Science.gov

Sample records for e6 transgenic mice

  1. Tumor prevention in HPV8 transgenic mice by HPV8-E6 DNA vaccination.

    PubMed

    Marcuzzi, Gian Paolo; Awerkiew, Sabine; Hufbauer, Martin; Schädlich, Lysann; Gissmann, Lutz; Eming, Sabine; Pfister, Herbert

    2014-06-01

    The genus beta human papillomavirus 8 (HPV8) is involved in the development of cutaneous squamous cell carcinomas (SCCs) in individuals with epidermodysplasia verruciformis. Immunosuppressed transplant recipients are prone to harbor particularly high betapapillomavirus DNA loads, which may contribute to their highly increased risk of SCC. Tumor induction in HPV8 transgenic mice correlates with increased expression of viral oncogenes E6 and E2. In an attempt to prevent skin tumor development, we evaluated an HPV8-E6-DNA vaccine, which was able to stimulate a detectable HPV8-E6-specific cell-mediated immune response in 8/15 immunized mice. When skin of HPV8 transgenic mice was grafted onto non-transgenic littermates, the grafted HPV8 transgenic tissue was not rejected and papillomas started to grow within 14 days all over the transplant of 9/9 non-vaccinated and 7/15 not successfully vaccinated mice. In contrast, no papillomas developed in 6/8 successfully vaccinated mice. In the other two of these eight mice, a large ulcerative lesion developed within the initial papilloma growth or papilloma development was highly delayed. As the vaccine completely or partially prevented papilloma development without rejecting the transplanted HPV8 positive skin, the immune system appears to attack only keratinocytes with increased levels of E6 protein, which would give rise to papillomas. PMID:24446083

  2. Increased Incidence of Squamous Cell Carcinomas in Mastomys natalensis Papillomavirus E6 Transgenic Mice during Two-Stage Skin Carcinogenesis

    PubMed Central

    Helfrich, Iris; Chen, Min; Schmidt, Rainer; Fürstenberger, Gerhard; Kopp-Schneider, Annette; Trick, David; Gröne, Hermann-Josef; zur Hausen, Harald; Rösl, Frank

    2004-01-01

    Papillomaviruses cause certain forms of human cancers, most notably carcinomas of the uterine cervix. In contrast to the well-established involvement of papillomavirus infection in the etiology of cervical carcinomas and in carcinomas of a rare hereditary condition, epidermodysplasia verruciformis, a causative role for cutaneous human papillomavirus types in the development of nonmelanoma skin cancer has not been proven. In order to better understand the functions of individual genes of cutaneous papillomavirus types, we generated transgenic mice carrying oncogene E6 of the Mastomys natalensis papillomavirus (MnPV), which causes keratoacanthomas of the skin in its natural host. In the present study, we demonstrate that under conditions of experimental two-stage skin carcinogenesis, fast-paced squamous cell carcinomas develop in nearly 100% of MnPV E6 transgenic mice in comparison to 10% in their nontransgenic littermates (log rank test; P < 0.0001). Therefore, we conclude that the MnPV E6 transgene favors the malignant progression of chemically induced tumors. Whereas an activating H-ras mutation is a consistent feature in benign and malignant tumors in wild-type mice, the majority of papillomas and keratoacanthomas and all squamous cell carcinomas obtained in MnPV E6 transgenic mice contain nonmutated ras alleles. These results indicate that the development of squamous cell carcinomas in MnPV E6 transgenic mice does not depend on an activated H-ras oncogene. PMID:15078961

  3. Tumorigenicity by human papillomavirus type 16 E6 and E7 in transgenic mice correlates with alterations in epithelial cell growth and differentiation.

    PubMed Central

    Griep, A E; Herber, R; Jeon, S; Lohse, J K; Dubielzig, R R; Lambert, P F

    1993-01-01

    The human papillomavirus type 16 (HPV-16) E6 and E7 oncogenes are thought to play a role in the development of most human cervical cancers. These E6 and E7 oncoproteins affect cell growth control at least in part through their association with and inactivation of the cellular tumor suppressor gene products, p53 and Rb. To study the biological activities of the HPV-16 E6 and E7 genes in epithelial cells in vivo, transgenic mice were generated in which expression of E6 and E7 was targeted to the ocular lens. Expression of the transgenes correlated with bilateral microphthalmia and cataracts (100% penetrance) resulting from an efficient impairment of lens fiber cell differentiation and coincident induction of cell proliferation. Lens tumors formed in 40% of adult mice from the mouse lineage with the highest level of E6 and E7 expression. Additionally, when lens cells from neonatal transgenic animals were placed in tissue culture, immortalized cell populations grew out and acquired a tumorigenic phenotype with continuous passage. These observations indicate that genetic changes in addition to the transgenes are likely necessary for tumor formation. These transgenic mice and cell lines provide the basis for further studies into the mechanism of action of E6 and E7 in eliciting the observed pathology and into the genetic alterations required for HPV-16-associated tumor progression. Images PMID:8382301

  4. Targeted expression of the E6 and E7 oncogenes of human papillomavirus type 16 in the epidermis of transgenic mice elicits generalized epidermal hyperplasia involving autocrine factors.

    PubMed Central

    Auewarakul, P; Gissmann, L; Cid-Arregui, A

    1994-01-01

    The E6 and E7 early genes of human papillomavirus type 16 have been shown in vitro to play a central role in the transforming capability of this virus. To explore their effects on differentiating epithelial cells in vivo, we used a bovine cytokeratin 10 (K10) promoter to target the expression of E6 and E7 to the suprabasal layers of the epidermis of transgenic mice. In two different lines of mice efficiently expressing the transgene, animals displayed generalized epidermal hyperplasia, hyperkeratosis and parakeratosis in the skin and the forestomach, both known to be sites of K10 expression. Northern (RNA) blot analysis revealed high levels of E6 and E7 transcripts, and in situ hybridizations localized these transcripts to the suprabasal strata of epidermis. In vivo labeling of proliferating cells showed two distinct effects of E6 and E7 expression in the epidermis: (i) an increase in the number of growing cells in the undifferentiated basal layer and (ii) abnormal proliferation of differentiated cells in the suprabasal strata. The expression of c-myc in the skin of transgenics was higher than that in control animals. The induction of c-myc transcription by topical application of tetradecanoyl phorbol acetate was prevented by simultaneous treatment with transforming growth factor beta 1 in nontransgenic skin but not in transgenic skin. In addition, transforming growth factor alpha was found to be overexpressed in the suprabasal layers of the transgenic epidermis. These findings suggest that autocrine mechanisms are involved in the development and maintenance of epidermal hyperplasia. Animals of both lines developed papillomas in skin sites exposed to mechanical irritation and wounding, suggesting that secondary events are necessary for progression to neoplasia. Collectively, these results provide new insights into the tumor promoter activities of human papillomavirus type 16 in epithelial cells in vivo. Images PMID:7969162

  5. Generation of Transgenic Mice

    PubMed Central

    Cho, Andrew; Haruyama, Naoto; Kulkarni, Ashok B.

    2009-01-01

    This unit describes detailed step-by-step protocols, reagents, and equipment required for successful generation of transgenic mice using pronuclear injection. The experimental methods and practical tips given here will help guide beginners in understanding what is required and what to avoid in these standard protocols for efficiently generating transgenic mice. PMID:19283729

  6. [Oncogenesis in transgenic mice].

    PubMed

    Shvemberger, I N; Ermilov, A N

    1994-01-01

    Oncogenesis in transgenic mice is at present a model, most adequately reflecting the natural conditions of tumor development. One of more important traits of this model is that it allows to study malignant growth simultaneously at all the structure-function levels in the context of the whole organism. This paper is a review of results of a series of experiments in which the localization of tumors was dependent or independent on the tissue specificity of a promoter, as well as development of multiple tumors with the use of viral regulatory sequences in genetic constructions. It has been shown that although a transgene is expressed in most of the tissues, tumors develop in some particular tissues only. These observations are interpreted by some authors in favour of the concept of multistep cancerogenesis. In this view, of primary importance are the results of studies on oncogenesis in transgenic mice, which contradict this concept and are regarded by their authors as an evidence of the possibility of a one-step transformation of normal cell into malignant one. The analysis of the obtained material enabled us to put forward an assumption that the key role in oncogenesis is played not only by certain genetic disturbances, but also by multi-level homeostatic mechanisms. Apparently, it is just the transgenic mice with cellular or viral oncogenes in their genome that represent a more adequate model for the detection of certain molecular-biological mechanisms underlying these disturbances. Also, of much importance is abundant material accumulated by now on oncogenesis of transgenic mice which shows a possibility of the effective use of various genetic constructions with prokaryotic and eukaryotic regulatory sequences, a possibility to induce not only tumors of some particular tissues, but also multiple hyperplastic and neoplastic changes in one and the same mouse. Development of tumors in such transgenic mice can be regarded as a model of different types of cancer disease

  7. A human papillomavirus type 18 E6/E7 transgene sensitizes mouse lens cells to human wild-type p53-mediated apoptosis.

    PubMed

    Nakamura, T; Williams-Simons, L; Westphal, H

    1997-06-26

    We have studied the concerted action of factors that influence the balance between cell proliferation and cell death in the developing lens of transgenic mice. We show that a human papillomavirus type 18 (HPV18) E6/E7 transgene that predominantly expresses the viral E7 gene product triggers apoptosis in a dose dependent manner, and causes retardation of lens growth or microphakia. E7 is known to inactivate pRB, the product of the retinoblastoma gene, and to enhance the action of p53. Our earlier work had demonstrated that over-expression of p53 itself can cause apoptosis of lens cells, and that a mutant p53 allele can interfere with this process. In the present study, we examined lenses that simultaneously express different constellations of the HPV18 E6/E7, wild-type and mutant human p53, and wild-type human pRB transgenes. We observed that lens cells expressing the HPV18 transgene are more sensitive to wild-type human p53 action than normal lens cells. As a result, there is severe microphakia in lenses that express both the HPV18 and the wild-type p53 transgenes. By contrast, apoptosis was reduced in lenses that co-expressed the HPV18 and either the pRB or the mutant p53 transgene. We conclude that levels of wild-type p53 are critical, and that any excess of p53 or suppression of pRB can cause cell death. Our results encourage attempts to counteract the deleterious action of human papillomaviruses in cervical cancer by a combination of measures that decrease cell proliferation and enhance apoptosis. PMID:9223662

  8. Overview: Engineering transgenic constructs and mice

    PubMed Central

    Haruyama, Naoto; Cho, Andrew; Kulkarni, Ashok B

    2009-01-01

    Cell biology research encompasses everything from single cells to whole animals. Recent discoveries concerning particular gene functions can be applied to the whole animal for understanding genotype-phenotype relationships underlying disease mechanisms. For this reason, genetically manipulated mouse models are now considered essential to correctly understand disease processes in whole animals. This unit provides the basic mouse technologies used to generate conventional transgenic mice, which represents gain-of-function approach. First, an overview of the transgenic construct design is presented. This unit then explains basic strategies for the identification and establishment of independent transgenic mouse lines, followed by comments on historical and emerging techniques, and then on typical problems that are encountered when researchers start to generate transgenic mice. PMID:19283728

  9. Studies of an expanded trinucleotide repeat in transgenic mice

    SciTech Connect

    Bingham, P.; Wang, S.; Merry, D.

    1994-09-01

    Spinal and bulbar muscular atrophy (SBMA) is a progressive motor neuron disease caused by expansion of a trinucleotide repeat in the androgen receptor gene (AR{sup exp}). AR{sup exp} repeats expand further or contract in approximately 25% of transmissions. Analogous {open_quotes}dynamic mutations{close_quotes} have been reported in other expanded trinucleotide repeat disorders. We have been developing a mouse model of this disease using a transgenic approach. Expression of the SBMA AR was documented in transgenic mice with an inducible promoter. No phenotypic effects of transgene expression were observed. We have extended our previous results on stability of the expanded trinucleotide repeat in transgenic mice in two lines carrying AR{sup exp}. Tail DNA was amplified by PCR using primers spanning the repeat on 60 AR{sup exp} transgenic mice from four different transgenic lines. Migration of the PCR product through an acrylamide gel showed no change of the 45 CAG repeat length in any progeny. Similarly, PCR products from 23 normal repeat transgenics showed no change from the repeat length of the original construct. Unlike the disease allele in humans, the expanded repeat AR cDNA in transgenic mice showed no change in repeat length with transmission. The relative stability of CAG repeats seen in the transgenic mice may indicate either differences in the fidelity of replicative enzymes, or differences in error identification and repair between mice and humans. Integration site or structural properties of the transgene itself might also play a role.

  10. Human prion strain selection in transgenic mice

    PubMed Central

    Giles, Kurt; Glidden, David V.; Patel, Smita; Korth, Carsten; Groth, Darlene; Lemus, Azucena; DeArmond, Stephen J.; Prusiner, Stanley B.

    2010-01-01

    Transgenic (Tg) mice expressing chimeras of mouse and human prion proteins (PrP) have shorter incubation periods for Creutzfeldt-Jakob disease (CJD) prions than mice expressing full-length human PrP. Increasing the sequence similarity of the chimeric PrP to mouse PrP, by reverting human residues to mouse, resulted in a Tg line, denoted Tg22372, which was susceptible to sporadic (s) CJD prions in ~110 days 1. Reversion of one additional residue (M111V) resulted in a new Tg line, termed Tg1014, susceptible to sCJD prions in ~75 days. Tg1014 mice also has shorter incubation periods for variant (v) CJD prions, providing a more tractable model for studying this prion strain. Transmission of vCJD prions to Tg1014 mice resulted in two different strains, determined by neuropathology and biochemical analysis, which correlated with the length of the incubation time. One strain had the biochemical, neuropathological, and transmission characteristics including longer incubation times of the inoculated vCJD strain; the second strain produced a phenotype resembling that of sCJD prions including relatively shorter incubation periods. Mice with intermediate incubation periods for vCJD prions had a mixture of the two strains. Both strains were serially transmitted in Tg1014 mice, which led to further reduction in incubation periods. Conversion of vCJD-like to sCJD-like strains was favored in Tg1014 mice more than in the Tg22372 line. The single amino acid difference therefore appears to offer selective pressure for propagation of the sCJD-like strain. These two Tg mouse lines provide relatively rapid models to study human prion diseases as well as the evolution of human prion strains. PMID:20695008

  11. Hepatic steatosis in transgenic mice overexpressing human histone deacetylase 1

    SciTech Connect

    Wang, Ai-Guo; Seo, Sang-Beom; Moon, Hyung-Bae; Shin, Hye-Jun; Kim, Dong Hoon; Kim, Jin-Man; Lee, Tae-Hoon; Kwon, Ho Jeong; Yu, Dae-Yeul . E-mail: dyyu10@kribb.re.kr; Lee, Dong-Seok . E-mail: lee10@kribb.re.kr

    2005-05-06

    It is generally thought that histone deacetylases (HDACs) play important roles in the transcriptional regulation of genes. However, little information is available concerning the specific functions of individual HDACs in disease states. In this study, two transgenic mice lines were established which harbored the human HDAC1 gene. Overexpressed HDAC1 was detected in the nuclei of transgenic liver cells, and HDAC1 enzymatic activity was significantly higher in the transgenic mice than in control littermates. The HDAC1 transgenic mice exhibited a high incidence of hepatic steatosis and nuclear pleomorphism. Molecular studies showed that HDAC1 may contribute to nuclear pleomorphism through the p53/p21 signaling pathway.

  12. Effect of transgene number of spontaneous and radiation-induced micronuclei in lacl transgenic mice

    SciTech Connect

    O`Loughlin, K.G.; Hamer, J.D.; Winegar, R.A.; Mirsalis, J.C.; Short, J.M.

    1994-12-31

    Lacl transgenic mice are widely used for the measurement of mutations in specific target issues. The lacl transgene is present in mice as 40 tandem repeats; this sequence is homozygous (contained in both copies of chromosome 5) in C57Bl/6 mice, and is hemizygous in B6C3F1 mice. Previous reports have indicated that tandem repeats can produce chromosome instability, fragile sites, and other effects. To determine whether the presence of the transgene effects micronucleus induction we compared the response of nontransgenic (NTR) to hemizygous (HEMI) transgenic B6C3F1 mice and to hemizygous and homozygous (HOMO) transgenic C57Bl/6 mice. Five mice/group were irradiated with 500 cGy from a {sup 137}Cs source. Bone marrow was harvested 24 hr after treatment and 2000 polychromatic erythrocytes (PCE) were analyzed per animal. The presence or absence of the lacl transgene had no effect in unirradiated mice on the percent of micronucleated PCE (MN) or on the ratio of PCE to total red blood cells for either strain: B6C3F1 mice had MN frequencies of 0.26% and 0.20% for NTR and HEMI mice, respectively; C57Bl/6 mice had MN frequencies of 0.34%, 0.32%, and 0.38% for NTR, HEMI, and HOMO mice, respectively. Radiation-induced micronucleus frequencies were significantly higher in HEMI lacl B6C3F1 mice (2.85%) than in NTR litter mates (1.59%); the converse was true in C57Bl/6 mice: NTR were 2.45%, HEMI were 1.25%, HOMO were 1.65%. These data suggest that the lacl transgene does not cause chromosome instability as measured by spontaneous micronucleus levels. However, the response of these transgenic mice to a variety of clastogenic agents needs to be investigated before they are integrated into standard in vivo assays for chromosome damage.

  13. Efficient Generation of Mice with Consistent Transgene Expression by FEEST.

    PubMed

    Gao, Lei; Jiang, Yonghua; Mu, Libing; Liu, Yanbin; Wang, Fengchao; Wang, Peng; Zhang, Aiqun; Tang, Nan; Chen, Ting; Luo, Minmin; Yu, Lei; Gao, Shaorong; Chen, Liang

    2015-01-01

    Transgenic mouse models are widely used in biomedical research; however, current techniques for producing transgenic mice are limited due to the unpredictable nature of transgene expression. Here, we report a novel, highly efficient technique for the generation of transgenic mice with single-copy integration of the transgene and guaranteed expression of the gene-of-interest (GOI). We refer to this technique as functionally enriched ES cell transgenics, or FEEST. ES cells harboring an inducible Cre gene enabled the efficient selection of transgenic ES cell clones using hygromycin before Cre-mediated recombination. Expression of the GOI was confirmed by assaying for the GFP after Cre recombination. As a proof-of-principle, we produced a transgenic mouse line containing Cre-activatable tTA (cl-tTA6). This tTA mouse model was able to induce tumor formation when crossed with a transgenic mouse line containing a doxycycline-inducible oncogene. We also showed that the cl-tTA6 mouse is a valuable tool for faithfully recapitulating the clinical course of tumor development. We showed that FEEST can be easily adapted for other genes by preparing a transgenic mouse model of conditionally activatable EGFR L858R. Thus, FEEST is a technique with the potential to generate transgenic mouse models at a genome-wide scale. PMID:26573149

  14. Enhanced human papillomavirus type 8 oncogene expression levels are crucial for skin tumorigenesis in transgenic mice

    SciTech Connect

    Hufbauer, M.; Lazic, D.; Akguel, B.; Brandsma, J.L.; Pfister, H.; Weissenborn, S.J.

    2010-08-01

    Human papillomavirus 8 (HPV8) is involved in skin cancer development in epidermodysplasia verruciformis patients. Transgenic mice expressing HPV8 early genes (HPV8-CER) developed papillomas, dysplasias and squamous cell carcinomas. UVA/B-irradiation and mechanical wounding of HPV8-CER mouse skin led to prompt papilloma induction in about 3 weeks. The aim of this study was to analyze the kinetics and level of transgene expression in response to skin irritations. Transgene expression was already enhanced 1 to 2 days after UVA/B-irradiation or tape-stripping and maintained during papilloma development. The enhanced transgene expression could be assigned to UVB and not to UVA. Papilloma development was thus always paralleled by an increased transgene expression irrespective of the type of skin irritation. A knock-down of E6 mRNA by tattooing HPV8-E6-specific siRNA led to a delay and a lower incidence of papilloma development. This indicates that the early increase of viral oncogene expression is crucial for induction of papillomatosis.

  15. Overexpression of mouse follistatin causes reproductive defects in transgenic mice.

    PubMed

    Guo, Q; Kumar, T R; Woodruff, T; Hadsell, L A; DeMayo, F J; Matzuk, M M

    1998-01-01

    Follistatin is an activin-binding protein that can act as an activin antagonist in vitro. Follistatin also binds heparin sulfate proteoglycans and may function as a reservoir for activins in vivo. In the mouse, follistatin mRNA is first detected in the deciduum on embryonic day 5.5 and later in the developing hindbrain, somites, vibrissae, teeth, epidermis, and muscle. We have previously shown that follistatin-deficient mice have numerous embryonic defects including shiny, taut skin, growth retardation, and cleft palate leading to death within hours of birth. To further define the roles of follistatin during mammalian reproduction and development, we created gain-of-function mutant mice in which mouse follistatin is overexpressed. The mouse metallothionein (MT)-I promoter was placed upstream of the six-exon mouse follistatin (FS) gene. To distinguish wild-type and transgenic follistatin mRNA, the 3'-untranslated region of the mouse follistatin gene was replaced with the SV40 untranslated and polyA sequences. Three male and two female founder transgenic mice were produced, were fertile, and transmitted the transgene to offspring. Northern blot analysis demonstrated that the transgene mRNA was expressed at varying levels in the livers of offspring from four of five of the transgenic lines and was expressed in the testes in all five lines. In MT-FS line 4, which had the highest expression of the transgene mRNA in the liver, the transgene transcripts were also present in multiple other tissues. Phenotypically, the MT-FS transgenic lines had defects in the testis, ovary, and hair. Mice from MT-FS lines 7 and 10 had slightly decreased testis size, whereas mice from lines 4, 5, and 9 had much smaller testes and shiny, somewhat irregular, fur. Histological analysis of the adult testes from line 5 and 9 males showed variable degrees of Leydig cell hyperplasia, an arrest of spermatogenesis, and seminiferous tubular degeneration leading to infertility. Female transgenic mice

  16. Non-invasive instant genotyping of fluorescently labelled transgenic mice.

    PubMed

    Fink, Dieter; Yau, Tien Yin; Kolbe, Thomas; Rülicke, Thomas

    2015-01-01

    Fluorescence proteins have been useful as genetic reporters for a wide range of applications in biomedical research and are frequently used for the analysis of transgene activity. Here, we show that expression levels of the ubiquitously expressed fluorescent proteins eGFP, mCherry, and tdTomato can be measured in transgenic mouse lines with random or targeted integrations. We identified the tail of the mouse as the tissue best suited for quantifying fluorescence intensity and show that expression levels in the tail correlate with gene dose. This allows for instant non-invasive determination of the genetic condition at the transgenic locus (hemizygous/heterozygous and homozygous), while simultaneously providing an objective comparison for transgene expression levels among different mouse lines. In summary, we demonstrate for the first time that the gene dose of a ubiquitously expressed fluorescence reporter can be reliably quantified and directly linked to the genotype of transgenic mice. Based on this information, animals with the appropriate genotype can be instantly selected without laborious analysis for establishing and breeding of new transgenic lines, reducing the number of "waste" animals. Furthermore, no tissue sampling is necessary, which is a significant refinement of genotyping procedures. Both aspects are important improvements for the genotyping of transgenic mice that follow the principles of the 3 Rs (reduction and refinement). PMID:25981046

  17. Severe iron deficiency anemia in transgenic mice expressing liver hepcidin.

    PubMed

    Nicolas, Gaël; Bennoun, Myriam; Porteu, Arlette; Mativet, Sandrine; Beaumont, Carole; Grandchamp, Bernard; Sirito, Mario; Sawadogo, Michèle; Kahn, Axel; Vaulont, Sophie

    2002-04-01

    We recently reported the hemochromatosis-like phenotype observed in our Usf2 knockout mice. In these mice, as in murine models of hemochromatosis and patients with hereditary hemochromatosis, iron accumulates in parenchymal cells (in particular, liver and pancreas), whereas the reticuloendothelial system is spared from this iron loading. We suggested that this phenotypic trait could be attributed to the absence, in the Usf2 knockout mice, of a secreted liver-specific peptide, hepcidin. We conjectured that the reverse situation, namely overexpression of hepcidin, might result in phenotypic traits of iron deficiency. This question was addressed by generating transgenic mice expressing hepcidin under the control of the liver-specific transthyretin promoter. We found that the majority of the transgenic mice were born with a pale skin and died within a few hours after birth. These transgenic animals had decreased body iron levels and presented severe microcytic hypochromic anemia. So far, three mosaic transgenic animals have survived. They were unequivocally identified by physical features, including reduced body size, pallor, hairless and crumpled skin. These pleiotropic effects were found to be associated with erythrocyte abnormalities, with marked anisocytosis, poikylocytosis and hypochromia, which are features characteristic of iron-deficiency anemia. These results strongly support the proposed role of hepcidin as a putative iron-regulatory hormone. The animal models devoid of hepcidin (the Usf2 knockout mice) or overexpressing the peptide (the transgenic mice presented in this paper) represent valuable tools for investigating iron homeostasis in vivo and for deciphering the molecular mechanisms of hepcidin action. PMID:11930010

  18. Severe iron deficiency anemia in transgenic mice expressing liver hepcidin

    PubMed Central

    Nicolas, Gaël; Bennoun, Myriam; Porteu, Arlette; Mativet, Sandrine; Beaumont, Carole; Grandchamp, Bernard; Sirito, Mario; Sawadogo, Michèle; Kahn, Axel; Vaulont, Sophie

    2002-01-01

    We recently reported the hemochromatosis-like phenotype observed in our Usf2 knockout mice. In these mice, as in murine models of hemochromatosis and patients with hereditary hemochromatosis, iron accumulates in parenchymal cells (in particular, liver and pancreas), whereas the reticuloendothelial system is spared from this iron loading. We suggested that this phenotypic trait could be attributed to the absence, in the Usf2 knockout mice, of a secreted liver-specific peptide, hepcidin. We conjectured that the reverse situation, namely overexpression of hepcidin, might result in phenotypic traits of iron deficiency. This question was addressed by generating transgenic mice expressing hepcidin under the control of the liver-specific transthyretin promoter. We found that the majority of the transgenic mice were born with a pale skin and died within a few hours after birth. These transgenic animals had decreased body iron levels and presented severe microcytic hypochromic anemia. So far, three mosaic transgenic animals have survived. They were unequivocally identified by physical features, including reduced body size, pallor, hairless and crumpled skin. These pleiotropic effects were found to be associated with erythrocyte abnormalities, with marked anisocytosis, poikylocytosis and hypochromia, which are features characteristic of iron-deficiency anemia. These results strongly support the proposed role of hepcidin as a putative iron-regulatory hormone. The animal models devoid of hepcidin (the Usf2 knockout mice) or overexpressing the peptide (the transgenic mice presented in this paper) represent valuable tools for investigating iron homeostasis in vivo and for deciphering the molecular mechanisms of hepcidin action. PMID:11930010

  19. Disproportionate growth in mice with Igf-2 transgenes.

    PubMed Central

    Ward, A; Bates, P; Fisher, R; Richardson, L; Graham, C F

    1994-01-01

    Injection transgenesis was used to study the long-term effects of excess insulin-like growth factor II on mouse growth and differentiation. By using a construct in which the coding region of the mouse insulin like growth factor II gene (Igf-2) was placed under the control of a keratin gene promoter, four transgenic lines were established, all of which displayed overgrowth of the skin as judged by wrinkling. In addition to high levels of expression in the skin, transgene transcripts were also present in the alimentary canal and uterus. At most of the sites of transgene expression the cell number (DNA content) was greatly increased, indicating a local action of the excess insulin-like growth factor II on cell multiplication. Adult total live weight was slightly increased and there was no macroscopic evidence of tumor formation. The characteristics of these transgenic mice indicate distinct local and systemic actions for insulin-like growth factor II. Images PMID:7524092

  20. SERPINB3 is associated with longer survival in transgenic mice

    PubMed Central

    Villano, Gianmarco; Ruvoletto, Mariagrazia; Ceolotto, Giulio; Quarta, Santina; Calabrese, Fiorella; Turato, Cristian; Tono, Natascia; Biasiolo, Alessandra; Cattelan, Arianna; Merkel, Carlo; Avogaro, Angelo; Gatta, Angelo; Pontisso, Patrizia

    2013-01-01

    The physiological roles of the protease inhibitor SERPINB3 (SB3) are still largely unknown. The study was addressed to assess the biological effects of this serpin in vivo using a SB3 transgenic mouse model. Two colonies of mice (123 transgenic for SB3 and 148 C57BL/6J controls) have been studied. Transgenic (TG) mice showed longer survival than controls and the difference was more remarkable in males than in females (18.5% vs 12.7% life span increase). In TG mice decreased IL-6 in serum and lower p66shc in the liver were observed. In addition, TG males showed higher expression of mTOR in the liver. Liver histology showed age-dependent increase of steatosis and decrease of glycogen storage in both groups and none of the animals developed neoplastic lesions. In conclusion, the gain in life span observed in SB3-transgenic mice could be determined by multiple mechanisms, including the decrease of circulating IL-6 and the modulation of ageing genes in the liver. PMID:24162160

  1. Progressive squamous epithelial neoplasia in K14-human papillomavirus type 16 transgenic mice.

    PubMed Central

    Arbeit, J M; Münger, K; Howley, P M; Hanahan, D

    1994-01-01

    To model human papillomavirus-induced neoplastic progression, expression of the early region of human papillomavirus type 16 (HPV16) was targeted to the basal cells of the squamous epithelium in transgenic mice, using a human keratin 14 (K14) enhancer/promoter. Twenty-one transgenic founder mice were produced, and eight lines carrying either wild-type or mutant HPV16 early regions that did not express the E1 or E2 genes were established. As is characteristic of human cancers, the E6 and E7 genes remained intact in these mutants. The absence of E1 or E2 function did not influence the severity of the phenotype that eventually developed in the transgenic mice. Hyperplasia, papillomatosis, and dysplasia appeared at multiple epidermal and squamous mucosal sites, including ear and truncal skin, face, snout and eyelids, and anus. The ears were the most consistently affected site, with pathology being present in all lines with 100% penetrance. This phenotype also progressed through discernible stages. An initial mild hyperplasia was followed by hyperplasia, which further progressed to dysplasia and papillomatosis. During histopathological progression, there was an incremental increase in cellular DNA synthesis, determined by 5-bromo-2'-deoxyuridine incorporation, and a profound perturbation in keratinocyte terminal differentiation, as revealed by immunohistochemistry to K5, K14, and K10 and filaggrin. These K14-HPV16 transgenic mice present an opportunity to study the role of the HPV16 oncogenes in the neoplastic progression of squamous epithelium and provide a model with which to identify genetic and epigenetic factors necessary for carcinogenesis. Images PMID:7515971

  2. Transmission of multiple system atrophy prions to transgenic mice

    PubMed Central

    Watts, Joel C.; Giles, Kurt; Oehler, Abby; Middleton, Lefkos; Dexter, David T.; Gentleman, Steve M.; DeArmond, Stephen J.; Prusiner, Stanley B.

    2013-01-01

    Prions are proteins that adopt alternative conformations, which become self-propagating. Increasing evidence argues that prions feature in the synucleinopathies that include Parkinson’s disease, Lewy body dementia, and multiple system atrophy (MSA). Although TgM83+/+ mice homozygous for a mutant A53T α-synuclein transgene begin developing CNS dysfunction spontaneously at ∼10 mo of age, uninoculated TgM83+/− mice (hemizygous for the transgene) remain healthy. To determine whether MSA brains contain α-synuclein prions, we inoculated the TgM83+/− mice with brain homogenates from two pathologically confirmed MSA cases. Inoculated TgM83+/− mice developed progressive signs of neurologic disease with an incubation period of ∼100 d, whereas the same mice inoculated with brain homogenates from spontaneously ill TgM83+/+ mice developed neurologic dysfunction in ∼210 d. Brains of MSA-inoculated mice exhibited prominent astrocytic gliosis and microglial activation as well as widespread deposits of phosphorylated α-synuclein that were proteinase K sensitive, detergent insoluble, and formic acid extractable. Our results provide compelling evidence that α-synuclein aggregates formed in the brains of MSA patients are transmissible and, as such, are prions. The MSA prion represents a unique human pathogen that is lethal upon transmission to Tg mice and as such, is reminiscent of the prion causing kuru, which was transmitted to chimpanzees nearly 5 decades ago. PMID:24218576

  3. Generation and characterization of transgenic mice expressing cobra venom factor.

    PubMed

    Andrä, Jörg; Halter, Roman; Kock, Michael A; Niemann, Heiner; Vogel, Carl-Wilhelm; Paul, Dieter

    2002-10-01

    Cobra venom factor (CVF), the anticomplementary protein in cobra venom, activates the alternative complement pathway, eventually leading to complement consumption. Here, we describe the development of a transgenic mouse model for CVF. We generated a DNA construct containing the full-length cDNA for single-chain pre-pro-CVF. Expression of CVF was controlled by the alpha(1)-antitrypsin promoter to achieve liver-specific expression. Linearized DNA was microinjected into murine ovary cells (strain CD(2)F(1) (BALB/cxDBA/2J)) and the newborn mice were analyzed for stable integration of CVF DNA. After establishing the transgene, mice were propagated in a BALB/c background. The CVF mRNA was detected in the liver and, in some animals, in the kidney. CVF protein was detected in small amounts in the serum. Serum complement hemolytic activity in CVF-transgenic mice was virtually absent. The concentration of plasma C3 was significantly reduced. The CVF-transgenic animals show no unusual phenotype. They provide an animal model to study the effect of long-term complement depletion by continued activation, as well as the role of complement in host immune response and pathogenesis of disease. PMID:12220893

  4. Visualization and genetic manipulation of adult neurogenesis using transgenic mice.

    PubMed

    Dhaliwal, Jagroop; Lagace, Diane C

    2011-03-01

    Many laboratories have focused efforts on the creation of transgenic mouse models to study adult neurogenesis. In the last decade several constitutive reporter, as well as inducible transgenic lines have been published that allowed for visualization, tracking and alteration of specific neurogenic cell populations in the adult brain. Given the popularity of this approach, multiple mouse lines are available, and this review summarizes the differences in the basic techniques that have been used to create these mice, highlighting the different constructs and reporter proteins used, as well as the strengths and limitations of each of these models. Representative examples from the literature demonstrate some of the diverse and seminal findings that have come to fruition through the laborious, yet highly rewarding work of creating transgenic mouse lines for adult neurogenesis research. PMID:21395845

  5. Generation of bacterial artificial chromosome (BAC) transgenic mice.

    PubMed

    Beil, Jane; Buch, Thorsten

    2014-01-01

    Transgenic mice are among the most helpful tools to study the role of genes in physiological conditions. In this protocol, we describe the generation of bacterial artificial chromosome (BACs) constructs, which are used to express a gene of interest under a particular promoter. BACs as driver of transgenes have the advantage that a characterization of transcriptional control elements is unnecessary and the construct's size usually reduces position effects from random integration. In the following, we firstly explain in detail the amplification of the BAC, the generation of the targeting construct as well as the recombination by ET-cloning, and the analysis of the recombined clones by Southern blot analysis. Finally, we also describe the preparation of the BACs for oocyte injection. In total, the construction of such BAC transgenes needs around 6-8 weeks. PMID:25064102

  6. Antitransferrin receptor antibody-RNase fusion protein expressed in the mammary gland of transgenic mice.

    PubMed

    Newton, D L; Pollock, D; DiTullio, P; Echelard, Y; Harvey, M; Wilburn, B; Williams, J; Hoogenboom, H R; Raus, J C; Meade, H M; Rybak, S M

    1999-12-10

    Antibodies fused to human enzymes offer an alternative to specifically targeting tumors with antibodies linked to plant or bacterial toxins. Since large amounts of these reagents can be administered without eliciting non-specific toxicities, efficient methods of production are needed. The goal of this work was to express a complex immunoenzyme fusion protein (immunotoxin) in the mammary gland of transgenic mice. A chimeric mouse/human antibody directed against the human transferrin receptor (E6) was fused at its CH2 domain to the gene for a human angiogenic ribonuclease, angiogenin (Ang). It was expressed in the mammary gland of mice and secreted into mouse milk. Expression levels in milk were approximately 0.8 g/l. The chimeric protein retained antibody binding activity and protein synthesis inhibitory activity equivalent to that of free Ang. It was specifically cytotoxic to human tumor cells in vitro. PMID:10648935

  7. Spi-1/PU.1 transgenic mice develop multistep erythroleukemias.

    PubMed Central

    Moreau-Gachelin, F; Wendling, F; Molina, T; Denis, N; Titeux, M; Grimber, G; Briand, P; Vainchenker, W; Tavitian, A

    1996-01-01

    Insertional mutagenesis of the spi-1 gene is associated with the emergence of malignant proerythroblasts during Friend virus-induced acute erythroleukemia. To determine the role of spi-1/PU.1 in the genesis of leukemia, we generated spi-1 transgenic mice. In one founder line the transgene was overexpressed as an unexpected-size transcript in various mouse tissues. Homozygous transgenic animals gave rise to live-born offspring, but 50% of the animals developed a multistep erythroleukemia within 1.5 to 6 months of birth whereas the remainder survived without evidence of disease. At the onset of the disease, mice became severely anemic. Their hematopoietic tissues were massively invaded with nontumorigenic proerythroblasts that express a high level of Spi-1 protein. These transgenic proerythroblasts are partially blocked in differentiation and strictly dependent on erythropoietin for their proliferation both in vivo and in vitro. A complete but transient regression of the disease was observed after erythrocyte transfusion, suggesting that the constitutive expression of spi-1 is related to the block of the differentiation of erythroid precursors. At relapse, erythropoietin-independent malignant proerythroblasts arose. Growth factor autonomy could be partially explained by the autocrine secretion of erythropoietin; however, other genetic events appear to be necessary to confer the full malignant phenotype. These results reveal that overexpression of spi-1 is essential for malignant erythropoiesis and does not alter other hematopoietic lineages. PMID:8628313

  8. Evaluating cerebellar functions using optogenetic transgenic mice

    NASA Astrophysics Data System (ADS)

    Welsh, John P.; Turecek, Josef; Turner, Eric E.

    2013-03-01

    We employed a transgenic mouse having conditional expression of ChR2(H134R) in neurons of the inferior olive to facilitate understanding of the role of electrical coupling and oscillation in central nervous system function. Two-photon excitation of ChR2-expressing neurons using 64 laser beams restricted to single inferior olive cell bodies depolarized neurons and evoked voltage deflections in neighboring neurons demonstrating electrical coupling. Broader illumination of neuronal ensembles using blue light induced an optical clamp of endogenous electrical rhythms in the inferior olive of acutely-prepared brain slices, which when applied in vivo directly modulated the local field potential activity and induced tremor. The experiments demonstrate novel methods to optically manipulate electrically coupled potentials and rhythmogenesis within a neuronal ensemble. From a functional perspective, the experiments shed light on the cellular and circuitry mechanisms of essential tremor, a prevalent neurological condition, by indicating time- and frequencydependence of tremor upon varying rhythms of inferior olive stimulation. The experiments indicate analog control of a brain rhythm that may be used to enhance our understanding of the functional consequences of central rhythmogenesis.

  9. Lymphoid hyperplasia and lymphoma in KSHV K1 transgenic mice.

    PubMed

    Berkova, Zuzana; Wang, Shu; Sehgal, Lalit; Patel, Keyur Pravinchandra; Prakash, Om; Samaniego, Felipe

    2015-05-01

    Growing evidence supports the involvement of human herpervirus 8, Kaposi's sarcoma associated herpesvirus (KSHV), in the pathology of primary effusion lymphoma, multicentric Castleman's disease, and Kaposi's sarcoma, but the exact mechanism of KSHV contribution to the oncogenic process remains elusive. We studied transgenic mice expressing the ORF K1 of KSHV, whose position in the KSHV genome corresponds to known lymphoproliferative genes of other herpesviruses. K1 protein was previously shown to contain a constitutively active ITAM domain, involved in activation of Akt and pro-survival signaling, and to inhibit Fas-mediated apoptosis by interfering with binding of FasL. All this pointed to a possible role of K1 in the pathogenesis of KSHV-associated cancers. K1 transgenic mice (80-90%) developed lymphoid hyperplasia and splenomegaly at 8 and 10 months of age, 25% had confirmed diagnosis of lymphoma, and 50% developed abdominal and/or hepatic tumors by 18 months of age. Histological examination showed loss of splenic architecture and increased cellularity. Lymph nodes showed disrupted architecture with effaced follicles and other pathological changes, including signs of angiofollicular lymphoid hyperplasia. One of the livers showed signs of angiosarcoma. In summary, our histology results revealed pathological changes in K1 transgenic mice similar to lymphoma, Castleman's disease, and angiosarcoma, suggesting that K1 may contribute to the development of KSHV-associated cancers. PMID:25301266

  10. Functional screening of an asthma QTL in YAC transgenic mice

    SciTech Connect

    Symula, Derek J.; Frazer, Kelly A.; Ueda, Yukihiko; Denefle, Patrice; Stevens, Mary E.; Wang, Zhi-En; Locksley, Richard; Rubin, Edward M.

    1999-07-02

    While large numbers of quantitative trait loci (QTLs) contributing to genetically complex conditions have been discovered, few causative genes have been identified. This is mainly due to the large size of QTLs and the subtle connection between genotype and quantitative phenotype associated with these conditions. While large numbers of quantitative trait loci (QTLs) contributing to genetically complex conditions have been discovered, few causative genes have been identified. This is mainly due to the large size of QTLs and the subtle connection between genotype and quantitative phenotype associated with these conditions. To screen for genes contributing to an asthma QTL mapped to human chromosome 5q33, the authors characterized a panel of large-insert 5q31 transgenics based on studies demonstrating that altering gene dosage frequently affects quantitative phenotypes normally influenced by that gene. This panel of human YAC transgenics, propagating a one megabase interva2048 chromosome 5q31 containing 23 genes, was screened for quantitative changes in several asthma-associated phenotypes. Multiple independent transgenic lines with altered IgE response to antigen treatment shared a 180 kb region containing 5 genes, including human interleukin 4 (IL4) and interleukin 13 (IL13), which induce IgE class switching in B cells5. Further analysis of these mice and mice transgenic for only murine Il4 and Il13 demonstrated that moderate changes in murine Il4 and Il13 expression affect asthma-associated phenotypes in vivo. This functional screen of large-insert transgenics enabled them to sift through multiple genes in the 5q3 asthma QTL without prior consideration of assumed individual gene function and identify genes that influence the QTL phenotype in vivo.

  11. Chimeric elk/mouse prion proteins in transgenic mice

    PubMed Central

    Tamgüney, Gültekin; Giles, Kurt; Oehler, Abby; Johnson, Natrina L.; DeArmond, Stephen J.

    2013-01-01

    Chronic wasting disease (CWD) of deer and elk is a highly communicable neurodegenerative disorder caused by prions. Investigations of CWD are hampered by slow bioassays in transgenic (Tg) mice. Towards the development of Tg mice that will be more susceptible to CWD prions, we created a series of chimeric elk/mouse transgenes that encode the N terminus of elk PrP (ElkPrP) up to residue Y168 and the C terminus of mouse PrP (MoPrP) beyond residue 169 (mouse numbering), designated Elk3M(SNIVVK). Between codons 169 and 219, six residues distinguish ElkPrP from MoPrP: N169S, T173N, V183I, I202V, I214V and R219K. Using chimeric elk/mouse PrP constructs, we generated 12 Tg mouse lines and determined incubation times after intracerebral inoculation with the mouse-passaged RML scrapie or Elk1P CWD prions. Unexpectedly, one Tg mouse line expressing Elk3M(SNIVVK) exhibited incubation times of <70 days when inoculated with RML prions; a second line had incubation times of <90 days. In contrast, mice expressing full-length ElkPrP had incubation periods of >250 days for RML prions. Tg(Elk3M,SNIVVK) mice were less susceptible to CWD prions than Tg(ElkPrP) mice. Changing three C-terminal mouse residues (202, 214 and 219) to those of elk doubled the incubation time for mouse RML prions and rendered the mice resistant to Elk1P CWD prions. Mutating an additional two residues from mouse to elk at codons 169 and 173 increased the incubation times for mouse prions to >300 days, but made the mice susceptible to CWD prions. Our findings highlight the role of C-terminal residues in PrP that control the susceptibility and replication of prions. PMID:23100369

  12. Genetic predisposition and parameters of malignant progression in K14-HPV16 transgenic mice.

    PubMed Central

    Coussens, L. M.; Hanahan, D.; Arbeit, J. M.

    1996-01-01

    Reproducible multi-stage progression to invasive squamous carcinoma of the epidermis has been achieved in transgenic mice expressing the HPV16 early-region genes, including the E6/E7 oncogenes, under the control of the human keratin-14 promoter/enhancer. Although 100% of K14-HPV16 transgenic animals develop hyperplastic and/or dysplastic lesions in several inbred backgrounds, including C57BL/6, BALB/c, and SSIN/SENCAR, only mice backcrossed into the FVB/n background progress to malignant squamous cell carcinomas of two pathological grades, well differentiated and moderate/poorly differentiated (WDSC or MPDSC, respectively), each displaying characteristic patterns of malignant behavior. WDSCs typically arise within the epidermis of the ear and invade deeply into the underlying dermis but fail to metastasize, whereas MPDSCs develop on the chest and truncal skin and invariably metastasize to regional lymph nodes. The transition to the malignant state, in 21% of FVB/n transgenic mice, is characterized by alteration of the repertoire of keratin intermediate filament proteins expressed within neoplastic epidermis, such that WDSCs maintain expression of keratins common to terminally differentiating stratified keratinocytes (K10), whereas MPDSCs are distinguished from WDSCs by activation of embryonic and mucosal keratins (K13, K8, and K19). Precursor hyperplastic and dysplastic lesions are characterized by a progressively increased proliferative index, striking morphological alterations in keratinocyte cell-cell and cell-matrix interactions, and extensive remodeling of the underlying dermal stroma. Remarkably, this extensive stromal remodeling, which may facilitate both angiogenesis and eventual tumor cell invasion, develops early at the dysplastic stage in all animals well before malignant conversion. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:8952526

  13. Mammary gland tumor formation in transgenic mice overexpressing stromelysin-1

    SciTech Connect

    Sympson, Carolyn J; Bissell, Mina J; Werb, Zena

    1995-06-01

    An intact basement membrane (BM) is essential for the proper function, differentiation and morphology of many epithelial cells. The disruption or loss of this BM occurs during normal development as well as in the disease state. To examine the importance of BM during mammary gland development in vivo, we generated transgenic mice that inappropriately express autoactivating isoforms of the matrix metalloproteinase stromelysin-1. The mammary glands from these mice are both functionally and morphologically altered throughout development. We have now documented a dramatic incidence of breast tumors in several independent lines of these mice. These data suggest that overexpression of stromelysin-1 and disruption of the BM may be a key step in the multi-step process of breast cancer.

  14. Acetaminophen-induced acute liver injury in HCV transgenic mice

    SciTech Connect

    Uehara, Takeki; Kosyk, Oksana; Jeannot, Emmanuelle; Bradford, Blair U.; Tech, Katherine; Macdonald, Jeffrey M.; Boorman, Gary A.; Chatterjee, Saurabh; Mason, Ronald P.; Melnyk, Stepan B.; Tryndyak, Volodymyr P.; Pogribny, Igor P.; Rusyn, Ivan

    2013-01-15

    The exact etiology of clinical cases of acute liver failure is difficult to ascertain and it is likely that various co-morbidity factors play a role. For example, epidemiological evidence suggests that coexistent hepatitis C virus (HCV) infection increased the risk of acetaminophen-induced acute liver injury, and was associated with an increased risk of progression to acute liver failure. However, little is known about possible mechanisms of enhanced acetaminophen hepatotoxicity in HCV-infected subjects. In this study, we tested a hypothesis that HCV-Tg mice may be more susceptible to acetaminophen hepatotoxicity, and also evaluated the mechanisms of acetaminophen-induced liver damage in wild type and HCV-Tg mice expressing core, E1 and E2 proteins. Male mice were treated with a single dose of acetaminophen (300 or 500 mg/kg in fed animals; or 200 mg/kg in fasted animals; i.g.) and liver and serum endpoints were evaluated at 4 and 24 h after dosing. Our results suggest that in fed mice, liver toxicity in HCV-Tg mice is not markedly exaggerated as compared to the wild-type mice. In fasted mice, greater liver injury was observed in HCV-Tg mice. In fed mice dosed with 300 mg/kg acetaminophen, we observed that liver mitochondria in HCV-Tg mice exhibited signs of dysfunction showing the potential mechanism for increased susceptibility. -- Highlights: ► Acetaminophen-induced liver injury is a significant clinical challenge. ► HCV-infected subjects may be at higher risk for acetaminophen-induced liver injury. ► We used HCV transgenics to test if liver injury due to acetaminophen is exacerbated.

  15. Multistage epidermal carcinogenesis in transgenic mice: cooperativity and paradox.

    PubMed

    Greenhalgh, D A; Wang, X J; Roop, D R

    1996-04-01

    Skin cancer is one of the most prevalent forms of human neoplasia with a frequency approaching that of all other neoplasms combined. Given this alarming statistic, which may be further exacerbated by increased ultraviolet B irradiation from ozone depletion, it is vital that realistic, relevant model systems are developed to increase our understanding of the underlying molecular mechanisms of carcinogenesis that result in or evaluate new treatment modalities. Toward this goal, the ability to stably introduce genes into the germline of mice has greatly enhanced prospects for generation of transgenic animal models of multistage molecular carcinogenesis. Moreover, when genes are combined with regulatory sequences that target their expression to specific tissues, investigators are able to study neoplasia both in the context of living organisms and in the tissues suspected of being the targets of these genes. The epidermis is an attractive tissue for targeted gene expression; not only is it a model for epithelial diseases in general, but the accessibility of the epidermis allows easy detection of progressive pathological changes that result from transgene expression and facilitates assessment of the potential role played by environmental factors. We have developed a targeting vector based on the human keratin gene (HK1), which is expressed exclusively in the epidermis of transgenic mice, at a late stage in development and in both basal and differentiated cells. Through the use of this targeting ability, rasHa, fos, and TGF alpha transgenic mice have been developed that exhibit preneoplastic epidermal hyperplasia and hyperkeratosis, and later benign, regression prone papillomas. Together, coexpression of two oncogenes cooperated to give autonomous papillomas, which possessed the phenotypic stability to allow assessment of a third genetic event, namely loss of the p53 tumor suppressor gene, via mating with p53 knockout mice. Loss of p53 expression, however, identified a

  16. Transgenic mice with overexpression of mutated human optineurin(E50K) in the retina.

    PubMed

    Meng, Qingfeng; Xiao, Zheng; Yuan, Huiping; Xue, Fei; Zhu, Yuanmao; Zhou, Xinrong; Yang, Binbin; Sun, Jingbo; Meng, Bo; Sun, Xian; Cheng, Fang

    2012-02-01

    In the present work, Site-directed mutagenesis to insert the Glu50Lys amino acid substitution was achieved by PCR using plasmid pBluescript-OPTN. Mutated human OPTN(E50K) gene-driven mouse c-kit promoter was constructed and confirmed by endonuclease digestion and sequence analysis. Transgenic mice were generated via the microinjection method. PCR and DNA dot blot were used to screen the positive transgenic mice. RT-PCR analyzed the RNA level and location of mutated human OPTN(E50K) mRNA expression in transgenic mice. Western blot and immunohistochemistry were used to detect the level and location of mutated human OPTN(E50K) expression in transgenic mice. A transgenic mouse model with overexpression of mutated human OPTN(E50K) in retina was successfully established. The transgene was integrated and transmitted into the chromosome of transgenic mice. Mutated human OPTN(E50K) gene was controlled by c-kit promoter and expressed in the retina in mice. Mutated human OPTN(E50K) in transgenic mice was higher than that of wild type C57BL/6J mice. Our studies had provided a new transgenic model for investigating the molecular properties of mutated human OPTN(E50K). PMID:21681420

  17. Sex-linked behavioural differences in mice expressing a human insulin transgene in the medial habenula.

    PubMed

    Douhet, P; Bertaina, V; Durkin, T; Calas, A; Destrade, C

    1997-12-01

    We previously reported that a human insulin transgene was specifically expressed in the medial habenula of the adult mouse brain, and that this expression was ascribed to the delta-168 transgene. The present study analyses the possible behavioural consequences of this insulin transgene expression using measures of food intake, spontaneous activity, emotional reactivity, learning and extinction performance of an operant task. The delta-168 transgenic mice did not differ from the C57BL/6 control mice as concerns food intake, behaviour in the open field, or emotional response in an elevated plus maze. On the other hand, measures of locomotor activity in a circular corridor revealed a significantly faster decline of spontaneous locomotor activity in male as compared to female delta-168 transgenic mice. Moreover, as compared to female transgenic mice, male transgenic mice exhibited a deficit in the rate of acquisition and an acceleration of the rate of extinction of a bar press response in a Skinner box. In contrast, the behaviour of female transgenic mice did not differ from either male or female C57BL/6 control mice. The results of the present study demonstrate that the behavioural modifications observed in delta-168 transgenic mice are sex-linked and suggest that these behavioural differences result from changes in the interaction (interface) between motivational and motor mechanisms mediated via the striato-habenulo-mesencephalic system. PMID:9475633

  18. Chronic activation of FXR in transgenic mice caused perinatal toxicity and sensitized mice to cholesterol toxicity.

    PubMed

    Cheng, Qiuqiong; Inaba, Yuka; Lu, Peipei; Xu, Meishu; He, Jinhan; Zhao, Yueshui; Guo, Grace L; Kuruba, Ramalinga; de la Vega, Rona; Evans, Rhobert W; Li, Song; Xie, Wen

    2015-04-01

    The nuclear receptor farnesoid X receptor (FXR) (nuclear receptor subfamily 1, group H, member 4, or NR1H4) is highly expressed in the liver and intestine. Previous reports have suggested beneficial functions of FXR in the homeostasis of bile acids, lipids, and glucose, as well as in promoting liver regeneration and inhibiting carcinogenesis. To investigate the effect of chronic FXR activation in vivo, we generated transgenic mice that conditionally and tissue specifically express the activated form of FXR in the liver and intestine. Unexpectedly, the transgenic mice showed several intriguing phenotypes, including partial neonatal lethality, growth retardation, and spontaneous liver toxicity. The transgenic mice also displayed heightened sensitivity to a high-cholesterol diet-induced hepatotoxicity but resistance to the gallstone formation. The phenotypes were transgene specific, because they were abolished upon treatment with doxycycline to silence the transgene expression. The perinatal toxicity, which can be rescued by a maternal vitamin supplement, may have resulted from vitamin deficiency due to low biliary bile acid output as a consequence of inhibition of bile acid formation. Our results also suggested that the fibroblast growth factor-inducible immediate-early response protein 14 (Fn14), a member of the proinflammatory TNF family, is a FXR-responsive gene. However, the contribution of Fn14 induction in the perinatal toxic phenotype of the transgenic mice remains to be defined. Because FXR is being explored as a therapeutic target, our results suggested that a chronic activation of this nuclear receptor may have an unintended side effect especially during the perinatal stage. PMID:25719402

  19. Chronic Activation of FXR in Transgenic Mice Caused Perinatal Toxicity and Sensitized Mice to Cholesterol Toxicity

    PubMed Central

    Cheng, Qiuqiong; Inaba, Yuka; Lu, Peipei; Xu, Meishu; He, Jinhan; Zhao, Yueshui; Guo, Grace L.; Kuruba, Ramalinga; de la Vega, Rona; Evans, Rhobert W.; Li, Song

    2015-01-01

    The nuclear receptor farnesoid X receptor (FXR) (nuclear receptor subfamily 1, group H, member 4, or NR1H4) is highly expressed in the liver and intestine. Previous reports have suggested beneficial functions of FXR in the homeostasis of bile acids, lipids, and glucose, as well as in promoting liver regeneration and inhibiting carcinogenesis. To investigate the effect of chronic FXR activation in vivo, we generated transgenic mice that conditionally and tissue specifically express the activated form of FXR in the liver and intestine. Unexpectedly, the transgenic mice showed several intriguing phenotypes, including partial neonatal lethality, growth retardation, and spontaneous liver toxicity. The transgenic mice also displayed heightened sensitivity to a high-cholesterol diet-induced hepatotoxicity but resistance to the gallstone formation. The phenotypes were transgene specific, because they were abolished upon treatment with doxycycline to silence the transgene expression. The perinatal toxicity, which can be rescued by a maternal vitamin supplement, may have resulted from vitamin deficiency due to low biliary bile acid output as a consequence of inhibition of bile acid formation. Our results also suggested that the fibroblast growth factor-inducible immediate-early response protein 14 (Fn14), a member of the proinflammatory TNF family, is a FXR-responsive gene. However, the contribution of Fn14 induction in the perinatal toxic phenotype of the transgenic mice remains to be defined. Because FXR is being explored as a therapeutic target, our results suggested that a chronic activation of this nuclear receptor may have an unintended side effect especially during the perinatal stage. PMID:25719402

  20. Specific ablation of thyroid follicle cells in adult transgenic mice.

    PubMed

    Wallace, H; Ledent, C; Vassart, G; Bishop, J O; al-Shawi, R

    1991-12-01

    The coding region of the herpes simplex type 1 virus thymidine kinase gene was coupled to the promoter of the bovine thyroglobulin gene and introduced into the genome of mice. The viral thymidine kinase (HSV1-TK) was expressed mainly in the thyroid glands and testis. Upon treatment of transgenic females with the antiherpetic agent Ganciclovir the thyroid regressed, while the parathyroid gland was unaffected. The number of thyroid follicle cells was greatly reduced after 3 days, and they were completely absent after 7 days of treatment. After 14 days, the levels of circulating T4 and T3 were below the limits of detection, total soluble protein recovered from the thyroid and parathyroid glands together was 10% of the control value, and the level of thyroid HSV1-TK was more than 100-fold lower than that in transgenic controls. Levels of circulating PTH and calcitonin remained normal. At the time of treatment the mice were adults. Thus, the thyroid follicle cells were selectively ablated after normal development with a functional thyroid gland. When treatment with Ganciclovir was terminated after 14 days, no circulating T4 or T3 or other indications of thyroid regeneration were detected for a subsequent period of 90 days. During this time the mice gained weight more slowly than controls, at a rate consistent with the suppression of GH synthesis by thyroid deficiency. The production of mouse major urinary protein (MUP) ceased in the treated mice and was completely restored by the administration of T4. MUP production was not restored by GH, demonstrating that the expression of the Mup genes requires T4 in addition to GH. PMID:1659524

  1. Pathogenesis of Chronic Wasting Disease in Cervidized Transgenic Mice

    PubMed Central

    Seelig, Davis M.; Mason, Gary L.; Telling, Glenn C.; Hoover, Edward A.

    2010-01-01

    Chronic wasting disease (CWD) is a fatal, endemic prion disease of wild and captive cervids, including deer, elk, and moose. Typical of prion diseases, CWD is characterized by the conversion of the native, protease-sensitive protein PrPC to a protease-resistant isoform, denoted as PrPRES. Here we have studied the expression of cervid PrPC and the pathogenesis of CWD infection in transgenic mice expressing the normal cervid prion protein (Tg[CerPrP] mice). Using tissue-based in situ immunohistochemistry protocols, we first identified cervid PrPC expression in the lymphoid, nervous, hemopoietic, endocrine, and certain epithelial tissues of Tg[CerPrP] mice. Tg[CerPrP] mice were then inoculated with CWD via one of four routes (intracerebral, intravenous, intraperitoneal, or oral); all groups developed spongiform encephalopathy, although the oral route required a larger infecting dose. Incubation periods were 184 ± 13, 218 ± 15, 200 ± 7, and 350 ± 27 days after inoculation, respectively. In longitudinal studies, we tracked the appearance of PrPRES in the brain, spleen, Peyer’s patches, lymph nodes, pancreatic islets of Langerhans, bone marrow, and salivary glands of preclinical and terminal mice. In addition, we documented horizontal transmission of CWD from inoculated mice and to un-inoculated cohabitant cage-mates. This work documents the multiroute susceptibility, pathogenesis, and lateral transmission of CWD infection in Tg[CerPrP] mice, affirming this model as a robust system to study this cervid transmissible spongiform encephalopathy. PMID:20395435

  2. Brain Phenotype of Transgenic Mice Overexpressing Cystathionine β-Synthase

    PubMed Central

    Régnier, Vinciane; Billard, Jean-Marie; Gupta, Sapna; Potier, Brigitte; Woerner, Stéphanie; Paly, Evelyne; Ledru, Aurélie; David, Sabrina; Luilier, Sabrina; Bizot, Jean-Charles; Vacano, Guido; Kraus, Jan P.; Patterson, David; Kruger, Warren D.; Delabar, Jean M.; London, Jaqueline

    2012-01-01

    Background The cystathionine β-synthase (CBS) gene, located on human chromosome 21q22.3, is a good candidate for playing a role in the Down Syndrome (DS) cognitive profile: it is overexpressed in the brain of individuals with DS, and it encodes a key enzyme of sulfur-containing amino acid (SAA) metabolism, a pathway important for several brain physiological processes. Methodology/Principal Findings Here, we have studied the neural consequences of CBS overexpression in a transgenic mouse line (60.4P102D1) expressing the human CBS gene under the control of its endogenous regulatory regions. These mice displayed a ∼2-fold increase in total CBS proteins in different brain areas and a ∼1.3-fold increase in CBS activity in the cerebellum and the hippocampus. No major disturbance of SAA metabolism was observed, and the transgenic mice showed normal behavior in the rotarod and passive avoidance tests. However, we found that hippocampal synaptic plasticity is facilitated in the 60.4P102D1 line. Conclusion/Significance We demonstrate that CBS overexpression has functional consequences on hippocampal neuronal networks. These results shed new light on the function of the CBS gene, and raise the interesting possibility that CBS overexpression might have an advantageous effect on some cognitive functions in DS. PMID:22253703

  3. Lymphopoiesis in transgenic mice over-expressing Artemis.

    PubMed

    Rivera-Munoz, P; Abramowski, V; Jacquot, S; André, P; Charrier, S; Lipson-Ruffert, K; Fischer, A; Galy, A; Cavazzana, M; de Villartay, J-P

    2016-02-01

    Artemis is a factor of the non-homologous end joining pathway involved in DNA double-strand break repair that has a critical role in V(D)J recombination. Mutations in DCLRE1C/ARTEMIS gene result in radiosensitive severe combined immunodeficiency in humans owing to a lack of mature T and B cells. Given the known drawbacks of allogeneic hematopoietic stem cell transplantation (HSCT), gene therapy appears as a promising alternative for these patients. However, the safety of an unregulated expression of Artemis has to be established. We developed a transgenic mouse model expressing human Artemis under the control of the strong CMV early enhancer/chicken beta actin promoter through knock-in at the ROSA26 locus to analyze this issue. Transgenic mice present a normal development, maturation and function of T and B cells with no signs of lymphopoietic malignancies for up to 15 months. These results suggest that the over-expression of Artemis in mice (up to 40 times) has no deleterious effects in early and mature lymphoid cells and support the safety of gene therapy as a possible curative treatment for Artemis-deficient patients. PMID:26361272

  4. Both core and F proteins of hepatitis C virus could enhance cell proliferation in transgenic mice

    SciTech Connect

    Hu, Wen-Ta; Li, Hui-Chun; Lee, Shen-Kao; Ma, Hsin-Chieh; Yang, Chee-Hing; Chen, Hung-Ling; Lo, Shih-Yen

    2013-05-24

    Highlights: •HCV core and F proteins could induce hepatocyte proliferation in the transgenic mice. •β-Catenin signaling pathway was activated by core protein in the transgenic mice. •β-Catenin signaling pathway was activated by myc-F protein in the transgenic mice. •Expression of SMA protein was enhanced by core but not myc-F protein. -- Abstract: The role of the protein encoded by the alternative open reading frame (ARF/F/core+1) of the Hepatitis C virus (HCV) genome in viral pathogenesis remains unknown. The different forms of ARF/F/core+1 protein were labile in cultured cells, a myc-tag fused at the N-terminus of the F protein made it more stable. To determine the role of core and F proteins in HCV pathogenesis, transgenic mice with either protein expression under the control of Albumin promoter were generated. Expression of core protein and F protein with myc tag (myc-F) could be detected by Western blotting analysis in the livers of these mice. The ratio of liver to body weight is increased for both core and myc-F transgenic mice compared to that of wild type mice. Indeed, the proliferating cell nuclear antigen protein, a proliferation marker, was up-regulated in the transgenic mice with core or myc-F protein. Further analyses by microarray and Western blotting suggested that β-catenin signaling pathway was activated by either core or myc-F protein in the transgenic mice. These transgenic mice were further treated with either Diethynitrosamine (a tumor initiator) or Phenobarbital (a tumor promoter). Phenobarbital but not Diethynitrosamine treatment could increase the liver/body weight ratio of these mice. However, no tumor formation was observed in these mice. In conclusion, HCV core and myc-F proteins could induce hepatocyte proliferation in the transgenic mice possibly through β-catenin signaling pathway.

  5. Multiple Cystic Sweat Gland Tumors in Transgenic Mice

    PubMed Central

    Matthias, Nadine; Lockworth, Cynthia R; Zhang, Fanmao; Lee, Mong-Hong; Yeung, Sai-Ching J; Tsai, Kenneth Y; Hamir, Amir N

    2012-01-01

    Here we describe gross and microscopic sweat gland tumors found in a transgenic mouse model of breast cancer, which had transforming growth factor α under the control of mouse mammary tumor virus promoter (MMTV–TGFα). Initially, 20% of the mice in the colony were affected. Cystic lesions formed on the phalanges, palmar surfaces of the metacarpals, and plantar surfaces of the metatarsals. The lesions were multifocal and nonulcerated with straw-colored fluid, ranging in size from 1 to 30 mm at the largest dimension. The colony was monitored for 6 mo; during that time, the prevalence of lesions increased to 52% of the mice. Histologically, in most cases the cyst walls were lined by 1 or 2 layers of normal-appearing epithelial cells that resembled basal cells, indicating adenoma. However, 2 cysts from 2 different mice had papillary proliferative projections and extensive disorganized glandular structures that protruded into the cyst cavities, indicating adenocarcinoma. In these 2 cases, the neoplastic cells revealed architectural and cytologic atypia with rare mitoses. Similar findings have previously been observed in sweat gland tumors; however, multiple sweat-gland tumors have not been reported in mice. PMID:22330648

  6. Extra-prostatic Transgene-associated Neoplastic Lesions in Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) Mice

    PubMed Central

    Berman-Booty, Lisa D.; Thomas-Ahner, Jennifer M.; Bolon, Brad; Oglesbee, Michael J.; Clinton, Steven K.; Kulp, Samuel K.; Chen, Ching-Shih; La Perle, Krista

    2014-01-01

    Male transgenic adenocarcinoma of the mouse prostate (TRAMP) mice are frequently used in prostate cancer research because their prostates consistently develop a series of pre-neoplastic and neoplastic lesions. Disease progression in TRAMP mouse prostates culminates in metastatic, poorly differentiated carcinomas with neuroendocrine features. The androgen dependence of the rat probasin promoter largely limits transgene expression to the prostatic epithelium. However, extra-prostatic transgene-positive lesions have been described in TRAMP mice, including renal tubulo-acinar carcinomas, neuroendocrine carcinomas of the urethra, and phyllodes-like tumors of the seminal vesicle. Here we describe the histologic and immunohistochemical features of two novel extra-prostatic lesions in TRAMP mice: primary anaplastic tumors of uncertain cell origin in the midbrain, and poorly differentiated adenocarcinomas of the submandibular salivary gland. These newly characterized tumors apparently result from transgene expression in extra-prostatic locations rather than representing metastatic prostate neoplasms because lesions were identified in both male and female mice as well as in male TRAMP mice without histologically apparent prostate tumors. In this paper we also calculate the incidences of the urethral carcinomas and renal tubulo-acinar carcinomas, further elucidate the biological behavior of the urethral carcinomas, and demonstrate the critical importance of complete necropsies even when evaluating presumably well characterized phenotypes in genetically engineered mice. PMID:24742627

  7. Extra-prostatic transgene-associated neoplastic lesions in transgenic adenocarcinoma of the mouse prostate (TRAMP) mice.

    PubMed

    Berman-Booty, Lisa D; Thomas-Ahner, Jennifer M; Bolon, Brad; Oglesbee, Michael J; Clinton, Steven K; Kulp, Samuel K; Chen, Ching-Shih; La Perle, Krista M D

    2015-02-01

    Male transgenic adenocarcinoma of the mouse prostate (TRAMP) mice are frequently used in prostate cancer research because their prostates consistently develop a series of preneoplastic and neoplastic lesions. Disease progression in TRAMP mouse prostates culminates in metastatic, poorly differentiated carcinomas with neuroendocrine features. The androgen dependence of the rat probasin promoter largely limits transgene expression to the prostatic epithelium. However, extra-prostatic transgene-positive lesions have been described in TRAMP mice, including renal tubuloacinar carcinomas, neuroendocrine carcinomas of the urethra, and phyllodes-like tumors of the seminal vesicle. Here, we describe the histologic and immunohistochemical features of 2 novel extra-prostatic lesions in TRAMP mice: primary anaplastic tumors of uncertain cell origin in the midbrain and poorly differentiated adenocarcinomas of the submandibular salivary gland. These newly characterized tumors apparently result from transgene expression in extra-prostatic locations rather than representing metastatic prostate neoplasms because lesions were identified in both male and female mice and in male TRAMP mice without histologically apparent prostate tumors. In this article, we also calculate the incidences of the urethral carcinomas and renal tubuloacinar carcinomas, further elucidate the biological behavior of the urethral carcinomas, and demonstrate the critical importance of complete necropsies even when evaluating presumably well characterized phenotypes in genetically engineered mice. PMID:24742627

  8. Divergent phenotypes in mutant TDP-43 transgenic mice highlight potential confounds in TDP-43 transgenic modeling.

    PubMed

    D'Alton, Simon; Altshuler, Marcelle; Cannon, Ashley; Dickson, Dennis W; Petrucelli, Leonard; Lewis, Jada

    2014-01-01

    The majority of cases of frontotemporal lobar degeneration and amyotrophic lateral sclerosis are pathologically defined by the cleavage, cytoplasmic redistribution and aggregation of TAR DNA binding protein of 43 kDa (TDP-43). To examine the contribution of these potentially toxic mechanisms in vivo, we generated transgenic mice expressing human TDP-43 containing the familial amyotrophic lateral sclerosis-linked M337V mutation and identified two lines that developed neurological phenotypes of differing severity and progression. The first developed a rapid cortical neurodegenerative phenotype in the early postnatal period, characterized by fragmentation of TDP-43 and loss of endogenous murine Tdp-43, but entirely lacking aggregates of ubiquitin or TDP-43. A second, low expressing line was aged to 25 months without a severe neurodegenerative phenotype, despite a 30% loss of mouse Tdp-43 and accumulation of lower molecular weight TDP-43 species. Furthermore, TDP-43 fragments generated during neurodegeneration were not C-terminal, but rather were derived from a central portion of human TDP-43. Thus we find that aggregation is not required for cell loss, loss of murine Tdp-43 is not necessarily sufficient in order to develop a severe neurodegenerative phenotype and lower molecular weight TDP-43 positive species in mouse models should not be inherently assumed to be representative of human disease. Our findings are significant for the interpretation of other transgenic studies of TDP-43 proteinopathy. PMID:24466128

  9. Gene expression profile in liver of hB1F transgenic mice

    PubMed Central

    Wang, Shui-Liang; Yang, Hua; Xie, You-Hua; Wang, Yuan; Li, Jian-Zhong; Wang, Long; Wang, Zhu-Gang; Fu, Ji-Liang

    2004-01-01

    AIM: To analyze the tissue morphologic phenotype and liver gene expression profile of hB1F transgenic mice. METHODS: Transgene expression was analyzed with RT-PCR and Western blotting. For one of the transgenic mouse lines, tissue expression pattern of the transgene was also examined with immunochemical methods. Pathological analysis was used to examine the tissue morphologic phenotype of established transgenic mice. The liver gene expression profile of transgenic mice was analyzed with microchip, and some of the differentially expressed genes were verified with RT-PCR. RESULTS: The expressions of hB1F were shown in livers from 6 of 7 transgenic mouse lines. The overexpression of hB1F transgene did not cause pathological changes. Expressions of three genes were up-regulated, while down-regulation was observed for 25 genes. CONCLUSION: The overexpression of hB1F transgene may cause changes of gene expression profiles in the liver of transgenic mice. PMID:15378783

  10. Strain-Specific Regulation of Striatal Phenotype in Drd2-eGFP BAC Transgenic Mice

    PubMed Central

    Chan, C. Savio; Peterson, Jayms D.; Gertler, Tracy S.; Glajch, Kelly E.; Quintana, Ruth E.; Cui, Qiaoling; Sebel, Luke E.; Plotkin, Joshua L.; Shen, Weixing; Heiman, Myriam; Heintz, Nathaniel; Greengard, Paul; Surmeier, D. James

    2012-01-01

    Mice carrying bacterial artificial chromosome (BAC) transgenes have become important tools for neuroscientists, providing a powerful means of dissecting complex neural circuits in the brain. Recently, it was reported that one popular line of these micemice possessing a BAC transgene with a D2 dopamine receptor (Drd2) promoter construct coupled to an enhanced green fluorescent protein (eGFP) reporter – had abnormal striatal gene expression, physiology and motor behavior. Unlike most of the work using BAC mice, this interesting study relied upon mice backcrossed on the outbred Swiss Webster strain that were homozygous for the Drd2-eGFP BAC transgene.The experiments reported here were conducted to determine whether mouse strain or zygosity was a factor in the reported abnormalities. As reported, SW mice were very sensitive to transgene expression. However, in more commonly used inbred strains of mice (C57BL/6, FVB/N) that were hemizygous for the transgene, the Drd2-eGFP BAC transgene did not alter striatal gene expression, physiology or motor behavior. Thus, the use of inbred strains of mice which are hemizygous for the Drd2 BAC transgene provide a reliable tool for studying basal ganglia function. PMID:22764222

  11. Evidence for lymphatic Aβ clearance in Alzheimer's transgenic mice.

    PubMed

    Pappolla, Miguel; Sambamurti, Kumar; Vidal, Ruben; Pacheco-Quinto, Javier; Poeggeler, Burkhard; Matsubara, Etsuro

    2014-11-01

    Evidence has shown that lymphatic drainage contributes to removal of debris from the brain but its role in the accumulation of amyloid β peptides (Aβ) has not been demonstrated. We examined the levels of various forms of Aβ in the brain, plasma and lymph nodes in a transgenic model of Alzheimer's disease (AD) at different ages. Herein, we report on the novel finding that Aβ is present in the cervical and axillary lymph nodes of AD transgenic mice and that Aβ levels in lymph nodes increase over time, mirroring the increase of Aβ levels observed in the brain. Aβ levels in lymph nodes were significantly higher than in plasma. At age 15.5months, there was a significant increase of monomeric soluble Aβ40 (p=0.003) and Aβ42 (p=0.05) in the lymph nodes over the baseline values measured at 6months of age. In contrast, plasma levels of Aβ40 showed no significant changes (p=0.68) and plasma levels Aβ42 significantly dropped (p=0.02) at the same age. Aβ concentration was low to undetectable in splenic lymphoid tissue and several other control tissues including heart, lung, liver, kidneys and intestine of the same animals, strongly suggesting that Aβ peptides in lymph nodes are derived from the brain. PMID:25102344

  12. Leukemia Inhibitory Factor Induces Neurotransmitter Switching in Transgenic Mice

    NASA Astrophysics Data System (ADS)

    Bamber, Bruce A.; Masters, Brian A.; Hoyle, Gary W.; Brinster, Ralph L.; Palmiter, Richard D.

