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T Doetschman

Publications and source records attributed to T Doetschman.

At least 19 recordsLinked to original sources
PubMed2001

Double-outlet right ventricle and overriding tricuspid valve reflect disturbances of looping, myocardialization, endocardial cushion differentiation, and apoptosis in TGF-beta(2)-knockout mice.

BACKGROUND: Transforming growth factor-beta(2) (TGF-beta(2)) is a member of a family of growth factors with the potential to modify multiple processes. Mice deficient in the TGF-beta(2) gene die around birth and show a variety of defects of different organs, including the heart. METHODS AND RESULTS: We studied the hearts of TGF-beta(2)-null mouse embryos from 11.5 to 18.5 days of gestation to analyze the types of defects and determine which processes of cardiac morphogenesis are affected by the absence of TGF-beta(2). Analysis of serial sections revealed malformations of the outflow tract (typically a double-outlet right ventricle) in 87.5%. There was 1 case of common arterial trunk. Abnormal thickening of the semilunar valves was seen in 4.2%. Associated malformations of the atrioventricular (AV) canal were found in 62.5% and were composed of perimembranous inlet ventricular septal defects (37.5%), AV valve thickening (33.3%), overriding tricuspid valve (25.0%), and complete AV septal defects (4.2%). Anomalies of the aorta and its branches were seen in 33.3%. Immunohistochemical staining showed failure of myocardialization of the mesenchyme of the atrial septum and the ventricular outflow tract as well as deficient valve differentiation. Morphometry documented this to be associated with absence of the normal decrease of total endocardial cushion volume in the older stages. Apoptosis in TGF-beta(2)-knockout mice was increased, although regional distribution was normal. CONCLUSIONS: TGF-beta(2)-knockout mice exhibited characteristic cardiovascular anomalies comparable to malformations seen in the human population.

Animals↗
PubMed2001

Salivary acinar cells from aquaporin 5-deficient mice have decreased membrane water permeability and altered cell volume regulation.

Aquaporins (AQPs) are channel proteins that regulate the movement of water through the plasma membrane of secretory and absorptive cells in response to osmotic gradients. In the salivary gland, AQP5 is the major aquaporin expressed on the apical membrane of acinar cells. Previous studies have shown that the volume of saliva secreted by AQP5-deficient mice is decreased, indicating a role for AQP5 in saliva secretion; however, the mechanism by which AQP5 regulates water transport in salivary acinar cells remains to be determined. Here we show that the decreased salivary flow rate and increased tonicity of the saliva secreted by Aqp5(-)/- mice in response to pilocarpine stimulation are not caused by changes in whole body fluid homeostasis, indicated by similar blood gas and electrolyte concentrations in urine and blood in wild-type and AQP5-deficient mice. In contrast, the water permeability in parotid and sublingual acinar cells isolated from Aqp5(-)/- mice is decreased significantly. Water permeability decreased by 65% in parotid and 77% in sublingual acinar cells from Aqp5(-)/- mice in response to hypertonicity-induced cell shrinkage and hypotonicity-induced cell swelling. These data show that AQP5 is the major pathway for regulating the water permeability in acinar cells, a critical property of the plasma membrane which determines the flow rate and ionic composition of secreted saliva.

Animals↗
PubMed2001

Cardiovascular abnormalities with normal blood pressure in tissue kallikrein-deficient mice.

Tissue kallikrein is a serine protease thought to be involved in the generation of bioactive peptide kinins in many organs like the kidneys, colon, salivary glands, pancreas, and blood vessels. Low renal synthesis and urinary excretion of tissue kallikrein have been repeatedly linked to hypertension in animals and humans, but the exact role of the protease in cardiovascular function has not been established largely because of the lack of specific inhibitors. This study demonstrates that mice lacking tissue kallikrein are unable to generate significant levels of kinins in most tissues and develop cardiovascular abnormalities early in adulthood despite normal blood pressure. The heart exhibits septum and posterior wall thinning and a tendency to dilatation resulting in reduced left ventricular mass. Cardiac function estimated in vivo and in vitro is decreased both under basal conditions and in response to beta-adrenergic stimulation. Furthermore, flow-induced vasodilatation is impaired in isolated perfused carotid arteries, which express, like the heart, low levels of the protease. These data show that tissue kallikrein is the main kinin-generating enzyme in vivo and that a functional kallikrein-kinin system is necessary for normal cardiac and arterial function in the mouse. They suggest that the kallikrein-kinin system could be involved in the development or progression of cardiovascular diseases.

