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At least 451 records · Page 25Linked to original sources
PubMed1997

Effects of cholecystokinin-B receptor antagonist on dopamine system in tenascin mutant mice.

The aim of this study was to investigate the effect of cholecystokinin (CCK) receptor antagonist on the abnormal behavior and dopamine (DA) transmission of tenascin (TN)-gene knockout mice. Recently, we demonstrated that TN-gene deficient mice show hyperlocomotion that is related to reduced DA transmission and tyrosine hydroxylase (TH) activities in the brain. In this report, we show that the intraperitoneal administration of a CCK-B receptor antagonist, PD135158 (0.1 mg/kg), but not a CCK-A receptor antagonist, lorglumide, inhibited hyperlocomotion. Moreover, PD135158 reversed the low levels of DA turnover rate and TH activities in the striatum of TN-gene knockout mouse brain. These results suggest that CCK-B receptor is involved in the behavior of TN-gene knockout mouse through striatal DA transmission.

3,4-Dihydroxyphenylacetic Acid↗
PubMed2002

Mice lacking the gene for inducible or endothelial nitric oxide are resistant to sporocyst induced Sarcocystis neurona infections.

Equine protozoal myeloencephalitis (EPM) is a neurologic syndrome in horses from the Americas and is usually caused by infection with the apicomplexan parasite, Sarcocystis neurona. Little is known about the role of immunobiological mediators to this parasite. Nitric oxide (NO) is important in resistance to many intracellular parasites. We, therefore, investigated the role of inducible and endothelial NO in resistance to clinical disease caused by S. neurona in mice. Groups of interferon-gamma gene knockout (IFN-gamma-KO) mice, inducible nitric oxide synthase gene knockout (iNOS-KO) mice, endothelial nitric oxide synthase gene knockout (eNOS-KO) and appropriate genetic background mice (BALB/c or C57BL/6) were orally fed sporocysts or Hanks balanced salt solution. Mice were observed for signs of clinical disease and examined at necropsy. Clinical disease and deaths occurred only in the IFN-gamma-KO mice. Microscopic lesions were seen only in the brains of IFN-gamma-KO mice. Results of this study indicate that iNOS and eNOS are not major mediators of resistance to S. neurona infections. Results of this study suggest that IFN-gamma mediated immunity to S. neurona may be mediated by non-NO-dependent mechanisms.

Animals↗
PubMed2003

The in vivo role of alpha-mannosidase IIx and its role in processing of N-glycans in spermatogenesis.

The surfaces of mammalian cells are covered by a variety of carbohydrates linked to proteins and lipids. N-glycans are commonly found carbohydrates in plasma membrane proteins. The structure and biosynthetic pathway of N-glycans have been analyzed extensively. However, functional analysis of cell surface N-glycans is just under way with recent studies of targeted disruption of genes involved in N-glycan synthesis. This review briefly introduces the potential role of processing alpha-mannosidases in N-glycan biosynthesis and recent findings derived from the alpha-mannosidase IIx (MX) gene knockout mouse, which shows male infertility. Thus, the MX gene knockout experiment unveiled a novel function of specific N-glycan, which is N-acetylglucosamine-terminated and fucosylated triantennary structure, in the adhesion between germ cells and Sertoli cells. Analysis of the MX gene knockout mouse is a good example of a multidisciplinary approach leading to a novel discovery in the emerging field of glycobiology.

Animals↗
PubMed2005

Identifying gene targets for the metabolic engineering of lycopene biosynthesis in Escherichia coli.

The identification of genetic targets that are effective in bringing about a desired phenotype change is still an open problem. While random gene knockouts have yielded improved strains in certain cases, it is also important to seek the guidance of cell-wide stoichiometric constraints in identifying promising gene knockout targets. To investigate these issues, we undertook a genome-wide stoichiometric flux balance analysis as an aid in discovering putative genes impacting network properties and cellular phenotype. Specifically, we calculated metabolic fluxes such as to optimize growth and then scanned the genome for single and multiple gene knockouts that yield improved product yield while maintaining acceptable overall growth rate. For the particular case of lycopene biosynthesis in Escherichia coli, we identified such targets that we subsequently tested experimentally by constructing the corresponding single, double and triple gene knockouts. While such strains are suggested (by the stoichiometric calculations) to increase precursor availability, this beneficial effect may be further impacted by kinetic and regulatory effects not captured by the stoichiometric model. For the case of lycopene biosynthesis, the so identified knockout targets yielded a triple knockout construct that exhibited a nearly 40% increase over an engineered, high producing parental strain.

