.INK4d] activity and the disruption of cell proliferation. Interestingly, in the absence of Msx1, the CNC-derived dental mesenchyme misdifferentiates and possesses properties consistent with a neuronal fate, possibly through a default mechanism. Attenuation of p[19.sup.INK4d] in Msx1 null mutant mandibular explants restores mitotic activity in the dental mesenchyme, demonstrating the functional significance of Msx1-mediated p[19.sup.INK4d] expression in regulating CNC cell proliferation during odontogenesis. Collectively, our results demonstrate that homeobox gene Msx1 regulates the fate of CNC cells by controlling the progression of the cell cycle. Genetic mutation of Msx1 may alternatively instruct the fate of these progenitor cells during craniofacial development. Keywords: Cranial neural crest (CNC) cells; Cell cycle; Msx1; p[19.sup.INK4d]; Tooth; Proliferation; Differentiation; Apoptosis

Mouse embryos lacking the retinoic acid receptor RXR[alpha] properly undergo the early steps of heart development, but then fail to initiate a proliferative expansion of cardiomyocytes that normally results in the formation of the compact zone of the ventricular chamber wall. [RXR[alpha].sup.-/-] embryos have a hypoplastic ventricular chamber and die in midgestation from cardiac insufficiency. In this study, we have investigated the underlying mechanistic basis of this phenotype. We find that interference with retinoic acid receptor function in the epicardium of transgenic embryos recapitulates the hypoplastic phenotype of RXR[alpha] deficient embryos. We further show that wild type primary epicardial cells, and an established epicardial Cell line (EMC cells), secrete trophic protein factors into conditioned media that stimulate thymidine incorporation in primary fetal cardiomyocytes, and thymidine incorporation, cell cycle progression, and induction of cyclin D1 and E activity in NIH3T3 cells. In contrast, primary epicardial cells derived from [RXR[alpha].sup.-/-] embryos and an EMC subline constitutively expressing a dominant negative receptor construct both fail to secrete activity into conditioned media. The production of trophic factors is induced by retinoic acid treatment and is inhibited by a retinoid receptor antagonist. Fetal atrial and ventricular myocytes both respond to epicardial-derived trophic signaling, although postnatal cardiomyocytes are nonresponsive. We therefore propose that the fetal epicardium, in response to retinoic acid and in a manner requiring the activity of RXR[alpha], secretes trophic factors which drive fetal cardiomyocyte proliferation and promote ventricular chamber morphogenesis.

During mammalian evolution, expansion of the cerebral hemispheres was accompanied by expansion of the frontal and parietal bones of the skull vault and deployment of the coronal (fronto-parietal) and sagittal (parietal-parietal) sutures as major growth centres. Using a transgenic mouse with a permanent neural crest cell lineage marker, Wnt1-Cre/R26R, we show that both sutures are formed at a neural crest-mesoderm interface: the frontal bones are neural crest-derived and the parietal bones mesodermal, with a tongue of neural crest between the two parietal bones. By detailed analysis of neural crest migration pathways using X-gal staining, and mesodermal tracing by DiI labelling, we show that the neural crest-mesodermal tissue juxtaposition that later forms the coronal suture is established at E9.5 as the caudal boundary of the frontonasal mesenchyme. As the cerebral hemispheres expand, they extend caudally, passing beneath the neural crest-mesodermal interface within the dermis, carrying with them a layer of neural crest cells that forms their meningeal covering. Exposure of embryos to retinoic acid at E10.0 reduces this meningeal neural crest and inhibits parietal ossification, suggesting that intramembranous ossification of this mesodermal bone requires interaction with neural crest-derived meninges, whereas ossification of the neural crest-derived frontal bone is autonomous. These observations provide new perspectives on skull evolution and on human genetic abnormalities of skull growth and ossification. c 2001 Elsevier Science Key Words: mouse development; neural crest; mesoderm; skull sutures; Wnt1; retinoic acid.

A developmental transition in definitive erythropoiesis has been studied with the finding that erythropoietin (Epo) expression is regulated sequentially by HNF4 and retinoic acid receptors. The Epo gene enhancer contains a DR2 sequence that represents a retinoic acid receptor-binding site and a retinoic acid receptor transcriptional response element. The transition may switch regulation of Epo expression from retinoic acid control to hypoxic control. The Epo gene is shown to be retinoic acid inducible in embryos.