Biomedical subjects

F Costantini

Publications and source records attributed to F Costantini.

At least 73 records · Page 4Linked to original sources

PubMed1995

Isolation and characterization of a chicken homolog of the c-ret proto-oncogene.

The c-ret proto-oncogene encodes a receptor tyrosine kinase that plays important roles in human disease and in normal mammalian development. Mutations in the human RET gene are associated with multiple endocrine neoplasia syndromes and Hirschsprung's disease in humans, while targeted mutagenesis of murine c-ret resulted in severe developmental abnormalities affecting the excretory and peripheral nervous systems. To examine the evolutionary conservation of the ret protein sequence and its developmental expression pattern, we isolated and sequenced cDNA clones of chicken c-ret and examined its expression in chick embryos and adult tissues. The cytoplasmic domains of chicken and human ret were relatively well conserved (91% similar), but the extracellular domains were more divergent (68% similar), although the conservation of cysteine residues in this region suggests a conserved secondary structure. As in mouse and human, chicken c-ret encodes two protein isoforms. The number and sizes of the transcripts were similar to those in human and mouse cells, and during chick embryogenesis, c-ret mRNA was observed in many of the same sites as in the mouse, including the Wolffian duct and ureteric bud, the enteric, dorsal root, sympathetic and facioacoustic ganglia, and the ventral spinal cord. Evolutionary differences in expression were observed in the trigeminal ganglion, the ventral roots of the spinal cord, the mesenchymal cells of the branchial arches and the adult testes. The results are discussed with regard to the role of the ret receptor in normal development and disease.

Amino Acid Sequence↗

PubMed1995

Vitamins E, C and lipid peroxidation in plasma and arterial tissue of smokers and non-smokers.

An imbalance between pro-oxidants and antioxidants is operative in atherosclerosis. Cigarette smoke is a major risk factor of atherosclerosis and has been reported to contain large amounts of oxidants. We assessed arterial (internal mammary artery) and plasma levels of vitamins E and C and lipid peroxides in 48 male patients, 24 smokers and 24 non-smokers, undergoing coronary bypass surgery. Lipid peroxidation was studied using fluorescent products of lipid peroxidation (FPLs). Tissue vitamins E and C levels were significantly lower and FPLs significantly higher in smokers than in non-smokers (P < 0.0006, 0.0005 and 0.0005, respectively). This pattern was associated with lower vitamin C and higher lipid peroxide plasma levels in smokers than in non-smokers (P < 0.0002 and 0.0005, respectively). Vitamins E and C plasma levels were strongly related to their tissue content both in smokers (r = 0.60, P < 0.005 and r = 0.57, P < 0.01) and in non-smokers (r = 0.42, P < 0.05 and r = 0.46, P < 0.05). Moreover, vitamin E content was significantly related to that of vitamin C only in the arterial tissue of both groups, pointing to the existence of a functional interaction between these antioxidants. In both groups, FPLs were significantly and inversely related to vitamin C in plasma and to vitamin E in tissue, suggesting the antioxidant primary of vitamin C and vitamin E in the plasma and arterial tissue compartments, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Arteries↗

PubMed1995

Phenotypic and molecular analysis of a transgenic insertional allele of the mouse Fused locus.

Spontaneous mutations at the mouse Fused (Fu) locus cause dominant skeletal and neurological defects and recessive lethal embryonic defects including neuroectodermal abnormalities and axial duplications. Here, we describe a new allele at the Fu locus caused by a transgenic insertional mutation, H epsilon 46. Embryos homozygous for the H epsilon 46 insertion die at day 9-10 post coitum and display phenotypic defects similar to those associated with Fu alleles. The H epsilon 46 locus was cloned and shown to contain a 20-kb deletion at the site of transgene insertion with no other detectable rearrangements. Genomic probes from the H epsilon 46 locus were mapped to a genetic locus closely linked to Fu on chromosome 17 and were hybridized to a YAC contig covering the FuKi critical region. Compound heterozygotes between H epsilon 46 and FuKi were inviable and displayed abnormalities at the same stage of embryogenesis as do homozygotes for either of the two mutations, demonstrating that these two recessive lethal mutations belong to the same complementation group. A genomic probe from the wild-type H epsilon 46 locus detected a transcript that is disrupted by the transgenic insertion, representing a candidate for the wild-type allele of Fused.

Alleles↗

PubMed1995

'White-coat' hypertension in patients with newly diagnosed hypertension: evaluation of prevalence by ambulatory monitoring and impact on cost of health care.