    1994-08-01

    Leukemia inhibitory factor (LIF) is a cytokine growth factor that induces rat sympathetic neurons to switch their neurotransmitter phenotype from noradrenergic to cholinergic in vitro. To test whether LIF can influence neuronal differentiation in vivo, we generated transgenic mice that expressed LIF in pancreatic islets under the control of the insulin promoter and evaluated the neurotransmitter phenotype of the pancreatic sympathetic innervation. We also used the insulin promoter to coexpress nerve growth factor in the islets, which greatly increased the density of sympathetic innervation and facilitated analysis of the effects of LIF. Our data demonstrate that tyrosine hydroxylase and catecholamines declined and choline acetyltransferase increased in response to LIF. We conclude that LIF can induce neurotransmitter switching of sympathetic neurons in vivo.

  13. Copper transport during lactation in transgenic mice expressing the human ATP7A protein

    PubMed Central

    Llanos, Roxana M.; Michalczyk, Agnes A.; Freestone, David J.; Currie, Scott; Linder, Maria C.; Ackland, M. Leigh; Mercer, Julian F.B.

    2008-01-01

    Both copper transporting ATPases, ATP7A and ATP7B, are expressed in mammary epithelial cells but their role in copper delivery to milk has not been clarified. We investigated the role of ATP7A in delivery of copper to milk using transgenic mice that over-express human ATP7A. In mammary gland of transgenic mice, human ATP7A protein was 10- to 20-fold higher than in control mice, and was localized to the basolateral membrane of mammary epithelial cells in lactating mice. The copper concentration in the mammary gland of transgenic dams and stomach contents of transgenic pups was significantly reduced compared to non-transgenic mice. The mRNA levels of endogenous Atp7a, Atp7b, and Ctr1 copper transporters in the mammary gland were not altered by the expression of the ATP7A transgene, and the protein levels of Atp7b and ceruloplasmin were similar in transgenic and non-transgenic mice. These data suggest that ATP7A plays a role in removing excess copper from the mammary epithelial cells rather than supplying copper to milk. PMID:18515074

  14. RNAi-mediated knockdown of IKK1 in transgenic mice using a transgenic construct containing the human H1 promoter.

    PubMed

    Moreno-Maldonado, Rodolfo; Murillas, Rodolfo; Navarro, Manuel; Page, Angustias; Suarez-Cabrera, Cristian; Alameda, Josefa P; Bravo, Ana; Casanova, M Llanos; Ramirez, Angel

    2014-01-01

    Inhibition of gene expression through siRNAs is a tool increasingly used for the study of gene function in model systems, including transgenic mice. To achieve perdurable effects, the stable expression of siRNAs by an integrated transgenic construct is necessary. For transgenic siRNA expression, promoters transcribed by either RNApol II or III (such as U6 or H1 promoters) can be used. Relatively large amounts of small RNAs synthesis are achieved when using RNApol III promoters, which can be advantageous in knockdown experiments. To study the feasibility of H1 promoter-driven RNAi-expressing constructs for protein knockdown in transgenic mice, we chose IKK1 as the target gene. Our results indicate that constructs containing the H1 promoter are sensitive to the presence of prokaryotic sequences and to transgene position effects, similar to RNApol II promoters-driven constructs. We observed variable expression levels of transgenic siRNA among different tissues and animals and a reduction of up to 80% in IKK1 expression. Furthermore, IKK1 knockdown led to hair follicle alterations. In summary, we show that constructs directed by the H1 promoter can be used for knockdown of genes of interest in different organs and for the generation of animal models complementary to knockout and overexpression models. PMID:24523631

  15. Early Axonopathy Preceding Neurofibrillary Tangles in Mutant Tau Transgenic Mice

    PubMed Central

    Leroy, Karelle; Bretteville, Alexis; Schindowski, Katharina; Gilissen, Emmanuel; Authelet, Michèle; De Decker, Robert; Yilmaz, Zehra; Buée, Luc; Brion, Jean-Pierre

    2007-01-01

    Neurodegenerative diseases characterized by brain and spinal cord involvement often show widespread accumulations of tau aggregates. We have generated a transgenic mouse line (Tg30tau) expressing in the forebrain and the spinal cord a human tau protein bearing two pathogenic mutations (P301S and G272V). These mice developed age-dependent brain and hippocampal atrophy, central and peripheral axonopathy, progressive motor impairment with neurogenic muscle atrophy, and neurofibrillary tangles and had decreased survival. Axonal spheroids and axonal atrophy developed early before neurofibrillary tangles. Neurofibrillary inclusions developed in neurons at 3 months and were of two types, suggestive of a selective vulnerability of neurons to form different types of fibrillary aggregates. A first type of tau-positive neurofibrillary tangles, more abundant in the forebrain, were composed of ribbon-like 19-nm-wide filaments and twisted paired helical filaments. A second type of tau and neurofilament-positive neurofibrillary tangles, more abundant in the spinal cord and the brainstem, were composed of 10-nm-wide neurofilaments and straight 19-nm filaments. Unbiased stereological analysis indicated that total number of pyramidal neurons and density of neurons in the lumbar spinal cord were not reduced up to 12 months in Tg30tau mice. This Tg30tau model thus provides evidence that axonopathy precedes tangle formation and that both lesions can be dissociated from overt neuronal loss in selected brain areas but not from neuronal dysfunction. PMID:17690183

  16. Transgenic mouse model for estrogen-regulated lipoprotein metabolism: studies on apoVLDL-II expression in transgenic mice.

    PubMed

    Zsigmond, E; Nakanishi, M K; Ghiselli, F E; Chan, L

    1995-07-01

    We have produced transgenic mice that express an estrogen-responsive avian apolipoprotein, apoVLDL-II. An apoVLDL-II natural gene construct containing 4.7 kb of 5' flanking and 19 bp of 3' flanking sequences together with the 4 exon/3 intron structural gene was expressed in a liver-specific manner in transgenic mice. A single injection of estrogen caused a 5.9- to 7.5-fold stimulation of apoVLDL-II mRNA in the liver. The transgene mRNA had the same initiation sites of transcription as the native mRNA isolated from laying hen liver, and the same sites were used before and after estrogen treatment. The number of hepatocytes that stain positive for immunoreactive apoVLDL-II increased from < 1% to 40-60% in 24 h after estrogen treatment. Thus, in trangenic mice as in the cockerel, hepatocytes are biochemically heterogeneous and induction of apoVLDL-II synthesis occurs by recruitment of hepatocytes. In the plamsa compartment, compared to controls, transgenic mice have a 3- to 5-fold higher basal total plasma triglyceride which was accounted for by a 5.4-fold high basal VLDL triglyceride. Estrogen treatment results in a approximately 2-fold increase in the VLDL triglycerides over basal levels and 8.5-fold increase over nontransgenic mice, which did not show any change in VLDL in response to estrogen. Transgenic mice with the integrated apoVLDL-II gene provide a useful model for the study of the regulation of lipoprotein metabolism by estrogen. PMID:7595069

  17. Enhanced spontaneous locomotor activity in bovine GH transgenic mice involves peripheral mechanisms.

    PubMed

    Bohlooly-Y, M; Olsson, B; Gritli-Linde, A; Brusehed, O; Isaksson, O G; Ohlsson, C; Söderpalm, B; Törnell, J; Ola, B

    2001-10-01

    Clinical and experimental studies indicate a role for GH in mechanisms related to anhedonia/hedonia, psychic energy, and reward. Recently we showed that transgenic mice with general overexpression of bovine GH display increased spontaneous locomotor activity. In the present study, we investigated whether this behavioral change is owing to a direct action of GH in the central nervous system or to peripheral GH actions. A transgenic construct, containing the glial fibrillary acidic protein promoter directing specific expression of bovine GH to the central nervous system, was designed. The central nervous system-specific expression of bovine GH in the glial fibrillary acidic protein-bovine GH transgenic mice was confirmed, but no effect on spontaneous locomotor activity was observed. Serum bovine GH levels were increased in glial fibrillary acidic protein-bovine GH transgenic mice but clearly lower than in transgenic mice with general overexpression of bovine GH. In contrast to the transgenic mice with general overexpression of bovine GH, glial fibrillary acidic protein-bovine GH mice did not display any difference in serum IGF-I levels. The levels of free T(3) and the conversion of the free T(4) to free T(3) were only increased in transgenic mice with general overexpression of bovine GH, but serum corticosterone levels were similarly increased in both transgenic models. These results suggest that free T(3) and/or IGF-I, affecting dopamine and serotonin systems in the central nervous system, may mediate the enhanced locomotor activity observed in transgenic mice with general overexpression of bovine GH. PMID:11564723

  18. INCREASED LIVER PATHOLOGY IN HEPATITIS C VIRUS TRANSGENIC MICE EXPRESSING THE HEPATITIS B VIRUS X PROTEIN

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenic mice expressing the full-length HCV coding sequence were crossed with mice that express the HBV X gene-encoded regulatory protein HBx (ATX mice) to test the hypothesis that HBx expression accelerates HCV-induced liver pathogenesis. At 16 months (mo) of age, hepatocellular carcinoma was id...

  19. Multiple effects of genetic background on variegated transgene expression in mice.

    PubMed Central

    Opsahl, Margaret L; McClenaghan, Margaret; Springbett, Anthea; Reid, Sarah; Lathe, Richard; Colman, Alan; Whitelaw, C Bruce A

    2002-01-01

    BLG/7 transgenic mice express an ovine beta-lactoglobulin transgene during lactation. Unusually, transgene expression levels in milk differ between siblings. This variable expression is due to variegated transgene expression in the mammary gland and is reminiscent of position-effect variegation. The BLG/7 line was created and maintained on a mixed CBA x C57BL/6 background. We have investigated the effect on transgene expression of backcrossing for 13 generations into these backgrounds. Variable transgene expression was observed in all populations examined, confirming that it is an inherent property of the transgene array at its site of integration. There were also strain-specific effects on transgene expression that appear to be independent of the inherent variegation. The transgene, compared to endogenous milk protein genes, is specifically susceptible to inbreeding depression. Outcrossing restored transgene expression levels to that of the parental population; thus suppression was not inherited. Finally, no generation-dependent decrease in mean expression levels was observed in the parental population. Thus, although the BLG/7 transgene is expressed in a variegated manner, there was no generation-associated accumulated silencing of transgene expression. PMID:11901126

  20. C9ORF72-ALS/FTD: Transgenic Mice Make a Come-BAC.

    PubMed

    Hayes, Lindsey R; Rothstein, Jeffrey D

    2016-05-01

    For five years, since the landmark discovery of the C9ORF72 hexanucleotide repeat expansion in ALS/FTD, a transgenic mouse model has remained elusive. Now, two laboratories (Liu et al., 2016; Jiang et al., 2016) report the development of BAC transgenic mice that recapitulate features of the human disease. PMID:27151634

  1. Enhanced UV-Induced Skin Carcinogenesis in Transgenic Mice Overexpressing Proprotein Convertases1

    PubMed Central

    Fu, Jian; Bassi, Daniel E; Zhang, Jirong; Li, Tianyu; Cai, Kathy Q; Testa, Courtney Lyons; Nicolas, Emmanuelle; Klein-Szanto, Andres J

    2013-01-01

    The proprotein convertases (PCs) furin and PACE4 process numerous substrates involved in tumor growth, invasion, and metastasis. We have previously shown that PCs increase the susceptibility to chemical skin carcinogenesis. Because of the human relevancy of UV radiation in the etiopathogenesis of human skin cancer, we investigated whether or not transgenic mice overexpressing either furin alone or both furin and PACE4 show increased susceptibility to UV carcinogenesis. After backcrossing our previously described furin and PACE4 transgenic lines, targeted to the epidermis, into a SKH-1 background, we exposed both single and double transgenic mice to UV radiation for 34 weeks. The results showed an increase in squamous cell carcinoma (SCC) multiplicity of approximately 70% in the single furin transgenic mouse line SF47 (P < .002) and a 30% increase in the other single transgenic line SF49 when compared to wild-type (WT) SKH-1 mice. Interestingly, there was also an increase in the percentage of high histologic grade SCCs in the transgenic lines compared to the WT mice, i.e., WT = 9%, SF47 = 15%, and SF49 = 26% (P < .02). Targeting both furin and PACE4 to the epidermis in double transgenic mice did not have an additive effect on tumor incidence/multiplicity but did enhance the tumor histopathologic grade, i.e., a significant increase in higher grade SCCs was seen in the bigenic mouse line SPF47 (P < .02). Thus, we observed an increased susceptibility to UV in single furin transgenic mice that was not substantially enhanced in the double furin/PACE4 transgenic mice. PMID:23441131

  2. Plasmacytoma induction in specific pathogen-free (SPF) bcl-2 transgenic BALB/c mice.

    PubMed

    Silva, S; Klein, G

    1999-01-01

    Germ-free (GF) and specific pathogen free (SPF) BALB/c mice are refractory to plasmacytoma induction by pristane (McIntire and Princler, 1969, Byrd et al 1991). It was therefore suggested that MPC development may depend on antigenic stimulation. If so, it may conceivably act by preventing the apoptotic elimination of tumor precursor cells. We have tested this idea by elevating the apoptotic threshold by the introduction of a bcl-2 transgene. We have found that MPCs could be induced by pristane oil in transgene carrying SPF mice. An E mu activated bcl-2 transgene was introduced into SPF BALB/c mice. The mice were used after two backcrosses (BC-2). Pristane oil treatment was started at 4 to 6 weeks of age (3 x 0.3 ml via i.p. at monthly intervals). For each transgene carrier a transgene negative littermate was used as control. Fifteen of 24 (63%) transgene carriers developed plasmacytomas after latency periods between 67 and 146 days (mean = 112 +/- 30 days) after the first pristane injection. Five additional transgene carriers developed lymphoma (3 cases) or mixed MPC and lymphoma (2 cases). In contrast, no tumors developed in 16 transgene negative littermates that were kept > 300 days under observation. Karyotyping showed that 10/15 (66%) of the MPCs carried a T(12;15) translocation, 4/15 (27%) carried both T(12;15) and T(6;15) translocations in the same metaphase plate, and 1/15 (7%) was translocation free. A T(12;15) translocation was also detected in one of the 2 mice with mixed tumor type. Pristane treated bcl-2 transgenic C57B1/6 mice remained tumor free, although T(12;15) translocation carrying cells were found in the peritoneal fluid of 4/20 mice 176 days after pristane. PMID:10396078

  3. Impaired growth of pancreatic exocrine cells in transgenic mice expressing human activin {beta}E subunit

    SciTech Connect

    Hashimoto, Osamu . E-mail: ohashim@vmas.kitasato-u.ac.jp; Ushiro, Yuuki; Sekiyama, Kazunari; Yamaguchi, Osamu; Yoshioka, Kazuki; Mutoh, Ken-Ichiro; Hasegawa, Yoshihisa

    2006-03-10

    Activins, TGF-{beta} superfamily members, have multiple functions in a variety of cells and tissues. Recently, additional activin {beta} subunit genes, {beta}C and {beta}E, have been identified. To explore the role of activin E, we created transgenic mice overexpressing human activin {beta}E subunit. There were pronounced differences in the pancreata of the transgenic animals as compared with their wild-type counterparts. Pancreatic weight, expressed relative to total body weight, was significantly reduced. Histologically, adipose replacement of acini in the exocrine pancreas was observed. There was a significant decrease in the number of PCNA-positive cells in the acinar cells, indicating reduced proliferation in the exocrine pancreas of the transgenic mice. However, quantitative pancreatic morphometry showed that the total number and mass of the islets of the transgenic mice were comparable with those of the nontransgenic control mice. Our findings suggest a role for activin E in regulating the proliferation of pancreatic exocrine cells.

  4. Rapid generation of dendritic cell specific transgenic mice by lentiviral vectors.

    PubMed

    Zhang, Jinyu; Zou, Liyun; Liu, Qin; Li, Jingyi; Zhou, Jingran; Wang, Yong; Li, Na; Liu, Ting; Wei, Hong; Wu, Min; Wan, Ying; Wu, Yuzhang

    2009-12-01

    Dendritic cell (DC) specific transgenic mice are a most important model for investigating dendritic cell functions in vivo. Recently, lentivirus mediated gene transfer has become a powerful and convenient method for generation of transgenic mice. We cloned a 1.2 kb CD11c promoter and constructed a lentiviral vector, which efficiently drove DC-specific expression in vitro. After microinjection of purified virus into the perivitelline space of single-cell embryo, more than 80% newborn mice were transgenic and 7 F0 founders were rapidly generated in 2 months. GFP was strictly expressed in CD11c+ cells in spleens, thymus and lymph nodes of the transgenic mice. Importantly, the physiological characteristics and functions of DCs in the transgenic mice were not altered by the specific expression. These results indicate that this vector could be used to rapidly prepare DC-specific transgenic mice. Thus, this lentiviral vector system may provide a convenient and useful tool to study the properties of DCs in vivo. PMID:19468852

  5. Microglial response to amyloid plaques in APPsw transgenic mice.

    PubMed Central

    Frautschy, S. A.; Yang, F.; Irrizarry, M.; Hyman, B.; Saido, T. C.; Hsiao, K.; Cole, G. M.

    1998-01-01

    Microglial activation is central to the inflammatory response in Alzheimer's Disease (AD). A recently described mouse line, Tg(HuAPP695.K670N/M671L)2576, expressing human amyloid precursor protein with a familial AD gene mutation, age-related amyloid deposits, and memory deficits, was found to develop a significant microglial response using Griffonia simplicifolia lectin or phosphotyrosine probe to identify microglia Both Griffonia simplicifolia lectin and phosphotyrosine staining showed increased numbers of intensely labeled, often enlarged microglia clustered in and around plaques, consistent with microglial activation related to beta-amyloid formation. Using quantitative image analysis of coronal phosphotyrosine-immunostained sections, transgene-positive 10- to 16-month-old, hemizygous, hybrid Tg2576 (APPsw) animals showed significantly increased microglial density and size in plaque-forming areas of hippocampus and frontal, entorhinal, and occipital cortex. Quantitative analysis of microglia as a function of distance from the center of plaques (double labeled for A beta peptide and microglia) revealed highly significant, two- to fivefold elevations in microglial number and area within plaques compared with neighboring regions. Tg2576 beta-amyloid-plaque-forming mice should be a useful system for assessing the consequences of the microglial-mediated inflammatory response to beta-amyloid and developing anti-inflammatory therapeutic strategies for Alzheimer's disease. These results provide the first quantitative link between beta-amyloid plaque formation and microglial activation in an animal model with neuritic plaques and memory deficits. Images Figure 1 Figure 2 PMID:9422548

  6. Generating Transgenic Mice by Lentiviral Transduction of Spermatozoa Followed by In Vitro Fertilization and Embryo Transfer.

    PubMed

    Chandrashekran, Anil; Casimir, Colin; Dibb, Nick; Readhead, Carol; Winston, Robert

    2016-01-01

    Most transgenic technologies rely on the oocyte as a substrate for genetic modification. Transgenics animals are usually generated by the injection of the gene constructs (including lentiviruses encoding gene constructs or modified embryonic stem cells) into the pronucleus of a fertilized egg followed by the transfer of the injected embryos into the uterus of a foster mother. Male germ cells also have potential as templates for transgenic development. We have previously shown that mature sperm can be utilized as template for lentiviral transduction and as such used to generate transgenic mice efficiently with germ line capabilities. We provide here a detailed protocol that is relatively simple, to establish transgenic mice using lentivirally transduced spermatozoa. This protocol employs a well-established lentiviral gene delivery system (usual for somatic cells) delivering a variety of transgenes to be directly used with sperm, and the subsequent use of these modified sperm in in vitro fertilization studies and embryo transfer into foster female mice, for the establishment of transgenic mice. PMID:27317176

  7. High Pathogenicity of Wild-Type Measles Virus Infection in CD150 (SLAM) Transgenic Mice

    PubMed Central

    Sellin, Caroline I.; Davoust, Nathalie; Guillaume, Vanessa; Baas, Dominique; Belin, Marie-Françoise; Buckland, Robin; Wild, T. Fabian; Horvat, Branka

    2006-01-01

    Measles virus (MV) infection causes an acute childhood disease, associated in certain cases with infection of the central nervous system and development of a severe neurological disease. We have generated transgenic mice ubiquitously expressing the human protein SLAM (signaling lymphocytic activation molecule), or CD150, recently identified as an MV receptor. In contrast to all other MV receptor transgenic models described so far, in these mice infection with wild-type MV strains is highly pathogenic. Intranasal infection of SLAM transgenic suckling mice leads to MV spread to different organs and the development of an acute neurological syndrome, characterized by lethargy, seizures, ataxia, weight loss, and death within 3 weeks. In addition, in this model, vaccine and wild-type MV strains can be distinguished by virulence. Furthermore, intracranial MV infection of adult transgenic mice generates a subclinical infection associated with a high titer of MV-specific antibodies in the serum. Finally, to analyze new antimeasles therapeutic approaches, we created a recombinant soluble form of SLAM and demonstrated its important antiviral activity both in vitro and in vivo. Taken together, our results show the high susceptibility of SLAM transgenic mice to MV-induced neurological disease and open new perspectives for the analysis of the implication of SLAM in the neuropathogenicity of other morbilliviruses, which also use this molecule as a receptor. Moreover, this transgenic model, in allowing a simple readout of the efficacy of an antiviral treatment, provides unique experimental means to test novel anti-MV preventive and therapeutic strategies. PMID:16775330

  8. Caspase 6 has a protective role in SOD1(G93A) transgenic mice.

    PubMed

    Hogg, Marion C; Mitchem, Mollie R; König, Hans-Georg; Prehn, Jochen H M

    2016-06-01

    In amyotrophic lateral sclerosis (ALS), it has been suggested that the process of neurodegeneration starts at the neuromuscular junction and is propagated back along axons towards motor neurons. Caspase-dependent pathways are well established as a cause of motor neuron death, and recent work in other disease models indicated a role for caspase 6 in axonal degeneration. Therefore we hypothesised that caspase 6 may be involved in motor neuron death in ALS. To investigate the role of caspase 6 in ALS we profiled protein levels of caspase-6 throughout disease progression in the ALS mouse model SOD1(G93A); this did not reveal differences in caspase 6 levels during disease. To investigate the role of caspase 6 further we generated a colony with SOD1(G93A) transgenic mice lacking caspase 6. Analysis of the transgenic SOD1(G93A); Casp6(-/-) revealed an exacerbated phenotype with motor dysfunction occurring earlier and a significantly shortened lifespan when compared to transgenic SOD1(G93A); Casp6(+/+) mice. Immunofluorescence analysis of the neuromuscular junction revealed no obvious difference between caspase 6(+/+) and caspase 6(-/-) in non-transgenic mice, while the SOD1(G93A) transgenic mice showed severe degeneration compared to non-transgenic mice in both genotypes. Our data indicate that caspase-6 does not exacerbate ALS pathogenesis, but may have a protective role. PMID:26976329

  9. Fertility comparison between wild type and transgenic mice by in vitro fertilization

    PubMed Central

    Vasudevan, Kuzhalini; Raber, James

    2011-01-01

    Transgenic mice are increasingly used as animal models for studies of gene function and regulation of mammalian genes. Although there has been continuous and remarkable progress in the development of transgenic technology over several decades, many aspects of the resulting transgenic model’s phenotype cannot be completely predicted. For example, it is well known that as a consequence of the random insertion of the injected DNA construct, several founder mice of the new line need to be analyzed for possible differences in phenotype secondary to different insertion sites. The Knock out technique for transgenic production disrupts a specific gene by insertion or homologous recombination creating a null expression or replacement of the gene with a marker to localize it expression. This modification could result in pleiotropic phenotype if the gene is also expressed in tissues other than the target organs. Although the future breeding performance of the newly created model is critical to many studies, it is rarely anticipated that the new integrations could modify the reproductive profile of the new transgenic line. To date, few studies have demonstrated the difference between the parent strain’s reproductive performance and the newly developed transgenic model. This study was designed to determine whether a genetic modification, knock out (KO) or transgenics, not anticipated to affect reproductive performance could affect the resulting reproductive profile of the newly developed transgenic mouse. More specifically, this study is designed to study the impact of the genetic modification on the ability of gametes to be fertilized in vitro. We analyzed the reproductive performance of mice with different background strains: FVB/N, C57BL/6 (129Sv/J × C57Bl/6)F1 and outbred CD1® and compared them to mice of the same strain carrying a transgene or KO which was not anticipated to affect fertility. In vitro Fertilization was used to analyze the fertility of the mice

  10. Interleukin-12 inhibits hepatitis B virus replication in transgenic mice.

    PubMed Central

    Cavanaugh, V J; Guidotti, L G; Chisari, F V

    1997-01-01

    Interleukin-12 (IL-12) is a heterodimeric cytokine produced by antigen-presenting cells that has the ability to induce gamma interferon (IFN-gamma) secretion by T and natural killer cells and to generate normal Th1 responses. These properties suggest that IL-12 may play an important role in the immune response to many viruses, including hepatitis B virus (HBV). Recently, we have shown that HBV-specific cytotoxic T lymphocytes inhibit HBV replication in the livers of transgenic mice by a noncytolytic process that is mediated in part by IFN-gamma. In the current study, we demonstrated that the same antiviral response can be initiated by recombinant murine IL-12 and we showed that the antiviral effect of IL-12 extends to extrahepatic sites such as the kidney. Southern blot analyses revealed the complete disappearance of HBV replicative intermediates from liver and kidney tissues at IL-12 doses that induce little or no inflammation in these tissues. In addition, immunohistochemical analysis demonstrated the disappearance of cytoplasmic hepatitis B core antigen from both tissues after IL-12 treatment, suggesting that IL-12 either prevents the assembly or triggers the degradation of the nucleocapsid particles within which HBV replication occurs. Importantly, we demonstrated that although IFN-gamma, tumor necrosis factor alpha, and IFN-alpha/beta mRNA are induced in the liver and kidney after IL-12 administration, the antiviral effect of IL-12 is mediated principally by its ability to induce IFN-gamma production in this model. These results suggest that IL-12, through its ability to induce IFN-gamma, probably plays an important role in the antiviral immune response to HBV during natural infection. Further, since relatively nontoxic doses of recombinant IL-12 profoundly inhibit HBV replication in the liver and extrahepatic sites in this model, IL-12 may have therapeutic value as an antiviral agent for the treatment of chronic HBV infection. PMID:9060687

  11. High-level hepatitis B virus replication in transgenic mice.

    PubMed Central

    Guidotti, L G; Matzke, B; Schaller, H; Chisari, F V

    1995-01-01

    Hepatitis B virus (HBV) transgenic mice whose hepatocytes replicate the virus at levels comparable to that in the infected livers of patients with chronic hepatitis have been produced, without any evidence of cytopathology. High-level viral gene expression was obtained in the liver and kidney tissues in three independent lineages. These animals were produced with a terminally redundant viral DNA construct (HBV 1.3) that starts just upstream of HBV enhancer I, extends completely around the circular viral genome, and ends just downstream of the unique polyadenylation site in HBV. In these animals, the viral mRNA is more abundant in centrilobular hepatocytes than elsewhere in the hepatic lobule. High-level viral DNA replication occurs inside viral nucleocapsid particles that preferentially form in the cytoplasm of these centrilobular hepatocytes, suggesting that an expression threshold must be reached for nucleocapsid assembly and viral replication to occur. Despite the restricted distribution of the viral replication machinery in centrilobular cytoplasmic nucleocapsids, nucleocapsid particles are detectable in the vast majority of hepatocyte nuclei throughout the hepatic lobule. The intranuclear nucleocapsid particles are empty, however, suggesting that viral nucleocapsid particle assembly occurs independently in the nucleus and the cytoplasm of the hepatocyte and implying that cytoplasmic nucleocapsid particles do not transport the viral genome across the nuclear membrane into the nucleus during the viral life cycle. This model creates the opportunity to examine the influence of viral and host factors on HBV pathogenesis and replication and to assess the antiviral potential of pharmacological agents and physiological processes, including the immune response. PMID:7666518

  12. Cardiac hypertrophy associated with myeloproliferative neoplasms in JAK2V617F transgenic mice

    PubMed Central

    2014-01-01

    Background Myeloproliferative neoplasms (MPNs) are blood malignancies manifested in increased production of red blood cells, white blood cells, and/or platelets. A major molecular lesion associated with the diseases is JAK2V617F, an activation mutation form of tyrosine kinase JAK2. Cardiovascular events represent the leading cause of morbidity and mortality associated MPNs, but the underlying mechanism is not well understood. Methods Previously, we generated JAK2V617F transgenic mice which displayed MPN-like phenotypes. In the present study, we further characterized these mice by analyzing the time course of MPN phenotype development and associated cardiac abnormalities. We performed detailed histochemical staining of cardiac sections. Results JAK2V617F transgenic mice developed cardiomegaly as a subsequent event of increased blood cell production during the course of MPN phenotype development. The cardiomegaly is manifested in increased ventricular wall thickness and enlarged cardiomyocytes. Trichrome and reticulin staining revealed extensive collagen fibrosis in the heart of JAK2V617F transgenic mice. Thrombosis in the coronary artery and inflammatory cell infiltration into cardiac muscle were also observed in JAK2V617F transgenic mice, and the latter event was accompanied by fibrosis. Conclusion JAK2V617F-induced blood disorders have a major impact on heart function and lead to cardiac hypertrophy. JAK2V617F transgenic mice represent an excellent model system to study both hematological malignancies and cardiovascular diseases. PMID:24646493

  13. Increased liver pathology in hepatitis C virus transgenic mice expressing the hepatitis B virus X protein

    SciTech Connect

    Keasler, Victor V.; Lerat, Herve; Madden, Charles R.; Finegold, Milton J.; McGarvey, Michael J.; Mohammed, Essam M.A.; Forbes, Stuart J.; Lemon, Stanley M.; Hadsell, Darryl L.; Grona, Shala J.; Hollinger, F. Blaine; Slagle, Betty L. . E-mail: bslagle@bcm.edu

    2006-04-10

    Transgenic mice expressing the full-length HCV coding sequence were crossed with mice that express the HBV X gene-encoded regulatory protein HBx (ATX mice) to test the hypothesis that HBx expression accelerates HCV-induced liver pathogenesis. At 16 months (mo) of age, hepatocellular carcinoma was identified in 21% of HCV/ATX mice, but in none of the single transgenic animals. Analysis of 8-mo animals revealed that, relative to HCV/WT mice, HCV/ATX mice had more severe steatosis, greater liver-to-body weight ratios, and a significant increase in the percentage of hepatocytes staining for proliferating cell nuclear antigen. Furthermore, primary hepatocytes from HCV, ATX, and HCV/ATX transgenic mice were more resistant to fas-mediated apoptosis than hepatocytes from nontransgenic littermates. These results indicate that HBx expression contributes to increased liver pathogenesis in HCV transgenic mice by a mechanism that involves an imbalance in hepatocyte death and regeneration within the context of severe steatosis.

  14. A foreign dihydrofolate reductase gene in transgenic mice acts as a dominant mutation.

    PubMed Central

    Gordon, J W

    1986-01-01

    We have produced 17 lines of transgenic mice by microinjecting a full-length cDNA clone of an altered dihydrofolate reductase (dhfr) gene. The protein specified by this gene carries a point mutation which triples its Km for dihydrofolate and reduces substrate turnover 20-fold relative to the wild-type enzyme. Transgenic mice from different pedigrees, several of which carry a single copy of this gene in different integration sites, manifest an array of similar developmental abnormalities including growth stunting, reduced fertility, pigmentation changes, and skeletal defects. These defects appear in animals heterozygous for the foreign gene. RNA analyses demonstrate significant expression of the cDNA in newborn mice and adult tissues. These findings show that the additional dhfr gene exerts its mutational effects in a dominant fashion, and therefore the data indicate that transgenic mice can serve as models for elucidating mechanisms of dominant mutagenesis. Images PMID:3785192

  15. Establishment of transgenic mice carrying the gene of human nuclear receptor NR5A2 (hB1F)

    PubMed Central

    Wang, Shui-Liang; Yang, Hua; Xie, You-Hua; Wang, Yuan; Li, Jian-Zhong; Wang, Long; Wang, Zhu-Gang; Fu, Ji-Liang

    2003-01-01

    AIM: Human hepatitis B virus enhancer II B1 binding factor (hB1F) was cloned and characterized as a novel member of the Ftz-F1 (NR5A) nuclear receptor subfamily. Although progresses have recently been made, its biological function remains largely unidentified. The aim of this study was to establish an hB1F transgenic mouse model to promote the functional study of hB1F. METHODS: Transgene fragments were microinjected into fertilized eggs of mice. The manipulated embryos were transferred into the oviducts of pseudopregnant female mice. The offsprings were identified by PCR and Southern blot analysis. Transgene expression was analyzed with RT-PCR and Western blot analysis. Transgenic founder mice were used to establish transgenic mouse lineages. The F1 and F2 mice were identified by PCR analysis. RESULTS: Seven mice were identified as carrying copies of transgene. RT-PCR and Western blotting results showed that the transgene was expressed in heart, liver, lung, kidney and stomach in one of the transgenic mouse lineages. Genetic analysis of the transgenic mice demonstrated that the transgene was integrated into the chromosome at a single site, and was transmitted stably. CONCLUSION: In this study we established an hB1F transgenic mouse model, which will facilitate the investigation of the biological function of hB1F in vivo. PMID:12800251

  16. Quantitative Genetics of Transgenic Mice: Components of Phenotypic Variation in Body Weights and Weight Gains

    PubMed Central

    Clutter, A. C.; Pomp, D.; Murray, J. D.

    1996-01-01

    Transgenic mice possessing an ovine growth hormone gene were used to study the effects of elevated growth hormone on quantitative genetic variation. Males hemizygous for the transgene were mated to wild-type females to produce half- and full-sib families in which approximately half the progeny were transgenic and half were wild type. Analyses of body weights at 3-10 weeks, and weight gains from 3 to 6, and 6 to 10 weeks produced estimates of the proportion of total variance due to additive genetic effects (h(2)) and common litter effects (c(2)), and the genetic correlation between transgenic and wild-type expression of each trait. At 10 weeks, body weight of transgenics exceeded that of wild types by 26 and 49% in males and females, respectively. Estimated genetic variances in the transgenic group were significantly greater than zero for body weights at most ages and for both measurements of gain. Common litter effects accounted for a similar proportion of variation in the wild-type and transgenic groups. Additive genetic correlations between wild-type and transgenic expression of body weights tended to decline with age, indicating that a partially different array of genes may have begun to affect body weight in the transgenic group. PMID:8844161

  17. CTRP9 transgenic mice are protected from diet-induced obesity and metabolic dysfunction

    PubMed Central

    Peterson, Jonathan M.; Wei, Zhikui; Seldin, Marcus M.; Byerly, Mardi S.; Aja, Susan

    2013-01-01

    CTRP9 is a secreted multimeric protein of the C1q family and the closest paralog of the insulin-sensitizing adipokine, adiponectin. The metabolic function of this adipose tissue-derived plasma protein remains largely unknown. Here, we show that the circulating levels of CTRP9 are downregulated in diet-induced obese mice and upregulated upon refeeding. Overexpressing CTRP9 resulted in lean mice that dramatically resisted weight gain induced by a high-fat diet, largely through decreased food intake and increased basal metabolism. Enhanced fat oxidation in CTRP9 transgenic mice resulted from increases in skeletal muscle mitochondrial content, expression of enzymes involved in fatty acid oxidation (LCAD and MCAD), and chronic AMPK activation. Hepatic and skeletal muscle triglyceride levels were substantially decreased in transgenic mice. Consequently, CTRP9 transgenic mice had a greatly improved metabolic profile with markedly reduced fasting insulin and glucose levels. The high-fat diet-induced obesity, insulin resistance, and hepatic steatosis observed in wild-type mice were prevented in transgenic mice. Consistent with the in vivo data, recombinant protein significantly enhanced fat oxidation in L6 myotubes via AMPK activation and reduced lipid accumulation in H4IIE hepatocytes. Collectively, these data establish CTRP9 as a novel metabolic regulator and a new component of the metabolic network that links adipose tissue to lipid metabolism in skeletal muscle and liver. PMID:23842676

  18. Transgenic knockout mice with exclusively human sickle hemoglobinand sickle cell disease

    SciTech Connect

    Paszty, C.; Brion, C.; Manci, E.; Witkowska, E.; Stevens, M.; Narla, M.; Rubin, E.

    1997-06-13

    To create mice expressing exclusively human sicklehemoglobin (HbS), transgenic mice expressing human alpha-, gamma-, andbeta[S]-globin were generated and bred with knockout mice that haddeletions of the murine alpha- and beta-globin genes. These sickle cellmice have the major features (irreversibly sickled red cells, anemia,multiorgan pathology) found in humans with sickle cell disease and, assuch, represent a useful in vivo system to accelerate the development ofimproved therapies for this common genetic disease.

  19. Overexpression of Thioredoxin in Transgenic Mice Attenuates Focal Ischemic Brain Damage

    NASA Astrophysics Data System (ADS)

    Takagi, Yasushi; Mitsui, Akira; Nishiyama, Akira; Nozaki, Kazuhiko; Sono, Hiroshi; Gon, Yasuhiro; Hashimoto, Nobuo; Yodoi, Junji

    1999-03-01

    Thioredoxin (TRX) plays important biological roles both in intra- and extracellular compartments, including in regulation of various intracellular molecules via thiol redox control. We produced TRX overexpressing mice and confirmed that there were no anatomical and physiological differences between wild-type (WT) mice and TRX transgenic (Tg) mice. In the present study we subjected mice to focal brain ischemia to shed light on the role of TRX in brain ischemic injury. At 24 hr after middle cerebral artery occlusion, infarct areas and volume were significantly smaller in Tg mice than in WT mice. Moreover neurological deficit was ameliorated in Tg mice compared with WT mice. Protein carbonyl content, a marker of cellular protein oxidation, in Tg mice showed less increase than did that of WT mice after the ischemic insult. Furthermore, c-fos expression in Tg mice was stronger than in WT mice 1 hr after ischemia. Our results suggest that transgene expression of TRX decreased ischemic neuronal injury and that TRX and the redox state modified by TRX play a crucial role in brain damage during stroke.

  20. Neurologic disease induced in transgenic mice by cerebral overexpression of interleukin 6.

    PubMed Central

    Campbell, I L; Abraham, C R; Masliah, E; Kemper, P; Inglis, J D; Oldstone, M B; Mucke, L

    1993-01-01

    Cytokines are thought to be important mediators in physiologic and pathophysiologic processes affecting the central nervous system (CNS). To explore this hypothesis, transgenic mice were generated in which the cytokine interleukin 6 (IL-6), under the regulatory control of the glial fibrillary acidic protein gene promoter, was overexpressed in the CNS. A number of transgenic founder mice and their offspring exhibited a neurologic syndrome the severity of which correlated with the levels of cerebral IL-6 expression. Transgenic mice with high levels of IL-6 expression developed severe neurologic disease characterized by runting, tremor, ataxia, and seizure. Neuropathologic manifestations included neuro-degeneration, astrocytosis, angiogenesis, and induction of acute-phase-protein production. These findings indicate that cytokines such as IL-6 can have a direct pathogenic role in inflammatory, infectious, and neurodegenerative CNS diseases. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:7694279

  1. APP transgenic mice for modelling behavioral and psychological symptoms of dementia (BPSD)

    PubMed Central

    Lalonde, R.; Fukuchi, K.; Strazielle, C.

    2012-01-01

    The discovery of gene mutations responsible for autosomal dominant Alzheimer's disease has enabled researchers to reproduce in transgenic mice several hallmarks of this disorder, notably Aβ accumulation, though in most cases without neurofibrillary tangles. Mice expressing mutated and wild-type APP as well as C-terminal fragments of APP exhibit variations in exploratory activity reminiscent of behavioral and psychological symptoms of Alzeimer dementia (BPSD). In particular, open-field, spontaneous alternation, and elevated plus-maze tasks as well as aggression are modified in several APP transgenic mice relative to non-transgenic controls. However, depending on the precise murine models, changes in open-field and elevated plus-maze exploration occur in either direction, either increased or decreased relative to controls. It remains to be determined which neurotransmitter changes are responsible for this variability, in particular with respect to GABA, 5HT, and dopamine. PMID:22373961

  2. Neural differentiation of adipose-derived stem cells isolated from GFP transgenic mice

    SciTech Connect

    Fujimura, Juri; E-mail: juri-f@nms.ac.jp; Ogawa, Rei; Mizuno, Hiroshi; Fukunaga, Yoshitaka; Suzuki, Hidenori

    2005-07-22

    Taking advantage of homogeneously marked cells from green fluorescent protein (GFP) transgenic mice, we have recently reported that adipose-derived stromal cells (ASCs) could differentiate into mesenchymal lineages in vitro. In this study, we performed neural induction using ASCs from GFP transgenic mice and were able to induce these ASCs into neuronal and glial cell lineages. Most of the neurally induced cells showed bipolar or multipolar appearance morphologically and expressed neuronal markers. Electron microscopy revealed their neuronal morphology. Some cells also showed glial phenotypes, as shown immunocytochemically. The present study clearly shows that ASCs derived from GFP transgenic mice differentiate into neural lineages in vitro, suggesting that these cells might provide an ideal source for further neural stem cell research with possible therapeutic application for neurological disorders.

  3. Severe B cell hyperplasia and autoimmune disease in TALL-1 transgenic mice

    PubMed Central

    Khare, Sanjay D.; Sarosi, Ildiko; Xia, Xing-Zhong; McCabe, Susan; Miner, Kent; Solovyev, Irina; Hawkins, Nessa; Kelley, Michael; Chang, David; Van, Gwyneth; Ross, Larry; Delaney, John; Wang, Ling; Lacey, David; Boyle, William J.; Hsu, Hailing

    2000-01-01

    TALL-1/Blys/BAFF is a member of the tumor necrosis factor (TNF) ligand superfamily that is functionally involved in B cell proliferation. Here, we describe B cell hyperplasia and autoimmune lupus-like changes in transgenic mice expressing TALL-1 under the control of a β-actin promoter. The TALL-1 transgenic mice showed severe enlargement of spleen, lymph nodes, and Peyer's patches because of an increased number of B220+ cells. The transgenic mice also had hypergammaglobulinemia contributed by elevations of serum IgM, IgG, IgA, and IgE. In addition, a phenotype similar to autoimmune lupus-like disease was also seen in TALL-1 transgenic mice, characterized by the presence of autoantibodies to nuclear antigens and immune complex deposits in the kidney. Prolonged survival and hyperactivity of transgenic B cells may contribute to the autoimmune lupus-like phenotype in these animals. Our studies further confirm TALL-1 as a stimulator of B cells that affect Ig production. Thus, TALL-1 may be a primary mediator in B cell-associated autoimmune diseases. PMID:10716715

  4. Effect of Hypertriglyceridemia on Beta Cell Mass and Function in ApoC3 Transgenic Mice.

    PubMed

    Liu, Yun-Zi; Cheng, Xiaoyun; Zhang, Ting; Lee, Sojin; Yamauchi, Jun; Xiao, Xiangwei; Gittes, George; Qu, Shen; Jiang, Chun-Lei; Dong, H Henry

    2016-07-01

    Hypertriglyceridemia results from increased production and decreased clearance of triglyceride-rich very low-density lipoproteins, a pathological condition that accounts for heightened risk of ischemic vascular diseases in obesity and type 2 diabetes. Despite its intimate association with insulin resistance, whether hypertriglyceridemia constitutes an independent risk for beta cell dysfunction in diabetes is unknown. Answering this fundamental question is stymied by the fact that hypertriglyceridemia is intertwined with hyperglycemia and insulin resistance in obese and diabetic subjects. To circumvent this limitation, we took advantage of apolipoprotein C3 (ApoC3)-transgenic mice, a model with genetic predisposition to hypertriglyceridemia. We showed that ApoC3-transgenic mice, as opposed to age/sex-matched wild-type littermates, develop hypertriglyceridemia with concomitant elevations in plasma cholesterol and non-esterified fatty acid levels. Anti-insulin and anti-glucagon dual immunohistochemistry in combination with morphometric analysis revealed that ApoC3-transgenic and wild-type littermates had similar beta cell and alpha cell masses as well as islet size and architecture. These effects correlated with similar amplitudes of glucose-stimulated insulin secretion and similar degrees of postprandial glucose excursion in ApoC3-transgenic versus wild-type littermates. Oil Red O histology did not visualize lipid infiltration into islets, correlating with the lack of ectopic triglyceride and cholesterol depositions in the pancreata of ApoC3-transgenic versus wild-type littermates. ApoC3-transgenic mice, despite persistent hypertriglyceridemia, maintained euglycemia under both fed and fasting conditions without manifestation of insulin resistance and fasting hyperinsulinemia. Thus, hypertriglyceridemia per se is not an independent risk factor for beta cell dysfunction in ApoC3 transgenic mice. PMID:27226540

  5. Tiam1 Transgenic Mice Display Increased Tumor Invasive and Metastatic Potential of Colorectal Cancer after 1,2-Dimethylhydrazine Treatment

    PubMed Central

    Yu, Li-Na; Zhang, Qing-Ling; Li, Xin; Hua, Xing; Cui, Yan-Mei; Zhang, Nian-Jie; Liao, Wen-Ting; Ding, Yan-Qing

    2013-01-01

    Background T lymphoma invasion and metastasis 1 (Tiam1) is a potential modifier of tumor development and progression. Our previous study in vitro and in nude mice suggested a promotion role of Tiam1 on invasion and metastasis of colorectal cancer (CRC). In the present study, we generated Tiam1/C1199-CopGFP transgenic mice to investigate the tumorigenetic, invasive and metastatic alterations in the colon and rectum of wild-type and Tiam1 transgenic mice under 1,2-dimethylhydrazine (DMH) treatment. Methods Transgenic mice were produced by the method of pronuclear microinlectlon. Whole-body fluorescence imaging (Lighttools, Edmonton, Alberta, Canada), PCR, and immunohistochemical techniques (IHC) were applied sequentially to identify the transgenic mice. The carcinogen DMH (20 mg/kg) was used to induce colorectal tumors though intraperitoneal (i.p.) injections once a week for 24 weeks from the age of 4 weeks on Tiam1 transgenic or non-transgenic mice. Results We successfully generated Tiam1/C1199-CopGFP transgenic mice and induced primary tumors in the intestine of both wild type and Tiam1 transgenic mice by DMH treatment. In addition, Tiam1 transgenic mice developed larger and more aggressive neoplasm than wild-type mice. Moreover, immunohistochemical staining revealed that upregulation of Tiam1 was correlated with increased expression of β-Catenin and Vimentin, and downregulation of E-Cadherin in these mice. Conclusions Our study has provided in vivo evidence supporting that Tiam1 promotes invasion and metastasis of CRC, most probably through activation of Wnt/β-catenin signaling pathway, in a Tiam1 transgenic mouse model. PMID:24069171

  6. [PHARMACOLOGICAL CORRECTION OF APOPTOSIS LEVEL OF CORTICAL NEURONS IN AGED HER2/NEU TRANSGENIC MICE].

    PubMed

    Bazhanova, E D; Kozlova, Yu O; Anisimov, V N; Sukhanov, D S; Teply, D L

    2016-01-01

    Neurodegenerative changes and neuronal death are the basis for development of the nervous system aging. We investigated the mechanism of apoptosis of the sensorimotor cortex neurons of transgenic mice HER2/neu during aging, changes in the cortex function and the participation of exogenous neurometabolites (cytoflavin, piracetam) in regulation of neuronal death and locomotor and psycho-emotional status of mice. The level of apoptosis and expression of apoptosis markers (TUNEL, immunohistochemistry, Western blotting) in HER2/neu transgenic mice as compared to wild type mice (FBV line) were determined. In aging FBV mice the basal activity was shown to decrease and anxiety to increase correlating with the high level of neuronal apoptosis. We identified behavioral characteristics of transgenic HER2/neu mice and found that their low basal activity does not change with aging. Previously we have shown that in this strain of mice the apoptosis level is low, without any age-related changes, due to the suppression, first of all, of the p53-dependent pathway by HER2 (tyrosine kinase receptor) overexpression. Cytoflavin and piracetam were revealed to possess a marked neuroprotective effect, preserving and restoring functions of the nervous system (improving locomotion and psychological status) in both strains of mice. The effect of neurometabolites studied on neuronal apoptosis is ambiguous. In case of its low level it is a moderate stumulation of apoptosis via the external p53-dependent pathways with activation of caspase-3 in transgenic HER2/neu mice with high carcinogenesis level that can possibly prevent tumor development. On the contrary, in old wild-type animals we observed a significant decrease of age-dependent apoptosis level (by stimulating expression of the anti-apoptotic protein Mcl-1), which prevents neurodegeneration. PMID:27220241

  7. Muscle-specific transgenic expression of porcine myostatin propeptide enhances muscle growth in mice.

    PubMed

    Wang, Kaiyun; Li, Zicong; Li, Yang; Zeng, Jinyong; He, Chang; Yang, Jinzeng; Liu, Dewu; Wu, Zhenfang

    2013-10-01

    Myostatin is a well-known negative regulator of skeletal muscle growth. Inhibition of myostatin activity results in increased muscle mass. Myostatin propeptide, as a myostatin antagonist, could be applied to promote meat production in livestock such as pigs. In this study, we generated a transgenic mouse model expressing porcine myostatin propeptide under the control of muscle-specific regulatory elements. The mean body weight of transgenic mice from a line expressing the highest level of porcine myostatin propeptide was increased by 5.4 % (P = 0.023) and 3.2 % (P = 0.031) in males and females, respectively, at 8 weeks of age. Weight of carcass, fore limb and hind limb was respectively increased by 6.0 % (P = 0.038), 9.0 % (P = 0.014), 8.7 % (P = 0.036) in transgenic male mice, compared to wild-type male controls at the age of 9 weeks. Similarly, carcass, fore limb and hind limb of transgenic female mice was 11.4 % (P = 0.002), 14.5 % (P = 0.006) and 14.5 % (P = 0.03) respectively heavier than that of wild-type female mice. The mean cross-section area of muscle fiber was increased by 17 % (P = 0.002) in transgenic mice, in comparison with wild-type controls. These results demonstrated that porcine myostatin propeptide is effective in enhancement of muscle growth. The present study provided useful information for future study on generation of transgenic pigs overexpressing porcine myostatin propeptide for improvement of muscle mass. PMID:23543410

  8. Generation of transgenic mice with liver-specific expression of human nuclear receptor nr5a2.

    PubMed

    Wang, Shui-Liang; Yang, Hua; Xie, You-Hua; Wang, Yuan; Li, Jian-Zhong; Wang, Long; Wang, Zhu-Gang; Fu, Ji-Liang

    2005-12-01

    Human nuclear receptor hb1 f(nr5a2) was cloned and characterized as a novel member of the Ftz-F1 (nr5a) nuclear receptor subfamily,whose its biological function remained largely unidentified. The aim of this study was to establish transgenic mouse model that specifically expressed hB1F in the liver to faciliate the functional study of hB1F. hb1f cDNA was placed downstream of mouse albumin gene enhancer/promoter to construct a liver-specific hb1f expression vector. Transgene fragments were microinjected into fertilized eggs of mice. The manipulated embryos were transferred into the oviducts of pseudopregnant female mice. Four offspring were identified as carrying the transgenes by PCR,from which one was also verified by Southern blotting. RT-PCR and Western blotting results showed that the transgene was expressed in the liver of the transgenic mice. Transgenic founder mice were used to establish transgenic mouse lineages. The F1 mice were identified by PCR analysis. Genetic analysis of the transgenic mice demonstrated that the transgene had been integrated into the chromosome at a single site and could be stably transmitted. PMID:16459652

  9. Expression of Cartilage Developmental Genes in Hoxc8- and Hoxd4-Transgenic Mice

    PubMed Central

    Kruger, Claudia; Kappen, Claudia

    2010-01-01

    Hox genes encode transcription factors, which regulate skeletal patterning and chondrocyte differentiation during the development of cartilage, the precursor to mature bone. Overexpression of the homeobox transcription factors Hoxc8 and Hoxd4 causes severe cartilage defects due to delay in cartilage maturation. Matrix metalloproteinases (MMPs), bone morphogenetic proteins (BMPs) and fibroblastic growth factors (FGFs) are known to play important roles in skeletal development and endochondral bone formation and remodeling. In order to investigate whether these molecules are aberrantly expressed in Hoxc8- and/or Hoxd4-transgenic cartilage, we performed quantitative RT-PCR on chondrocytes from Hox-transgenic mice. Gene expression levels of Bmp4, Fgf8, Fgf10, Mmp9, Mmp13, Nos3, Timp3, Wnt3a and Wnt5a were altered in Hoxc8-transgenic chondrocytes, and Fgfr3, Ihh, Mmp8, and Wnt3a expression levels were altered in Hoxd4-transgenic chondrocytes, respectively. Notably, Wnt3a expression was elevated in Hoxc8- and reduced in Hoxd4-transgenic cartilage. These results suggest that both transcription factors affect cartilage maturation through different molecular mechanisms, and provide the basis for future studies into the role of these genes and possible interactions in pathogenesis of cartilage defects in Hoxc8- and Hoxd4-transgenic mice. PMID:20126390

  10. Tissue-specific expression of human CD4 in transgenic mice.

    PubMed

    Gillespie, F P; Doros, L; Vitale, J; Blackwell, C; Gosselin, J; Snyder, B W; Wadsworth, S C

    1993-05-01

    The gene for the human CD4 glycoprotein, which serves as the receptor for human immunodeficiency virus type 1, along with approximately 23 kb of sequence upstream of the translational start site, was cloned. The ability of 5' flanking sequences to direct tissue-specific expression was tested in cell culture and in transgenic mice. A 5' flanking region of 6 kb was able to direct transcription of the CD4 gene in NIH 3T3 cells but did not result in detectable expression in the murine T-cell line EL4 or in four lines of transgenic mice. A larger 5' flanking region of approximately 23 kb directed high-level CD4 transcription in the murine T-cell line EL4 and in three independent lines of transgenic mice. Human CD4 expression in all tissues analyzed was tightly correlated with murine CD4 expression; the highest levels of human CD4 RNA expression were found in the thymus and spleen, with relatively low levels detected in other tissues. Expression of human CD4 protein in peripheral blood mononuclear cells was examined by flow cytometry in these transgenic animals and found to be restricted to the murine CD4+ subset of lymphocytes. Human CD4 protein, detected with an anti-human CD4 monoclonal antibody, was present on the surface of 45 to 50% of the peripheral blood mononuclear cells from all transgenic lines. PMID:8474453

  11. Transgenic mice expressing human glucocerebrosidase variants: utility for the study of Gaucher disease.

    PubMed

    Sanders, Angela; Hemmelgarn, Harmony; Melrose, Heather L; Hein, Leanne; Fuller, Maria; Clarke, Lorne A

    2013-08-01

    Gaucher disease is an autosomal recessively inherited storage disorder caused by deficiency of the lysosomal hydrolase, acid β-glucosidase. The disease manifestations seen in Gaucher patients are highly heterogeneous as is the responsiveness to therapy. The elucidation of the precise factors responsible for this heterogeneity has been challenging as the development of clinically relevant animal models of Gaucher disease has been problematic. Although numerous murine models for Gaucher disease have been described each has limitations in their specific utility. We describe here, transgenic murine models of Gaucher disease that will be particularly useful for the study of pharmacological chaperones. We have produced stable transgenic mouse strains that individually express wild type, N370S and L444P containing human acid β-glucosidase and show that each of these transgenic lines rescues the lethal phenotype characteristic of acid β-glucosidase null mice. Both the N370S and L444P transgenic models show early and progressive elevations of tissue sphingolipids with L444P mice developing progressive splenic Gaucher cell infiltration. We demonstrate the potential utility of these new transgenic models for the study of Gaucher disease pathogenesis. In addition, since these mice produce only human enzyme, they are particularly relevant for the study of pharmacological chaperones that are specifically targeted to human acid β-glucosidase and the common mutations underlying Gaucher disease. PMID:23642305

  12. The effects of enhanced zinc on spatial memory and plaque formation in transgenic mice

    USGS Publications Warehouse

    Linkous, D.H.; Adlard, P.A.; Wanschura, P.B.; Conko, K.M.; Flinn, J.M.

    2009-01-01

    There is considerable evidence suggesting that metals play a central role in the pathogenesis of Alzheimer's disease. Reports suggest that elevated dietary metals may both precipitate and potentiate an Alzheimer's disease phenotype. Despite this, there remain few studies that have examined the behavioral consequences of elevated dietary metals in wild type and Alzheimer's disease animals. To further investigate this in the current study, two separate transgenic models of AD (Tg2576 and TgCRND8), together with wild type littermates were administered 10 ppm (0.153 mM) Zn. Tg2576 animals were maintained on a zinc-enriched diet both pre- and postnatally until 11 months of age, while TgCRND8 animals were treated for five months following weaning. Behavioral testing, consisting of "Atlantis" and "moving" platform versions of the Morris water maze, were conducted at the end of the study, and tissues were collected for immunohistochemical analysis of amyloid-β burden. Our data demonstrate that the provision of a zinc-enriched diet potentiated Alzheimer-like spatial memory impairments in the transgenic animals and was associated with reduced hippocampal amyloid-β plaque deposits. Zinc-related behavioral deficits were also demonstrated in wild type mice, which were sometimes as great as those present in the transgenic animals. However, zinc-related cognitive impairments in transgenic mice were greater than the summation of zinc effects in the wild type mice and the transgene effects.

  13. A Cyclosporine-Sensitive Psoriasis-Like Disease Produced in Tie2 Transgenic Mice

    PubMed Central

    Voskas, Daniel; Jones, Nina; Van Slyke, Paul; Sturk, Celina; Chang, Wing; Haninec, Alex; Babichev, Yael Olya; Tran, Jennifer; Master, Zubin; Chen, Stephen; Ward, Nicole; Cruz, Maribelle; Jones, Jamie; Kerbel, Robert S.; Jothy, Serge; Dagnino, Lina; Arbiser, Jack; Klement, Giannoula; Dumont, Daniel J.

    2005-01-01

    Psoriasis is a common, persistent skin disorder characterized by recurrent erythematous lesions thought to arise as a result of inflammatory cell infiltration and activation of keratinocyte proliferation. Unscheduled angiogenic growth has also been proposed to mediate the pathogenesis of psoriasis although the cellular and molecular basis for this response remains unclear. Recently, a role for the angiopoietin signaling system in psoriasis has been suggested by studies that demonstrate an up-regulation of the tyrosine kinase receptor Tie2 (also known as Tek) as well as angiopoietin-1 and angiopoietin-2 in human psoriatic lesions. To examine temporal expression of Tie2, we have developed a binary transgenic approach whereby expression of Tie2 can be conditionally regulated by the presence of tetracycline analogs in double-transgenic mice. A psoriasis-like phenotype developed in double-transgenic animals within 5 days of birth and persisted throughout adulthood. The skin of affected mice exhibited many cardinal features of human psoriasis including epidermal hyperplasia, inflammatory cell accumulation, and altered dermal angiogenesis. These skin abnormalities resolved completely with tetracycline-mediated suppression of transgene expression, thereby illustrating a complete dependence on Tie2 signaling for disease maintenance and progression. Furthermore, the skin lesions in double-transgenic mice markedly improved after administration of the immunosuppressive anti-psoriatic agent cyclosporine, thus demonstrating the clinical significance of this new model. PMID:15743796

  14. Enhanced neurofibrillary degeneration in transgenic mice expressing mutant tau and APP.

    PubMed

    Lewis, J; Dickson, D W; Lin, W L; Chisholm, L; Corral, A; Jones, G; Yen, S H; Sahara, N; Skipper, L; Yager, D; Eckman, C; Hardy, J; Hutton, M; McGowan, E

    2001-08-24

    JNPL3 transgenic mice expressing a mutant tau protein, which develop neurofibrillary tangles and progressive motor disturbance, were crossed with Tg2576 transgenic mice expressing mutant beta-amyloid precursor protein (APP), thus modulating the APP-Abeta (beta-amyloid peptide) environment. The resulting double mutant (tau/APP) progeny and the Tg2576 parental strain developed Abeta deposits at the same age; however, relative to JNPL3 mice, the double mutants exhibited neurofibrillary tangle pathology that was substantially enhanced in the limbic system and olfactory cortex. These results indicate that either APP or Abeta influences the formation of neurofibrillary tangles. The interaction between Abeta and tau pathologies in these mice supports the hypothesis that a similar interaction occurs in Alzheimer's disease. PMID:11520987

  15. High levels of circulating beta-amyloid peptide do not cause cerebral beta-amyloidosis in transgenic mice.

    PubMed Central

    Fukuchi, K.; Ho, L.; Younkin, S. G.; Kunkel, D. D.; Ogburn, C. E.; LeBoeuf, R. C.; Furlong, C. E.; Deeb, S. S.; Nochlin, D.; Wegiel, J.; Wisniewski, H. M.; Martin, G. M.

    1996-01-01

    We have established transgenic mice that constitutively overproduce the signal sequence and the 99-amino-acid carboxyl-terminal region of the human beta-amyloid precursor protein. The transgenic mice strongly expressed the transgene in multiple tissues under the control of a cytomegalovirus enhancer/chick beta-actin promoter. There were exceptionally high levels of beta-amyloid peptides in the plasma (approximately 17 times or more compared with the human plasma level). Although some transgenic mice from one founder line developed amyloidosis in the intestine, no neuropathology was found in transgenic mice up to age 29 months. Given the absence of cerebral beta-amyloidosis despite extremely high levels of circulating beta-amyloid peptides in the transgenic mice, the results suggest that local cerebral metabolism of beta-amyloid precursor protein may play a predominant role in cerebral beta-amyloidosis in transgenic mice. Such transgenic mice may be useful for the investigation of the etiology of the disease and for the establishment of therapeutic strategies. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:8686746

  16. Expression of human protamine P1 in sperm of transgenic mice

    SciTech Connect

    Wyrobek, A.J.; Keith, C.; Stilwell, J.; Lowe, X.; Anderson, G.

    1994-12-31

    Transgenic mice were produced by pronuclear injection with DNA constructs containing human protamine P1 cDNA recombined with a murine protamine P1 promoter, and were identified by PCR. Expression of human P1 was investigated using huplm, a monoclonal antibody specific for human P1, applied to murine testicular cells, smears of epididymal sperm, and smears of detergent-isolated sperm nuclei. Various antibodies and nontransgenic littermates were used as controls. Two male founders (T3 and T7) sired more than five generations of transgenic offspring each with continued expression of human P1 in their sperm. Transgenic animals appear of normal fertility with sperm of typical nuclear morphology. The human P1 transgene was expressed postmeioticly in both lines, as expected. Nearly 100% of sperm of T3 and T7 hemizygotes labeled with huplm, consistent with complete diffusion of human P1 protein through the intercellular bridge of spermatogenic cells. Human P1 labeling of sperm nuclei was not visibly affected by sonication or by treatment with the detergent MATAB or the reducing agent DTT. A third founder female (T5) showed a transmission pattern consistent with insertion of the transgene into an X chromosome; her transgenic offspring expressed human P1 in only a small fraction of sperm. Human P1 transgenes may serve as efficient targets for germinal mutations and transgenicmice may provide promising models for investigating the DNA complexes.

  17. Search Strategies Used by "APP" Transgenic Mice during Navigation in the Morris Water Maze

    ERIC Educational Resources Information Center

    Janus, Christopher

    2004-01-01

    TgCRND8 mice represent a transgenic mouse model of Alzheimer's disease, with onset of cognitive impairment and increasing amyloid-[beta] plaques in their brains at 12 weeks of age. In this study, the spatial memory in 25- to 30-week-old TgCRND8 mice was analyzed in two reference and one working memory Morris water maze (MWM) tests. In reference…

  18. Rescue of NGF-deficient mice I: transgenic expression of NGF in skin rescues mice lacking endogenous NGF.

    PubMed

    Harrison, Susan M W; Davis, Brian M; Nishimura, Merry; Albers, Kathryn M; Jones, Marc E; Phillips, Heidi S

    2004-03-30

    Mice lacking a functional NGF gene (ngf-/- mice) have less than one third of the normal complement of sensory neurons, few sympathetic postganglionic neurons and die shortly after birth. We report here that transgenic expression of NGF under control of the K14 keratin promoter can rescue some elements of the peripheral nervous system and restore normal growth and viability to ngf-/- mice. While hybrid transgenic-ngf-/- mice (ngfTKOs) displayed marginal rescue of trigeminal ganglion neurons, the percentage of CGRP-positive neurons was restored to normal. Restoration of CGRP-positive terminals in skin and spinal cord was also found and accompanied by recovery of behavioral responses to noxious stimuli. ngfTKO mice displayed a normal number of superior cervical ganglion neurons and recovery of sympathetic innervation of skin. These results demonstrate that substitution of a functional NGF locus by a transgene directing expression largely to skin can result in normal growth and viability. Thus, the most vital functions of NGF are not dependent on faithful recapitulation of the normal spatiotemporal pattern of gene expression. PMID:15010204

  19. Traumatic Brain Injury Precipitates Cognitive Impairment and Extracellular Aβ Aggregation in Alzheimer's Disease Transgenic Mice

    PubMed Central

    Shimizu, Toru; Arendash, Gary W.; Borlongan, Cesar V.

    2013-01-01

    Traumatic brain injury (TBI) has become a signature wound of the wars in Iraq and Afghanistan. Many American soldiers, even those undiagnosed but likely suffering from mild TBI, display Alzheimer's disease (AD)-like cognitive impairments, suggesting a pathological overlap between TBI and AD. This study examined the cognitive and neurohistological effects of TBI in presymptomatic APP/PS1 AD-transgenic mice. AD mice and non-transgenic (NT) mice received an experimental TBI on the right parietal cortex using the controlled cortical impact model. Animals were trained in a water maze task for spatial memory before TBI, and then reevaluated in the same task at two and six weeks post-TBI. The results showed that AD mice with TBI made significantly more errors in the task than AD mice without TBI and NT mice regardless of TBI. A separate group of AD mice and NT mice were evaluated neurohistologically at six weeks after TBI. The number of extracellular beta-amyloid (Aβ)-deposits significantly increased by at least one fold in the cortex of AD mice that received TBI compared to the NT mice that received TBI or the AD and NT mice that underwent sham surgery. A significant decrease in MAP2 positive cells, indicating neuronal loss, was observed in the cortex of both the AD and NT mice that received TBI compared to the AD and NT mice subjected to sham surgery. Similar changes in extracellular Aβ deposits and MAP2 positive cells were also seen in the hippocampus. These results demonstrate for the first time that TBI precipitates cognitive impairment in presymptomatic AD mice, while also confirming extracellular Aβ deposits following TBI. The recognition of this pathological link between TBI and AD should aid in developing novel treatments directed at abrogating cellular injury and extracellular Aβ deposition in the brain. PMID:24223856

  20. Traumatic brain injury precipitates cognitive impairment and extracellular Aβ aggregation in Alzheimer's disease transgenic mice.

    PubMed

    Tajiri, Naoki; Kellogg, S Leilani; Shimizu, Toru; Arendash, Gary W; Borlongan, Cesar V

    2013-01-01

    Traumatic brain injury (TBI) has become a signature wound of the wars in Iraq and Afghanistan. Many American soldiers, even those undiagnosed but likely suffering from mild TBI, display Alzheimer's disease (AD)-like cognitive impairments, suggesting a pathological overlap between TBI and AD. This study examined the cognitive and neurohistological effects of TBI in presymptomatic APP/PS1 AD-transgenic mice. AD mice and non-transgenic (NT) mice received an experimental TBI on the right parietal cortex using the controlled cortical impact model. Animals were trained in a water maze task for spatial memory before TBI, and then reevaluated in the same task at two and six weeks post-TBI. The results showed that AD mice with TBI made significantly more errors in the task than AD mice without TBI and NT mice regardless of TBI. A separate group of AD mice and NT mice were evaluated neurohistologically at six weeks after TBI. The number of extracellular beta-amyloid (Aβ)-deposits significantly increased by at least one fold in the cortex of AD mice that received TBI compared to the NT mice that received TBI or the AD and NT mice that underwent sham surgery. A significant decrease in MAP2 positive cells, indicating neuronal loss, was observed in the cortex of both the AD and NT mice that received TBI compared to the AD and NT mice subjected to sham surgery. Similar changes in extracellular Aβ deposits and MAP2 positive cells were also seen in the hippocampus. These results demonstrate for the first time that TBI precipitates cognitive impairment in presymptomatic AD mice, while also confirming extracellular Aβ deposits following TBI. The recognition of this pathological link between TBI and AD should aid in developing novel treatments directed at abrogating cellular injury and extracellular Aβ deposition in the brain. PMID:24223856

  1. Generation and characterization of mice transgenic for human IL-18-binding protein isoform a.

    PubMed

    Fantuzzi, Giamila; Banda, Nirmal K; Guthridge, Carla; Vondracek, Andrea; Kim, Soo-Hyun; Siegmund, Britta; Azam, Tania; Sennello, Joseph A; Dinarello, Charles A; Arend, William P

    2003-11-01

    Interleukin (IL)-18 binding protein (IL-18BP) is a natural inhibitor of the pleiotropic cytokine IL-18. To study the role of IL-18BP in modulating inflammatory responses in vivo, mice transgenic for human IL-18BP isoform a (IL-18BP-Tg) were generated. The transgene was expressed at high levels in each organ examined. High levels of bioactive human IL-18BPa were detectable in the circulation of IL-18BP-Tg mice, which were viable, fertile, and had no tissue or organ abnormality. The high levels of IL-18BP in the transgenic mice were able to completely neutralize the interferon-gamma (IFN-gamma)-inducing activity of exogenously administered IL-18. Following administration of endotoxin, with or without prior sensitization with heat-inactivated Propionibacterium acnes, IL-18BP-Tg mice produced significantly lower serum levels of IFN-gamma and macrophage-inflammatory protein-2 compared with nontransgenic littermates. Significantly reduced production of IFN-gamma in response to endotoxin was also observed in cultures of IL-18BP-Tg splenocytes. Finally, IL-18BP-Tg mice were completely protected in a model of hepatotoxicity induced by administration of concanavalin A. These results indicate that high endogenous levels of IL-18BP in trangenic mice effectively neutralize IL-18 and are protective in response to different inflammatory stimuli. PMID:12960225

  2. Progranulin reduction is associated with increased tau phosphorylation in P301L tau transgenic mice.

    PubMed

    Hosokawa, Masato; Arai, Tetsuaki; Masuda-Suzukake, Masami; Kondo, Hiromi; Matsuwaki, Takashi; Nishihara, Masugi; Hasegawa, Masato; Akiyama, Haruhiko

    2015-02-01

    Granulin (GRN) mutations have been identified in familial frontotemporal lobar degeneration patients with ubiquitin pathology. GRN transcript haploinsufficiency is proposed as a disease mechanism that leads to the loss of functional progranulin (PGRN) protein. Thus, these mutations are strongly involved in frontotemporal lobar degeneration pathogenesis. Moreover, recent findings indicate that GRN mutations are associated with other neurodegenerative disorders with tau pathology, including Alzheimer disease and corticobasal degeneration. To investigate the potential influence of a decline in PGRN protein on tau accumulation, P301L tau transgenic mice were interbred with GRN-deficient mice, producing P301L tau transgenic mice harboring the GRN hemizygote. Brains were collected from 13- and 19-month-old mice, and sequential extraction of proteins, immunoblotting, and immunohistochemical analyses were performed. Immunoblotting analysis revealed that tau phosphorylation was accelerated in the Tris-saline soluble fraction of 13-month-old and in the sarkosyl-insoluble fraction of 19-month-old P301L tau/GRN hemizygotes compared with those in fractions from P301L tau transgenic mice. Activity of cyclin-dependent kinases was also upregulated in the brains of P301L tau/GRN hemizygote mice. Although the mechanisms involved in these findings remain unknown, our data suggest that a reduction in PGRN protein might contribute to phosphorylation and intraneuronal accumulation of tau. PMID:25575133

  3. Pkd1 transgenic mice: adult model of polycystic kidney disease with extrarenal and renal phenotypes

    PubMed Central

    Kurbegovic, Almira; Côté, Olivier; Couillard, Martin; Ward, Christopher J.; Harris, Peter C.; Trudel, Marie

    2010-01-01

    While high levels of Pkd1 expression are detected in tissues of patients with autosomal dominant polycystic kidney disease (ADPKD), it is unclear whether enhanced expression could be a pathogenetic mechanism for this systemic disorder. Three transgenic mouse lines were generated from a Pkd1-BAC modified by introducing a silent tag via homologous recombination to target a sustained wild-type genomic Pkd1 expression within the native tissue and temporal regulation. These mice specifically overexpressed the Pkd1 transgene in extrarenal and renal tissues from ∼2- to 15-fold over Pkd1 endogenous levels in a copy-dependent manner. All transgenic mice reproducibly developed tubular and glomerular cysts leading to renal insufficiency. Interestingly, Pkd1TAG mice also exhibited renal fibrosis and calcium deposits in papilla reminiscent of nephrolithiasis as frequently observed in ADPKD. Similar to human ADPKD, these mice consistently displayed hepatic fibrosis and ∼15% intrahepatic cysts of the bile ducts affecting females preferentially. Moreover, a significant proportion of mice developed cardiac anomalies with severe left-ventricular hypertrophy, marked aortic arch distention and/or valvular stenosis and calcification that had profound functional impact. Of significance, Pkd1TAG mice displayed occasional cerebral lesions with evidence of ruptured and unruptured cerebral aneurysms. This Pkd1TAG mouse model demonstrates that overexpression of wild-type Pkd1 can trigger the typical adult renal and extrarenal phenotypes resembling human ADPKD. PMID:20053665

  4. Seeded strain-like transmission of β-amyloid morphotypes in APP transgenic mice

    PubMed Central

    Heilbronner, Götz; Eisele, Yvonne S; Langer, Franziska; Kaeser, Stephan A; Novotny, Renata; Nagarathinam, Amudha; Åslund, Andreas; Hammarström, Per; Nilsson, K Peter R; Jucker, Mathias

    2013-01-01

    The polymorphic β-amyloid lesions present in individuals with Alzheimer's disease are collectively known as cerebral β-amyloidosis. Amyloid precursor protein (APP) transgenic mouse models similarly develop β-amyloid depositions that differ in morphology, binding of amyloid conformation-sensitive dyes, and Aβ40/Aβ42 peptide ratio. To determine the nature of such β-amyloid morphotypes, β-amyloid-containing brain extracts from either aged APP23 brains or aged APPPS1 brains were intracerebrally injected into the hippocampus of young APP23 or APPPS1 transgenic mice. APPPS1 brain extract injected into young APP23 mice induced β-amyloid deposition with the morphological, conformational, and Aβ40/Aβ42 ratio characteristics of β-amyloid deposits in aged APPPS1 mice, whereas APP23 brain extract injected into young APP23 mice induced β-amyloid deposits with the characteristics of β-amyloid deposits in aged APP23 mice. Injecting the two extracts into the APPPS1 host revealed a similar difference between the induced β-amyloid deposits, although less prominent, and the induced deposits were similar to the β-amyloid deposits found in aged APPPS1 hosts. These results indicate that the molecular composition and conformation of aggregated Aβ in APP transgenic mice can be maintained by seeded conversion. PMID:23999102

  5. {alpha}-Lipoic acid prevents lipotoxic cardiomyopathy in acyl CoA-synthase transgenic mice

    SciTech Connect

    Lee, Young; Naseem, R. Haris; Park, Byung-Hyun; Garry, Daniel J.; Richardson, James A.; Schaffer, Jean E.; Unger, Roger H. . E-mail: roger.unger@utsouthwestern.edu

    2006-05-26

    {alpha}-Lipoic acid ({alpha}-LA) mimics the hypothalamic actions of leptin on food intake, energy expenditure, and activation of AMP-activated protein kinase (AMPK). To determine if, like leptin, {alpha}-LA protects against cardiac lipotoxicity, {alpha}-LA was fed to transgenic mice with cardiomyocyte-specific overexpression of the acyl CoA synthase (ACS) gene. Untreated ACS-transgenic mice died prematurely with increased triacylglycerol content and dilated cardiomyopathy, impaired systolic function and myofiber disorganization, apoptosis, and interstitial fibrosis on microscopy. In {alpha}-LA-treated ACS-transgenic mice heart size, echocardiogram and TG content were normal. Plasma TG fell 50%, hepatic-activated phospho-AMPK rose 6-fold, sterol regulatory element-binding protein-1c declined 50%, and peroxisome proliferator-activated receptor-{gamma} cofactor-1{alpha} mRNA rose 4-fold. Since food restriction did not prevent lipotoxicity, we conclude that {alpha}-LA treatment, like hyperleptinemia, protects the heart of ACS-transgenic mice from lipotoxicity.