Carotenoids↗
PubMed2003

Combined analysis of oligonucleotide microarray data from transgenic and knockout mice identifies direct SREBP target genes.

The synthesis of fatty acids and cholesterol, the building blocks of membranes, is regulated by three membrane-bound transcription factors: sterol regulatory element-binding proteins (SREBP)-1a, -1c, and -2. Their function in liver has been characterized in transgenic mice that overexpress each SREBP isoform and in mice that lack all three nuclear SREBPs as a result of gene knockout of SREBP cleavage-activating protein (SCAP), a protein required for nuclear localization of SREBPs. Here, we use oligonucleotide arrays hybridized with RNA from livers of three lines of mice (transgenic for SREBP-1a, transgenic for SREBP-2, and knockout for SCAP) to identify genes that are likely to be direct targets of SREBPs in liver. A total of 1,003 genes showed statistically significant increased expression in livers of transgenic SREBP-1a mice, 505 increased in livers of transgenic SREBP-2 mice, and 343 showed decreased expression in Scap-/- livers. A subset of 33 genes met the stringent combinatorial criteria of induction in both SREBP transgenics and decreased expression in SCAP-deficient mice. Of these 33 genes, 13 were previously identified as direct targets of SREBP action. Of the remaining 20 genes, 13 encode enzymes or carrier proteins involved in cholesterol metabolism, 3 participate in fatty acid metabolism, and 4 have no known connection to lipid metabolism. Through application of stringent combinatorial criteria, the transgenic/knockout approach allows identification of genes whose activities are likely to be controlled directly by one family of transcription factors, in this case the SREBPs.

Animals↗
PubMed2005

Triple knockouts reveal gene interactions affecting fertility of male mice.

Triple knockout mice were used to investigate the interactions of five genes that were expressed in meiotic and haploid spermatogenic cells in mice, transition protein 2 (Tnp2), proacrosin (Acr), histone H1.1 (H1.1), histone H1t (H1t), and sperm mitochondria-associated cysteine-rich protein (Smcp). TNP2 functions in the replacement of histones and the initial condensation of the spermatid nucleus. The linker histone subtypes H1.1 and H1t are expressed at high levels in meiotic and early haploid cells. ACR, a protease that is stored as a proenzyme in the acrosome, is activated during the acrosome reaction and functions in binding of sperm to the zona pellucida. SMCP is a structural protein in the outer membranes of sperm mitochondria that functions in motility. Previous work demonstrates that homozygous knockout mice lacking each of these proteins individually exhibit no defect in fertility on mixed genetic backgrounds. In contrast, the present study demonstrates that five triple knockout lines, Acr/H1.1/Smcp, Acr/Tnp2/Smcp, Tnp2/H1.1/Smcp, Acr/H1t/Smcp, Tnp2/H1t/Smcp, exhibit drastic reductions in fertility on mixed genetic backgrounds. Analysis of fertility parameters reveal that the decreased fertility is due to line-dependent defects in sperm motility in vitro correlated with reduced migration in the female reproductive tract, and decreased fertilization due to defects in adhesion of sperm to the zona pellucida, the membrane surrounding the egg. It was also found that triple knockout males, that are hemizygous for one locus and homozygous for two other loci, are as subfertile as homozygous triple knockout males, a phenomenon known as haploinsufficiency. These findings demonstrate that male fertility involves synergistic interactions of genes that function in sperm motility and sperm-egg adhesion during fertilization.

Acrosome Reaction↗
PubMed2001

Enhanced formation of azoxymethane-induced colorectal adenocarcinoma in gammadelta T lymphocyte-deficient mice.

T cell receptor (TCR) gammadelta-positive T lymphocytes, which are localized mostly within the intraepithelial space of intestinal epithelium, have been suggested to play a role in maintaining the normal configuration of intestinal epithelium. However, the role of TCRgammadelta-positive T lymphocytes in the formation and progression of colorectal adenocarcinoma that originates from colorectal epithelial cells remains to be elucidated. In this study, TCRalphabeta and TCRgammadelta-positive T lymphocyte-deficient mice (homozygous TCRa and TCRd-gene knockout mice) and the background wild-type mice were administered azoxymethane, and the formation of macroscopic tumors and microscopic aberrant crypt foci in colorectal mucosa were compared among the three types of mice. Well-differentiated adenocarcinoma appeared 5 months after 5 administrations of azoxymethane (10 mg/kg weight) only in a few TCRd -gene knockout mice and the frequency of the carcinoma-bearing mice was increased at 7 and 9 months after the administration. Aberrant crypt foci were also detected in the colorectal mucosa of TCRd -gene knockout mice to a greater extent than in colorectal mucosa of TCRa-gene knockout mice 1 month after the azoxymethane administration. These results suggest that TCRgammadelta-positive T lymphocytes, which are present mainly in the intraepithelial space, play a role in suppression of the formation and progression of colorectal adenocarcinoma in mice.

