Loss of functional endothelial connexin40 results in exercise-induced hypertension in mice.

During activity, coordinated vasodilation of microcirculatory networks with upstream supply vessels increases blood flow to skeletal and cardiac muscles and reduces peripheral resistance. Endothelial dysfunction in humans attenuates activity-dependent vasodilation, resulting in exercise-induced hypertension in otherwise normotensive individuals. Underpinning activity-dependent hyperemia is an ascending vasodilation in which the endothelial gap junction protein, connexin (Cx)40, plays an essential role.

Dual proinflammatory and antiviral properties of pulmonary eosinophils in respiratory syncytial virus vaccine-enhanced disease.

Unlabelled: Human respiratory syncytial virus (RSV) is a major cause of morbidity and severe lower respiratory tract disease in the elderly and very young, with some infants developing bronchiolitis, recurrent wheezing, and asthma following infection. Previous studies in humans and animal models have shown that vaccination with formalin-inactivated RSV (FI-RSV) leads to prominent airway eosinophilic inflammation following RSV challenge; however, the roles of pulmonary eosinophilia in the antiviral response and in disease pathogenesis are inadequately understood. In vivo studies in mice with eotaxin and/or interleukin 5 (IL-5) deficiency showed that FI-RSV vaccination did not lead to enhanced pulmonary disease, where following challenge there were reduced pulmonary eosinophilia, inflammation, Th2-type cytokine responses, and altered chemokine (TARC and CCL17) responses.

Perturbation of chemical coupling by an endothelial Cx40 mutant attenuates endothelium-dependent vasodilation by KCa channels and elevates blood pressure in mice.

Mutant forms of connexin40 (Cx40) exist in the human population and predispose carriers to atrial fibrillation. Since endothelial expression of Cx40 is important for electrical and chemical communication within the arterial wall, carriers of mutant Cx40 proteins may be predisposed to peripheral arterial dysfunction and dysregulation of blood pressure. We have therefore studied mice expressing either a chemically dysfunctional mutant, Cx40T202S, or wild-type Cx40, with native Cx40, specifically in the endothelium.

Nox1 upregulates the function of vascular T-type calcium channels following chronic nitric oxide deficit.

Cardiovascular disease is characterised by reduced nitric oxide bioavailability resulting from oxidative stress. Our previous studies have shown that nitric oxide deficit per se increases the contribution of T-type calcium channels to vascular tone through increased superoxide from NADPH oxidase (Nox). The aim of the present study was therefore to identify the Nox isoform responsible for modulating T-type channel function, as T-type channels are implicated in several pathophysiological conditions involving oxidative stress.

The kinematics of cytotoxic lymphocytes influence their ability to kill target cells.

Cytotoxic lymphocytes (CTL) have been reported to show a range of motility patterns from rapid long-range tracking to complete arrest, but how and whether these kinematics affect their ability to kill target cells is not known. Many in vitro killing assays utilize cell lines and tumour-derived cells as targets, which may be of limited relevance to the kinetics of CTL-mediated killing of somatic cells. Here, live-cell microscopy is used to examine the interactions of CTL and primary murine skin cells presenting antigens.

Regulation of endothelial-specific transgene expression by the LacI repressor protein in vivo.

Genetically modified mice have played an important part in elucidating gene function in vivo. However, conclusions from transgenic studies may be compromised by complications arising from the site of transgene integration into the genome and, in inducible systems, the non-innocuous nature of inducer molecules. The aim of the present study was to use the vascular system to validate a technique based on the bacterial lac operon system, in which transgene expression can be repressed and de-repressed by an innocuous lactose analogue, IPTG.

Mice deficient in heparanase exhibit impaired dendritic cell migration and reduced airway inflammation.

Heparanase is a β-d-endoglucuronidase that cleaves heparan sulphate, a key component of the ECM and basement membrane. The remodelling of the ECM by heparanase has been proposed to regulate both normal physiological and pathological processes, including wound healing, inflammation, tumour angiogenesis and cell migration. Heparanase is also known to exhibit non-enzymatic functions by regulating cell adhesion, cell signalling and differentiation.

Internal ribosome entry site-based attenuation of a flavivirus candidate vaccine and evaluation of the effect of beta interferon coexpression on vaccine properties.

Infectious clone technologies allow the rational design of live attenuated viral vaccines with the possibility of vaccine-driven coexpression of immunomodulatory molecules for additional vaccine safety and efficacy. The latter could lead to novel strategies for vaccine protection against infectious diseases where traditional approaches have failed. Here we show for the flavivirus Murray Valley encephalitis virus (MVEV) that incorporation of the internal ribosome entry site (IRES) of Encephalomyocarditis virus between the capsid and prM genes strongly attenuated virulence and that the resulting bicistronic virus was both genetically stable and potently immunogenic.