  6. Human SCARB2 Transgenic Mice as an Infectious Animal Model for Enterovirus 71

    PubMed Central

    Lin, Yi-Wen; Yu, Shu-Ling; Shao, Hsiao-Yun; Lin, Hsiang-Yin; Liu, Chia-Chyi; Hsiao, Kuang-Nan; Chitra, Ebenezer; Tsou, Yueh-Liang; Chang, Hsuen-Wen; Sia, Charles; Chong, Pele; Chow, Yen-Hung

    2013-01-01

    Enterovirus 71 (EV71) and coxsackievirus (CVA) are the most common causative factors for hand, foot, and mouth disease (HFMD) and neurological disorders in children. Lack of a reliable animal model is an issue in investigating EV71-induced disease manifestation in humans, and the current clinical therapies are symptomatic. We generated a novel EV71-infectious model with hSCARB2-transgenic mice expressing the discovered receptor human SCARB2 (hSCARB2). The challenge of hSCARB2-transgenic mice with clinical isolates of EV71 and CVA16 resulted in HFMD-like and neurological syndromes caused by E59 (B4) and N2838 (B5) strains, and lethal paralysis caused by 5746 (C2), N3340 (C4), and CVA16. EV71 viral loads were evident in the tissues and CNS accompanied the upregulated pro-inflammatory mediators (CXCL10, CCL3, TNF-α, and IL-6), correlating to recruitment of the infiltrated T lymphocytes that result in severe diseases. Transgenic mice pre-immunized with live E59 or the FI-E59 vaccine was able to resist the subsequent lethal challenge with EV71. These results indicate that hSCARB2-transgenic mice are a useful model for assessing anti-EV71 medications and for studying the pathogenesis induced by EV71. PMID:23451246

  7. Curcumin relieves TPA-induced Th1 inflammation in K14-VEGF transgenic mice.

    PubMed

    Sun, Jun; Zhao, Yi; Jin, Hairong; Hu, Jinhong

    2015-04-01

    Curcumin has been confirmed to have anti-inflammatory properties in addition to the ability to decrease the expression of pro-inflammatory cytokines in keratinocytes. It was suggested that the interleukin-23 (IL-23)/IL-17A cytokine axis played a critical role in the pathogenesis of 12-O-tetradecanoyl phorbol 12-myristate 13-acetate (TPA)-induced K14-VEGF transgenic psoriasis-like mice model. Here, we report that topical use of a curcumin gel formulation inhibited TPA-induced Th1 inflammation in K14-VEGF transgenic mice ears but not Th17 inflammation as expected. Real-time PCR showed that mRNA levels of IL-23, IL-17A, IL-22, IL-6 and TNFα cytokines failed to increase after TPA-induction in K14-VEGF transgenic mice ear skin; but the mRNA level of IFNγ increased significantly at the same time. Furthermore, TPA-induction up-regulated the TCRγδ protein but failed to impact the CCR6 protein, which means that the proliferation of γδ T cells is incapable of IL-17A production. We find that curcumin is capable of relieving TPA-induced inflammation by directly down-regulating IFNγ production. In conclusion, curcumin inhibits TPA-induced Th1 inflammation in K14-VEGF transgenic mice which has not been previously described. PMID:25682767

  8. Bovine growth hormone transgenic mice display alterations in locomotor activity and brain monoamine neurochemistry.

    PubMed

    Söderpalm, B; Ericson, M; Bohlooly, M; Engel, J A; Törnell, J

    1999-12-01

    Recent clinical and experimental data indicate a role for GH in mechanisms related to anhedonia/hedonia, psychic energy, and reward. In the present study we have investigated whether bovine GH (bGH) transgenic mice and nontransgenic controls differ in spontaneous locomotor activity, a behavioral response related to brain dopamine (DA) and reward mechanisms, as well as in locomotor activity response to drugs of abuse known to interfere with brain DA systems. The animals were tested for locomotor activity once a week for 4 weeks. When first exposed to the test apparatus, bGH transgenic animals displayed significantly more locomotor activity than controls during the entire registration period (1 h). One week later, after acute pretreatment with saline, the two groups did not differ in locomotor activity, whereas at the third test occasion, bGH mice were significantly more stimulated by d-amphetamine (1 mg/kg, ip) than controls. At the fourth test, a tendency for a larger locomotor stimulatory effect of ethanol (2.5 g/kg, ip) was observed in bGH transgenic mice. bGH mice displayed increased tissue levels of serotonin and 5-hydroxyindoleacetic acid in several brain regions, decreased DA levels in the brain stem, and decreased levels of the DA metabolite 3,4-dihydroxyphenylacetic acid in the mesencephalon and diencephalon, compared with controls. In conclusion, bGH mice display more spontaneous locomotor activity than nontransgenic controls in a novel environment and possibly also a disturbed habituation process. The finding that bGH mice were also more sensitive to d-amphetamine-induced locomotor activity may suggest that the behavioral differences observed are related to differences in brain DA systems, indicating a hyperresponsiveness of these systems in bGH transgenic mice. These findings may constitute a neurochemical basis for the reported psychic effects of GH in humans. PMID:10579325

  9. APP/PS1 transgenic mice treated with aluminum: an update of Alzheimer's disease model.

    PubMed

    Zhang, Q L; Jia, L; Jiao, X; Guo, W L; Ji, J W; Yang, H L; Niu, Q

    2012-01-01

    There is still no animal model available that can mimic all the cognitive, behavioral, biochemical, and histopathological abnormalities observed in patients with Alzheimer's disease (AD). We undertook to consider the interaction between genetic factors, including amyloid precursor protein (APP) and presenilin-1 (PS1), and environmental factors, such as Aluminum (Al) in determining susceptibility outcomes when studying the pathogenesis of AD. In this article, we provide an AD model in APP/PS1 transgenic mice triggered by Al. The animal model was established via intracerebral ventricular microinjection of aluminum chloride once a day for 5 days in APP/PS1 transgenic mice. Twenty wild type (WT) mice and 20 APP/PS1 transgenic (TG) mice were separately divided into 2 groups (control and Al group), and a stainless steel injector with stopper was used for microinjection into the left-lateral cerebral ventricle of each mouse. The Morris water maze task was used to evaluate behavioral function of learning and memory ability on the 20th day after the last injection. This AD model's brain was analyzed by: (1) amyloid beta immunohistochemical staining; (2) Tunnel staining; (3) apoptotic rates; (4) caspase-3 gene expression. Here, decrease of cognitive ability and neural cells loss were shown in APP/PS1 transgenic mice exposed to Al, which were more extensive than those in APP/PS1 TG alone and WT mice exposed to Al alone. These findings indicate that there is a close relationship between over-expression of APP and PS1 genes and Al overload. It is also suggested that APP/PS1 TG mice exposed to Al have potential value for improving AD models. PMID:22507317

  10. Induction of proteinuria by cannabinoid receptors 1 signaling activation in CB1 transgenic mice.

    PubMed

    Hsu, Yung-Chien; Lei, Chen-Chou; Shih, Ya-Hsueh; Ho, Cheng; Lin, Chun-Liang

    2015-02-01

    Proteinuria is not only a sign of kidney damage but is also involved in the progression of renal disease as an independent pathologic factor. Although patients with mutated type 1 cannabinoid receptors (CB1) polymorphism are associated with renal microvascular damage, the biologic role of CB1 signaling in proteinuria remains uncharacterized till now. Herein, we investigate whether CB1 participates in glomerular proteinuria in CB1 transgenic mice and treatment with CB1 agonist WIN55212-2 rat, neither of which are diabetic models. The CB1 transgenic mice and rats treated with CB1 agonist WIN55212-2 had higher kidney weight and urinary protein concentrations but not blood glucose levels compared with the wild-type group. A combination of laser-capture microsdissection, quantitative reverse transcription-polymerase chain reaction, immunoblotting and immunohistochemical validation revealed that CB1 transgenic mice and rats treated with CB1 agonist WIN55212-2 had higher vascular endothelial growth factor (VEGF) expression in renal glomeruli than that of the wild-type group. Geneticorpharmacological activation of CB1 by transgenic CB1 mice or treatment with WIN55212-2 reduced nephrin expression in the renal glomeruli compared with that of the wild-type group in the glomerular mesanglium. Taken together, CB1 transgenic mice and rats treated with CB1 agonist WIN55212-2 induced proteinuria with upregulation of CB1 resulting in impaired nephrin expression, by inducing excess VEGF reaction in the renal glomeruli. Genetic and pharmacological manipulation of CB1 signaling revealed VEGF-dependent nephrin depression of glomerulopathy. Controlling CB1 activity can be used an alternative strategy for sustaining renal function in the presence of CB1 activation. PMID:25474224

  11. Eμ-BRD2 transgenic mice develop B-cell lymphoma and leukemia

    PubMed Central

    Greenwald, Rebecca J.; Tumang, Joseph R.; Sinha, Anupama; Currier, Nicolas; Cardiff, Robert D.; Rothstein, Thomas L.; Faller, Douglas V.; Denis, Gerald V.

    2010-01-01

    Transgenic mice with lymphoid-restricted overexpression of the double bromodomain protein bromodomain-containing 2 (Brd2) develop splenic B-cell lymphoma and, upon transplantation, B-cell leukemia with leukemic infiltrates in liver and lung. Brd2 is a nuclear-localized transcription factor kinase that is most closely related to TATA box binding protein–associated factor, 250 kDa (TAFII250) and the Drosophila developmental protein female sterile homeotic. Constitutive expression of BRD2 in the lymphoid compartment increases cyclin A transcription, “priming” transgenic B cells for proliferation. Mice stochastically develop an aggressive B-cell lymphoma with the features of B-1 cells, including CD5 and surface IgM expression. The B-cell lymphoma is monoclonal for immunoglobulin gene rearrangement and is phenotypically stable. The lymphoblasts are very large and express a transcriptome that is similar to human non-Hodgkin lymphomas. Both a wild-type BRD2 transgene and a kinase-null point mutant drive lymphomagenesis; therefore we propose that, rather than kinase activity, Brd2-mediated recruitment of E2 promoter binding factors (E2Fs) and a specific histone acetyltransferase to the cyclin A promoter by both types of transgene is a mechanistic basis for neoplasia. This report is the first to describe a transgenic mouse model for constitutive expression of a protein with more than one bromodomain. PMID:14563639

  12. RasGRP1 Transgenic Mice Develop Cutaneous Squamous Cell Carcinomas in Response to Skin Wounding

    PubMed Central

    Diez, Federico R.; Garrido, Ann A.; Sharma, Amrish; Luke, Courtney T.; Stone, James C.; Dower, Nancy A.; Cline, J. Mark; Lorenzo, Patricia S.

    2009-01-01

    Models of epidermal carcinogenesis have demonstrated that Ras is a critical molecule involved in tumor initiation and progression. Previously, we have shown that RasGRP1 increases the susceptibility of mice to skin tumorigenesis when overexpressed in the epidermis by a transgenic approach, related to its ability to activate Ras. Moreover, RasGRP1 transgenic mice develop spontaneous papillomas and cutaneous squamous cell carcinomas, some of which appear to originate in sites of injury, suggesting that RasGRP1 may be responding to signals generated during the wound-healing process. In this study, we examined the response of the RasGRP1 transgenic animals to full-thickness incision wounding of the skin, and demonstrated that they respond by developing tumors along the wounded site. The tumors did not present mutations in the H-ras gene, but Rasgrp1 transgene dosage correlated with tumor susceptibility and size. Analysis of serum cytokines showed increased levels of granulocyte colony-stimulating factor in transgenic animals after wounding. Furthermore, in vitro experiments with primary keratinocytes showed that granulocyte colony-stimulating factor stimulated Ras activation, although RasGRP1 was dispensable for this effect. Since granulocyte colony-stimulating factor has been recently associated with proliferation of skin cancer cells, our results may help in the elucidation of pathways that activate Ras in the epidermis during tumorigenesis in the absence of oncogenic ras mutations. PMID:19497993

  13. Fluorescent transgenic mice suitable for multi-color aggregation chimera studies.

    PubMed

    Ohtsuka, Masato; Miura, Hiromi; Gurumurthy, Channabasavaiah B; Kimura, Minoru; Inoko, Hidetoshi; Yoshimura, Shinichi; Sato, Masahiro

    2012-11-01

    We recently reported a novel method of mouse transgenesis called Pronuclear Injection-based Targeted Transgenisis (PITT) using which a series of fluorescent transgenic (Tg) mice lines were generated. These lines, unlike those generated using conventional random integration methods, express the transgenes faithfully and reproducibly generation after generation. Because of this superior nature, these lines are ideal for the generation of multi-colored aggregation chimeras that can be used to study cell-cell interactions and lineage analyses in living embryos/organs, where the transgenes can be detected and the clonal origin of a given cell population easily traced by its distinct fluorescence. In this study, to verify if Tg fluorescent mice generated through PITT were suitable for such applications, we sought to generate chimeric blastocysts and chimeric-Tg mice by aggregating two- or three-colored 8-cell embryos. Our analyses using these models led to the following observations. First, we noticed that cell mixing was infrequent during the stages of morula to early blastocyst. Second, chimeric fetuses obtained after aggregation of the two-colored 8-cell embryos exhibited uniform cell mixing. And third, in the organs of adult chimeric mice, the mode of cell distribution could be either clonal or polyclonal, as previously pointed out by others. Implications of our novel and improved Tg-chimeric mice approach for clonal cell lineage and developmental studies are discussed. PMID:22868913

  14. Identification of a prion protein epitope modulating transmission of bovine spongiform encephalopathy prions to transgenic mice

    PubMed Central

    Scott, Michael R.; Safar, Jiri; Telling, Glenn; Nguyen, Oanh; Groth, Darlene; Torchia, Marilyn; Koehler, Ruth; Tremblay, Patrick; Walther, Dirk; Cohen, Fred E.; DeArmond, Stephen J.; Prusiner, Stanley B.

    1997-01-01

    There is considerable concern that bovine prions from cattle with bovine spongiform encephalopathy (BSE) may have been passed to humans (Hu), resulting in a new form of Creutzfeldt–Jakob disease (CJD). We report here the transmission of bovine (Bo) prions to transgenic (Tg) mice expressing BoPrP; one Tg line exhibited incubation times of ≈200 days. Like most cattle with BSE, vacuolation and astrocytic gliosis were confined in the brainstems of these Tg mice. Unexpectedly, mice expressing a chimeric Bo/Mo PrP transgene were resistant to BSE prions whereas mice expressing Hu or Hu/Mo PrP transgenes were susceptible to Hu prions. A comparison of differences in Mo, Bo, and Hu residues within the C terminus of PrP defines an epitope that modulates conversion of PrPC into PrPSc and, as such, controls prion transmission across species. Development of susceptible Tg(BoPrP) mice provides a means of measuring bovine prions that may prove critical in minimizing future human exposure. PMID:9405603

  15. Measles virus replication in lymphatic cells and organs of CD150 (SLAM) transgenic mice

    PubMed Central

    Welstead, G. Grant; Iorio, Caterina; Draker, Ryan; Bayani, Jane; Squire, Jeremy; Vongpunsawad, Sompong; Cattaneo, Roberto; Richardson, Christopher D.

    2005-01-01

    A transgenic mouse containing the complete human SLAM (hSLAM/CD150) gene, including its endogenous promoter for transcription, was generated by using human genomic DNA cloned into a bacterial artificial chromosome. hSLAM, the primary receptor for measles viruses (MV), was expressed on activated B, T, and dendritic cells with an expression profile equivalent to that of humans. We demonstrated that hSLAM+ cells obtained from the transgenic mouse, including activated B, T, and dendritic cells, were susceptible to MV infection in a receptor-dependent manner. Evidence was provided for transient infection in the nasal lymph nodes of hSLAM+ mice after intranasal inoculation. Virus was rapidly cleared without signs of secondary replication. To improve the efficiency of MV production, the hSLAM+ mice were bred with mice having a Stat1-deficient background. These mice were more susceptible to MV infection and produced more virus particles. After intranasal and intraperitoneal inoculation of these mice with MV, infections of the thymus, spleen, nasal, mesenteric, and leg lymph nodes were detected. Upon necropsy, enlarged lymph nodes and spleen were apparent. Flow cytometric analysis showed that abnormally large numbers of mature neutrophils and natural killer cells caused the splenomegaly. The hSLAM transgenic mouse constitutes an improved rodent model for studying the interaction of MV with immune cells that more accurately reflects the infection pattern found in humans. PMID:16260741

  16. Myodegeneration in EDA-A2 Transgenic Mice Is Prevented by XEDAR Deficiency

    PubMed Central

    Newton, Kim; French, Dorothy M.; Yan, Minhong; Frantz, Gretchen D.; Dixit, Vishva M.

    2004-01-01

    EDA-A1 and EDA-A2 are members of the tumor necrosis factor family of ligands. The products of alternative splicing of the ectodysplasin (EDA) gene, EDA-A1 and EDA-A2 differ by an insertion of two amino acids and bind to distinct receptors. The longer isoform, EDA-A1, binds to EDAR and plays an important role in sweat gland, hair, and tooth development; mutations in EDA, EDAR, or the downstream adaptor EDARADD cause hypohidrotic ectodermal dysplasia. EDA-A2 engages the receptor XEDAR, but its role in the whole organism is less clear. We have generated XEDAR-deficient mice by gene targeting and transgenic mice expressing secreted forms of EDA-A1 or EDA-A2 downstream of the skeletal muscle-specific myosin light-chain 2 or skin-specific keratin 5 promoter. Mice lacking XEDAR were indistinguishable from their wild-type littermates, but EDA-A2 transgenic mice exhibited multifocal myodegeneration. This phenotype was not observed in the absence of XEDAR. Skeletal muscle in EDA-A1 transgenic mice was unaffected, but their sebaceous glands were hypertrophied and hyperplastic, consistent with a role for EDA-A1 in the development of these structures. These data indicate that XEDAR-transduced signals are dispensable for development of ectoderm-derived organs but might play a role in skeletal muscle homeostasis. PMID:14749376

  17. Enhanced penetration of exogenous EPCs into brains of APP/PS1 transgenic mice

    PubMed Central

    Yuan, Xiaoyang; Mei, Bin; Zhang, Le; Zhang, Cuntai; Zheng, Miao; Liang, Huifang; Wang, Wei; Zheng, Jie; Ding, Ling; Zheng, Kai

    2016-01-01

    The aim of this study was to investigate the repair function of exogenous Endothelial progenitor cells (EPCs) for brain microvascular damage of the APP/PS1 transgenic mouse model of Alzheimer’s disease (AD). This study used a density-gradient centrifugation method to isolate mononuclear cells (MNCs) from mouse bone marrow, which were subsequently seeded and cultured. Cells were characterized by morphology and detection of the surface markers CD34 and CD133 at different time points by immunofluorescence (IF) and flow cytometry (FCM). Then, EPCs were transfected with GFP adenoviral vectors (GFP-EPCs). Wild-type (WT) and APP/PS1 transgenic mice both received GFP-EPCs injection through the tail vein, and using a PBS buffer injection as the control. Seven days later, the animals’ brain tissue was isolated. Expression of GFP was detected by quantitative polymerase chain reaction (qPCR) and western-blot (WB), while the fluorescence of GFP within the brains of mice was observed under a fluorescence microscope. Higher mRNA and protein expression of GFP, accompanied with increased green fluorescence, were detected in the brain of GFP-EPCs-injected APP/PS1 mice, as compared with GFP-EPCs-injected WT mice. The results show that the APP/PS1 transgenic mouse model of AD exhibited enhanced penetration of exogenous EPCs into brains than the WT mice. PMID:27186272

  18. Enhanced penetration of exogenous EPCs into brains of APP/PS1 transgenic mice.

    PubMed

    Yuan, Xiaoyang; Mei, Bin; Zhang, Le; Zhang, Cuntai; Zheng, Miao; Liang, Huifang; Wang, Wei; Zheng, Jie; Ding, Ling; Zheng, Kai

    2016-01-01

    The aim of this study was to investigate the repair function of exogenous Endothelial progenitor cells (EPCs) for brain microvascular damage of the APP/PS1 transgenic mouse model of Alzheimer's disease (AD). This study used a density-gradient centrifugation method to isolate mononuclear cells (MNCs) from mouse bone marrow, which were subsequently seeded and cultured. Cells were characterized by morphology and detection of the surface markers CD34 and CD133 at different time points by immunofluorescence (IF) and flow cytometry (FCM). Then, EPCs were transfected with GFP adenoviral vectors (GFP-EPCs). Wild-type (WT) and APP/PS1 transgenic mice both received GFP-EPCs injection through the tail vein, and using a PBS buffer injection as the control. Seven days later, the animals' brain tissue was isolated. Expression of GFP was detected by quantitative polymerase chain reaction (qPCR) and western-blot (WB), while the fluorescence of GFP within the brains of mice was observed under a fluorescence microscope. Higher mRNA and protein expression of GFP, accompanied with increased green fluorescence, were detected in the brain of GFP-EPCs-injected APP/PS1 mice, as compared with GFP-EPCs-injected WT mice. The results show that the APP/PS1 transgenic mouse model of AD exhibited enhanced penetration of exogenous EPCs into brains than the WT mice. PMID:27186272

  19. Park2-null/tau transgenic mice reveal a functional relationship between parkin and tau.

    PubMed

    Guerrero, Rosa; Navarro, Paloma; Gallego, Eva; Avila, Jesus; de Yebenes, Justo G; Sanchez, Marina P

    2008-03-01

    Mutations, haplotypes, and polymorphisms of tau and Park-2 genes constitute risk factors for developing tauopathies. In order to analyze the possible relationship between parkin and tau we generated a double-mutant mouse deficient for Park-2 expression and overexpressing a mutant tau protein (hTauVLW). Mice develop normally, although the median survival rate is considerably reduced with respect to wild type (45%). Aggregates of phosphorylated tau in neurons and reactive gliosis are quite abundant in cortex and hippocampus of these mice. Moreover, while in young transgenic mice the hTauVLW immunostained transgene product is observed in both cell bodies and dendrites, the hTauVLW mutant protein is only detected in the neuronal cell bodies when Park-2 gene is additionally deleted. Moreover, DNA fragmentation was detected by the TUNEL method, and cerebral atrophy is also present in these regions. The levels of phosphorylated tau and Hsp70 are increased in the double-mutant mice, while CHIP expression in hippocampus is lower when the Park-2 gene is deleted. Thus, the combination of Park-2 gene deletion with hTauVLW transgene overexpression in mice produces serious neuropathological effects, which reflect the existence of some relationship between both proteins. PMID:18376058

  20. Overexpression of transforming growth factor-alpha causes liver enlargement and increased hepatocyte proliferation in transgenic mice.

    PubMed Central

    Webber, E. M.; Wu, J. C.; Wang, L.; Merlino, G.; Fausto, N.

    1994-01-01

    Transforming growth factor-alpha (TGF-alpha) expression is associated with hepatocyte DNA replication both in vivo and in culture. Our previous work using TGF-alpha transgenic mice showed that constitutive overexpression of this growth factor in the liver causes hepatic tumors in 75 to 80% of the animals at 12 to 15 months of age. To understand the cellular events by which TGF-alpha overexpression leads to abnormal liver growth, we examined hepatocyte proliferative activity in young and old TGF-alpha transgenic mice and hepatocyte ploidy in normal, dysplastic, and neoplastic livers of these animals. At 4 weeks of age, transgenic mice had higher liver weights and liver weight/body weight ratios than non-transgenic mice of the same age and hepatocyte proliferative activity, measured by 3H-thymidine incorporation after 3- and 7-day infusion, proliferating cell nuclear antigen staining, and mitotic index determination, was 2 to 3 times higher than in controls. In both transgenic and non-transgenic mice hepatocyte proliferation declined with age but the decrease was much more pronounced in control animals, so that at 8 months of age, hepatocyte replication was 8 to 10 times higher in transgenic animals. Surprisingly, however, transgenic and non-transgenic mice at this age had similar liver weight/body weight ratios. Labeling studies done in 3-month-old animals revealed that hepatocyte turnover was much faster in transgenic than in control animals, suggesting that a homeostatic compensatory mechanism involving cell death tended to restore normal liver weight/body weight ratios in older transgenic mice. Ploidy analyses showed that at 4 weeks of age transgenic mice had a higher proportion of diploid and tetraploid hepatocytes and that the hepatocellular tumors which developed in TGF-alpha transgenic mice at 13 months of age contained a higher fraction of diploid hepatocytes than that present in adjacent tissue or in dysplastic livers. The results demonstrate that

  1. Visualization of Signaling Molecules During Neutrophil Recruitment in Transgenic Mice Expressing FRET Biosensors.

    PubMed

    Mizuno, Rei; Kamioka, Yuji; Sakai, Yoshiharu; Matsuda, Michiyuki

    2016-01-01

    A number of chemical mediators regulate neutrophil recruitment to inflammatory sites either positively or negatively. Although the actions of each chemical mediator on the intracellular signaling networks controlling cell migration have been studied with neutrophils cultured in vitro, how such chemical mediators act cooperatively or counteractively in vivo remains largely unknown. To understand the mechanisms regulating neutrophil recruitment to the inflamed intestine in vivo, we recently generated transgenic mice expressing biosensors based on FRET (Förster resonance energy transfer) and set up two-photon excitation microscopy to observe the gastrointestinal tract in living mice. By measuring FRET in neutrophils, we showed activity changes of protein kinases in the neutrophils recruited to inflamed intestines. In this chapter, we describe the protocol used to visualize the protein kinase activities in neutrophils of the inflamed intestine of transgenic mice expressing the FRET biosensors. PMID:27246030

  2. Welfare assessment and phenotype characterisation of transgenic mice.

    PubMed

    Mertens, Claudia; Rülicke, Thomas

    2007-01-01

    Induced mutations can cause new and unpredictable phenotypes and may impact the health and welfare of animals. Impairments may arise within normal husbandry and breeding regimes i.e. before starting to do experiments. In order to apply the 3R principles and to use transgenic animals under high scientific and welfare standards, two structured forms for individual health monitoring and strain characterisation have been developed. They are available at: www.vu-wien.ac.at/labortierkunde or www.altex.ch. PMID:19835056

  3. Skeletal Phenotype of Transgenic Mice Expressing the Beta1 Integrin Cytoplasmic Tail In Osteoblasts

    NASA Technical Reports Server (NTRS)

    Globus, R. K.; vanderMeulen, M. C. H.; Damsky, D.; Kim, J.-B.; Amblard, D.; Amblard, D.; Nishimura, Y.; Almeida, E.; Iwaniec, U. T.; Wronski, T. J.; Dalton, Bonnie (Technical Monitor)

    2002-01-01

    To define the physiologic role of beta1 integrin in bone formation and mechanical loading, transgenic mice were generated by expressing the cytoplasmic tall and transmembrane domain of Beta1 integrin under the control of the osteocalcin promoter. In cultured cells, this truncated fragment of Beta1 can act as a dominant negative. Previously, the matrix of calvariae was shown to be abnormal in transgenic (TG) compared to wildtype (WT) mice. In this study, we analyzed appendicular bone in TG and WT, male and female mice at 14, 35, 63, 90 and 365 days old (n=8-12/gp). To assess beta1 integrin function in mechanical loading, a pilot study using hindlimb unloading by tail suspension was performed. 35d old TG and WT females were hindlimb unloaded for 4 wks (n=3-5). Body mass, bone mineral content, histomorphometric (distal femur) and biomechanical parameters were analyzed. Statistical significance (P less than.05) was defined by ANOVA using the Tukey-Kramer post-hoc test. We confirmed transgene expression by immunoprecipitating then immunoblotting bone lysates using an antibody against the beta1 tail. Body masses of TG mice at 63, 90 and 365d old were greater (16-25%) than WT. Some TG female mice at 365d appeared obese; mean abdominal fat mass was 415% greater in TG than WT mice. Tibiae were longer (5-7%) in TG than WT mice at 63 and 90d. Tibial mineral mass of 35d males was 7% lower in TG than WT mice, but at 63d was 21% higher. The % osteoblast surface in 35d TG mice was 20% higher than WT, and at 63d was 17% lower, while % osteoclast surface did not differ. In 365d mice, cancellous bone volume (125%) and endocortical mineral apposition rate (40%) were greater in TG than WT males but not females. In WT mice, hindlimb unloading caused a reduction in mineral mass of tibiae (-20%) and lumbar vertebrae (-22%) relative to normally loaded controls. Surprisingly, hindlimb unloading also caused a relative reduction (-13%) in humerus mass. The effects of hindlimb unloading on

  4. Regulatory region in choline acetyltransferase gene directs developmental and tissue-specific expression in transgenic mice.

    PubMed Central

    Lönnerberg, P; Lendahl, U; Funakoshi, H; Arhlund-Richter, L; Persson, H; Ibáñez, C F

    1995-01-01

    Acetylcholine, one of the main neurotransmitters in the nervous system, is synthesized by the enzyme choline acetyltransferase (ChAT; acetyl-CoA:choline O-acetyltransferase, EC 2.3.1.6). The molecular mechanisms controlling the establishment, maintenance, and plasticity of the cholinergic phenotype in vivo are largely unknown. A previous report showed that a 3800-bp, but not a 1450-bp, 5' flanking segment from the rat ChAT gene promoter directed cell type-specific expression of a reporter gene in cholinergic cells in vitro. Now we have characterized a distal regulatory region of the ChAT gene that confers cholinergic specificity on a heterologous downstream promoter in a cholinergic cell line and in transgenic mice. A 2342-bp segment from the 5' flanking region of the ChAT gene behaved as an enhancer in cholinergic cells but as a repressor in noncholinergic cells in an orientation-independent manner. Combined with a heterologous basal promoter, this fragment targeted transgene expression to several cholinergic regions of the central nervous system of transgenic mice, including basal forebrain, cortex, pons, and spinal cord. In eight independent transgenic lines, the pattern of transgene expression paralleled qualitatively and quantitatively that displayed by endogenous ChAT mRNA in various regions of the rat central nervous system. In the lumbar enlargement of the spinal cord, 85-90% of the transgene expression was targeted to the ventral part of the cord, where cholinergic alpha-motor neurons are located. Transgene expression in the spinal cord was developmentally regulated and responded to nerve injury in a similar way as the endogenous ChAT gene, indicating that the 2342-bp regulatory sequence contains elements controlling the plasticity of the cholinergic phenotype in developing and injured neurons. Images Fig. 1 Fig. 2 PMID:7732028

  5. Aspects of achondroplasia in the skulls of dwarf transgenic mice: a cephalometric study.

    PubMed

    Bloom, Melissa Wadler; Murakami, Shunichi; Cody, Dianna; Montufar-Solis, Dina; Duke, Pauline Jackie

    2006-03-01

    Achondroplasia, the most common short-limbed dwarfism in humans, results from a single nucleotide substitution in the gene for fibroblast growth factor receptor 3 (FGFR3). FGFR3 regulates bone growth in part via the mitogen-activated protein kinase pathway (MAPK). To examine the role of this pathway in chondrocyte differentiation, a transgenic mouse was generated that expresses a constitutively active mutant of MEK1 in chondrocytes and exhibits dwarfing characteristics typical of human achondroplasia, i.e., shortened axial and appendicular skeletons, mid-facial hypoplasia, and dome-shaped cranium. In this study, cephalometrics of the MEK1 mutant skulls were assessed to determine if the MEK1 mice are a good model of achondroplasia. Skull length, arc of the cranial vault, and area, maximum and minimum diameters of the brain case were measured on digitized radiographs of skulls of MEK1 and control mice. Cranial base and nasal bone length and foramen magnum diameter were measured on midsagittal micro-CT sections. Data were normalized by dividing by the cube root of each animal's weight. Transgenic mice exhibited a domed skull, deficient midface, and (relatively) prognathic mandible and had a shorter cranial base and nasal bone than the wild-type. Skull length was significantly less in transgenic mice, but cranial arc was significantly greater. The brain case was larger and more circular and minimum diameter of the brain case was significantly greater in transgenic mice. The foramen magnum was displaced anteriorly but not narrowed. MEK1 mouse cephalometrics confirm these mice as a model for achondroplasia, demonstrating that the MAP kinase signaling pathway is involved in FGF signaling in skeletal development. PMID:16463380

  6. Transcriptional insulation of the human keratin 18 gene in transgenic mice.

    PubMed Central

    Neznanov, N; Thorey, I S; Ceceña, G; Oshima, R G

    1993-01-01

    Expression of the 10-kb human keratin 18 (K18) gene in transgenic mice results in efficient and appropriate tissue-specific expression in a variety of internal epithelial organs, including liver, lung, intestine, kidney, and the ependymal epithelium of brain, but not in spleen, heart, or skeletal muscle. Expression at the RNA level is directly proportional to the number of integrated K18 transgenes. These results indicate that the K18 gene is able to insulate itself both from the commonly observed cis-acting effects of the sites of integration and from the potential complications of duplicated copies of the gene arranged in head-to-tail fashion. To begin to identify the K18 gene sequences responsible for this property of transcriptional insulation, additional transgenic mouse lines containing deletions of either the 5' or 3' distal end of the K18 gene have been characterized. Deletion of 1.5 kb of the distal 5' flanking sequence has no effect upon either the tissue specificity or the copy number-dependent behavior of the transgene. In contrast, deletion of the 3.5-kb 3' flanking sequence of the gene results in the loss of the copy number-dependent behavior of the gene in liver and intestine. However, expression in kidney, lung, and brain remains efficient and copy number dependent in these transgenic mice. Furthermore, herpes simplex virus thymidine kinase gene expression is copy number dependent in transgenic mice when the gene is located between the distal 5'- and 3'-flanking sequences of the K18 gene. Each adult transgenic male expressed the thymidine kinase gene in testes and brain and proportionally to the number of integrated transgenes. We conclude that the characteristic of copy number-dependent expression of the K18 gene is tissue specific because the sequence requirements for transcriptional insulation in adult liver and intestine are different from those for lung and kidney. In addition, the behavior of the transgenic thymidine kinase gene in testes and

  7. Virus-Neutralizing Monoclonal Antibody Expressed in Milk of Transgenic Mice Provides Full Protection against Virus-Induced Encephalitis

    PubMed Central

    Kolb, Andreas F.; Pewe, Lecia; Webster, John; Perlman, Stanley; Whitelaw, C. Bruce A.; Siddell, Stuart G.

    2001-01-01

    Neutralizing antibodies represent a major host defense mechanism against viral infections. In mammals, passive immunity is provided by neutralizing antibodies passed to the offspring via the placenta or the milk as immunoglobulin G and secreted immunoglobulin A. With the long-term goal of producing virus-resistant livestock, we have generated mice carrying transgenes that encode the light and heavy chains of an antibody that is able to neutralize the neurotropic JHM strain of murine hepatitis virus (MHV-JHM). MHV-JHM causes acute encephalitis and acute and chronic demyelination in susceptible strains of mice and rats. Transgene expression was targeted to the lactating mammary gland by using the ovine β-lactoglobulin promoter. Milk from these transgenic mice contained up to 0.7 mg of recombinant antibody/ml. In vitro analysis of milk derived from different transgenic lines revealed a linear correlation between antibody expression and virus-neutralizing activity, indicating that the recombinant antibody is the major determinant of MHV-JHM neutralization in murine milk. Offspring of transgenic and control mice were challenged with a lethal dose of MHV-JHM. Litters suckling nontransgenic dams succumbed to fatal encephalitis, whereas litters suckling transgenic dams were fully protected against challenge, irrespective of whether they were transgenic. This demonstrates that a single neutralizing antibody expressed in the milk of transgenic mice is sufficient to completely protect suckling offspring against MHV-JHM-induced encephalitis. PMID:11222704

  8. Epidermal Expression of Intercellular Adhesion Molecule 1 is Not a Primary Inducer of Cutaneous Inflammation in Transgenic Mice

    NASA Astrophysics Data System (ADS)

    Williams, Ifor R.; Kupper, Thomas S.

    1994-10-01

    Keratinocytes at sites of cutaneous inflammation have increased expression of intercellular adhesion molecule 1 (ICAM-1), a cytokine-inducible adhesion molecule which binds the leukocyte integrins LFA-1 and Mac-1. Transgenic mice were prepared in which the expression of mouse ICAM-1 was targeted to basal keratinocytes by using the human K14 keratin promoter. The level of constitutive expression attained in the transgenic mice exceeded the peak level of ICAM-1 expression induced on nontransgenic mouse keratinocytes in vitro by optimal combinations of interferon γ and tumor necrosis factor α or in vivo by proinflammatory stimuli such as phorbol 12-myristate 13-acetate. In vitro adhesion assays demonstrated that cultured transgenic keratinocytes were superior to normal keratinocytes as a substrate for the LFA-1-dependent binding of mouse T cells, confirming that the transgene-encoded ICAM-1 was expressed in a functional form. However, the high level of constitutive ICAM-1 expression achieved on keratinocytes in vivo in these transgenic mice did not result in additional recruitment of CD45^+ leukocytes into transgenic epidermis, nor did it elicit dermal inflammation. Keratinocyte ICAM-1 expression also did not potentiate contact-hypersensitivity reactions to epicutaneous application of haptens. The absence of a spontaneous phenotype in these transgenic mice was not the result of increased levels of soluble ICAM-1, since serum levels of soluble ICAM-1 were equal in transgenic mice and controls. We conclude that elevated ICAM-1 expression on keratinocytes cannot act independently to influence leukocyte trafficking and elicit cutaneous inflammation.

  9. Tetracycline-inducible system for regulation of skeletal muscle-specific gene expression in transgenic mice

    NASA Technical Reports Server (NTRS)

    Grill, Mischala A.; Bales, Mark A.; Fought, Amber N.; Rosburg, Kristopher C.; Munger, Stephanie J.; Antin, Parker B.

    2003-01-01

    Tightly regulated control of over-expression is often necessary to study one aspect or time point of gene function and, in transgenesis, may help to avoid lethal effects and complications caused by ubiquitous over-expression. We have utilized the benefits of an optimized tet-on system and a modified muscle creatine kinase (MCK) promoter to generate a skeletal muscle-specific, doxycycline (Dox) controlled over-expression system in transgenic mice. A DNA construct was generated in which the codon optimized reverse tetracycline transactivator (rtTA) was placed under control of a skeletal muscle-specific version of the mouse MCK promoter. Transgenic mice containing this construct expressed rtTA almost exclusively in skeletal muscles. These mice were crossed to a second transgenic line containing a bi-directional promoter centered on a tet responder element driving both a luciferase reporter gene and a tagged gene of interest; in this case the calpain inhibitor calpastatin. Compound hemizygous mice showed high level, Dox dependent muscle-specific luciferase activity often exceeding 10,000-fold over non-muscle tissues of the same mouse. Western and immunocytochemical analysis demonstrated similar Dox dependent muscle-specific induction of the tagged calpastatin protein. These findings demonstrate the effectiveness and flexibility of the tet-on system to provide a tightly regulated over-expression system in adult skeletal muscle. The MCKrtTA transgenic lines can be combined with other transgenic responder lines for skeletal muscle-specific over-expression of any target gene of interest.

  10. Premature lethality, hyperactivity, and aberrant phosphorylation in transgenic mice expressing a constitutively active form of Fyn

    PubMed Central

    Xia, Di; Götz, Jürgen

    2014-01-01

    The kinase Fyn, the microtubule-associated protein tau and the peptide amyloid-β (Aβ) constitute a toxic triad in Alzheimer's disease (AD). Tau's subcellular localization is mainly regulated by phosphorylation whereas Fyn's localization is dictated by palmitoylation targeting it to the plasma membrane in a reversible manner. We have previously shown that tau is required for Fyn to be targeted to the dendritic spine. We had also shown that a truncated form of tau (Δtau) that accumulates in the cell soma is capable of trapping Fyn and preventing it from entering the spine. Here we determined that palmitoylation is required for Fyn's membrane and spine localization. We further evaluated the functional consequences of neuronal over-expression of the constitutively active Y531F mutant form of Fyn (FynCA) in transgenic mice. We found that the FynCA transgenic mice displayed a reduced weight, a massively reduced lifespan and a high level of hyperactivity. The lifespan of the FynCA mice was only slightly extended by crossing them with Δtau transgenic mice, possibly reflecting differences in expression patterns of the transgenes and high levels of transgenic FynCA compared to endogenous Fyn. Analysis of synaptosomes revealed that FynCA accumulated at high levels in the spine, resulting in increased levels of the NMDA receptor subunit NR2b phosphorylated at residue Y1472. Tau was strongly phosphorylated at the AT8 epitope S202/T205 as shown by Western blot and immunohistochemistry indicating that an increased tyrosine kinase activity of Fyn has down-stream consequences for serine/threonine-directed phosphorylation. PMID:24860422

  11. Expression of a Chimeric Antigen Receptor in Multiple Leukocyte Lineages in Transgenic Mice.

    PubMed

    Yong, Carmen S M; Westwood, Jennifer A; Schröder, Jan; Papenfuss, Anthony T; von Scheidt, Bianca; Moeller, Maria; Devaud, Christel; Darcy, Phillip K; Kershaw, Michael H

    2015-01-01

    Genetically modified CD8+ T lymphocytes have shown significant anti-tumor effects in the adoptive immunotherapy of cancer, with recent studies highlighting a potential role for a combination of other immune subsets to enhance these results. However, limitations in present genetic modification techniques impose difficulties in our ability to fully explore the potential of various T cell subsets and assess the potential of other leukocytes armed with chimeric antigen receptors (CARs). To address this issue, we generated a transgenic mouse model using a pan-hematopoietic promoter (vav) to drive the expression of a CAR specific for a tumor antigen. Here we present a characterization of the immune cell compartment in two unique vav-CAR transgenic mice models, Founder 9 (F9) and Founder 38 (F38). We demonstrate the vav promoter is indeed capable of driving the expression of a CAR in cells from both myeloid and lymphoid lineage, however the highest level of expression was observed in T lymphocytes from F38 mice. Lymphoid organs in vav-CAR mice were smaller and had reduced cell numbers compared to the wild type (WT) controls. Furthermore, the immune composition of F9 mice differed greatly with a significant reduction in lymphocytes found in the thymus, lymph node and spleen of these mice. To gain insight into the altered immune phenotype of F9 mice, we determined the chromosomal integration site of the transgene in both mouse strains using whole genome sequencing (WGS). We demonstrated that compared to the 7 copies found in F38 mice, F9 mice harbored almost 270 copies. These novel vav-CAR models provide a ready source of CAR expressing myeloid and lymphoid cells and will aid in facilitating future experiments to delineate the role for other leukocytes for adoptive immunotherapy against cancer. PMID:26505904

  12. Expression of a Chimeric Antigen Receptor in Multiple Leukocyte Lineages in Transgenic Mice

    PubMed Central

    Yong, Carmen S. M.; Westwood, Jennifer A.; Schröder, Jan; Papenfuss, Anthony T.; von Scheidt, Bianca; Moeller, Maria; Devaud, Christel

    2015-01-01

    Genetically modified CD8+ T lymphocytes have shown significant anti-tumor effects in the adoptive immunotherapy of cancer, with recent studies highlighting a potential role for a combination of other immune subsets to enhance these results. However, limitations in present genetic modification techniques impose difficulties in our ability to fully explore the potential of various T cell subsets and assess the potential of other leukocytes armed with chimeric antigen receptors (CARs). To address this issue, we generated a transgenic mouse model using a pan-hematopoietic promoter (vav) to drive the expression of a CAR specific for a tumor antigen. Here we present a characterization of the immune cell compartment in two unique vav-CAR transgenic mice models, Founder 9 (F9) and Founder 38 (F38). We demonstrate the vav promoter is indeed capable of driving the expression of a CAR in cells from both myeloid and lymphoid lineage, however the highest level of expression was observed in T lymphocytes from F38 mice. Lymphoid organs in vav-CAR mice were smaller and had reduced cell numbers compared to the wild type (WT) controls. Furthermore, the immune composition of F9 mice differed greatly with a significant reduction in lymphocytes found in the thymus, lymph node and spleen of these mice. To gain insight into the altered immune phenotype of F9 mice, we determined the chromosomal integration site of the transgene in both mouse strains using whole genome sequencing (WGS). We demonstrated that compared to the 7 copies found in F38 mice, F9 mice harbored almost 270 copies. These novel vav-CAR models provide a ready source of CAR expressing myeloid and lymphoid cells and will aid in facilitating future experiments to delineate the role for other leukocytes for adoptive immunotherapy against cancer. PMID:26505904

  13. Expression of Growth Hormone Genes in Transgenic Mice

    PubMed Central

    Palmiter, Richard D.; Hammer, Robert E.; Brinster, Ralph L.

    2016-01-01

    OVERVIEW Human or rat growth hormone (GH) genes have been introduced into all cells of a mouse by microinjection of fertilized eggs but they were not expressed under their own promoters. However, substitution of a mouse metallothionein (MT) promoter allowed expression and regulation comparable to that of the endogenous MT genes. These fusion genes have been used to stimulate the growth of both normal mice and dwarf mice that lack sufficient GH. Substitution of a rat elastase-I promoter directed expression of GH exclusively to the acinar cells of the pancreas. Progress has been made towards developing the hGH gene into a vector that is not expressed in vivo unless an enhancer element is inserted. Recombination between overlapping DNA fragments derived from a MThGH gene, each of which is nonfunctional, has been observed when they are coinjected into mouse eggs. In some cases, functional hGH was produced as evidenced by enhanced growth of the mice.

  14. Glucose homeostasis and insulin sensitivity in growth hormone-transgenic mice: a cross-sectional analysis.

    PubMed

    Boparai, Ravneet K; Arum, Oge; Khardori, Romesh; Bartke, Andrzej

    2010-10-01

    In contrast to its stimulatory effects on musculature, bone, and organ development, and its lipolytic effects, growth hormone (GH) opposes insulin effects on glucose metabolism. Chronic GH overexposure is thought to result in insulin insensitivity and decreased blood glucose homeostatic control. Yet, despite the importance of this concept for basic biology, as well as human conditions of GH excess or deficiency, no systematic assessment of the impact of GH over- expression on glucose homeostasis and insulin sensitivity has been conducted. We report that male and female adult GH transgenic mice have enhanced glucose tolerance compared to littermate controls and this effect is not dependent on age or on the particular heterologous GH transgene used. Furthermore, increased glucose-stimulated insulin secretion, augmented insulin sensitivity, and muted gluconeogenesis were also observed in bovine GH overexpressing mice. These results show that markedly increased systemic GH concentration in GH-transgenic mice exerts unexpected beneficial effects on glucose homeostasis, presumably via a compensatory increase in insulin release. The counterintuitive nature of these results challenges previously held presumptions of the physiology of these mice and other states of GH overexpression or suppression. In addition, they pose intriguing queries about the relationships between GH, endocrine control of metabolism, and aging. PMID:20707609

  15. Chemopreventive effect of Curcuma longa Linn on liver pathology in HBx transgenic mice.

    PubMed

    Kim, Jungsun; Ha, Hye-Lin; Moon, Hyung-Bae; Lee, Yeon-Weol; Cho, Chong-Kwan; Yoo, Hwa-Seung; Yu, Dae-Yeul

    2011-06-01

    Unlike other forms of hepatocellular carcinoma (HCC), HCC induced by hepatitis B virus (HBV) infection shows a poor prognosis after conventional therapies. HBV induces liver cirrhosis and HCC. Many researchers have made efforts to find new substances that suppress the activity of HBV. Curcuma longa Linn (CLL) has been used for traditional medicine and food in Asia, especially in India, and has shown chemopreventive effects in a HBV-related in vitro model. This in vivo study was designed to seek the chemopreventive effects of CLL and its mechanisms. CLL mixture concentrated with dextrose water by boiling was lyophilized. CLL extracts were administrated to HBV X protein (HBx) transgenic mice aged 4 weeks for 2 to 4 weeks and aged 6 months for 3 months. After administration, histological changes in the liver tissue and expression of HBx-related genes were investigated. CLL-treated mice showed less visceral fat, a smaller liver/body weight ratio and delayed liver pathogenesis. Proliferating cell nuclear antigen (PCNA) expression was also increased in CLL-treated HBx transgenic mice, indicating regeneration of damaged liver tissue. CLL treatment decreased expression of HBx and increased p21 and cyclin D1 in livers of HBx transgenic mice. In addition, p-p53 was increased after CLL treatment. These results suggest that CLL can have beneficial effects on the early and late stages of liver pathogenesis, preventing and delaying liver carcinogenesis. This drug should be considered as a potential chemopreventive agent for HBV-related hepatocarcinogenesis. PMID:21190953

  16. Overexpression of urokinase-type plasminogen activator in transgenic mice is correlated with impaired learning.

    PubMed Central

    Meiri, N; Masos, T; Rosenblum, K; Miskin, R; Dudai, Y

    1994-01-01

    Transgenic mice designated alpha MUPA overproduce in the brain murine urokinase-type plasminogen activator (uPA), an extracellular protease implicated in tissue remodeling. We have now localized, by in situ hybridization, extensive signal of uPA mRNA in the alpha MUPA cortex, hippocampus, and amygdala, sites that were not labeled in counterpart wild-type mice. Furthermore, biochemical measurements reveal a remarkably high level of enzymatic activity of uPA in the cortex and hippocampus of alpha MUPA compared with wild-type mice. We have used the alpha MUPA mice to examine whether the abnormal level of uPA in the cortex and the limbic system affects learning ability. We report that alpha MUPA mice perform poorly in tasks of spatial, olfactory, and taste-aversion learning, while displaying normal sensory and motor capabilities. Our results suggest that uPA is involved in neural processes subserving a variety of learning types. Images PMID:8159723

  17. Transgenic expression of Dspp partially rescued the long bone defects of Dmp1-null mice.

    PubMed

    Jani, Priyam H; Gibson, Monica P; Liu, Chao; Zhang, Hua; Wang, Xiaofang; Lu, Yongbo; Qin, Chunlin

    2016-01-01

    Dentin matrix protein 1 (DMP1) and dentin sialophosphoprotein (DSPP) belong to the Small Integrin-Binding Ligand N-linked Glycoprotein (SIBLING) family. In addition to the features common to all SIBLING members, DMP1 and DSPP share several unique similarities in chemical structure, proteolytic activation and tissue localization. Mutations in, or deletion of DMP1, cause autosomal recessive hypophosphatemic rickets along with dental defects; DSPP mutations or its ablation are associated with dentinogenesis imperfecta. While the roles and functional mechanisms of DMP1 in osteogenesis have been extensively studied, those of DSPP in long bones have been studied only to a limited extent. Previous studies by our group revealed that transgenic expression of Dspp completely rescued the dentin defects of Dmp1-null (Dmp1(-/-)) mice. In this investigation, we assessed the effects of transgenic Dspp on osteogenesis by analyzing the formation and mineralization of the long bones in Dmp1(-/-) mice that expresses a transgene encoding full-length DSPP driven by a 3.6-kb rat Col1a1 promoter (referred as "Dmp1(-/-);Dspp-Tg mice"). We characterized the long bones of the Dmp1(-/-);Dspp-Tg mice at different ages and compared them with those from Dmp1(-/-) and Dmp1(+/-) (normal control) mice. Our analyses showed that the long bones of Dmp1(-/-);Dspp-Tg mice had a significant increase in cortical bone thickness, bone volume and mineral density along with a remarkable restoration of trabecular thickness compared to those of the Dmp1(-/-) mice. The long bones of Dmp1(-/-);Dspp-Tg mice underwent a dramatic reduction in the amount of osteoid, significant improvement of the collagen fibrillar network, and better organization of the lacunocanalicular system, compared to the Dmp1(-/-) mice. The elevated levels of biglycan, bone sialoprotein and osteopontin in Dmp1(-/-) mice were also noticeably corrected by the transgenic expression of Dspp. These findings suggest that DSPP and DMP1 may function

  18. The maedi-visna virus Tat protein induces multiorgan lymphoid hyperplasia in transgenic mice.

    PubMed Central

    Vellutini, C; Philippon, V; Gambarelli, D; Horschowski, N; Nave, K A; Navarro, J M; Auphan, M; Courcoul, M A; Filippi, P

    1994-01-01

    Sheep infected with maedi-visna virus experience immunological disorders leading to progressive chronic diseases involving the brain, lung, spleen, and lymph nodes. To study the biological activity of the viral transactivating Tat protein, we generated transgenic mice carrying the tat gene. Analysis of the transgenic mouse tissues for tat mRNA revealed that while low at the messenger level, the expression of the transgene correlated with dramatic follicular lymphoproliferative disorders involving the lung, spleen, lymph nodes, and skin. This finding suggests that the viral protein possesses a high pathological potency. Our findings show that the maedi-visna virus tat gene product contributes to the pathogenesis of multiorgan proliferative disorders associated with maedi-visna virus infection. Images PMID:8035494

  19. Oncogenic potential of guanine nucleotide stimulatory factor alpha subunit in thyroid glands of transgenic mice.

    PubMed Central

    Michiels, F M; Caillou, B; Talbot, M; Dessarps-Freichey, F; Maunoury, M T; Schlumberger, M; Mercken, L; Monier, R; Feunteun, J

    1994-01-01

    Transgenic mice have been used to address the issue of the oncogenic potential of mutant guanine nucleotide stimulatory factor (Gs) alpha subunit in the thyroid gland. The expression of the mutant Arg-201-->His Gs alpha subunit transgene has been directed to murine thyroid epithelial cells by bovine thyroglobulin promoter. The transgenic animals develop hyperfunctioning thyroid adenomas with increased intracellular cAMP levels and high uptake of [125I]iodine and produced elevated levels of circulating triiodothyronine and thyroxine. These animals demonstrate that the mutant form of Gs alpha subunit carries an oncogenic activity, thus supporting the model that deregulation of cAMP level alters growth control in thyroid epithelium. These animals represent models for humans with autonomously functioning thyroid nodules. Images PMID:7937980

  20. Decreased early atherosclerotic lesions in hypertriglyceridemic mice expressing cholesteryl ester transfer protein transgene.

    PubMed Central

    Hayek, T; Masucci-Magoulas, L; Jiang, X; Walsh, A; Rubin, E; Breslow, J L; Tall, A R

    1995-01-01

    The human cholesteryl ester transfer protein (CETP) facilitates the transfer of cholesteryl ester from HDL to triglyceride-rich lipoproteins. The activity of CETP results in a reduction in HDL cholesterol levels, but CETP may also promote reverse cholesterol transport. Thus, the net impact of CETP expression on atherogenesis is uncertain. The influence of hypertriglyceridemia and CETP on the development of atherosclerotic lesions in the proximal aorta was assessed by feeding transgenic mice a high cholesterol diet for 16 wk. 13 out of 14 (93%) hypertriglyceridemic human apo CIII (HuCIII) transgenic (Tg) mice developed atherosclerotic lesions, compared to 18 out of 29 (62%) controls. In HuCIII/CETPTg, human apo AI/CIIITg and HuAI/CIII/CETPTg mice, 7 of 13 (54%), 5 of 10 (50%), and 5 of 13 (38%), respectively, developed lesions in the proximal aorta (P < .05 compared to HuCIIITg). The average number of aortic lesions per mouse in HuCIIITg and controls was 3.4 +/- 0.8 and 2.7 +/- 0.6, respectively in HuCIII/CETPTg, HuAI/CIIIg, and HuAI/CIII/CETPTg mice the number of lesions was significantly lower than in HuCIIITg and control mice: 0.9 +/- 0.4, 1.5 +/- 0.5, and 0.9 +/- 0.4, respectively. There were parallel reductions in mean lesion area. In a separate study, we found an increased susceptibility to dietary atherosclerosis in nonhypertriglyceridemic CETP transgenic mice compared to controls. We conclude that CETP expression inhibits the development of early atherosclerotic lesions but only in hypertriglyceridemic mice. PMID:7560101

  1. In vivo regulation of the heme oxygenase-1 gene in humanized transgenic mice

    PubMed Central

    Kim, Junghyun; Zarjou, Abolfazl; Traylor, Amie M.; Bolisetty, Subhashini; Jaimes, Edgar A.; Hull, Travis D.; George, James F.; Mikhail, Fady M.; Agarwal, Anupam

    2012-01-01

    Heme oxygenase-1 (HO-1) catalyzes the rate-limiting step in heme degradation producing equimolar amounts of carbon monoxide, iron, and biliverdin. Induction of HO-1 is a beneficial response to tissue injury in diverse animal models of diseases including acute kidney injury. In vitro analysis has shown that the human HO-1 gene is transcriptionally regulated by changes in chromatin conformation but whether such control occurs in vivo is not known. To enable such analysis, we generated transgenic mice, harboring an 87-kb bacterial artificial chromosome expressing human HO-1 mRNA and protein and bred these mice with HO-1 knockout mice to generate humanized BAC transgenic mice. This successfully rescued the phenotype of the knockout mice including reduced birth rates, tissue iron overload, splenomegaly, anemia, leukocytosis, dendritic cell abnormalities and survival after acute kidney injury induced by rhabdomyolysis or cisplatin nephrotoxicity. Transcription factors such as USF1/2, JunB, Sp1, and CTCF were found to associate with regulatory regions of the human HO-1 gene in the kidney following rhabdomyolysis. Chromosome Conformation Capture and ChIP-loop assays confirmed this in the formation of chromatin looping in vivo. Thus, these bacterial artificial chromosome humanized HO-1 mice are a valuable model to study the human HO-1 gene providing insight to the in vivo architecture of the gene in acute kidney injury and other diseases. PMID:22495295

  2. Coenzyme Q10 attenuates beta-amyloid pathology in the aged transgenic mice with Alzheimer presenilin 1 mutation.

    PubMed

    Yang, Xifei; Yang, Ying; Li, Geng; Wang, Jianzhi; Yang, Edward S

    2008-02-01

    One of the neuropathological features of Alzheimer's disease (AD) is the deposition of senile plaques containing beta-amyloid (A beta). There is limited evidence for the treatment to arrest A beta pathology of AD. In our present study, we tested the effect of coenzyme Q10 (CoQ10), an endogenous antioxidant and a powerful free radical scavenger, on A beta in the aged transgenic mice overexpressing Alzheimer presenilin 1-L235P (leucine-to-proline mutation at codon 235, 16-17 months old). The treatment by feeding the transgenic mice with CoQ10 for 60 days (1,200 mg kg(-1) day(-1)) partially attenuated A beta overproduction and intracellular A beta deposit in the cortex of the transgenic mice compared with the age-matched untreated transgenic mice. Meanwhile, an increased oxidative stress reaction was detected as evidenced by elevated level of malondialdehyde (MDA) and decreased activity of superoxide dismutase (SOD) in the transgenic mice relative to the wild-type mice, and supplementation of CoQ10 partially decreased MDA level and upregulated the activity of SOD. The results indicate that oxidative stress is enhanced in the brain of the transgenic mice, that this enhancement may further promote A beta 42 overproduction in a vicious formation, and that CoQ10 would be beneficial for the therapy of AD. PMID:18181031

  3. HDL from apoA1 transgenic mice expressing the 4WF isoform is resistant to oxidative loss of function.

    PubMed

    Berisha, Stela Z; Brubaker, Greg; Kasumov, Takhar; Hung, Kimberly T; DiBello, Patricia M; Huang, Ying; Li, Ling; Willard, Belinda; Pollard, Katherine A; Nagy, Laura E; Hazen, Stanley L; Smith, Jonathan D

    2015-03-01

    HDL functions are impaired by myeloperoxidase (MPO), which selectively targets and oxidizes human apoA1. We previously found that the 4WF isoform of human apoA1, in which the four tryptophan residues are substituted with phenylalanine, is resistant to MPO-mediated loss of function. The purpose of this study was to generate 4WF apoA1 transgenic mice and compare functional properties of the 4WF and wild-type human apoA1 isoforms in vivo. Male mice had significantly higher plasma apoA1 levels than females for both isoforms of human apoA1, attributed to different production rates. With matched plasma apoA1 levels, 4WF transgenics had a trend for slightly less HDL-cholesterol versus human apoA1 transgenics. While 4WF transgenics had 31% less reverse cholesterol transport (RCT) to the plasma compartment, equivalent RCT to the liver and feces was observed. Plasma from both strains had similar ability to accept cholesterol and facilitate ex vivo cholesterol efflux from macrophages. Furthermore, we observed that 4WF transgenic HDL was partially (∼50%) protected from MPO-mediated loss of function while human apoA1 transgenic HDL lost all ABCA1-dependent cholesterol acceptor activity. In conclusion, the structure and function of HDL from 4WF transgenic mice was not different than HDL derived from human apoA1 transgenic mice. PMID:25561462

  4. Urinary Bladder Dysfunction in Transgenic Sickle Cell Disease Mice

    PubMed Central

    Claudino, Mário Angelo; Leiria, Luiz Osório Silveira; da Silva, Fábio Henrique; Alexandre, Eduardo Costa; Renno, Andre; Mónica, Fabiola Zakia; de Nucci, Gilberto; Fertrin, Kleber Yotsumoto; Antunes, Edson; Costa, Fernando Ferreira; Franco-Penteado, Carla Fernanda

    2015-01-01

    Background Urological complications associated with sickle cell disease (SCD), include nocturia, enuresis, urinary infections and urinary incontinence. However, scientific evidence to ascertain the underlying cause of the lower urinary tract symptoms in SCD is lacking. Objective Thus, the aim of this study was to evaluate urinary function, in vivo and ex vivo, in the Berkeley SCD murine model (SS). Methods Urine output was measured in metabolic cage for both wild type and SS mice (25-30 g). Bladder strips and urethra rings were dissected free and mounted in organ baths. In isolated detrusor smooth muscle (DSM), relaxant response to mirabegron and isoproterenol (1nM-10μM) and contractile response to (carbachol (CCh; 1 nM-100μM), KCl (1 mM-300mM), CaCl2 (1μM-100mM), α,β-methylene ATP (1, 3 and 10 μM) and electrical field stimulation (EFS; 1-32 Hz) were measured. Phenylephrine (Phe; 10nM-100μM) was used to evaluate the contraction mechanism in the urethra rings. Cystometry and histomorphometry were also performed in the urinary bladder. Results SS mice present a reduced urine output and incapacity to produce typical bladder contractions and bladder emptying (ex vivo), compared to control animals. In DSM, relaxation in response to a selective β3-adrenergic agonist (mirabegron) and to a non-selective β-adrenergic (isoproterenol) agonist were lower in SS mice. Additionally, carbachol, α, β-methylene ATP, KCl, extracellular Ca2+ and electrical-field stimulation promoted smaller bladder contractions in SS group. Urethra contraction induced by phenylephrine was markedly reduced in SS mice. Histological analyses of SS mice bladder revealed severe structural abnormalities, such as reductions in detrusor thickness and bladder volume, and cell infiltration. Conclusions Taken together, our data demonstrate, for the first time, that SS mice display features of urinary bladder dysfunction, leading to impairment in urinary continence, which may have an important role in

  5. Pressure overload-induced hypertrophy in transgenic mice selectively overexpressing AT2 receptors in ventricular myocytes.

    PubMed

    Yan, Xinhua; Schuldt, Adam J T; Price, Robert L; Amende, Ivo; Liu, Fen-Fen; Okoshi, Katashi; Ho, Kalon K L; Pope, Adèle J; Borg, Thomas K; Lorell, Beverly H; Morgan, James P

    2008-03-01

    The role of the angiotensin II type 2 (AT2) receptor in cardiac hypertrophy remains controversial. We studied the effects of AT2 receptors on chronic pressure overload-induced cardiac hypertrophy in transgenic mice selectively overexpressing AT2 receptors in ventricular myocytes. Left ventricular (LV) hypertrophy was induced by ascending aorta banding (AS). Transgenic mice overexpressing AT2 (AT2TG-AS) and nontransgenic mice (NTG-AS) were studied after 70 days of aortic banding. Nonbanded NTG mice were used as controls. LV function was determined by catheterization via LV puncture and cardiac magnetic resonance imaging. LV myocyte diameter and interstitial collagen were determined by confocal microscopy. Atrial natriuretic polypeptide (ANP) and brain natriuretic peptide (BNP) were analyzed by Northern blot. Sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA)2, inducible nitric oxide synthase (iNOS), endothelial NOS, ERK1/2, p70S6K, Src-homology 2 domain-containing protein tyrosine phosphatase-1, and protein serine/threonine phosphatase 2A were analyzed by Western blot. LV myocyte diameter and collagen were significantly reduced in AT2TG-AS compared with NTG-AS mice. LV anterior and posterior wall thickness were not different between AT2TG-AS and NTG-AS mice. LV systolic and diastolic dimensions were significantly higher in AT2TG-AS than in NTG-AS mice. LV systolic pressure and end-diastolic pressure were lower in AT2TG-AS than in NTG-AS mice. ANP, BNP, and SERCA2 were not different between AT2TG-AS and NTG-AS mice. Phospholamban (PLB) and the PLB-to-SERCA2 ratio were significantly higher in AT2TG-AS than in NTG-AS mice. iNOS was higher in AT2TG-AS than in NTG-AS mice but not significantly different. Our results indicate that AT2 receptor overexpression modified the pathological hypertrophic response to aortic banding in transgenic mice. PMID:18178728

  6. Targeted overexpression of luteinizing hormone in transgenic mice leads to infertility, polycystic ovaries, and ovarian tumors.

    PubMed Central

    Risma, K A; Clay, C M; Nett, T M; Wagner, T; Yun, J; Nilson, J H

    1995-01-01

    Hypersecretion of luteinizing hormone (LH) is implicated in infertility and miscarriages in women. A lack of animal models has limited progress in determining the mechanisms of LH toxicity. We have recently generated transgenic mice expressing a chimeric LH beta subunit (LH beta) in gonadotropes. The LH beta chimera contains the C-terminal peptide of the human chorionic gonadotropin beta subunit. Addition of this peptide to bovine LH beta resulted in a hormone with a longer half-life. Furthermore, targeted expression of the LH beta chimera led to elevated LH levels and infertility in female transgenics. These mice ovulated infrequently, maintained a prolonged luteal phase, and developed pathologic ovarian changes such as cyst formation, marked enlargement of ovaries, and granulosa cell tumors. Testosterone and estradiol levels were increased compared to nontransgenic littermates. An unusual extragonadal phenotype was also observed: transgenic females developed hydronephropathy and pyelonephritis. The pathology observed demonstrates a direct association between abnormal secretion of LH and infertility and underscores the utility of the transgenic model for studying how excess LH leads to cyst formation, ovarian tumorigenesis, and infertility. Images Fig. 2 Fig. 3 Fig. 5 PMID:7877975

  7. Pathogenesis of axonal dystrophy and demyelination in αA-crystallin-expressing transgenic mice

    PubMed Central

    Van Rijk, AF; Sweers, MAM; Merkx, GFM; Lammens, M; Bloemendal, H

    2003-01-01

    We recently described a transgenic mouse strain overexpressing hamster αA-crystallin, a small heat shock protein, under direction of the hamster vimentin promoter. As a result myelin was degraded and axonal dystrophy in both central nervous system (especially spinal cord) and peripheral nervous system occurred. Homozygous transgenic mice developed hind limb paralysis after 8 weeks of age and displayed progressive loss of myelin and axonal dystrophy in both the central and peripheral nervous system with ongoing age. Pathologically the phenotype resembled, to a certain extent, neuroaxonal dystrophy. The biochemical findings presented in this paper (activity of the enzymes superoxide dismutase, catalase and transglutamase, myelin protein zero expression levels and blood sugar levels) confirm this pathology and exclude other putative pathologies like Amyothrophic Lateral Sclerosis and Hereditary Motor and Sensory Neuropathy. Consequently, an excessive cytoplasmic accumulation of the transgenic protein or a disturbance of the normal metabolism are considered to cause the observed neuropathology. Therefore, extra-ocular αA-crystallin-expressing transgenic mice may serve as a useful animal model to study neuroaxonal dystrophy. PMID:12801283

  8. Cyclic hair-loss and regrowth in transgenic mice overexpressing an intermediate filament gene.

    PubMed Central

    Powell, B C; Rogers, G E

    1990-01-01

    We have produced transgenic mice containing up to 250 copies of a sheep wool intermediate filament keratin gene to study the effect of its expression on hair structure and development. Several transgenic lines expressed the gene and in the one containing 250 transgenes, a pattern of hair-loss and regrowth was stably established. Successive waves of hair growth follow periods of denuding like the natural progression of hairs in the mouse hair cycle. By in situ hybridization we have shown that the sheep transgenes are expressed at the correct stage in mouse hair development and at a high level. The transgenic hairs contain not only an elevated level of intermediate filament keratin protein but also a decreased level of the filament-associated proteins. This imbalance disrupts the normal ordered array of these proteins in the cells of the hair cortex and leads to weakened fibres which are prematurely lost. Images Fig. 2. Fig. 3. Fig. 4. Fig. 5. Fig. 6. Fig. 7. Fig. 8. PMID:1691707

  9. The a”MAZE”ing world of lung specific transgenic mice

    PubMed Central

    Rawlins, Emma L.; Perl, Anne-Karina

    2013-01-01

    The purpose of this review is to give a comprehensive overview of transgenic mouse lines suitable for studying gene function and cellular lineage relationships in lung development, homeostasis, injury and repair. Many of the mouse strains reviewed in this article have been widely shared within the lung research community and new strains are continuously being developed. There are many useful transgenic lines that work to target subsets of lung cells, but it remains a challenge for investigators to select the correct transgenic modules for their experiment. This review covers both the tetracycline and tamoxifen inducible systems and will primarily focus on conditional lines that target the epithelial cells. We point out the limitations of each strain so investigators can choose the system that will work best for their scientific question. Current mesenchymal and endothelial lines are limited by the fact that they are not lung specific. These lines will be summarized in a brief overview. In addition, useful transgenic reporter mice for studying lineage relationships, promoter activity and signaling pathways will complete our lung specific conditional transgenic mouse-shopping list. PMID:22180870

  10. Abnormal differentiation, hyperplasia and embryonic/perinatal lethality in BK5-T/t transgenic mice

    PubMed Central

    Chen, Xin; Schneider-Broussard, Robin; Hollowell, Debra; McArthur, Mark; Jeter, Collene R.; Benavides, Fernando; DiGiovanni, John; Tang, Dean G.