Animals↗
PubMed2000

Stomachs of mice lacking the gastric H,K-ATPase alpha -subunit have achlorhydria, abnormal parietal cells, and ciliated metaplasia.

The H,K-ATPase of the gastric parietal cell is the most critical component of the ion transport system mediating acid secretion in the stomach. To study the requirement of this enzyme in the development, maintenance, and function of the gastric mucosa, we used gene targeting to prepare mice lacking the alpha-subunit. Homozygous mutant (Atp4a(-/-)) mice appeared healthy and exhibited normal systemic electrolyte and acid-base status but were achlorhydric and hypergastrinemic. Immunocytochemical, histological, and ultrastructural analyses of Atp4a(-/-) stomachs revealed the presence of chief cells, demonstrating that the lack of acid secretion does not interfere with their differentiation. Parietal cells were also present in normal numbers, and despite the absence of alpha-subunit mRNA and protein, the beta-subunit was expressed. However, Atp4a(-/-) parietal cells had dilated canaliculi and lacked typical canalicular microvilli and tubulovesicles, and subsets of these cells contained abnormal mitochondria and/or massive glycogen stores. Stomachs of adult Atp4a(-/-) mice exhibited metaplasia, which included the presence of ciliated cells. We conclude that ablation of the H,K-ATPase alpha-subunit causes achlorhydria and hypergastrinemia, severe perturbations in the secretory membranes of the parietal cell, and metaplasia of the gastric mucosa; however, the absence of the pump appears not to perturb parietal cell viability or chief cell differentiation.

Achlorhydria↗
PubMed2000

Delayed wound healing in immunodeficient TGF-beta 1 knockout mice.

Previous studies showed that full-thickness wounds in transforming growth factor-beta1-deficient mice initially heal normally. Unfortunately, transforming growth factor-beta1 deficiency leads to a multifocal inflammatory disease affecting most organs of the body, which ultimately interferes with later stages of wound healing in these mice. As this inflammatory disease is eliminated in transforming growth factor-beta1-deficient mice lacking T and B cells (Tgfb1-/- Scid-/- mice), we hypothesized that wound repair in the latter would proceed normally, even at later stages of healing. Unexpectedly, Tgfb1-/- Scid-/- mice demonstrate a major delay of approximately 1 wk in each of the major phases of wound healing: inflammation, proliferation, and maturation. Immuno- deficient Scid-/- mice that have the wild-type Tgfb1 allele do not experience this delay in wound healing. One interpretation of these findings is that lymphocytes and transforming growth factor-beta1 affect compensatory pathways in wound healing. An alternative interpretation is that the delayed expression of Tgfb2 and Tgfb3 that occurs in the absence of transforming growth factor-beta1 results in the delayed wound healing, suggesting that transforming growth factor-beta2 and/or transforming growth factor-beta3 play important parts in wound healing.

Animals↗
PubMed2000

Disruption of the fibroblast growth factor-2 gene results in decreased bone mass and bone formation.

Basic fibroblast growth factor (FGF-2), an important modulator of cartilage and bone growth and differentiation, is expressed and regulated in osteoblastic cells. To investigate the role of FGF-2 in bone, we examined mice with a disruption of the Fgf2 gene. Measurement of trabecular bone architecture of the femoral metaphysis of Fgf2(+/+) and Fgf2(-/-) adult mice by micro-CT revealed that the platelike trabecular structures were markedly reduced and many of the connecting rods of trabecular bone were lost in the Fgf2(-/-) mice. Dynamic histomorphometry confirmed a significant decrease in trabecular bone volume, mineral apposition, and bone formation rates. In addition, there was a profound decreased mineralization of bone marrow stromal cultures from Fgf2(-/-) mice. This study provides strong evidence that FGF-2 helps determine bone mass as well as bone formation.

Alkaline Phosphatase↗
PubMed1999

Transforming growth factor beta1 enhances platelet aggregation through a non-transcriptional effect on the fibrinogen receptor.

