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1.
PLoS One ; 15(6): e0233007, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32492036

RESUMO

BACKGROUND: In humans, stillbirth describes the death of a fetus before birth after 28 weeks gestation, and accounts for approximately 2.6 million deaths worldwide annually. In high-income countries, up to half of stillbirths have an unknown cause and are described as "unexplained stillbirths"; this lack of understanding impairs efforts to prevent stillbirth. There are also few animal models of stillbirth, but those that have been described usually have significant placental abnormalities. This study describes a novel mutant murine model of fetal death with atrial conduction block due to an ErbB2 missense mutation which is not associated with abnormal placental morphology. METHODS: Phenotypic characterisation and histological analysis of the mutant mouse model was conducted. The mRNA distribution of the early cardiomyocyte marker Nkx2-5 was assessed via in situ hybridisation. Cardiac structure was quantified and cellular morphology evaluated by electron microscopy. Immunostaining was employed to quantify placental structure and cell characteristics on matched heterozygous and homozygous mutant placental samples. RESULTS: There were no structural abnormalities observed in hearts of mutant embryos. Comparable Nkx2-5 expression was observed in hearts of mutants and controls, suggesting normal cardiac specification. Additionally, there was no significant difference in the weight, placenta dimensions, giant cell characteristics, labyrinth tissue composition, levels of apoptosis, proliferation or vascularisation between placentas of homozygous mutant mice and controls. CONCLUSION: Embryonic lethality in the ErbB2 homozygous mutant mouse cannot be attributed to placental pathology. As such, we conclude the ErbB2M802R mutant is a model of stillbirth with a non-placental cause of death. The mechanism of the atrial block resulting from ErbB2 mutation and its role in embryonic death is still unclear. Studying this mutant mouse model could identify candidate genes involved in stillbirth associated with structural or functional cardiac defects.

2.
J Invest Dermatol ; 139(5): 1171-1181.e6, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30684552

RESUMO

Cellular senescence can be broadly defined as a stable, but essentially irreversible, loss of proliferative capacity. Historically, senescence has been described as a negative outcome of advanced cellular age. It is now clear, however, that senescence represents a dynamic autonomous stress response, integral to long-term tumor suppression. Transient induction of a senescent phenotype has actually been suggested to promote regeneration in both liver and skin. Here, we explored the role of senescence in pathological aged and diabetic murine wound healing. Aged and diabetic wounds had greater numbers of senescent cells, and diabetic macrophages maintained altered retention of polarization and produced a CXCR2-enriched senescence-associated secretory phenotype (i.e., SASP). Of translational relevance, targeted expression of CXCR2 in primary human dermal fibroblasts led to paracrine induction of nuclear p21. Furthermore, a selective agonist to CXCR2 was able to reverse delayed healing in diabetic mice and accelerate ex vivo human skin wound healing. Collectively, these data suggest a hitherto unappreciated role for CXCR2 in mediating cellular senescence in pathological wound repair.


Assuntos
Envelhecimento/genética , Senescência Celular/genética , Receptores de Interleucina-8B/genética , Cicatrização/genética , Análise de Variância , Animais , Biópsia por Agulha , Células Cultivadas , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/terapia , Modelos Animais de Doenças , Fibroblastos/citologia , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Interleucina-8B/metabolismo , Valores de Referência , Úlcera Cutânea/genética , Úlcera Cutânea/patologia , Cicatrização/fisiologia , Ferimentos e Lesões/genética , Ferimentos e Lesões/patologia
3.
PLoS Genet ; 13(10): e1007068, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29084269

RESUMO

The coronary vasculature is an essential vessel network providing the blood supply to the heart. Disruptions in coronary blood flow contribute to cardiac disease, a major cause of premature death worldwide. The generation of treatments for cardiovascular disease will be aided by a deeper understanding of the developmental processes that underpin coronary vessel formation. From an ENU mutagenesis screen, we have isolated a mouse mutant displaying embryonic hydrocephalus and cardiac defects (EHC). Positional cloning and candidate gene analysis revealed that the EHC phenotype results from a point mutation in a splice donor site of the Myh10 gene, which encodes NMHC IIB. Complementation testing confirmed that the Myh10 mutation causes the EHC phenotype. Characterisation of the EHC cardiac defects revealed abnormalities in myocardial development, consistent with observations from previously generated NMHC IIB null mouse lines. Analysis of the EHC mutant hearts also identified defects in the formation of the coronary vasculature. We attribute the coronary vessel abnormalities to defective epicardial cell function, as the EHC epicardium displays an abnormal cell morphology, reduced capacity to undergo epithelial-mesenchymal transition (EMT), and impaired migration of epicardial-derived cells (EPDCs) into the myocardium. Our studies on the EHC mutant demonstrate a requirement for NMHC IIB in epicardial function and coronary vessel formation, highlighting the importance of this protein in cardiac development and ultimately, embryonic survival.