Adenocarcinoma↗
PubMed2004

Intratracheal double-stranded RNA plus interferon-gamma: a model for analysis of the acute phase response to respiratory viral infections.

Double-stranded (ds)RNA is made as a by-product of viral replication. Synthetic dsRNA induces virtually all of the same systemic symptoms as acute viral infections, such as fever and malaise. In order to develop a model of respiratory viral infections (such as influenza) suitable for use in gene knockout mice (where the deleted gene may affect viral replication), we examined C57BL/6 mouse body temperature and locomotor activity responses to the synthetic dsRNA polyriboinosinic.polyribocytidylic acid (poly[rI.rC]) introduced via the intratracheal (IT) route. We compared the IT poly[rI.rC] responses to the well-characterized intraperitoneal (IP) poly[rI.rC] responses. IT poly[rI.rC] failed to induce an acute phase response (APR) in mice, in contrast to IP poly[rI.rC]. However, addition of interferon (IFN)gamma to the IT poly[rI.rC] inoculum induced sustained hypothermia and suppressed locomotor activity responses with similar kinetics to those responses seen in acute mouse influenza. We further examined cytokine, antiviral, muscarinic M2 receptor and inducible nitric oxide synthase gene expression at 5 hr in the lungs of IT challenged mice. These studies suggested that priming the lung with IFNgamma could enhance proinflammatory (IL1beta, IL6, TNFalpha) cytokine gene expression and suppress interferon gene expression compared to IT poly[rI.rC] alone. No differences were detected for the other genes examined. While further molecular characterization of the model is required, we demonstrate that IT challenge with combined poly[rI.rC] and IFNgamma closely simulates the APR to an acute respiratory virus, and may serve as a suitable model for analyzing the molecular basis of the viral APR in gene knockout mice.

Acute-Phase Reaction↗
PubMed2006

Selective cognitive dysfunction in mice lacking histamine H1 and H2 receptors.

Previous pharmacological experiments provide conflicting findings that describe both facilitatory and inhibitory effects of neuronal histamine on learning and memory. Here, we examined learning and memory and synaptic plasticity in mice with a null mutation of gene coding histamine H1 or H2 receptor in order to clarify the role of these receptors in learning and memory processes. Learning and memory were evaluated by several behavioral tasks including object recognition, Barnes maze and fear conditioning. These behavioral tasks are highly dependent on the function of prefrontal cortex, hippocampus or amygdala. Object recognition and Barnes maze performance were significantly impaired in both H1 receptor gene knockout (H1KO) and H2 receptor gene knockout (H2KO) mice when compared to the respective wild-type (WT) mice. Conversely, both H1KO and H2KO mice showed better auditory and contextual freezing acquisition than their respective WT mice. Furthermore, we also examined long-term potentiation (LTP) in the CA1 area of hippocampus in H1KO and H2KO mice and their respective WT mice. LTP in the CA1 area of hippocampus was significantly reduced in both H1KO and H2KO mice when compared with their respective WT mice. In conclusion, our results demonstrate that both H1 and H2 receptors are involved in learning and memory processes for which the frontal cortex, amygdala and hippocampus interact.

Analysis of Variance↗
PubMed2001

The psychopharmacology-molecular biology interface: exploring the behavioural roles of dopamine receptor subtypes using targeted gene deletion ('knockout').

In the absence of selective agonists and antagonists able to discriminate between individual members of the D1-like and D2-like families of dopamine receptor subtypes, functional parcellation has remained problematic. 'Knockout' of these subtypes by targeted gene deletion offers a new approach to evaluating their roles in the regulation of behaviour. Like any new technique, 'knockout' has associated with it a number of methodological limitations that are now being addressed in a systematic manner. Studies on the phenotype of D1(A/1), D(1B/5), D2, D3 and D4 'knockouts' at the level of spontaneous and agonist/antagonist-induced behaviour are reviewed, in terms of methodological issues, neuronal implications and potential clinical relevance. Dopamine receptor subtype 'knockout' is a nascent technology that is now beginning to fulfil its potential. It is being complemented by more systematic phenotypic characterisation at the level of behaviour and additional, molecular biologically-based approaches.

Animals↗
PubMed2004

Endothelial nitric oxide synthase in the control of osteoblastic mineralizing activity and bone integrity.