To evaluate the prevalence of 'white-coat' hypertension in patients with newly diagnosed hypertension, 255 subjects (131 males and 124 females) underwent 24-h ambulatory blood pressure monitoring. Patients with 24-h systolic and diastolic blood pressure < 135/85 mmHg were classified as white-coat hypertensives and the remaining as sustained hypertensives. On the assumption that white-coat hypertensives may not need to take antihypertensive medication, we evaluated the impact on cost of health care of two strategies based essentially on treating all patients according to casual blood pressure, or ambulatory blood pressure monitoring, followed by drug treatment in sustained hypertensives only. Of the 255 hypertensives studied, 54 (21%), confidence interval 16%, 26%, were classified as white-coat hypertensives. The age, sex-ratio and body mass index did not differ between the white-coat and the sustained hypertensive subjects. The strategy of monitoring all patients and of treating only the sustained hypertensives resulted in a substantial coat saving, which was calculated to be about 110,000 U.S.A. dollars over a period of 6 years. In conclusion, white-coat hypertensives are frequent among patients with newly diagnosed hypertension, and they do not differ from sustained hypertensives as regards demographic data. Ambulatory blood pressure monitoring, when used to decide whether or not to treat pharmacologically, increases the cost-effectiveness of treatment for hypertension and reduces the cost of health care.

Adult↗

PubMed1995

RET-deficient mice: an animal model for Hirschsprung's disease and renal agenesis.

Receptor tyrosine kinases play a critical role in transducing signals involved in cell growth and differentiation. The c-ret proto-oncogene is a member of the receptor tyrosine kinase gene superfamily originally identified by its transforming ability. Somatic mutations of c-ret are responsible for a large proportion of thyroid papillary carcinomas, while germ-line mutations are responsible for multiple endocrine neoplasia types 2A and 2B, dominantly inherited cancer syndromes characterized by multiple tumours of neuroectodermal origin. In addition to its role in tumour formation. c-ret is thought to have a developmental role since mutations of the gene have been implicated in the aetiology of Hirschsprung's syndrome (congenital megacolon). A targeted mutation in the murine c-ret locus shows that the ret receptor is required for normal development of two lineally unrelated systems, the excretory system and the enteric nervous system.

Animals↗

PubMed1995

In vivo demonstration of red cell-endothelial interaction, sickling and altered microvascular response to oxygen in the sickle transgenic mouse.

Intravascular sickling, red cell-endothelium interaction, and altered microvascular responses have been suggested to contribute to the pathophysiology of human sickle cell disease, but have never been demonstrated under in vivo flow. To address this issue, we have examined a transgenic mouse line, alphaHbetaSbetaS-Antilles [betaMDD] which has a combined high (78%) expression of beta S and beta S-Antilles globins. In vivo microcirculatory studies using the cremaster muscle preparation showed adhesion of red cells, restricted to postcapillary venules, in transgenic mice but not in control mice. Electron microscopy revealed distinct contacts between the red cell membrane and the endothelium surface. Some red cells exhibiting sickling were regularly observed in the venular flow. Infusion of transgenic mouse red cells into the ex vivo mesocecum vasculature also showed adhesion of mouse red cells exclusively in venules. Under resting conditions (pO2, 15-20 mmHg), there were no differences in the cremaster microvascular diameters of control and transgenic mice; however, transgenic mice showed a drastic reduction in microvascular red cell velocities (Vrbc) with maximal Vrbc decrease (> 60%) occurring in venules, the sites of red cell adhesion and sickling. Local, transient hyperoxia (pO2, 150 mmHg) resulted in striking differences between control and transgenic mice. In controls, oxygen caused a 69% arteriolar constriction, accompanied by 75% reduction in Vrbc. In contrast, in transgenic mice, hyperoxia resulted in only 8% decrease in the arteriolar diameter and in 68% increase in VrBC; the latter is probably due to an improved flow behavior of red cells as a consequence of unsickling. In summary, the high expression of human sickle hemoglobin in the mouse results not only in intravascular sickling but also red cell-endothelium interaction. The altered microvascular response to oxygen could be secondary to blood rheological changes, although possible intrinsic differences in the endothelial cell/vascular smooth muscle function in the transgenic mouse may also contribute. These sickle transgenic mice could serve as a useful model to investigate vasoocclusive mechanisms, as well as to test potential therapies.

Anemia, Sickle Cell↗

PubMed1995

Development of hematopoietic cells lacking transcription factor GATA-1.