Assuntos
Vasos Coronários/crescimento & desenvolvimento , Desenvolvimento Embrionário/genética , Cadeias Pesadas de Miosina/genética , Miosina não Muscular Tipo IIB/genética , Pericárdio/crescimento & desenvolvimento , Animais , Diferenciação Celular/genética , Vasos Coronários/metabolismo , Embrião de Mamíferos , Transição Epitelial-Mesenquimal/genética , Humanos , Hidrocefalia/genética , Hidrocefalia/metabolismo , Hidrocefalia/patologia , Camundongos , Camundongos Knockout , Mutação , Miocárdio/metabolismo , Pericárdio/metabolismo
4.
PLoS One ; 9(9): e107041, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25269082

RESUMO

The heart is the first organ required to function during embryonic development and is absolutely necessary for embryo survival. Cardiac activity is dependent on both the sinoatrial node (SAN), which is the pacemaker of heart's electrical activity, and the cardiac conduction system which transduces the electrical signal though the heart tissue, leading to heart muscle contractions. Defects in the development of cardiac electrical function may lead to severe heart disorders. The Erbb2 (Epidermal Growth Factor Receptor 2) gene encodes a member of the EGF receptor family of receptor tyrosine kinases. The Erbb2 receptor lacks ligand-binding activity but forms heterodimers with other EGF receptors, stabilising their ligand binding and enhancing kinase-mediated activation of downstream signalling pathways. Erbb2 is absolutely necessary in normal embryonic development and homozygous mouse knock-out Erbb2 embryos die at embryonic day (E)10.5 due to severe cardiac defects. We have isolated a mouse line, l11Jus8, from a random chemical mutagenesis screen, which carries a hypomorphic missense mutation in the Erbb2 gene. Homozygous mutant embryos exhibit embryonic lethality by E12.5-13. The l11Jus8 mutants display cardiac haemorrhage and a failure of atrial function due to defects in atrial electrical signal propagation, leading to an atrial-specific conduction block, which does not affect ventricular conduction. The l11Jus8 mutant phenotype is distinct from those reported for Erbb2 knockout mouse mutants. Thus, the l11Jus8 mouse reveals a novel function of Erbb2 during atrial conduction system development, which when disrupted causes death at mid-gestation.


Assuntos
Átrios do Coração/metabolismo , Cardiopatias Congênitas/genética , Receptor ErbB-2/genética , Potenciais de Ação , Animais , Função Atrial , Átrios do Coração/embriologia , Átrios do Coração/fisiopatologia , Sistema de Condução Cardíaco/embriologia , Sistema de Condução Cardíaco/fisiopatologia , Cardiopatias Congênitas/fisiopatologia , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação de Sentido Incorreto , Receptor ErbB-2/metabolismo
5.
Genesis ; 52(8): 713-37, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24866031

RESUMO

Genes required for an organism to develop to maturity (for which no other gene can compensate) are considered essential. The continuing functional annotation of the mouse genome has enabled the identification of many essential genes required for specific developmental processes including cardiac development. Patterns are now emerging regarding the functional nature of genes required at specific points throughout gestation. Essential genes required for development beyond cardiac progenitor cell migration and induction include a small and functionally homogenous group encoding transcription factors, ligands and receptors. Actions of core cardiogenic transcription factors from the Gata, Nkx, Mef, Hand, and Tbx families trigger a marked expansion in the functional diversity of essential genes from midgestation onwards. As the embryo grows in size and complexity, genes required to maintain a functional heartbeat and to provide muscular strength and regulate blood flow are well represented. These essential genes regulate further specialization and polarization of cell types along with proliferative, migratory, adhesive, contractile, and structural processes. The identification of patterns regarding the functional nature of essential genes across numerous developmental systems may aid prediction of further essential genes and those important to development and/or progression of disease.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento/genética , Genes Essenciais/genética , Coração/crescimento & desenvolvimento , Mamíferos/genética , Animais , Diferenciação Celular , Movimento Celular , Coração/embriologia , Mamíferos/embriologia , Mamíferos/crescimento & desenvolvimento , Camundongos , Células-Tronco
6.
Cell Stress Chaperones ; 15(6): 835-49, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20428984