It has been shown previously that osteoblast differentiation and maintenance of bone mass are impaired in endothelial nitric oxide synthase gene knockout mice. The present study shows by analysis of messenger RNA expression that the transcription factor Cbfa-1/Runx-2 and the bone matrix protein osteocalcin, which are fundamental to osteoblast differentiation, are significantly reduced in neonatal calvarial osteoblasts from these gene knockout mice. Expression of these genes could be restored to wild-type levels by exogenous supply of the photoactivatable nitric oxide donor potassium nitrosylpentachlorouthenate, but this was dependent on the timing of its activation and recovery in gene expression was only evident during the latter stages of osteoblast differentiation associated with its mineralizing activity. Calvarial, femoral/pelvic, spinal, and total bone mineral density, together with bone microhardness and expression of osteocalcin in whole femurs, were all reduced significantly in gene knockout mice at 8 weeks of age, but not at 12 weeks, where all of these indices of bone integrity were comparable to wild type. In accordance with these temporal effects, reduced bone mineral density, bone microhardness, and osteocalcin expression could be restored to normal, wild-type values after 21 days in vivo administration of the nitric oxide donor glyceryl trinitrate to 4-week-old endothelial nitric oxide synthase knockout mice, but there was no significant effect in older animals. Taken together, these results further demonstrate the importance of endothelial nitric oxide synthase in the regulation of osteoblast metabolism. In particular, they show that nitric oxide is involved in co-ordinating specific phases of osteoblast differentiation and bone formation: this could be relevant to its therapeutic actions on bone turnover.

Animals↗
PubMed1997

[Catecholamine and dopamine].

Almost all the genes of the enzymes which synthesize and metabolize the catecholamines (dopamine, norepinephrine, epinephrine) have been cloned and the gene targeting technology have been applied to introduce the gene knockout mouse such as thyrosine hydroxylase and dopamine beta hydroxylase. At least nine adrenergic receptors and five dopamine receptors have been cloned, which include alpha 1A-, alpha 1 B-, alpha 1 D-, alpha 2 A-, alpha 2B-, alpha 2C-, beta 1-, beta 2-, beta 3-adrenergic receptors and D1-, D2-, D3-, D4-, D5-dopamine receptors. Transgenic mouse as well as gene knockout mouse of these genes have been also produced. Furthermore, intracellular signal transduction systems of the catecholamines have been clarified using molecular techniques, including nine subtypes of adenylyl cyclase. Using these cloned genes and transgenic and gene knockout mouse, more detailed features of the catecholamine systems and those receptors and intracellular signal transduction systems will be clarified in near future.

Adenylyl Cyclases↗
PubMed1998

Developmental biology at the Howard Florey Institute.

1. A brief account of the history of developmental biology at the Howard Florey Institute, as well as in a wider general context, is presented and some of the results of developmental studies performed at the Institute over the past 7 years are discussed. 2. In the foetal rat, parathyroid hormone rather than parathyroid hormone-related protein is produced by the parathyroid anlage. This contrasts with results reported by others for the sheep. 3. The control of imprinting of H19 and insulin-like growth factor-II has been studied in the mouse using gene knockout of the coding region of the H19 gene. Bi-allelic expression of H19 in some liver and gut cells was observed. 4. Gene knockout of the homeobox gene Cdx2 produced dramatic effects. Homozygous embryos did not implant in the uterus. Heterozygotes exhibited homeosis involving the gut and skeleton. Animals developed intestinal tumours within a few weeks of birth.

Academies and Institutes↗
PubMed2002

Maternal behavior and developmental psychopathology.

This paper reviews recent developments in the phenomenology, neurobiology, and genetics of maternal behavior in animal model systems from an evolutionary perspective on psychopathology. Following a review of the phenomenology and neurobiology of maternal behavior, recent studies addressing the role of genetic factors in the maternal behavior of rodents were identified in a search of literature in peer-reviewed journals. Gene knockout studies were evaluated with regard to mouse strain background, method of behavioral phenotyping, and quantification of the behavioral deficits. Gene knockout data were then analyzed using a cluster analysis technique. At least nine genes have been identified that are necessary for the expression of one or more aspects of maternal behavior. These genes encode for three transcription factors: three enzymes, including dopamine beta hydroxylase and neuronal nitric oxide synthase; two receptors, including the prolactin and the estrogen alpha receptor; and one neuropeptide, oxytocin. Cluster analysis suggested possible relationships between specific genes. Gene knockout technology has provided new insights into the molecular basis of maternal behavior that are congruent with the existing neurobiological literature. Future studies of genetic and environmental influences on maternal behavior have the potential to inform models of disease pathogenesis.