Teratoma formation in immunocompetent mice after syngeneic and allogeneic implantation of germline capable mouse embryonic stem cells.

Background: Embryonic stem cells (ESCs) have the potential to form teratomas when implanted into immunodeficient mice, but data in immunocompetent mice are limited. We therefore investigated teratoma formation after implantation of three different mouse ESC (mESC) lines into immunocompetent mice.

Materials And Methods: BALB/c mice were injected with three highly germline competent mESCs (129Sv, BALB/c and C57BL/6) subcutaneously or under the kidney capsule.

Molecular pathways involved in the improvement of non-alcoholic fatty liver disease.

Unlabelled: Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis are components of the metabolic syndrome. Serum leptin levels are elevated in obesity, but the role of leptin in the pathophysiology of the liver involvement is still unclear. To identify the effects and mechanisms by which leptin influences the pathogenesis of NAFLD, we performed epididymal white adipose tissue (eWAT) transplantation from congenic wild-type mice into the subcutaneous dorsal area of Lep(ob/ob) recipient mice and compared the results with those of the Lep(ob/ob) sham-operated mice.

Polymorphism in endothelial connexin40 enhances sensitivity to intraluminal pressure and increases arterial stiffness.

Objective: To determine whether impairment of endothelial connexin40 (Cx40), an effect that can occur in hypertension and aging, contributes to the arterial dysfunction and stiffening in these conditions.

Approach And Results: A new transgenic mouse strain, expressing a mutant Cx40, (Cx40T202S), specifically in the vascular endothelium, has been developed and characterized. This mutation produces nonfunctional hemichannels, whereas gap junctions containing the mutant are electrically, but not chemically, patent.

Spreading vasodilatation in the murine microcirculation: attenuation by oxidative stress-induced change in electromechanical coupling.

Regulation of blood flow in microcirculatory networks depends on spread of local vasodilatation to encompass upstream arteries; a process mediated by endothelial conduction of hyperpolarization. Given that endothelial coupling is reduced in hypertension, we used hypertensive Cx40ko mice, in which endothelial coupling is attenuated, to investigate the contribution of the renin-angiotensin system and reduced endothelial cell coupling to conducted vasodilatation of cremaster arterioles in vivo. When the endothelium was disrupted by light dye treatment, conducted vasodilatation, following ionophoresis of acetylcholine, was abolished beyond the site of endothelial damage.

The human parasite Loa loa in cytokine and cytokine receptor gene knock out BALB/c mice: survival, development and localization.

Background: Immunological mechanisms involved in the survival and development of human filarial species in the vertebrate host are poorly known due to the lack of suitable experimental models. In order to understand the role of cytokines in the survival and development of filarial larvae in the vertebrate host, we infected different strains of BALB/c mice deficient in a number of cytokine or cytokine receptor genes with Loa loa. The survival and development of larvae were monitored.

Gene dose influences cellular and calcium channel dysregulation in heterozygous and homozygous T4826I-RYR1 malignant hyperthermia-susceptible muscle.

Malignant hyperthermia susceptibility (MHS) is primarily conferred by mutations within ryanodine receptor type 1 (RYR1). Here we address how the MHS mutation T4826I within the S4-S5 linker influences excitation-contraction coupling and resting myoplasmic Ca(2+) concentration ([Ca(2+)](rest)) in flexor digitorum brevis (FDB) and vastus lateralis prepared from heterozygous (Het) and homozygous (Hom) T4826I-RYR1 knock-in mice (Yuen, B. T.

Mice expressing T4826I-RYR1 are viable but exhibit sex- and genotype-dependent susceptibility to malignant hyperthermia and muscle damage.

Mutation T4825I in the type 1 ryanodine receptor (RYR1(T4825I/+)) confers human malignant hyperthermia susceptibility (MHS). We report a knock-in mouse line that expresses the isogenetic mutation T4826I. Heterozygous RYR1(T4826I/+) (Het) or homozygous RYR1(T4826I/T4826I) (Hom) mice are fully viable under typical rearing conditions but exhibit genotype- and sex-dependent susceptibility to environmental conditions that trigger MH.

Glutathione transferase kappa deficiency causes glomerular nephropathy without overt oxidative stress.

