Murine tissue factor disulfide mutation causes a bleeding phenotype with sex specific organ pathology and lethality.

Tissue factor is highly expressed in sub-endothelial tissue. The extracellular allosteric disulfide bond Cys186-Cys209 of human tissue factor shows high evolutionary conservation and in vitro evidence suggests that it significantly contributes to tissue factor procoagulant activity. To investigate the role of this allosteric disulfide bond in vivo, we generated a C213G mutant tissue factor mouse by replacing Cys213 of the corresponding disulfide Cys190-Cys213 in murine tissue factor.

MFAP4 Promotes Vascular Smooth Muscle Migration, Proliferation and Accelerates Neointima Formation.

Objective: Arterial injury stimulates remodeling responses that, when excessive, lead to stenosis. These responses are influenced by integrin signaling in vascular smooth muscle cells (VSMCs). Microfibrillar-associated protein 4 (MFAP4) is an integrin ligand localized to extracellular matrix fibers in the vascular wall.

Analysis of mammalian gene function through broad-based phenotypic screens across a consortium of mouse clinics.

The function of the majority of genes in the mouse and human genomes remains unknown. The mouse embryonic stem cell knockout resource provides a basis for the characterization of relationships between genes and phenotypes. The EUMODIC consortium developed and validated robust methodologies for the broad-based phenotyping of knockouts through a pipeline comprising 20 disease-oriented platforms.

Heart-Specific Knockout of the Mitochondrial Thioredoxin Reductase (Txnrd2) Induces Metabolic and Contractile Dysfunction in the Aging Myocardium.

Background: Ubiquitous deletion of thioredoxin reductase 2 (Txnrd2) in mice is embryonically lethal and associated with abnormal heart development, while constitutive, heart-specific Txnrd2 inactivation leads to dilated cardiomyopathy and perinatal death. The significance of Txnrd2 in aging cardiomyocytes, however, has not yet been examined.

Methods And Results: The tamoxifen-inducible heart-specific αMHC-MerCreMer transgene was used to inactivate loxP-flanked Txnrd2 alleles in adult mice.

MTO1-deficient mouse model mirrors the human phenotype showing complex I defect and cardiomyopathy.

Recently, mutations in the mitochondrial translation optimization factor 1 gene (MTO1) were identified as causative in children with hypertrophic cardiomyopathy, lactic acidosis and respiratory chain defect. Here, we describe an MTO1-deficient mouse model generated by gene trap mutagenesis that mirrors the human phenotype remarkably well. As in patients, the most prominent signs and symptoms were cardiovascular and included bradycardia and cardiomyopathy.

Standardized, systemic phenotypic analysis of Slc12a1I299F mutant mice.

Background: Type I Bartter syndrome is a recessive human nephropathy caused by loss-of-function mutations in the SLC12A1 gene coding for the Na+-K+-2Cl- cotransporter NKCC2. We recently established the mutant mouse line Slc12a1I299F exhibiting kidney defects highly similar to the late-onset manifestation of this hereditary human disease. Besides the kidney defects, low blood pressure and osteopenia were revealed in the homozygous mutant mice which were also described in humans.

Standardized, systemic phenotypic analysis of Umod(C93F) and Umod(A227T) mutant mice.

Uromodulin-associated kidney disease (UAKD) summarizes different clinical features of an autosomal dominant heritable disease syndrome in humans with a proven uromodulin (UMOD) mutation involved. It is often characterized by hyperuricemia, gout, alteration of urine concentrating ability, as well as a variable rate of disease progression inconstantly leading to renal failure and histological alterations of the kidneys. We recently established the two Umod mutant mouse lines Umod(C93F) and Umod(A227T) on the C3H inbred genetic background both showing kidney defects analogous to those found in human UAKD patients.

Innovations in phenotyping of mouse models in the German Mouse Clinic.

Under the label of the German Mouse Clinic (GMC), a concept has been developed and implemented that allows the better understanding of human diseases on the pathophysiological and molecular level. This includes better understanding of the crosstalk between different organs, pleiotropy of genes, and the systemic impact of envirotypes and drugs. In the GMC, experts from various fields of mouse genetics and physiology, in close collaboration with clinicians, work side by side under one roof.

Cytochrome c oxidase subunit 4 isoform 2-knockout mice show reduced enzyme activity, airway hyporeactivity, and lung pathology.

Cytochrome c oxidase (COX) is the terminal enzyme of the mitochondrial electron transport chain. The purpose of this study was to analyze the function of lung-specific cytochrome c oxidase subunit 4 isoform 2 (COX4i2) in vitro and in COX4i2-knockout mice in vivo. COX was isolated from cow lung and liver as control and functionally analyzed.

Large-scale phenotyping of an accurate genetic mouse model of JNCL identifies novel early pathology outside the central nervous system.

Cln3(Δex7/8) mice harbor the most common genetic defect causing juvenile neuronal ceroid lipofuscinosis (JNCL), an autosomal recessive disease involving seizures, visual, motor and cognitive decline, and premature death. Here, to more thoroughly investigate the manifestations of the common JNCL mutation, we performed a broad phenotyping study of Cln3(Δex7/8) mice. Homozygous Cln3(Δex7/8) mice, congenic on a C57BL/6N background, displayed subtle deficits in sensory and motor tasks at 10-14 weeks of age.

Cardiopulmonary dysfunction in the Osteogenesis imperfecta mouse model Aga2 and human patients are caused by bone-independent mechanisms.