Upon activation, platelets store and release large amounts of the peptide transforming growth factor beta1 (TGFbeta1). The released TGFbeta1 can then act on nearby vascular cells to mediate subsequent vessel repair. In addition, TGFbeta1 may circulate to bone marrow and regulate megakaryocyte activity. It is not known what effect, if any, TGFbeta1 has on platelets. Adult TGFbeta1-deficient mice exhibit thrombocythemia and a mild bleeding disorder that is shown to result from faulty platelet aggregation. TGFbeta1-deficient platelets are shown to contain functional receptors, and preincubation with recombinant TGFbeta1 improves aggregation, demonstrating that TGFbeta1 plays an active role in platelet aggregation. TGFbeta1-deficient platelets fail to retain bound fibrinogen in response to aggregation agonists, but they possess normal levels of the alpha(IIb)/beta(3) fibrinogen receptor. Signaling from agonist receptors is normal because the platelets change shape, produce thromboxane A(2), and present P-selectin in response to stimulation. Consequently, activation and maintenance of alpha(IIb)/beta(3) into a fibrinogen-binding conformation is impaired in the absence of TGFbeta1. 4-Phorbol 12-myristate 13-acetate treatment and protein kinase C activity measurements suggest a defect downstream of protein kinase C in its activation cascade. Because platelets lack nuclei, these data demonstrate for the first time a non-transcriptionally mediated TGFbeta1 signaling pathway that enhances the activation and maintenance of integrin function.

Adenosine Diphosphate↗
PubMed1999

Mice lacking the basolateral Na-K-2Cl cotransporter have impaired epithelial chloride secretion and are profoundly deaf.

In chloride-secretory epithelia, the basolateral Na-K-2Cl cotransporter (NKCC1) is thought to play a major role in transepithelial Cl(-) and fluid transport. Similarly, in marginal cells of the inner ear, NKCC1 has been proposed as a component of the entry pathway for K(+) that is secreted into the endolymph, thus playing a critical role in hearing. To test these hypotheses, we generated and analyzed an NKCC1-deficient mouse. Homozygous mutant (Nkcc1(-/-)) mice exhibited growth retardation, a 28% incidence of death around the time of weaning, and mild difficulties in maintaining their balance. Mean arterial blood pressure was significantly reduced in both heterozygous and homozygous mutants, indicating an important function for NKCC1 in the maintenance of blood pressure. cAMP-induced short circuit currents, which are dependent on the CFTR Cl(-) channel, were reduced in jejunum, cecum, and trachea of Nkcc1(-/-) mice, indicating that NKCC1 contributes to cAMP-induced Cl(-) secretion. In contrast, secretion of gastric acid in adult Nkcc1(-/-) stomachs and enterotoxin-stimulated fluid secretion in the intestine of suckling Nkcc1(-/-) mice were normal. Finally, homozygous mutants were deaf, and histological analysis of the inner ear revealed a collapse of the membranous labyrinth, consistent with a critical role for NKCC1 in transepithelial K(+) movements involved in generation of the K(+)-rich endolymph and the endocochlear potential.

Animals↗
PubMed1999

The TGF-beta2 isoform is both a required and sufficient inducer of murine hair follicle morphogenesis.

Hair follicle development serves as an excellent model to study control of organ morphogenesis. Three specific isoforms of TGF-beta exist which exhibit a distinct pattern of expression during hair follicle morphogenesis. To clarify the still elusive role of these factors in hair follicle development, we have used a combined genetic and functional approach: analysis of hair follicle development in mice with disruptions of the TGF-beta1, 2, and 3 genes was coupled with a direct functional test of the effect of added purified factors on fetal hair follicle development in skin organ cultures. TGF-beta2 null mice exhibited a profound delay of hair follicle morphogenesis, with a 50% reduced number of hair follicles. In contrast to hair follicle development, growth and differentiation of interfollicular keratinocytes proceeded unimpaired. Unlike TGF-beta2-/- mice, mice with a disruption of the TGF-beta1 gene showed slightly advanced hair follicle formation, while lack of the TGF-beta3 gene did not have any effects. Treatment of wild-type, embryonic skin explants (E14.5 or E15.5) with TGF-beta2 protein in either soluble form or slow release beads induced hair follicle development and epidermal hyperplasia, while similar TGF-beta1 treatment exerted suppressive effects. Thus, the TGF-beta2 isoform plays a specific role, not shared by the other TGF-beta isoforms, as an inducer of hair follicle morphogenesis and is both required and sufficient to promote this process.

Animals↗
PubMed1999

Transforming growth factor beta1 suppresses nonmetastatic colon cancer at an early stage of tumorigenesis.