GATA-1 is a zinc-finger transcription factor believed to play an important role in gene regulation during the development of erythroid cells, megakaryocytes and mast cells. Other members of the GATA family, which can bind to the same DNA sequence motif, are co-expressed in several of these hemopoietic lineages, raising the possibility of overlap in function. To examine the specific roles of GATA-1 in hematopoietic cell differentiation, we have tested the ability of embryonic stem cells, carrying a targeted mutation in the X-linked GATA-1 gene, to contribute to various blood cell types when used to produce chimeric embryos or mice. Previously, we reported that GATA-1- mutant cells failed to contribute to the mature red blood cell population, indicating a requirement for this factor at some point in the erythroid lineage (L. Pevny et al., (1991) Nature 349, 257-260). In this study, we have used in vitro colony assays to identify the stage at which mutant erythroid cells are affected, and to examine the requirement for GATA-1 in other lineages. We found that the development of erythroid progenitors in embryonic yolk sacs was unaffected by the mutation, but that the cells failed to mature beyond the proerythroblast stage, an early point in terminal differentiation. GATA-1- colonies contained phenotypically normal macrophages, neutrophils and megakaryocytes, indicating that GATA-1 is not required for the in vitro differentiation of cells in these lineages. GATA-1- megakaryocytes were abnormally abundant in chimeric fetal livers, suggesting an alteration in the kinetics of their formation or turnover. The lack of a block in terminal megakaryocyte differentiation was shown by the in vivo production of platelets expressing the ES cell-derived GPI-1C isozyme. The role of GATA-1 in mast cell differentiation was examined by the isolation of clonal mast cell cultures from chimeric fetal livers. Mutant and wild-type mast cells displayed similar growth and histochemical staining properties after culture under conditions that promote the differentiation of cells resembling mucosal or serosal mast cells. Thus, the mast and megakaryocyte lineages, in which GATA-1 and GATA-2 are co-expressed, can complete their maturation in the absence of GATA-1, while erythroid cells, in which GATA-1 is the predominant GATA factor, are blocked at a relatively early stage of maturation.

Animals↗

PubMed1994

Interleukin-6 deficient mice are protected from bone loss caused by estrogen depletion.

Interleukin-6 (IL-6) is a multifunctional cytokine whose circulating levels are under physiological conditions below detection, but whose production is rapidly and strongly induced by several pathological and inflammatory stimuli. IL-6 has been implicated in a number of cell functions connected to immunity and hematopoiesis. Recently, it has been proposed to act as a stimulator of osteoclast formation and activity, in particular following estrogen depletion. The purpose of this study was to gain additional insights into the role of IL-6 during development, as well as in physiological and pathological conditions. We report here that IL-6 deficient mice generated by gene targeting are viable and do not present any evident phenotypic abnormality. However, analysis of bone metabolism revealed a specific bone phenotype. IL-6 deficient female mice have a normal amount of trabecular bone, but higher rates of bone turnover than control littermates. Estrogen deficiency induced by ovariectomy causes in wild type animals a significant loss of bone mass together with an increase in bone turnover rates. Strikingly, ovariectomy does not induce any change in either bone mass or bone remodeling rates in the IL-6 deficient mice. These findings indicate that IL-6 plays an important role in the local regulation of bone turnover and, at least in mice, appears to be essential for the bone loss caused by estrogen deficiency.

Animals↗

PubMed1994

Defects in the kidney and enteric nervous system of mice lacking the tyrosine kinase receptor Ret.

Receptor tyrosine kinases (RTKs) are cell-surface molecules that transduce signals for cell growth and differentiation. The RTK encoded by the c-ret proto-oncogene is rearranged and constitutively activated in a large proportion of thyroid papillary carcinomas, and germ-line point mutations in c-ret seem to be responsible for the dominantly inherited cancer syndromes multiple endocrine neoplasia (MEN) types 2A and B. The gene is expressed in the developing central and peripheral nervous systems (sensory, autonomic and enteric ganglia) and the excretory system (Wolffian duct and ureteric bud epithelium) of mice, indicating that it may play a role in normal development. Here we show that mice homozygous for a targeted mutation in c-ret develop to term, but die soon after birth, showing renal agenesis or severe dysgenesis, and lacking enteric neurons throughout the digestive tract. Ret is thus an essential component of a signalling pathway required for renal organogenesis and enteric neurogenesis.

Animals↗

PubMed1994

Analysis of the developmental and transcriptional potentiation functions of 5'HS2 of the murine beta-globin locus control region in transgenic mice.