Osteogenesis imperfecta (OI) is an inherited connective tissue disorder with skeletal dysplasia of varying severity, predominantly caused by mutations in the collagen I genes (COL1A1/COL1A2). Extraskeletal findings such as cardiac and pulmonary complications are generally considered to be significant secondary features. Aga2, a murine model for human OI, was systemically analyzed in the German Mouse Clinic by means of in vivo and in vitro examinations of the cardiopulmonary system, to identify novel mechanisms accounting for perinatal lethality.

Requirement of the RNA-editing enzyme ADAR2 for normal physiology in mice.

ADAR2, an RNA editing enzyme that converts specific adenosines to inosines in certain pre-mRNAs, often leading to amino acid substitutions in the encoded proteins, is mainly expressed in brain. Of all ADAR2-mediated edits, a single one in the pre-mRNA of the AMPA receptor subunit GluA2 is essential for survival. Hence, early postnatal death of mice lacking ADAR2 is averted when the critical edit is engineered into both GluA2 encoding Gria2 alleles.

Post-stroke inhibition of induced NADPH oxidase type 4 prevents oxidative stress and neurodegeneration.

Ischemic stroke is the second leading cause of death worldwide. Only one moderately effective therapy exists, albeit with contraindications that exclude 90% of the patients. This medical need contrasts with a high failure rate of more than 1,000 pre-clinical drug candidates for stroke therapies.

Mouse phenotyping.

Model organisms like the mouse are important tools to learn more about gene function in man. Within the last 20 years many mutant mouse lines have been generated by different methods such as ENU mutagenesis, constitutive and conditional knock-out approaches, knock-down, introduction of human genes, and knock-in techniques, thus creating models which mimic human conditions. Due to pleiotropic effects, one gene may have different functions in different organ systems or time points during development.

Mutation of the Na(+)-K(+)-2Cl(-) cotransporter NKCC2 in mice is associated with severe polyuria and a urea-selective concentrating defect without hyperreninemia.

The bumetanide-sensitive Na(+)-K(+)-2Cl(-) cotransporter NKCC2, located in the thick ascending limb of Henle's loop, plays a critical role in the kidney's ability to concentrate urine. In humans, loss-of-function mutations of the solute carrier family 12 member 1 gene (SLC12A1), coding for NKCC2, cause type I Bartter syndrome, which is characterized by prenatal onset of a severe polyuria, salt-wasting tubulopathy, and hyperreninemia. In this study, we describe a novel chemically induced, recessive mutant mouse line termed Slc12a1(I299F) exhibiting late-onset manifestation of type I Bartter syndrome.

Novel missense mutation of uromodulin in mice causes renal dysfunction with alterations in urea handling, energy, and bone metabolism.

Uromodulin-associated kidney disease is a heritable renal disease in humans caused by mutations in the uromodulin (UMOD) gene. The pathogenesis of the disease is mostly unknown. In this study, we describe a novel chemically induced mutant mouse line termed Umod(A227T) exhibiting impaired renal function.

Dll1 haploinsufficiency in adult mice leads to a complex phenotype affecting metabolic and immunological processes.

Background: The Notch signaling pathway is an evolutionary conserved signal transduction pathway involved in embryonic patterning and regulation of cell fates during development and self-renewal. Recent studies have demonstrated that this pathway is integral to a complex system of interactions, involving as well other signal transduction pathways, and implicated in distinct human diseases. Delta-like 1 (Dll1) is one of the known ligands of the Notch receptors.

A humanized version of Foxp2 affects cortico-basal ganglia circuits in mice.

It has been proposed that two amino acid substitutions in the transcription factor FOXP2 have been positively selected during human evolution due to effects on aspects of speech and language. Here, we introduce these substitutions into the endogenous Foxp2 gene of mice. Although these mice are generally healthy, they have qualitatively different ultrasonic vocalizations, decreased exploratory behavior and decreased dopamine concentrations in the brain suggesting that the humanized Foxp2 allele affects basal ganglia.

Systemic first-line phenotyping.

With the completion of the mouse genome sequence an essential task for biomedical sciences in the twenty-first century will be the generation and functional analysis of mouse models for every gene in the mammalian genome. More than 30,000 mutations in ES cells will be engineered and thousands of mouse disease models will become available over the coming years by the collaborative effort of the International Mouse Knockout Consortium. In order to realize the full value of the mouse models proper characterization, archiving and dissemination of mouse disease models to the research community have to be performed.

Pleiotropic effects in Eya3 knockout mice.

Background: In Drosophila, mutations in the gene eyes absent (eya) lead to severe defects in eye development. The functions of its mammalian orthologs Eya1-4 are only partially understood and no mouse model exists for Eya3. Therefore, we characterized the phenotype of a new Eya3 knockout mouse mutant.

"Sighted C3H" mice--a tool for analysing the influence of vision on mouse behaviour?

It is unclear what role vision plays in guiding mouse behaviour, since the mouse eye is of comparably low optical quality, and mice are considered to rely primarily on other senses. All C3H substrains are homozygous for the Pde6b(rd1) mutation and get blind by weaning age. To study the impact of the Pde6b(rd1) mutation on mouse behaviour and physiology, sighted C3H (C3H.

Atrial natriuretic peptide and osteopontin are useful markers of cardiac disorders in mice.

Biomarkers are not established for cardiovascular phenotyping in mice. We compared the use of echocardiography with the determination of N-terminal propeptide of the atrial natriuretic peptide (Nt-proANP) and osteopontin (Opn). We measured plasma Nt-proANP and Opn levels in (1) the inbred strains C57BL/6, BALB/c, C3H/He, DBA/2, FVB/N, 129S1/Sv; (2) a surgical model of nonischemic myocardial infarction; and (3) delta-sarcoglycan (Sgcd) and calsarcin 1 [also known as myozenin 2 (Myoz2)] knockout models of cardiomyopathy.