    2009-01-01

    The cell-of-origin has a great impact on the types of tumors that develop and the stem/progenitor cells have long been considered main targets of malignant transformation. The SV40 large T and small t antigens (T/t), have been targeted to multiple differentiated cellular compartments in transgenic mice. In most of these studies, transgenic animals develop tumors without apparent defects in animal development. In this study, we used the bovine keratin 5 (BK5) promoter to target the T/t antigens to stem/progenitor cell-containing cytokeratin 5 (CK5) cellular compartment. A transgene construct, BK5-T/t, was made and microinjected into the male pronucleus of FVB/N mouse oocytes. After implanting ∼1700 embryos, only 7 transgenics were obtained, including 4 embryos (E9.5, E13, E15, and E20) and 3 postnatal animals, which died at P1, P2, and P18, respectively. Immunohistological analysis revealed aberrant differentiation and prominent hyperplasia in several transgenic CK5 tissues, especially the upper digestive organs (tongue, oral mucosa, esophagus, and forestomach) and epidermis, the latter of which also showed focal dysplasia. Altogether, these results indicate that constitutive expression of the T/t antigens in CK5 cellular compartment results in abnormal epithelial differentiation and leads to embryonic/perinatal animal lethality. PMID:19272531

  11. Expression of the human growth hormone variant gene in cultured fibroblasts and transgenic mice

    SciTech Connect

    Selden, R.F.; Wagner, T.E.; Blethen, S.; Yun, J.S.; Rowe, M.E.; Goodman, H.M. )

    1988-11-01

    The nucleotide sequence of the human growth hormone variant gene, one of the five members of the growth hormone gene family, predicts that it encodes a growth hormone-like protein. As a first step in determining whether this gene is functional in humans, the authors have expressed a mouse methallothionein I/human growth hormone variant fusion gene in mouse L cells and in transgenic mice. The growth hormone variant protein expressed in transiently transfected L cells is distinct from growth hormone itself with respect to reactivity with anti-growth hormone monoclonal antibodies, behavior during column chromatography, and isoelectric point. Transgenic mice expressing the growth hormone variant protein are 1.4- to 1.9-fold larger than nontransgenic controls, suggesting that the protein has growth-promoting properties.

  12. Imaging Neural Activity Using Thy1-GCaMP Transgenic mice

    PubMed Central

    Chen, Qian; Cichon, Joseph; Wang, Wenting; Qiu, Li; Lee, Seok-Jin R.; Campbell, Nolan R.; DeStefino, Nicholas; Goard, Michael J.; Fu, Zhanyan; Yasuda, Ryohei; Looger, Loren L.; Arenkiel, Benjamin R.; Gan, Wen-Biao; Feng, Guoping

    2014-01-01

    Summary The ability to chronically monitor neuronal activity in the living brain is essential for understanding the organization and function of the nervous system. The genetically encoded green fluorescent protein based calcium sensor GCaMP provides a powerful tool for detecting calcium transients in neuronal somata, processes, and synapses that are triggered by neuronal activities. Here we report the generation and characterization of transgenic mice that express improved GCaMPs in various neuronal subpopulations under the control of the Thy1 promoter. In vitro and in vivo studies show that calcium transients induced by spontaneous and stimulus-evoked neuronal activities can be readily detected at the level of individual cells and synapses in acute brain slices, as well as chronically in awake behaving animals. These GCaMP transgenic mice allow investigation of activity patterns in defined neuronal populations in the living brain, and will greatly facilitate dissecting complex structural and functional relationships of neural networks. PMID:23083733

  13. Overexpression of motor protein KIF17 enhances spatial and working memory in transgenic mice.

    PubMed

    Wong, Richard Wing-Chuen; Setou, Mitsutoshi; Teng, Junlin; Takei, Yosuke; Hirokawa, Nobutaka

    2002-10-29

    The kinesin superfamily proteins (KIFs) play essential roles in receptor transportation along the microtubules. KIF17 transports the N-methyl-d-aspartate receptor NR2B subunit in vitro, but its role in vivo is unknown. To clarify this role, we generated transgenic mice overexpressing KIF17 tagged with GFP. The KIF17 transgenic mice exhibited enhanced learning and memory in a series of behavioral tasks, up-regulated NR2B expression with the potential involvement of a transcriptional factor, the cAMP-dependent response element-binding protein, and increased phosphorylation of the cAMP-dependent response element-binding protein. Our results suggest that the motor protein KIF17 contributes to neuronal events required for learning and memory by trafficking fundamental N-methyl-d-aspartate-type glutamate receptors. PMID:12391294

  14. Chemopreventive effect of silymarin on liver pathology in HBV X protein transgenic mice.

    PubMed

    Wu, Yi-Fang; Fu, Shu-Ling; Kao, Cheng-Heng; Yang, Chu-Wen; Lin, Chao-Hsiung; Hsu, Ming-Ta; Tsai, Ting-Fen

    2008-03-15

    There are currently limited therapeutic regimens available for effective treatment of hepatocellular carcinoma (HCC). Silymarin is a naturally derived polyphenolic antioxidant with hepatoprotective properties and is very widely used in clinical application; however, effect of silymarin on spontaneous HCC has not been studied. Silymarin was evaluated for its efficacy against spontaneous carcinogenesis using the HBV X protein (HBx) transgenic model. Silymarin was p.o. given to the HBx transgenic mice from 4 to 6 weeks of age. Our data indicated that silymarin has therapeutic effects on the early stages of liver damage, reversing fatty changes and recovering liver histopathology in a dose-dependent manner. To study the chemopreventive effects on the later stages of carcinogenesis, the mice at 13 months were split into a precancerous group and a group with significant liver carcinogenesis. After silymarin was given to the precancerous mice from 13 to 16 months of age, in contrast to an 80% incidence of HCC development in the untreated transgenic mice, no HCC was detected in any of these mice. Nonetheless, small hyperplastic nodules were detected in 86% of these precancerous mice. In the second group with notable HCC, silymarin was unable to block cancer progression. Although silymarin did not affect HBx expression, intracellular reactive oxygen species levels were decreased, cell proliferation was stimulated, and hepatocyte ultrastructure was found to significantly recover. In conclusion, silymarin exerts beneficial effects on the early stages of liver pathogenesis, preventing and delaying liver carcinogenesis. This drug should be considered as a potential chemopreventive agent for HBV-related hepatocarcinogenesis. PMID:18339886

  15. Excessive Growth Hormone Expression in Male GH Transgenic Mice Adversely Alters Bone Architecture and Mechanical Strength

    PubMed Central

    Lim, S. V.; Marenzana, M.; Hopkinson, M.; List, E. O.; Kopchick, J. J.; Pereira, M.; Javaheri, B.; Roux, J. P.; Chavassieux, P.; Korbonits, M.

    2015-01-01

    Patients with acromegaly have a higher prevalence of vertebral fractures despite normal bone mineral density (BMD), suggesting that GH overexpression has adverse effects on skeletal architecture and strength. We used giant bovine GH (bGH) transgenic mice to analyze the effects of high serum GH levels on BMD, architecture, and mechanical strength. Five-month-old hemizygous male bGH mice were compared with age- and sex-matched nontransgenic littermates controls (NT; n=16/group). Bone architecture and BMD were analyzed in tibia and lumbar vertebrae using microcomputed tomography. Femora were tested to failure using three-point bending and bone cellular activity determined by bone histomorphometry. bGH transgenic mice displayed significant increases in body weight and bone lengths. bGH tibia showed decreases in trabecular bone volume fraction, thickness, and number compared with NT ones, whereas trabecular pattern factor and structure model index were significantly increased, indicating deterioration in bone structure. Although cortical tissue perimeter was increased in transgenic mice, cortical thickness was reduced. bGH mice showed similar trabecular BMD but reduced trabecular thickness in lumbar vertebra relative to controls. Cortical BMD and thickness were significantly reduced in bGH lumbar vertebra. Mechanical testing of femora confirmed that bGH femora have decreased intrinsic mechanical properties compared with NT ones. Bone turnover is increased in favor of bone resorption in bGH tibia and vertebra compared with controls, and serum PTH levels is also enhanced in bGH mice. These data collectively suggest that high serum GH levels negatively affect bone architecture and quality at multiple skeletal sites. PMID:25646711

  16. Impaired Immunogenicity of Meningococcal Neisserial Surface Protein A in Human Complement Factor H Transgenic Mice.

    PubMed

    Lujan, Eduardo; Pajon, Rolando; Granoff, Dan M

    2015-01-01

    Neisserial surface protein A (NspA) is a highly conserved outer membrane protein previously investigated as a meningococcal vaccine candidate. Despite eliciting serum bactericidal activity in mice, a recombinant NspA vaccine failed to elicit serum bactericidal antibodies in a phase 1 clinical trial in humans. The discordant results may be explained by the recent discovery that NspA is a human-specific ligand of the complement inhibitor factor H (FH). Therefore, in humans but not mice, NspA would be expected to form a complex with FH, which could impair human anti-NspA protective antibody responses. To investigate this question, we immunized human FH transgenic BALB/c mice with three doses of recombinant NspA expressed in Escherichia coli microvesicles, with each dose being separated by 3 weeks. Three of 12 (25%) transgenic mice and 13 of 14 wild-type mice responded with bactericidal titers of ≥1:10 in postimmunization sera (P = 0.0008, Fisher's exact test). In contrast, human FH transgenic and wild-type mice immunized with a control meningococcal native outer membrane vesicle vaccine had similar serum bactericidal antibody responses directed at PorA, which is not known to bind human FH, and a mutant factor H binding protein (FHbp) antigen with a >50-fold lower level of FH binding than wild-type FHbp antigen binding.Thus, human FH can impair anti-NspA serum bactericidal antibody responses, which may explain the poor immunogenicity of the NspA vaccine previously tested in humans. A mutant NspA vaccine engineered to have decreased binding to human FH may increase protective antibody responses in humans. PMID:26597984

  17. FHL1 Reduces Dystrophy in Transgenic Mice Overexpressing FSHD Muscular Dystrophy Region Gene 1 (FRG1)

    PubMed Central

    Feeney, Sandra J.; McGrath, Meagan J.; Sriratana, Absorn; Gehrig, Stefan M.; Lynch, Gordon S.; D’Arcy, Colleen E.; Price, John T.; McLean, Catriona A.; Tupler, Rossella; Mitchell, Christina A.

    2015-01-01

    Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal-dominant disease with no effective treatment. The genetic cause of FSHD is complex and the primary pathogenic insult underlying the muscle disease is unknown. Several disease candidate genes have been proposed including DUX4 and FRG1. Expression analysis studies of FSHD report the deregulation of genes which mediate myoblast differentiation and fusion. Transgenic mice overexpressing FRG1 recapitulate the FSHD muscular dystrophy phenotype. Our current study selectively examines how increased expression of FRG1 may contribute to myoblast differentiation defects. We generated stable C2C12 cell lines overexpressing FRG1, which exhibited a myoblast fusion defect upon differentiation. To determine if myoblast fusion defects contribute to the FRG1 mouse dystrophic phenotype, this strain was crossed with skeletal muscle specific FHL1-transgenic mice. We previously reported that FHL1 promotes myoblast fusion in vitro and FHL1-transgenic mice develop skeletal muscle hypertrophy. In the current study, FRG1 mice overexpressing FHL1 showed an improvement in the dystrophic phenotype, including a reduced spinal kyphosis, increased muscle mass and myofiber size, and decreased muscle fibrosis. FHL1 expression in FRG1 mice, did not alter satellite cell number or activation, but enhanced myoblast fusion. Primary myoblasts isolated from FRG1 mice showed a myoblast fusion defect that was rescued by FHL1 expression. Therefore, increased FRG1 expression may contribute to a muscular dystrophy phenotype resembling FSHD by impairing myoblast fusion, a defect that can be rescued by enhanced myoblast fusion via expression of FHL1. PMID:25695429

  18. Compensation of the AKT signaling by ERK signaling in transgenic mice hearts overexpressing TRIM72

    SciTech Connect

    Ham, Young-Mi; Mahoney, Sarah Jane

    2013-06-10

    The AKT and ERK signaling pathways are known to be involved in cell hypertrophy, proliferation, survival and differentiation. Although there is evidence for crosstalk between these two signaling pathways in cellulo, there is less evidence for cross talk in vivo. Here, we show that crosstalk between AKT and ERK signaling in the hearts of TRIM72-overexpressing transgenic mice (TRIM72-Tg) with alpha-MHC promoter regulates and maintains their heart size. TRIM72, a heart- and skeletal muscle-specific protein, downregulates AKT-mTOR signaling via IRS-1 degradation and reduces the size of rat cardiomyocytes and the size of postnatal TRIM72-Tg hearts. TRIM72 expression was upregulated by hypertrophic inducers in cardiomyocytes, while IRS-1 was downregulated by IGF-1. TRIM72 specifically regulated IGF-1-dependent AKT-mTOR signaling, resulting in a reduction of the size of cardiomyocytes. Postnatal TRIM72-Tg hearts were smaller than control-treated hearts with inhibition of AKT-mTOR signaling. However, adult TRIM72-Tg hearts were larger than of control despite the suppression of AKT-mTOR signaling. Activation of ERK, PKC-α, and JNK were observed to be elevated in adult TRIM72-Tg, and these signals were mediated by ET-1 via the ET receptors A and B. Altogether, these results suggest that AKT signaling regulates cardiac hypertrophy in physiological conditions, and ERK signaling compensates for the absence of AKT signaling during TRIM72 overexpression, leading to pathological hypertrophy. -- Highlights: • TRIM72 inhibits AKT signaling through ubiquitination of IRS-1 in cardiac cells. • TRIM72 regulates the size of cardiac cells. • TRIM72 regulates size of postnatal TRIM72-overexpressing transgenic mice hearts. • Adult TRIM72-overexpressing transgenic mice hearts showed cardiac dysfunction. • Adult TRIM72 transgenic mice hearts showed higher expression of endothelin receptors.

  19. Gut microbes define liver cancer risk in mice exposed to chemical and viral transgenic hepatocarcinogens

    PubMed Central

    Fox, J G; Feng, Y; Theve, E J; Raczynski, A R; Fiala, J L A; Doernte, A L; Williams, M; McFaline, J L; Essigmann, J M; Schauer, D B; Tannenbaum, S R; Dedon, P C; Weinman, S A; Lemon, S M; Fry, R C; Rogers, A B

    2013-01-01

    Background and aims Hepatocellular carcinoma (HCC) frequently results from synergism between chemical and infectious liver carcinogens. Worldwide, the highest incidence of HCC is in regions endemic for the foodborne contaminant aflatoxin B1 (AFB1) and hepatitis B virus (HBV) infection. Recently, gut microbes have been implicated in multisystemic diseases including obesity and diabetes. Here, the hypothesis that specific intestinal bacteria promote liver cancer was tested in chemical and viral transgenic mouse models. Methods Helicobacter-free C3H/HeN mice were inoculated with AFB1 and/or Helicobacter hepaticus. The incidence, multiplicity and surface area of liver tumours were quantitated at 40 weeks. Molecular pathways involved in tumourigenesis were analysed by microarray, quantitative real-time PCR, liquid chromatography/mass spectrometry, ELISA, western blot and immunohistochemistry. In a separate experiment, C57BL/6 FL-N/35 mice harbouring a full-length hepatitis C virus (HCV) transgene were crossed with C3H/HeN mice and cancer rates compared between offspring with and without H hepaticus. Results Intestinal colonisation by H hepaticus was sufficient to promote aflatoxin- and HCV transgene-induced HCC. Neither bacterial translocation to the liver nor induction of hepatitis was necessary. From its preferred niche in the intestinal mucus layer, H hepaticus activated nuclear factor-κB (NF-κB)-regulated networks associated with innate and T helper 1 (Th1)-type adaptive immunity both in the lower bowel and liver. Biomarkers indicative of tumour progression included hepatocyte turnover, Wnt/β-catenin activation and oxidative injury with decreased phagocytic clearance of damaged cells. Conclusions Enteric microbiota define HCC risk in mice exposed to carcinogenic chemicals or hepatitis virus transgenes. These results have implications for human liver cancer risk assessment and prevention. PMID:19850960

  20. Transgenic human C-reactive protein is not proatherogenic in apolipoprotein E-deficient mice

    PubMed Central

    Hirschfield, Gideon M.; Gallimore, J. Ruth; Kahan, Melvyn C.; Hutchinson, Winston L.; Sabin, Caroline A.; Benson, G. Martin; Dhillon, Amar P.; Tennent, Glenys A.; Pepys, Mark B.

    2005-01-01

    The association between circulating concentrations of C-reactive protein (CRP) and future atherothrombotic events has provoked speculation about a possible pathogenetic role of CRP. However, we show here that transgenic expression of human CRP had no effect on development, progression, or severity of spontaneous atherosclerosis, or on morbidity or mortality, in male apolipoprotein E (apoE)-deficient C57BL/6 mice up to 56 weeks, despite deposition of human CRP and mouse complement component 3 in the plaques. Although female apoE knockouts develop atherosclerosis more rapidly than males, the human CRP transgene is under sex hormone control and is expressed at human levels only in males. We therefore studied only male mice. The concentration of mouse serum amyloid P component, an extremely sensitive systemic marker of inflammation, remained normal throughout except for transient spikes in response to fighting in a few animals, indicating that atherogenesis in this model is not associated with an acute-phase response. However, among human CRP transgenic mice, the circulating CRP concentration was higher in apoE knockouts than in wild-type controls. The higher CRP values were associated with substantially lower estradiol concentrations in the apoE-deficient animals. Human CRP transgene expression is thus up-regulated in apoE-deficient mice, apparently reflecting altered estrogen levels, despite the absence of other systemic signs of inflammation. Extrapolation to human pathology from this xenogeneic combination of human CRP with apoE deficiency-mediated mouse atherosclerosis must be guarded. Nevertheless, the present results do not suggest that human CRP is either proatherogenic or atheroprotective in vivo. PMID:15919817

  1. Transgenic mice over-expressing carbonic anhydrase I showed aggravated joint inflammation and tissue destruction

    PubMed Central

    2012-01-01

    Background Studies have demonstrated that carbonic anhydrase I (CA1) stimulates calcium salt precipitation and cell calcification, which is an essential step in new bone formation. Our study had reported that CA1 encoding gene has a strong association with rheumatoid arthritis (RA) and ankylosing spondylitis (AS), two rheumatic diseases with abnormal new bone formation and bone resorption in joints. This study investigated the effect of CA1 on joint inflammation and tissue destruction in transgenic mice that over-express CA1 (CA1-Tg). Methods CA1-Tg was generated with C57BL/6J mice by conventional methods. CA1-Tg was treated with collagen-II to induce arthritis (CIA). Wild-type mice, CA1-Tg treated with bovine serum albumin (BSA) and transgenic mice over-expressing PADI4 (PADI4-Tg), a gene known to be involved in rheumatoid arthritis, were used as controls. Histochemistry and X-ray radiographic assay were used to examine joint destruction. Western blotting and real time-PCR were used to examine CA1 expression. Results CIA was observed in 60% of CA1-Tg, 20% of PADI4-Tg and 20% of wild-type mice after collagen injections. No CIA was found in CA1-Tg mice that received injections of BSA. The arthritic score was 5.5 ± 0.84 in the CA1-Tgs but the score was less than 2 in the injected wild-type mice and the PADI4-Tgs. The thickness of the hind paws in the CA1-Tgs was 3.46 ± 0.11 mm, which was thicker than that of PADI4-Tgs (2.23 ± 0.08 mm), wild-type mice (2.08 ± 0.06 mm) and BSA-treated CA1-Tgs (2.04 ± 0.07 mm). Histochemistry showed obvious inflammation, synovial hyperplasia and bone destruction in the joints of CA1-Tg that was not detected in PADI4-Tgs or wild-type mice. X-ray assays showed bone fusion in the paws and spines of CA1-Tg mice. Conclusion Over-expression of CA1 may aggravate joint inflammation and tissue destruction in the transgenic mice. PMID:23256642

  2. Trichostatin A suppresses lung adenocarcinoma development in Grg1 overexpressing transgenic mice

    SciTech Connect

    Liu, Ju; Li, Yan; Dong, Fengyun; Li, Liqun; Masuda, Takahiro; Allen, Thaddeus D.; Lobe, Corrinne G.

    2015-08-07

    Trichostatin A (TSA) is a histone deacetylase inhibitor and a potential therapeutic for various malignancies. The in vivo effect of TSA, however, has not been investigated in a transgenic lung cancer model. Previously, we generated transgenic mice with overexpression of Groucho-related-gene 1 (Grg1) and these mice all developed mucinous lung adenocarcinoma. Grg1 is a transcriptional co-repressor protein, the function of which is thought to depend on HDAC activity. However, functions outside the nucleus have also been proposed. We tested the supposition that Grg1-induced tumorigenesis is HDAC-dependent by assaying the therapeutic effect of TSA in the Grg1 transgenic mouse model. We found that TSA significantly inhibited lung tumorigenesis in Grg1 transgenic mice (p < 0.01). TSA did not affect overall Grg1 protein levels, but instead reduced ErbB1 and ErbB2 expression, which are upregulated by Grg1 in the absence of TSA. We confirmed this effect in A549 cells. Furthermore, lapatinib, an inhibitor of both ErbB1 and ErbB2, effectively masked the effect of TSA on the inhibition of A549 cell proliferation and migration, suggesting TSA does work, at least in part, by downregulating ErbB receptors. We additionally found that TSA reduced the expression of VEGF and VEGFR2, but not basic FGF and FGFR1. Our findings indicate that TSA effectively inhibits Grg1-induced lung tumorigenesis through the down-regulation of ErbB1 and ErbB2, as well as reduced VEGF signaling. This suggests TSA and other HDAC inhibitors could have therapeutic value in the treatment of lung cancers with Grg1 overexpression. - Highlights: • TSA suppresses lung tumorigenesis in Grg1 overexpressing transgenic mice. • TSA does not affect overall Grg1 protein levels in the mice and in A549 cells. • TSA reduces ErbB1 and ErbB2 expression in the mice and in A549 cells. • Lapatinib masks TSA-induced inhibition of A549 cell proliferation and migration. • TSA inhibits VEGF signaling, but not basic FGF

  3. To B or not to B: Role of B cells in pathogenesis of arthritis in HLA transgenic mice

    PubMed Central

    Behrens, Marshall; Smart, Michele; Luckey, David; Luthra, Harvinder; Taneja, Veena

    2011-01-01

    Population studies have shown that amongst all the genetic factors linked with autoimmune disease development, MHC class II genes are the most significant. Experimental autoimmune arthritis resembling human rheumatoid arthritis (RA) can be induced in susceptible strains of mice following immunization with type II collagen (CIA). We generated transgenic mice lacking endogenous class II molecules and expressing various HLA genes including RA-associated, HLA-DRB1*0401 and HLA-DQ8, and RA-resistant, DRB1*0402, genes. The HLA molecules in these mice are expressed on the cell surface and can positively select CD4+ T cells expressing various Vβ T cell receptors. Endogenous class II invariant chain is required for proper functioning of the class II transgene. Arthritis development in transgenic mice is CD4+ and B cells dependent. Studies in humanized mice showed that B cells are required as antigen presenting cells in addition to antibody producing cells for the development of CIA. The transgenic mice expressing *0401 and *0401/DQ8 genes developed sex-biased arthritis with predominantly females being affected, similar to that of human RA. Further, the transgenic mice produced autoantibodies like rheumatoid factor and anti-cyclic antibodies. Antigen presentation by B cells leads to a sex specific immune response in DRB1*0401 mice suggesting a role of B cells and HLA-DR in rendering susceptibility to develop arthritis in females. PMID:21665435

  4. Reversible suppression of an essential gene in adult mice using transgenic RNA interference

    PubMed Central

    McJunkin, Katherine; Mazurek, Anthony; Premsrirut, Prem K.; Zuber, Johannes; Dow, Lukas E.; Simon, Janelle; Stillman, Bruce; Lowe, Scott W.

    2011-01-01

    RNAi has revolutionized loss-of-function genetics by enabling sequence-specific suppression of virtually any gene. Furthermore, tetracycline response elements (TRE) can drive expression of short hairpin RNAs (shRNAs) for inducible and reversible target gene suppression. Here, we demonstrate the feasibility of transgenic inducible RNAi for suppression of essential genes. We set out to directly target cell proliferation by screening an RNAi library against DNA replication factors and identified multiple shRNAs against Replication Protein A, subunit 3 (RPA3). We generated transgenic mice with TRE-driven Rpa3 shRNAs whose expression enforced a reversible cell cycle arrest. In adult mice, the block in cell proliferation caused rapid atrophy of the intestinal epithelium which led to weight loss and lethality within 8–11 d of shRNA induction. Upon shRNA withdrawal, villus atrophy and weight loss were fully reversible. Thus, shRpa3 transgenic mice provide an interesting tool to study tissue maintenance and regeneration. Overall, we have established a robust system that serves the purpose of temperature-sensitive alleles in other model organisms, enabling inducible and reversible suppression of essential genes in a mammalian system. PMID:21482754

  5. The production of transgenic mice expressing human cystathionine beta-synthase to study Down syndrome.

    PubMed

    Butler, Christine; Knox, Aaron J; Bowersox, Jeffrey; Forbes, Stacy; Patterson, David

    2006-05-01

    Down syndrome (DS) is the most common genetic cause of significant cognitive disability. We hypothesize that by identifying metabolic alterations associated with cognitive impairment, it may be possible to develop medical or dietary interventions to ameliorate cognitive disabilities in persons with DS. Evidence suggests that one-carbon/transsulfuration (1C-TS) metabolism is abnormal in persons with DS. Cystathionine beta-synthase (CBS) plays a critical role in this metabolic system. The gene for CBS is on human chromosome 21, and there is evidence of elevated CBS enzyme activity in tissues and cells from individuals with DS. To analyze the possible role of CBS in Down syndrome, we have produced several lines of transgenic mice expressing the human CBS gene. We describe the use of Florescence Situ Hybridization (FISH) analysis to characterize the transgene insertion site for each line. Our initial expression analysis of each transgenic line by RT-PCR shows that the tissue specificity of human CBS mRNA levels in these mice may differ from the tissue specificity of mouse CBS mRNA levels in the same animals. These mice will be invaluable for assessing the regulation of the CBS gene and the role of CBS in cognition. They can also be used to develop therapies that target abnormalities in 1C-TS metabolism to improve cognition in persons with DS. PMID:16541333

  6. Loss of renal microvascular integrity in postnatal Crim1 hypomorphic transgenic mice.

    PubMed

    Wilkinson, Lorine; Gilbert, Thierry; Sipos, Arnold; Toma, Ildiko; Pennisi, David J; Peti-Peterdi, Janos; Little, Melissa H

    2009-12-01

    Crim1 is a cell-surface, transmembrane protein that binds to a variety of cystine knot-containing growth factors, including vascular endothelial growth factor A. In the developing renal glomerulus, Crim1 acts to tether vascular endothelial growth factor A to the podocyte cell surface, thus regulating its release to glomerular endothelial cells. The hypomorphic transgenic mouse (Crim1(KST264/KST264)) has glomerular cysts and severe glomerular vascular defects because of the lack of functional Crim1 in the glomerulus. Adult transgenic mice have a reduced glomerular filtration rate and glomerular capillary defects. We now show that, in these adult transgenic mice, renal vascular defects are not confined to the glomerulus but also extend to the peritubular microvasculature, as live imaging revealed leakiness of both glomerular and peritubular capillaries. An ultrastructural analysis of the microvasculature showed an abnormal endothelium and collagen deposition between the endothelium and the tubular basement membrane, present even in juvenile mice. Overt renal disease, including fibrosis and renin recruitment, was not evident until adulthood. Our study suggests that Crim1 is involved in endothelial maintenance and integrity and its loss contributes to a primary defect in the extraglomerular vasculature. PMID:19776720

  7. Mice Transgenic for a Human Porcine Endogenous Retrovirus Receptor Are Susceptible to Productive Viral Infection†

    PubMed Central

    Martina, Y.; Marcucci, K. T.; Cherqui, S.; Szabo, A.; Drysdale, T.; Srinivisan, U.; Wilson, C. A.; Patience, C.; Salomon, D. R.

    2006-01-01

    Porcine endogenous retrovirus (PERV) is considered one of the major risks in xenotransplantation. No valid animal model has been established to evaluate the risks associated with PERV transmission to human patients by pig tissue xenotransplantation or to study the potential pathogenesis associated with PERV infection. In previous work we isolated two genes encoding functional human PERV receptors and proved that introduction of these into mouse fibroblasts allowed the normally nonpermissive mouse cells to become productively infected (T. A. Ericsson, Y. Takeuchi, C. Templin, G. Quinn, S. F. Farhadian, J. C. Wood, B. A. Oldmixon, K. M. Suling, J. K. Ishii, Y. Kitagawa, T. Miyazawa, D. R. Salomon, R. A. Weiss, and C. Patience, Proc. Natl. Acad. Sci. USA 100:6759-6764, 2003). In the present study we created mice transgenic for human PERV-A receptor 2 (HuPAR-2). After inoculation of transgenic animals with infectious PERV supernatants, viral DNA and RNA were detected at multiple time points, indicating productive replication. This establishes the role of HuPAR-2 in PERV infection in vivo; in addition, these transgenic mice represent a new model for determining the risk of PERV transmission and potential pathogenesis. These mice also create a unique opportunity to study the immune response to PERV infection and test potential therapeutic or preventative modalities. PMID:16537582

  8. Antiviral effects of Stichopus japonicus acid mucopolysaccharide on hepatitis B virus transgenic mice

    NASA Astrophysics Data System (ADS)

    Xin, Yongning; Li, Wei; Lu, Linlin; Zhou, Li; Victor, David W.; Xuan, Shiying

    2016-08-01

    Hepatitis B virus (HBV) is a significant global pathogen and efficient cure for HBV patients is still a challenging goal. We previously reported that acidic mucopolysaccharide from stichopus japonicus selenka (SJAMP) could inhibit HBsAg and HBeAg expression in vitro. However, the potential anti-HBV effects of SJAMP in vivo have not yet been explored. In this study, we show that SJAMP exhibits potent anti-HBV activity in HBV transgenic mice in a dose-dependent manner. Specifically, sixty HBV transgenic male BALB/c mice were randomly selected to receive the treatment of PBS, low dose SJAMP (30 mg kg-1), middle dose SJAMP (40 mg kg-1), high dose SJAMP (50 mg kg-1) and IFN (45 IU kg-1) for 30 d. SJAMP treatment suppressed serum HBV-DNA, and liver HBsAg and HBcAg levels in HBV-transgenic mice. The present study highlights the potential application of SJAMP in HBV therapy.

  9. Evaluation of APP695 Transgenic Mice Bone Marrow Mesenchymal Stem Cells Neural Differentiation for Transplantation

    PubMed Central

    Li, Qian; Jia, Yanjie; Zhang, John; Yang, Jun

    2015-01-01

    Objective. Even though there is a therapeutic potential to treat Alzheimer's disease (AD) with neural cell replenishment and replacement, immunological rejections of stem cell transplantation remain a challenging risk. Autologous stem cells from AD patients however may prove to be a promising candidate. Therefore, we studied the neuronal differentiation efficiency of bone marrow mesenchymal stem cells (MSCs) from APP695 transgenic mice, which share features of human AD. Method. Cultured MSCs from APP695 transgenic mice are used; neuronal differentiation was assessed by immunocytochemistry and Western blot. Correlation with Notch signaling was examined. Autophage flux was assessed by western blot analysis. Results. MSCs from APP695 mice have higher neuronal differentiation efficiency than MSCs from wild type mice (WT MSCs). The expression of Notch-1 signaling decreased during the differentiation process. However, autophagy flux, which is essential for neuronal cell survival and neuronal function, was impaired in the neuronally differentiated counterparts of APP695 MSCs (APP695 MSCs–n). Conclusion. These results suggested autologous MSCs of APP690 mice may not be a good candidate for cell transplantation. PMID:26491658

  10. Regulation of expression of a sheep metallothionein 1a-sheep growth hormone fusion gene in transgenic mice.

    PubMed Central

    Shanahan, C M; Rigby, N W; Murray, J D; Marshall, J T; Townrow, C A; Nancarrow, C D; Ward, K A

    1989-01-01

    Transgenic mice containing a sheep metallothionein 1a-sheep growth hormone fusion gene exhibited low, tissue-specific basal levels of transgene mRNA expression, resulting in slightly elevated levels of circulating growth hormone that did not lead to a detectable increase in growth. After zinc stimulation, high levels of transgene mRNA expression were induced in a number of tissues; these levels correlated with increased levels of circulating growth hormone, resulting in growth increases of up to 1.5 times the levels of controls and unstimulated transgenic mice. After removal of the zinc stimulus, transgene expression and circulating growth hormone concentrations returned to basal levels. Additional evidence from the pattern of developmental expression of the transgene suggests that zinc is the main regulator of this promoter in mice. The demonstrated regulation and low basal level of expression of the sheep metallothionein 1a promoter make it a candidate for use in other mouse transgenic studies and for use in transgenic livestock, in which regulation of expression is essential. Images PMID:2479830

  11. Genetic Biomarkers for ALS Disease in Transgenic SOD1G93A Mice

    PubMed Central

    Calvo, Ana C.; Manzano, Raquel; Atencia-Cibreiro, Gabriela; Oliván, Sara; Muñoz, María J.; Zaragoza, Pilar; Cordero-Vázquez, Pilar; Esteban-Pérez, Jesús; García-Redondo, Alberto; Osta, Rosario

    2012-01-01

    The pathophysiological mechanisms of both familial and sporadic Amyotrophic Lateral Sclerosis (ALS) are unknown, although growing evidence suggests that skeletal muscle tissue is a primary target of ALS toxicity. Skeletal muscle biopsies were performed on transgenic SOD1G93A mice, a mouse model of ALS, to determine genetic biomarkers of disease longevity. Mice were anesthetized with isoflurane, and three biopsy samples were obtained per animal at the three main stages of the disease. Transcriptional expression levels of seventeen genes, Ankrd1, Calm1, Col19a1, Fbxo32, Gsr, Impa1, Mef2c, Mt2, Myf5, Myod1, Myog, Nnt, Nogo A, Pax7, Rrad, Sln and Snx10, were tested in each muscle biopsy sample. Total RNA was extracted using TRIzol Reagent according to the manufacturer's protocol, and variations in gene expression were assayed by real-time PCR for all of the samples. The Pearson correlation coefficient was used to determine the linear correlation between transcriptional expression levels throughout disease progression and longevity. Consistent with the results obtained from total skeletal muscle of transgenic SOD1G93A mice and 74-day-old denervated mice, five genes (Mef2c, Gsr, Col19a1, Calm1 and Snx10) could be considered potential genetic biomarkers of longevity in transgenic SOD1G93A mice. These results are important because they may lead to the exploration of previously unexamined tissues in the search for new disease biomarkers and even to the application of these findings in human studies. PMID:22412900

  12. Postnatal Male Germ Cell Expression of Cre Recombinase in Tex101-iCre Transgenic Mice

    PubMed Central

    Lei, Zhenmin; Lin, Jing; Li, Xian; Li, Shengqiang; Zhou, Huaxin; Araki, Yoshihiko; Lan, Zi-Jian

    2010-01-01

    We have generated a transgenic mouse line which expresses improved Cre recombinase (iCre) under the control of the testis-expressed gene 101 (Tex101) promoter. This transgenic mouse line was named Tex101-iCre. Using the floxed ROSA reporter mice, we found that robust Cre recombinase activity was detected in postnatal testes with weak or no activity in other tissues. Within the testis, Cre recombinase was active in spermatogenic cells as early as the prospermatogonia stage at day 1 after birth. In 30- and 60-day-old mice, positive Cre recombinase activity was detected not only in prospermatogonia but also in spermatogenic cells at later stages of spermatogenesis. There was little or no Cre activity in interstitial cells. Breeding wild-type females with homozygous floxed fibroblast growth factor receptor 2 (Fgfr2) males carrying the Tex101-iCre transgene did not produce any progeny with the floxed Fgfr2 allele. All of the progeny inherited a recombined Fgfr2 allele, indicating that complete deletion of the floxed Fgfr2 allele can be achieved in the male germline by Tex101-iCre mice. Furthermore, FGFR2 protein was not detected in spermatocytes and spermatids of adult Fgfr2fl/fl;Tex101-iCre mice. Taken together, our results suggest that the Tex101-iCre mouse line allows the inactivation of a floxed gene in spermatogenic cells in adult mice, which will facilitate the functional characterization of genes in normal spermatogenesis and male fertility. PMID:20853429

  13. Studies on the thymus of non-obese diabetic (NOD) mice: effect of transgene expression.

    PubMed Central

    O'Reilly, L A; Healey, D; Simpson, E; Chandler, P; Lund, T; Ritter, M A; Cooke, A

    1994-01-01

    The non-obese diabetic (NOD) mouse is a good model of insulin-dependent diabetes mellitus. Autoreactive T cells may play a fundamental role in disease initiation in this model, while disregulation of such cells may result from an abnormal thymic microenvironment. Diabetes is prevented in NOD mice by direct introduction of an E alpha d transgene (NOD-E) or a modified I-A beta chain of NOD origin (NOD-PRO or NOD-ASP). To investigate if disease pathology in NOD mice, protection from disease in transgenic NOD-E and NOD-PRO and partial protection from disease in NOD-ASP can be attributed to alterations in the thymic microenvironment, immunohistochemical and flow cytometric analysis of the thymi of these mouse strains was studied. Thymi from NOD and NOD-E mice showed a progressive increase in thymic B-cell percentage from 12 weeks of age. This was accompanied by a concomitant loss in thymic epithelial cells with the appearance of large epithelial-free areas mainly at the corticomedullary junction, which increased in size and number with age and contained the B-cell clusters. Such thymic B cells did not express CD5 and were absent in CBA, NOD-ASP and NOD-PRO mice as were the epithelial cell-free spaces, even at 5 months of age. Therefore the mechanisms of disease protection in the transgenic NOD-E and NOD-ASP/NOD-PRO mice may differ if these thymic abnormalities are related to disease. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:7523287

  14. Quantitative analysis of lentiviral transgene expression in mice over seven generations.

    PubMed

    Wang, Yong; Song, Yong-tao; Liu, Qin; Liu, Cang'e; Wang, Lu-lu; Liu, Yu; Zhou, Xiao-yang; Wu, Jun; Wei, Hong

    2010-10-01

    Lentiviral transgenesis is now recognized as an extremely efficient and cost-effective method to produce transgenic animals. Transgenes delivered by lentiviral vectors exhibited inheritable expression in many species including those which are refractory to genetic modification such as non-human primates. However, epigenetic modification was frequently observed in lentiviral integrants, and transgene expression found to be inversely correlated with methylation density. Recent data showed that about one-third lentiviral integrants exhibited hypermethylation and low expression, but did not demonstrate whether those integrants with high expression could remain constant expression and hypomethylated during long term germline transmission. In this study, using lentiviral eGFP transgenic mice as the experimental animals, lentiviral eGFP expression levels and its integrant numbers in genome were quantitatively analyzed by fluorescent quantitative polymerase-chain reaction (FQ-PCR), using the house-keeping gene ribosomal protein S18 (Rps18) and the single copy gene fatty acid binding protein of the intestine (Fabpi) as the internal controls respectively. The methylation densities of the integrants were quantitatively analyzed by bisulfite sequencing. We found that the lentiviral integrants with high expression exhibited a relative constant expression level per integrant over at least seven generations. Besides, the individuals containing these integrants exhibited eGFP expression levels which were positively and almost linearly correlated with the integrant numbers in their genomes, suggesting that no remarkable position effect on transgene expression of the integrants analyzed was observed. In addition, over seven generations the methylation density of these integrants did not increase, but rather decreased remarkably, indicating that these high expressing integrants were not subjected to de novo methylation during at least seven generations of germline transmission. Taken

  15. Ultrastructural morphometry of capillary basement membrane thickness in normal and transgenic diabetic mice.

    PubMed

    Carlson, Edward C; Audette, Janice L; Veitenheimer, Nicole J; Risan, Jessica A; Laturnus, Donna I; Epstein, Paul N

    2003-04-01

    Capillary basement membrane (CBM) thickening is an ultrastructural hallmark in diabetic patients and in animal models of diabetes. However, the wide variety of tissues sampled and diverse methods employed have made the interpretation of thickness data difficult. We showed previously that acellular glomerular BMs in OVE26 transgenic diabetic mice were thickened beyond normal age-related thickening, and in the current study we hypothesized that other microvascular BMs likewise would show increased widths relative to age-matched controls. Accordingly, a series of tissues, including skeletal and cardiac muscle, ocular retina and choriod, peripheral nerve, lung, pancreas, and renal glomerulus was collected from 300-350-day-old normal and transgenic mice. Transmission electron micrographs of cross sections through capillary walls were prepared, and CBM thickness (CBMT) was determined by the "orthogonal intercept" method. Morphometric analyses showed highly variable transgene-related BMT increases in the sampled tissues, with glomerular BM showing by far the greatest increase (+87%). Significant thickness increases were also seen in the retina, pulmonary alveolus, and thoracoabdominal diaphragm. BMT increases were not universal; however, most were modestly widened, and those that were thickest in controls generally showed the greatest increase. Although the pathogenesis of diabetes-related increases in CBM is poorly understood, data in the current study showed that in OVE26 transgenic mice increased BMT was a frequent concomitant of hyperglycemia. Accordingly, it seems likely that hyperglycemia-induced microvascular damage may be a contributing factor in diabetic BM disease, and that microvessel cellular and extracellular heterogeneity may limit the extent of CBM thickening in diverse tissues. PMID:12629676

  16. Overexpression of Heparanase Lowers the Amyloid Burden in Amyloid-β Precursor Protein Transgenic Mice*

    PubMed Central

    Jendresen, Charlotte B.; Cui, Hao; Zhang, Xiao; Vlodavsky, Israel; Nilsson, Lars N. G.; Li, Jin-Ping

    2015-01-01

    Heparan sulfate (HS) and HS proteoglycans (HSPGs) colocalize with amyloid-β (Aβ) deposits in Alzheimer disease brain and in Aβ precursor protein (AβPP) transgenic mouse models. Heparanase is an endoglycosidase that specifically degrades the unbranched glycosaminoglycan side chains of HSPGs. The aim of this study was to test the hypothesis that HS and HSPGs are active participators of Aβ pathogenesis in vivo. We therefore generated a double-transgenic mouse model overexpressing both human heparanase and human AβPP harboring the Swedish mutation (tgHpa*Swe). Overexpression of heparanase did not affect AβPP processing because the steady-state levels of Aβ1–40, Aβ1–42, and soluble AβPP β were the same in 2- to 3-month-old double-transgenic tgHpa*Swe and single-transgenic tgSwe mice. In contrast, the Congo red-positive amyloid burden was significantly lower in 15-month-old tgHpa*Swe brain than in tgSwe brain. Likewise, the Aβ burden, measured by Aβx-40 and Aβx-42 immunohistochemistry, was reduced significantly in tgHpa*Swe brain. The intensity of HS-stained plaques correlated with the Aβx-42 burden and was reduced in tgHpa*Swe mice. Moreover, the HS-like molecule heparin facilitated Aβ1–42-aggregation in an in vitro Thioflavin T assay. The findings suggest that HSPGs contribute to amyloid deposition in tgSwe mice by increasing Aβ fibril formation because heparanase-induced fragmentation of HS led to a reduced amyloid burden. Therefore, drugs interfering with Aβ-HSPG interactions might be a potential strategy for Alzheimer disease treatment. PMID:25548284

  17. Transgenic mice expressing an intracellular fluorescent fusion of angiotensin II demonstrate renal thrombotic microangiopathy and elevated blood pressure

    PubMed Central

    Redding, K. M.; Chen, B. L.; Singh, A.; Re, R. N.; Navar, L. G.; Seth, D. M.; Sigmund, C. D.; Tang, W. W.

    2010-01-01

    We have generated transgenic mice that express angiotensin II (ANG II) fused downstream of enhanced cyan fluorescent protein, expression of which is regulated by the mouse metallothionein promoter. The fusion protein, which lacks a secretory signal, is retained intracellularly. In the present study, RT-PCR, immunoblot analyses, whole-animal fluorescent imaging, and fluorescent microscopy of murine embryonic fibroblasts confirm expression of the fusion protein in vivo and in vitro. The transgene is expressed in all tissues tested (including brain, heart, kidney, liver, lung, and testes), and radioimmunoassay of plasma samples obtained from transgenic mice indicate no increase in circulating ANG II over wild-type levels, consistent with intracellular retention of the transgene product. Kidneys from transgenic and corresponding wild-type littermates were histologically evaluated, and abnormalities in transgenic mice consistent with thrombotic microangiopathy were observed; microthrombosis was frequently observed within the glomerular capillaries and small vessels. In addition, systolic and diastolic blood pressures, measured by telemetry (n = 8 for each group), were significantly higher in transgenic mice compared with wild-type littermates. Blood pressure of line A male transgenic mice was 125 ± 1.7 over 97 ± 1.6 compared with 109 ± 1.7 over 83 ± 1.4 mmHg in wild-type littermates (systolic over diastolic). In summary, overexpression of an intracellular fluorescent fusion protein of ANG II correlates with elevated blood pressure and kidney pathology. This transgenic model may be useful to further explore the intracellular renin-angiotensin system and its implication in abnormal kidney function and hypertension. PMID:20363893

  18. Ablation of the Locus Coeruleus Increases Oxidative Stress in Tg-2576 Transgenic but Not Wild-Type Mice

    PubMed Central

    Hurko, Orest; Boudonck, Kurt; Gonzales, Cathleen; Hughes, Zoe A.; Jacobsen, J. Steve; Reinhart, Peter H.; Crowther, Daniel

    2010-01-01

    Mice transgenic for production of excessive or mutant forms of beta-amyloid differ from patients with Alzheimer's disease in the degree of inflammation, oxidative damage, and alteration of intermediary metabolism, as well as the paucity or absence of neuronal atrophy and cognitive impairment. Previous observers have suggested that differences in inflammatory response reflect a discrepancy in the state of the locus coeruleus (LC), loss of which is an early change in Alzheimer's disease but which is preserved in the transgenic mice. In this paper, we extend these observations by examining the effects of the LC on markers of oxidative stress and intermediary metabolism. We compare four groups: wild-type or Tg2576 Aβ transgenic mice injected with DSP4 or vehicle. Of greatest interest were metabolites different between ablated and intact transgenics, but not between ablated and intact wild-type animals. The Tg2576_DSP4 mice were distinguished from the other three groups by oxidative stress and altered energy metabolism. These observations provide further support for the hypothesis that Tg2576 Aβ transgenic mice with this ablation may be a more congruent model of Alzheimer's disease than are transgenics with an intact LC. PMID:20981353

  19. Redox Proteomic Profiling of Specifically Carbonylated Proteins in the Serum of Triple Transgenic Alzheimer's Disease Mice.

    PubMed

    Shen, Liming; Chen, Youjiao; Yang, Aochu; Chen, Cheng; Liao, Liping; Li, Shuiming; Ying, Ming; Tian, Jing; Liu, Qiong; Ni, Jiazuan

    2016-01-01

    Oxidative stress is a key event in the onset and progression of neurodegenerative diseases, including Alzheimer's disease (AD). To investigate the role of oxidative stress in AD and to search for potential biomarkers in peripheral blood, serums were collected in this study from the 3-, 6-, and 12-month-old triple transgenic AD mice (3×Tg-AD mice) and the age- and sex-matched non-transgenic (non-Tg) littermates. The serum oxidized proteins were quantified by slot-blot analysis and enzyme-linked immunosorbent assay (ELISA) to investigate the total levels of serum protein carbonyl groups. Western blotting, in conjunction with two-dimensional gel electrophoresis (2D-Oxyblot), was employed to identify and quantify the specifically-carbonylated proteins in the serum of 3×Tg-AD mice. The results showed that the levels of serum protein carbonyls were increased in the three month old 3×Tg-AD mice compared with the non-Tg control mice, whereas no significant differences were observed in the six and 12 months old AD mice, suggesting that oxidative stress is an early event in AD progression. With the application of 2D-Oxyblot analysis, (immunoglobin) Ig gamma-2B chain C region (IGH-3), Ig lambda-2 chain C region (IGLC2), Ig kappa chain C region (IGKC), and Ig kappa chain V-V region HP R16.7 were identified as significantly oxidized proteins compared with the control. Among them IGH-3 and IGKC were validated via immunoprecipitation and Western blot analysis. Identification of oxidized proteins in the serums of 3×Tg-AD mice can not only reveal potential roles of those proteins in the pathogenesis of AD but also provide potential biomarkers of AD at the early stage. PMID:27077851

  20. Expression and immune recognition of SV40 Tag in transgenic mice that develop metastatic osteosarcomas.

    PubMed

    Marton, I; Johnson, S E; Damjanov, I; Currier, K S; Sundberg, J P; Knowles, B B

    2000-04-01

    Mature adult mice of the C57BL/6-TgN(Amy1TAg)501Knw transgenic mouse lineage, 501, containing a liver alpha-amylase promoted-SV40 Tag hybrid gene, routinely develop SV40 Tag-induced metastatic osteosarcomas. This form of alpha-amylase was known to be expressed in the liver, salivary glands, pancreas, and fat. Cells in the normal rib adjacent to the periosteum also express alpha-amylase suggesting that transgene expression is correctly targeted to generate osteosarcomas. 501 mice express SV40 Tag in the salivary glands but do not develop abnormalities in these organs by the time of their death from SV40-induced osteosarcomas. Mice of the C57BL/6 strain make a strong and effective anti-tumor immune response to SV40 Tag immunization. However, immunization of 501 mice with SV40 Tag early in life does not alter or prevent SV40 Tag-induced osteosarcomagenesis. 501 mice mount a significantly less effective cytotoxic T-lymphocyte response following SV40 Tag immunization while 501 osteosarcoma-derived cells are fully susceptible to SV40 Tag-specific T-cell lysis. This suggests that partial tolerance, not loss of antigen presentation by tumor cells, characterizes this mouse model of endogenous bone tumor development. To determine whether the immune recognition of endogenous SV40 Tag could influence tumorigenesis, the metastatic potential and time of death from tumor was investigated in CD4-null mutant 501 mice and beta-2 microglobulin-null mutant 501 mice. The size and number of metastases in these strains and longevity of these strains varied. We suggest that components of both the innate and adaptive immune response control tumor appearance and progression. PMID:10951695

  1. Transgene Expression Levels Determine the Immunogenicity of Transduced Hematopoietic Grafts in Partially Myeloablated Mice

    PubMed Central

    Eixarch, Herena; Gómez, Alba; Kádár, Elisabeth; George, Mónica; Martínez, Nuria; Espejo, Carmen; Pétriz, Jordi; Gimeno, Ramon; Barquinero, Jordi

    2009-01-01

    We investigated whether transgene expression levels influence the immunogenicity of transduced hematopoietic grafts upon transplantation into partially myeloablated mice. To this aim, bone marrow cells (BMCs) transduced with retroviral vectors driving green fluorescent protein (GFP) expression either at high (high-EGFP) or low levels (low-EGFP) were transplanted into congenic recipients conditioned with sublethal doses of total body irradiation (TBI) or busulfan. Virtually all recipients showed evidence of donor engraftment 4 weeks after transplantation. However, as opposed to recipients receiving low-EGFP transduced grafts, the risk of rejecting the EGFP+ cells by 30 days after transplantation was significantly higher in mice conditioned with busulfan and receiving high-EGFP transduced grafts. Anti-EGFP cellular immune responses were demonstrated in high-EGFP-treated mice conditioned with busulfan by interferon-γ (IFN-γ), enzyme-linked immunospot assay (ELISPOT), and cytotoxic T lymphocyte (CTL) assays, in contrast to that observed in mice transplanted with low-EGFP BMC. These results show for the first time that transgene expression levels can be critical for the immunogenicity of gene-modified hematopoietic grafts, especially in immunocompetent or in partially immunosuppressed recipients. These results have profound implications in vector choice and in the design of gene therapy (GT) protocols. PMID:19707185

  2. Effect of dihydrotestosterone on gastrointestinal tract of male Alzheimer's disease transgenic mice.

    PubMed

    Karri, Sritulasi; Acosta-Martinez, Veronica; Coimbatore, Gopalakrishnan

    2010-05-01

    The cause of Alzheimer's disease (AD) is still unknown. While research contributions identifying brain as locus of the disease is growing, evidence of severely impaired gastrointestinal (GI) functions with ageing too is accumulating, there is an equal dearth of information on GI tract in AD condition. The aim of this study was to assess the molecular, histological, morphological and microflora alterations of GI tract in male Alzheimer's transgenic mice. The present study also investigates the effect of dihydrotestosterone (DHT) treatment (1 mg/kg) on AD mice. Histoarchitecture data revealed a significant decrease in the villi number, muscular layer thickness, villi length, width, crypt length, enterocyte length and nuclei length. A shift in colon feces microbial community composition was observed by fatty acid methyl ester analysis. Amyloid precursor protein (APP) expression levels in intestine significantly increased in AD mice revealing its toxicity. DHT treatment attenuated the effect caused by AD on GI morphometrics, APP expression and colon micro flora population. These results for the first time reveal the quantitative and qualitative characteristics of GI tract in male Alzheimer's disease transgenic mice. PMID:20795362

  3. Biaryl Amides and Hydrazones as Therapeutics for Prion Disease in Transgenic Mice

    PubMed Central

    Lu, Duo; Giles, Kurt; Li, Zhe; Rao, Satish; Dolghih, Elena; Gever, Joel R.; Geva, Michal; Elepano, Manuel L.; Oehler, Abby; Bryant, Clifford; Renslo, Adam R.; Jacobson, Matthew P.; DeArmond, Stephen J.; Silber, B. Michael

    2013-01-01

    The only small-molecule compound demonstrated to substantially extend survival in prion-infected mice is a biaryl hydrazone termed “Compd B” (4-pyridinecarboxaldehyde,2-[4-(5-oxazolyl)phenyl]hydrazone). However, the hydrazone moiety of Compd B results in toxic metabolites, making it a poor candidate for further drug development. We developed a pharmacophore model based on diverse antiprion compounds identified by high-throughput screening; based on this model, we generated biaryl amide analogs of Compd B. Medicinal chemistry optimization led to multiple compounds with increased potency, increased brain concentrations, and greater metabolic stability, indicating that they could be promising candidates for antiprion therapy. Replacing the pyridyl ring of Compd B with a phenyl group containing an electron-donating substituent increased potency, while adding an aryl group to the oxazole moiety increased metabolic stability. To test the efficacy of Compd B, we applied bioluminescence imaging (BLI), which was previously shown to detect prion disease onset in live mice earlier than clinical signs. In our studies, Compd B showed good efficacy in two lines of transgenic mice infected with the mouse-adapted Rocky Mountain Laboratory (RML) strain of prions, but not in transgenic mice infected with human prions. The BLI system successfully predicted the efficacies in all cases long before extension in survival could be observed. Our studies suggest that this BLI system has good potential to be applied in future antiprion drug efficacy studies. PMID:23965382

  4. In Vivo Magnetic Resonance Microimaging of Individual Amyloid Plaques in Alzheimer’s Transgenic Mice

    PubMed Central

    Jack, Clifford R.; Wengenack, Thomas M.; Reyes, Denise A.; Garwood, Michael; Curran, Geoffrey L.; Borowski, Bret J.; Lin, Joseph; Preboske, Gregory M.; Holasek, Silvina S.; Adriany, Gregor; Poduslo, Joseph F.

    2009-01-01

    The ability to detect individual Alzheimer’s amyloid plaques in vivo by magnetic resonance microimaging (MRI) should improve diagnosis and also accelerate discovery of effective therapeutic agents for Alzheimer’s disease (AD). Here we perform in vivo and ex vivo MRI on double transgenic AD mice as well as wild type mice at varying ages and correlate these with Thioflavin-S and iron staining histology. Quantitative counts of individual plaques on MRI increase with age and correlate with histologically determined plaque burden. Plaques 20 μm in diameter can be detected in AD mice as young as 3 months of age with ex vivo MRI. Plaques 35 μm in diameter can be detected by 9 months of age with in vivo MRI. In vivo MRI of individual Alzheimer’s amyloid plaques provides a non-invasive estimate of plaque burden in transgenic AD mice which might be useful in assessing the efficacy of amyloid reduction therapies. PMID:16251453

  5. Transgenic Mice Harboring SV40 T-Antigen Genes Develop Characteristic Brain Tumors

    PubMed Central

    Brinster, Ralph L.; Chen, Howard Y.; Messing, Albee; van Dyke, Terry; Levine, Arnold J.; Palmiter, Richard D.

    2016-01-01

    Summary A high percentage of transgenic mice developing from eggs microinjected with plasmids containing the SV40 early region genes and a metallothionein fusion gene develop tumors within the choroid plexus. A line of mice has been established in which nearly every affected animal succumbs to this brain tumor. Thymic hypertrophy and kidney pathology are also observed in some mice. SV40 T-antigen mRNA and protein are readily detected in affected tissues; however, SV40 T-antigen gene expression is barely detectable in unaffected tissues or in susceptible tissues prior to overt pathology, suggesting that tumorigenesis depends upon activation of the SV40 genes. Comparison of DNA from tumor tissue (or cell lines derived from tumors) with DNA from unaffected tissues reveals structural rearrangements as well as changes in DNA methylation of the foreign DNA. The SV40 genes are frequently amplified in tumor tissue, which further indicates that their expression is intimately involved in tumorigenesis in transgenic mice. PMID:6327063

  6. [The transgenic mice specifically express Cre recombinase in central nerve system].

    PubMed

    Sheng, Ji-Po; Hou, Ning; Cheng, Xuan; Yang, Xiao; Deng, Ji-Xian

    2004-12-01

    For specific expressing Cre recombinase in central nerve system (CNS), a transgenic construct (pGFAP-Cre-hGH), containing the beta-globin insulators, 1.8 kb of glial fibrillary acidic protein gene (GFAP) 5' end regulation region, Cre gene and polyA of human growth hormone gene (hGH) was generated, in which the 5' end regulation region of GFAP was isolated from a 129sv mouse genomic DNA library with PCR-screening. 7.6 kb of pGFAP-Cre-hGH DNA fragment was introduced into 191 fertilized eggs by microinjection. 176 injected eggs were implanted into the oviducts of eight female mice respectively, from which 25 offspring were obtained. Seven mice carry the Cre genes by the identification of PCR and Southern blotting, and the integration efficiency is 28%. The GFAP-Cre transgenic mice were crossed with ROSA26 mice whose genomic DNA is integrated by LoxP sites and LacZ expression frame to check the activity and the tissue-specific expression of Cre recombinase and recombination with its mediation between two LoxP sites. The results of LacZ dying showed that the Cre recombinase was expressed only in CNS and successfully mediated the recombination between the LoxP sites in vivo. PMID:15633637

  7. Utilization of myoblasts from transgenic mice to evaluate the efficacy of myoblast transplantation.

    PubMed

    Kinoshita, I; Huard, J; Tremblay, J P

    1994-09-01

    A possible treatment for Duchenne muscular dystrophy is the injection of normal myoblasts into dystrophic muscles to induce the formation of new, healthy, and dystrophin-positive muscle fibers. To develop this therapy, it is important to identify the muscle fibers formed by the injected myoblasts in the host muscles. In this study, we used myoblasts from transgenic mice which have a gene expressing beta-galactosidase under the control of the promoter of quail fast skeletal muscle troponin I. This transgene is expressed in myotubes and muscle fibers, but not in myoblasts. Twenty-eight days after myoblast transplantation in nude and in mdx mice, muscle fibers containing of beta-galactosidase were identified by x-gal staining. In mdx mice, most of the beta-galactosidase-positive muscle fibers resulting from the myoblast transplantation were also dystrophin positive. This technique could make it possible to follow the success of myoblast transplantation even in mice that are not depleted of dystrophin. PMID:8065399

  8. Neuron-specific expression and physiological regulation of bovine vasopressin transgenes in mice.

    PubMed Central

    Ang, H L; Carter, D A; Murphy, D

    1993-01-01

    We have used transgenic mice to analyse the regulation of the bovine vasopressin (BVP) gene. We find that the restriction of BVP gene expression to anatomically and functionally distinct hypothalamic neuronal groups is achieved, in part, by selective repression. The expression of a 1.25 kb BVP proximal promoter, which on its own confers general expression of a reporter to most peripheral and brain tissues, was limited by sequences in the BVP structural gene to neural cells in the adrenal medulla and brain. Transgene expression in the hypothalamus was shown to be regulated by the physiological stimulus of dehydration in parallel with the endogenous gene. The expression of a larger 13.4 kb BVP transgene, containing 9 kb of 5' upstream sequence, the VP structural gene and 1.5 kb 3' of the transcription unit, was even more restricted and resembles that of the endogenous mouse gene. Hypothalamic expression of the 13.4 kb BVP transgene was regulated appropriately in response to an osmotic challenge. Images PMID:7685275

  9. The food additive vanillic acid controls transgene expression in mammalian cells and mice

    PubMed Central

    Gitzinger, Marc; Kemmer, Christian; Fluri, David A.; Daoud El-Baba, Marie; Weber, Wilfried; Fussenegger, Martin

    2012-01-01

    Trigger-inducible transcription-control devices that reversibly fine-tune transgene expression in response to molecular cues have significantly advanced the rational reprogramming of mammalian cells. When designed for use in future gene- and cell-based therapies the trigger molecules have to be carefully chosen in order to provide maximum specificity, minimal side-effects and optimal pharmacokinetics in a mammalian organism. Capitalizing on control components that enable Caulobacter crescentus to metabolize vanillic acid originating from lignin degradation that occurs in its oligotrophic freshwater habitat, we have designed synthetic devices that specifically adjust transgene expression in mammalian cells when exposed to vanillic acid. Even in mice transgene expression was robust, precise and tunable in response to vanillic acid. As a licensed food additive that is regularly consumed by humans via flavoured convenience food and specific fresh vegetable and fruits, vanillic acid can be considered as a safe trigger molecule that could be used for diet-controlled transgene expression in future gene- and cell-based therapies. PMID:22187155

  10. Autoantigen-Specific B Cell Activation in FAS-Deficient Rheumatoid Factor Immunoglobulin Transgenic Mice

    PubMed Central

    Wang, Haowei; Shlomchik, Mark J.

    1999-01-01

    In systemic autoimmune disease, self-tolerance fails, leading to autoantibody production. A central issue in immunology is to understand the origins of activated self-reactive B cells. We have used immunoglobulin (Ig) transgenic mice to investigate the regulation of autoreactive B cells with specificity for self-IgG2a (the rheumatoid factor [RF] specificity) to understand how normal mice regulate RF autoantibodies and how this fails in autoimmune mice. We previously showed that normal mice do not tolerize the AM14 RF clone, nor do they appear to activate it. Here we show that in Fas-deficient autoimmune mice, the picture is quite different. RF B cells are activated to divide and secrete, but only when the autoantigen is present. Thus, B cells that are ignored rather than anergized in normal mice can be stimulated to produce autoantibody in Fas-deficient mice. This demonstrates a novel developmental step at which intact Fas–Fas ligand signaling is required to regulate B cells in order to prevent autoimmunity. These data also establish the relevance of ignorant self-specific B cells to autoantibody production in disease and prove that in the case of the RF specificity, the nominal autoantigen IgG2a is the driving autoantigen in vivo. PMID:10477549

  11. Transgenic sickle cell disease mice have high mortality and dysregulated immune responses after vaccination

    PubMed Central

    Szczepanek, Steven M.; Secor, Eric R.; Bracken, Sonali J.; Guernsey, Linda; Rafti, Ektor; Matson, Adam; Thrall, Roger S.; Andemariam, Biree

    2014-01-01

    Background Children with sickle cell disease (SCD) are susceptible to recurrent infections, which are often life threatening and necessitate frequent vaccinations. Given the altered baseline immunity and proinflammatory state associated with SCD, we sought to determine the relative safety and efficacy of vaccination in transgenic SCD mice. Methods Eight week-old SCD mice were vaccinated with ovalbumin (OVA) and aluminum hydroxide weekly for three weeks by the intraperitoneal (IP) or intramuscular (IM) route. One week after the third vaccination, serum cytokines/chemokines, immunoglobulins, and bronchoalveolar lavage (BAL) fluid cytokines were measured. Results Only SCD mice were prone to mortality associated with vaccination as 40% of the animals died after the IP vaccinations and 50% died after the IM vaccinations. Serum IgG2b and IgM were significantly lower in SCD than C57Bl/6 mice after vaccination, but OVA-specific IgE was significantly higher. Serum interleukin 1 alpha (IL-1α), IL-2, IL-5, macrophage inflammatory protein 1 alpha (MIP-1α), and granulocyte macrophage-colony stimulating factor (GM-CSF) were significantly lower in SCD mice than C57Bl/6 mice after vaccination, whereas BAL fluid IL-1β and IL-6 were elevated. Conclusions Mice with SCD appear to have a dysregulated immune response to vaccination. Thus, the relative safety and immunogenicity of vaccination should be studied in greater detail in the context of SCD. PMID:23728384

  12. FIAT represses ATF4-mediated transcription to regulate bone mass in transgenic mice.

    PubMed

    Yu, Vionnie W C; Ambartsoumian, Gourgen; Verlinden, Lieve; Moir, Janet M; Prud'homme, Josée; Gauthier, Claude; Roughley, Peter J; St-Arnaud, René

    2005-05-23

    We report the characterization of factor inhibiting activating transcription factor 4 (ATF4)-mediated transcription (FIAT), a leucine zipper nuclear protein. FIAT interacted with ATF4 to inhibit binding of ATF4 to DNA and block ATF4-mediated transcription of the osteocalcin gene in vitro. Transgenic mice overexpressing FIAT in osteoblasts also had reduced osteocalcin gene expression and decreased bone mineral density, bone volume, mineralized volume, trabecular thickness, trabecular number, and decreased rigidity of long bones. Mineral homeostasis, osteoclast number and activity, and osteoblast proliferation and apoptosis were unchanged in transgenics. Expression of osteoblastic differentiation markers was largely unaffected and type I collagen synthesis was unchanged. Mineral apposition rate was reduced in transgenic mice, suggesting that the lowered bone mass was due to a decline in osteoblast activity. This cell-autonomous decrease in osteoblast activity was confirmed by measuring reduced alkaline phosphatase activity and mineralization in primary osteoblast cultures. These results show that FIAT regulates bone mass accrual and establish FIAT as a novel transcriptional regulator of osteoblastic function. PMID:15911876

  13. Cosmetics-triggered percutaneous remote control of transgene expression in mice.

    PubMed

    Wang, Hui; Ye, Haifeng; Xie, Mingqi; Daoud El-Baba, Marie; Fussenegger, Martin

    2015-08-18

    Synthetic biology has significantly advanced the rational design of trigger-inducible gene switches that program cellular behavior in a reliable and predictable manner. Capitalizing on genetic componentry, including the repressor PmeR and its cognate operator OPmeR, that has evolved in Pseudomonas syringae pathovar tomato DC3000 to sense and resist plant-defence metabolites of the paraben class, we have designed a set of inducible and repressible mammalian transcription-control devices that could dose-dependently fine-tune transgene expression in mammalian cells and mice in response to paraben derivatives. With an over 60-years track record as licensed preservatives in the cosmetics industry, paraben derivatives have become a commonplace ingredient of most skin-care products including shower gels, cleansing toners and hand creams. As parabens can rapidly reach the bloodstream of mice following topical application, we used this feature to percutaneously program transgene expression of subcutaneous designer cell implants using off-the-shelf commercial paraben-containing skin-care cosmetics. The combination of non-invasive, transdermal and orthogonal trigger-inducible remote control of transgene expression may provide novel opportunities for dynamic interventions in future gene and cell-based therapies. PMID:25943548

  14. A pipeline for the generation of shRNA transgenic mice

    PubMed Central

    Dow, Lukas E; Premsrirut, Prem K; Zuber, Johannes; Fellmann, Christof; McJunkin, Katherine; Miething, Cornelius; Park, Youngkyu; Dickins, Ross A; Hannon, Gregory J; Lowe, Scott W

    2013-01-01

    RNA interference (RNAi) is an extremely effective tool for studying gene function in almost all metazoan and eukaryotic model systems. RNAi in mice, through the expression of short hairpin RNA s (shRNA s), offers something not easily achieved with traditional genetic approaches—inducible and reversible gene silencing. However, technical variability associated with the production of shRNA transgenic strains has so far limited their widespread use. Here we describe a pipeline for the generation of miR30-based shRNA transgenic mice that enables efficient and consistent targeting of doxycycline-regulated, fluorescence-linked shRNA s to the Col1a1 locus. Notably, the protocol details crucial steps in the design and testing of miR30-based shRNA s to maximize the potential for developing effective transgenic strains. In all, this 14-week procedure provides a fast and cost-effective way for any laboratory to investigate gene function in vivo in the mouse. PMID:22301776

  15. Endoplasmic reticulum stress response in P104L mutant caveolin-3 transgenic mice.

    PubMed

    Kuga, Atsushi; Ohsawa, Yutaka; Okada, Tadashi; Kanda, Fumio; Kanagawa, Motoi; Toda, Tatsushi; Sunada, Yoshihide

    2011-08-01

    Mutations in the caveolin-3 gene cause autosomal dominant limb-girdle muscular dystrophy 1C (LGMD1C). However, the precise molecular pathogenesis of caveolin-3-related muscular dystrophy remains uncertain. Here, we demonstrate the effect of gene dosage on the severity of the myopathic phenotype in P104L mutant caveolin-3 (mCav3(P104L)) transgenic mice, a model of LGMD1C. We analyzed the endoplasmic reticulum (ER) stress response in the transgenic mice and found upregulated transcription of the molecular chaperone, glucose-regulated protein (GRP78). Moreover, signaling downstream of GRP78 in the myofibers was activated toward apoptosis. However, terminal transferase dUTP nick end labeling assays detected a few apoptotic nuclei in transgenic mouse skeletal muscle, probably due to the transcriptional activation of Dad1, an anti-apoptotic factor in the ER. These findings suggest that the ER stress response caused by mCav3(P104L) plays a role in the pathogenesis of LGMD1C as a toxic gain of function effect. PMID:21610159

  16. Long Tract of Untranslated CAG Repeats Is Deleterious in Transgenic Mice

    PubMed Central

    Lin, Min-Jon; Li, Chui-Yen; Wang, Li-Chun; Chen, Luen-Kui; Pan, Huichin

    2011-01-01

    The most frequent trinucleotide repeat found in human disorders is the CAG sequence. Expansion of CAG repeats is mostly found in coding regions and is thought to cause diseases through a protein mechanism. Recently, expanded CAG repeats were shown to induce toxicity at the RNA level in Drosophila and C. elegans. These findings raise the possibility that CAG repeats may trigger RNA-mediated pathogenesis in mammals. Here, we demonstrate that transgenic mice expressing EGFP transcripts with long CAG repeats in the 3′ untranslated region develop pathogenic features. Expression of the transgene was directed to the muscle in order to compare the resulting phenotype to that caused by the CUG expansion, as occurs in myotonic dystrophy. Transgenic mice expressing 200, but not those expressing 0 or 23 CAG repeats, showed alterations in muscle morphology, histochemistry and electrophysiology, as well as abnormal behavioral phenotypes. Expression of the expanded CAG repeats in testes resulted in reduced fertility due to defective sperm motility. The production of EGFP protein was significantly reduced by the 200 CAG repeats, and no polyglutamine-containing product was detected, which argues against a protein mechanism. Moreover, nuclear RNA foci were detected for the long CAG repeats. These data support the notion that expanded CAG repeat RNA can cause deleterious effects in mammals. They also suggest the possible involvement of an RNA mechanism in human diseases with long CAG repeats. PMID:21283659

  17. Green Tea Polyphenols Control Dysregulated Glutamate Dehydrogenase in Transgenic Mice by Hijacking the ADP Activation Site

    SciTech Connect

    Li, Changhong; Li, Ming; Chen, Pan; Narayan, Srinivas; Matschinsky, Franz M.; Bennett, Michael J.; Stanley, Charles A.; Smith, Thomas J.

    2012-05-09

    Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of L-glutamate and, in animals, is extensively regulated by a number of metabolites. Gain of function mutations in GDH that abrogate GTP inhibition cause the hyperinsulinism/hyperammonemia syndrome (HHS), resulting in increased pancreatic {beta}-cell responsiveness to leucine and susceptibility to hypoglycemia following high protein meals. We have previously shown that two of the polyphenols from green tea (epigallocatechin gallate (EGCG) and epicatechin gallate (ECG)) inhibit GDH in vitro and that EGCG blocks GDH-mediated insulin secretion in wild type rat islets. Using structural and site-directed mutagenesis studies, we demonstrate that ECG binds to the same site as the allosteric regulator, ADP. Perifusion assays using pancreatic islets from transgenic mice expressing a human HHS form of GDH demonstrate that the hyperresponse to glutamine caused by dysregulated GDH is blocked by the addition of EGCG. As observed in HHS patients, these transgenic mice are hypersensitive to amino acid feeding, and this is abrogated by oral administration of EGCG prior to challenge. Finally, the low basal blood glucose level in the HHS mouse model is improved upon chronic administration of EGCG. These results suggest that this common natural product or some derivative thereof may prove useful in controlling this genetic disorder. Of broader clinical implication is that other groups have shown that restriction of glutamine catabolism via these GDH inhibitors can be useful in treating various tumors. This HHS transgenic mouse model offers a highly useful means to test these agents in vivo.