We analyze the role of 5'HS2 of the mouse beta-globin LCR in the transcriptional and developmental regulation of beta-globin gene expression. Previous studies have shown that the human beta-globin gene behaves as an adult gene in transgenic mice, being expressed in fetal liver and bone marrow-derived erythroblasts but not in yolk sac-derived embryonic erythroid cells. We show that linkage of mLCR5'HS2 to a human beta-globin gene alters this pattern of expression during ontogeny, resulting in expression of the linked beta-globin gene at all stages of murine erythroid development. Expression was independent of integration position and correlated with transgene copy number. Our results provide the first test of a phylogenetically homologous LCR in transgenic mice and demonstrate evolutionary conservation of both developmental and transcriptional potentiation functions between mammalian beta-globin LCRs.

Animals↗

PubMed1994

Retinal and choroidal neovascularization in a transgenic mouse model of sickle cell disease.

A complication of sickle cell disease is proliferative retinopathy. We investigated the eyes from a transgenic mouse model of sickle cell disease (alpha H beta S[beta MDD] type) to determine if pathological changes occurred in their retinas and choroids. One retina from each animal was processed by flat-embedding adenosine diphosphatase-reacted retinas in glycol methacrylate. The fellow eye from each animal was embedded whole in glycol methacrylate for histopathological analysis of all ocular structures. Retinal vascular occlusions resulted in nonperfused areas of retina and arterio-venous anastomoses. Intra- and extraretinal neovascularization was observed adjacent to nonperfused areas. Retinal pigmented lesions were formed by the migration of retinal pigment epithelial cells into sensory retina, often ensheathing choroidal neovascularization. The incidence of this bilateral chorioretinopathy was 30% in animals older than 15 months of age. The ocular histopathological changes we observed in the mouse model mimicked many aspects of human proliferative sickle cell retinopathy. Furthermore, this is the first genetically derived animal model for chorio-retinal neovascularization.

Anemia, Sickle Cell↗

PubMed1993

Long-term high-level expression of human beta-globin occurs following transplantation of transgenic marrow into irradiated mice.

When the human beta-globin gene is transferred into the bone marrow cells of live mice, its expression is very low. To investigate the reason for this, we transferred the bone marrow of transgenic mice containing and expressing the human beta-globin into irradiated recipients. We demonstrate that long-term high level expression of the human beta-globin gene can be maintained in the marrow and blood of irradiated recipients following transplantation. Although expression decreased over time in most animals because of host marrow reconstitution, the ratio of human beta-globin transgene expression to endogenous mouse beta-globin gene expression in donor-derived erythroid cells remained constant over time. We conclude that there is no inherent limitation to efficient expression of an exogenous human beta-globin gene in mouse bone marrow cells following marrow transplantation.

Animals↗

PubMed1993

Morula decompaction (mdn), a preimplantation recessive lethal defect in a transgenic mouse line.

The beta S12 transgenic line carries an insertion of exogenous DNA and a deletion of approximately 2 cM on chromosome 1. Transgenic heterozygotes (beta S12/+) were viable and fertile, but homozygous mice (beta S12/beta S12) could not be produced. To determine the stage at which developmental arrest occurred in homozygotes, embryos derived from intercrosses among heterozygotes or from control crosses were examined at various stages. No homozygotes could be detected at postimplantation stages, suggesting that the mutation affected embryogenesis before implantation. Recovery of embryos at 1.5 or 3.5 days postcoitum, followed by culture in vitro, revealed an abnormal class of embryos which constituted one-fourth of the progeny from intercrosses, and thus appeared to represent the beta S12/beta S12 homozygotes. These embryos cleaved normally to the 8-cell stage and formed compacted morulae, but then decompacted without forming a blastocyst, and ceased dividing at approximately the 16-cell stage. Nearly all blastomeres were viable at the time of decompaction, as demonstrated by trypan blue exclusion, indicating that the loss of compaction is not simply a consequence of cell death. When labeled with a short-term lineage marker and aggregated with normal embryos at the early 8-cell stage, homozygous mutant embryos failed to contribute to the resulting blastocysts, indicating that the defect in the maintenance of compaction is cell autonomous. Based on the mutant phenotype, we conclude that this genetic lesion affects a gene (or genes) required for the maintenance of compaction, and propose that it be named morula decompaction (mdn). The phenotype of mdn/mdn embryos appears to be distinct from those caused by other previously described preimplantation lethal mutations or chromosomal deletions.

Animals↗

PubMed1993

Expression of the c-ret proto-oncogene during mouse embryogenesis.