Glutathione transferase kappa (GSTK1-1) is a highly conserved, mitochondrial enzyme potentially involved in redox reactions. GSTK1-1-deficient mice were generated to further study the enzyme's biological role. Reduced and total glutathione levels in liver and kidney were unchanged by GSTK1-1 deficiency and NADPH quinone oxidoreductase 1 expression was not elevated indicating that there is no general underlying oxidative stress in Gstk1(-/-) mice.

Mouse strains for the ubiquitous or conditional overexpression of the Flii gene.

The gelsolin related actin binding protein, Flii, is able to regulate wound healing; mice with decreased Flii expression show improved wound healing whereas mice with elevated Flii expression exhibit impaired wound healing. In both mice and humans Flii expression increases with age and amelioration of FLII activity represents a possible therapeutic strategy for improved wound healing in humans. Despite analysis of Flii function in a variety of organisms we know little of the molecular mechanisms underlying Flii action.

Plasmacytoid dendritic cells promote host defense against acute pneumovirus infection via the TLR7-MyD88-dependent signaling pathway.

Human respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infection in infants. In human infants, plasmacytoid dendritic cells (pDC) are recruited to the nasal compartment during infection and initiate host defense through the secretion of type I IFN, IL-12, and IL-6. However, RSV-infected pDC are refractory to TLR7-mediated activation.

Interleukin-13 (IL-13)/IL-13 receptor alpha1 (IL-13Ralpha1) signaling regulates intestinal epithelial cystic fibrosis transmembrane conductance regulator channel-dependent Cl- secretion.

Interleukin-13 (IL-13) has been linked to the pathogenesis of inflammatory diseases of the gastrointestinal tract. It is postulated that IL-13 drives inflammatory lesions through the modulation of both hematopoietic and nonhematopoietic cell function in the intestine. To delineate the relevant contribution of elevated levels of intestinal IL-13 to intestinal structure and function, we generated an intestinal IL-13 transgenic mouse (iIL-13Tg).

Evaluating vaccinia virus cytokine co-expression in TLR GKO mice.

Using Toll-like receptor (TLR) and MyD88 gene knock-out (GKO) mice the effect of TLRs and MyD88 on virus replication, interferon (IFN)-β production, natural killer (NK) cell and CD8T cell responses were assessed following ectromelia virus (ECTV) and recombinant vaccinia virus (rVV) infection. The capacity for rVVs encoding cytokines to restore immune function in MyD88(-/-) mice was clearly demonstrated. Results showed that TLR2(-/-), TLR4(-/-)and TLR7(-/-) mice survived ECTV infection whereas MyD88(-/-) and TLR9(-/-)mice, in contrast, were highly susceptible.

Expression of distinct RNAs from 3' untranslated regions.

The 3' untranslated regions (3'UTRs) of eukaryotic genes regulate mRNA stability, localization and translation. Here, we present evidence that large numbers of 3'UTRs in human, mouse and fly are also expressed separately from the associated protein-coding sequences to which they are normally linked, likely by post-transcriptional cleavage. Analysis of CAGE (capped analysis of gene expression), SAGE (serial analysis of gene expression) and cDNA libraries, as well as microarray expression profiles, demonstrate that the independent expression of 3'UTRs is a regulated and conserved genome-wide phenomenon.

Generation and characterization of mice with null mutation of the chloride intracellular channel 1 gene.

CLIC1 belongs to a family of highly conserved and widely expressed intracellular chloride ion channel proteins existing in both soluble and membrane integrated forms. To study the physiological and biological role of CLIC1 in vivo, we undertook conditional gene targeting to engineer Clic1 gene knock-out mice. This represents creation of the first gene knock-out of a vertebrate CLIC protein family member.

Phenylalanine-induced leucopenia in genetic and dichloroacetic acid generated deficiency of glutathione transferase Zeta.

Glutathione transferase Zeta (GSTZ1-1) is identical to maleylacetoacetate isomerase and catalyses a significant step in the catabolism of phenylalanine and tyrosine. Exposure of GSTZ1-1 deficient mice to high dietary phenylalanine causes a rapid loss of circulating white blood cells (WBCs). The loss was significant (P<0.

Identification of therapeutic drug targets through genetically manipulated mice: are we getting it right?

A significant contribution to the identification of therapeutic targets in disease control and management has been made through the use of genetically modified mice in which specific gene function was altered. However, the genetic background of these mice can have a strong influence on the observed phenotype and, if not properly controlled, can lead to misinterpretation of data and incorrect conclusions as to the importance of particular genes in disease states. Nevertheless, it is becoming apparent that advantages may also ensue, since analysis of gene function in mixed (or a number of different) genetic backgrounds may more closely approximate the human genome and provide the potential to identify modifier genes and polymorphisms which influence the resulting phenotype of the target gene.