  18. Utrophin Up-Regulation by an Artificial Transcription Factor in Transgenic Mice

    PubMed Central

    Mattei, Elisabetta; Corbi, Nicoletta; Di Certo, Maria Grazia; Strimpakos, Georgios; Severini, Cinzia; Onori, Annalisa; Desantis, Agata; Libri, Valentina; Buontempo, Serena; Floridi, Aristide; Fanciulli, Maurizio; Baban, Dilair; Davies, Kay E.; Passananti, Claudio

    2007-01-01

    Duchenne Muscular Dystrophy (DMD) is a severe muscle degenerative disease, due to absence of dystrophin. There is currently no effective treatment for DMD. Our aim is to up-regulate the expression level of the dystrophin related gene utrophin in DMD, complementing in this way the lack of dystrophin functions. To this end we designed and engineered several synthetic zinc finger based transcription factors. In particular, we have previously shown that the artificial three zinc finger protein named Jazz, fused with the appropriate effector domain, is able to drive the transcription of a test gene from the utrophin promoter “A”. Here we report on the characterization of Vp16-Jazz-transgenic mice that specifically over-express the utrophin gene at the muscular level. A Chromatin Immunoprecipitation assay (ChIP) demonstrated the effective access/binding of the Jazz protein to active chromatin in mouse muscle and Vp16-Jazz was shown to be able to up-regulate endogenous utrophin gene expression by immunohistochemistry, western blot analyses and real-time PCR. To our knowledge, this is the first example of a transgenic mouse expressing an artificial gene coding for a zinc finger based transcription factor. The achievement of Vp16-Jazz transgenic mice validates the strategy of transcriptional targeting of endogenous genes and could represent an exclusive animal model for use in drug discovery and therapeutics. PMID:17712422

  19. Cosmetics-triggered percutaneous remote control of transgene expression in mice

    PubMed Central

    Wang, Hui; Ye, Haifeng; Xie, Mingqi; Daoud El-Baba, Marie; Fussenegger, Martin

    2015-01-01

    Synthetic biology has significantly advanced the rational design of trigger-inducible gene switches that program cellular behavior in a reliable and predictable manner. Capitalizing on genetic componentry, including the repressor PmeR and its cognate operator OPmeR, that has evolved in Pseudomonas syringae pathovar tomato DC3000 to sense and resist plant-defence metabolites of the paraben class, we have designed a set of inducible and repressible mammalian transcription-control devices that could dose-dependently fine-tune transgene expression in mammalian cells and mice in response to paraben derivatives. With an over 60-years track record as licensed preservatives in the cosmetics industry, paraben derivatives have become a commonplace ingredient of most skin-care products including shower gels, cleansing toners and hand creams. As parabens can rapidly reach the bloodstream of mice following topical application, we used this feature to percutaneously program transgene expression of subcutaneous designer cell implants using off-the-shelf commercial paraben-containing skin-care cosmetics. The combination of non-invasive, transdermal and orthogonal trigger-inducible remote control of transgene expression may provide novel opportunities for dynamic interventions in future gene and cell-based therapies. PMID:25943548

  20. Transgenic mice with a constitutively active aryl hydrocarbon receptor display a gender-specific bone phenotype.

    PubMed

    Wejheden, Carolina; Brunnberg, Sara; Larsson, Sune; Lind, P Monica; Lind, Pia M; Andersson, Göran; Hanberg, Annika

    2010-03-01

    Bone tissue homeostasis is governed by hormones, growth factors, and cytokines and can be distorted by environmental pollutants, such as ligands to the aryl hydrocarbon receptor (AhR). A transgenic mouse expressing a constitutively active aryl hydrocarbon receptor (CA-AhR), mimicking continuous low-dose exposure to AhR ligands, was used to explore potential long-term effects of these ligands on bone. The density, content, and dimensions of cortical and trabecular bone, as well as physical properties, were significantly altered in female transgenic mice, while almost no alterations were detected in males. Osteoclast volume density and serum level of C-telopeptide of type I collagen (CTX), reflecting osteoclast activity, were both increased by approximately 60% in female CA-AhR mice, while serum tartrate-resistant acid phosphatase (TRAP) 5b, reflecting osteoclast numbers, was unchanged. Subsequently, the resorption index (CTX/TRAP 5b) was increased by 90%, indicating increased osteoclast activity in female CA-AhR. Moreover, the protein level of the osteoclast collagenase cathepsin K was increased by 40% in bone extracts of female CA-AhR mice. The messenger RNA expression of several osteoclast- and osteoblast-associated genes was altered in female transgenic mice but not in males. Notably, early markers for osteoclast and osteoblast differentiation were normal, while the expression of functional markers of osteoclasts and osteoblasts were reduced. In conclusion, a low continuous activation of the AhR leads to a skeletal phenotype with increased bone resorption associated with more ductile bones in females but not in males. The results indicate the presence of an interaction between the AhR and a female-specific mechanism implicated in inhibition of osteoclast development and function. Female bone tissue appears more susceptible to dioxins and other AhR ligands than male bone tissue. PMID:19934163

  1. Transgenic mice support replication of hepatitis delta virus RNA in multiple tissues, particularly in skeletal muscle.

    PubMed Central

    Polo, J M; Jeng, K S; Lim, B; Govindarajan, S; Hofman, F; Sangiorgi, F; Lai, M M

    1995-01-01

    Hepatitis delta virus (HDV) is hepatotropic and frequently causes fulminant hepatitis in both human and nonhuman primate hosts. To understand the molecular basis of HDV tissue tropism and the mechanism of pathogenesis, transgenic mice in which replication-competent HDV dimeric RNA is expressed under the control of either liver-specific or universal transcriptional promoters were developed. The expressed RNA replicated efficiently in the liver and several tissues of nonhepatic origin. Surprisingly, maximal replication of HDV RNA occurred in skeletal muscle and was almost 100-fold greater than in the liver. These findings suggest that the hepatotropism of HDV is most likely a receptor-mediated restriction and that muscle-specific factors may facilitate HDV RNA replication. No evidence of cytopathology was apparent in most of the tissues examined, including the liver, supporting the contention that hepatocellular disease is not mediated by direct cytopathological effects associated with HDV RNA replication and gene expression. However, mild muscle atrophy in some of the transgenic mice was noted. Delta antigen was detected in the nuclei of myocytes. Only the small form, not the large form, of delta antigen was detected, suggesting that the RNA editing event which causes the conversion of delta antigen did not occur in transgenic mice. Furthermore, the 0.8-kb antigenomic RNA species, which is postulated to be the mRNA for delta antigen, was not detected in mice. The preferential replication of HDV RNA in skeletal muscle suggests that HDV RNA replication can be facilitated by certain muscle-specific factors. PMID:7609056

  2. Diet-induced hyperlipoproteinemia and atherosclerosis in apolipoprotein E3-Leiden transgenic mice.

    PubMed Central

    van Vlijmen, B J; van den Maagdenberg, A M; Gijbels, M J; van der Boom, H; HogenEsch, H; Frants, R R; Hofker, M H; Havekes, L M

    1994-01-01

    Apolipoprotein E3-Leiden (APOE*3-Leiden) transgenic mice have been used to study the effect of different cholesterol-containing diets on the remnant lipoprotein levels and composition and on the possible concurrent development of atherosclerotic plaques. On high fat/cholesterol (HFC) diet, the high expressing lines 2 and 181 developed severe hypercholesterolemia (up to 40 and 60 mmol/liter, respectively), whereas triglyceride levels remained almost normal when compared with regular mouse diet. The addition of cholate increased the hypercholesterolemic effect of this diet. In lines 2 and 181, serum levels of apo E3-Leiden also increased dramatically upon cholesterol feeding (up to 107 and 300 mg/dl, respectively). In these high expressing APOE*3-Leiden transgenic mice, the increase in both serum cholesterol and apo E3-Leiden occurred mainly in the VLDL/LDL-sized fractions, whereas a considerable increase in large, apo E-rich HDL particles also occurred. In contrast to the high expressing lines, the low expressing line 195 reacted only mildly upon HFC diet. On HFC diets, the high expresser APOE*3-Leiden mice developed atherosclerotic lesions in the aortic arch, the descending aorta, and the carotid arteries, varying from fatty streaks containing foam cells to severe atherosclerotic plaques containing cholesterol crystals, fibrosis, and necrotic calcified tissue. Quantitative evaluation revealed that the atherogenesis is positively correlated with the serum level of cholesterol-rich VLDL/LDL particles. In conclusion, with APOE*3-Leiden transgenic mice, factors can be studied that influence the metabolism of remnant VLDL and the development of atherosclerosis. Images PMID:8163645

  3. Isoflurane Exposure during Mid-Adulthood Attenuates Age-Related Spatial Memory Impairment in APP/PS1 Transgenic Mice

    PubMed Central

    Xu, Huan; Wang, Beilei; Chen, Xuemei; Chen, Jie; Wang, Xiangrui

    2012-01-01

    Many in vitro findings suggest that isoflurane exposure might accelerate the process of Alzheimer Disease (AD); however, no behavioral evidence exists to support this theory. In the present study, we hypothesized that exposure of APP/PS1 transgenic mice to isoflurane during mid-adulthood, which is the pre-symptomatic phase of amyloid beta (Abeta) deposition, would alter the progression of AD. Seven-month-old Tg(APPswe,PSEN1dE9)85Dbo/J transgenic mice and their wild-type littermates were exposed to 1.1% isoflurane for 2 hours per day for 5 days. Learning and memory ability was tested 48 hours and 5 months following isoflurane exposure using the Morris Water Maze and Y maze, respectively. Abeta deposition and oligomers in the hippocampus were measured by immunohistochemistry or Elisa 5 months following isoflurane exposure. We found that the performance of both the transgenic and wild-type mice in the Morris Water Maze significantly improved 48 hours following isoflurane exposure. The transgenic mice made significantly fewer discrimination errors in the Y maze following isoflurane exposure, and no differences were found between wild-type littermates 5 months following isoflurane exposure. For the transgenic mice, the Abeta plaque and oligomers in the hippocampus was significantly decreased in the 5 months following isoflurane exposure. In summary, repeated isoflurane exposure during the pre-symptomatic phase not only improved spatial memory in both the APP/PS1 transgenic and wild-type mice shortly after the exposure but also prevented age-related decline in learning and memory and attenuated the Abeta plaque and oligomers in the hippocampus of transgenic mice. PMID:23185565

  4. Induction of robust cellular immunity against HPV6 and HPV11 in mice by DNA vaccine encoding for E6/E7 antigen

    PubMed Central

    Shin, Thomas; Pankhong, Panyupa; Yan, Jian; Khan, Amir S.; Sardesai, Niranjan Y.; Weiner, David B.

    2012-01-01

    Due to the strong relationship between the Human Papillomavirus (HPV) “high-risk” subtypes and cervical cancers, most HPV-related studies have been focusing on the “high-risk” HPV subtypes 16 and 18. However, it has been suggested that the “low-risk” subtypes of HPV, HPV6 and HPV11, are the major cause of recurrent respiratory papillomatosis and genital warts. In addition, HPV 6 and 11 are also associated with otolaryngologic malignancies, carcinoma of the lung, tonsil, larynx and low-grade cervical lesions. Therefore, development of HPV therapeutic vaccines targeting on subtypes 6 and 11 E6 or E7 are in great need. In this report, we describe two novel engineered DNA vaccines that encode HPV 6 and 11 consensus E6/E7 fusion proteins (p6E6E7 and p11E6E7) by utilizing a multi-phase strategy. Briefly, after generating consensus sequences, several modifications were performed to increase the expression of both constructs, including codon/RNA optimization, addition of a Kozak sequence and a highly efficient leader sequence. An endoproteolytic cleavage site was also introduced between E6 and E7 protein for proper protein folding and for better CTL processing. The expressions of both constructs were confirmed by western blot analysis and immunofluorescence assay. Vaccination with these DNA vaccines could elicit robust cellular immune responses. The epitope mapping assay was performed to further characterize the cellular immune responses induced by p6E6E7 and p11E6E7. The HPV 6 and 11 E6 or E7-specific immunodominant and subdominant epitopes were verified, respectively. The intracellular cytokine staining revealed that the magnitude of IFN-γ and TNF-α secretion in antigen-specific CD8+ cells was significantly enhanced, indicating that the immune responses elicited by p6E6E7 and p11E6E7 was heavily skewed toward driving CD8+ T cells. Such DNA immunogens are interesting candidates for further studies on HPV 6 and 11-associated diseases. PMID:22336879

  5. Altered temporal patterns of anxiety in aged and amyloid precursor protein (APP) transgenic mice

    PubMed Central

    Bedrosian, Tracy A.; Herring, Kamillya L.; Weil, Zachary M.; Nelson, Randy J.

    2011-01-01

    Both normal aging and dementia are associated with dysregulation of the biological clock, which contributes to disrupted circadian organization of physiology and behavior. Diminished circadian organization in conjunction with the loss of cholinergic input to the cortex likely contributes to impaired cognition and behavior. One especially notable and relatively common circadian disturbance among the aged is “sundowning syndrome,” which is characterized by exacerbated anxiety, agitation, locomotor activity, and delirium during the hours before bedtime. Sundowning has been reported in both dementia patients and cognitively intact elderly individuals living in institutions; however, little is known about temporal patterns in anxiety and agitation, and the neurobiological basis of these rhythms remains unspecified. In the present study, we explored the diurnal pattern of anxiety-like behavior in aged and amyloid precursor protein (APP) transgenic mice. We then attempted to treat the observed behavioral disturbances in the aged mice using chronic nightly melatonin treatment. Finally, we tested the hypothesis that time-of-day differences in acetylcholinesterase and choline acetyltransferase expression and general neuronal activation (i.e., c-Fos expression) coincide with the behavioral symptoms. Our results show a temporal pattern of anxiety-like behavior that emerges in elderly mice. This behavioral pattern coincides with elevated locomotor activity relative to adult mice near the end of the dark phase, and with time-dependent changes in basal forebrain acetylcholinesterase expression. Transgenic APP mice show a similar behavioral phenomenon that is not observed among age-matched wild-type mice. These results may have useful applications to the study and treatment of age- and dementia-related circadian behavioral disturbances, namely, sundowning syndrome. PMID:21709248

  6. Endogenous galectin-3 expression levels modulate immune responses in galectin-3 transgenic mice.

    PubMed

    Chaudhari, Aparna D; Gude, Rajiv P; Kalraiya, Rajiv D; Chiplunkar, Shubhada V

    2015-12-01

    Galectin-3 (Gal-3), a β-galactoside-binding mammalian lectin, is involved in cancer progression and metastasis. However, there is an unmet need to identify the underlying mechanisms of cancer metastasis mediated by endogenous host galectin-3. Galectin-3 is also known to be an important regulator of immune responses. The present study was aimed at analysing how expression of endogenous galectin-3 regulates host immunity and lung metastasis in B16F10 murine melanoma model. Transgenic Gal-3(+/-) (hemizygous) and Gal-3(-/-) (null) mice exhibited decreased levels of Natural Killer (NK) cells and lower NK mediated cytotoxicity against YAC-1 tumor targets, compared to Gal-3(+/+) (wild-type) mice. On stimulation, Gal-3(+/-) and Gal-3(-/-) mice splenocytes showed increased T cell proliferation than Gal-3(+/+) mice. Intracellular calcium flux was found to be lower in activated T cells of Gal-3(-/-) mice as compared to T cells from Gal-3(+/+) and Gal-3(+/-) mice. In Gal-3(-/-) mice, serum Th1, Th2 and Th17 cytokine levels were found to be lowest, exhibiting dysregulation of pro-inflammatory and anti-inflammatory cytokines balance. Marked decrease in serum IFN-γ levels and splenic IFN-γR1 (IFN-γ Receptor 1) expressing T and NK cell percentages were observed in Gal-3(-/-) mice. On recombinant IFN-γ treatment of splenocytes in vitro, Suppressor of Cytokine Signaling (SOCS) 1 and SOCS3 protein expression was higher in Gal-3(-/-) mice compared to that in Gal-3(+/+) and Gal-3(+/-) mice; suggesting possible attenuation of Signal Transducer and Activator of Transcription (STAT) 1 mediated IFN-γ signaling in Gal-3(-/-) mice. The ability of B16F10 melanoma cells to form metastatic colonies in the lungs of Gal-3(+/+) and Gal-3(-/-) mice remained comparable, whereas it was found to be reduced in Gal-3(+/-) mice. Our data indicates that complete absence of endogenous host galectin-3 facilitates lung metastasis of B16F10 cells in mice, which may be contributed by dysregulated immune

  7. Stromelysin-1 (MMP-3) expression driven by a macrophage-specific promoter results in reduced viability in transgenic mice.

    PubMed

    Fabunmi, R P; Moore, K J; Libby, P; Freeman, M W

    2000-02-01

    Macrophage expression of matrix degrading metalloproteinases (MMPs) in human atheroma has been found to occur in rupture-prone areas of plaques. To investigate the effect of metalloproteinase activity on plaque stability, we attempted to generate mice that expressed a stromelysin-1 (MMP-3) transgene specifically in macrophages. Promoter sequences taken from a macrophage-tropic lentivirus (visna) were used to drive transgene expression. The transgene construct was expressed in macrophages in vitro and its autoactivation was established by casein zymography. Transgenic mice generated with this construct died at or before birth. No gross anatomical changes were observed in these mice. Embryos arising from a second round of oocyte injections with the transgene were examined at day 16 of gestation. Of the products of conception, approximately 40% resulted in vacant conceptuses. Only one animal of 38 examined carried the transgene and its expression of MMP-3 mRNA at E16 was faintly detected by RT-PCR. When a non-toxic reporter gene, luciferase, was substituted for the MMP-3 cDNA, healthy transgenic mice were produced that expressed the reporter gene in a wide variety of tissue macrophages, including those located in the brain, testis, lung, and thymus. These studies suggest that constitutive expression of MMP-3 in diverse populations of tissue macrophages leads to prenatal or neonatal death in the mouse. It appears likely that more sophisticated transcriptional control of MMP-3 expression will be required in order to generate stromelysin-1 transgenic mice that could be useful models for studying overexpression of this metalloproteinase's activity in the lesional macrophages of atherosclerotic plaques. PMID:10657574

  8. Transmissibility of H-Type Bovine Spongiform Encephalopathy to Hamster PrP Transgenic Mice

    PubMed Central

    Okada, Hiroyuki; Masujin, Kentaro; Miyazawa, Kohtaro; Yokoyama, Takashi

    2015-01-01

    Two distinct forms of atypical bovine spongiform encephalopathies (H-BSE and L-BSE) can be distinguished from classical (C-) BSE found in cattle based on biochemical signatures of disease-associated prion protein (PrPSc). H-BSE is transmissible to wild-type mice—with infected mice showing a long survival period that is close to their normal lifespan—but not to hamsters. Therefore, rodent-adapted H-BSE with a short survival period would be useful for analyzing H-BSE characteristics. In this study, we investigated the transmissibility of H-BSE to hamster prion protein transgenic (TgHaNSE) mice with long survival periods. Although none of the TgHaNSE mice manifested the disease during their lifespan, PrPSc accumulation was observed in some areas of the brain after the first passage. With subsequent passages, TgHaNSE mice developed the disease with a mean survival period of 220 days. The molecular characteristics of proteinase K-resistant PrPSc (PrPres) in the brain were identical to those observed in first-passage mice. The distribution of immunolabeled PrPSc in the brains of TgHaNSE mice differed between those infected with H-BSE as compared to C-BSE or L-BSE, and the molecular properties of PrPres in TgHaNSE mice infected with H-BSE differed from those of the original isolate. The strain-specific electromobility, glycoform profiles, and proteolytic cleavage sites of H-BSE in TgHaNSE mice were indistinguishable from those of C-BSE, in which the diglycosylated form was predominant. These findings indicate that strain-specific pathogenic characteristics and molecular features of PrPres in the brain are altered during cross-species transmission. Typical H-BSE features were restored after back passage from TgHaNSE to bovinized transgenic mice, indicating that the H-BSE strain was propagated in TgHaNSE mice. This could result from the overexpression of the hamster prion protein. PMID:26466381

  9. Effects of chronic stress on the onset and progression of Huntington's disease in transgenic mice.

    PubMed

    Mo, Christina; Renoir, Thibault; Hannan, Anthony J

    2014-11-01

    Huntington's disease (HD) is a neurodegenerative disease caused by a tandem repeat mutation encoding an expanded polyglutamine tract. Our previous work showed that memory deficits in HD transgenic mice could be accelerated by increased levels of stress hormone, while memory in WT mice remained unaffected. HD patients experience higher levels of stress compared to the general population and symptoms of HD also include motor, cognitive, psychiatric, sexual and olfactory abnormalities, and an associated decline in activities of daily living. Therefore we investigated the impact of a robust stressor (i.e. restraint) on the onset and progression of a range of behavioral phenotypes in R6/1 transgenic HD mice. Restraint was administered for 1h daily from 6weeks of age and continued until R6/1 mice were clearly motor symptomatic at 14weeks of age. Serum corticosterone levels in both R6/1 and WT littermates were elevated immediately after the last restraint session and weight gain was suppressed in restrained animals throughout the treatment period. Motor coordination and locomotor activity were enhanced by chronic restraint in males, regardless of genotype. However, there was no effect of restraint on motor performances in female animals. At 8weeks of age, olfactory sensitivity was impaired by restraint in R6/1 HD female mice, but not in WT mice. In male R6/1 mice, the olfactory deficit was exacerbated by restraint and olfaction was also impaired in male WT mice. The development of deficits in saccharin preference, Y-maze memory, nest-building and sexually-motivated vocalizations was unaffected by chronic restraint in R6/1 and had little impact on such behavioral performances in WT animals. We provide evidence that chronic stress can negatively modulate specific endophenotypes in HD mice, while the same functions were affected to a lesser extent in WT mice. This vulnerability in HD animals seems to be sex-specific depending on the stress paradigm used. It is hoped that our

  10. GFAP expression and social deficits in transgenic mice overexpressing human sAPPα.

    PubMed

    Bailey, Antoinette R; Hou, Huayan; Song, Min; Obregon, Demian F; Portis, Samantha; Barger, Steven; Shytle, Doug; Stock, Saundra; Mori, Takashi; Sanberg, Paul G; Murphy, Tanya; Tan, Jun

    2013-09-01

    Autistic individuals display impaired social interactions and language, and restricted, stereotyped behaviors. Elevated levels of secreted amyloid precursor protein-alpha (sAPPα), the product of α-secretase cleavage of APP, are found in the plasma of some individuals with autism. The sAPPα protein is neurotrophic and neuroprotective and recently showed a correlation to glial differentiation in human neural stem cells (NSCs) via the IL-6 pathway. Considering evidence of gliosis in postmortem autistic brains, we hypothesized that subsets of patients with autism would exhibit elevations in CNS sAPPα and mice generated to mimic this observation would display markers suggestive of gliosis and autism-like behavior. Elevations in sAPPα levels were observed in brains of autistic patients compared to controls. Transgenic mice engineered to overexpress human sAPPα (TgsAPPα mice) displayed hypoactivity, impaired sociability, increased brain glial fibrillary acidic protein (GFAP) expression, and altered Notch1 and IL-6 levels. NSCs isolated from TgsAPPα mice, and those derived from wild-type mice treated with sAPPα, displayed suppressed β-tubulin III and elevated GFAP expression. These results suggest that elevations in brain sAPPα levels are observed in subsets of individuals with autism and TgsAPPα mice display signs suggestive of gliosis and behavioral impairment. PMID:23840007

  11. GFAP expression and social deficits in transgenic mice overexpressing human sAPPα

    PubMed Central

    Bailey, Antoinette R; Hou, Huayan; Song, Min; Obregon, Demian F; Portis, Samantha; Barger, Steven; Shytle, Doug; Stock, Saundra; Mori, Takashi; Sanberg, Paul G; Murphy, Tanya; Tan, Jun

    2013-01-01

    Autistic individuals display impaired social interactions and language, and restricted, stereotyped behaviors. Elevated levels of secreted amyloid precursor protein-alpha (sAPPα), the product of α-secretase cleavage of APP, are found in the plasma of some individuals with autism. The sAPPα protein is neurotrophic and neuroprotective and recently showed a correlation to glial differentiation in human neural stem cells (NSCs) via the IL-6 pathway. Considering evidence of gliosis in postmortem autistic brains, we hypothesized that subsets of patients with autism would exhibit elevations in CNS sAPPα and mice generated to mimic this observation would display markers suggestive of gliosis and autism-like behavior. Elevations in sAPPα levels were observed in brains of autistic patients compared to controls. Transgenic mice engineered to overexpress human sAPPα (TgsAPPα mice) displayed hypoactivity, impaired sociability, increased brain glial fibrillary acidic protein (GFAP) expression, and altered Notch1 and IL-6 levels. NSCs isolated from TgsAPPα mice, and those derived from wild-type mice treated with sAPPα, displayed suppressed β-tubulin III and elevated GFAP expression. These results suggest that elevations in brain sAPPα levels are observed in subsets of individuals with autism and TgsAPPα mice display signs suggestive of gliosis and behavioral impairment. PMID:23840007

  12. Decreased Plasma Aβ in Hyperlipidemic APPSL Transgenic Mice Is Associated with BBB Dysfunction

    PubMed Central

    Löffler, Tina; Flunkert, Stefanie; Temmel, Magdalena; Hutter-Paier, Birgit

    2016-01-01

    Besides the continued focus on Aβ and Tau in Alzheimer's disease (AD), it is increasingly evident that other pathologic characteristics, such as vascular alterations or inflammation, are associated with AD. Whether these changes are an initial cause for the onset of AD or occur as a result of the disease in late stages is still under debate. In the present study, the impact of the high-fat diet (HFD) induced vascular risk factor hyperlipidemia on Aβ levels and clearance as well as cerebral vasculature and blood-brain barrier (BBB) integrity was examined in mice. For this purpose, human APP transgenic (APPSL) and wildtype (WT) mice were fed a HFD for 12 weeks. Plasma and tissues were subsequently investigated for Aβ distribution and concentrations of several vascular markers. Decreased plasma Aβ together with increased levels of insoluble Aβ and amyloid plaques in the brains of HFD fed APPSL mice point toward impaired Aβ clearance due to HFD. Additionally, HFD induced manifold alterations in the cerebral vasculature and BBB integrity exclusively in human APP overexpressing mice but not in wildtype mice. Therefore, HFD appears to enhance Aβ dependent vascular/BBB dysfunction in combination with an increased proportion of cerebral to plasma Aβ in APPSL mice. PMID:27313503

  13. Overexpression of IGF-I in skeletal muscle of transgenic mice does not prevent unloading-induced atrophy

    NASA Technical Reports Server (NTRS)

    Criswell, D. S.; Booth, F. W.; DeMayo, F.; Schwartz, R. J.; Gordon, S. E.; Fiorotto, M. L.

    1998-01-01

    This study examined the association between local insulin-like growth factor I (IGF-I) overexpression and atrophy in skeletal muscle. We hypothesized that endogenous skeletal muscle IGF-I mRNA expression would decrease with hindlimb unloading (HU) in mice, and that transgenic mice overexpressing human IGF-I (hIGF-I) specifically in skeletal muscle would exhibit less atrophy after HU. Male transgenic mice and nontransgenic mice from the parent strain (FVB) were divided into four groups (n = 10/group): 1) transgenic, weight-bearing (IGF-I/WB); 2) transgenic, hindlimb unloaded (IGF-I/HU); 3) nontransgenic, weight-bearing (FVB/WB); and 4) nontransgenic, hindlimb unloaded (FVB/HU). HU groups were hindlimb unloaded for 14 days. Body mass was reduced (P < 0.05) after HU in both IGF-I (-9%) and FVB mice (-13%). Contrary to our hypothesis, we found that the relative abundance of mRNA for the endogenous rodent IGF-I (rIGF-I) was unaltered by HU in the gastrocnemius (GAST) muscle of wild-type FVB mice. High-level expression of hIGF-I peptide and mRNA was confirmed in the GAST and tibialis anterior (TA) muscles of the transgenic mice. Nevertheless, masses of the GAST and TA muscles were reduced (P < 0.05) in both FVB/HU and IGF-I/HU groups compared with FVB/WB and IGF-I/WB groups, respectively, and the percent atrophy in mass of these muscles did not differ between FVB and IGF-I mice. Therefore, skeletal muscle atrophy may not be associated with a reduction of endogenous rIGF-I mRNA level in 14-day HU mice. We conclude that high local expression of hIGF-I mRNA and peptide in skeletal muscle alone cannot attenuate unloading-induced atrophy of fast-twitch muscle in mice.

  14. GABAB Receptor Constituents Revealed by Tandem Affinity Purification from Transgenic Mice*

    PubMed Central

    Bartoi, Tudor; Rigbolt, Kristoffer T. G.; Du, Dan; Köhr, Georg; Blagoev, Blagoy; Kornau, Hans-Christian

    2010-01-01

    GABAB receptors function as heterodimeric G-protein-coupled receptors for the neurotransmitter γ-aminobutyric acid (GABA). Receptor subtypes, based on isoforms of the ligand-binding subunit GABAB1, are thought to involve a differential set of associated proteins. Here, we describe two mouse lines that allow a straightforward biochemical isolation of GABAB receptors. The transgenic mice express GABAB1 isoforms that contain sequences for a two-step affinity purification, in addition to their endogenous subunit repertoire. Comparative analyses of purified samples from the transgenic mice and wild-type control animals revealed two novel components of the GABAB1 complex. One of the identified proteins, potassium channel tetramerization domain-containing protein 12, associates with heterodimeric GABAB receptors via the GABAB2 subunit. In transfected hippocampal neurons, potassium channel tetramerization domain-containing protein 12 augmented axonal surface targeting of GABAB2. The mice equipped with tags on GABAB1 facilitate validation and identification of native binding partners of GABAB receptors, providing insight into the molecular mechanisms of synaptic modulation. PMID:20406808

  15. GABAB receptor constituents revealed by tandem affinity purification from transgenic mice.

    PubMed

    Bartoi, Tudor; Rigbolt, Kristoffer T G; Du, Dan; Köhr, Georg; Blagoev, Blagoy; Kornau, Hans-Christian

    2010-07-01

    GABA(B) receptors function as heterodimeric G-protein-coupled receptors for the neurotransmitter gamma-aminobutyric acid (GABA). Receptor subtypes, based on isoforms of the ligand-binding subunit GABA(B1), are thought to involve a differential set of associated proteins. Here, we describe two mouse lines that allow a straightforward biochemical isolation of GABA(B) receptors. The transgenic mice express GABA(B1) isoforms that contain sequences for a two-step affinity purification, in addition to their endogenous subunit repertoire. Comparative analyses of purified samples from the transgenic mice and wild-type control animals revealed two novel components of the GABA(B1) complex. One of the identified proteins, potassium channel tetramerization domain-containing protein 12, associates with heterodimeric GABA(B) receptors via the GABA(B2) subunit. In transfected hippocampal neurons, potassium channel tetramerization domain-containing protein 12 augmented axonal surface targeting of GABA(B2). The mice equipped with tags on GABA(B1) facilitate validation and identification of native binding partners of GABA(B) receptors, providing insight into the molecular mechanisms of synaptic modulation. PMID:20406808

  16. Sex dimorphisms of crossbridge cycling kinetics in transgenic hypertrophic cardiomyopathy mice.

    PubMed

    Birch, Camille L; Behunin, Samantha M; Lopez-Pier, Marissa A; Danilo, Christiane; Lipovka, Yulia; Saripalli, Chandra; Granzier, Henk; Konhilas, John P

    2016-07-01

    Familial hypertrophic cardiomyopathy (HCM) is a disease of the sarcomere and may lead to hypertrophic, dilated, restrictive, and/or arrhythmogenic cardiomyopathy, congestive heart failure, or sudden cardiac death. We hypothesized that hearts from transgenic HCM mice harboring a mutant myosin heavy chain increase the energetic cost of contraction in a sex-specific manner. To do this, we assessed Ca(2+) sensitivity of tension and crossbridge kinetics in demembranated cardiac trabeculas from male and female wild-type (WT) and HCM hearts at an early time point (2 mo of age). We found a significant effect of sex on Ca(2+) sensitivity such that male, but not female, HCM mice displayed a decrease in Ca(2+) sensitivity compared with WT counterparts. The HCM transgene and sex significantly impacted the rate of force redevelopment by a rapid release-restretch protocol and tension cost by the ATPase-tension relationship. In each of these measures, HCM male trabeculas displayed a gain-of-function when compared with WT counterparts. In addition, cardiac remodeling measured by echocardiography, histology, morphometry, and posttranslational modifications demonstrated sex- and HCM-specific effects. In conclusion, female and male HCM mice display sex dimorphic crossbridge kinetics accompanied by sex- and HCM-dependent cardiac remodeling at the morphometric, histological, and cellular level. PMID:27199124

  17. Dendritic spinopathy in transgenic mice expressing ALS/dementia-linked mutant UBQLN2.

    PubMed

    Gorrie, George H; Fecto, Faisal; Radzicki, Daniel; Weiss, Craig; Shi, Yong; Dong, Hongxin; Zhai, Hong; Fu, Ronggen; Liu, Erdong; Li, Sisi; Arrat, Hasan; Bigio, Eileen H; Disterhoft, John F; Martina, Marco; Mugnaini, Enrico; Siddique, Teepu; Deng, Han-Xiang

    2014-10-01

    Mutations in the gene encoding ubiquilin2 (UBQLN2) cause amyotrophic lateral sclerosis (ALS), frontotemporal type of dementia, or both. However, the molecular mechanisms are unknown. Here, we show that ALS/dementia-linked UBQLN2(P497H) transgenic mice develop neuronal pathology with ubiquilin2/ubiquitin/p62-positive inclusions in the brain, especially in the hippocampus, recapitulating several key pathological features of dementia observed in human patients with UBQLN2 mutations. A major feature of the ubiquilin2-related pathology in these mice, and reminiscent of human disease, is a dendritic spinopathy with protein aggregation in the dendritic spines and an associated decrease in dendritic spine density and synaptic dysfunction. Finally, we show that the protein inclusions in the dendritic spines are composed of several components of the proteasome machinery, including Ub(G76V)-GFP, a representative ubiquitinated protein substrate that is accumulated in the transgenic mice. Our data, therefore, directly link impaired protein degradation to inclusion formation that is associated with synaptic dysfunction and cognitive deficits. These data imply a convergent molecular pathway involving synaptic protein recycling that may also be involved in other neurodegenerative disorders, with implications for development of widely applicable rational therapeutics. PMID:25246588

  18. Tanshinone IIA Alleviates the AD Phenotypes in APP and PS1 Transgenic Mice

    PubMed Central

    Li, Fengling; Han, Guosheng; Wu, Kexiang

    2016-01-01

    Therapeutic approach for Alzheimer's disease (AD) is still deficient. To find active compounds from herbal medicine is of interest in the alleviation of AD symptoms. This study aimed to investigate the protective effects of Tanshinone IIA (TIIA) on memory performance and synaptic plasticity in a transgenic AD model at the early phase. 25–100 mg/kg TIIA (intraperitoneal injection, i.p.) was administered to the six-month-old APP and PS1 transgenic mice for 30 consecutive days. After treatment, spatial memory, synaptic plasticity, and related mechanisms were investigated. Our result showed that memory impairment in AD mice was mitigated by 50 and 100 mg/kg TIIA treatments. Hippocampal long-term potentiation was impaired in AD model but rescued by 100 mg/kg TIIA treatment. Mechanically, TIIA treatment reduced the accumulations of beta-amyloid 1–42, C-terminal fragments (CTFs), and p-Tau in the AD model. TIIA did not affect basal BDNF but promoted depolarization-induced BDNF synthesis in the AD mice. Taken together, TIIA repairs hippocampal LTP and memory, likely, through facilitating the clearance of AD-related proteins and activating synaptic BDNF synthesis. TIIA might be a candidate drug for AD treatment. PMID:27274990

  19. Mantle cell lymphoma in cyclin D1 transgenic mice with Bim-deficient B cells

    PubMed Central

    Katz, Samuel G.; LaBelle, James L.; Meng, Hailong; Valeriano, Regina P.; Fisher, Jill K.; Sun, Heather; Rodig, Scott J.; Kleinstein, Steven H.

    2014-01-01

    Mantle cell lymphoma (MCL) is a highly aggressive B-cell lymphoma resistant to conventional chemotherapy. Although defined by the characteristic t(11;14) translocation, MCL has not been recapitulated in transgenic mouse models of cyclin D1 overexpression alone. Indeed, several genetic aberrations have been identified in MCL that may contribute to its pathogenesis and chemoresistance. Of particular interest is the frequent biallelic deletion of the proapoptotic BCL-2 family protein BIM. BIM exerts its pro-death function via its α-helical BH3 death domain that has the dual capacity to inhibit antiapoptotic proteins such as BCL-2 and MCL-1 and directly trigger proapoptotic proteins such as the mitochondrial executioner protein BAX. To evaluate a functional role for Bim deletion in the pathogenesis of MCL, we generated cyclin D1–transgenic mice harboring Bim-deficient B cells. In response to immunization, EμCycD1CD19CREBimfl/fl mice manifested selective expansion of their splenic mantle zone compartment. Three distinct immune stimulation regimens induced lymphomas with histopathologic and molecular features of human MCL in a subset of mice. Thus, deletion of Bim in B cells, in the context of cyclin D1 overexpression, disrupts a critical control point in lymphoid maturation and predisposes to the development of MCL. This genetic proof of concept for MCL pathogenesis suggests an opportunity to reactivate the death pathway by pharmacologic mimicry of proapoptotic BIM. PMID:24352880

  20. RAP-011, an activin receptor ligand trap, increases hemoglobin concentration in Hepcidin transgenic mice

    PubMed Central

    Langdon, Jacqueline M.; Barkataki, Sangjucta; Berger, Alan E.; Cheadle, Chris; Xue, Qian-Li; Sung, Victoria; Roy, Cindy N.

    2014-01-01

    Over expression of hepcidin antimicrobial peptide is a common feature of iron-restricted anemia in humans. We investigated the erythroid response to either erythropoietin or RAP-011, a “murinized” ortholog of sotatercept, in C57BL/6 mice and in hepcidin antimicrobial peptide over expressing mice. Sotatercept, a soluble, activin receptor type IIA ligand trap, is currently being evaluated for the treatment of anemias associated with chronic renal disease, myelodysplastic syndrome, β-thalassemia, and Diamond Blackfan anemia and acts by inhibiting signaling downstream of activin and other Transforming Growth Factor-β superfamily members. We found that erythropoietin and RAP-011 increased hemoglobin concentration in C57BL/6 mice and in hepcidin antimicrobial peptide over expressing mice. While erythropoietin treatment depleted splenic iron stores in C57BL/6 mice, RAP-011 treatment did not deplete splenic iron stores in mice of either genotype. Bone marrow erythroid progenitors from erythropoietin-treated mice exhibited iron-restricted erythropoiesis, as indicated by increased median fluorescence intensity of transferrin receptor immunostaining by flow cytometry. In contrast, RAP-011-treated mice did not exhibit the same degree of iron-restricted erythropoiesis. In conclusion, we have demonstrated that RAP-011 can improve hemoglobin concentration in hepcidin antimicrobial peptide transgenic mice. Our data support the hypothesis that RAP-011 has unique biologic effects which prevent or circumvent depletion of mouse splenic iron stores. RAP-011 may, therefore, be an appropriate therapeutic for trials in human anemias characterized by increased expression of hepcidin antimicrobial peptide and iron-restricted erythropoiesis. PMID:25236856

  1. RAP-011, an activin receptor ligand trap, increases hemoglobin concentration in hepcidin transgenic mice.

    PubMed

    Langdon, Jacqueline M; Barkataki, Sangjucta; Berger, Alan E; Cheadle, Chris; Xue, Qian-Li; Sung, Victoria; Roy, Cindy N

    2015-01-01

    Over expression of hepcidin antimicrobial peptide is a common feature of iron-restricted anemia in humans. We investigated the erythroid response to either erythropoietin or RAP-011, a "murinized" ortholog of sotatercept, in C57BL/6 mice and in hepcidin antimicrobial peptide 1 over expressing mice. Sotatercept, a soluble, activin receptor type IIA ligand trap, is currently being evaluated for the treatment of anemias associated with chronic renal disease, myelodysplastic syndrome, β-thalassemia, and Diamond Blackfan anemia and acts by inhibiting signaling downstream of activin and other Transforming Growth Factor-β superfamily members. We found that erythropoietin and RAP-011 increased hemoglobin concentration in C57BL/6 mice and in hepcidin antimicrobial peptide 1 over expressing mice. While erythropoietin treatment depleted splenic iron stores in C57BL/6 mice, RAP-011 treatment did not deplete splenic iron stores in mice of either genotype. Bone marrow erythroid progenitors from erythropoietin-treated mice exhibited iron-restricted erythropoiesis, as indicated by increased median fluorescence intensity of transferrin receptor immunostaining by flow cytometry. In contrast, RAP-011-treated mice did not exhibit the same degree of iron-restricted erythropoiesis. In conclusion, we have demonstrated that RAP-011 can improve hemoglobin concentration in hepcidin antimicrobial peptide 1 transgenic mice. Our data support the hypothesis that RAP-011 has unique biologic effects which prevent or circumvent depletion of mouse splenic iron stores. RAP-011 may, therefore, be an appropriate therapeutic for trials in human anemias characterized by increased expression of hepcidin antimicrobial peptide and iron-restricted erythropoiesis. PMID:25236856

  2. Chronic wasting disease of deer and elk in transgenic mice: oral transmission and pathobiology.

    PubMed

    Trifilo, Matthew J; Ying, Ge; Teng, Chao; Oldstone, Michael B A

    2007-08-15

    To study the pathogenesis of chronic wasting disease (CWD) in deer and elk, transgenic (tg) mice were generated that expressed the prion protein (PrP) of deer containing a glycine at amino acid (aa) 96 and a serine at aa 225 under transcriptional control of the murine PrP promoter. This construct was introduced into murine PrP-deficient mice. As anticipated, neither non-tg mice nor PrP ko mice were susceptible when inoculated intracerebrally (i.c.) or orally with CWD brain material (scrapie pool from six mule deer) and followed for 600+ days (dpi). Deer PrP tg mice were not susceptible to i.c. inoculation with murine scrapie. In contrast, a fatal neurologic disease occurred accompanied by conversion of deer PrPsen to PrPres by western blot and immunohistochemistry after either i.c. inoculation with CWD brain into two lines of tg mice studied (312+32 dpi [mean+2 standard errors] for the heterozygous tg line 33, 275+46 dpi for the heterozygous tg line 39 and 210 dpi for the homozygous tg line 33) or after oral inoculation (381+55 dpi for the homozygous tg line 33 and 370+26 dpi for the homozygous tg line 39). Kinetically, following oral inoculation of CWD brain, PrPres was observed by day 200 when mice were clinically healthy in the posterior surface of the dorsum of the tongue primarily in serous and mucous glands, in the intestines, in large cells at the splenic marginal zone that anatomically resembled follicular dendritic cells and macrophages and in the olfactory bulb and brain stem but did not occur in the cerebellum, cerebral cortex or hippocampus or in hearts, lungs and livers of infected mice. After 350 days when mice become clinically ill the cerebellum, cerebral cortex and hippocampus became positive for PrPres and displayed massive spongiosis, neuronal drop out, gliosis and florid plaques. PMID:17451773

  3. Hepatic overexpression of bovine scavenger receptor type I in transgenic mice prevents diet-induced hyperbetalipoproteinemia.

    PubMed Central

    Wölle, S; Via, D P; Chan, L; Cornicelli, J A; Bisgaier, C L

    1995-01-01

    Hepatic scavenger receptors (SR) may play a protective role by clearing modified lipoproteins before they target the artery wall. To gain insight into this hypothesized function, transgenic mice expressing hepatic bovine SR (TgSR) were created and studied when fed chow, and during diet-induced hyperlipidemia. SR overexpression resulted in extensive hepatic parenchymal cell uptake of fluorescently labeled acetylated human low density lipoprotein (DiI ac-hLDL) and a twofold increase in 125I-acetylated-LDL clearance. Food intake and cholesterol absorption was indistinguishable between control and TgSR mice. In chow-fed mice, lipoprotein cholesterol was similar in control and TgSR mice. However, on a 3-wk high fat/cholesterol (HFHC) diet, the rise in apoB containing lipoproteins was suppressed in TgSR+/- and TgSR+/+ mice. The rise in HDL was similar in control and TgSR+/- mice, but significantly elevated in the TgSR+/+ mice. Overall, on chow, the ratio of apo-B containing lipoprotein cholesterol to HDL cholesterol was similar for all groups (control = 0.33; TgSR+/- = 0.32; TgSR+/+ = 0.38). However, after 3 wk on the HFHC diet, this ratio was markedly higher in control (2.34 +/- 0.21) than in either TgSR+/- (1.00 +/- 0.24) or TgSR+/+ (1.00 +/- 0.19) mice. In TgSR+/- mice, hepatic cholesteryl esters were reduced by 59%, 7 alpha-hydroxylase mRNA levels were elevated twofold, and a significant increase in fecal bile acid flux was observed after the 3-wk HFHC diet. These results suggest SR may play a protective role in liver by preventing diet-induced increases in apoB containing lipoproteins. Images PMID:7615795

  4. Osteopenia in transgenic mice with osteoblast-targeted expression of the inducible cAMP early repressor

    PubMed Central

    Chandhoke, Taranpreet K.; Huang, Yu-Feng; Liu, Fei; Gronowicz, Gloria A.; Adams, Douglas J.; Harrison, John R.; Kream, Barbara E.

    2008-01-01

    ICER is a member of the CREM family of basic leucine zipper transcription factors that acts as a dominant negative regulator of gene transcription. Four different isoforms of ICER (I, Iγ, II and IIγ) are transcribed from the P2 promoter of the Crem gene. We previously found that each of the ICER isoforms is induced by parathyroid hormone in osteoblasts. The goal of the present study was to assess the function of ICER in bone by overexpressing ICER in osteoblasts of transgenic mice. ICER I and ICER II cDNAs, each containing an N-terminal FLAG epitope tag, were cloned downstream of a fragment containing 3.6 kb of the rat Col1a1 promoter and most of the rat Col1a1 first intron to produce pOBCol3.6-ICER I and pOBCol3.6-ICER II transgenes, respectively. Multiple lines of mice were generated bearing the ICER I and ICER II transgenes. At 8 weeks of age, ICER I and ICER II transgenic mice had lower body weights and decreased bone mineral density of femurs and vertebrae. Further studies were done with ICER I transgenic mice, which had had greatly reduced trabecular bone volume and a markedly decreased bone formation rate in femurs. Osteoblast differentiation and osteocalcin expression were reduced in ex vivo bone marrow cultures from ICER I transgenic mice. ICER I antagonized the activity of ATF4 at its consensus DNA binding site in the osteocalcin promoter in vitro. Thus, transgenic mice with osteoblast-targeted overexpression of ICER resulted in osteopenia caused primarily by reduced bone formation. We speculate that ICER regulates the activity and/or expression of ATF/CREB factors required for normal bone formation. PMID:18460422

  5. A soluble form of Siglec-9 provides an antitumor benefit against mammary tumor cells expressing MUC1 in transgenic mice

    SciTech Connect

    Tomioka, Yukiko; Morimatsu, Masami; Nishijima, Ken-ichi; Usui, Tatsufumi; Yamamoto, Sayo; Suyama, Haruka; Ozaki, Kinuyo; Ito, Toshihiro; and others

    2014-07-18

    Highlights: • Tumor-associated antigen MUC1 binds to Siglec-9. • Soluble Siglec-9 reduced proliferation of MUC1-positive tumor in transgenic mice. • Soluble Siglec-9 and MUC1 on tumor cells were colocalized in transgenic mice. • MUC1 expression on tumor cells were reduced in soluble Siglec-9 transgenic mice. - Abstract: Tumor-associated MUC1 binds to Siglec-9, which is expected to mediate tumor cell growth and negative immunomodulation. We hypothesized that a soluble form of Siglec-9 (sSiglec-9) competitively inhibits a binding of MUC1 to its receptor molecules like human Siglec-9, leading to provide antitumor benefit against MUC1-expressing tumor, and generated transgenic mouse lines expressing sSiglec-9 (sSiglec-9 Tg). When mammary tumor cells expressing MUC1 were intraperitoneally transplanted into sSiglec-9 Tg, tumor proliferation was slower with the lower histological malignancy as compared with non-transgenic mice. The sSiglec-9 was detected in the ascites caused by the tumor in the sSiglec-9 Tg, and sSiglec-9 and MUC1 were often colocalized on surfaces of the tumor cells. PCNA immunohistochemistry also revealed the reduced proliferation of the tumor cells in sSiglec-9 Tg. In sSiglec-9 Tg with remarkable suppression of tumor proliferation, MUC1 expressions were tend to be reduced. In the ascites of sSiglec-9 Tg bearing the tumor, T cells were uniformly infiltrated, whereas aggregations of degenerative T cells were often observed in the non-transgenic mice. These results suggest that sSiglec-9 has an antitumor benefit against MUC1-expressing tumor in the transgenic mice, which may avoid the negative immunomodulation and/or suppress tumor-associated MUC1 downstream signal transduction, and subsequent tumor proliferation.

  6. Increased microvascular density and enhanced leukocyte rolling and adhesion in the skin of VEGF transgenic mice.

    PubMed

    Detmar, M; Brown, L F; Schön, M P; Elicker, B M; Velasco, P; Richard, L; Fukumura, D; Monsky, W; Claffey, K P; Jain, R K

    1998-07-01

    Vascular endothelial growth factor (VEGF) has been implicated in the pathologic angiogenesis observed in psoriasis and other chronic inflammatory skin diseases that are characterized by enhanced expression of VEGF by epidermal keratinocytes and of VEGF receptors by tortuous microvessels in the upper dermis. To investigate the functional importance of chronic VEGF overexpression in vivo, we used a keratin 14 promoter expression cassette containing the gene for murine VEGF164 to selectively target VEGF expression to basal epidermal keratinocytes in transgenic mice. These mice demonstrated an increased density of tortuous cutaneous blood capillaries with elevated expression levels of the high affinity VEGF receptors, VEGFR-1 and VEGFR-2, most prominently during the neonatal period. In contrast, no abnormalities of lymphatic vessels were detected. In addition, the number of mast cells in the upper dermis was significantly increased in transgenic skin. Intravital fluorescence microscopy revealed highly increased leukocyte rolling and adhesion in postcapillary skin venules that were both inhibited after injection of blocking antibodies against E- and P-selectin. Combined blocking antibodies against intercellular adhesion molecule-1 and lymphocyte function-associated antigen-1 were without effect, whereas an anti-vascular cell adhesion molecule-1/VLA-4 antibody combination almost completely normalized the enhanced leukocyte adhesion in transgenic mice. This study reveals VEGF as a growth factor specific for blood vessels, but not lymphatic vessels, and demonstrates that chronic orthotopic overexpression of VEGF in the epidermis is sufficient to induce cardinal features of chronic skin inflammation, providing a molecular link between angiogenesis, mast cell accumulation, and leukocyte recruitment to sites of inflammation. PMID:9665379

  7. Respiratory epithelial cell expression of human transforming growth factor-alpha induces lung fibrosis in transgenic mice.

    PubMed Central

    Korfhagen, T R; Swantz, R J; Wert, S E; McCarty, J M; Kerlakian, C B; Glasser, S W; Whitsett, J A

    1994-01-01

    Increased production of EGF or TGF-alpha by the respiratory epithelial cells has been associated with the pathogenesis of various forms of lung injury. Growth factors and cytokines are thought to act locally, via paracrine and autocrine mechanisms, to stimulate cell proliferation and matrix deposition by interstitial lung cells resulting in pulmonary fibrosis. To test whether TGF-alpha mediates pulmonary fibrotic responses, we have generated transgenic mice expressing human TGF-alpha under control of regulatory regions of the human surfactant protein C (SP-C) gene. Human TGF-alpha mRNA was expressed in pulmonary epithelial cells in the lungs of the transgenic mice. Adult mice bearing the SP-C-TGF-alpha transgene developed severe pulmonary fibrosis. Fibrotic lesions were observed in peribronchial, peribronchiolar, and perivascular regions, as well as subjacent to pleural surfaces. Lesions consisted of fibrous tissue that included groups of epithelial cells expressing endogenous SP-C mRNA, consistent with their identification as distal respiratory epithelial cells. Peripheral fibrotic regions consisted of thickened pleura associated with extensive collagen deposition. Alveolar architecture was disrupted in the transgenic mice with loss of alveoli in the lung parenchyma. Pulmonary epithelial cell expression of TGF-alpha in transgenic mice disrupts alveolar morphogenesis and produces fibrotic lesions mediated by paracrine signaling between respiratory epithelial and interstitial cells of the lung. Images PMID:8163670

  8. Multiple Renal Cyst Development but Not Situs Abnormalities in Transgenic RNAi Mice against Inv::GFP Rescue Gene

    PubMed Central

    Kamijho, Yuki; Shiozaki, Yayoi; Sakurai, Eiki; Hanaoka, Kazunori; Watanabe, Daisuke

    2014-01-01

    In this study we generated RNA interference (RNAi)-mediated gene knockdown transgenic mice (transgenic RNAi mice) against the functional Inv gene. Inv mutant mice show consistently reversed internal organs (situs inversus), multiple renal cysts and neonatal lethality. The Inv::GFP-rescue mice, which introduced the Inv::GFP fusion gene, can rescue inv mutant mice phenotypes. This indicates that the Inv::GFP gene is functional in vivo. To analyze the physiological functions of the Inv gene, and to demonstrate the availability of transgenic RNAi mice, we introduced a short hairpin RNA expression vector against GFP mRNA into Inv::GFP-rescue mice and analyzed the gene silencing effects and Inv functions by examining phenotypes. Transgenic RNAi mice with the Inv::GFP-rescue gene (Inv-KD mice) down-regulated Inv::GFP fusion protein and showed hypomorphic phenotypes of inv mutant mice, such as renal cyst development, but not situs abnormalities or postnatal lethality. This indicates that shRNAi-mediated gene silencing systems that target the tag sequence of the fusion gene work properly in vivo, and suggests that a relatively high level of Inv protein is required for kidney development in contrast to left/right axis determination. Inv::GFP protein was significantly down-regulated in the germ cells of Inv-KD mice testis compared with somatic cells, suggesting the existence of a testicular germ cell-specific enhanced RNAi system that regulates germ cell development. The Inv-KD mouse is useful for studying Inv gene functions in adult tissue that are unable to be analyzed in inv mutant mice showing postnatal lethality. In addition, the shRNA-based gene silencing system against the tag sequence of the fusion gene can be utilized as a new technique to regulate gene expression in either in vitro or in vivo experiments. PMID:24586938

  9. Studies on induction of lamotrigine metabolism in transgenic UGT1 mice

    PubMed Central

    Argikar, U. A.; Senekeo-Effenberger, K.; Larson, E. E.; Tukey, R. H.; Remmel, R. P.

    2010-01-01

    A transgenic ‘knock-in’ mouse model expressing a human UGT1 locus (Tg-UGT1) was recently developed and validated. Although these animals express mouse UGT1A proteins, UGT1A4 is a pseudo-gene in mice. Therefore, Tg-UGT1 mice serve as a ‘humanized’ UGT1A4 animal model.Lamotrigine (LTG) is primarily metabolized to its N-glucuronide (LTGG) by hUGT1A4. This investigation aimed at examining the impact of pregnane X receptor (PXR), constitutive androstane receptor (CAR) and peroxisome proliferator-activated receptor (PPAR) activators on LTG glucuronidation in vivo and in vitro. Tg-UGT1 mice were administered the inducers phenobarbital (CAR), pregnenolone-16α-carbonitrile (PXR), WY-14643 (PPAR-α), ciglitazone (PPAR-γ), or L-165041 (PPAR-β), once daily for 3 or 4 days. Thereafter, LTG was administered orally and blood samples were collected over 24 h. LTG was measured in blood and formation of LTGG was measured in pooled microsomes made from the livers of treated animals.A three-fold increase in in vivo LTG clearance was seen after phenobarbital administration. In microsomes prepared from phenobarbital-treated Tg-UGT1 animals, 13-fold higher CLint (Vmax/Km) value was observed as compared with the untreated transgenic mice. A trend toward induction of catalytic activity in vitro and in vivo was also observed following pregnenolone-16α-carbonitrile and WY-14643 treatment. This study demonstrates the successful application of Tg-UGT1 mice as a novel tool to study the impact of induction and regulation on metabolism of UGT1A4 substrates. PMID:19845433

  10. Generation and characterization of KsprtTA and KsptTA transgenic mice

    PubMed Central

    Pan, Xinchao; Small, Erin V.; Igarashi, Peter; Carroll, Thomas J.

    2013-01-01

    The advent of technologies that allow tissue specific expression or ablation of genes has contributed enormously to our knowledge of the mechanism regulating organ development and maintenance in mice. The tetracycline inducible system allows reversible regulation of gene products upon administration of Doxycycline. Here we describe the generation and activity of two transgenic lines expressing the cDNAs for the Tet responsive transcription factors rtTA and tTA (Tet-on and off) respectively under the control of an element that drives expression in the epithelium of the developing and adult kidney. Both lines show inducible and reversible activity in the embryonic and adult organ. PMID:23420736

  11. The Long Journey of TCL1 Transgenic Mice: Lessons Learned in the Last 15 Years

    PubMed Central

    Pekarsky, Yuri; Drusco, Alessandra; Kumchala, Prasanthi; Croce, Carlo M.; Zanesi, Nicola

    2016-01-01

    The first transgenic mouse of the TCL1 oncogene was described more than 15 years ago, and since then, the overexpression of the gene in T- and B-cells in vivo has been extensively studied to reveal the molecular details in the pathogenesis of some lymphocytic leukemias. This review discusses the main features of the original TCL1 models and the different lines of research successively developed with particular attention to genetically compound mice and the therapeutic applications in drug development. PMID:25700368

  12. Assessing the Susceptibility of Transgenic Mice Overexpressing Deer Prion Protein to Bovine Spongiform Encephalopathy

    PubMed Central

    Vickery, Christopher M.; Lockey, Richard; Holder, Thomas M.; Thorne, Leigh; Beck, Katy E.; Wilson, Christina; Denyer, Margaret; Sheehan, John; Marsh, Sarah; Webb, Paul R.; Dexter, Ian; Norman, Angela; Popescu, Emma; Schneider, Amanda; Holden, Paul; Griffiths, Peter C.; Plater, Jane M.; Dagleish, Mark P.; Martin, Stuart; Telling, Glenn C.; Simmons, Marion M.

    2014-01-01

    Several transgenic mouse models have been developed which facilitate the transmission of chronic wasting disease (CWD) of cervids and allow prion strain discrimination. The present study was designed to assess the susceptibility of the prototypic mouse line, Tg(CerPrP)1536+/−, to bovine spongiform encephalopathy (BSE) prions, which have the ability to overcome species barriers. Tg(CerPrP)1536+/− mice challenged with red deer-adapted BSE resulted in 90% to 100% attack rates, and BSE from cattle failed to transmit, indicating agent adaptation in the deer. PMID:24257620

  13. Tissue-specific and developmentally regulated expression of a chimeric actin-globin gene in transgenic mice.

    PubMed Central

    Shani, M

    1986-01-01

    A chimeric plasmid containing about 2/3 of the rat skeletal muscle actin gene plus 730 base pairs of its 5' flanking sequences fused to the 3' end of a human embryonic globin gene (D. Melloul, B. Aloni, J. Calvo, D. Yaffe, and U. Nudel, EMBO J. 3:983-990, 1984) was inserted into mice by microinjection into fertilized eggs. Eleven transgenic mice carrying the chimeric gene with or without plasmid pBR322 DNA sequences were identified. The majority of these mice transmitted the injected DNA to about 50% of their progeny. However, in transgenic mouse CV1, transmission to progeny was associated with amplification or deletion of the injected DNA sequences, while in transgenic mouse CV4 transmission was distorted, probably as a result of insertional mutagenesis. Tissue-specific expression was dependent on the removal of the vector DNA sequences from the chimeric gene sequences prior to microinjection. None of the transgenic mice carrying the chimeric gene together with plasmid pBR322 sequences expressed the introduced gene in striated muscles. In contrast, the six transgenic mice carrying the chimeric gene sequences alone expressed the inserted gene specifically in skeletal and cardiac muscles. Moreover, expression of the chimeric gene was not only tissue specific, but also developmentally regulated. Similar to the endogenous skeletal muscle actin gene, the chimeric gene was expressed at a relatively high level in cardiac muscle of neonatal mice and at a significantly lower level in adult cardiac muscle. These results indicate that the injected DNA included sufficient cis-acting control elements for its tissue-specific and developmentally regulated expression in transgenic mice. Images PMID:3023942

  14. Morphine Tolerance and Physical Dependence Are Altered in Conditional HIV-1 Tat Transgenic Mice.

    PubMed

    Fitting, Sylvia; Stevens, David L; Khan, Fayez A; Scoggins, Krista L; Enga, Rachel M; Beardsley, Patrick M; Knapp, Pamela E; Dewey, William L; Hauser, Kurt F

    2016-01-01

    Despite considerable evidence that chronic opiate use selectively affects the pathophysiologic consequences of human immunodeficiency virus type 1 (HIV-1) infection in the nervous system, few studies have examined whether neuro-acquired immune deficiency syndrome (neuroAIDS) might intrinsically alter the pharmacologic responses to chronic opiate exposure. This is an important matter because HIV-1 and opiate abuse are interrelated epidemics, and HIV-1 patients are often prescribed opiates as a treatment of HIV-1-related neuropathic pain. Tolerance and physical dependence are inevitable consequences of frequent and repeated administration of morphine. In the present study, mice expressing HIV-1 Tat in a doxycycline (DOX)-inducible manner [Tat(+)], their Tat(-) controls, and control C57BL/6 mice were chronically exposed to placebo or 75-mg morphine pellets to explore the effects of Tat induction on morphine tolerance and dependence. Antinociceptive tolerance and locomotor activity tolerance were assessed using tail-flick and locomotor activity assays, respectively, and physical dependence was measured with the platform-jumping assay and recording of other withdrawal signs. We found that Tat(+) mice treated with DOX [Tat(+)/DOX] developed an increased tolerance in the tail-flick assay compared with control Tat(-)/DOX and/or C57/DOX mice. Equivalent tolerance was developed in all mice when assessed by locomotor activity. Further, Tat(+)/DOX mice expressed reduced levels of physical dependence to chronic morphine exposure after a 1-mg/kg naloxone challenge compared with control Tat(-)/DOX and/or C57/DOX mice. Assuming the results seen in Tat transgenic mice can be generalized to neuroAIDS, our findings suggest that HIV-1-infected individuals may display heightened analgesic tolerance to similar doses of opiates compared with uninfected individuals and show fewer symptoms of physical dependence. PMID:26542403

  15. Effects of Environmental Enrichment and Physical Activity on Neurogenesis in Transgenic PS1/APP Mice

    PubMed Central

    Catlow, Briony J; Rowe, Amanda R; Clearwater, Courtney R; Mamcarz, Maggie; Arendash, Gary W; Sanchez-Ramos, Juan

    2009-01-01

    Rodents exposed to environmental enrichment show many differences, including improved cognitive performance, when compared to those living in standard (impoverished) housing. The purpose of the present study was to determine if a selective increase in neurogenesis occurred in cognitively-protected Tg mice raised in an enriched environment compared to those reared in physical activity housing. At weaning, double Tg APP+PS1 mice were placed into one of three environments: complete environmental enrichment (CE), enhanced physical activity (PA), or individual, impoverished housing (IMP). At 9–10 months of age, Tg mice were injected with BrdU (100 mg/kg BID) followed by euthanasia either 24 hrs or two weeks after the last injection. Unbiased estimates of BrdU positive cells in the hippocampal subgranular zone revealed a significant increase in cellular proliferation in Tg mice raised in CE or PA compared to Tg mice reared in IMP housing. However, counts of BrdU birth-dated cells two weeks after labeling showed no difference among the three groups, indicating decreased survival of cells in those groups (CE and PA) with higher cellular proliferation rates in the neurogenic niche. Counts of calretinin-expressing cells, a marker of immature neurons, also indicated no difference among the three groups of mice. In view of our prior study showing that enhanced cognitive activity (but not enhanced physical activity) protects Tg mice against cognitive impairment, the present results indicate that increased generation and survival of new neurons in the hippocampal dentate gyrus is not involved with the cognitively-protective effects of complete CE in Alzheimer’s transgenic mice. PMID:19135431

  16. Elevated systolic blood pressure in male GH transgenic mice is age dependent.

    PubMed

    Jara, Adam; Benner, Chance M; Sim, Don; Liu, Xingbo; List, Edward O; Householder, Lara A; Berryman, Darlene E; Kopchick, John J

    2014-03-01

    Acromegaly is associated with an increased incidence of cardiovascular disease. Transgenic mice expressing bovine GH (bGH) gene have previously been used to examine the effects of chronic GH stimulation on cardiovascular function. Results concerning systolic blood pressure (SBP) in bGH mice are conflicting. We hypothesized that these discrepancies may be the result of the various ages of the mice used in previous studies. In the current study, SBP was assessed monthly in male bGH mice from 3-12 months of age. Factors known to alter blood pressure were assessed during this time and included: levels of brain natriuretic peptide (BNP) and glucose homeostasis markers, and renal levels of angiotensin-converting enzyme 2 and endothelial nitric oxide synthase. Beginning at 6 months of age bGH had increased SBP compared with wild-type controls, which remained elevated through 12 months of age. Despite having increased blood pressure and cardiac BNP mRNA, bGH mice had decreased circulating levels of BNP. Additionally, bGH mice had an age-dependent decline in insulin levels. For example, they were hyperinsulinemic at 3 months, but by 11 months of age were hypoinsulinemic relative to wild-type controls. This decrease in insulin was accompanied by improved glucose tolerance at 11 months. Finally, both angiotensin-converting enzyme 2 and endothelial nitric oxide synthase expression were severely depressed in kidneys of 11-month-old bGH mice. These results indicate that elevated SBP in bGH mice is dependent on age, independent of insulin resistance, and related to alterations in both the natriuretic peptide and renin-angiotensin systems. PMID:24424040

  17. Monoclonal gammopathies in aging mu, kappa-transgenic mice: involvement of the B-1 cell lineage.

    PubMed

    van Arkel, C; Hopstaken, C M; Zurcher, C; Bos, N A; Kroese, F G; Savelkoul, H F; Benner, R; Radl, J

    1997-09-01

    Monoclonal gammopathies (MG) are monoclonal proliferative disorders of B cells at the differentiation stage of Ig production. They can be detected in the serum, either as transient or as persistent homogenous immunoglobulin (H-Ig) components. The exact phenotype, localization, and cell lineage origin of the precursor cells of MG are unknown, but may be crucial for both the correct diagnosis and for timely efficient treatment of the malignant forms. We used for the first time transgenic (Tg) mice (Sp6; mu/kappa) to study the origin of MG. In the mu, kappa Tg mice a small proportion of B cells can still produce endogenous IgM. These cells are of B-1 cell origin. The MG in Tg mice showed a later onset and a lower frequency than those in littermate control mice, mainly due to a four times lower frequency of benign monoclonal gammopathy. The 10% of B-1 cells that were able to produce endogenous Ig led to the development of MG in a frequency that was half the number of MG found in normal littermates. None of the MG in Tg mice produced an Ig of the Tg origin and therefore it can be concluded that they originated from B-1 cells. PMID:9341790

  18. Deposition of BACE-1 Protein in the Brains of APP/PS1 Double Transgenic Mice

    PubMed Central

    Luo, Gang; Xu, Hongxia; Huang, Yinuo; Mo, Dapeng; Song, Ligang; Jia, Baixue; Wang, Bo; Jin, Zhanqiang; Miao, Zhongrong

    2016-01-01

    The main causes of Alzheimer's disease remain elusive. Previous data have implicated the BACE-1 protein as a central player in the pathogenesis of Alzheimer's disease. However, many inhibitors of BACE-1 have failed during preclinical and clinical trials for AD treatment. Therefore, uncovering the exact role of BACE-1 in AD may have significant impact on the future development of therapeutic agents. Three- and six-month-old female APP/PS1 double transgenic mice were used to study abnormal accumulation of BACE-1 protein in brains of mice here. Immunofluorescence, immunohistochemistry, and western blot were performed to measure the distributing pattern and expression level of BACE-1. We found obvious BACE-1 protein accumulation in 3-month-old APP/PS1 mice, which had increased by the time of 6 months. Coimmunostaining results showed BACE-1 surrounded amyloid plaques in brain sections. The abnormal protein expression might not be attributable to the upregulation of BACE-1 protein, as no significant difference of protein expression was observed between wild-type and APP/PS1 mice. With antibodies against BACE-1 and CD31, we found a high immunoreactive density of BACE-1 protein on the outer layer of brain blood vessels. The aberrant distribution of BACE-1 in APP/PS1 mice suggests BACE-1 may be involved in the microvascular abnormality of AD. PMID:27294139

  19. An Efficient and Versatile System for Visualization and Genetic Modification of Dopaminergic Neurons in Transgenic Mice

    PubMed Central

    Kramer, Edgar R.

    2015-01-01

    Background & Aims The brain dopaminergic (DA) system is involved in fine tuning many behaviors and several human diseases are associated with pathological alterations of the DA system such as Parkinson’s disease (PD) and drug addiction. Because of its complex network integration, detailed analyses of physiological and pathophysiological conditions are only possible in a whole organism with a sophisticated tool box for visualization and functional modification. Methods & Results Here, we have generated transgenic mice expressing the tetracycline-regulated transactivator (tTA) or the reverse tetracycline-regulated transactivator (rtTA) under control of the tyrosine hydroxylase (TH) promoter, TH-tTA (tet-OFF) and TH-rtTA (tet-ON) mice, to visualize and genetically modify DA neurons. We show their tight regulation and efficient use to overexpress proteins under the control of tet-responsive elements or to delete genes of interest with tet-responsive Cre. In combination with mice encoding tet-responsive luciferase, we visualized the DA system in living mice progressively over time. Conclusion These experiments establish TH-tTA and TH-rtTA mice as a powerful tool to generate and monitor mouse models for DA system diseases. PMID:26291828

  20. Abeta42 gene vaccine prevents Abeta42 deposition in brain of double transgenic mice.

    PubMed

    Qu, Bao-Xi; Xiang, Qun; Li, Liping; Johnston, Stephen Albert; Hynan, Linda S; Rosenberg, Roger N

    2007-09-15

    Abeta42 peptide aggregation and deposition is an important component of the neuropathology of Alzheimer's disease (AD). Gene-gun mediated gene vaccination targeting Abeta42 is a potential method to prevent and treat AD. APPswe/PS1DeltaE9 transgenic (Tg) mice were immunized with an Abeta42 gene construct delivered by the gene gun. The vaccinated mice developed Th2 antibodies (IgG1) against Abeta42. The Abeta42 levels in brain were decreased by 41% and increased in plasma 43% in the vaccinated compared with control mice as assessed by ELISA analysis. Abeta42 plaque deposits in cerebral cortex and hippocampus were reduced by 51% and 52%, respectively, as shown by quantitative immunolabeling. Glial cell activation was also significantly attenuated in vaccinated compared with control mice. One rhesus monkey was vaccinated and developed anti-Abeta42 antibody. These new findings advance significantly our knowledge that gene-gun mediated Abeta42 gene immunization effectively induces a Th2 immune response and reduces the Abeta42 levels in brain in APPswe/PS1DeltaE9 mice. Abeta42 gene vaccination may be safe and efficient immunotherapy for AD. PMID:17574274

  1. Brain trauma in aged transgenic mice induces regression of established abeta deposits.

    PubMed

    Nakagawa, Y; Reed, L; Nakamura, M; McIntosh, T K; Smith, D H; Saatman, K E; Raghupathi, R; Clemens, J; Saido, T C; Lee, V M; Trojanowski, J Q

    2000-05-01

    Traumatic brain injury (TBI) increases susceptibility to Alzheimer's disease (AD), but it is not known if TBI affects the progression of AD. To address this question, we studied the neuropathological consequences of TBI in transgenic (TG) mice with a mutant human Abeta precursor protein (APP) mini-gene driven by a platelet-derived (PD) growth factor promoter resulting in overexpression of mutant APP (V717F), elevated brain Abeta levels, and AD-like amyloidosis. Since brain Abeta deposits first appear in 6-month-old TG (PDAPP) mice and accumulate with age, 2-year-old PDAPP and wild-type (WT) mice were subjected to controlled cortical impact (CCI) TBI or sham treatment. At 1, 9, and 16 weeks after TBI, neuron loss, gliosis, and atrophy were most prominent near the CCI site in PDAPP and WT mice. However, there also was a remarkable regression in the Abeta amyloid plaque burden in the hippocampus ipsilateral to TBI compared to the contralateral hippocampus of the PDAPP mice by 16 weeks postinjury. Thus, these data suggest that previously accumulated Abeta plaques resulting from progressive amyloidosis in the AD brain also may be reversible. PMID:10785464

  2. Overexpression of human loricrin in transgenic mice produces a normal phenotype.

    PubMed Central

    Yoneda, K; Steinert, P M

    1993-01-01

    The cornified cell envelope (CE) of terminally differentiating stratified squamous epithelial cells is a complex multiprotein assembly about 15 nm thick of which loricrin is a major component. We have produced transgenic mice bearing the human loricrin transgene in order to study the role of loricrin in CE assembly, structure, and function. By analyses of RNA and protein, we show that the human transgene is expressed in mouse epithelial tissues in an appropriate developmental manner but at an overall level about twice that of endogenous mouse loricrin. Thus the 20-kbp construct used contains all necessary regulatory elements. By immunogold electron microscopy, all of the expressed protein is incorporated into the CE. That no alternations were noted indicates that overproduction of the loricrin component of the CE does not affect the flexible structure or function of the epithelial tissues. Furthermore, these data imply that loricrin may be the last protein to be deposited onto, and thus lines, the intracellular surface of the CE, where it may be accessible to interact with the subjacent keratin intermediate-filament network. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 PMID:8248167

  3. Papillomavirus E6 proteins

    SciTech Connect

    Howie, Heather L.; Katzenellenbogen, Rachel A.; Galloway, Denise A.

    2009-02-20

    The papillomaviruses are small DNA viruses that encode approximately eight genes, and require the host cell DNA replication machinery for their viral DNA replication. Thus papillomaviruses have evolved strategies to induce host cell DNA synthesis balanced with strategies to protect the cell from unscheduled replication. While the papillomavirus E1 and E2 genes are directly involved in viral replication by binding to and unwinding the origin of replication, the E6 and E7 proteins have auxillary functions that promote proliferation. As a consequence of disrupting the normal checkpoints that regulate cell cycle entry and progression, the E6 and E7 proteins play a key role in the oncogenic properties of human papillomaviruses with a high risk of causing anogenital cancers (HR HPVs). As a consequence, E6 and E7 of HR HPVs are invariably expressed in cervical cancers. This article will focus on the E6 protein and its numerous activities including inactivating p53, blocking apoptosis, activating telomerase, disrupting cell adhesion, polarity and epithelial differentiation, altering transcription and reducing immune recognition.