The c-ret proto-oncogene encodes a receptor tyrosine kinase whose normal function has yet to be determined. To begin to investigate the potential role of this gene in vertebrate development, we have isolated cDNA clones representing the murine c-ret gene, and have analyzed the pattern of expression during mouse embryogenesis, using northern blotting, in situ hybridization to histological sections and whole-mount hybridization histochemistry. c-ret transcripts were detected beginning at day 8.5 of embryogenesis, and were observed in a number of cell lineages in the developing peripheral and central nervous systems, as well as in the excretory system. In the cranial region at day 8.5-9.5, c-ret mRNA was restricted to a population of neural crest cells migrating from rhombomere 4 and forming the anlage of the facioacoustic ganglion, as well as to a closely associated domain of surface ectoderm and pharyngeal endoderm. At later stages (10.5-14.5 days), c-ret mRNA was observed in all cranial ganglia. In the peripheral nervous system of the trunk, c-ret was expressed in the autonomic ganglia and in subsets of cells in the dorsal root ganglia. In the enteric nervous system, c-ret was expressed in the presumptive enteric neuroblasts of the vagal crest (day 9.0-11.5), and in the myenteric ganglia of the gut (day 13.5-14.5). c-ret mRNA was observed in several regions of the central nervous system, including the undifferentiated neuroepithelial cells of the ventral neural tube (8.5 days), the motor neurons in the spinal cord and the hindbrain (10.5-14.5 days), the embryonic neuroretina (day 13.5) and the layers of the postnatal retina containing ganglion, amacrine and horizontal cells. Outside the nervous system, c-ret was expressed in the nephric (Wolffian) duct at day 8.5-10.5, the ureteric bud epithelium (but not the surrounding metanephric mesenchyme) at day 11.0-11.5, and the growing tips of the renal collecting ducts (but not the previously formed, subcortical portions of the collecting ducts, or the mesenchyme-derived renal vesicles) at day 13.5-17.5. Our results suggest that the c-ret gene may encode the receptor for a factor involved in the proliferation, migration, differentiation or survival of a variety of neuronal cell lineages, as well as in inductive interactions during organogenesis of the kidney.

Animals↗

PubMed1992

High expression of human beta S- and alpha-globins in transgenic mice: hemoglobin composition and hematological consequences.

A line of transgenic mice (alpha H beta S-11; where alpha H is human alpha-globin) was created in which the human beta S and human alpha 2 globin genes, each linked to the beta-globin locus control region, were cointegrated into the mouse genome. On a normal genetic background, the transgenic mice produced 36% human beta S-globin chains with an alpha H/beta S ratio of 1.3. Higher levels of beta S were achieved by breeding the transgenic mice with mutant mice carrying a mouse beta major-globin gene deletion. Mice heterozygous for the beta major deletion (alpha H beta S[beta MD]; MD, mouse deletion) had 54% beta S with an alpha H/beta S ratio of 1.0; mice homozygous for the beta major deletion (alpha H beta S[beta MDD]) had 72.5% beta S and an alpha H/beta S ratio of 0.73. Because mouse alpha chains inhibit hemoglobin (Hb) S polymerization, we bred the mice to heterozygosity for a mouse alpha-globin deletion. These mice (alpha H beta S[alpha MD beta MDD]) had an increased alpha H/beta S ratio of 0.89 but expressed 65% beta S. Expression of the human genes cured the thalassemic phenotype associated with the murine beta major deletion. Transgenic alpha H beta S[beta MDD] mice had normal hematocrit and Hb and somewhat elevated reticulocytes (6% vs. 3% for control), whereas the mice carrying the alpha-globin deletion (alpha H beta S[alpha MD beta MDD]) had a normal hematocrit and Hb and more elevated reticulocytes (10.3 +/- 7.6% vs. 3.4 +/- 1.0%). Expression of the transgene restored a normal distribution of erythrocyte densities when compared to thalassemic mice; however, the average mean corpuscular Hb concentration of alpha H beta S[beta MDD] mice increased to 35.7 g/dl (vs. control 33.7 g/dl) whereas that of alpha H beta S[alpha MD beta MDD] mice was further elevated to 36.3 g/dl. The intrinsic oxygen affinity was increased in transgenic mouse erythrocytes at 280 milliosmolal, and the PO2 at midsaturation of alpha H beta S[alpha MD beta MDD] erythrocytes was higher than that of alpha H beta S[beta MDD] cells (37.4 +/- 2 vs. 33.5 +/- 1 mmHg). The higher values of the mean corpuscular Hb concentration and intrinsic PO2 at midsaturation, which favor in vivo sickling, may explain the slightly more severe hematological picture in alpha H beta S[alpha MD beta MDD] mice. We conclude that the transgenic mouse with high Hb S expression does not exhibit adult anemia but does have abnormal hematological features: increased erythrocyte density, high oxygen affinity, and reticulocytosis with increased stress reticulocytes.