The transforming growth factor beta (TGF-beta) pathway is known to play an important role in both human and urine colon cancer. However, the staging, ligand specificity, and mechanism underlying the tumor suppressive activity of this pathway are unknown. We developed a mouse model for colon cancer that identifies an early role for TGF-beta1 in tumor suppression and implicates TGF-beta2 or TGF-beta3 in the prevention of metastasis. Analysis of the development of colon cancer in TGF-beta1 knockout mice pinpoints the defect to the hyperplasty/adenoma transition and reveals that the mechanism involves an inability to maintain epithelial tissue organization and not a loss of growth control, increased inflammatory activity, or increased genetic instability. These mice provide a unique opportunity to investigate the specific role of TGF-beta1 at this critical transition in the development of colon cancer.

Adenocarcinoma↗
PubMed1999

TGF-beta3, but not TGF-beta1, protects keratinocytes against 12-O-tetradecanoylphorbol-13-acetate-induced cell death in vitro and in vivo.

We have examined the role that individual TGF-beta isoforms, and in particular TGF-beta3, play in control of epidermal homeostasis. Mice with a knockout mutation of the TGF-beta3 gene die a few hours after birth. A full-thickness skin grafting approach was used to investigate the postnatal development and homeostatic control of the skin of these mice. Grafted skin of mice with a disruption of the TGF-beta3 gene developed similarly to grafts of wild type and TGF-beta1 knockout animals. However, a strikingly different response was observed after acute treatment with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). When exposed to TPA, the grafted skin of wild type and TGF-beta1 knockout mice underwent a hyperplastic response similar to that of normal mouse skin. In marked contrast, TPA treatment of TGF-beta3 knockout grafts induced widespread areas of keratinocyte cell death. Analysis of cultured keratinocytes treated with purified TGF-beta isoforms revealed that TGF-beta3 plays a direct and specific function in protecting keratinocytes against TPA-induced cell death. The protective function of TGF-beta3 on TPA-induced cell death was not because of general suppression of the signaling pathways triggered by this agent, as ERK1/2 activation occurred to a similar if not greater extent in TGF-beta3-treated versus control keratinocytes. Instead, TGF-beta3 treatment led to a significant reduction in TPA-induced c-Jun N-terminal kinase activity, which was associated and possibly explained by specific counteracting effects of TGF-beta3 on TPA-induced disruption of keratinocyte focal adhesions.

Actins↗
PubMed1999

Impaired cardiac performance in heterozygous mice with a null mutation in the sarco(endo)plasmic reticulum Ca2+-ATPase isoform 2 (SERCA2) gene.

The sarco(endo)plasmic reticulum Ca2+-ATPase isoform 2 (SERCA2) gene encodes both SERCA2a, the cardiac sarcoplasmic reticulum Ca2+ pump, and SERCA2b, which is expressed in all tissues. To gain a better understanding of the physiological functions of SERCA2, we used gene targeting to develop a mouse in which the promoter and 5' end of the gene were eliminated. Mating of heterozygous mutant mice yielded wild-type and heterozygous offspring; homozygous mutants were not observed. RNase protection, Western blotting, and biochemical analysis of heart samples showed that SERCA2 mRNA was reduced by approximately 45% in heterozygous mutant hearts and that SERCA2 protein and maximal velocity of Ca2+ uptake into the sarcoplasmic reticulum were reduced by approximately 35%. Measurements of cardiovascular performance via transducers in the left ventricle and right femoral artery of the anesthetized mouse revealed reductions in mean arterial pressure, systolic ventricular pressure, and the absolute values of both positive and negative dP/dt in heterozygous mutants. These results demonstrate that two functional copies of the SERCA2 gene are required to maintain normal levels of SERCA2 mRNA, protein, and Ca2+ sequestering activity, and that the deficit in Ca2+ sequestering activity due to the loss of one copy of the SERCA2 gene impairs cardiac contractility and relaxation.

Animals↗
PubMed1999

Strain dependency of TGFbeta1 function during embryogenesis.

There is incomplete penetrance to Tgfb1 knockout phenotypes. About 50% of Tgfb1 homozygous mutant (Tgfb1-/-) and 25% of Tgfb1 heterozygous (Tgfb1+/-) embryos die during embryogenesis. In a mixed NIH/Ola x C57BL/6J/Ola x 129 background partial embryonic lethality of the Tgfb1-/-embryos occurs due to defective yolk sac vasculopoiesis and/or hematopoiesis. We show here that on a predominantly CF-1 genetic background, lack of TGFbeta1 causes a pre-morula lethality in about 50% of the null embryos. This partial lethality is not reversed by transfer of Tgfb1-/- embryos to Tgfb1-/+ hosts. The extent of embryonic lethality in Tgfb1-/- embryos ranges in a background dependent manner from 20% to 100%. Based on these and other studies it is clear that TGFbeta1 acts at two distinct phases of embryogenesis: pre-implantation development and yolk sac vasculogenesis/hematopoiesis. The susceptibility for the pre-implantation lethality depends on a small number of genetic modifiers since a small number of backcrosses onto the high susceptibility strain C57BL/6 leads to complete penetrance of the lethality.