  4. miR-483-3p regulates hyperglycaemia-induced cardiomyocyte apoptosis in transgenic mice.

    PubMed

    Qiao, Yu; Zhao, Yanli; Liu, Yan; Ma, Ning; Wang, Chuxuan; Zou, Jiaqi; Liu, Zhiyan; Zhou, Zhongqiu; Han, Dong; He, Jun; Sun, Qian; Liu, Yicong; Xu, Changqing; Du, Zhimin; Huang, Hui

    2016-09-01

    Diabetic cardiomyopathy represents severe heart complications, and is the leading cause of morbidity and mortality among patients with diabetes. Although a few microRNAs (miRNAs) have been implicated in diabetes-related complications, a functional association between miRNAs and cardiac dysfunction in diabetic cardiomyopathy remains to be demonstrated. Our results show that miR-483-3p is upregulated in streptozotocin-induced diabetic mice, and cultured cardiomyocytes mimicking hyperglycemia. Overexpressing miR-483-3p in transgenic mice with diabetes mellitus (DM) exacerbated cardiomyocyte apoptosis by transcriptionally repressing insulin growth factor 1 (IGF1). Therefore, we have uncovered a novel signaling pathway, involving miR-483-3p-IGF1, that promotes myocardial cell apoptosis under high blood-glucose condition. Further, our study indicates that miR-483-3p could be a potential therapeutic target for managing diabetes-associated heart complications. PMID:27346130

  5. Tissue expression profile of human neonatal Fc receptor (FcRn) in Tg32 transgenic mice.

    PubMed

    Fan, Yao-Yun; Avery, Lindsay B; Wang, Mengmeng; O'Hara, Denise M; Leung, Sheldon; Neubert, Hendrik

    2016-07-01

    The neonatal Fc receptor (FcRn) is a homeostatic receptor responsible for prolonging immunoglobulin G (IgG) half-life by protecting it from lysosomal degradation and recycling it to systemic circulation. Tissue-specific FcRn expression is a critical parameter in physiologically-based pharmacokinetic (PBPK) modeling for translational pharmacokinetics of Fc-containing biotherapeutics. Using online peptide immuno-affinity chromatography coupled with high resolution mass spectrometry, we established a quantitative FcRn tissue protein expression profile in human FcRn (hFcRn) transgenic mice, Tg32 homozygous and hemizygous strains. The concentration of hFcRn across 14 tissues ranged from 3.5 to 111.2 pmole per gram of tissue. Our hFcRn quantification data from Tg32 mice will enable a more refined PBPK model to improve the accuracy of human PK predictions for Fc-containing biotherapeutics. PMID:27104806

  6. Restoration of dystrophin-associated proteins in skeletal muscle of mdx mice transgenic for dystrophin gene.

    PubMed

    Matsumura, K; Lee, C C; Caskey, C T; Campbell, K P

    1993-04-12

    Duchenne muscular dystrophy (DMD) patients and mdx mice are characterized by the absence of dystrophin, a membrane cytoskeletal protein. Dystrophin is associated with a large oligomeric complex of sarcolemmal glycoproteins, including dystroglycan which provides a linkage to the extracellular matrix component, laminin. The finding that all of the dystrophin-associated proteins (DAPs) are drastically reduced in DMD and mdx skeletal muscle supports the primary function of dystrophin as an anchor of the sarcolemmal glycoprotein complex to the subsarcolemmal cytoskeleton. These findings indicate that the efficacy of dystrophin gene therapy will depend not only on replacing dystrophin but also on restoring all of the DAPs in the sarcolemma. Here we have investigated the status of the DAPs in the skeletal muscle of mdx mice transgenic for the dystrophin gene. Our results demonstrate that transfer of dystrophin gene restores all of the DAPs together with dystrophin, suggesting that dystrophin gene therapy should be effective in restoring the entire dystrophin-glycoprotein complex. PMID:8462697

  7. Growth hormone (GH) binding and effects of GH analogs in transgenic mice

    SciTech Connect

    Bartke, A.; Steger, R.W.; Turyn, D.

    1994-12-31

    Overexpression of human (h) or bovine (b) growth hormone (GH) in transgenic mice is associated with marked (2- to 12-fold) and significant increase in hepatic binding of GH and prolactin (PRL). This is due to an increase in the number of GH and PRL receptors (GHR, PRLR) per mg of microsomal protein without changes in binding affinity. Comparison of results obtained in transgenic animals expressing bGH with a mouse metallothionein (MT) or a rat phosphoenolpyruvate carboxykinase (PEPCK) promoter suggests that effects of bGH on hepatic GHR and PRLR do not require GH overexpression during fetal life and, within the dose range tested, the effects on PRLR are not dose dependent. The increase in hepatic GHR was accompanied by significant increases in plasma GH-binding protein (GHBP) and in mean residence time of injected GH. Thus life-long elevation of peripheral GH levels alters the availability of both free GH and GHR. Site-directed in vitro mutagenesis was used to produce hGH and bGH analogs mutated within one of the sites involved in binding to GHR and PRLR. Mutating hGH to produce amino acid identity with bGH at Position 11, 18 (within Helix 1), 57, or 60 (within the loop between Helix 1 and 2) did not affect binding to GHR in vitro, or somatotropic activity in transgenic mice in vivo but reduced lactogenic activity in Nb{sub 2} cells by 22%-45%. Mutations of bGH designed to produce amino acid identity with hGH at one to four of the corresponding positions in the bGH molecule did not interfere with binding to GHR or somatotropic activity in vivo, and failed to produce significant binding to PRLR but resulted in alterations in the effects on the hypothalamic and anterior pituitary function in transgenic mice. Apparently region(s) outside the domains examined are essential for lactogenic activity of hGH, and different portions of the GH molecule are responsible for its diverse actions in vivo. 35 refs.

  8. Expression of LIF in transgenic mice results in altered thymic epithelium and apparent interconversion of thymic and lymph node morphologies.

    PubMed Central

    Shen, M M; Skoda, R C; Cardiff, R D; Campos-Torres, J; Leder, P; Ornitz, D M

    1994-01-01

    Leukemia inhibitory factor (LIF) is a cytokine involved in embryonic and hematopoietic development. To investigate the effects of LIF on the lymphoid system, we generated a line of transgenic mice that expresses diffusible LIF protein specifically in T cells. These mice display two categories of phenotype that were not previously attributed to LIF overexpression. First, they display B cell hyperplasia, polyclonal hypergammaglobulinemia and mesangial proliferative glomerulonephritis, defects similar to those described for transgenic mice overexpressing the functionally related cytokine, interleukin-6. Secondly, the LIF transgenic mice display novel thymic and lymph node abnormalities. In the thymus, cortical CD4+CD8+ lymphocytes are lost, while numerous B cell follicles develop. Peripheral lymph nodes contain a vastly expanded CD4+CD8+ lymphocyte population. Furthermore, the thymic epithelium is profoundly disorganized, suggesting that disruption of stroma-lymphocyte interactions is responsible for many observed defects. Transplantation of transgenic bone marrow into wild type recipients transfers both the thymic and lymph node defects. However, transplantation of wild type marrow into transgenic recipients rescues the lymph node abnormality, but not the thymic defect, indicating the thymic epithelium is irreversibly altered. Our observations are consistent with a role for LIF in maintaining a functional thymic epithelium that will support proper T cell maturation. Images PMID:8137821

  9. Function and distribution of circulating human PCSK9 expressed extrahepatically in transgenic mice

    PubMed Central

    Luo, Yi; Warren, Laurie; Xia, Donghui; Jensen, Heather; Sand, Thomas; Petras, Stephen; Qin, Wenning; Miller, Kenneth S.; Hawkins, Julie

    2009-01-01

    Proprotein convertase subtilisin/kexin type 9 (PCSK9) is predominantly expressed in liver and regulates cholesterol metabolism by down regulating liver LDL receptor (LDLR) proteins. Here we report transgenic overexpression of human PCSK9 in kidney increased plasma levels of PCSK9 and subsequently led to a dramatic reduction in liver LDLR proteins. The regulation of LDLR by PCSK9 displayed tissue specificity, with liver being the most responsive tissue. Even though the PCSK9 transgene was highly expressed in kidney, LDLR proteins were suppressed to a lower extent in this tissue than in liver. Adrenal LDLR proteins were not regulated by elevated plasma PCSK9. hPCSK9 transgene expression and subsequent reduction of liver LDLR led to increases in plasma total cholesterol, LDL cholesterol, and ApoB, which were further increased by a high-fat, high-cholesterol diet. We also observed that the size distribution of hPCSK9 in transgenic mouse plasma was heterogeneous. In chow-fed mice, the majority of PCSK9 proteins were in free forms; however, feeding a high-fat, high-cholesterol diet resulted in a shift of hPCSK9 distribution toward larger complexes. PCSK9 distribution in human plasma also exhibited heterogeneity and individual variability in the percentage of PCSK9 in free form and in large complexes. We provide strong evidence to support that human PCSK9 proteins secreted from extrahepatic tissue are able to promote LDLR degradation in liver and increase plasma LDL. Our data also suggest that LDLR protein regulation by PCSK9 has tissue specificity, with liver being the most responsive tissue. PMID:19060325

  10. Alu sequence involvement in transcriptional insulation of the keratin 18 gene in transgenic mice.

    PubMed Central

    Thorey, I S; Ceceña, G; Reynolds, W; Oshima, R G

    1993-01-01

    The human keratin 18 (K18) gene is expressed in a variety of adult simple epithelial tissues, including liver, intestine, lung, and kidney, but is not normally found in skin, muscle, heart, spleen, or most of the brain. Transgenic animals derived from the cloned K18 gene express the transgene in appropriate tissues at levels directly proportional to the copy number and independently of the sites of integration. We have investigated in transgenic mice the dependence of K18 gene expression on the distal 5' and 3' flanking sequences and upon the RNA polymerase III promoter of an Alu repetitive DNA transcription unit immediately upstream of the K18 promoter. Integration site-independent expression of tandemly duplicated K18 transgenes requires the presence of either an 825-bp fragment of the 5' flanking sequence or the 3.5-kb 3' flanking sequence. Mutation of the RNA polymerase III promoter of the Alu element within the 825-bp fragment abolishes copy number-dependent expression in kidney but does not abolish integration site-independent expression when assayed in the absence of the 3' flanking sequence of the K18 gene. The characteristics of integration site-independent expression and copy number-dependent expression are separable. In addition, the formation of the chromatin state of the K18 gene, which likely restricts the tissue-specific expression of this gene, is not dependent upon the distal flanking sequences of the 10-kb K18 gene but rather may depend on internal regulatory regions of the gene. Images PMID:7692231

  11. Elimination of GD3 synthase improves memory and reduces amyloid-beta plaque load in transgenic mice.

    PubMed

    Bernardo, Alexandra; Harrison, Fiona E; McCord, Meghan; Zhao, Jiali; Bruchey, Aleksandra; Davies, Sean S; Jackson Roberts, L; Mathews, Paul M; Matsuoka, Yasuji; Ariga, Toshio; Yu, Robert K; Thompson, Rebecca; McDonald, Michael P

    2009-11-01

    Gangliosides have been shown to be necessary for beta-amyloid (Abeta) binding and aggregation. GD3 synthase (GD3S) is responsible for biosynthesis of the b- and c-series gangliosides, including two of the four major brain gangliosides. We examined Abeta-ganglioside interactions in neural tissue from mice lacking the gene coding for GD3S (St8sia1), and in a double-transgenic (APP/PSEN1) mouse model of Alzheimer's disease cross-bred with GD3S-/- mice. In primary neurons and astrocytes lacking GD3S, Abeta-induced cell death and Abeta aggregation were inhibited. Like GD3S-/- and APP/PSEN1 double-transgenic mice, APP/PSEN1/GD3S-/- "triple-mutant" mice are indistinguishable from wild-type mice on casual examination. APP/PSEN1 double-transgenics exhibit robust impairments on a number of reference-memory tasks. In contrast, APP/PSEN1/GD3S-/- triple-mutant mice performed as well as wild-type control and GD3S-/- mice. Consistent with the behavioral improvements, both aggregated and unaggregated Abeta and associated neuropathology were almost completely eliminated in triple-mutant mice. These results suggest that GD3 synthase may be a novel therapeutic target to combat the cognitive deficits, amyloid plaque formation, and neurodegeneration that afflict Alzheimer's patients. PMID:18258340

  12. Hepatic expression of mature transforming growth factor beta 1 in transgenic mice results in multiple tissue lesions.

    PubMed

    Sanderson, N; Factor, V; Nagy, P; Kopp, J; Kondaiah, P; Wakefield, L; Roberts, A B; Sporn, M B; Thorgeirsson, S S

    1995-03-28

    Aberrant expression of transforming growth factor beta 1 (TGF-beta 1) has been implicated in a number of disease processes, particularly those involving fibrotic and inflammatory lesions. To determine the in vivo effects of overexpression of TGF-beta 1 on the function and structure of hepatic as well as extrahepatic tissues, transgenic mice were generated containing a fusion gene (Alb/TGF-beta 1) consisting of modified porcine TGF-beta 1 cDNA under the control of the regulatory elements of the mouse albumin gene. Five transgenic lines were developed, all of which expressed the Alb/TGF-beta 1 transgene selectively in hepatocytes. The transgenic line 25 expressing the highest level of the transgene in the liver also had high (> 10-fold over control) plasma levels of TGF-beta 1. Hepatic fibrosis and apoptotic death of hepatocytes developed in all the transgenic lines but was more pronounced in line 25. The fibrotic process was characterized by deposition of collagen around individual hepatocytes and within the space of Disse in a radiating linear pattern. Several extrahepatic lesions developed in line 25, including glomerulonephritis and renal failure, arteritis and myocarditis, as well as atrophic changes in pancreas and testis. The results from this transgenic model strongly support the proposed etiological role for TGF-beta 1 in a variety of fibrotic and inflammatory disorders. The transgenic model may also provide an appropriate paradigm for testing therapeutic interventions aimed at neutralizing the detrimental effects of this important cytokine. PMID:7708687

  13. Apolipoprotein D Transgenic Mice Develop Hepatic Steatosis through Activation of PPARγ and Fatty Acid Uptake.

    PubMed

    Labrie, Marilyne; Lalonde, Simon; Najyb, Ouafa; Thiery, Maxime; Daneault, Caroline; Des Rosiers, Chrisitne; Rassart, Eric; Mounier, Catherine

    2015-01-01

    Transgenic mice (Tg) overexpressing human apolipoprotein D (H-apoD) in the brain are resistant to neurodegeneration. Despite the use of a neuron-specific promoter to generate the Tg mice, they expressed significant levels of H-apoD in both plasma and liver and they slowly develop hepatic steatosis and insulin resistance. We show here that hepatic PPARγ expression in Tg mice is increased by 2-fold compared to wild type (WT) mice. Consequently, PPARγ target genes Plin2 and Cide A/C are overexpressed, leading to increased lipid droplets formation. Expression of the fatty acid transporter CD36, another PPARgamma target, is also increased in Tg mice associated with elevated fatty acid uptake as measured in primary hepatocytes. Elevated expression of AMPK in the liver of Tg leads to phosphorylation of acetyl CoA carboxylase, indicating a decreased activity of the enzyme. Fatty acid synthase expression is also induced but the hepatic lipogenesis measured in vivo is not significantly different between WT and Tg mice. In addition, expression of carnitine palmitoyl transferase 1, the rate-limiting enzyme of beta-oxidation, is slightly upregulated. Finally, we show that overexpressing H-apoD in HepG2 cells in presence of arachidonic acid (AA), the main apoD ligand, increases the transcriptional activity of PPARγ. Supporting the role of apoD in AA transport, we observed enrichment in hepatic AA and a decrease in plasmatic AA concentration. Taken together, our results demonstrate that the hepatic steatosis observed in apoD Tg mice is a consequence of increased PPARγ transcriptional activity by AA leading to increased fatty acid uptake by the liver. PMID:26083030

  14. Transgenic mice expressing the Peripherin-EGFP genomic reporter display intrinsic peripheral nervous system fluorescence.

    PubMed

    McLenachan, Samuel; Goldshmit, Yona; Fowler, Kerry J; Voullaire, Lucille; Holloway, Timothy P; Turnley, Ann M; Ioannou, Panos A; Sarsero, Joseph P

    2008-12-01

    The development of homologous recombination methods for the precise modification of bacterial artificial chromosomes has allowed the introduction of disease causing mutations or fluorescent reporter genes into human loci for functional studies. We have introduced the EGFP gene into the human PRPH-1 locus to create the Peripherin-EGFP (hPRPH1-G) genomic reporter construct. The hPRPH1-G reporter was used to create transgenic mice with an intrinsically fluorescent peripheral nervous system (PNS). During development, hPRPH1-G expression was concomitant with the acquisition of neuronal cell fate and growing axons could be observed in whole embryo mounts. In the adult, sensory neurons were labeled in both the PNS and central nervous system, while motor neurons in the spinal cord had more limited expression. The fusion protein labeled long neuronal processes, highlighting the peripheral circuitry of hPRPH1-G transgenic mice to provide a useful resource for a range of neurobiological applications. PMID:18709437

  15. Inhibition of Intimal Hyperplasia in Transgenic Mice Conditionally Expressing the Chemokine-Binding Protein M3

    PubMed Central

    Pyo, Robert; Jensen, Kristian K.; Wiekowski, Maria T.; Manfra, Denise; Alcami, Antonio; Taubman, Mark B.; Lira, Sergio A.

    2004-01-01

    Chemokines have been implicated in the pathogenesis of a wide variety of diseases. This report describes the generation of transgenic mice that conditionally express M3, a herpesvirus protein that binds and inhibits chemokines. In response to doxycycline, M3 expression was induced in a variety of tissues and M3 was detectable in the blood by Western blotting. No gross or histological abnormalities were seen in mice expressing M3. To determine whether M3 expression could modify a significant pathophysiological response, we examined its effect on the development of intimal hyperplasia in response to femoral arterial injury. Intimal hyperplasia is thought to play a critical role in the development of restenosis after percutaneous transluminal coronary angioplasty and in the progression of atherosclerosis. Induction of M3 expression resulted in a 67% reduction in intimal area and a 68% reduction in intimal/medial ratio after femoral artery injury. These data support a role for chemokines in regulating intimal hyperplasia and suggest that M3 may be effective in attenuating this process. This transgenic mouse model should be a valuable tool for investigating the role of chemokines in a variety of pathological states. PMID:15161661

  16. Inhibition of intimal hyperplasia in transgenic mice conditionally expressing the chemokine-binding protein M3.

    PubMed

    Pyo, Robert; Jensen, Kristian K; Wiekowski, Maria T; Manfra, Denise; Alcami, Antonio; Taubman, Mark B; Lira, Sergio A

    2004-06-01

    Chemokines have been implicated in the pathogenesis of a wide variety of diseases. This report describes the generation of transgenic mice that conditionally express M3, a herpesvirus protein that binds and inhibits chemokines. In response to doxycycline, M3 expression was induced in a variety of tissues and M3 was detectable in the blood by Western blotting. No gross or histological abnormalities were seen in mice expressing M3. To determine whether M3 expression could modify a significant pathophysiological response, we examined its effect on the development of intimal hyperplasia in response to femoral arterial injury. Intimal hyperplasia is thought to play a critical role in the development of restenosis after percutaneous transluminal coronary angioplasty and in the progression of atherosclerosis. Induction of M3 expression resulted in a 67% reduction in intimal area and a 68% reduction in intimal/medial ratio after femoral artery injury. These data support a role for chemokines in regulating intimal hyperplasia and suggest that M3 may be effective in attenuating this process. This transgenic mouse model should be a valuable tool for investigating the role of chemokines in a variety of pathological states. PMID:15161661

  17. Transgenic kallikrein 5 mice reproduce major cutaneous and systemic hallmarks of Netherton syndrome.

    PubMed

    Furio, Laetitia; de Veer, Simon; Jaillet, Madeleine; Briot, Anais; Robin, Aurelie; Deraison, Celine; Hovnanian, Alain

    2014-03-10

    Netherton syndrome (NS) is a severe genetic skin disease in which absence of a key protease inhibitor causes congenital exfoliative erythroderma, eczematous-like lesions, and atopic manifestations. Several proteases are overactive in NS, including kallikrein-related peptidase (KLK) 5, KLK7, and elastase-2 (ELA2), which are suggested to be part of a proteolytic cascade initiated by KLK5. To address the role of KLK5 in NS, we have generated a new transgenic murine model expressing human KLK5 in the granular layer of the epidermis (Tg-KLK5). Transgene expression resulted in increased proteolytic activity attributable to KLK5 and its downstream targets KLK7, KLK14, and ELA2. Tg-KLK5 mice developed an exfoliative erythroderma with scaling, growth delay, and hair abnormalities. The skin barrier was defective and the stratum corneum was detached through desmosomal cleavage. Importantly, Tg-KLK5 mice displayed cutaneous and systemic hallmarks of severe inflammation and allergy with pruritus. The skin showed enhanced expression of inflammatory cytokines and chemokines, infiltration of immune cells, and markers of Th2/Th17/Th22 T cell responses. Moreover, serum IgE and Tslp levels were elevated. Our study identifies KLK5 as an important contributor to the NS proteolytic cascade and provides a new and viable model for the evaluation of future targeted therapies for NS or related diseases such as atopic dermatitis. PMID:24534191

  18. Morpholino treatment improves muscle function and pathology of Pitx1 transgenic mice.

    PubMed

    Pandey, Sachchida Nand; Lee, Yi-Chien; Yokota, Toshifumi; Chen, Yi-Wen

    2014-02-01

    Paired-like homeodomain transcription factor 1 (PITX1) was proposed to be part of the disease mechanisms of facioscapulohumeral muscular dystrophy (FSHD). We generated a tet-repressible muscle-specific Pitx1 transgenic mouse model which develops phenotypes of muscular dystrophy after the PITX1 expression is induced. In this study, we attempted to block the translation of PITX1 protein using morpholinos. Three groups of the transgenic mice received intravenous injections of phosphorodiamidate morpholino oligomers (PMO) (100 mg/kg), octaguanidinium dendrimer-conjugated morpholino (vivo-morpholino) (10 mg/kg), or phosphate-buffered saline (PBS) after the PITX1 expression was induced. Immunoblotting data showed that PITX1 expression in the triceps and quadriceps was significantly reduced 70% and 63% by the vivo-morpholino treatment, respectively. Muscle pathology of the mice treated with the vivo-morpholino was improved by showing 44% fewer angular-shaped atrophic myofibers. Muscle function determined by grip strength was significantly improved by the vivo-morpholino treatment. The study showed that systemic delivery of the vivo-morpholino reduced the PITX1 expression and improved the muscle phenotypes. Aberrant expression of DUX4 from the last unit of the D4Z4 array has been proposed to be the cause of FSHD. The findings of this study suggest that the same principle may be applied to suppress the aberrantly expressed DUX4 in FSHD. PMID:24232919

  19. Olfactory marker protein gene: its structure and olfactory neuron-specific expression in transgenic mice.

    PubMed Central

    Danciger, E; Mettling, C; Vidal, M; Morris, R; Margolis, F

    1989-01-01

    Olfactory marker protein (OMP) genomic clones were isolated from a Charon 4A phage lambda rat genomic library. A 16.5-kilobase (kb) fragment of the rat genome containing the gene was isolated and characterized. Sequence analysis of the gene showed the absence of introns and the lack of CAAT and TATA boxes in the 5' flanking region. The transcription initiation site was mapped, and two sites 55 and 58 base pairs upstream of the ATG were observed. The 5' flanking region is rich in G+C residues and contains a G+C-rich motif as well as direct and inverted repeats. Functional OMP regulatory sequences were demonstrated in transgenic mice. An 11-kb chimeric gene was constructed in which the coding region for OMP was replaced with that for Thy-1.1. In Thy-1.2 mice carrying this transgene, Thy-1.1 was expressed solely by olfactory receptor neurons and their axons and terminals in the olfactory bulb. Images PMID:2701951

  20. Transgenic reporter mice with promoter region of murine LRAT specifically marks lens and meiosis spermatocytes.

    PubMed

    Prukova, D; Ileninova, Z; Antosova, B; Kasparek, P; Gregor, M; Sedlacek, R

    2015-01-01

    Lecithin:retinol acyltransferase (LRAT) is the major enzyme responsible for retinol esterification in the mammalian body. LRAT exhibits specific activity in the cells with active retinol metabolism where it converts retinols into retinyl esters, which represents the major storage form of retinol. Besides hepatic stellate cells in the liver, LRAT appears to have a key physiologic role in several other tissues. In this study, we generated a transgenic reporter mouse expressing green fluorescence protein (EGFP) under the control of region containing -1166 bps from promoter upstream from the putative transcriptional start site and 262 bps downstream of this start. Transgenic reporter mice exhibited specific expression in eyes and testes. In eyes, expression of EGFP-reporter is found in lens and lens epithelium and fibers from embryo to adulthood. In testes, LRAT-EGFP reporter is expressed both in Sertoli and in spermatocytes marking initiation of spermatogenesis in prepubertal mice. Our data show that the examined LRAT regulatory region is sufficient to achieve strong and selective expression in the eye and testes but not in liver and other organs. PMID:25317684

  1. Morpholino Treatment Improves Muscle Function and Pathology of Pitx1 Transgenic Mice

    PubMed Central

    Pandey, Sachchida Nand; Lee, Yi-Chien; Yokota, Toshifumi; Chen, Yi-Wen

    2014-01-01

    Paired-like homeodomain transcription factor 1 (PITX1) was proposed to be part of the disease mechanisms of facioscapulohumeral muscular dystrophy (FSHD). We generated a tet-repressible muscle-specific Pitx1 transgenic mouse model which develops phenotypes of muscular dystrophy after the PITX1 expression is induced. In this study, we attempted to block the translation of PITX1 protein using morpholinos. Three groups of the transgenic mice received intravenous injections of phosphorodiamidate morpholino oligomers (PMO) (100 mg/kg), octaguanidinium dendrimer-conjugated morpholino (vivo-morpholino) (10 mg/kg), or phosphate-buffered saline (PBS) after the PITX1 expression was induced. Immunoblotting data showed that PITX1 expression in the triceps and quadriceps was significantly reduced 70% and 63% by the vivo-morpholino treatment, respectively. Muscle pathology of the mice treated with the vivo-morpholino was improved by showing 44% fewer angular-shaped atrophic myofibers. Muscle function determined by grip strength was significantly improved by the vivo-morpholino treatment. The study showed that systemic delivery of the vivo-morpholino reduced the PITX1 expression and improved the muscle phenotypes. Aberrant expression of DUX4 from the last unit of the D4Z4 array has been proposed to be the cause of FSHD. The findings of this study suggest that the same principle may be applied to suppress the aberrantly expressed DUX4 in FSHD. PMID:24232919

  2. Introduction of yeast artificial chromosomes containing mutant human amyloid precursor protein genes into transgenic mice

    SciTech Connect

    Call, L.M.; Lamb, B.T.; Boese, K.F.

    1994-09-01

    Several hypothetical mechanisms have been proposed for the generation and deposition of the amyloid beta (A{beta}) peptide in Alzheimer`s disease (AD). These include overexpression of the amyloid precursor protein (APP) gene, as suggested by Down Syndrome (DS, trisomy 21), and mutation of APP, as suggested by mutations associated with the presence of disease/amyloid deposition in some cases of familial AD (FAD). Although numerous in vitro studies have lead to certain insights into the molecular basis for amyloid deposition, the mechanisms(s) of amyloidogenesis in vivo remains poorly defined. To examine the effect of FAD mutations on amyloidogenesis in an animal model, we have focused on producing APP YAC transgenic mice containing the human APP gene with FAD mutations. These APP YAC transgenics are being produced by introduction of a 650 kb APP YAC through lipid-mediated transfection of ES cells. This strategy has two principal advantages: the APP genomic sequences contain transcriptional regulatory elements required for proper spatial and temporal expression and contain appropriate splice donor and acceptor sites needed to generate the entire spectrum of alternatively spliced APP transcripts. As a first step, we cloned the genomic regions surrounding APP exons 16 and 17 from an APP YAC sublibrary. Both the Swedish and the 717 mutations were then introduced into exons 16 and 17, respectively, by PCR mutagenesis, and subsequently transferred into the 650 kb APP YAC by a two step gene replacement in yeast. The mutant YACs have been introduced into ES cells, and we have determined that these cells are expressing human mutant APP mRNA and protein. These cells are being used to generate transgenic mice. This paradigm should provide the appropriate test of whether a mutant APP gene is capable of producing AD-like pathology in a mouse model.

  3. SHP2E76K mutant promotes lung tumorigenesis in transgenic mice

    PubMed Central

    Schneeberger, Valentina E.; Luetteke, Noreen; Ren, Yuan; Berns, Hartmut; Chen, Liwei; Foroutan, Parastou; Martinez, Gary V.; Haura, Eric B.; Chen, Jiandong; Coppola, Domenico; Wu, Jie

    2014-01-01

    Lung cancer is a major disease carrying heterogeneous molecular lesions and many of them remain to be analyzed functionally in vivo. Gain-of-function (GOF) SHP2 (PTPN11) mutations have been found in various types of human cancer, including lung cancer. However, the role of activating SHP2 mutants in lung cancer has not been established. We generated transgenic mice containing a doxycycline (Dox)-inducible activating SHP2 mutant (tetO-SHP2E76K) and analyzed the role of SHP2E76K in lung tumorigenesis in the Clara cell secretory protein (CCSP)-reverse tetracycline transactivator (rtTA)/tetO-SHP2E76K bitransgenic mice. SHP2E76K activated Erk1/Erk2 (Erk1/2) and Src, and upregulated c-Myc and Mdm2 in the lungs of bitransgenic mice. Atypical adenomatous hyperplasia and small adenomas were observed in CCSP-rtTA/tetO-SHP2E76K bitransgenic mice induced with Dox for 2–6 months and progressed to larger adenoma and adenocarcinoma by 9 months. Dox withdrawal from bitransgenic mice bearing magnetic resonance imaging-detectable lung tumors resulted in tumor regression. These results show that the activating SHP2 mutant promotes lung tumorigenesis and that the SHP2 mutant is required for tumor maintenance in this mouse model of non-small cell lung cancer. SHP2E76K was associated with Gab1 in the lung of transgenic mice. Elevated pGab1 was observed in the lung of Dox-induced CCSP-rtTA/tetO-SHP2E76K mice and in cell lines expressing SHP2E76K, indicating that the activating SHP2 mutant autoregulates tyrosine phosphorylation of its own docking protein. Gab1 tyrosine phosphorylation is sensitive to inhibition by the Src inhibitor dasatinib in GOF SHP2-mutant-expressing cells, suggesting that Src family kinases are involved in SHP2 mutant-induced Gab1 tyrosine phosphorylation. PMID:24480804

  4. SHP2E76K mutant promotes lung tumorigenesis in transgenic mice.

    PubMed

    Schneeberger, Valentina E; Luetteke, Noreen; Ren, Yuan; Berns, Hartmut; Chen, Liwei; Foroutan, Parastou; Martinez, Gary V; Haura, Eric B; Chen, Jiandong; Coppola, Domenico; Wu, Jie

    2014-08-01

    Lung cancer is a major disease carrying heterogeneous molecular lesions and many of them remain to be analyzed functionally in vivo. Gain-of-function (GOF) SHP2 (PTPN11) mutations have been found in various types of human cancer, including lung cancer. However, the role of activating SHP2 mutants in lung cancer has not been established. We generated transgenic mice containing a doxycycline (Dox)-inducible activating SHP2 mutant (tetO-SHP2(E76K)) and analyzed the role of SHP2(E76K) in lung tumorigenesis in the Clara cell secretory protein (CCSP)-reverse tetracycline transactivator (rtTA)/tetO-SHP2(E76K) bitransgenic mice. SHP2(E76K) activated Erk1/Erk2 (Erk1/2) and Src, and upregulated c-Myc and Mdm2 in the lungs of bitransgenic mice. Atypical adenomatous hyperplasia and small adenomas were observed in CCSP-rtTA/tetO-SHP2(E76K) bitransgenic mice induced with Dox for 2-6 months and progressed to larger adenoma and adenocarcinoma by 9 months. Dox withdrawal from bitransgenic mice bearing magnetic resonance imaging-detectable lung tumors resulted in tumor regression. These results show that the activating SHP2 mutant promotes lung tumorigenesis and that the SHP2 mutant is required for tumor maintenance in this mouse model of non-small cell lung cancer. SHP2(E76K) was associated with Gab1 in the lung of transgenic mice. Elevated pGab1 was observed in the lung of Dox-induced CCSP-rtTA/tetO-SHP2(E76K) mice and in cell lines expressing SHP2(E76K), indicating that the activating SHP2 mutant autoregulates tyrosine phosphorylation of its own docking protein. Gab1 tyrosine phosphorylation is sensitive to inhibition by the Src inhibitor dasatinib in GOF SHP2-mutant-expressing cells, suggesting that Src family kinases are involved in SHP2 mutant-induced Gab1 tyrosine phosphorylation. PMID:24480804

  5. High incidence of lung, bone, and lymphoid tumors in transgenic mice overexpressing mutant alleles of the p53 oncogene.

    PubMed Central

    Lavigueur, A; Maltby, V; Mock, D; Rossant, J; Pawson, T; Bernstein, A

    1989-01-01

    We have investigated the role of the p53 gene in oncogenesis in vivo by generating transgenic mice carrying murine p53 genomic fragments isolated from a mouse Friend erythroleukemia cell line or BALB/c mouse liver DNA. Elevated levels of p53 mRNA were detected in several tissues of two transgenic lines tested. Increased levels of p53 protein were also detected in most of the tissues analyzed by Western blotting (immunoblotting). Because both transgenes encoded p53 proteins that were antigenically distinct from wild-type p53, it was possible to demonstrate that overexpression of the p53 protein was mostly, if not entirely, due to the expression of the transgenes. Neoplasms developed in 20% of the transgenic mice, with a high incidence of lung adenocarcinomas, osteosarcomas, and lymphomas. Tissues such as ovaries that expressed the transgene at high levels were not at higher risk of malignant transformation than tissues expressing p53 protein at much lower levels. The long latent period and low penetrance suggest that overexpression of p53 alone is not sufficient to induce malignancies and that additional events are required. These observations provide direct evidence that mutant alleles of the p53 oncogene have oncogenic potential in vivo and that different cell types show intrinsic differences in susceptibility to malignant transformation by p53. Since recent data suggest that p53 may be a recessive oncogene, it is possible that the elevated tumor incidence results from functional inactivation of endogenous p53 by overexpression of the mutant transgene. The high incidence of lung and bone tumors suggests that p53 transgenic mice may provide a useful model to investigate the molecular events that underlie these malignancies in humans. Images PMID:2476668

  6. Group III secreted phospholipase A2 transgenic mice spontaneously develop inflammation

    PubMed Central

    Sato, Hiroyasu; Taketomi, Yoshitaka; Isogai, Yuki; Masuda, Seiko; Kobayashi, Tetsuyuki; Yamamoto, Kei; Murakami, Makoto

    2009-01-01

    PLA2 (phospholipase A2) group III is an atypical sPLA2 (secretory PLA2) that is homologous with bee venom PLA2 rather than with other mammalian sPLA2s. In the present paper, we show that endogenous group III sPLA2 (PLA2G3) is expressed in mouse skin and that Tg (transgenic) mice overexpressing human PLA2G3 spontaneously develop skin inflammation. Pla2g3-Tg mice over 9 months of age frequently developed dermatitis with hyperkeratosis, acanthosis, parakeratosis, erosion, ulcer and sebaceous gland hyperplasia. The dermatitis was accompanied by infiltration of neutrophils and macrophages and by elevated levels of pro-inflammatory cytokines, chemokines and prostaglandin E2. In addition, Pla2g3-Tg mice had increased lymph aggregates and mucus in the airway, lymphocytic sialadenitis, hepatic extramedullary haemopoiesis, splenomegaly with increased populations of granulocytes and monocytes/macrophages, and increased serum IgG1. Collectively, these observations provide the first demonstration of spontaneous development of inflammation in mice with Tg overexpression of mammalian sPLA2. PMID:19371233

  7. Linalool reverses neuropathological and behavioral impairments in old triple transgenic Alzheimer's mice.

    PubMed

    Sabogal-Guáqueta, Angélica Maria; Osorio, Edison; Cardona-Gómez, Gloria Patricia

    2016-03-01

    Alzheimer's disease (AD) is an age-related progressive neurodegenerative disorder. Several types of treatments have been tested to block or delay the onset of the disease, but none have been completely successful. Diet, lifestyle and natural products are currently the main scientific focuses. Here, we evaluate the effects of oral administration of the monoterpene linalool (25 mg/kg), every 48 h for 3 months, on aged (21-24 months old) mice with a triple transgenic model of AD (3xTg-AD) mice. Linalool-treated 3xTg-AD mice showed improved learning and spatial memory and greater risk assessment behavior during the elevated plus maze. Hippocampi and amygdalae from linalool-treated 3xTg-AD mice exhibited a significant reduction in extracellular β-amyloidosis, tauopathy, astrogliosis and microgliosis as well as a significant reduction in the levels of the pro-inflammatory markers p38 MAPK, NOS2, COX2 and IL-1β. Together, our findings suggest that linalool reverses the histopathological hallmarks of AD and restores cognitive and emotional functions via an anti-inflammatory effect. Thus, linalool may be an AD prevention candidate for preclinical studies. PMID:26549854

  8. Organ-targeted mutagenicity of nitrofurantoin in Big Blue transgenic mice.

    PubMed

    Quillardet, Philippe; Arrault, Xavier; Michel, Valérie; Touati, Eliette

    2006-09-01

    Nitrofurans are widely used in human medicine, as nitrofurantoin and nifuroxazide, still prescribed for long-term antimicrobial prophylaxis of urinary tract and gastrointestinal infection in humans respectively. Recent experiments in mammals, as well as reports mentioning toxic effects in humans associated with a long-term use, specially in the case of nitrofurantoin, raised the need for reevaluating their genotoxicity. The objective of this study was to determine whether these two compounds induce a mutagenic effect in the Big Blue transgenic mouse mutation assay. Mice were orally treated either with nitrofurantoin or nifuroxazide for five consecutive days and sacrificed 3 weeks later. In order to optimize the genotoxic response, the doses used for each compound were 25-fold higher as the posology in humans. They corresponded to 50% of the highest doses tolerated by mice. The mutant frequency was determined from kidney, lung, bladder, caecum, colon, small intestine, spleen and stomach. A weak mutagenic response of nitrofurantoin-treated mice specifically in the kidney was observed. As in the case of other nitrofuran compounds, the mutation spectra determined from treated samples exhibited slightly more GC-->TA transversions as compared with untreated conditions. These data are relevant to the targeted action of nitrofurantoin as a urinary antimicrobial agent. No significant increase of mutants was detected in the case of nifuroxazide-treated mice whatever the organs analysed. PMID:16895946

  9. Long-term dantrolene treatment reduced intraneuronal amyloid in aged Alzheimer triple transgenic mice.

    PubMed

    Wu, Zhen; Yang, Bin; Liu, Chunxia; Liang, Ge; Liu, Weixia; Pickup, Stephen; Meng, Qingcheng; Tian, Yuke; Li, Shitong; Eckenhoff, Maryellen F; Wei, Huafeng

    2015-01-01

    In this study, we investigated the long-term treatment of dantrolene on amyloid and tau neuropathology, brain volume, and cognitive function in aged triple transgenic Alzheimer (3xTg-AD) mice. Fifteen-month old 3xTg-AD mice and wild-type controls were treated with oral dantrolene (5 mg/kg) or vehicle control twice a week for 6 months. Learning and memory were examined using the Morris Water Maze at 21 and 22 months of age. After the behavioral testing, hippocampal and cortical brain volumes were calculated with magnetic resonance imaging and motor function was evaluated using the rotorod. The amyloid burden and tau neurofibrillary tangles in the hippocampus were determined using immunohistochemistry. We found that dantrolene significantly decreased the intraneuronal amyloid accumulation by as much as 76% compared with its corresponding vehicle control, together with a trend to reduce phosphorylated tau in the hippocampus. No significant differences could be detected in hippocampal or cortical brain volume, motor function or cognition among all experimental groups, indicating that the mice were still presymptomatic for Alzheimer disease. Thus, presymptomatic and long-term dantrolene treatment significantly decreased the intraneuronal amyloid burden in aged 3xTg-AD mice before significant changes in brain volume, or cognition. PMID:25650693

  10. Transgenic mice carrying the human poliovirus receptor: new animal models for study of poliovirus neurovirulence.

    PubMed Central

    Horie, H; Koike, S; Kurata, T; Sato-Yoshida, Y; Ise, I; Ota, Y; Abe, S; Hioki, K; Kato, H; Taya, C

    1994-01-01

    Recombinant viruses between the virulent Mahoney and attenuated Sabin 1 strains of poliovirus type 1 were subjected to neurovirulence tests using a transgenic (Tg) mouse line, ICR-PVRTg1, that carried the human poliovirus receptor gene. The Tg mice were inoculated intracerebrally with these recombinant viruses and observed for clinical signs, histopathological lesions, and viral antigens as parameters of neurovirulence of the viruses. These parameters observed in the Tg mice were different for different inoculated viruses. Dose-dependent incidences of paralysis and of death were observed in the Tg mice inoculated with any viruses used. This indicates that values of 50% lethal dose are useful to score a wide range of neurovirulence of poliovirus. The neurovirulence of individual viruses estimated by the Tg mouse model had a strong correlation with those estimated by monkey model. Consequently, the mouse tests identified the neurovirulence determinants on the genome of poliovirus that had been identified by monkey tests. In addition, the mouse tests revealed new neurovirulence determinants, that is, different nucleotides between the two strains at positions 189 and 21 and/or 935 in the 5'-proximal 1,122 nucleotides. The Tg mice used in this study may be suitable for replacing monkeys for investigating poliovirus neurovirulence. Images PMID:8289371

  11. Whole-Body In Vivo Monitoring of Inflammatory Diseases Exploiting Human Interleukin 6-Luciferase Transgenic Mice.

    PubMed

    Hayashi, Makiko; Takai, Jun; Yu, Lei; Motohashi, Hozumi; Moriguchi, Takashi; Yamamoto, Masayuki

    2015-10-01

    Chronic inflammation underlies the pathological progression of various diseases, and thus many efforts have been made to quantitatively evaluate the inflammatory status of the diseases. In this study, we generated a highly sensitive inflammation-monitoring mouse system using a bacterial artificial chromosome (BAC) clone containing extended flanking sequences of the human interleukin 6 gene (hIL6) locus, in which the luciferase (Luc) reporter gene is integrated (hIL6-BAC-Luc). We successfully monitored lipopolysaccharide-induced systemic inflammation in various tissues of the hIL6-BAC-Luc mice using an in vivo bioluminescence imaging system. When two chronic inflammatory disease models, i.e., a genetic model of atopic dermatitis and a model of experimental autoimmune encephalomyelitis (EAE), were applied to the hIL6-BAC-Luc mice, luciferase bioluminescence was specifically detected in the atopic skin lesion and central nervous system, respectively. Moreover, the Luc activities correlated well with the disease severity. Nrf2 is a master transcription factor that regulates antioxidative and detoxification enzyme genes. Upon EAE induction, the Nrf2-deficient mice crossed with the hIL6-BAC-Luc mice exhibited enhanced neurological symptoms concomitantly with robust luciferase luminescence in the neuronal tissue. Thus, whole-body in vivo monitoring using the hIL6-BAC-Luc transgenic system (WIM-6 system) provides a new and powerful diagnostic tool for real-time in vivo monitoring of inflammatory status in multiple different disease models. PMID:26283726

  12. Pituitary Changes in Prop1 Transgenic Mice: Hormone Producing Tumors and Signet-ring Type Gonadotropes

    PubMed Central

    Egashira, Noboru; Minematsu, Takeo; Miyai, Syunsuke; Takekoshi, Susumu; Camper, Sally A.; Osamura, Robert Y.

    2008-01-01

    Prophet of Pit-1 (Prop1) is an early transcription factor that delays the appearance of gonadotropin in the developing pituitaries. Prop1 transgenic (Tg) mice have been shown to generate pituitary tumors that either produce TSH or are non-hormone producing. In our series of Prop1 Tg mice, only 5 out of 9 female mice produced pituitary adenomas, and the adenomas were only GH, PRL, GH and PRL, PRL and gonadotropin or TSH producing. The pituitary cells that surrounded these adenomas showed hyperplasia of the corresponding hormone producing cells; i.e. the GH cells were increased in the pituitary that contained GH producing adenoma. In addition, although the adenomas lacked the expression of Prop1, the non-neoplastic pituitary cells showed expression of Prop1. The Prop1 Tg mice also showed vacuolated cells with eccentric nuclei, which are characteristic of “signet-ring hypertrophic cells”. Using immunohistochemistry, these signet ring hypertrophic cells were found to be positive for gonadotropin. Taken together, our results suggest a (1) tumorigenic effect of Prop1 in the pituitaries, and (2) causative effects of signet ring-type gonadotropes. PMID:18636109

  13. A deregulated immune response to gliadin causes a decreased villus height in DQ8 transgenic mice.

    PubMed

    D'Arienzo, Rossana; Stefanile, Rosita; Maurano, Francesco; Luongo, Diomira; Bergamo, Paolo; Mazzarella, Giuseppe; Troncone, Riccardo; Auricchio, Salvatore; David, Chella; Rossi, Mauro

    2009-12-01

    Celiac disease (CD) is an enteropathy triggered by gluten and mediated by CD4+ T cells. A complete understanding of CD immunopathogenesis has been hindered due to the lack of adequate in vivo models. Here, we explored the effect of the inhibition of COX by indomethacin in wheat gliadin-sensitized transgenic mice expressing the HLA-DQ8 heterodimer, a molecule associated with CD. Treated mice showed a gliadin-specific immune response with a significant reduction of villus height, not linked to crypt hyperplasia and to expansion of intraepithelial T cells. Notably, treated mice showed increased numbers of CD25+ and apoptotic cells in the lamina propria, whereas high basal levels of IFN-gamma secretion, along with a reduced gliadin-specific IL-2 expression were detected in MLN. Biochemical assessment of the lesion revealed increased mRNA of Lamb3 and Adamts2, encoding for ECM proteins, and enhanced activities of metalloproteinases MMP1, 2 and 7. We conclude that an intestinal sensitivity to gliadin, in connection with COX inhibition, caused a decreased villus height in DQ8 tg mice. The lesion was induced by a deregulated mucosal cell immunity to gliadin, thus triggering activation of a specific ECM protein pathway responsible for lamina propria remodeling. PMID:19795413

  14. Tcf4 transgenic female mice display delayed adaptation in an auditory latent inhibition paradigm.

    PubMed

    Brzózka, M M; Rossner, M J; de Hoz, L

    2016-09-01

    Schizophrenia (SZ) is a severe mental disorder affecting about 1 % of the human population. Patients show severe deficits in cognitive processing often characterized by an improper filtering of environmental stimuli. Independent genome-wide association studies confirmed a number of risk variants for SZ including several associated with the gene encoding the transcription factor 4 (TCF4). TCF4 is widely expressed in the central nervous system of mice and humans and seems to be important for brain development. Transgenic mice overexpressing murine Tcf4 (Tcf4tg) in the adult brain display cognitive impairments and sensorimotor gating disturbances. To address the question of whether increased Tcf4 gene dosage may affect cognitive flexibility in an auditory associative task, we tested latent inhibition (LI) in female Tcf4tg mice. LI is a widely accepted translational endophenotype of SZ and results from a maladaptive delay in switching a response to a previously unconditioned stimulus when this becomes conditioned. Using an Audiobox, we pre-exposed Tcf4tg mice and their wild-type littermates to either a 3- or a 12-kHz tone before conditioning them to a 12-kHz tone. Tcf4tg animals pre-exposed to a 12-kHz tone showed significantly delayed conditioning when the previously unconditioned tone became associated with an air puff. These results support findings that associate TCF4 dysfunction with cognitive inflexibility and improper filtering of sensory stimuli observed in SZ patients. PMID:26404636

  15. C9orf72 BAC Transgenic Mice Display Typical Pathologic Features of ALS/FTD.

    PubMed

    O'Rourke, Jacqueline G; Bogdanik, Laurent; Muhammad, A K M G; Gendron, Tania F; Kim, Kevin J; Austin, Andrew; Cady, Janet; Liu, Elaine Y; Zarrow, Jonah; Grant, Sharday; Ho, Ritchie; Bell, Shaughn; Carmona, Sharon; Simpkinson, Megan; Lall, Deepti; Wu, Kathryn; Daughrity, Lillian; Dickson, Dennis W; Harms, Matthew B; Petrucelli, Leonard; Lee, Edward B; Lutz, Cathleen M; Baloh, Robert H

    2015-12-01

    Noncoding expansions of a hexanucleotide repeat (GGGGCC) in the C9orf72 gene are the most common cause of familial amyotrophic lateral sclerosis and frontotemporal dementia. Here we report transgenic mice carrying a bacterial artificial chromosome (BAC) containing the full human C9orf72 gene with either a normal allele (15 repeats) or disease-associated expansion (∼100-1,000 repeats; C9-BACexp). C9-BACexp mice displayed pathologic features seen in C9orf72 expansion patients, including widespread RNA foci and repeat-associated non-ATG (RAN) translated dipeptides, which were suppressed by antisense oligonucleotides targeting human C9orf72. Nucleolin distribution was altered, supporting that either C9orf72 transcripts or RAN dipeptides promote nucleolar dysfunction. Despite early and widespread production of RNA foci and RAN dipeptides in C9-BACexp mice, behavioral abnormalities and neurodegeneration were not observed even at advanced ages, supporting the hypothesis that RNA foci and RAN dipeptides occur presymptomatically and are not sufficient to drive neurodegeneration in mice at levels seen in patients. PMID:26637796

  16. Extraneural manifestations of prion infection in GPI-anchorless transgenic mice

    SciTech Connect

    Lee, Andrew M.; Paulsson, Johan F.; Cruite, Justin; Andaya, Abegail A.; Trifilo, Matthew J.; Oldstone, Michael B.A.

    2011-03-01

    Earlier studies indicated that transgenic (tg) mice engineered to express prion protein (PrP) lacking the glycophosphatidylinositol (GPI{sup -/-}) membrane anchor formed abnormal proteinase-resistant prion (PrPsc) amyloid deposits in their brains and hearts when infected with the RML strain of murine scrapie. In contrast, RML scrapie infection of normal mice with a GPI-anchored PrP did not deposit amyloid with PrPsc in the brain or the heart. Here we report that scrapie-infected GPI{sup -/-} PrP tg mice also deposit PrP and transmissible infectious material in the gut, kidneys, and islets of Langerhans. Similar to previously reported amyloid deposits in the brain and heart, amyloid deposits were found in the gut; however, no amyloid deposited in the islets. By high-resolution electron microscopy, we show PrP is located primarily in {alpha} cells and also {beta} cells. Islets contain abundant insulin and there is no abnormality in glucose metabolism in infected GPI{sup -/-} PrP tg mice.

  17. N17 Modifies Mutant Huntingtin Nuclear Pathogenesis and Severity of Disease in HD BAC Transgenic Mice

    PubMed Central

    Gu, Xiaofeng; Cantle, Jeffrey P.; Greiner, Erin R.; Lee, C.Y. Daniel; Barth, Albert M.; Gao, Fuying; Park, Chang Sin; Zhang, Zhiqiang; Sandoval-Miller, Susana; Zhang, Richard L.; Diamond, Marc; Mody, Istvan; Coppola, Giovanni

    2015-01-01

    SUMMARY The nucleus is a critical subcellular compartment for the pathogenesis of polyglutamine disorders, including Huntington’s disease (HD). Recent studies suggest the first 17-amino-acid domain (N17) of mutant Huntingtin (mHTT) mediates its nuclear exclusion in cultured cells. Here, we test whether N17 could be a molecular determinant of nuclear mHTT pathogenesis in vivo. BAC transgenic mice expressing mHTT lacking the N17 domain (BACHD-ΔN17) show dramatically accelerated mHTT pathology exclusively in the nucleus, which is associated with HD-like transcriptionopathy. Interestingly, BACHD-ΔN17 mice manifest more overt disease-like phenotypes than the original BACHD mice, including body weight loss, movement deficits, robust striatal neuronal loss, and neuroinflammation. Mechanistically, N17 is necessary for nuclear exclusion of small mHTT fragments that are part of nuclear pathology in HD. Together, our study suggests that N17 modifies nuclear pathogenesis and disease severity in HD mice by regulating subcellular localization of known nuclear pathogenic mHTT species. PMID:25661181

  18. Muscle-directed gene therapy for phenylketonuria (PKU): Development of transgenic mice with muscle-specific phenylalanine hydroxylase expression

    SciTech Connect

    Harding, C.O.; Messing, A.; Wolff, J.A.

    1994-09-01

    Phenylketonuria (PKU) is an attractive target for gene therapy because of shortcomings in current therapy including lifelong commitment to a difficult and expensive diet, persistent mild cognitive deficits in some children despite adequate dietary therapy, and maternal PKU syndrome. Phenylalanine hydroxylase (PAH) is normally expressed only in liver, but we propose to treat PKU by introducing the gene for PAH into muscle. In order to evaluate both the safety and efficacy of this approach, we have a developed a trangenic mouse which expresses PAH in both cardiac and skeletal muscle. The transgene includes promoter and enhancer sequences from the mouse muscle creatine kinase (MCK) gene fused to the mouse liver PAH cDNA. Mice which have inherited the transgene are healthy, active, and do not exhibit any signs of muscle weakness or wasting. Ectopic PAH expression in muscle is not detrimental to the health, neurologic function, or reproduction of the mice. Pah{sup enu2} hyperphenylalaninemic mice, a model of human PAH deficiency, bred to carry the transgene have substantial PAH expression in cardiac and skeletal muscle but none in liver. Muscle PAH expression alone does not complement the hyperphenylalaninemic phenotype of Pah{sup enu2} mice. However, administration of reduced tetrahydrobiopterin to transgenic Pah{sup enu2} mice is associated with a 25% mean decrease in serum phenylalanine levels. We predict that ectopic expression of PAH in muscle along with adequate muscle supplies of reduced biopterin cofactor will decrease hyperphenylalaninemia in PKU.

  19. Genome scan identifies a locus affecting gamma-globin expression in human beta-cluster YAC transgenic mice

    SciTech Connect

    Lin, S.D.; Cooper, P.; Fung, J.; Weier, H.U.G.; Rubin, E.M.

    2000-03-01

    Genetic factors affecting post-natal g-globin expression - a major modifier of the severity of both b-thalassemia and sickle cell anemia, have been difficult to study. This is especially so in mice, an organism lacking a globin gene with an expression pattern equivalent to that of human g-globin. To model the human b-cluster in mice, with the goal of screening for loci affecting human g-globin expression in vivo, we introduced a human b-globin cluster YAC transgene into the genome of FVB mice . The b-cluster contained a Greek hereditary persistence of fetal hemoglobin (HPFH) g allele resulting in postnatal expression of human g-globin in transgenic mice. The level of human g-globin for various F1 hybrids derived from crosses between the FVB transgenics and other inbred mouse strains was assessed. The g-globin level of the C3HeB/FVB transgenic mice was noted to be significantly elevated. To map genes affecting postnatal g-globin expression, a 20 centiMorgan (cM) genome scan of a C3HeB/F VB transgenics [prime] FVB backcross was performed, followed by high-resolution marker analysis of promising loci. From this analysis we mapped a locus within a 2.2 cM interval of mouse chromosome 1 at a LOD score of 4.2 that contributes 10.4% of variation in g-globin expression level. Combining transgenic modeling of the human b-globin gene cluster with quantitative trait analysis, we have identified and mapped a murine locus that impacts on human g-globin expression in vivo.

  20. Noninvasive Assessments of Optic Nerve Neurodegeneration in Transgenic Mice With Isolated Optic Neuritis

    PubMed Central

    Talla, Venu; Yang, Cui; Shaw, Gerry; Porciatti, Vittorio; Koilkonda, Rajeshwari D.; Guy, John

    2013-01-01

    Purpose. To determine if phosphorylated neurofilament heavy chain (pNF-H) released into the bloodstream and the pattern ERG are noninvasive indicators of neurodegeneration in experimental optic neuritis. Methods. Serum from Myelin oligodendrocyte glycoprotein (MOG)-specific T cell receptor–positive (TCR+) transgenic mice that develop isolated optic neuritis usually without any other characteristic lesions of inflammation or demyelination in the spinal cord and littermates negative for the transgene were assayed for the presence of serum phosphorylated neurofilament H (pNF-H). In vivo measurements of optic nerve and retinal ganglion cell injury were assessed by magnetic resonance imaging (MRI), optical coherence tomography (OCT), and pattern electroretinogram (PERG). Automated two dimensional fluorescence differential in-gel electrophoresis (2D-DIGE) of pooled optic nerve samples, light, and transmission electron micrographs were used to evaluate optic atrophy postmortem. Results. We found an almost 3-fold elevation in serum pNF-H levels in MOG+ mice relative to MOG-littermates (P = 0.02). 2D-DIGE revealed a 3-fold reduction in optic nerve neurofilaments. Visual function assessed by the PERG was reduced by one-quarter (P = 0.033) and latencies increased by 38% (P = 0.036). MOG+ mice with the lowest PERG amplitudes had optic nerve atrophy visualized by MRI. Optic nerve diameters were reduced by one-third (P = 0.0001) and axon counts reduced by more than two-thirds. Histopathology of the spinal cords was normal. Conclusions. Elevated serum pNF-H levels and the PERG are useful markers of neurodegeneration of the optic nerve in isolated experimental optic neuritis. Our findings suggest that elevations of this axonal protein in patients with optic neuritis who had a poor visual outcome are likely also due to demise of optic nerve axons. PMID:23722393

  1. Studies on the correlation with olfactory dysfunction in a transgenic mice model of Alzheimer's disease

    NASA Astrophysics Data System (ADS)

    Rasheed, Ameer; Lee, Ji Hye; Suh, Yoo-Hun; Moon, Cheil

    2013-05-01

    Alzheimer's disease (AD) is a progressively debilitating neurodegenerative disorder characterized by the presence of proteinaceous deposits in the brain. AD often results in olfactory dysfunction and impaired olfactory perceptual acuity may be a potential biomarker for early diagnosis of AD. Until recently, there is no Alzheimer's nanoscope or any other high-end microscope developed to be capable of seeing buried feature of AD clearly. Modern neuroimaging techniques are more effective only after the occurrence of cognitive impairment. Therefore, early detection of Alzheimer's disease is critical in developing effective treatment of AD. H and E (Haematoxyline and Eosin) staining is performed for examining gross morphological changes, while TUNEL (transferase (TdT)-mediated dUTP nick end labeling) staining for monitoring neuronal death in the olfactory epithelium (OE). Furthermore, immunohistochemistry and western blot are performed to examine β-amyloid protein expression. AD model animals were Tg2576 (transgenic mice that overexpress a mutated form of the Aβ precursor protein), and 6 month (before onset of AD symptoms) and 14 month (after onset of AD symptoms) old WT (wild type) and transgenic mice were compared in their olfactory system. We found that in OE of Tg2576 mice, thickness and total number of cells were decreased, while the numbers of TUNEL-positive neurons, caspase-3 activation were significantly increased compared with age-matched WT. Our results demonstrate that the olfactory system may get deteriorated before onset of AD symptoms. Our findings imply that an olfactory biopsy could be served as an early and relatively simple diagnostic tool for potential AD patients.

  2. Characterization of Fam20C expression in odontogenesis and osteogenesis using transgenic mice

    PubMed Central

    Du, Er-Xia; Wang, Xiao-Fang; Yang, Wu-Chen; Kaback, Deborah; Yee, Siu-Pok; Qin, Chun-Lin; George, Anne; Hao, Jian-Jun

    2015-01-01

    Our previous studies have demonstrated that Fam20C promotes differentiation and mineralization of odontoblasts, ameloblasts, osteoblasts and osteocytes during tooth and bone development. Ablation of the Fam20C gene inhibits bone and tooth growth by increasing fibroblast growth factor 23 in serum and causing hypophosphatemia in conditional knockout mice. However, control and regulation of the expression of Fam20C are still unknown. In this study, we generated a transgenic reporter model which expresses green fluorescence protein (GFP) driven by the Fam20C promoter. Recombineering was used to insert a 16 kb fragment of the mouse Fam20C gene (containing the 15 kb promoter and 1.1 kb of exon 1) into a pBluescript SK vector with the topaz variant of GFP and a bovine growth hormone polyadenylation sequence. GFP expression was subsequently evaluated by histomorphometry on cryosections from E14 to adult mice. Fluorescence was evident in the bone and teeth as early as E17.5. The GFP signal was maintained stably in odontoblasts and osteoblasts until 4 weeks after birth. The expression of GFP was significantly reduced in teeth, alveolar bone and muscle by 8 weeks of age. We also observed colocalization of the GFP signal with the Fam20C antibody in postnatal 1- and 7-day-old animals. Successful generation of Fam20C-GFP transgenic mice will provide a unique model for studying Fam20C gene expression and the biological function of this gene during odontogenesis and osteogenesis. PMID:25537657

  3. Characterization of Fam20C expression in odontogenesis and osteogenesis using transgenic mice.

    PubMed

    Du, Er-Xia; Wang, Xiao-Fang; Yang, Wu-Chen; Kaback, Deborah; Yee, Siu-Pok; Qin, Chun-Lin; George, Anne; Hao, Jian-Jun

    2015-06-01

    Our previous studies have demonstrated that Fam20C promotes differentiation and mineralization of odontoblasts, ameloblasts, osteoblasts and osteocytes during tooth and bone development. Ablation of the Fam20C gene inhibits bone and tooth growth by increasing fibroblast growth factor 23 in serum and causing hypophosphatemia in conditional knockout mice. However, control and regulation of the expression of Fam20C are still unknown. In this study, we generated a transgenic reporter model which expresses green fluorescence protein (GFP) driven by the Fam20C promoter. Recombineering was used to insert a 16 kb fragment of the mouse Fam20C gene (containing the 15 kb promoter and 1.1 kb of exon 1) into a pBluescript SK vector with the topaz variant of GFP and a bovine growth hormone polyadenylation sequence. GFP expression was subsequently evaluated by histomorphometry on cryosections from E14 to adult mice. Fluorescence was evident in the bone and teeth as early as E17.5. The GFP signal was maintained stably in odontoblasts and osteoblasts until 4 weeks after birth. The expression of GFP was significantly reduced in teeth, alveolar bone and muscle by 8 weeks of age. We also observed colocalization of the GFP signal with the Fam20C antibody in postnatal 1- and 7-day-old animals. Successful generation of Fam20C-GFP transgenic mice will provide a unique model for studying Fam20C gene expression and the biological function of this gene during odontogenesis and osteogenesis. PMID:25537657

  4. Lethal cutaneous disease in transgenic mice conditionally expressing type I human T cell leukemia virus Tax.

    PubMed

    Kwon, Hakju; Ogle, Louise; Benitez, Bobby; Bohuslav, Jan; Montano, Mauricio; Felsher, Dean W; Greene, Warner C

    2005-10-21

    Type I human T cell leukemia virus (HTLV-I) is etiologically linked with adult T cell leukemia, an aggressive and usually fatal expansion of activated CD4+ T lymphocytes that frequently traffic to skin. T cell transformation induced by HTLV-I involves the action of the 40-kDa viral Tax transactivator protein. Tax both stimulates the HTLV-I long terminal repeat and deregulates the expression of select cellular genes by altering the activity of specific host transcription factors, including cyclic AMP-responsive element-binding protein (CREB)/activating transcription factor, NF-kappaB/Rel, and serum response factor. To study initiating events involved in HTLV-I Tax-induced T cell transformation, we generated "Tet-off" transgenic mice conditionally expressing in a lymphocyte-restricted manner (EmuSR alpha promoter-enhancer) either wild-type Tax or mutant forms of Tax that selectively compromise the NF-kappaB (M22) or CREB/activating transcription factor (M47) activation pathways. Wild-type Tax and M47 Tax-expressing mice, but not M22-Tax expressing mice, developed progressive alopecia, hyperkeratosis, and skin lesions containing profuse activated CD4 T cell infiltrates with evidence of deregulated inflammatory cytokine production. In addition, these animals displayed systemic lymphadenopathy and splenomegaly. These findings suggest that Tax-mediated activation of NF-kappaB plays a key role in the development of this aggressive skin disease that shares several features in common with the skin disease occurring during the preleukemic stage in HTLV-I-infected patients. Of note, this skin disease completely resolved when Tax transgene expression was suppressed by administration of doxycycline, emphasizing the key role played by this viral oncoprotein in the observed pathology. PMID:16105841

  5. Cooperation of ETV6/RUNX1 and BCL2 enhances immunoglobulin production and accelerates glomerulonephritis in transgenic mice.

    PubMed

    Bauer, Eva; Schlederer, Michaela; Scheicher, Ruth; Horvath, Jaqueline; Aigner, Petra; Schiefer, Ana-Iris; Kain, Renate; Regele, Heinz; Hoermann, Gregor; Steiner, Günter; Kenner, Lukas; Sexl, Veronika; Villunger, Andreas; Moriggl, Richard; Stoiber, Dagmar

    2016-03-15

    The t(12;21) translocation generating the ETV6/RUNX1 fusion gene represents the most frequent chromosomal rearrangement in childhood leukemia. Presence of ETV6/RUNX1 alone is usually not sufficient for leukemia onset, and additional genetic alterations have to occur in ETV6/RUNX1-positive cells to cause transformation. We have previously generated an ETV6/RUNX1 transgenic mouse model where the expression of the fusion gene is restricted to CD19-positive B cells. Since BCL2 family members have been proposed to play a role in leukemogenesis, we investigated combined effects of ETV6/RUNX1 with exogenous expression of the antiapoptotic protein BCL2 by crossing ETV6/RUNX1 transgenic animals with Vav-BCL2 transgenic mice. Strikingly, co-expression of ETV6/RUNX1 and BCL2 resulted in significantly shorter disease latency in mice, indicating oncogene cooperativity. This was associated with faster development of follicular B cell lymphoma and exacerbated immune complex glomerulonephritis. ETV6/RUNX1-BCL2 double transgenic animals displayed increased B cell numbers and immunoglobulin titers compared to Vav-BCL2 transgenic mice. This led to pronounced deposition of immune complexes in glomeruli followed by accelerated development of immune complex glomerulonephritis. Thus, our study reveals a previously unrecognized synergism between ETV6/RUNX1 and BCL2 impacting on malignant disease and autoimmunity. PMID:26919255

  6. Cooperation of ETV6/RUNX1 and BCL2 enhances immunoglobulin production and accelerates glomerulonephritis in transgenic mice

    PubMed Central

    Bauer, Eva; Schlederer, Michaela; Scheicher, Ruth; Horvath, Jaqueline; Aigner, Petra; Schiefer, Ana-Iris; Kain, Renate; Regele, Heinz; Hoermann, Gregor; Steiner, Günter; Kenner, Lukas; Sexl, Veronika; Villunger, Andreas; Moriggl, Richard; Stoiber, Dagmar

    2016-01-01

    The t(12;21) translocation generating the ETV6/RUNX1 fusion gene represents the most frequent chromosomal rearrangement in childhood leukemia. Presence of ETV6/RUNX1 alone is usually not sufficient for leukemia onset, and additional genetic alterations have to occur in ETV6/RUNX1-positive cells to cause transformation. We have previously generated an ETV6/RUNX1 transgenic mouse model where the expression of the fusion gene is restricted to CD19-positive B cells. Since BCL2 family members have been proposed to play a role in leukemogenesis, we investigated combined effects of ETV6/RUNX1 with exogenous expression of the antiapoptotic protein BCL2 by crossing ETV6/RUNX1 transgenic animals with Vav-BCL2 transgenic mice. Strikingly, co-expression of ETV6/RUNX1 and BCL2 resulted in significantly shorter disease latency in mice, indicating oncogene cooperativity. This was associated with faster development of follicular B cell lymphoma and exacerbated immune complex glomerulonephritis. ETV6/RUNX1-BCL2 double transgenic animals displayed increased B cell numbers and immunoglobulin titers compared to Vav-BCL2 transgenic mice. This led to pronounced deposition of immune complexes in glomeruli followed by accelerated development of immune complex glomerulonephritis. Thus, our study reveals a previously unrecognized synergism between ETV6/RUNX1 and BCL2 impacting on malignant disease and autoimmunity. PMID:26919255

  7. Regulated tissue-specific alternative splicing of enhanced green fluorescent protein transgenes conferred by alpha-tropomyosin regulatory elements in transgenic mice.

    PubMed

    Ellis, Peter D; Smith, Christopher W J; Kemp, Paul

    2004-08-27

    The mutually exclusive exons 2 and 3 of alpha-tropomyosin (alphaTM) have been used as a model system for strictly regulated alternative splicing. Exon 2 inclusion is only observed at high levels in smooth muscle (SM) tissues, whereas striated muscle and non-muscle cells use predominantly exon 3. Experiments in cell culture have shown that exon 2 selection results from repression of exon 3 and that this repression is mediated by regulatory elements flanking exon 3. We have now tested the cell culture-derived model in transgenic mice. We show that by harnessing the intronic splicing regulatory elements, expression of an enhanced green fluorescent protein transgene with a constitutively active promoter can be restricted to SM cells. Splicing of both endogenous alphaTM and a series of transgenes carrying regulatory element mutations was analyzed by reverse transcriptasePCR. These studies indicated that although SM-rich tissues are equipped to regulate splicing of high levels of endogenous or transgene alphaTM RNA, other non-SM tissues such as spleen, which express lower amounts of alphaTM, also splice significant proportions of exon 2, and this splicing pattern can be recapitulated by transgenes expressed at low levels. We confirm the importance in vivo of the negatively acting regulatory elements for regulated skipping of exon 3. Moreover, we provide evidence that some of the regulatory factors responsible for exon 3 skipping appear to be titratable, with loss of regulated splicing sometimes being associated with high transgene expression levels. PMID:15194683

  8. E6 Gamma Decay

    SciTech Connect

    Brown, B. Alex; Rae, W. D. M.

    2011-05-06

    Rare electric hexacontatetrapole (E6) transitions are studied in the full (f{sub 7/2},f{sub 5/2},p{sub 3/2},p{sub 1/2}) shell-model basis. Comparison of theory to the results from the gamma decay in {sup 53}Fe and from inelastic electron scattering on {sup 52}Cr provides unique and interesting tests of the valence wavefunctions, the models used for energy density functionals and into the origin of effective charge.

  9. Contribution by synaptic zinc to the gender-disparate plaque formation in human Swedish mutant APP transgenic mice

    PubMed Central

    Lee, Joo-Yong; Cole, Toby B.; Palmiter, Richard D.; Suh, Sang Won; Koh, Jae-Young

    2002-01-01

    Endogenous metals may contribute to the accumulation of amyloid plaques in Alzheimer's disease. To specifically examine the role of synaptic zinc in the plaque accumulation, Tg2576 (also called APP2576) transgenic mice (hAPP+) expressing cerebral amyloid plaque pathology were crossed with mice lacking zinc transporter 3 (ZnT3−/−), which is required for zinc transport into synaptic vesicles. With aging, female hAPP+:ZnT3+/+ mice manifested higher levels of synaptic zinc, insoluble amyloid β, and plaques than males; these sex differences disappeared in hAPP+:ZnT3−/− mice. Both sexes of hAPP+:ZnT3−/− mice had markedly reduced plaque load and less insoluble amyloid β compared with hAPP+:ZnT3+/+ mice. Hence, of endogenous metals, synaptic zinc contributes predominantly to amyloid deposition in hAPP+ mice. PMID:12032347

  10. Bone turnover in wild type and pleiotrophin-transgenic mice housed for three months in the International Space Station (ISS).

    PubMed

    Tavella, Sara; Ruggiu, Alessandra; Giuliani, Alessandra; Brun, Francesco; Canciani, Barbara; Manescu, Adrian; Marozzi, Katia; Cilli, Michele; Costa, Delfina; Liu, Yi; Piccardi, Federica; Tasso, Roberta; Tromba, Giuliana; Rustichelli, Franco; Cancedda, Ranieri

    2012-01-01

    Bone is a complex dynamic tissue undergoing a continuous remodeling process. Gravity is a physical force playing a role in the remodeling and contributing to the maintenance of bone integrity. This article reports an investigation on the alterations of the bone microarchitecture that occurred in wild type (Wt) and pleiotrophin-transgenic (PTN-Tg) mice exposed to a near-zero gravity on the International Space Station (ISS) during the Mice Drawer System (MDS) mission, to date, the longest mice permanence (91 days) in space. The transgenic mouse strain over-expressing pleiotrophin (PTN) in bone was selected because of the PTN positive effects on bone turnover. Wt and PTN-Tg control animals were maintained on Earth either in a MDS payload or in a standard vivarium cage. This study revealed a bone loss during spaceflight in the weight-bearing bones of both strains. For both Tg and Wt a decrease of the trabecular number as well as an increase of the mean trabecular separation was observed after flight, whereas trabecular thickness did not show any significant change. Non weight-bearing bones were not affected. The PTN-Tg mice exposed to normal gravity presented a poorer trabecular organization than Wt mice, but interestingly, the expression of the PTN transgene during the flight resulted in some protection against microgravity's negative effects. Moreover, osteocytes of the Wt mice, but not of Tg mice, acquired a round shape, thus showing for the first time osteocyte space-related morphological alterations in vivo. The analysis of specific bone formation and resorption marker expression suggested that the microgravity-induced bone loss was due to both an increased bone resorption and a decreased bone deposition. Apparently, the PTN transgene protection was the result of a higher osteoblast activity in the flight mice. PMID:22438896

  11. Electrophysiology of Arcuate Neurokinin B Neurons in Female Tac2-EGFP Transgenic Mice

    PubMed Central

    Cholanian, Marina; Krajewski-Hall, Sally J.; Levine, Richard B.; McMullen, Nathaniel T.