Anemia, Sickle Cell↗

PubMed1992

High expression of human beta S- and alpha-globins in transgenic mice: erythrocyte abnormalities, organ damage, and the effect of hypoxia.

A line of transgenic mice with two cointegrated transgenes, the human beta S- and alpha 2-globin genes, linked to the beta-globin locus control region was produced and bred with mice carrying a deletion of the mouse beta major-globin gene. In transgenic mice homozygous for the beta major deletion (alpha H beta S[beta MDD]; where alpha H is human alpha-globin and MD is mouse deletion), 72.5 +/- 2.4% (mean +/- SD) of the beta-chains are beta S and the ratio of alpha H- to beta S-globin was 0.73. Introduction of a heterozygous mouse alpha-globin deletion into mice homozygous for the beta major deletion (alpha H beta S[alpha MD beta MDD]) resulted in 65.1 +/- 8.5% beta S and a human alpha/beta ratio of 0.89 +/- 0.2. Sickling occurs in 95% of erythrocytes from alpha H beta S[beta MDD] mice after slow deoxygenation. Transmission electron microscopy revealed polymer fiber formation but not fascicles of fiber. Increased organ weight was noted in lung, spleen, and kidney of transgenic mice vs. controls that may be due to hypertrophy or increased blood volume in the lungs and/or increased tissue water content. The hemoglobin content of lung, spleen, and kidney was also elevated in transgenic animals due to trapped hemoglobin and/or increased blood volume. When transgenic and control mice were examined by magnetic resonance imaging at 9.4 tesla, some transgenic animals had enlarged kidneys with prolonged relaxation time, consistent with increased organ weight and water content. The glomerular filtration rate was elevated in transgenic animals, which is characteristic of young sickle cell patients. Furthermore, exposure to hypoxia resulted in significantly decreased hematocrit, increased erythrocyte density, and induced a urine-concentrating defect. We conclude that the transgenic mouse line reported here has chronic organ damage and further hematological and organ dysfunction can be induced by hypoxia.

Anemia, Sickle Cell↗

PubMed1992

Use of embryonic stem cells to study mutations affecting postimplantation development in the mouse.

The generation and analysis of insertional mutations that perturb early postimplantation development provide a means to identify genes required at this stage of embryogenesis. We have been studying two independently generated insertional mutations termed 413.d and H beta 58 that result in early postimplantation lethality. Each mutation is associated with a distinct phenotype. 413.d mutant embryos become profoundly abnormal around the time of gastrulation: no identifiable embryonic axis or mesodermal structures are formed. H beta 58 mutant embryos proceed further in development, forming a relatively normal anteroposterior axis before developmental arrest occurs. We isolated embryonic stem cell lines homozygous for each of these mutations and assessed their differentiation abilities and developmental potential in vitro and after their introduction into wild-type blastocysts. From these studies we conclude that the 413.d mutation acts in a non-cell-autonomous fashion: mutant cells appear capable of participating, in conjunction with wild-type cells, in the formation of derivatives of all three primary cell lineages of the embryo. H beta 58 mutant embryonic stem cells are clearly pluripotent but they appear to be more restricted in their developmental potential, suggesting that the H beta 58 gene product may be required by specific tissues of the embryo.

Animals↗

PubMed1992

Rescue of erythroid development in gene targeted GATA-1- mouse embryonic stem cells.

Development of definitive (fetal liver-derived) red cells is blocked by a targeted mutation in the gene encoding the transcription factor GATA-1. We used in vitro differentiation of GATA-1- mouse embryonic stem (ES) cells to reveal a requirement for GATA-1 during primitive (yolk sac-derived) erythropoiesis and to establish a rescue assay. We show that the block to development includes primitive, as well as definitive, erythroid cells and is complete at the level of globin RNA expression; that the introduction of a normal GATA-1 gene restores developmental potential both in vivo and in vitro; and that efficient rescue is dependent on a putative autoregulatory GATA-motif in the distal promoter. Use of in vitro differentiated ES cells bridges a gap between conventional approaches to gene function in cell lines and analysis of loss of function mutations in the whole animal.

Animals↗