Animals↗
PubMed1999

Changes in cerebral cortex size are governed by fibroblast growth factor during embryogenesis.

We show that fibroblast growth factor 2 (FGF2) and FGF receptors are transiently expressed by cells of the pseudostratified ventricular epithelium (PVE) during early neurogenesis. A single microinjection of FGF2 into cerebral ventricles of rat embryos at E15.5 increased the volume and total number of neurons in the adult cerebral cortex by 18% and 87%, respectively. Microinjection of FGF2 by the end of neurogenesis, at E20.5, selectively increased the number of glia. Mice lacking the FGF2 gene had fewer cortical neurons and glia at maturity. BrdU studies in FGF2-microinjected and FGF2-null animals suggested that FGF2 increases the proportion of dividing cells in the PVE without affecting the cell-cycle length. Thus, FGF2 increases the number of rounds of division of cortical progenitors.

Animals↗
PubMed1999

Fibroblast growth factor-2 mediates pressure-induced hypertrophic response.

In vitro, fibroblast growth factor-2 (FGF2) has been implicated in cardiomyocyte growth and reexpression of fetal contractile genes, both markers of hypertrophy. However, its in vivo role in cardiac hypertrophy during pressure overload is not well characterized. Mice with or without FGF2 (Fgf2(+/+) and Fgf2(-/-), respectively) were subjected to transverse aortic coarctation (AC). Left ventricular (LV) mass and wall thickness were assessed by echocardiography preoperatively and once a week postoperatively for 10 weeks. In vivo LV function during dobutamine stimulation, cardiomyocyte cross-sectional area, and recapitulation of fetal cardiac genes were also measured. AC Fgf2(-/-) mice develop significantly less hypertrophy (4-24% increase) compared with AC Fgf2(+/+) mice (41-52% increase). Cardiomyocyte cross-sectional area is significantly reduced in AC Fgf2(-/-) mice. Noncoarcted (NC) and AC Fgf2(-/-) mice have similar beta-adrenergic responses, but those of AC Fgf2(+/+) mice are blunted. A lack of mitotic growth in both AC Fgf2(+/+) and Fgf2(-/-) hearts indicates a hypertrophic response of cardiomyocytes. Consequently, FGF2 plays a major role in cardiac hypertrophy. Comparison of alpha- and beta-cardiac myosin heavy chain mRNA and protein levels in NC and AC Fgf2(+/+) and Fgf2(-/-) mice indicates that myosin heavy chain composition depends on hemodynamic stress rather than on FGF2 or hypertrophy, and that isoform switching is transcriptionally, not posttranscriptionally, regulated.

Animals↗
PubMed1999

Cardiac hypertrophy with preserved contractile function after selective deletion of GLUT4 from the heart.

Glucose enters the heart via GLUT1 and GLUT4 glucose transporters. GLUT4-deficient mice develop striking cardiac hypertrophy and die prematurely. Whether their cardiac changes are caused primarily by GLUT4 deficiency in cardiomyocytes or by metabolic changes resulting from the absence of GLUT4 in skeletal muscle and adipose tissue is unclear. To determine the role of GLUT4 in the heart we used cre-loxP recombination to generate G4H(-/-) mice in which GLUT4 expression is abolished in the heart but is present in skeletal muscle and adipose tissue. Life span and serum concentrations of insulin, glucose, FFAs, lactate, and beta-hydroxybutyrate were normal. Basal cardiac glucose transport and GLUT1 expression were both increased approximately 3-fold in G4H(-/-) mice, but insulin-stimulated glucose uptake was abolished. G4H(-/-) mice develop modest cardiac hypertrophy associated with increased myocyte size and induction of atrial natriuretic and brain natriuretic peptide gene expression in the ventricles. Myocardial fibrosis did not occur. Basal and isoproterenol-stimulated isovolumic contractile performance was preserved. Thus, selective ablation of GLUT4 in the heart initiates a series of events that results in compensated cardiac hypertrophy.

Animals↗