    2014-01-01

    Neurons in the arcuate nucleus that coexpress kisspeptin, neurokinin B (NKB), and dynorphin (KNDy neurons) play an important role in the modulation of reproduction by estrogens. Here, we study the anatomical and electrophysiological properties of arcuate NKB neurons in heterozygous female transgenic mice with enhanced green fluorescent protein (EGFP) under the control of the Tac2 (NKB) promoter (Tac2-EGFP mice). The onset of puberty, estrous cyclicity, and serum LH were comparable between Tac2-EGFP and wild-type mice. The location of EGFP-immunoreactive neurons was consistent with previous descriptions of Tac2 mRNA-expressing neurons in the rodent. In the arcuate nucleus, nearly 80% of EGFP neurons expressed pro-NKB-immunoreactivity. Moreover, EGFP fluorescent intensity in arcuate neurons was increased by ovariectomy and reduced by 17β-estradiol (E2) treatment. Electrophysiology of single cells in tissue slices was used to examine the effects of chronic E2 treatment on Tac2-EGFP neurons in the arcuate nucleus of ovariectomized mice. Whole-cell recordings revealed arcuate NKB neurons to be either spontaneously active or silent in both groups. E2 had no significant effect on the basic electrophysiological properties or spontaneous firing frequencies. Arcuate NKB neurons exhibited either tonic or phasic firing patterns in response to a series of square-pulse current injections. Notably, E2 reduced the number of action potentials evoked by depolarizing current injections. This study demonstrates the utility of the Tac2-EGFP mouse for electrophysiological and morphological studies of KNDy neurons in tissue slices. In parallel to E2 negative feedback on LH secretion, E2 decreased the intensity of the EGFP signal and reduced the excitability of NKB neurons in the arcuate nucleus of ovariectomized Tac2-EGFP mice. PMID:24735328

  12. Hyperhomocysteinemia promotes inflammatory monocyte generation and accelerates atherosclerosis in transgenic cystathionine β-synthase deficient mice

    PubMed Central

    Zhang, Daqing; Jiang, Xiaohua; Fang, Pu; Yan, Yan; Song, Jian; Gupta, Sapna; Schafer, Andrew I.; Durante, William; Kruger, Warren D.; Yang, Xiaofeng; Wang, Hong

    2009-01-01

    Background Hyperhomocysteinemia (HHcy) is an independent risk factor for cardiovascular diseases. Monocytes display inflammatory and resident subsets, and commit to specific functions in atherogenesis. In this study, we examined the hypothesis that HHcy modulates monocyte heterogeneity and leads to atherosclerosis. Methods and Results We established a novel atherosclerosis susceptible mouse model with both severe HHcy and hypercholesterolemia, in which the mouse cystathionine β-synthase (CBS) and apolipoprotein E (apoE) genes are deficient, and an inducible human CBS transgene is introduced to circumvent the neonatal lethality of the CBS deficiency (Tg-hCBS apoE−/− Cbs−/− mice). Severe HHcy accelerated atherosclerosis and inflammatory monocyte/macrophage accumulation in lesions and increased plasma TNFα and MCP-1 levels in Tg-hCBS apoE−/− Cbs−/− mice fed a high fat diet. Furthermore, we characterized monocyte heterogeneity in Tg-hCBS apoE−/− Cbs−/− mice and another severe HHcy mouse model (Tg-S466L Cbs−/−) with a disease relevant mutation (Tg-S466L) that lacks hyperlipidemia. HHcy increased monocyte population and selective expansion of inflammatory Ly-6Chi and Ly-6Cmid monocyte subsets in blood, spleen and bone marrow of Tg-S466L Cbs−/− and Tg-hCBS apoE−/− Cbs−/− mice. These changes were exacerbated in Tg-S466L Cbs−/− mice with aging. Addition of L-homocysteine (100–500 μM), but not L-cysteine, maintained the Ly-6Chi subset and induced the Ly-6Cmid subset in cultured mouse primary splenocytes. Homocysteine-induced differentiation of Ly-6Cmid subset was prevented by catalase plus SOD, and the NAD(P)H oxidase inhibitor, apocynin. Conclusions HHcy promotes differentiation of inflammatory monocyte subsets and their accumulation in atherosclerotic lesions via NAD(P)H oxidase-mediated oxidant stress. PMID:19858416

  13. Nicastrin overexpression in transgenic mice induces aberrant behavior and APP processing.

    PubMed

    Goo, Jun Seo; Kim, Yong Bin; Shim, Sun Bo; Jee, Seung Wan; Lee, Su Hae; Kim, Ji Een; Hwang, In Sik; Lee, Young Ju; Kwak, Moon Hwa; Lim, Chuel Ju; Hong, Jin Tae; Hwang, Dae Youn

    2013-08-01

    Nicastrin (NCT) is a component of the presenilin protein complex, which is involved in the cleavage of β-amyloid precursor protein (βAPP) and Notch. The aim of this study was to determine the manner in which overexpression of wild-type human nicastrin (hNCTw) or mutant human nicastrin (hNCTm, D336A/Y337A) regulates brain functions and amyloid precusor protein (APP) processing. For this, we created transgenic (Tg) mice expressing neuron-specific enolase (NSE)-controlled hNCTw or hNCTm and measured their phenotypes as time passed. The NSE/hNCTw and NSE/hNCTm Tg groups exhibited greater behavioral dysfunction from 10 months of age than the non-Tg group, although their severities differed. Further, activity and component levels of the γ-secretase complex were significantly elevated in NSE/hNCTw Tg mice, expect for PEN-2. These alterations induced stimulation of APP processing, resulting in overproduction of Aβ-42 peptide in the NSE/hNCTw Tg group, whereas the NSE/hNCTm Tg group showed a comparatively weaker effect. Furthermore, the highest expression levels of β-secretase and NICD were observed in the NSE/hNCTw Tg group, similar to other phenotypes. Especially, a significances interference on the interaction between NCT and γ-secretase substrates was detected in NSE/hNCTm Tg groups compare with NSE/hNCTw Tg group. These results indicate that hNCTw overexpression in Tg mice promoted active assembly of the γ-secretase complex through modulation of APP processing and behavior, whereas the lesser effect in NSE/hNCTm Tg mice was due to reduced expression of hNCTm. These Tg mice could be useful for the development and application of therapeutic drugs in an animal model of Alzheimer's disease. PMID:23595812

  14. Decreased severity of collagen antibody and lipopolysaccharide-induced arthritis in human IL-32β overexpressed transgenic mice

    PubMed Central

    Ban, Jung Ok; Kim, Dae Hwan; Lee, Dong Hun; Song, Sukgil; Kim, Youngsoo; Han, Sang-Bae; Lee, Hee Pom; Hong, Jin Tae

    2015-01-01

    Interleukin (IL)-32, mainly produced by T-lymphocytes, natural killer cells, epithelial cells, and blood monocytes, is dominantly known as a pro-inflammatory cytokine. However, the role of IL-32 on inflammatory disease has been doubtful according to diverse conflicting results. This study was designed to examine the role of IL-32β on the development of collagen antibody (CAIA) and lipopolysaccharide (LPS)-induced inflammatory arthritis. Our data showed that the paw swelling volume and clinical score were significantly reduced in the CAIA and LPS-treated IL-32β transgenic mice compared with non-transgenic mice. The populations of cytotoxic T, NK and dendritic cells was inhibited and NF-κB and STAT3 activities were significantly lowered in the CAIA and LPS-treated IL-32β transgenic mice. The expression of pro-inflammatory proteins was prevented in the paw tissues of CAIA and LPS-treated IL-32β transgenic mice. In addition, IL-32β altered several cytokine levels in the blood, spleen and paw joint. Our data indicates that IL-32β comprehensively inhibits the inflammation responses in the CAIA and LPS-induced inflammatory arthritis model. PMID:26497686

  15. Calcium imaging of vomeronasal organ response using slice preparations from transgenic mice expressing G-CaMP2.

    PubMed

    Yu, C Ron

    2013-01-01

    The vomeronasal organ (VNO) in vertebrate animals detects pheromones and interspecies chemical signals. We describe in this chapter a Ca(2+) imaging approach using transgenic mice that express the genetically encoded Ca(2+) sensor G-CaMP2 in VNO tissue. This approach allows us to analyze the complex patterns of the vomeronasal neuron response to large number of chemosensory stimuli. PMID:24014364

  16. Parkinson-like phenotype in insulin-resistant PED/PEA-15 transgenic mice

    PubMed Central

    Perruolo, Giuseppe; Viggiano, Davide; Fiory, Francesca; Cassese, Angela; Nigro, Cecilia; Liotti, Antonietta; Miele, Claudia; Beguinot, Francesco; Formisano, Pietro

    2016-01-01

    Neurological abnormalities, such as Parkinson-like disorders (PlD), are often co-morbidities of Type 2 Diabetic (T2D) patients, although the epidemiological link between these two disorders remains controversial. The PED/PEA-15 protein represents a possible candidate linking T2D and PD, because it is increased in subjects with T2D and is highly expressed in the brain. To test this hypothesis, we have analyzed the neurological and neurochemical phenotype of transgenic mice overexpressing PED/PEA-15 (tgPED). These mice develop impaired glucose tolerance and insulin resistance, accompanied by neurological features resembling PlD: feet clasping, slow and delayed locomotor movements in different behavioral tests in absence of clear cognitive deficits, ataxia or anxiety. Morphological analysis of the brains showed selective modifications of metabolic activity in the striatal region. In the same region, we have observed 26% decrease of dopamine fibers, confirmed by immunohistochemistry and Western Blot for tyrosine hydroxylase. Moreover, they also showed 48% reduction of dopamine levels in the striatum. Thus the tgPED mice may represent a genetic animal model of neurological disease linked to T2D. PMID:27426254

  17. Parkinson-like phenotype in insulin-resistant PED/PEA-15 transgenic mice.

    PubMed

    Perruolo, Giuseppe; Viggiano, Davide; Fiory, Francesca; Cassese, Angela; Nigro, Cecilia; Liotti, Antonietta; Miele, Claudia; Beguinot, Francesco; Formisano, Pietro

    2016-01-01

    Neurological abnormalities, such as Parkinson-like disorders (PlD), are often co-morbidities of Type 2 Diabetic (T2D) patients, although the epidemiological link between these two disorders remains controversial. The PED/PEA-15 protein represents a possible candidate linking T2D and PD, because it is increased in subjects with T2D and is highly expressed in the brain. To test this hypothesis, we have analyzed the neurological and neurochemical phenotype of transgenic mice overexpressing PED/PEA-15 (tgPED). These mice develop impaired glucose tolerance and insulin resistance, accompanied by neurological features resembling PlD: feet clasping, slow and delayed locomotor movements in different behavioral tests in absence of clear cognitive deficits, ataxia or anxiety. Morphological analysis of the brains showed selective modifications of metabolic activity in the striatal region. In the same region, we have observed 26% decrease of dopamine fibers, confirmed by immunohistochemistry and Western Blot for tyrosine hydroxylase. Moreover, they also showed 48% reduction of dopamine levels in the striatum. Thus the tgPED mice may represent a genetic animal model of neurological disease linked to T2D. PMID:27426254

  18. Life extension factor klotho prevents mortality and enhances cognition in hAPP transgenic mice.

    PubMed

    Dubal, Dena B; Zhu, Lei; Sanchez, Pascal E; Worden, Kurtresha; Broestl, Lauren; Johnson, Erik; Ho, Kaitlyn; Yu, Gui-Qiu; Kim, Daniel; Betourne, Alexander; Kuro-O, Makoto; Masliah, Eliezer; Abraham, Carmela R; Mucke, Lennart

    2015-02-11

    Aging is the principal demographic risk factor for Alzheimer disease (AD), the most common neurodegenerative disorder. Klotho is a key modulator of the aging process and, when overexpressed, extends mammalian lifespan, increases synaptic plasticity, and enhances cognition. Whether klotho can counteract deficits related to neurodegenerative diseases, such as AD, is unknown. Here we show that elevating klotho expression decreases premature mortality and network dysfunction in human amyloid precursor protein (hAPP) transgenic mice, which simulate key aspects of AD. Increasing klotho levels prevented depletion of NMDA receptor (NMDAR) subunits in the hippocampus and enhanced spatial learning and memory in hAPP mice. Klotho elevation in hAPP mice increased the abundance of the GluN2B subunit of NMDAR in postsynaptic densities and NMDAR-dependent long-term potentiation, which is critical for learning and memory. Thus, increasing wild-type klotho levels or activities improves synaptic and cognitive functions, and may be of therapeutic benefit in AD and other cognitive disorders. PMID:25673831

  19. Memantine improves cognition and reduces Alzheimer's-like neuropathology in transgenic mice.

    PubMed

    Martinez-Coria, Hilda; Green, Kim N; Billings, Lauren M; Kitazawa, Masashi; Albrecht, Miriam; Rammes, Gerhard; Parsons, Chris G; Gupta, Sandeep; Banerjee, Pradeep; LaFerla, Frank M

    2010-02-01

    Memantine is an N-methyl-d-aspartate receptor antagonist that is approved for the treatment of moderate to severe Alzheimer's disease (AD). In this study, three groups of triple-transgenic (3xTg-AD) mice with differing levels of AD-like pathology (6, 9, and 15 months of age) were treated for 3 months with doses of memantine equivalent to those used in humans. After the treatment, memantine-treated mice had restored cognition and significantly reduced the levels of insoluble amyloid-beta (Abeta), Abeta dodecamers (Abeta*56), prefibrillar soluble oligomers, and fibrillar oligomers. The effects on pathology were stronger in older, more impaired animals. Memantine treatment also was associated with a decline in the levels of total tau and hyperphosphorylated tau. Finally, memantine pre-incubation prevented Abeta-induced inhibition of long-term potentiation in hippocampal slices of cognitively normal mice. These results suggest that the effects of memantine treatment on AD brain include disease modification and prevention of synaptic dysfunction. PMID:20042680

  20. Life Extension Factor Klotho Prevents Mortality and Enhances Cognition in hAPP Transgenic Mice

    PubMed Central

    Zhu, Lei; Sanchez, Pascal E.; Worden, Kurtresha; Broestl, Lauren; Johnson, Erik; Ho, Kaitlyn; Yu, Gui-Qiu; Kim, Daniel; Betourne, Alexander; Kuro-o, Makoto; Masliah, Eliezer; Abraham, Carmela R.

    2015-01-01

    Aging is the principal demographic risk factor for Alzheimer disease (AD), the most common neurodegenerative disorder. Klotho is a key modulator of the aging process and, when overexpressed, extends mammalian lifespan, increases synaptic plasticity, and enhances cognition. Whether klotho can counteract deficits related to neurodegenerative diseases, such as AD, is unknown. Here we show that elevating klotho expression decreases premature mortality and network dysfunction in human amyloid precursor protein (hAPP) transgenic mice, which simulate key aspects of AD. Increasing klotho levels prevented depletion of NMDA receptor (NMDAR) subunits in the hippocampus and enhanced spatial learning and memory in hAPP mice. Klotho elevation in hAPP mice increased the abundance of the GluN2B subunit of NMDAR in postsynaptic densities and NMDAR-dependent long-term potentiation, which is critical for learning and memory. Thus, increasing wild-type klotho levels or activities improves synaptic and cognitive functions, and may be of therapeutic benefit in AD and other cognitive disorders. PMID:25673831

  1. Transgenic mice expressing yellow fluorescent protein under control of the human tyrosine hydroxylase promoter.

    PubMed

    Choi, Eun Yang; Yang, Jae Won; Park, Myung Sun; Sun, Woong; Kim, Hyun; Kim, Seung U; Lee, Myung Ae

    2012-10-01

    Pathogenesis of Parkinson's disease and related catecholaminergic neurological disorders is closely associated with changes in the levels of tyrosine hydroxylase (TH). Therefore, investigation of the regulation of the TH gene system should assist in understanding the pathomechanisms involved in these neurological disorders. To identify regulatory domains that direct human TH expression in the central nervous system (CNS), we generated two transgenic mouse lines in which enhanced yellow fluorescent protein (EYFP) is expressed under the control of either 3.2-kb (hTHP-EYFP construct) human TH promoter or 3.2-kb promoter with 2-kb 3'-flanking regions (hTHP-ex3-EYFP construct) of the TH gene. In the adult transgenic mouse brain, the hTHP-EYFP construct directs neuron-specific EYFP expression in various CNS areas, such as olfactory bulb, striatum, interpeduncular nucleus, cerebral cortex, hippocampus, and particularly dentate gyrus. Although these EYFP-positive cells were identified as mature neurons, few EYFP-positive cells were TH-positive neurons. On the other hand, we could detect the EYFP mRNA expression in a subset of neurons in the olfactory bulb, midbrain, and cerebellum, in which expression of endogenous TH is enriched, with hTHP-ex3-EYFP transgenic mice. These results indicate that the 3.2-kb sequence upstream of the TH gene is not sufficient for proper expression and that the 2-kb sequence from the translation start site to exon 3 is necessary for expression of EYFP in a subset of catecholaminergic neurons. PMID:22714400

  2. An acetylcholine receptor alpha subunit promoter confers intrathymic expression in transgenic mice. Implications for tolerance of a transgenic self-antigen and for autoreactivity in myasthenia gravis.

    PubMed

    Salmon, A M; Bruand, C; Cardona, A; Changeux, J P; Berrih-Aknin, S

    1998-06-01

    Myasthenia gravis (MG) is an autoimmune disease targeting the skeletal muscle acetylcholine receptor (AChR). Although the autoantigen is present in the thymus, it is not tolerated in MG patients. In addition, the nature of the cell bearing the autoantigen is controversial. To approach these questions, we used two lineages of transgenic mice in which the beta-galactosidase (beta-gal) gene is under the control of a 842-bp (Tg1) or a 3300-bp promoter fragment (Tg2) of the chick muscle alpha subunit AChR gene. In addition to expression in muscle cells, thymic expression was observed in both mouse lines (mainly in myoid cells in Tg1 and myoid cells and epithelial cells in Tg2). After challenge with beta-gal, Tg1 mice produced Th2-dependent anti-beta-gal antibodies, while Tg2 mice were almost unresponsive. By contrast, in a proliferation assay both Tg lines were unresponsive to beta-gal. Cells from Tg1 mice produce Th2-dependent cytokine whereas cells from Tg2 mice were nonproducing in response to beta-gal. These data indicate that the level of expression in Tg1 mice could be sufficient to induce tolerance of Th1 cells but not of Th2 cells, while both populations are tolerated in Tg2 mice. These findings are compatible with the hypothesis that AChR expression is not sufficiently abundant in MG thymus to induce a full tolerance. PMID:9616205

  3. Pathological hallmarks of amyotrophic lateral sclerosis/frontotemporal lobar degeneration in transgenic mice produced with TDP-43 genomic fragments.

    PubMed

    Swarup, Vivek; Phaneuf, Daniel; Bareil, Christine; Robertson, Janice; Rouleau, Guy A; Kriz, Jasna; Julien, Jean-Pierre

    2011-09-01

    Transactive response DNA-binding protein 43 ubiquitinated inclusions are a hallmark of amyotrophic lateral sclerosis and of frontotemporal lobar degeneration with ubiquitin-positive inclusions. Yet, mutations in TARDBP, the gene encoding these inclusions are associated with only 3% of sporadic and familial amyotrophic lateral sclerosis. Recent transgenic mouse studies have revealed a high degree of toxicity due to transactive response DNA-binding protein 43 proteins when overexpressed under the control of strong neuronal gene promoters, resulting in early paralysis and death, but without the presence of amyotrophic lateral sclerosis-like ubiquitinated transactive response DNA-binding protein 43-positive inclusions. To better mimic human amyotrophic lateral sclerosis, we generated transgenic mice that exhibit moderate and ubiquitous expression of transactive response DNA-binding protein 43 species using genomic fragments that encode wild-type human transactive response DNA-binding protein 43 or familial amyotrophic lateral sclerosis-linked mutant transactive response DNA-binding protein 43 (G348C) and (A315T). These novel transgenic mice develop many age-related pathological and biochemical changes reminiscent of human amyotrophic lateral sclerosis including ubiquitinated transactive response DNA-binding protein 43-positive inclusions, transactive response DNA-binding protein 43 cleavage fragments, intermediate filament abnormalities, axonopathy and neuroinflammation. All three transgenic mouse models (wild-type, G348C and A315T) exhibited impaired learning and memory capabilities during ageing, as well as motor dysfunction. Real-time imaging with the use of biophotonic transactive response DNA-binding protein 43 transgenic mice carrying a glial fibrillary acidic protein-luciferase reporter revealed that the behavioural defects were preceded by induction of astrogliosis, a finding consistent with a role for reactive astrocytes in amyotrophic lateral sclerosis

  4. Virally mediated optogenetic excitation and inhibition of pain in freely moving non-transgenic mice

    PubMed Central

    Iyer, Shrivats Mohan; Montgomery, Kate L.; Towne, Chris; Lee, Soo Yeun; Ramakrishnan, Charu; Deisseroth, Karl; Delp, Scott L.

    2014-01-01

    Primary nociceptors are the first neurons involved in the complex processing system that regulates normal and pathological pain1. Our ability to excite and inhibit these neurons has been limited by pharmacological and electrical stimulation constraints; non-invasive excitation and inhibition of these neurons in freely moving non-transgenic animals has not been possible. Here we use an optogenetic2 strategy to bidirectionally control nociceptors of non-transgenic mice. Intra-sciatic nerve injection of adeno-associated viruses encoding an excitatory opsin enabled light-inducible stimulation of acute pain, place aversion, and optogenetically mediated reductions in withdrawal thresholds to mechanical and thermal stimuli. In contrast, viral delivery of an inhibitory opsin enabled light-inducible inhibition of acute pain perception, and reversed mechanical allodynia and thermal hyperalgesia in a model of neuropathic pain. Light was delivered transdermally enabling these behaviors to be induced in freely moving animals. This approach may have utility in basic and translational pain research, and enable rapid drug screening and testing of newly engineered opsins. PMID:24531797

  5. Pathogenesis of dilated cardiomyopathy: molecular, structural, and population analyses in tropomodulin-overexpressing transgenic mice.

    PubMed

    Sussman, M A; Welch, S; Gude, N; Khoury, P R; Daniels, S R; Kirkpatrick, D; Walsh, R A; Price, R L; Lim, H W; Molkentin, J D

    1999-12-01

    Dilated cardiomyopathy is characterized by decreased contractile function and loss of myofibril organization. Previously unexplored structural and molecular events that precede and initiate dilation can now be studied in tropomodulin-overexpressing transgenic (TOT) mice exhibiting progressive dilated cardiomyopathy. Onset of dilation did not correspond to a change in transgene expression levels, which were more than threefold above normal at birth and remained elevated throughout postnatal life. Similarly, mitogen-activated protein kinase activation (p38, ERK1/ERK2, JNK1/JNK2) was not associated with dilation. In contrast, calcineurin was activated before dilation, presumably due to doubling of intracellular diastolic calcium levels in TOT cardiomyocytes. Amplitude of systolic calcium transients was greatly increased as well, demonstrating the novel and unique calcium handling profile of TOT cardiomyocytes. Loss of myofibril organization was not apparent by confocal microscopy until over 1 week after birth, although neonatal sarcomeric abnormalities were revealed by ultrastructural analysis. Rapid postnatal increases in heart:body weight ratio at 1.5 weeks were followed by two waves of mortality between 2 and 3 weeks after birth coincident with maturational stress. Ultimately, TOT pathogenesis is a compensatory response to altered sarcomeric structure driven by calcineurin activation within days after birth, making TOTs an excellent paradigm for studying the role of calcium overload in dilated cardiomyopathy. PMID:10595939

  6. Intrachoroidal Neovascularization in Transgenic Mice Overexpressing Vascular Endothelial Growth Factor in the Retinal Pigment Epithelium

    PubMed Central

    Schwesinger, Catherine; Yee, Charles; Rohan, Richard M.; Joussen, Antonia M.; Fernandez, Antonio; Meyer, Tobias N.; Poulaki, Vassiliki; Ma, Joseph J. K.; Redmond, T. Michael; Liu, Suyan; Adamis, Anthony P.; D’Amato, Robert J.

    2001-01-01

    Choroidal neovascularization in age-related macular degeneration is a frequent and poorly treatable cause of vision loss in elderly Caucasians. This choroidal neovascularization has been associated with the expression of vascular endothelial growth factor (VEGF). In current animal models choroidal neovascularization is induced by subretinal injection of growth factors or vectors encoding growth factors such as VEGF, or by disruption of the Bruch’s membrane/retinal pigment epithelium complex with laser treatment. We wished to establish a transgenic murine model of age-related macular degeneration, in which the overexpression of VEGF by the retinal pigment epithelium induces choroidal neovascularization. A construct consisting of a tissue-specific murine retinal pigment epithelium promoter (RPE65 promoter) coupled to murine VEGF164 cDNA with a rabbit β-globin-3′ UTR was introduced into the genome of albino mice. Transgene mRNA was expressed in the retinal pigment epithelium at all ages peaking at 4 months. The expression of VEGF protein was increased in both the retinal pigment epithelium and choroid. An increase of intravascular adherent leukocytes and vessel leakage was observed. Histopathology revealed intrachoroidal neovascularization that did not penetrate through an intact Bruch’s membrane. These results support the hypothesis that additional insults to the integrity of Bruch’s membrane are required to induce growth of choroidal vessels into the subretinal space as seen in age-related macular degeneration. This model may be useful to screen for inhibitors of choroidal vessel growth. PMID:11238064

  7. Nuclear microscopy of diffuse plaques in the brains of transgenic mice

    NASA Astrophysics Data System (ADS)

    Rajendran, Reshmi; Ren, Minqin; Casadesus, Gemma; Smith, Mark A.; Perry, George; Huang, En; Ong, Wei Yi; Halliwell, Barry; Watt, Frank

    2005-04-01

    Using nuclear microscopy, extracellular diffuse amyloid deposits in fresh unstained brain tissue from Alzheimer's disease transgenic mice Tg2576 have been identified and analyzed for trace element content. Off-axis scanning transmission ion microscopy (STIM) images can be obtained which are similar to the images produced using direct STIM. Since the proton beam current required for off-axis STIM is compatible with PIXE and RBS, we can identify the plaque location and analyze for trace elements simultaneously. Analysis of the diffuse plaques showed an increase in the transition metals iron and zinc compared with the surrounding area of comparable areal density. This supports the theory that redox interactions between Aβ and metals could be at the heart of a pathological feedback system wherein Aβ amyloidosis and oxidative stress promote each other, possibly via Fenton chemistry.

  8. Focally Elevated Creatine Detected in Amyloid Precursor Protein (APP) Transgenic Mice and Alzheimer Disease Brain Tissue

    SciTech Connect

    Gallant,M.; Rak, M.; Szeghalmi, A.; Del Bigio, M.; Westaway, D.; Yang, J.; Julian, R.; Gough, K.

    2006-01-01

    The creatine/phosphocreatine system, regulated by creatine kinase, plays an important role in maintaining energy balance in the brain. Energy metabolism and the function of creatine kinase are known to be affected in Alzheimer diseased brain and in cells exposed to the {beta}-amyloid peptide. We used infrared microspectroscopy to examine hippocampal, cortical, and caudal tissue from 21-89-week-old transgenic mice expressing doubly mutant (K670N/M671L and V717F) amyloid precursor protein and displaying robust pathology from an early age. Microcrystalline deposits of creatine, suggestive of perturbed energetic status, were detected by infrared microspectroscopy in all animals with advanced plaque pathology. Relatively large creatine deposits were also found in hippocampal sections from post-mortem Alzheimer diseased human brain, compared with hippocampus from non-demented brain. We therefore speculate that this molecule is a marker of the disease process.

  9. Development of neuroendocrine tumors in the gastrointestinal tract of transgenic mice. Heterogeneity of hormone expression.

    PubMed Central

    Rindi, G.; Grant, S. G.; Yiangou, Y.; Ghatei, M. A.; Bloom, S. R.; Bautch, V. L.; Solcia, E.; Polak, J. M.

    1990-01-01

    Expression of hormones in endocrine tumors and derived cell lines of transgenic mice carrying insulin-promoted oncogenes has been investigated by histochemical, immunohistochemical, ultrastructural, and radioimmunologic means. Tumors of the pancreas, small intestine, mesentery, and liver were examined. Insulin-immunoreactive cells were prevalent in pancreatic tumors, with a significant subpopulation of pancreatic polypeptide-immunoreactive elements. Conventional ultrastructural and immunogold analysis identified insulin-storing beta granules in pancreatic tumor cells. In contrast, the largest immunoreactive subpopulation of intestinal tumors expressed secretin (53% of total cells), followed by proglucagon-related peptides (15%), glucose-dependent insulinotropic polypeptide (7%), gastrin (7%), pancreatic polypeptide (2%), neurotensin (2%), and somatostatin (1%). No detectable immunoreactivity for either insulin or serotonin was observed. Electron microscopy and immunogold labeling showed that intestinal tumor cells contained secretin-storing S-type granules. Lymph node and liver tumors contained secretin-immunoreactive cells with ultrastructural features similar to those of intestinal tumors. In addition, high levels of circulating insulinlike and secretinlike immunoreactants were detectable. Analogous hormone profiles were identified in tumor cell lines and culture media. Large T-antigen immunoreactivity was detected in all the nuclei of neoplastic cells, as well as in insulin-immunoreactive elements of non-neoplastic islets and pancreatic ducts and in some secretin-immunoreactive cells of small intestinal mucosa. These data indicate that neuroendocrine tumors arise both in beta cell and S-cell subpopulations of transgenic mice. Images Figure 5 Figure 1 Figure 2 Figure 3 Figure 4 Figure 6 PMID:2162628

  10. Disrupted Differentiation and Oncogenic Transformation of Lymphoid Progenitors in E2A-HLF Transgenic Mice

    PubMed Central

    Smith, Kevin S.; Rhee, Joon Whan; Naumovski, Louie; Cleary, Michael L.

    1999-01-01

    The hepatic leukemia factor (HLF) gene codes for a basic region-leucine zipper (bZIP) protein that is disrupted by chromosomal translocations in a subset of pediatric acute lymphoblastic leukemias. HLF undergoes fusions with the E2A gene, resulting in chimeric E2a-Hlf proteins containing the E2a transactivation domains and the Hlf bZIP DNA binding and dimerization motifs. To investigate the in vivo role of this chimeric bZIP protein in oncogenic transformation, its expression was directed to the lymphoid compartments of transgenic mice. Within the thymus, E2a-Hlf induced profound hypoplasia, premature involution, and progressive accumulation of a T-lineage precursor population arrested at an early stage of maturation. In the spleen, mature T cells were present but in reduced numbers, and they lacked expression of the transgene, suggesting further that E2a-Hlf expression was incompatible with T-cell differentiation. In contrast, mature splenic B cells expressed E2a-Hlf but at lower levels and without apparent adverse or beneficial effects on their survival. Approximately 60% of E2A-HLF mice developed lymphoid malignancies with a mean latency of 10 months. Tumors were monoclonal, consistent with a requirement for secondary genetic events, and displayed phenotypes of either mid-thymocytes or, rarely, B-cell progenitors. We conclude that E2a-Hlf disrupts the differentiation of T-lymphoid progenitors in vivo, leading to profound postnatal thymic depletion and rendering B- and T-cell progenitors susceptible to malignant transformation. PMID:10330184

  11. Everolimus prolonged survival in transgenic mice with EGFR-driven lung tumors.

    PubMed

    Yasugi, Masayuki; Takigawa, Nagio; Ochi, Nobuaki; Ohashi, Kadoaki; Harada, Daijiro; Ninomiya, Takashi; Murakami, Toshi; Honda, Yoshihiro; Ichihara, Eiki; Tanimoto, Mitsune; Kiura, Katsuyuki

    2014-08-15

    Everolimus is an orally administered mTOR inhibitor. The effect, and mechanism of action, of everolimus on lung cancers with an epidermal growth factor receptor (EGFR) mutation remain unclear. Four gefitinib-sensitive and -resistant cell lines were used in the present work. Growth inhibition was determined using the MTT assay. Transgenic mice carrying the EGFR L858R mutation were treated with everolimus (10 mg/kg/day), or vehicle alone, from 5 to 20 weeks of age, and were then sacrificed. To evaluate the efficacy of everolimus in prolonging survival, everolimus (10 mg/kg/day) or vehicle was administered from 5 weeks of age. The four cell lines were similarly sensitive to everolimus. Expression of phosphorylated (p) mTOR and pS6 were suppressed upon treatment with everolimus in vitro, whereas the pAKT level increased. The numbers of lung tumors with a long axis exceeding 1mm in the everolimus-treated and control groups were 1.9 ± 0.9 and 9.4 ± 3.2 (t-test, p<0.001), respectively. pS6 was suppressed during eve r olimus treatment. Although apoptosis and autophagy were not induced in everolimus-treated EGFR transgenic mice, angiogenesis was suppressed. The median survival time in the everolimus-treated group (58.0 weeks) was significantly longer than that in the control group (31.2 weeks) (logrank test, p<0.001). These findings suggest that everolimus had an indirect effect on tumor formation by inhibiting angiogenesis and might be effective to treat lung tumors induced by an activating EGFR gene mutation. PMID:24768699

  12. ABCA1 overexpression leads to hyperalphalipoproteinemia and increased biliary cholesterol excretion in transgenic mice

    PubMed Central

    Vaisman, Boris L.; Lambert, Gilles; Amar, Marcelo; Joyce, Charles; Ito, Toshimitsu; Shamburek, Robert D.; Cain, William J.; Fruchart-Najib, Jamila; Neufeld, Edward D.; Remaley, Alan T.; Brewer, H. Bryan; Santamarina-Fojo, Silvia

    2001-01-01

    The discovery of the ABCA1 lipid transporter has generated interest in modulating human plasma HDL levels and atherogenic risk by enhancing ABCA1 gene expression. To determine if increased ABCA1 expression modulates HDL metabolism in vivo, we generated transgenic mice that overexpress human ABCA1 (hABCA1-Tg). Hepatic and macrophage expression of hABCA1 enhanced macrophage cholesterol efflux to apoA-I; increased plasma cholesterol, cholesteryl esters (CEs), free cholesterol, phospholipids, HDL cholesterol, and apoA-I and apoB levels; and led to the accumulation of apoE-rich HDL1. ABCA1 transgene expression delayed 125I-apoA-I catabolism in both liver and kidney, leading to increased plasma apoA-I levels, but had no effect on apoB secretion after infusion of Triton WR1339. Although the plasma clearance of HDL-CE was not significantly altered in hABCA1-Tg mice, the net hepatic delivery of exogenous 3H-CEt-HDL, which is dependent on the HDL pool size, was increased 1.5-fold. In addition, the cholesterol and phospholipid concentrations in hABCA1-Tg bile were increased 1.8-fold. These studies show that steady-state overexpression of ABCA1 in vivo (a) raises plasma apoB levels without altering apoB secretion and (b) raises plasma HDL-C and apoA-I levels, facilitating hepatic reverse cholesterol transport and biliary cholesterol excretion. Similar metabolic changes may modify atherogenic risk in humans. PMID:11457883

  13. Silencing Mutant Ataxin-3 Rescues Motor Deficits and Neuropathology in Machado-Joseph Disease Transgenic Mice

    PubMed Central

    Onofre, Isabel; Albuquerque, David; Hirai, Hirokazu; Déglon, Nicole; de Almeida, Luís Pereira

    2013-01-01

    Machado-Joseph disease (MJD) or spinocerebellar ataxia type 3 (SCA3) is an autosomal dominantly-inherited neurodegenerative disorder caused by the over-repetition of a CAG codon in the MJD1 gene. This expansion translates into a polyglutamine tract that confers a toxic gain-of-function to the mutant protein – ataxin-3, leading to neurodegeneration in specific brain regions, with particular severity in the cerebellum. No treatment able to modify the disease progression is available. However, gene silencing by RNA interference has shown promising results. Therefore, in this study we investigated whether lentiviral-mediated allele-specific silencing of the mutant ataxin-3 gene, after disease onset, would rescue the motor behavior deficits and neuropathological features in a severely impaired transgenic mouse model of MJD. For this purpose, we injected lentiviral vectors encoding allele-specific silencing-sequences (shAtx3) into the cerebellum of diseased transgenic mice expressing the targeted C-variant of mutant ataxin-3 present in 70% of MJD patients. This variation permits to discriminate between the wild-type and mutant forms, maintaining the normal function of the wild-type allele and silencing only the mutant form. Quantitative analysis of rotarod performance, footprint and activity patterns revealed significant and robust alleviation of gait, balance (average 3-fold increase of rotarod test time), locomotor and exploratory activity impairments in shAtx3-injected mice, as compared to control ones injected with shGFP. An important improvement of neuropathology was also observed, regarding the number of intranuclear inclusions, calbindin and DARPP-32 immunoreactivity, fluorojade B and Golgi staining and molecular and granular layers thickness. These data demonstrate for the first time the efficacy of gene silencing in blocking the MJD-associated motor-behavior and neuropathological abnormalities after the onset of the disease, supporting the use of this strategy

  14. Analysis of the effects of overexpression of metallothionein-I in transgenic mice on the reproductive toxicology of cadmium.

    PubMed

    Dalton, T; Fu, K; Enders, G C; Palmiter, R D; Andrews, G K

    1996-01-01

    Exposure to low levels of cadmium reduces fertility. In male mice spermatogenesis is highly sensitive to cadmium, whereas in females the peri-implantation period of pregnancy is sensitive. To examine the potential roles of the cadmium-binding protein, metallothionein (MT), in the reproductive toxicology of cadmium, we examined a transgenic mouse strain that overexpresses metallothionein-I (MT-I). These mice had dramatically increased steady-state levels of MT-I mRNA and MT in the testes and in the female reproductive tract during the peri-implantation period of pregnancy, and this overexpression occurred in a cell-specific and temporally regulated manner similar to that of the endogenous MT-I gene. Transgenic and control males were injected with cadmium, and the histology of the testes was examined. An injection of 7.5 mumol Cd/kg had no effect on histology of the testes in either transgenic or control mice. In contrast, an injection of 10 mumol Cd/kg caused rapid changes in the histology of the testes and resulted in pronounced testicular necrosis in both control and transgenic mice. Female transgenic and control mice were mated and then injected with cadmium (30-45 mumol Cd/kg) on the day of blastocyst implantation (day 4). In both of these groups, injection of cadmium reduced pregnancy rate, and no dramatic protection was afforded by maternal and/or embryonic overexpression of MT. Thus, overexpression of MT-I does not significantly protect against either of these cadmium-induced effects on fertility. PMID:8834864

  15. Increased astrocyte expression of IL-6 or CCL2 in transgenic mice alters levels of hippocampal and cerebellar proteins

    PubMed Central

    Gruol, Donna L.; Vo, Khanh; Bray, Jennifer G.

    2014-01-01

    Emerging research has identified that neuroimmune factors are produced by cells of the central nervous system (CNS) and play critical roles as regulators of CNS function, directors of neurodevelopment and responders to pathological processes. A wide range of neuroimmune factors are produced by CNS cells, primarily the glial cells, but the role of specific neuroimmune factors and their glial cell sources in CNS biology and pathology have yet to be fully elucidated. We have used transgenic mice that express elevated levels of a specific neuroimmune factor, the cytokine IL-6 or the chemokine CCL2, through genetic modification of astrocyte expression to identify targets of astrocyte produced IL-6 or CCL2 at the protein level. We found that in non-transgenic mice constitutive expression of IL-6 and CCL2 occurs in the two CNS regions studied, the hippocampus and cerebellum, as measured by ELISA. In the CCL2 transgenic mice elevated levels of CCL2 were evident in the hippocampus and cerebellum, whereas in the IL-6 transgenic mice, elevated levels of IL-6 were only evident in the cerebellum. Western blot analysis of the cellular and synaptic proteins in the hippocampus and cerebellum of the transgenic mice showed that the elevated levels of CCL2 or IL-6 resulted in alterations in the levels of specific proteins and that these actions differed for the two neuroimmune factors and for the two brain regions. These results are consistent with cell specific profiles of action for IL-6 and CCL2, actions that may be an important aspect of their respective roles in CNS physiology and pathophysiology. PMID:25177271

  16. DHA diet reduces AD pathology in young APPswe/PS1 Delta E9 transgenic mice: possible gender effects.

    PubMed

    Perez, Sylvia E; Berg, Brian M; Moore, Kenneth A; He, Bin; Counts, Scott E; Fritz, Jason J; Hu, Yuan-Shih; Lazarov, Orly; Lah, James J; Mufson, Elliott J

    2010-04-01

    Epidemiological and clinical trial findings suggest that consumption of docosahexaenoic acid (DHA) lowers the risk of Alzheimer's disease (AD). We examined the effects of short-term (3 months) DHA enriched diet on plaque deposition and synaptic defects in forebrain of young APPswe/PS1 Delta E9 transgenic (tg) and non-transgenic (ntg) mice. Gas chromatography revealed a significant increase in DHA concomitant with a decrease of arachidonic acid in both brain and liver in mice fed with DHA. Female tg mice consumed relatively more food daily than ntg female mice, independent of diet. Plaque load was significantly reduced in the cortex, ventral hippocampus and striatum of female APPswe/PS1 Delta E9 tg mice on DHA diet compared to female tg mice on control diet. Immunoblot quantitation of the APOE receptor, LR11, which is involved in APP trafficking and A beta production, were unchanged in mice on DHA or control diets. Moreover drebrin levels were significantly increased in the hippocampus of tg mice on the DHA diet. Finally, in vitro DHA treatment prevented amyloid toxicity in cell cultures. Our findings support the concept that increased DHA consumption may play and important role in reducing brain insults in female AD patients. PMID:19859965

  17. Regulation of Pulmonary and Systemic Bacterial Lipopolysaccharide Responses in Transgenic Mice Expressing Human Elafin

    PubMed Central

    Sallenave, J.-M.; Cunningham, G. A.; James, R. M.; McLachlan, G.; Haslett, C.

    2003-01-01

    The control of lung inflammation is of paramount importance in a variety of acute pathologies, such as pneumonia, the acute respiratory distress syndrome, and sepsis. It is becoming increasingly apparent that local innate immune responses in the lung are negatively influenced by systemic inflammation. This is thought to be due to a local deficit in cytokine responses by alveolar macrophages and neutrophils following systemic bacterial infection and the development of a septic response. Recently, using an adenovirus-based strategy which overexpresses the human elastase inhibitor elafin locally in the lung, we showed that elafin is able to prime lung innate immune responses. In this study, we generated a novel transgenic mouse strain expressing human elafin and studied its response to bacterial lipopolysaccharide (LPS) when the LPS was administered locally in the lungs and systemically. When LPS was delivered to the lungs, we found that mice expressing elafin had lower serum-to-bronchoalveolar lavage ratios of proinflammatory cytokines, including tumor necrosis factor alpha (TNF-α), macrophage inflammatory protein 2, and monocyte chemoattractant protein 1, than wild-type mice. There was a concomitant increase in inflammatory cell influx, showing that there was potential priming of innate responses in the lungs. When LPS was given systemically, the mice expressing elafin had reduced levels of serum TNF-α compared to the levels in wild-type mice. These results indicate that elafin may have a dual function, promoting up-regulation of local lung innate immunity while simultaneously down-regulating potentially unwanted systemic inflammatory responses in the circulation. PMID:12819058

  18. Regulation of pulmonary and systemic bacterial lipopolysaccharide responses in transgenic mice expressing human elafin.

    PubMed

    Sallenave, J-M; Cunningham, G A; James, R M; McLachlan, G; Haslett, C

    2003-07-01

    The control of lung inflammation is of paramount importance in a variety of acute pathologies, such as pneumonia, the acute respiratory distress syndrome, and sepsis. It is becoming increasingly apparent that local innate immune responses in the lung are negatively influenced by systemic inflammation. This is thought to be due to a local deficit in cytokine responses by alveolar macrophages and neutrophils following systemic bacterial infection and the development of a septic response. Recently, using an adenovirus-based strategy which overexpresses the human elastase inhibitor elafin locally in the lung, we showed that elafin is able to prime lung innate immune responses. In this study, we generated a novel transgenic mouse strain expressing human elafin and studied its response to bacterial lipopolysaccharide (LPS) when the LPS was administered locally in the lungs and systemically. When LPS was delivered to the lungs, we found that mice expressing elafin had lower serum-to-bronchoalveolar lavage ratios of proinflammatory cytokines, including tumor necrosis factor alpha (TNF-alpha), macrophage inflammatory protein 2, and monocyte chemoattractant protein 1, than wild-type mice. There was a concomitant increase in inflammatory cell influx, showing that there was potential priming of innate responses in the lungs. When LPS was given systemically, the mice expressing elafin had reduced levels of serum TNF-alpha compared to the levels in wild-type mice. These results indicate that elafin may have a dual function, promoting up-regulation of local lung innate immunity while simultaneously down-regulating potentially unwanted systemic inflammatory responses in the circulation. PMID:12819058

  19. Accelerated fracture healing in transgenic mice overexpressing an anabolic isoform of fibroblast growth factor 2.

    PubMed

    Hurley, Marja M; Adams, Douglas J; Wang, Liping; Jiang, Xi; Burt, Patience Meo; Du, Erxia; Xiao, Liping

    2016-03-01

    The effect of targeted expression of an anabolic isoform of basic fibroblast growth factor (FGF2) in osteoblastic lineage on tibial fracture healing was assessed in mice. Closed fracture of the tibiae was performed in Col3.6-18 kDaFgf2-IRES-GFPsaph mice in which a 3.6 kb fragment of type I collagen promoter (Col3.6) drives the expression of only the 18 kD isoform of FGF2 (18 kDaFgf2/LMW) with green fluorescent protein-sapphire (GFPsaph) as well as Vector mice (Col3.6-IRES-GFPsaph, Vector) that did not harbor the FGF2 transgene. Radiographic, micro-CT, DEXA, and histologic analysis of fracture healing of tibiae harvested at 3, 10 and 20 days showed a smaller fracture callus but accelerated fracture healing in LMWTg compared with Vector mice. At post fracture day 3, FGF receptor 3 and Sox 9 mRNA were significantly increased in LMWTg compared with Vector. Accelerated fracture healing was associated with higher FGF receptor 1, platelet derived growth factors B, C, and D, type X collagen, vascular endothelial cell growth factor, matrix metalloproteinase 9, tartrate resistant acid phosphatase, cathepsin K, runt-related transcription factor-2, Osterix and Osteocalcin and lower Sox9, and type II collagen expression at 10 days post fracture. We postulate that overexpression of LMW FGF2 accelerated the fracture healing process due to its effects on factors that are important in chondrocyte and osteoblast differentiation and vascular invasion. PMID:26252425

  20. Intraductal Delivery of Adenoviruses Targets Pancreatic Tumors in Transgenic Ela-myc Mice and Orthotopic Xenografts

    PubMed Central

    José, Anabel; Sobrevals, Luciano; Camacho-Sánchez, Juan Miguel; Huch, Meritxell; Andreu, Núria; Ayuso, Eduard; Navarro, Pilar; Alemany, Ramon; Fillat, Cristina

    2013-01-01

    Gene-based anticancer therapies delivered by adenoviruses are limited by the poor viral distribution into the tumor. In the current work we have explored the feasibility of targeting pancreatic tumors through a loco-regional route. We have taken advantage of the ductal network in the pancreas to retrogradelly inject adenoviruses through the common bile duct in two different mouse models of pancreatic carcinogenesis: The transgenic Ela-myc mice that develop mixed neoplasms displaying both acinar-like and duct-like neoplastic cells affecting the whole pancreas; and mice bearing PANC-1 and BxPC-3 orthotopic xenografts that constitute a model of localized human neoplastic tumors. We studied tumor targeting and the anticancer effects of newly thymidine kinase-engineered adenoviruses both in vitro and in vivo, and conducted comparative studies between intraductal or intravenous administration. Our data indicate that the intraductal delivery of adenovirus efficiently targets pancreatic tumors in the two mouse models. The in vivo application of AduPARTKT plus ganciclovir (GCV) treatment induced tumor regression in Ela-myc mice. Moreover, the intraductal injection of ICOVIR15-TKT oncolytic adenoviruses significantly improved mean survival of mice bearing PANC-1 and BxPC-3 pancreatic xenografts from 30 to 52 days and from 20 to 68 days respectively (p<0.0001) when combined with GCV. Of notice, both AduPARTKT and ICOVIR15-TKT antitumoral responses were stronger by ductal viral application than intravenously, in line with the 38-fold increase in pancreas transduction observed upon ductal administration. In summary our data show that cytotoxic adenoviruses retrogradelly injected to the pancreas can be a feasible approach to treat localized pancreatic tumors. PMID:23328228

  1. Ventilator-induced lung injury is reduced in transgenic mice that overexpress endothelial nitric oxide synthase.

    PubMed

    Takenaka, Kaori; Nishimura, Yoshihiro; Nishiuma, Teruaki; Sakashita, Akihiro; Yamashita, Tomoya; Kobayashi, Kazuyuki; Satouchi, Miyako; Ishida, Tatsuro; Kawashima, Seinosuke; Yokoyama, Mitsuhiro

    2006-06-01

    Although mechanical ventilation (MV) is an important supportive strategy for patients with acute respiratory distress syndrome, MV itself can cause a type of acute lung damage termed ventilator-induced lung injury (VILI). Because nitric oxide (NO) has been reported to play roles in the pathogenesis of acute lung injury, the present study explores the effects on VILI of NO derived from chronically overexpressed endothelial nitric oxide synthase (eNOS). Anesthetized eNOS-transgenic (Tg) and wild-type (WT) C57BL/6 mice were ventilated at high or low tidal volume (Vt; 20 or 7 ml/kg, respectively) for 4 h. After MV, lung damage, including neutrophil infiltration, water leakage, and cytokine concentration in bronchoalveolar lavage fluid (BALF) and plasma, was evaluated. Some mice were given N(omega)-nitro-L-arginine methyl ester (L-NAME), a potent NOS inhibitor, via drinking water (1 mg/ml) for 1 wk before MV. Histological analysis revealed that high Vt ventilation caused severe VILI, whereas low Vt ventilation caused minimal VILI. Under high Vt conditions, neutrophil infiltration and lung water content were significantly attenuated in eNOS-Tg mice compared with WT animals. The concentrations of macrophage inflammatory protein-2 in BALF and plasma, as well as plasma tumor necrosis factor-alpha and monocyte chemoattractant protein-1, also were decreased in eNOS-Tg mice. L-NAME abrogated the beneficial effect of eNOS overexpression. In conclusion, chronic eNOS overexpression may protect the lung from VILI by inhibiting the production of inflammatory chemokines and cytokines that are associated with neutrophil infiltration into the air space. PMID:16399791

  2. Connexin26 expression in brain parenchymal cells demonstrated by targeted connexin ablation in transgenic mice.

    PubMed

    Nagy, J I; Lynn, B D; Tress, O; Willecke, K; Rash, J E

    2011-07-01

    Astrocytes are known to express the gap junction forming proteins connexin30 (Cx30) and connexin43 (Cx43), but it has remained controversial whether these cells also express connexin26 (Cx26). To investigate this issue further, we examined immunofluorescence labelling of glial connexins in wild-type vs. transgenic mice with targeted deletion of Cx26 in neuronal and glial cells (Cx26fl/fl:Nestin-Cre mice). The Cx26 antibodies utilized specifically recognized Cx26 and lacked cross reaction with highly homologous Cx30, as demonstrated by immunoblotting and immunofluorescence in Cx26-transfected and Cx30-transfected C6 glioma cells. Punctate immunolabelling of Cx26 with these antibodies was observed in leptomeninges and subcortical brain regions. This labelling was absent in subcortical areas of Cx26fl/fl:Nestin-Cre mice, but persisted in leptomeningeal tissues of these mice, thereby distinguishing localization of Cx26 between parenchymal and non-parenchymal tissue. In subcortical brain parenchyma, Cx26-positive puncta were often co-localized with astrocytic Cx43, and some were localized along astrocyte cell bodies and processes immunolabelled for glial fibrillary acidic protein. Cx26-positive puncta were also co-localized with punctate labelling of Cx47 around oligodendrocyte somata. Comparisons of Cx26 labelling in rodent species revealed a lower density of Cx26-positive puncta and a more restricted distribution in subcortical regions of mouse compared with rat brain, perhaps partly explaining reported difficulties in detection of Cx26 in mouse brain parenchyma using antibodies or Cx26 gene reporters. These results support our earlier observations of Cx26 expression in astrocytes and its ultrastructural localization in individual gap junction plaques formed between astrocytes as well as in heterotypic gap junctions between astrocytes and oligodendrocytes. PMID:21714813

  3. Characterization of Transgenic Gfrp Knock-In Mice: Implications for Tetrahydrobiopterin in Modulation of Normal Tissue Radiation Responses

    PubMed Central

    Pathak, Rupak; Pawar, Snehalata A.; Fu, Qiang; Gupta, Prem K.; Berbée, Maaike; Garg, Sarita; Sridharan, Vijayalakshmi; Wang, Wenze; Biju, Prabath G.; Krager, Kimberly J.; Boerma, Marjan; Ghosh, Sanchita P.; Cheema, Amrita K.; Hendrickson, Howard P.; Aykin-Burns, Nukhet

    2014-01-01

    Abstract Aims: The free radical scavenger and nitric oxide synthase cofactor, 5,6,7,8-tetrahydrobiopterin (BH4), plays a well-documented role in many disorders associated with oxidative stress, including normal tissue radiation responses. Radiation exposure is associated with decreased BH4 levels, while BH4 supplementation attenuates aspects of radiation toxicity. The endogenous synthesis of BH4 is catalyzed by the enzyme guanosine triphosphate cyclohydrolase I (GTPCH1), which is regulated by the inhibitory GTP cyclohydrolase I feedback regulatory protein (GFRP). We here report and characterize a novel, Cre-Lox-driven, transgenic mouse model that overexpresses Gfrp. Results: Compared to control littermates, transgenic mice exhibited high transgene copy numbers, increased Gfrp mRNA and GFRP expression, enhanced GFRP–GTPCH1 interaction, reduced BH4 levels, and low glutathione (GSH) levels and differential mitochondrial bioenergetic profiles. After exposure to total body irradiation, transgenic mice showed decreased BH4/7,8-dihydrobiopterin ratios, increased vascular oxidative stress, and reduced white blood cell counts compared with controls. Innovation and Conclusion: This novel Gfrp knock-in transgenic mouse model allows elucidation of the role of GFRP in the regulation of BH4 biosynthesis. This model is a valuable tool to study the involvement of BH4 in whole body and tissue-specific radiation responses and other conditions associated with oxidative stress. Antioxid. Redox Signal. 20, 1436–1446. PMID:23521531

  4. Mimp/Mtch2, an Obesity Susceptibility Gene, Induces Alteration of Fatty Acid Metabolism in Transgenic Mice

    PubMed Central

    Tsarfaty, Galia; Kaufman, Dafna; Horev, Judith; Resau, James H.; Tsarfaty, Ilan

    2016-01-01

    Objective Metabolic dysfunctions, such as fatty liver, obesity and insulin resistance, are among the most common contemporary diseases worldwide, and their prevalence is continuously rising. Mimp/Mtch2 is a mitochondrial carrier protein homologue, which localizes to the mitochondria and induces mitochondrial depolarization. Mimp/Mtch2 single-nucleotide polymorphism is associated with obesity in humans and its loss in mice muscle protects from obesity. Our aim was to study the effects of Mimp/Mtch2 overexpression in vivo. Methods Transgenic mice overexpressing Mimp/Mtch2-GFP were characterized and monitored for lipid accumulation, weight and blood glucose levels. Transgenic mice liver and kidneys were used for gene expression analysis. Results Mimp/Mtch2-GFP transgenic mice express high levels of fatty acid synthase and of β-oxidation genes and develop fatty livers and kidneys. Moreover, high-fat diet–fed Mimp/Mtch2 mice exhibit high blood glucose levels. Our results also show that Mimp/Mtch2 is involved in lipid accumulation and uptake in cells and perhaps in human obesity. Conclusions Mimp/Mtch2 alters lipid metabolism and may play a role in the onset of obesity and development of insulin resistance. PMID:27359329

  5. Neonatal motoneurons overexpressing the bcl-2 protooncogene in transgenic mice are protected from axotomy-induced cell death.

    PubMed Central

    Dubois-Dauphin, M; Frankowski, H; Tsujimoto, Y; Huarte, J; Martinou, J C

    1994-01-01

    In vitro, the overexpression of the bcl-2 protooncogene in cultured neurons has been shown to prevent apoptosis induced by neurotrophic factor deprivation. We have generated transgenic mice overexpressing the Bcl-2 protein in neurons, including motoneurons of the facial nucleus. We have tested whether Bcl-2 could protect these motoneurons from experimentally induced cell death in new born mice. To address this question, we performed unilateral lesion of the facial nerve of wild-type and transgenic 2-day-old mice. In wild-type mice, the lesioned nerve and the corresponding motoneuron cell bodies in the facial nucleus underwent rapid degeneration. In contrast, in transgenic mice, facial motoneurons survived axotomy. Not only their cell bodies but also their axons were protected up to the lesion site. These results demonstrate that in vivo Bcl-2 protects neonatal motoneurons from degeneration after axonal injury. A better understanding of the mechanisms by which Bcl-2 prevents neuronal cell death in vivo could lead to the development of strategies for the treatment of motoneuron degenerative diseases. Images PMID:8159744

  6. Reduced wheel running and blunted effects of voluntary exercise in LPA1-null mice: The importance of assessing the amount of running in transgenic mice studies

    PubMed Central

    Castilla-Ortega, Estela; Rosell-Valle, Cristina; Blanco, Eduardo; Pedraza, Carmen; Chun, Jerold; de Fonseca, Fernando Rodríguez; Estivill-Torrús, Guillermo; Santín, Luis J.

    2014-01-01

    This work was aimed to assess whether voluntary exercise rescued behavioral and hippocampal alterations in mice lacking the lysophosphatidic acid LPA1 receptor (LPA1-null mice), studying the potential relationship between the amount of exercise performed and its effects. Normal and LPA1-null mice underwent 23 days of free wheel running and were tested for open-field behavior and adult hippocampal neurogenesis (cell proliferation, immature neurons, cell survival). Running decreased anxiety-like behavior in both genotypes but increased exploration only in the normal mice. While running affected all neurogenesis-related measures in normal mice (especially in the suprapyramidal blade of the dentate gyrus), only a moderate increase in cell survival was found in the mutants. Importantly, the LPA1-nulls showed notably reduced running. Analysis suggested that defective running in the LPA1-null mice could contribute to explain the scarce benefit of the voluntary exercise treatment. On the other hand, a literature review revealed that voluntary exercise is frequently used to modulate behavior and the hippocampus in transgenic mice, but half of the studies did not assess the quantity of running, overlooking any potential running impairments. This study adds evidence to the relevance of the quantity of exercise performed, emphasizing the importance of its assessment in transgenic mice research. PMID:24055600

  7. Receptor-specific in vivo desensitization by the G protein-coupled receptor kinase-5 in transgenic mice.

    PubMed Central

    Rockman, H A; Choi, D J; Rahman, N U; Akhter, S A; Lefkowitz, R J; Koch, W J

    1996-01-01

    Transgenic mice were generated with cardiac-specific overexpression of the G protein-coupled receptor kinase-5 (GRK5), a serine/threonine kinase most abundantly expressed in the heart compared with other tissues. Animals overexpressing GRK5 showed marked beta-adrenergic receptor desensitization in both the anesthetized and conscious state compared with nontransgenic control mice, while the contractile response to angiotensin II receptor stimulation was unchanged. In contrast, the angiotensin II-induced rise in contractility was significantly attenuated in transgenic mice overexpressing the beta-adrenergic receptor kinase-1, another member of the GRK family. These data suggest that myocardial overexpression of GRK5 results in selective uncoupling of G protein-coupled receptors and demonstrate that receptor specificity of the GRKs may be important in determining the physiological phenotype. Images Fig. 1 PMID:8790438

  8. Combined Micro-PET/Micro-CT Imaging of Lung Tumours in SPC-raf and SPC-myc Transgenic Mice

    PubMed Central

    Rodt, Thomas; Luepke, Matthias; Boehm, Claudia; Hueper, Katja; Halter, Roman; Glage, Silke; Hoy, Ludwig; Wacker, Frank; Borlak, Juergen; von Falck, Christian

    2012-01-01

    Introduction SPC-raf and SPC-myc transgenic mice develop disseminated and circumscribed lung adenocarcinoma respectively, allowing for assessment of carcinogenesis and treatment strategies. The purpose of this study was to investigate the technical feasibility, the correlation of initial findings to histology and the administered radiation dose of combined micro-PET/micro-CT in these animal models. Material and Methods 14 C57BL/6 mice (4 nontransgenic, 4 SPC-raf transgenic, 6 SPC-myc transgenic) were examined using micro-CT and 18F-Fluoro-deoxyglucose micro-PET in-vivo. Micro-PET data was corrected for random events and scatter prior to reconstruction with a 3D-FORE/2D-OSEM iterative algorithm. Rigid micro-PET/micro-CT registration was performed. Tumour-to-non-tumour ratios were calculated for different lung regions and focal lesions. Diffuse tumour growth was quantified using a semiautomated micro-CT segmentation routine reported earlier. Regional histologic tumour load was assessed using a 4-point rating scale. Gamma radiation dose was determined using thermoluminescence dosimeters. Results Micro-CT allowed visualisation of diffuse and circumscribed tumours in SPC-raf and SPC-myc transgenic animals along with morphology, while micro-PET provided information on metabolism, but lacked morphologic detail. Mean tumour-to-non-tumour ratio was 2.47 for circumscribed lesions. No significant correlation could be shown between histological tumour load and tumour-to-nontumour ratio for diffuse tumours in SPC-raf transgenic animals. Calculation of the expected dose based on gamma dosimetry yielded approximately 140 mGy/micro-PET examination additional to approximately 200 mGy due to micro-CT. Conclusions Combined micro-PET/micro-CT imaging allows for in-vivo assessment of lung tumours in SPC-raf and SPC-myc transgenic mice. The technique has potential for the evaluation of carcinogenesis and treatment strategies in circumscribed lung tumours. PMID:23028537

  9. Targeted expression of SV40 T antigen in the hair follicle of transgenic mice produces an aberrant hair phenotype.

    PubMed

    Keough, R; Powell, B; Rogers, G

    1995-03-01

    Directed expression of SV40 large T antigen (TAg) in transgenic mice can induce tissue-specific tumorigenesis and useful cell lines exhibiting differentiated characteristics can be established from resultant tumor cells. In an attempt to produce an immortalised mouse hair follicle cortical cell line for the study of hair keratin gene control, SV40 TAg expression was targeted to the hair follicles of transgenic mice using a sheep hair gene promoter. Expression of SV40 TAg in the follicle cortex disrupted normal fiber ultrastructure, producing a marked phenotypic effect. Affected hairs were wavy or severely kinked (depending on the severity of the phenotype) producing an appearance ranging from a ruffled coat to a stubble covering the back of the mouse. The transgenic hairs appeared to be weakened at the base of the fibers, leading to premature hair-loss and a thinner pelage, or regions of temporary nudity. No follicle tumors or neoplasia were apparent and immortalisation of cortical cells could not be established in culture. In situ hybridisation studies in the hair follicle using histone H3 as a cell proliferation marker suggested that cell proliferation had ceased prior to commencement of K2.10-TAg expression and was not re-established in the differentiating cortical cells. Hence, TAg was unable to induce cell immortalisation at that stage of cortical cell differentiation. However, transgenic mice developed various other abnormalities including vertebral abnormalities and bladder, liver and intestinal tumors, which resulted in reduced life expectancy. PMID:7542671

  10. Studies in transgenic mice reveal potential relationships between secretin-producing cells and other endocrine cell types.

    PubMed

    Lopez, M J; Upchurch, B H; Rindi, G; Leiter, A B

    1995-01-13

    We have produced transgenic mice expressing fusion genes consisting of 1.6 kilobase pairs of the secretin gene 5' flanking region to direct the expression of human growth hormone (hGH) or simian virus 40 large T antigen to secretin-producing cells. Analysis of different mouse tissues for hGH transcripts revealed expression in each of the major secretin-producing tissues, namely the intestine and endocrine pancrease. Multiple label immunohistochemistry demonstrated that the transgene was correctly directed to secretin cells in the intestinal tract, including a previously unrecognized population of secretin cells in the colon of adult and developing mice. In the small intestine, subpopulations of hGH-containing cells frequently coexpressed substance P, serotonin, and cholecystokinin, whereas in the colon, cells expressing hGH frequently coexpressed glucagon, peptide YY, or neurotensin. Transgenic mice expressing large T antigen in secretin cells developed poorly differentiated neuroendocrine tumors of the small intestine, well differentiated colonic tumors containing glucagon-expressing cells, and insulin-producing tumors in pancreas. These studies indicate that the major cis-regulatory sequences necessary for secretin expression in enteroendocrine cells and fetal islets are localized with 1.6 kilobase pairs of the transcriptional start site. Coexpression of reporter transgenes with several gastrointestinal hormones suggests a potential relationships between secretin cells and other enteroendocrine cell types, as well as pancreatic beta cells. PMID:7822327

  11. Visualization of the Epiblast and Visceral Endodermal Cells Using Fgf5-P2A-Venus BAC Transgenic Mice and Epiblast Stem Cells

    PubMed Central

    Tsukiyama, Tomoyuki; Matsushita, Jun; Tsukiyama-Fujii, Setsuko; Takahashi, Satoru; Ema, Masatsugu

    2016-01-01

    Fibroblast growth factor 5 (Fgf5) has been widely used as a marker for the epiblast in the postimplantation embryo and epiblast stem cells (mEpiSCs) in the mouse, making it valuable for study of differentiation of various tissues and epiblast cells in vivo and in vitro. Here, we report for the first time the generation of Fgf5-P2A-Venus BAC transgenic (Tg) mice and show that the BAC Tg can recapitulate endogenous Fgf5 expression in epiblast and visceral endodermal cells of E6.5 and 7.5 embryos. We also show that Fgf5-P2A-Venus BAC Tg mEpiSCs in the undifferentiated state expressed abundant Venus, and upon reprogramming into naïve state, Venus was suppressed. Furthermore, while most Tg mEpiSCs expressed Venus abundantly, surprisingly the Tg mEpiSCs contained a minor subpopulation of Venus-negative cells that were capable of conversion to Venus-positive cells, indicating that even Fgf5 expression shows dynamic heterogeneity in mEpiSCs. Taken together, Fgf5-P2A-Venus BAC Tg mice and mEpiSCs generated in this study will be useful for developmental biology as well as stem cell biology research. PMID:27409080

  12. Chronic administration of R-flurbiprofen attenuates learning impairments in transgenic amyloid precursor protein mice

    PubMed Central

    Kukar, Thomas; Prescott, Sonya; Eriksen, Jason L; Holloway, Vallie; Murphy, M Paul; Koo, Edward H; Golde, Todd E; Nicolle, Michelle M

    2007-01-01

    ability to selectively target Aβ42 production and improve cognitive impairments in transgenic APP mice, as well as promising data from a phase 2 human clinical trial, future studies are needed to investigate the utility of R-flurbiprofen as an AD therapeutic and its possible mechanisms of action. PMID:17650315

  13. Impairment of T cell development and acute inflammatory response in HIV-1 Tat transgenic mice

    PubMed Central

    Fiume, Giuseppe; Scialdone, Annarita; Albano, Francesco; Rossi, Annalisa; Maria Tuccillo, Franca; Rea, Domenica; Palmieri, Camillo; Caiazzo, Elisabetta; Cicala, Carla; Bellevicine, Claudio; Falcone, Cristina; Vecchio, Eleonora; Pisano, Antonio; Ceglia, Simona; Mimmi, Selena; Iaccino, Enrico; Laurentiis, Annamaria de; Pontoriero, Marilena; Agosti, Valter; Troncone, Giancarlo; Mignogna, Chiara; Palma, Giuseppe; Arra, Claudio; Mallardo, Massimo; Maria Buonaguro, Franco; Scala, Giuseppe; Quinto, Ileana

    2015-01-01

    Immune activation and chronic inflammation are hallmark features of HIV infection causing T-cell depletion and cellular immune dysfunction in AIDS. Here, we addressed the issue whether HIV-1 Tat could affect T cell development and acute inflammatory response by generating a transgenic mouse expressing Tat in lymphoid tissue. Tat-Tg mice showed thymus atrophy and the maturation block from DN4 to DP thymic subpopulations, resulting in CD4+ and CD8+ T cells depletion in peripheral blood. In Tat-positive thymus, we observed the increased p65/NF-κB activity and deregulated expression of cytokines/chemokines and microRNA-181a-1, which are involved in T-lymphopoiesis. Upon LPS intraperitoneal injection, Tat-Tg mice developed an abnormal acute inflammatory response, which was characterized by enhanced lethality and production of inflammatory cytokines. Based on these findings, Tat-Tg mouse could represent an animal model for testing adjunctive therapies of HIV-1-associated inflammation and immune deregulation. PMID:26343909

  14. Lactobacillus plantarum NCIMB8826 ameliorates inflammation of colon and skin in human APOC1 transgenic mice.

    PubMed

    Mariman, R; Reefman, E; Tielen, F; Persoon-Deen, C; van de Mark, K; Worms, N; Koning, F; Nagelkerken, L

    2016-03-11

    Genetic predisposition and environmental factors, including the gut microbiota, have been suggested as major factors in the development and progression of atopic dermatitis. Hyperlipidemic human APOC1(+/+) transgenic mice display many features of human atopic dermatitis, such as scaling, lichenification, excoriations, and pruritus, along with a disturbed skin barrier function. Cytokine analysis of serum shows an increase of various pro-inflammatory cytokines, including interleukin (IL)-12p40, IL-6, and IL-1α, but lower levels of interferon-γ. These mice also display aspects of colitis evident from macroscopic and histological abnormalities. Genome-wide transcriptome analysis of the intestine shows up-regulation of several genes associated with mast cells and eosinophils and this observation was confirmed by demonstrating increased numbers of IgE(+) and FcRε(+) mast cells in the colon and in the skin. Oral treatment with Lactobacillus plantarum NCIMB8826 resulted in decreased numbers of mast cells in the colon. Moreover, this L. plantarum strain ameliorated skin pathology, evident from improved skin barrier integrity, absence of skin thickening, and less excoriations. These results suggest that modulation of intestinal immune homeostasis contributes to the suppression of atopic dermatitis. PMID:26689228

  15. Impairment of T cell development and acute inflammatory response in HIV-1 Tat transgenic mice.

    PubMed

    Fiume, Giuseppe; Scialdone, Annarita; Albano, Francesco; Rossi, Annalisa; Tuccillo, Franca Maria; Rea, Domenica; Palmieri, Camillo; Caiazzo, Elisabetta; Cicala, Carla; Bellevicine, Claudio; Falcone, Cristina; Vecchio, Eleonora; Pisano, Antonio; Ceglia, Simona; Mimmi, Selena; Iaccino, Enrico; de Laurentiis, Annamaria; Pontoriero, Marilena; Agosti, Valter; Troncone, Giancarlo; Mignogna, Chiara; Palma, Giuseppe; Arra, Claudio; Mallardo, Massimo; Buonaguro, Franco Maria; Scala, Giuseppe; Quinto, Ileana

    2015-01-01

    Immune activation and chronic inflammation are hallmark features of HIV infection causing T-cell depletion and cellular immune dysfunction in AIDS. Here, we addressed the issue whether HIV-1 Tat could affect T cell development and acute inflammatory response by generating a transgenic mouse expressing Tat in lymphoid tissue. Tat-Tg mice showed thymus atrophy and the maturation block from DN4 to DP thymic subpopulations, resulting in CD4(+) and CD8(+) T cells depletion in peripheral blood. In Tat-positive thymus, we observed the increased p65/NF-κB activity and deregulated expression of cytokines/chemokines and microRNA-181a-1, which are involved in T-lymphopoiesis. Upon LPS intraperitoneal injection, Tat-Tg mice developed an abnormal acute inflammatory response, which was characterized by enhanced lethality and production of inflammatory cytokines. Based on these findings, Tat-Tg mouse could represent an animal model for testing adjunctive therapies of HIV-1-associated inflammation and immune deregulation. PMID:26343909

  16. Amniotic Fluid Stem Cells from EGFP Transgenic Mice Attenuate Hyperoxia-Induced Acute Lung Injury

    PubMed Central

    Lai, Cheng-Wei; Yen, Chih-Ching; Lee, Kun-Hsiung; Wu, Shinn-Chih; Chen, Chuan-Mu

    2013-01-01

    High concentrations of oxygen aggravate the severity of lung injury in patients requiring mechanical ventilation. Although mesenchymal stem cells have been shown to effectively attenuate various injured tissues, there is limited information regarding a role for amniotic fluid stem cells (AFSCs) in treating acute lung injury. We hypothesized that intravenous delivery of AFSCs would attenuate lung injury in an experimental model of hyperoxia-induced lung injury. AFSCs were isolated from EGFP transgenic mice. The in vitro differentiation, surface markers, and migration of the AFSCs were assessed by specific staining, flow cytometry, and a co-culture system, respectively. The in vivo therapeutic potential of AFSCs was evaluated in a model of acute hyperoxia-induced lung injury in mice. The administration of AFSCs significantly reduced the hyperoxia-induced pulmonary inflammation, as reflected by significant reductions in lung wet/dry ratio, neutrophil counts, and the level of apoptosis, as well as reducing the levels of inflammatory cytokine (IL-1β, IL-6, and TNF-α) and early-stage fibrosis in lung tissues. Moreover, EGFP-expressing AFSCs were detected and engrafted into a peripheral lung epithelial cell lineage by fluorescence microscopy and DAPI stain. Intravenous administration of AFSCs may offer a new therapeutic strategy for acute lung injury (ALI), for which efficient treatments are currently unavailable. PMID:24040409

  17. Late onset distal axonal swelling in YFP-H transgenic mice.

    PubMed

    Bridge, Katherine E; Berg, Nicola; Adalbert, Robert; Babetto, Elisabetta; Dias, Tatyana; Spillantini, Maria-Grazia; Ribchester, Richard R; Coleman, Michael P

    2009-02-01

    Axonal swellings, or spheroids, are a feature of central nervous system (CNS) axon degeneration during normal aging and in many disorders. The direct cause and mechanism are unknown. The use of transgenic mouse line YFP-H, which expresses yellow-fluorescent protein (YFP) in a subset of neurons, greatly facilitates longitudinal imaging and live imaging of axonal swellings, but it has not been established whether long-term expression of YFP itself contributes to axonal swelling. Using conventional methods to compare YFP-H mice with their YFP negative littermates, we found an age-related increase in swellings in discrete CNS regions in both genotypes, but the presence of YFP caused significantly more swellings in mice aged 8 months or over. Increased swelling was found in gracile tract, gracile nucleus and dorsal roots but not in lateral columns, olfactory bulb, motor cortex, ventral roots or peripheral nerve. Thus, long-term expression of YFP accelerates age-related axonal swelling in some axons and data reliant on the presence of YFP in these CNS regions in older animals needs to be interpreted carefully. The ability of a foreign protein to exacerbate age-related axon pathology is an important clue to the mechanisms by which such pathology can arise. PMID:17658198

  18. Maternal dexamethasone exposure ameliorates cognition and tau pathology in the offspring of triple transgenic AD mice.