Animals↗
PubMed2003

HFE mutation and dietary iron content interact to increase ischemia/reperfusion injury of the heart in mice.

Hereditary hemochromatosis is an inherited pathological condition characterized by iron overload in several vital organs including heart. To increase our understanding of the underlying pathogenic mechanisms of hereditary hemochromatosis, we used a HFE gene knockout mouse model that replicates hereditary hemochromatosis. A group of mice with no copies of HFE gene and corresponding wild-type mice were maintained either on low-iron (30 ppm) or high-iron (300 ppm) diet since birth. The results of our study revealed that HFE gene knockout mouse hearts were susceptible to ischemia-reperfusion injury as evidenced by increased postischemic ventricular dysfunction, increased myocardial infarct size and cardiomyocyte apoptosis compared with wild-type control hearts. The degree of injury increased in the hearts of the mice fed high-iron diet. The hearts of the HFE knockout mice showed increased iron deposition, increased content of reactive oxygen species (ROS) as evidenced by the increased formation of malondialdehyde, and reduced antioxidant enzymes including superoxide dismutase, catalase, and glutathione peroxidase. The results suggest that increased amount of ROS and reduced antioxidant reserve secondary to iron overloading may be instrumental for the susceptibility of the HFE gene knockout mice to cardiac injury.

Administration, Oral↗
PubMed2002

The mifepristone-inducible gene regulatory system in mouse models of disease and gene therapy.

The mifepristone (Mfp)-inducible gene regulatory system is designed to allow control of the spatiotemporal expression of transgenes in vivo in a ligand-dependent manner. This regulatory system is composed of two components: (1) a chimeric transactivator protein that activates transgene transcription only in the presence of the progesterone antagonist Mfp, and (2) a target transgene placed in the context of a promoter which is responsive only to the Mfp-bound chimeric transactivator. Incorporation of the components of the Mfp-inducible gene regulatory system into transgenic mice has resulted in the establishment of several novel, Mfp-dependent models of disease. Similarly, adaptation of the Mfp-inducible system for use in gene knockout models has resulted in the development of new gene ablation technology which is both tissue-specific and Mfp-dependent. Additionally, the Mfp-inducible gene regulatory system has been used in animal experiments involving somatic gene therapy, where it has shown considerable promise in the regulation of both reporter and therapeutic gene expression. This review focuses on recent application of the Mfp-inducible system to transgenic models, gene knockout models, and somatic gene therapy experiments. In so doing, it demonstrates the considerable promise that future use of this system holds for better understanding and treatment of human disease.

Animals↗
PubMed2004

Transposon mutagenesis of the obligate intracellular pathogen Rickettsia prowazekii.

Genetic analysis of Rickettsia prowazekii has been hindered by the lack of selectable markers and efficient mechanisms for generating rickettsial gene knockouts. We have addressed these problems by adapting a gene that codes for rifampin resistance for expression in R. prowazekii and by incorporating this selection into a transposon mutagenesis system suitable for generating rickettsial gene knockouts. The arr-2 gene codes for an enzyme that ADP-ribosylates rifampin, thereby destroying its antibacterial activity. Based on the published sequence, this gene was synthesized by PCR with overlapping primers that contained rickettsial codon usage base changes. This R. prowazekii-adapted arr-2 gene (Rparr-2) was placed downstream of the strong rickettsial rpsL promoter (rpsL(P)), and the entire construct was inserted into the Epicentre EZ::TN transposome system. A purified transposon containing rpsL(P)-Rparr-2 was combined with transposase, and the resulting DNA-protein complex (transposome) was electroporated into competent rickettsiae. Following selection with rifampin, rickettsiae with transposon insertions in the genome were identified by PCR and Southern blotting and the insertion sites were determined by rescue cloning and inverse PCR. Multiple insertions into widely spaced areas of the R. prowazekii genome were identified. Three insertions were identified within gene coding sequences. Transposomes provide a mechanism for generating random insertional mutations in R. prowazekii, thereby identifying nonessential rickettsial genes.

Animals↗
PubMed2001

Knockout of renin-angiotensin system genes: effects on vascular development.

Pharmacologic inhibition of the renin-angiotensin system (RAS) is a widely accepted and effective treatment for hypertension. However, in the past several years, much attention has been focused on additional roles of the RAS including the possibility that its end-product, angiotensin II, could elicit end-organ pathologies independent of its effect on blood pressure. The ability to selectively delete genes in mice (by homologous recombination or gene knockouts) has led to new--and sometimes surprising--insights into the roles of the RAS in the developmental modeling and pathologic remodeling of the heart and blood vessels.

Angiotensinogen↗