RESUMO

Multiple epiphyseal dysplasia (MED) can result from mutations in matrilin-3, a structural protein of the cartilage extracellular matrix. We have previously shown that in a mouse model of MED the tibia growth plates were normal at birth but developed a progressive dysplasia characterised by the intracellular retention of mutant matrilin-3 and abnormal chondrocyte morphology. By 3 weeks of age, mutant mice displayed a significant decrease in chondrocyte proliferation and dysregulated apoptosis. The aim of this current study was to identify the initial post-natal stages of the disease. We confirmed that the disease phenotype is seen in rib and xiphoid cartilage and, like tibia growth plate cartilage is characterised by the intracellular retention of mutant matrilin-3. Gene expression profiling showed a significant activation of classical unfolded protein response (UPR) genes in mutant chondrocytes at 5 days of age, which was still maintained by 21 days of age. Interestingly, we also noted the upregulation of arginine-rich, mutated in early stage of tumours (ARMET) and cysteine-rich with EGF-like domain protein 2 (CRELD2) are two genes that have only recently been implicated in the UPR. This endoplasmic reticulum (ER) stress and UPR did not lead to increased chondrocyte apoptosis in mutant cartilage by 5 days of age. In an attempt to alleviate ER stress, mutant mice were fed with a chemical chaperone, 4-sodium phenylbutyrate (SPB). SPB at the dosage used had no effect on chaperone expression at 5 days of age but modestly decreased levels of chaperone proteins at 3 weeks. However, this did not lead to increased secretion of mutant matrilin-3 and in the long term did not improve the disease phenotype. We performed similar studies with a mouse model of Schmid metaphyseal chondrodysplasia, but again this treatment did not improve the phenotype.


Assuntos
Proteínas da Matriz Extracelular/genética , Osteocondrodisplasias/metabolismo , Resposta a Proteínas não Dobradas , Animais , Apoptose , Cartilagem/patologia , Cartilagem/ultraestrutura , Condrócitos/metabolismo , Modelos Animais de Doenças , Retículo Endoplasmático/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Perfilação da Expressão Gênica , Proteínas Matrilinas , Camundongos , Mutação , Fatores de Crescimento Neural/genética , Fatores de Crescimento Neural/metabolismo , Osteocondrodisplasias/genética , Osteocondrodisplasias/patologia , Fenótipo , Fenilbutiratos/farmacologia , Costelas/citologia , Fatores de Tempo , Regulação para Cima
7.
Dev Biol ; 342(2): 146-56, 2010 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-20347762

RESUMO

During development, the mammalian embryo must integrate signals to control growth and proliferation. A failure in the ability to respond to mitogenic stimuli can cause embryonic growth restriction. We have identified a mouse mutant, l11Jus15, from a mutagenesis screen that exhibits growth defects and late-gestation lethality. Here we demonstrate that this phenotype results from a mutation in the Mediator complex gene Med31, which causes degradation of Med31 protein. The Med31 mutant phenotype is not similar to other Mediator complex mouse mutants, and target genes of other Mediator proteins are expressed normally in Med31 mutants, suggesting that Med31 has distinct target genes required for mammalian development. Med31 mutant embryos have fewer proliferating cells than controls, especially in regions that expand rapidly during development such as the forelimb buds. Likewise, embryonic fibroblast cells cultured from mutant embryos have a severe proliferation defect, as well as reduced levels of the cell cycle protein Cdc2. Med31 mutants have normal limb bud patterning but defective or delayed chondrogenesis due to a lack of Sox9 and Col2a1 expression. As the Mediator complex is a transcriptional co-activator, our results suggest that Med31 functions to promote the transcription of genes required for embryonic growth and cell proliferation.


Assuntos
Proliferação de Células , Extremidades/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Complexo Mediador/metabolismo , Animais , Linhagem Celular , Clonagem Molecular , Humanos , Complexo Mediador/genética , Camundongos , Mutação
8.
Hum Mutat ; 29(12): 1405-11, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-18561327

RESUMO

Hair morphology differs dramatically between human populations: people of East Asian ancestry typically have a coarse hair texture, with individual fibers being straight, of large diameter, and cylindrical when compared to hair of European or African origin. Ectodysplasin-A receptor (EDAR) is a cell surface receptor of the tumor necrosis factor receptor (TNFR) family involved in the development of hair follicles, teeth, and sweat glands. Analyses of genome-wide polymorphism data from multiple human populations suggest that EDAR experienced strong positive selection in East Asians. It is likely that a nonsynonymous SNP in EDAR, rs3827760, was the direct target of selection as the derived p.Val370Ala variant is seen at high frequencies in populations of East Asian and Native American origin but is essentially absent from European and African populations. Here we demonstrate that the derived EDAR370A common in East Asia has a more potent signaling output than the ancestral EDAR370 V in vitro. We show that elevation of Edar activity in transgenic mice converts their hair phenotype to the typical East Asian morphology. The coat texture becomes coarse, with straightening and thickening of individual hairs and conversion of fiber cross-sectional profile to a circular form. These thick hair fibers are produced by enlarged hair follicles, which in turn develop from enlarged embryonic organ primordia. This work shows that the multiple differences in hair form between East Asian and other human populations can be explained by the simplest of genetic alterations.


Assuntos
Grupo com Ancestrais do Continente Asiático/genética , Receptor Edar/genética , Receptor Edar/metabolismo , Cabelo/química , Polimorfismo de Nucleotídeo Único , Sequência de Aminoácidos , Substituição de Aminoácidos , Animais , Linhagem Celular , Receptor Edar/química , Cabelo/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Alinhamento de Sequência
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