    PubMed

    Di Meco, A; Joshi, Y B; Lauretti, E; Praticò, D

    2016-03-01

    Dysregulation of stress hormones, such as glucocorticoids, in adult life increases the risk to develop Alzheimer's disease (AD). However, the effect of prenatal glucocorticoids exposure on AD development in the offspring remains unknown. We studied how gestational dexamethasone exposure influences the AD-like phenotype in the offspring of triple transgenic AD mice (3 × Tg). To this end, female mice received dexamethasone or vehicle during the entire pregnancy time in the drinking water. Offspring from vehicle-treated 3 × Tg (controls) were compared with offspring from dexamethasone-treated 3 × Tg later in life for their memory, learning ability and brain pathology. Compared with controls, offspring from dexamethasone-treated mothers displayed improvement in their memory as assessed by fear conditioning test, both in the cue and recall phases. The same animals had a significant reduction in the insoluble fraction of tau, which was associated with an increase in autophagy. In addition, they showed an activation of the transcription factor cellular response element-binding protein and an increase in brain-derived neurotrophic factor and c-FOS protein levels, key regulators of synaptic plasticity and memory. We conclude that dexamethasone exposure during pregnancy provides long-lasting protection against the onset and development of the AD-like phenotype by improving cognition and tau pathology. PMID:26077691

  19. Non-invasive fluorescent imaging of gliosis in transgenic mice for profiling developmental neurotoxicity

    SciTech Connect

    Ho, Gideon; Zhang Chunyan; Zhuo Lang . E-mail: lzhuo@ibn.a-star.edu.sg

    2007-05-15

    Gliosis is a universal response of Brain to almost all types of neural insults, including neurotoxicity, neurodegeneration, viral infection, and stroke. A hallmark of gliotic reaction is the up-regulation of the astrocytic biomarker GFAP (glial fibrillary acidic protein), which often precedes the anatomically apparent damages in Brain. In this study, neonatal transgenic mice at postnatal day (PD) 4 expressing GFP (green fluorescent protein) under the control of a widely used 2.2-kb human GFAP promoter in Brain are treated with two model neurotoxicants, 1-methyl-4(2'-methylphenyl)-1,2,3,6-tetrahydropyridine (2'-CH{sub 3}-MPTP), and kainic acid (KA), respectively, to induce gliosis. Here we show that the neurotoxicant-induced acute gliosis can be non-invasively imaged and quantified in Brain of conscious (un-anesthetized) mice in real-time, at 0, 2, 4, 6, and 8 h post-toxicant dosing. Therefore the current methodology could be a useful tool for studying the developmental aspects of neuropathies and neurotoxicity.

  20. Estrogen and progesterone receptors have distinct roles in the establishment of the hyperplastic phenotype in PR-A transgenic mice

    SciTech Connect

    Simian, Marina; Bissell, Mina J.; Barcellos-Hoff, Mary Helen; Shyamala, Gopalan

    2009-05-11

    Expression of the A and B forms of progesterone receptor (PR) in an appropriate ratio is critical for mammary development. Mammary glands of PR-A transgenic mice, carrying an additional A form of PR as a transgene, exhibit morphological features associated with the development of mammary tumors. Our objective was to determine the roles of estrogen (E) and progesterone (P) in the genesis of mammary hyperplasias/preneoplasias in PR-A transgenics. We subjected PR-A mice to hormonal treatments and analyzed mammary glands for the presence of hyperplasias and used BrdU incorporation to measure proliferation. Quantitative image analysis was carried out to compare levels of latency-associated peptide and transforming growth factor beta 1 (TGF{beta}1) between PR-A and PR-B transgenics. Basement membrane disruption was examined by immunofluorescence and proteolytic activity by zymography. The hyperplastic phenotype of PR-A transgenics is inhibited by ovariectomy, and is reversed by treatment with E + P. Studies using the antiestrogen ICI 182,780 or antiprogestins RU486 or ZK 98,299 show that the increase in proliferation requires signaling through E/estrogen receptor alpha but is not sufficient to give rise to hyperplasias, whereas signaling through P/PR has little impact on proliferation but is essential for the manifestation of hyperplasias. Increased proliferation is correlated with decreased TGF{beta}1 activation in the PR-A transgenics. Analysis of basement membrane integrity showed loss of laminin-5, collagen III and collagen IV in mammary glands of PR-A mice, which is restored by ovariectomy. Examination of matrix metalloproteases (MMPs) showed that total levels of MMP-2 correlate with the steady-state levels of PR, and that areas of laminin-5 loss coincide with those of activation of MMP-2 in PR-A transgenics. Activation of MMP-2 is dependent on treatment with E and P in ovariectomized wild-type mice, but is achieved only by treatment with P in PR-A mice. These data

  1. Expression of human hormone-sensitive lipase in white adipose tissue of transgenic mice increases lipase activity but does not enhance in vitro lipolysis.

    PubMed

    Lucas, Stéphanie; Tavernier, Geneviève; Tiraby, Claire; Mairal, Aline; Langin, Dominique

    2003-01-01

    Hormone-sensitive lipase (HSL) catalyzes the hydrolysis of acylglycerols and cholesteryl esters (CEs). The enzyme is highly expressed in adipose tissues (ATs), where it is thought to play an important role in fat mobilization. The purpose of the present work was to study the effect of a physiological increase of HSL expression in vivo. Transgenic mice were produced with a 21 kb human genomic fragment encompassing the exons encoding the adipocyte form of HSL. hHSL mRNA was expressed at 3-fold higher levels than murine HSL mRNA in white adipocytes. Transgene expression was also observed in brown adipose tissue (BAT) and skeletal muscle. The human protein was detected in ATs of transgenic (Tg) mice. The hydrolytic activities against triacylglycerol (TG), diacylglycerol (DG) analog, and CE were increased in transgenic mouse AT. However, cAMP-inducible adipocyte lipolysis was lower in transgenic animals. In the B6CBA genetic background, transgenic mice up to 14 weeks of age showed lower body weight and fat mass. The phenotype was not observed in older animals and in mice fed a high-fat diet (HFD). In the OF1 genetic background, there was no difference in fat mass of mice fed ad libitum. However, transgenic mice became leaner than their wild-type (WT) littermates after a 4 day calorie restriction. The data show that overexpression of HSL, despite increased lipase activity, does not lead to enhanced lipolysis. PMID:12518034

  2. Decreased glial and synaptic glutamate uptake in the striatum of HIV-1 gp120 transgenic mice.

    PubMed

    Melendez, Roberto I; Roman, Cristina; Capo-Velez, Coral M; Lasalde-Dominicci, Jose A

    2016-06-01

    The mechanisms leading to the neurocognitive deficits in humans with immunodeficiency virus type 1 (HIV-1) are not well resolved. A number of cell culture models have demonstrated that the HIV-envelope glycoprotein 120 (gp120) decreases the reuptake of glutamate, which is necessary for learning, memory, and synaptic plasticity. However, the impact of brain HIV-1 gp120 on glutamate uptake systems in vivo remains unknown. Notably, alterations in brain glutamate uptake systems are implicated in a number of neurodegenerative and neurocognitive disorders. We characterized the kinetic properties of system XAG (sodium-dependent) and systems xc- (sodium-independent) [3H]-L-glutamate uptake in the striatum and hippocampus of HIV-1 gp120 transgenic mice, an established model of HIV neuropathology. We determined the kinetic constant Vmax (maximal velocity) and Km (affinity) of both systems XAG and xc- using subcellular preparations derived from neurons and glial cells. We show significant (30-35 %) reductions in the Vmax of systems XAG and xc- in both neuronal and glial preparations derived from the striatum, but not from the hippocampus of gp120 mice relative to wild-type (WT) controls. Moreover, immunoblot analysis showed that the protein expression of glutamate transporter subtype-1 (GLT-1), the predominant brain glutamate transporter, was significantly reduced in the striatum but not in the hippocampus of gp120 mice. These extensive and region-specific deficits of glutamate uptake likely contribute to the development and/or severity of HIV-associated neurocognitive disorders. Understanding the role of striatal glutamate uptake systems in HIV-1 gp120 may advance the development of new therapeutic strategies to prevent neuronal damage and improve cognitive function in HIV patients. PMID:26567011

  3. Prediabetes Linked to Excess Glucagon in Transgenic Mice with Pancreatic Active AKT1

    PubMed Central

    Albury-Warren, Toya M.; Pandey, Veethika; Spinel, Lina P.; Masternak, Michal M.; Altomare, Deborah A

    2015-01-01

    Protein Kinase B/AKT, has three isoforms (AKT1-3) and is renowned for its central role in the regulation of cell growth and proliferation, due to its constitutive activation in various cancers. AKT2, which is highly expressed in insulin responsive tissues, has been identified as a primary regulator of glucose metabolism as Akt2 knockout mice (Akt2−/−) are glucose intolerant and insulin resistant. However, the role of AKT1 in glucose metabolism is not as clearly defined. We previously showed that mice with myristoylated Akt1 (AKT1Myr) expressed through a bicistronic Pdx1-TetA and TetO-MyrAkt1 system were susceptible to islet cell carcinomas, and in this study we characterized an early onset, prediabetic phenotype. Beginning at weaning (3 weeks of age), the glucose intolerant AKT1Myr mice exhibited non-fasted hyperglycemia, which progressed to fasted hyperglycemia by 5 months of age. The glucose intolerance was attributed to a fasted hyperglucagonemia, and hepatic insulin resistance detectable by reduced phosphorylation of the insulin receptor following insulin injection into the inferior vena cava. In contrast, treatment with doxycycline diet to turn-off the transgene, caused attenuation of the non-fasted and fasted hyperglycemia, thus affirming AKT1 hyperactivation as the trigger. Collectively, this model highlights a novel glucagon-mediated mechanism by which AKT1 hyperactivation affects glucose homeostasis, and provides an avenue to better delineate the molecular mechanisms responsible for diabetes mellitus and the potential association with pancreatic cancer. PMID:26487674

  4. Cytochrome P450 2J2 Is Protective against Global Cerebral Ischemia in Transgenic Mice

    PubMed Central

    Li, Rui; Xu, Xizhen; Chen, Chen; Yu, Xuefeng; Edin, Matthew L.; Degraff, Laura Miller; Lee, Craig R.; Zeldin, Darryl C.; Wang, Dao Wen

    2012-01-01

    Background and Purpose Cytochrome P450 epoxygenase metabolites of arachidonic acid (EETs) have multiple cardiovascular effects, including reduction of blood pressure, protection against myocardial ischemia-reperfusion injury, and attenuation of endothelial inflammation and apoptosis. The present study was aimed to determine potential neuroprotective roles for EETs in cerebral ischemia. Methods Transgenic mice with endothelial overexpression of CYP2J2 (Tie2-CYP2J2-Tr) were subjected to global cerebral ischemia induced by bilateral common carotid artery occlusion (BCCAO) for 10 minutes, Cerebral EET production, infarct size, and apoptosis were examined after 24 hours of reperfusion. The action mechanisms of EETs on cerebral ischemia was also studied in cultures of astrocytes and Neuro-2a cells exposed to oxygen-glucose deprivation (OGD). Results In Tie2-CYP2J2-Tr mice, CYP2J2 expression and 14, 15-EET production in both brain tissue and plasma significantly increased while brain infarct size and apoptosis after ischemia decreased, accompanied increased activation of the PI3K/AKT and ERK1/2 pathways, decreased activation of JNK, and higher ratios of Bcl-2/Bax and Bcl-xl/Bax in ischemic brain compared to wild type mice. In cells, addition of exogenous EETs or CYP2J2 transfection attenuated OGD-induced apoptosis by activation of ERK1/2 and PI3K/AKT pathways, inhibition of JNK, which were reduced by pretreatments with inhibitors of the PI3K (LY294002), the MAPK (PD98059) and EETs (EEZE), respectively. Conclusions We conclude that CYP2J2 overexpression exerts marked neuroprotective effects against ischemic injury by a mechanism linked to increased level of circulating EETs and reduction of apoptosis. These data suggests the possibility for clinical therapy of cerebral ischemia by enhancing EET levels. PMID:23041291

  5. Prediabetes linked to excess glucagon in transgenic mice with pancreatic active AKT1.

    PubMed

    Albury-Warren, Toya M; Pandey, Veethika; Spinel, Lina P; Masternak, Michal M; Altomare, Deborah A

    2016-01-01

    Protein kinase B/AKT has three isoforms (AKT1-3) and is renowned for its central role in the regulation of cell growth and proliferation, due to its constitutive activation in various cancers. AKT2, which is highly expressed in insulin-responsive tissues, has been identified as a primary regulator of glucose metabolism as Akt2 knockout mice (Akt2(-/-)) are glucose-intolerant and insulin-resistant. However, the role of AKT1 in glucose metabolism is not as clearly defined. We previously showed that mice with myristoylated Akt1 (AKT1(Myr)) expressed through a bicistronic Pdx1-TetA and TetO-MyrAkt1 system were susceptible to islet cell carcinomas, and in this study we characterized an early onset, prediabetic phenotype. Beginning at weaning (3 weeks of age), the glucose-intolerant AKT1(Myr) mice exhibited non-fasted hyperglycemia, which progressed to fasted hyperglycemia by 5 months of age. The glucose intolerance was attributed to a fasted hyperglucagonemia, and hepatic insulin resistance detectable by reduced phosphorylation of the insulin receptor following insulin injection into the inferior vena cava. In contrast, treatment with doxycycline diet to turn off the transgene caused attenuation of the non-fasted and fasted hyperglycemia, thus affirming AKT1 hyperactivation as the trigger. Collectively, this model highlights a novel glucagon-mediated mechanism by which AKT1 hyperactivation affects glucose homeostasis and provides an avenue to better delineate the molecular mechanisms responsible for diabetes mellitus and the potential association with pancreatic cancer. PMID:26487674

  6. Copper and Zinc Metallation Status of Copper Zinc Superoxide Dismutase form Amyotrophic Lateral Sclerosis Transgenic Mice

    SciTech Connect

    Lelie, H.L.; Miller, L.; Liba, A.; Bourassa, M.W.; Chattopadhyay, M.; Chan, P.K.; Gralla, E.B.; Borchelt, D.R.; et al

    2010-09-24

    Mutations in the metalloenzyme copper-zinc superoxide dismutase (SOD1) cause one form of familial amyotrophic lateral sclerosis (ALS), and metals are suspected to play a pivotal role in ALS pathology. To learn more about metals in ALS, we determined the metallation states of human wild-type or mutant (G37R, G93A, and H46R/H48Q) SOD1 proteins from SOD1-ALS transgenic mice spinal cords. SOD1 was gently extracted from spinal cord and separated into insoluble (aggregated) and soluble (supernatant) fractions, and then metallation states were determined by HPLC inductively coupled plasma MS. Insoluble SOD1-rich fractions were not enriched in copper and zinc. However, the soluble mutant and WT SOD1s were highly metallated except for the metal-binding-region mutant H46R/H48Q, which did not bind any copper. Due to the stability conferred by high metallation of G37R and G93A, it is unlikely that these soluble SOD1s are prone to aggregation in vivo, supporting the hypothesis that immature nascent SOD1 is the substrate for aggregation. We also investigated the effect of SOD1 overexpression and disease on metal homeostasis in spinal cord cross-sections of SOD1-ALS mice using synchrotron-based x-ray fluorescence microscopy. In each mouse genotype, except for the H46R/H48Q mouse, we found a redistribution of copper between gray and white matters correlated to areas of high SOD1. Interestingly, a disease-specific increase of zinc was observed in the white matter for all mutant SOD1 mice. Together these data provide a picture of copper and zinc in the cell as well as highlight the importance of these metals in understanding SOD1-ALS pathology.

  7. Tg-SwDI Transgenic Mice Exhibit Novel Alterations in AβPP Processing, Aβ Degradation, and Resilient Amyloid Angiopathy

    PubMed Central

    Van Vickle, Gregory D.; Esh, Chera L.; Daugs, Ian D.; Kokjohn, Tyler A.; Kalback, Walter M.; Patton, R. Lyle; Luehrs, Dean C.; Walker, Douglas G.; Lue, Lih-Fen; Beach, Thomas G.; Davis, Judianne; Van Nostrand, William E.; Castaño, Eduardo M.; Roher, Alex E.

    2008-01-01

    Alzheimer’s disease (AD) is characterized by the accumulation of extracellular insoluble amyloid, primarily derived from polymerized amyloid-β (Aβ) peptides. We characterized the chemical composition of the Aβ peptides deposited in the brain parenchyma and cerebrovascular walls of triple transgenic Tg-SwDI mice that produce a rapid and profuse Aβ accumulation. The processing of the N- and C-terminal regions of mutant AβPP differs substantially from humans because the brain parenchyma accumulates numerous, diffuse, nonfibrillar plaques, whereas the thalamic microvessels harbor overwhelming amounts of compact, fibrillar, thioflavine-S- and apolipoprotein E-positive amyloid deposits. The abundant accretion of vascular amyloid, despite low AβPP transgene expression levels, suggests that inefficient Aβ proteolysis because of conformational changes and dimerization may be key pathogenic factors in this animal model. The disruption of amyloid plaque cores by immunotherapy is accompanied by increased perivascular deposition in both humans and transgenic mice. This analogous susceptibility and response to the disruption of amyloid deposits suggests that Tg-SwDI mice provide an excellent model in which to study the functional aftermath of immunotherapeutic interventions. These mice might also reveal new avenues to promote amyloidogenic AβPP processing and fundamental insights into the faulty degradation and clearance of Aβ in AD, pivotal issues in understanding AD pathophysiology and the assessment of new therapeutic agents. PMID:18599612

  8. Ectopic expression of the agouti gene in transgenic mice causes obesity, features of type II diabetes, and yellow fur

    SciTech Connect

    Klebig, M.L.; Woychik, R.P.; Wilkinson, J.E.; Geisler, J.G. |

    1995-05-23

    Mice that carry the lethal yellow (A{sup y}) or viable yellow (A{sup vy}) mutation, two dominant mutations of the agouti (a) gene in mouse chromosome 2, exhibit a phenotype that includes yellow fur, marked obesity, a form of type II diabetes associated with insulin resistance, and an increased susceptibility to tumor development. Molecular analyses of these and several other dominant {open_quotes}obese yellow{close_quotes} a-locus mutations suggested that ectopic expression of the normal agouti protein gives rise to this complex pleiotropic phenotype. We have now tested this hypothesis directly by generating transgenic mice that ectopically express an agouti cDNA clone encoding the normal agouti protein in all tissues examined. Transgenic mice of both sexes have yellow fur, become obese, and develop hyperinsulinemia. In addition, male transgenic mice develop hyperglycemia by 12-20 weeks of age. These results demonstrate conclusively that the ectopic agouti expression is responsible for most, if not all, of the phenotypic traits of the dominant, obese yellow mutants. 42 refs., 5 figs.

  9. Embryonic Lethality in Homozygous Human Her-2 Transgenic Mice Due to Disruption of the Pds5b Gene

    PubMed Central

    Yong, Carmen S. M.; Sharkey, Janelle; Duscio, Belinda; Venville, Ben; Wei, Wei-Zen; Jones, Richard F.; Slaney, Clare Y.; Mir Arnau, Gisela; Papenfuss, Anthony T.

    2015-01-01

    The development of antigen-targeted therapeutics is dependent on the preferential expression of tumor-associated antigens (TAA) at targetable levels on the tumor. Tumor-associated antigens can be generated de novo or can arise from altered expression of normal basal proteins, such as the up-regulation of human epidermal growth factor receptor 2 (Her2/ErbB2). To properly assess the development of Her2 therapeutics in an immune tolerant model, we previously generated a transgenic mouse model in which expression of the human Her2 protein was present in both the brain and mammary tissue. This mouse model has facilitated the development of Her2 targeted therapies in a clinically relevant and suitable model. While heterozygous Her2+/- mice appear to develop in a similar manner to wild type mice (Her2-/-), it has proven difficult to generate homozygous Her2+/+ mice, potentially due to embryonic lethality. In this study, we performed whole genome sequencing to determine if the integration site of the Her2 transgene was responsible for this lethality. Indeed, we report that the Her2 transgene had integrated into the Pds5b (precocious dissociation of sisters) gene on chromosome 5, as a 162 copy concatemer. Furthermore, our findings demonstrate that Her2+/+ mice, similar to Pds5b-/- mice, are embryonic lethal and confirm the necessity for Pds5b in embryonic development. This study confirms the value of whole genome sequencing in determining the integration site of transgenes to gain insight into associated phenotypes. PMID:26334628

  10. Cadmium-induced effects on cellular signaling pathways in the liver of transgenic estrogen reporter mice.

    PubMed

    Ali, Imran; Damdimopoulou, Pauliina; Stenius, Ulla; Adamsson, Annika; Mäkelä, Sari I; Åkesson, Agneta; Berglund, Marika; Håkansson, Helen; Halldin, Krister

    2012-05-01

    Estrogen-like effects of cadmium (Cd) have been reported in several animal studies, and recent epidemiological findings suggest increased risk of hormone-dependent cancers after Cd exposure. The mechanisms underlying these effects are still under investigation. Our aim was to study the effects of Cd on cellular signaling pathways in vivo with special focus on estrogen signaling and to perform benchmark dose analysis on the effects. Transgenic adult ERE-luciferase male mice were exposed subcutaneously to 0.5-500 μg CdCl(2) per kg body weight (bw) or 17α-ethinylestradiol (EE2) for 3 days. These doses had no effects on organ and bw or testicular histology, indicating subtoxic exposure levels. The transgene luciferase, reporting genomic estrogen response, was significantly increased by EE2 but not by Cd. However, Cd significantly affected kinase phosphorylation and endogenous gene expression. Interestingly, gene expression changes displayed a traditional dose-response relationship, with benchmark dose levels for the expression of Mt1, Mt2, p53, c-fos, and Mdm2 being 92.9, 19.9, 7.6, 259, and 25.9 μg/kg bw, respectively, but changes in kinase phosphorylation were only detected at low exposure levels. Phosphorylation of Erk1/2 was significantly increased even in the lowest dose group, 0.5 μg/kg bw, rendering pErk1/2 a more sensitive sensor of exposure than changes in gene expression. Collectively, our data suggest that the effects triggered by Cd in vivo are markedly concentration dependent. Furthermore, we conclude that the estrogen-like effects of Cd are likely to result from a mechanism different from steroidal estrogens. PMID:22314386

  11. Copper Deficiency in the Lungs of TNF-α Transgenic Mice.

    PubMed

    Liu, Liu; Geng, Xiangrong; McDermott, Joseph; Shen, Jian; Corbin, Cody; Xuan, Stephanie; Kim, Jae; Zuo, Li; Liu, Zijuan

    2016-01-01

    Tumor necrosis factor (TNF)-α is a well-known pro-inflammatory cytokine. Increased expression of Tnf-α is a feature of inflammatory lung diseases, such as asthma, emphysema, fibrosis, and smoking-induced chronic obstructive pulmonary disease (COPD). Using a mouse line with lung-specific Tnf-α overexpression (SPC-TNF-α) to mimic TNF-α-associated lung diseases, we investigated the role of chronic inflammation in the homeostasis of lung trace elements. We performed a quantitative survey of micronutrients and biometals, including copper (Cu), zinc (Zn), and selenium (Se), in the transgenic mice tissues. We also examined the expression of Cu-dependent proteins in the inflammatory lung tissue to determine whether they were affected by the severe Cu deficiency, including cuproenzymes, Cu transporters, and Cu chaperones. We found consistent lung-specific reduction of the metal Cu, with a mean decrease of 70%; however, Zn and Se were unaffected in all other tissues. RT-PCR showed that two Cu enzymes associated with lung pathology were downregulated: amine oxidase, Cu containing 3 (Aoc3) and lysyl oxidase (Lox). Two factors, vascular endothelial growth factor (Vegf) and focal adhesion kinase (Fak), related with Cu deficiency treatment, showed decreased expression in the transgenic inflammatory lung. We concluded that Cu deficiency occurs following chronic TNF-α-induced lung inflammation and this likely plays an essential role in the inflammation-induced lung damage. These results suggest the restoration of lung Cu status as a potential strategy in both treatment and prevention of chronic lung inflammation and related disorders. PMID:27378943

  12. Copper Deficiency in the Lungs of TNF-α Transgenic Mice

    PubMed Central

    Liu, Liu; Geng, Xiangrong; McDermott, Joseph; Shen, Jian; Corbin, Cody; Xuan, Stephanie; Kim, Jae; Zuo, Li; Liu, Zijuan

    2016-01-01

    Tumor necrosis factor (TNF)-α is a well-known pro-inflammatory cytokine. Increased expression of Tnf-α is a feature of inflammatory lung diseases, such as asthma, emphysema, fibrosis, and smoking-induced chronic obstructive pulmonary disease (COPD). Using a mouse line with lung-specific Tnf-α overexpression (SPC-TNF-α) to mimic TNF-α-associated lung diseases, we investigated the role of chronic inflammation in the homeostasis of lung trace elements. We performed a quantitative survey of micronutrients and biometals, including copper (Cu), zinc (Zn), and selenium (Se), in the transgenic mice tissues. We also examined the expression of Cu-dependent proteins in the inflammatory lung tissue to determine whether they were affected by the severe Cu deficiency, including cuproenzymes, Cu transporters, and Cu chaperones. We found consistent lung-specific reduction of the metal Cu, with a mean decrease of 70%; however, Zn and Se were unaffected in all other tissues. RT-PCR showed that two Cu enzymes associated with lung pathology were downregulated: amine oxidase, Cu containing 3 (Aoc3) and lysyl oxidase (Lox). Two factors, vascular endothelial growth factor (Vegf) and focal adhesion kinase (Fak), related with Cu deficiency treatment, showed decreased expression in the transgenic inflammatory lung. We concluded that Cu deficiency occurs following chronic TNF-α-induced lung inflammation and this likely plays an essential role in the inflammation-induced lung damage. These results suggest the restoration of lung Cu status as a potential strategy in both treatment and prevention of chronic lung inflammation and related disorders. PMID:27378943

  13. Transgenic expression of an expanded (GCG)13 repeat PABPN1 leads to weakness and coordination defects in mice.

    PubMed

    Dion, Patrick; Shanmugam, Vijayalakshmi; Gaspar, Claudia; Messaed, Christiane; Meijer, Inge; Toulouse, André; Laganiere, Janet; Roussel, Julie; Rochefort, Daniel; Laganiere, Simon; Allen, Carol; Karpati, George; Bouchard, Jean-Pierre; Brais, Bernard; Rouleau, Guy A

    2005-04-01

    Oculopharyngeal muscular dystrophy (OPMD) is a late-onset disorder caused by a (GCG)n trinucleotide repeat expansion in the poly(A) binding protein nuclear-1 (PABPN1) gene, which in turn leads to an expanded polyalanine tract in the protein. We generated transgenic mice expressing either the wild type or the expanded form of human PABPN1, and transgenic animals with the expanded form showed clear signs of abnormal limb clasping, muscle weakness, coordination deficits, and peripheral nerves alterations. Analysis of mitotic and postmitotic tissues in those transgenic animals revealed ubiquitinated PABPN1-positive intranuclear inclusions (INIs) in neuronal cells. This latter observation led us to test and confirm the presence of similar INIs in postmortem brain sections from an OPMD patient. Our results indicate that expanded PABPN1, presumably via the toxic effects of its polyalanine tract, can lead to inclusion formation and neurodegeneration in both the mouse and the human. PMID:15755680

  14. Male infertility caused by epididymal dysfunction in transgenic mice expressing a dominant negative mutation of retinoic acid receptor alpha 1.

    PubMed

    Costa, S L; Boekelheide, K; Vanderhyden, B C; Seth, R; McBurney, M W

    1997-04-01

    Retinoids are thought to be required for the normal development and maturation of a number of tissues, including most epithelia. The action of retinoids appears to be mediated through the binding to retinoic acid receptors (RARs) in the nucleus. The activity of retinoic acid can be inhibited in cells carrying dominant negative mutations of RAR alpha. We created transgenic mice expressing a dominant negative mutant of RAR alpha driven by the murine mammary tumor virus promoter. Expression of the transgene was evident in the epididymis and vas deferens in transgenic males. These males were either infertile or had reduced fertility, and the epithelium lining the ducts of the epididymis and vas deferens had undergone squamous metaplasia. Sperm developed normally in the testis but degenerated in the epididymis and vas deferens because inspissated ductal fluid blocked the normal passage of the sperm. PMID:9096882

  15. Motor deficits associated with Huntington's disease occur in the absence of striatal degeneration in BACHD transgenic mice.

    PubMed

    Mantovani, Susanna; Gordon, Richard; Li, Rui; Christie, Daniel C; Kumar, Vinod; Woodruff, Trent M

    2016-05-01

    Huntington's disease (HD) is an incurable neurodegenerative condition characterized by progressive motor and cognitive dysfunction, and depletion of neurons in the striatum. Recently, BACHD transgenic mice expressing the full-length human huntingtin gene have been generated, which recapitulate some of the motor and cognitive deficits seen in HD. In this study, we carried out a series of extensive behavioural and neuropathological tests on BACHD mice, to validate this mouse for preclinical research. Transgenic C57BL/6J BACHD and litter-matched wild-type mice were examined in a battery of motor and cognitive function tests at regular intervals up to 12 months of age. Brains from these mice were also analysed for signs of neurodegeneration and striatal and cortical volume sizes compared using anatomic 16.4T magnetic resonance imaging (MRI) brain scans. BACHD mice showed progressive motor impairments on rotarod and balance beam tests starting from 3 months of age, were hypoactive in the open field tests starting from 6 months of age, however, showed no alterations in gait and grip strength at any age. Surprisingly, despite these distinct motor deficits, no signs of neuronal loss, gliosis or blood-brain barrier degeneration were observed in the striatum of 12-month-old mice. MRI brain scans confirmed no reduction in striatal or cortical volumes at 12 months of age, and BACHD mice had a normal lifespan. These results demonstrate that classical Huntington's-like motor impairments seen in this transgenic model, do not occur due to degeneration of the striatum, and thus caution against the use of this model for preclinical studies into HD. PMID:26908618

  16. Amyloid-β and tau synergistically impair the oxidative phosphorylation system in triple transgenic Alzheimer's disease mice

    PubMed Central

    Rhein, Virginie; Song, Xiaomin; Wiesner, Andreas; Ittner, Lars M.; Baysang, Ginette; Meier, Fides; Ozmen, Laurence; Bluethmann, Horst; Dröse, Stefan; Brandt, Ulrich; Savaskan, Egemen; Czech, Christian; Götz, Jürgen; Eckert, Anne

    2009-01-01

    Alzheimer's disease (AD) is characterized by amyloid-beta (Aβ)-containing plaques, neurofibrillary tangles, and neuron and synapse loss. Tangle formation has been reproduced in P301L tau transgenic pR5 mice, whereas APPswPS2N141I double-transgenic APP152 mice develop Aβ plaques. Cross-breeding generates triple transgenic (tripleAD) mice that combine both pathologies in one model. To determine functional consequences of the combined Aβ and tau pathologies, we performed a proteomic analysis followed by functional validation. Specifically, we obtained vesicular preparations from tripleAD mice, the parental strains, and nontransgenic mice, followed by the quantitative mass-tag labeling proteomic technique iTRAQ and mass spectrometry. Within 1,275 quantified proteins, we found a massive deregulation of 24 proteins, of which one-third were mitochondrial proteins mainly related to complexes I and IV of the oxidative phosphorylation system (OXPHOS). Notably, deregulation of complex I was tau dependent, whereas deregulation of complex IV was Aβ dependent, both at the protein and activity levels. Synergistic effects of Aβ and tau were evident in 8-month-old tripleAD mice as only they showed a reduction of the mitochondrial membrane potential at this early age. At the age of 12 months, the strongest defects on OXPHOS, synthesis of ATP, and reactive oxygen species were exhibited in the tripleAD mice, again emphasizing synergistic, age-associated effects of Aβ and tau in perishing mitochondria. Our study establishes a molecular link between Aβ and tau protein in AD pathology in vivo, illustrating the potential of quantitative proteomics. PMID:19897719

  17. Overexpression of spermidine/spermine N1-acetyltransferase elevates the threshold to pentylenetetrazol-induced seizure activity in transgenic mice.

    PubMed

    Kaasinen, Selma K; Gröhn, Olli H J; Keinänen, Tuomo A; Alhonen, Leena; Jänne, Juhani

    2003-10-01

    Activation of polyamine catabolism in transgenic mice through an overexpression of spermidine/spermine N(1)-acetyltransferase (SSAT) results in a massive overaccumulation of the diamine putrescine in most tissues including brain. Putrescine pool in transgenic animals was strikingly expanded in every six brain regions analyzed at present. Pons (23-fold), cerebellum (37-fold), cerebrum (34-fold), and hippocampus (16-fold) showed the greatest increases in putrescine levels. Moreover, the molar ratio of putrescine to spermidine was increased in the different brain regions of the transgenic animals on an average of nearly 40-fold. Upon an exposure of the animals to pentylenetetrazol (PTZ) infusions, a compound known to induce epilepsy-like seizure activity, the SSAT transgenic mice showed significantly elevated seizure threshold to both clonic and tonic convulsions in comparison with their syngenic littermates. This difference, however, disappeared when the animals were treated with ifenprodil prior to PTZ infusions. The latter compound acts as an antagonist of N-methyl-D-aspartate receptor by binding to the polyamine site of the receptor. Overexpression of SSAT likewise appeared to protect the transgenic animals from PTZ-induced neuron loss in the hippocampus. As putrescine is known to serve as a precursor to gamma-aminobutyric acid (GABA), we carried out (1)H NMR analyses the results of which revealed that the levels of the inhibitory amino acid GABA and its excitatory counterpart glutamate were indistinguishable in syngenic and transgenic animals in all brain regions analyzed. The present results suggest that the frequently observed enhanced accumulation of putrescine in response to brain insults belongs to neuroprotective measures rather than being a cause of the subsequent injury. PMID:14552906

  18. HDL from apoA1 transgenic mice expressing the 4WF isoform is resistant to oxidative loss of function[S

    PubMed Central

    Berisha, Stela Z.; Brubaker, Greg; Kasumov, Takhar; Hung, Kimberly T.; DiBello, Patricia M.; Huang, Ying; Li, Ling; Willard, Belinda; Pollard, Katherine A.; Nagy, Laura E.; Hazen, Stanley L.; Smith, Jonathan D.

    2015-01-01

    HDL functions are impaired by myeloperoxidase (MPO), which selectively targets and oxidizes human apoA1. We previously found that the 4WF isoform of human apoA1, in which the four tryptophan residues are substituted with phenylalanine, is resistant to MPO-mediated loss of function. The purpose of this study was to generate 4WF apoA1 transgenic mice and compare functional properties of the 4WF and wild-type human apoA1 isoforms in vivo. Male mice had significantly higher plasma apoA1 levels than females for both isoforms of human apoA1, attributed to different production rates. With matched plasma apoA1 levels, 4WF transgenics had a trend for slightly less HDL-cholesterol versus human apoA1 transgenics. While 4WF transgenics had 31% less reverse cholesterol transport (RCT) to the plasma compartment, equivalent RCT to the liver and feces was observed. Plasma from both strains had similar ability to accept cholesterol and facilitate ex vivo cholesterol efflux from macrophages. Furthermore, we observed that 4WF transgenic HDL was partially (∼50%) protected from MPO-mediated loss of function while human apoA1 transgenic HDL lost all ABCA1-dependent cholesterol acceptor activity. In conclusion, the structure and function of HDL from 4WF transgenic mice was not different than HDL derived from human apoA1 transgenic mice. PMID:25561462

  19. Partial rescue of taste responses of alpha-gustducin null mice by transgenic expression of alpha-transducin.

    PubMed

    He, Wei; Danilova, Vicktoria; Zou, Shiying; Hellekant, Göran; Max, Marianna; Margolskee, Robert F; Damak, Sami

    2002-10-01

    The transduction of responses to bitter and sweet compounds utilizes guanine nucleotide binding proteins (G proteins) and their coupled receptors. Alpha-gustducin, a transducin-like G protein alpha-subunit, and rod alpha-transducin are expressed in taste receptor cells. Alpha-gustducin knockout mice have profoundly diminished behavioral and electrophysiological responses to many bitter and sweet compounds, although these mice retain residual responses to these compounds. Alpha-gustducin and rod alpha-transducin are biochemically indistinguishable in their in vitro interactions with retinal phosphodiesterase, rhodopsin and G protein betagamma-subunits. To determine if alpha-transducin can function in taste receptor cells and to compare the function of alpha-gustducin versus alpha-transducin in taste transduction in vivo, we generated transgenic mice that express alpha-transducin under the control of the alpha-gustducin promoter in the alpha-gustducin null background. Immunohistochemistry showed that the alpha-transducin transgene was expressed in about two-thirds of the alpha-gustducin lineage of taste receptor cells. Two-bottle preference tests showed that transgenic expression of rod alpha-transducin partly rescued responses to denatonium benzoate, sucrose and the artificial sweetener SC45647, but not to quinine sulfate. Gustatory nerve recordings showed a partial rescue by the transgene of the response to sucrose, SC45647 and quinine, but not to denatonium. These results demonstrate that alpha-transducin can function in taste receptor cells and transduce some taste cell responses. Our results also suggest that alpha-transducin and alpha-gustducin may differ, at least in part, in their function in these cells, although this conclusion must be qualified because of the limited fidelity of the transgene expression. PMID:12379596

  20. Low level expression of glycine receptor beta subunit transgene is sufficient for phenotype correction in spastic mice.

    PubMed Central

    Hartenstein, B; Schenkel, J; Kuhse, J; Besenbeck, B; Kling, C; Becker, C M; Betz, H; Weiher, H

    1996-01-01

    Mutations in inhibitory glycine receptor (GlyR) subunit genes are associated with neuromotor diseases in man and mouse. To use the potential of the mouse mutants as animal models of human disease, we altered GlyR levels in mutant mice and studied their phenotype. A transgene coding for the beta subunit of the rat GlyR was introduced into the genetic background of the spa mutation, which is characterized by low endogenous expression levels of the beta subunit and a dramatic neuromotor phenotype. The resulting transgenic mice expressed the beta subunit mRNA at intermediate levels, and their phenotype was rescued. This provides formal proof for the casual relationship between GlyR beta gene mutation and motor disease, and indicates that a low level of beta gene expression (25% of normal) is sufficient for proper functioning of glycinergic synapses. Images PMID:8635460

  1. Cardiac overexpression of monocyte chemoattractant protein-1 in transgenic mice prevents cardiac dysfunction and remodeling after myocardial infarction.

    PubMed

    Morimoto, Hajime; Takahashi, Masafumi; Izawa, Atsushi; Ise, Hirohiko; Hongo, Minoru; Kolattukudy, Pappachan E; Ikeda, Uichi

    2006-10-13

    Myocardial infarction (MI) is accompanied by inflammatory responses that lead to the recruitment of leukocytes and subsequent myocardial damage, healing, and scar formation. Because monocyte chemoattractant protein-1 (MCP-1) (also known as CCL2) regulates monocytic inflammatory responses, we investigated the effect of cardiac MCP-1 overexpression on left ventricular (LV) dysfunction and remodeling in a murine MI model. Transgenic mice expressing the mouse JE-MCP-1 gene under the control of the alpha-cardiac myosin heavy chain promoter (MHC/MCP-1 mice) were used for this purpose. MHC/MCP-1 mice had reduced infarct area and scar formation and improved LV dysfunction after MI. These mice also showed induction of macrophage infiltration and neovascularization; however, few bone marrow-derived endothelial cells were detected in MHC/MCP-1 mice whose bone marrow was replaced with that of Tie2/LacZ transgenic mice. Flow cytometry analysis showed no increase in endothelial progenitor cells (CD34+/Flk-1+ cells) in MHC/MCP-1 mice. Marked myocardial interleukin (IL)-6 secretion, STAT3 activation, and LV hypertrophy were observed after MI in MHC/MCP-1 mice. Furthermore, cardiac myofibroblasts accumulated after MI in MHC/MCP-1 mice. In vitro experiments revealed that a combination of IL-6 with MCP-1 synergistically stimulated and sustained STAT3 activation in cardiomyocytes. MCP-1, IL-6, and hypoxia directly promoted the differentiation of cardiac fibroblasts into myofibroblasts. Our results suggest that cardiac overexpression of MCP-1 induced macrophage infiltration, neovascularization, myocardial IL-6 secretion, and accumulation of cardiac myofibroblasts, thereby resulting in the prevention of LV dysfunction and remodeling after MI. They also provide a new insight into the role of cardiac MCP-1 in the pathophysiology of MI. PMID:16990567

  2. An immunological renal disease in transgenic mice that overexpress Fli-1, a member of the ets family of transcription factor genes

    SciTech Connect

    Zhang, Liqun; Teng, Yen-Tung; Melet, F.

    1995-12-01

    This report describes how expression of the proto-oncogene Fli-1 is involved in the regulation of lymphopoiesis. Transgenic mice were generated which overexpressed the oncogene, leading to a high incidence of immunological renal diseases and death. The data suggests that overexpression of Fli-1 perturbs normal lymphoid cell function and programmed cell death, making these transgenic mice suitable as a biological model for autoimmune disease in humans. 35 refs., 6 figs., 4 tabs.

  3. Pattern of tau hyperphosphorylation and neurotransmitter markers in the brainstem of senescent tau filament forming transgenic mice.

    PubMed

    Morcinek, Kerstin; Köhler, Christoph; Götz, Jürgen; Schröder, Hannsjörg

    2013-02-25

    The early occurrence of brainstem-related symptoms, e.g. gait and balance impairment, apathy and depression in Alzheimer's disease patients suggests brainstem involvement in the initial pathogenesis. To address the question whether tau filament forming mice expressing mutated human tau mirror histopathological changes observed in Alzheimer brainstem, the degree and distribution of neurofibrillary lesions as well as the pattern of cholinergic and monoaminergic neurons were investigated. The expression of the human tau transgene was observed in multiple brainstem nuclei, particularly in the magnocellular reticular formation, vestibular nuclei, cranial nerve motor nuclei, sensory trigeminal nerve nuclei, inferior and superior colliculi, periaqueductal and pontine gray matter, and the red nucleus. Most of the human tau-immunoreactive cell groups also showed tau hyperphosphorylation at the epitopes Thr231/Ser235 and Ser202/Thr205, while abnormal tau phosphorylation at the epitope Ser422 or silver stained structures were almost totally lacking. We found no obvious differences in distribution and density of cholinergic and monoaminergic neurons between tau-transgenic and wild type mice. Although numerous brainstem nuclei in our model expressed human tau protein, the development of neurofibrillary tangles, neuropil threads and ghost tangles was rare and likewise its distribution differed largely from Alzheimer's disease pattern. The number of monoaminergic neurons remained unchanged in the transgenic mice, while monoaminergic nuclei in Alzheimer brainstem showed a distinct neuronal loss. However, the distribution of pretangle-affected neurons in the tau-transgenic mice partly resembled those seen in progressive supranuclear palsy, presenting these animals as a model to examine brainstem pathogenesis of progressive supranuclear palsy. PMID:23261664

  4. Lentiviral vector encoding ubiquitinated hepatitis B core antigen induces potent cellular immune responses and therapeutic immunity in HBV transgenic mice.

    PubMed

    Dai, Shenglan; Zhuo, Meng; Song, Linlin; Chen, Xiaohua; Yu, Yongsheng; Zang, Guoqing; Tang, Zhenghao

    2016-07-01

    Predominant T helper cell type 1 (Th1) immune responses accompanied by boosted HBV-specific cytotoxic T lymphocyte (CTL) activity are essential for the clearance of hepatitis B virus (HBV) in chronic hepatitis B (CHB) patients. Ubiquitin (Ub) serves as a signal for the target protein to be recognized and degraded through the ubiquitin-proteasome system (UPS). Ubiquitinated hepatitis B core antigen (Ub-HBcAg) has been proved to be efficiently degraded into the peptides, which can be presented by major histocompatibility complex (MHC) class I resulting in stimulating cell-mediated responses. In the present study, lentiviral vectors encoding Ub-HBcAg (LV-Ub-HBcAg) were designed and constructed as a therapeutic vaccine for immunotherapy. HBcAg-specific cellular immune responses and anti-viral effects induced by LV-Ub-HBcAg were evaluated in HBV transgenic mice. We demonstrated that immunization with LV-Ub-HBcAg promoted the secretion of cytokines interleukin-2 (IL-2), interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α), generated remarkably high percentages of IFN-γ-secreting CD8(+) T cells and CD4(+) T cells, and enhanced HBcAg-specific CTL activity in HBV transgenic mice. More importantly, vaccination with LV-Ub-HBcAg could efficiently decreased the levels of serum hepatitis B surface antigen (HBsAg), HBV DNA and the expression of HBsAg and HBcAg in liver tissues of HBV transgenic mice. In addition, LV-Ub-HBcAg could upregulate the expression of T cell-specific T-box transcription factor (T-bet) and downregulate the expression of GATA-binding protein 3 (GATA-3) in spleen T lymphocytes. The therapeutic vaccine LV-Ub-HBcAg could break immune tolerance, and induce potent HBcAg specific cellular immune responses and therapeutic effects in HBV transgenic mice. PMID:26874581

  5. Development of early postnatal peripheral nerve abnormalities in Trembler-J and PMP22 transgenic mice

    PubMed Central

    ROBERTSON, A. M.; HUXLEY, C.; KING, R. H. M.; THOMAS, P. K.

    1999-01-01

    Mutations in the gene for peripheral myelin protein 22 (PMP22) are associated with peripheral neuropathy in mice and humans. Although PMP22 is strongly expressed in peripheral nerves and is localised largely to the myelin sheath, a dual role has been suggested as 2 differentially expressed promoters have been found. In this study we compared the initial stages of postnatal development in transgenic mouse models which have, in addition to the murine pmp22 gene, 7 (C22) and 4 (C61) copies of the human PMP22 gene and in homozygous and heterozygous Trembler-J (TrJ) mice, which have a point mutation in the pmp22 gene. The number of axons that were singly ensheathed by Schwann cells was the same in all groups indicating that PMP22 does not function in the initial ensheathment and separation of axons. At both P4 and P12 all mutants had an increased proportion of fibres that were incompletely surrounded by Schwann cell cytoplasm indicating that this step is disrupted in PMP22 mutants. C22 and homozygous TrJ animals could be distinguished by differences in the Schwann cell morphology at the initiation of myelination. In homozygous TrJ animals the Schwann cell cytoplasm had failed to make a full turn around the axon whereas in the C22 strain most fibres had formed a mesaxon. It is concluded that PMP22 functions in the initiation of myelination and probably involves the ensheathment of the axon by the Schwann cell, and the extension of this cell along the axon. Abnormalities may result from a failure of differentiation but more probably from defective interactions between the axon and the Schwann cell. PMID:10580849

  6. BAC transgenic mice provide evidence that p53 expression is highly regulated in vivo

    PubMed Central

    Chen, L; Zhang, G X; Zhou, Y; Zhang, C X; Xie, Y Y; Xiang, C; He, X Y; Zhang, Q; Liu, G

    2015-01-01

    p53 is an important tumor suppressor and stress response mediator. Proper control of p53 level and activity is tightly associated with its function. Posttranslational modifications and the interactions with Mdm2 and Mdm4 are major mechanisms controlling p53 activity and stability. As p53 protein is short-lived and hardly detectable in unstressed situations, less is known on its basal level expression and the corresponding controlling mechanisms in vivo. In addition, it also remains obscure how p53 expression might contribute to its functional regulation. In this study, we established bacterial artificial chromosome transgenic E.coli β-galactosidase Z gene reporter mice to monitor p53 expression in mouse tissues and identify important regulatory elements critical for the expression in vivo. We revealed preferentially high level of p53 reporter expressions in the proliferating, but not the differentiated compartments of the majority of tissues during development and tissue homeostasis. In addition, tumors as well as regenerating tissues in the p53 reporter mice also expressed high level of β-gal. Furthermore, both the enhancer box sequence (CANNTG) in the p53 promoter and the 3′ terminal untranslated region element were critical in mediating the high-level expression of the reporter. We also provided evidence that cellular myelocytomatosis oncogene was a critical player regulating p53 mRNA expression in proliferating cells and tissues. Finally, we found robust p53 activation preferentially in the proliferating compartment of mouse tissues upon DNA damage and the proliferating cells exhibited an enhanced p53 response as compared with cells in a quiescent state. Together, these results suggested a highly regulated expression pattern of p53 in the proliferating compartment controlled by both transcriptional and posttranscriptional mechanisms, and such regulated p53 expression may impose functional significance upon stress by setting up a precautionary mode in

  7. Hepatocyte nuclear phenotype: the cross-talk between anabolic androgenic steroids and exercise in transgenic mice.

    PubMed

    Fontana, Karina; Aldrovani, Marcela; de Paoli, Flávia; Oliveira, Helena C F; de Campos Vidal, Benedicto; da Cruz-Höfling, Maria Alice

    2008-11-01

    The growing and indiscriminate use of high doses of anabolic androgenic steroid (AAS) among youth and athletes has raised serious concerns about its hepatotoxic effects. Herein, the influence of AAS in the nuclear phenotype of hepatocytes was investigated in sedentary and trained mice heterozygous for the human CETP (cholesteryl ester transfer protein) transgene and for LDL-receptor null allele (CETP+/-LDLr+/-) by image analysis. Five groups were assayed comprising treadmill exercised (Ex) and sedentary (Sed) mice, administered mesterolone (AAS) or gum arabic (GA) and a sedentary blank control: G1(SedAAS), G2(SedGA), G3(ExAAS), G4(ExGA), and G5(SedBL). To assess nuclear phenotypes, the state of chromatin supraorganization, DNA content and fragmentation (TUNEL assay), area and perimeter of hepatocytes were determined in Feulgen-stained liver imprints. In addition, the activity of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) hepatic transaminases were measured. SedAAS-G1 showed the lowest chromatin condensation and highest Feulgen-DNA content, polyploid nuclei frequency, nuclear area and perimeter, suggesting gene activation. Contrarily, ExAAS-G3 showed a highest chromatin condensation, and a significant decrease of Feulgen-DNA content and decreased frequency of polyploid nuclei, which suggest gene silencing. Image analysis of the nuclear phenotype offered a coherent descriptive picture of the changing patterns of chromatin organization, which were shown to be congruent with the levels of Feulgen-DNA content, geometric nuclear parameters and hepatocyte activity. In this study, the image analysis permitted the monitoring of the nuclear response to mesterolone and physical exercise action in liver cells, the molecular mechanism of which is in prospect. PMID:18785119

  8. Progressive inflammatory pathology in the retina of aluminum-fed 5xFAD transgenic mice.

    PubMed

    Pogue, A I; Dua, P; Hill, J M; Lukiw, W J

    2015-11-01

    At least 57 murine transgenic models for Alzheimer's disease (Tg-AD) have been developed to overexpress the 42 amino acid amyloid-beta (Aβ42) peptide in the central nervous system (CNS). These 'humanized murine Tg-AD models' have greatly expanded our understanding of the contribution of Aβ42 peptide-mediated pro-inflammatory neuropathology to the AD process. A number of independent laboratories using different amyloid-overexpressing Tg-AD models have shown that supplementation of murine Tg-AD diets and/or drinking water with aluminum significantly enhances Aβ42 peptide-mediated inflammatory pathology and AD-type cognitive change compared to animals receiving control diets. In humans AD-type pathology appears to originate in the limbic system and progressively spreads into primary processing and sensory regions such as the retina. In these studies, for the first time, we assess the propagation of Aβ42 and inflammatory signals into the retina of 5xFAD Tg-AD amyloid-overexpressing mice whose diets were supplemented with aluminum. The two most interesting findings were (1) that similar to other Tg-AD models, there was a significantly accelerated development of Aβ42 and inflammatory pathology in 5xFAD Tg-AD mice fed aluminum; and (2) in aluminum-supplemented animals, markers for inflammatory pathology appeared in both the brain and the retina as evidenced by an evolving presence of Aβ42 peptides, and accompanied by inflammatory markers - cyclooxygenase-2 (COX-2) and C-reactive protein (CRP). The results indicate that in the 5xFAD Tg-AD model aluminum not only enhances an Aβ42-mediated inflammatory degeneration of the brain but also appears to induce AD-type pathology in an anatomically-linked primary sensory area that involves vision. PMID:26213226

  9. In Vivo Visualization of Alzheimer’s Amyloid Plaques by MRI in Transgenic Mice Without a Contrast Agent

    PubMed Central

    Jack, Clifford R.; Garwood, Michael; Wengenack, Thomas M.; Borowski, Bret; Curran, Geoffrey L.; Lin, Joseph; Adriany, Gregor; Grohn, Olli H.J.; Grimm, Roger; Poduslo, Joseph F.

    2009-01-01

    One of the cardinal pathologic features of Alzheimer’s disease (AD) is formation of senile, or amyloid, plaques. Transgenic mice have been developed that express one or more of the genes responsible for familial AD in humans. Doubly transgenic mice develop “human-like” plaques, providing a mechanism to study amyloid plaque biology in a controlled manner. Imaging of labeled plaques has been accomplished with other modalities, but only MRI has sufficient spatial and contrast resolution to visualize individual plaques non-invasively. Methods to optimize visualization of plaques in vivo in transgenic mice at 9.4 T using a spin echo sequence based on adiabatic pulses are described. Preliminary results indicate that a spin echo acquisition more accurately reflects plaque size, while a T2* weighted gradient echo sequence reflects plaque iron content not plaque size. In vivo MRI – ex vivo MRI – in vitro histological correlations are provided. Histologically verified plaques as small as 50 μm in diameter were visualized in the living animal. To our knowledge this work represents the first demonstration of non-invasive in vivo visualization of individual AD plaques without the use of a contrast agent. PMID:15562496

  10. Quantitative assessment of aortic atherosclerosis in APOE*3 Leiden transgenic mice and its relationship to serum cholesterol exposure.

    PubMed

    Groot, P H; van Vlijmen, B J; Benson, G M; Hofker, M H; Schiffelers, R; Vidgeon-Hart, M; Havekes, L M

    1996-08-01

    Transgenic mice overexpressing the human dysfunctional apolipoprotein E variant, APOE*3 Leiden, develop hyperlipidemia and are highly susceptible to diet-induced atherosclerosis. In the present study, we investigated the effects of diet composition and feeding period on serum cholesterol exposure and the amount of atherosclerosis in the aortic sinus in these mice, using quantitative image analysis. On each of the three diets tested--a low-fat diet, a high-saturated-fat/cholesterol diet, and a high saturated-fat/high-cholesterol/0.5%-cholate diet--transgenic animals showed a marked hyperlipidemia compared with nontransgenic littermates. Measurement of the atherosclerotic lesion areas in cross sections of the aortic sinus in animals exposed to these three diets for up to 6 months showed a 5 to 10 times greater lesion area in transgenic mice compared with nontransgenic controls. Highly significant positive correlations were found between the log-transformed data on lesion area and serum cholesterol exposure (r = .82 to .85 for the 1-, 2-, and 3-month treatment groups), indicating that the hyperlipidemia is likely to be a major determinant in lesion formation. On the basis of these findings, we suggest that the APOE*3 Leiden mouse represents a promising model for intervention studies with hypolipidemic and antiatherosclerotic drugs. PMID:8696955

  11. Tissue-specific expression in the salivary glands of transgenic mice.

    PubMed Central

    Mikkelsen, T R; Brandt, J; Larsen, H J; Larsen, B B; Poulsen, K; Ingerslev, J; Din, N; Hjorth, J P

    1992-01-01

    Using a DNA construct, named Lama, derived from the murine parotid secretory protein (PSP) gene, we have obtained salivary gland specific gene expression in transgenic mice. Lama is a PSP minigene and allows analysis of the PSP gene 5' regulatory region by transgenesis. We show here that the regulatory region included in Lama with 4.6 kb of 5' flanking sequence is sufficient to direct expression specifically to the salivary glands. The expression level in the parotid gland is only about one percent of the PSP mRNA level, while that of the sublingual gland is near the PSP mRNA level. This suggests significant differences in the PSP gene regulation in the two glands. In addition, Lama is a secretory expression vector in which cDNAs or genomic fragments can be inserted. We demonstrate that the Lama construct can direct the expression of a heterologous cDNA encoding the C-terminal peptide of human factor VIII to salivary glands and that the corresponding peptide is secreted into saliva. Images PMID:1594444

  12. Congenital hydrocephalus and abnormal subcommissural organ development in Sox3 transgenic mice.

    PubMed

    Lee, Kristie; Tan, Jacqueline; Morris, Michael B; Rizzoti, Karine; Hughes, James; Cheah, Pike See; Felquer, Fernando; Liu, Xuan; Piltz, Sandra; Lovell-Badge, Robin; Thomas, Paul Q

    2012-01-01

    Congenital hydrocephalus (CH) is a life-threatening medical condition in which excessive accumulation of CSF leads to ventricular expansion and increased intracranial pressure. Stenosis (blockage) of the Sylvian aqueduct (Aq; the narrow passageway that connects the third and fourth ventricles) is a common form of CH in humans, although the genetic basis of this condition is unknown. Mouse models of CH indicate that Aq stenosis is associated with abnormal development of the subcommmissural organ (SCO) a small secretory organ located at the dorsal midline of the caudal diencephalon. Glycoproteins secreted by the SCO generate Reissner's fibre (RF), a thread-like structure that descends into the Aq and is thought to maintain its patency. However, despite the importance of SCO function in CSF homeostasis, the genetic program that controls SCO development is poorly understood. Here, we show that the X-linked transcription factor SOX3 is expressed in the murine SCO throughout its development and in the mature organ. Importantly, overexpression of Sox3 in the dorsal diencephalic midline of transgenic mice induces CH via a dose-dependent mechanism. Histological, gene expression and cellular proliferation studies indicate that Sox3 overexpression disrupts the development of the SCO primordium through inhibition of diencephalic roof plate identity without inducing programmed cell death. This study provides further evidence that SCO function is essential for the prevention of hydrocephalus and indicates that overexpression of Sox3 in the dorsal midline alters progenitor cell differentiation in a dose-dependent manner. PMID:22291885

  13. Congenital Hydrocephalus and Abnormal Subcommissural Organ Development in Sox3 Transgenic Mice

    PubMed Central

    Lee, Kristie; Tan, Jacqueline; Morris, Michael B.; Rizzoti, Karine; Hughes, James; Cheah, Pike See; Felquer, Fernando; Liu, Xuan; Piltz, Sandra; Lovell-Badge, Robin; Thomas, Paul Q.

    2012-01-01

    Congenital hydrocephalus (CH) is a life-threatening medical condition in which excessive accumulation of CSF leads to ventricular expansion and increased intracranial pressure. Stenosis (blockage) of the Sylvian aqueduct (Aq; the narrow passageway that connects the third and fourth ventricles) is a common form of CH in humans, although the genetic basis of this condition is unknown. Mouse models of CH indicate that Aq stenosis is associated with abnormal development of the subcommmissural organ (SCO) a small secretory organ located at the dorsal midline of the caudal diencephalon. Glycoproteins secreted by the SCO generate Reissner's fibre (RF), a thread-like structure that descends into the Aq and is thought to maintain its patency. However, despite the importance of SCO function in CSF homeostasis, the genetic program that controls SCO development is poorly understood. Here, we show that the X-linked transcription factor SOX3 is expressed in the murine SCO throughout its development and in the mature organ. Importantly, overexpression of Sox3 in the dorsal diencephalic midline of transgenic mice induces CH via a dose-dependent mechanism. Histological, gene expression and cellular proliferation studies indicate that Sox3 overexpression disrupts the development of the SCO primordium through inhibition of diencephalic roof plate identity without inducing programmed cell death. This study provides further evidence that SCO function is essential for the prevention of hydrocephalus and indicates that overexpression of Sox3 in the dorsal midline alters progenitor cell differentiation in a dose-dependent manner. PMID:22291885

  14. Therapeutic potential of human amniotic membrane-derived mesenchymal stem cells in APP transgenic mice

    PubMed Central

    Jiao, Hongliang; Shi, Ke; Zhang, Weijie; Yang, Liang; Yang, Lu; Guan, Fangxia; Yang, Bo

    2016-01-01

    Growing evidence indicates that the presence of extensive oxidative stress plays an essential role in the initiation and progression of Alzheimer's disease (AD). Amyloid-β (Aβ) aggregation is involved in the elevation of oxidative stress, contributing to mitochondrial dysfunction and lipid peroxidation. In the present study, human placenta amniotic membrane-derived mesenchymal stem cells (hAMMSCs) were intravenously injected into C57BL/6J-APP transgenic mice. hAMMSCs significantly ameliorated spatial learning and memory function, and were associated with a decreased amount of amyloid plaques of the brain. The correlation of oxidative stress with Aβ levels was lower in the hAMMSCs-injected group than in the phosphate-buffered saline (PBS)-injected group, as indicated by the increased level of antioxidative enzymes and the decreased level of lipid peroxidation product. The glutathione (GSH) level and ratio of GSH to glutathione disulfide were higher in the hAMMSC group than in the PBS group. The superoxide dismutase activity and malonaldehyde level were improved significantly as the level of Aβ decreased, but there was no such trend in the PBS group. As a result, our findings represent evidence that hAMMSC treatment might improve the pathology of AD and memory function through the regulation of oxidative stress. PMID:27588134

  15. Longitudinal noninvasive magnetic resonance imaging of brain microhemorrhages in BACE inhibitor-treated APP transgenic mice.

    PubMed

    Beckmann, Nicolau; Doelemeyer, Arno; Zurbruegg, Stefan; Bigot, Karine; Theil, Diethilde; Frieauff, Wilfried; Kolly, Carine; Moulin, Pierre; Neddermann, Daniel; Kreutzer, Robert; Perrot, Ludovic; Brzak, Irena; Jacobson, Laura H; Staufenbiel, Matthias; Neumann, Ulf; Shimshek, Derya R

    2016-09-01

    Currently, several immunotherapies and BACE (Beta Site APP Cleaving Enzyme) inhibitor approaches are being tested in the clinic for the treatment of Alzheimer's disease. A crucial mechanism-related safety concern is the exacerbation of microhemorrhages, which are already present in the majority of Alzheimer patients. To investigate potential safety liabilities of long-term BACE inhibitor therapy, we used aged amyloid precursor protein (APP) transgenic mice (APP23), which robustly develop cerebral amyloid angiopathy. T2*-weighted magnetic resonance imaging (MRI), a translational method applicable in preclinical and clinical studies, was used for the detection of microhemorrhages throughout the entire brain, with subsequent histological validation. Three-dimensional reconstruction based on in vivo MRI and serial Perls' stained sections demonstrated a one-to-one matching of the lesions thus allowing for their histopathological characterization. MRI detected small Perls' positive areas with a high spatial resolution. Our data demonstrate that volumetric assessment by noninvasive MRI is well suited to monitor cerebral microhemorrhages in vivo. Furthermore, 3 months treatment of aged APP23 with the potent BACE-inhibitor NB-360 did not exacerbate microhemorrhages in contrast to Aβ-antibody β1. These results substantiate the safe use of BACE inhibitors regarding microhemorrhages in long-term clinical studies for the treatment of Alzheimer's disease. PMID:27459925

  16. Impaired plasticity of cortical dendritic spines in P301S tau transgenic mice

    PubMed Central

    2013-01-01

    Background Illuminating the role of the microtubule-associated protein tau in neurodegenerative diseases is of increasing importance, supported by recent studies establishing novel functions of tau in synaptic signalling and cytoskeletal organization. In severe dementias like Alzheimer’s disease (AD), synaptic failure and cognitive decline correlate best with the grade of tau-pathology. To address synaptic alterations in tauopathies, we analyzed the effects of mutant tau expression on excitatory postsynapses in vivo. Results Here we followed the fate of single dendritic spines in the neocortex of a tauopathy mouse model, expressing human P301S mutated tau, for a period of two weeks. We observed a continuous decrease in spine density during disease progression, which we could ascribe to a diminished fraction of gained spines. Remaining spines were enlarged and elongated, thus providing evidence for morphological reorganization in compensation for synaptic dysfunction. Remarkably, loss of dendritic spines in cortical pyramidal neurons occurred in the absence of neurofibrillary tangles (NFTs). Therefore, we consider prefibrillar tau species as causative for the observed impairment in spine plasticity. Conclusions Dendritic spine plasticity and morphology are altered in layer V cortical neurons of P301S tau transgenic mice in vivo. This does not coincide with the detection of hyperphosphorylated tau in dendritic spines. PMID:24344647

  17. Evaluating Human T-Cell Therapy of Cytomegalovirus Organ Disease in HLA-Transgenic Mice

    PubMed Central

    Thomas, Simone; Klobuch, Sebastian; Podlech, Jürgen; Plachter, Bodo; Hoffmann, Petra; Renzaho, Angelique; Theobald, Matthias

    2015-01-01

    Reactivation of human cytomegalovirus (HCMV) can cause severe disease in recipients of hematopoietic stem cell transplantation. Although preclinical research in murine models as well as clinical trials have provided 'proof of concept' for infection control by pre-emptive CD8 T-cell immunotherapy, there exists no predictive model to experimentally evaluate parameters that determine antiviral efficacy of human T cells in terms of virus control in functional organs, prevention of organ disease, and host survival benefit. We here introduce a novel mouse model for testing HCMV epitope-specific human T cells. The HCMV UL83/pp65-derived NLV-peptide was presented by transgenic HLA-A2.1 in the context of a lethal infection of NOD/SCID/IL-2rg-/- mice with a chimeric murine CMV, mCMV-NLV. Scenarios of HCMV-seropositive and -seronegative human T-cell donors were modeled by testing peptide-restimulated and T-cell receptor-transduced human T cells, respectively. Upon transfer, the T cells infiltrated host tissues in an epitope-specific manner, confining the infection to nodular inflammatory foci. This resulted in a significant reduction of viral load, diminished organ pathology, and prolonged survival. The model has thus proven its potential for a preclinical testing of the protective antiviral efficacy of HCMV epitope-specific human T cells in the evaluation of new approaches to an immunotherapy of CMV disease. PMID:26181057

  18. Gata3 Hypomorphic Mutant Mice Rescued with a Yeast Artificial Chromosome Transgene Suffer a Glomerular Mesangial Cell Defect.

    PubMed

    Moriguchi, Takashi; Yu, Lei; Otsuki, Akihito; Ainoya, Keiko; Lim, Kim-Chew; Yamamoto, Masayuki; Engel, James Douglas

    2016-09-01

    GATA3 is a zinc finger transcription factor that plays a crucial role in embryonic kidney development, while its precise functions in the adult kidney remain largely unexplored. Here, we demonstrate that GATA3 is specifically expressed in glomerular mesangial cells and plays a critical role in the maintenance of renal glomerular function. Newly generated Gata3 hypomorphic mutant mice exhibited neonatal lethality associated with severe renal hypoplasia. Normal kidney size was restored by breeding the hypomorphic mutant with a rescuing transgenic mouse line bearing a 662-kb Gata3 yeast artificial chromosome (YAC), and these animals (termed G3YR mice) survived to adulthood. However, most of the G3YR mice showed degenerative changes in glomerular mesangial cells, which deteriorated progressively during postnatal development. Consequently, the G3YR adult mice suffered severe renal failure. We found that the 662-kb Gata3 YAC transgene recapitulated Gata3 expression in the renal tubules but failed to direct sufficient GATA3 activity to mesangial cells. Renal glomeruli of the G3YR mice had significantly reduced amounts of platelet-derived growth factor receptor (PDGFR), which is known to participate in the development and maintenance of glomerular mesangial cells. These results demonstrate a critical role for GATA3 in the maintenance of mesangial cells and its absolute requirement for prevention of glomerular disease. PMID:27296697

  19. The MRI marker gene MagA attenuates the oxidative damage induced by iron overload in transgenic mice.

    PubMed

    Guan, Xiaoying; Jiang, Xinhua; Yang, Chuan; Tian, Xiumei; Li, Li

    2016-06-01

    We aimed to create transgenic (Tg) mice engineered for magnetic resonance imaging (MRI). To ascertain if MagA expression contributes to oxidative stress and iron metabolism, we report the generation of Tg mice in which ubiquitous expression of MagA can be detected by MRI in vivo. Expression of MagA in diverse tissues of Tg mice was assessed, and iron accumulation and deposition of nanoparticles in tissues were analyzed. Levels of antioxidant enzymes, lipid peroxidation and cytokine production were determined, and iron metabolism-related proteins were also detected. MagA Tg showed no apparent pathologic symptoms and no histologic changes compared with wild-type (WT) mice. Overexpression of MagA resulted in specific alterations of the transverse relaxation rate (R2) of water. Transgene-dependent changes in R2 were detectable by MRI in iron-overloaded mice. We also evaluated antioxidant abilities between WT and Tg groups or two iron-overloaded groups. Together with the data of cytokines and iron metabolism-related proteins, we inferred that MagA could regulate nanoparticle production and thus attenuate the oxidative damage induced by iron overload. The novel MagA Tg mouse, which expresses an MRI reporter in many tissues, would be a valuable model of MagA molecular imaging in which to study diseases related to iron metabolism. PMID:26488480

  20. Over-Expression of Porcine Myostatin Missense Mutant Leads to A Gender Difference in Skeletal Muscle Growth between Transgenic Male and Female Mice.

    PubMed

    Ma, Dezun; Gao, Pengfei; Qian, Lili; Wang, Qingqing; Cai, Chunbo; Jiang, Shengwang; Xiao, Gaojun; Cui, Wentao

    2015-01-01

    Myostatin, a transforming growth factor-β family member, is a negative regulator of skeletal muscle development and growth. Piedmontese cattle breeds have a missense mutation, which results in a cysteine to tyrosine substitution in the mature myostatin protein (C313Y). This loss-of-function mutation in myostatin results in a double-muscled phenotype in cattle. Myostatin propeptide is an inhibitor of myostatin activity and is considered a potential agent to stimulate muscle growth in livestock. In this study, we generated transgenic mice overexpressing porcine myostatin missense mutant (pmMS), C313Y, and wild-type porcine myostatin propeptide (ppMS), respectively, to examine their effects on muscle growth in mice. Enhanced muscle growth was observed in both pmMS and ppMS transgenic female mice and also in ppMS transgenic male mice. However, there was no enhanced muscle growth observed in pmMS transgenic male mice. To explore why there is such a big difference in muscle growth between pmMS and ppMS transgenic male mice, the expression level of androgen receptor (AR) mutant AR45 was measured by Western blot. Results indicated that AR45 expression significantly increased in pmMS transgenic male mice while it decreased dramatically in ppMS transgenic male mice. Our data demonstrate that both pmMS and ppMS act as myostatin inhibitors in the regulation of muscle growth, but the effect of pmMS in male mice is reversed by an increased AR45 expression. These results provide useful insight and basic theory to future studies on improving pork quality by genetically manipulating myostatin expression or by regulating myostatin activity. PMID:26305245

  1. Over-Expression of Porcine Myostatin Missense Mutant Leads to A Gender Difference in Skeletal Muscle Growth between Transgenic Male and Female Mice

    PubMed Central

    Ma, Dezun; Gao, Pengfei; Qian, Lili; Wang, Qingqing; Cai, Chunbo; Jiang, Shengwang; Xiao, Gaojun; Cui, Wentao

    2015-01-01

    Myostatin, a transforming growth factor-β family member, is a negative regulator of skeletal muscle development and growth. Piedmontese cattle breeds have a missense mutation, which results in a cysteine to tyrosine substitution in the mature myostatin protein (C313Y). This loss-of-function mutation in myostatin results in a double-muscled phenotype in cattle. Myostatin propeptide is an inhibitor of myostatin activity and is considered a potential agent to stimulate muscle growth in livestock. In this study, we generated transgenic mice overexpressing porcine myostatin missense mutant (pmMS), C313Y, and wild-type porcine myostatin propeptide (ppMS), respectively, to examine their effects on muscle growth in mice. Enhanced muscle growth was observed in both pmMS and ppMS transgenic female mice and also in ppMS transgenic male mice. However, there was no enhanced muscle growth observed in pmMS transgenic male mice. To explore why there is such a big difference in muscle growth between pmMS and ppMS transgenic male mice, the expression level of androgen receptor (AR) mutant AR45 was measured by Western blot. Results indicated that AR45 expression significantly increased in pmMS transgenic male mice while it decreased dramatically in ppMS transgenic male mice. Our data demonstrate that both pmMS and ppMS act as myostatin inhibitors in the regulation of muscle growth, but the effect of pmMS in male mice is reversed by an increased AR45 expression. These results provide useful insight and basic theory to future studies on improving pork quality by genetically manipulating myostatin expression or by regulating myostatin activity. PMID:26305245

  2. Melanocyte expression of Survivin promotes development and metastasis of UV-induced melanoma in HGF-transgenic mice

    PubMed Central

    Thomas, Joshua; Liu, Tong; Cotter, Murray A.; Florell, Scott R.; Robinette, Kyle; Hanks, Adrianne N.; Grossman, Douglas

    2008-01-01

    We previously found the apoptosis inhibitor Survivin to be expressed in melanocytic nevi and melanoma, but not in normal melanocytes. To investigate the role of Survivin in melanoma development and progression, we examined the consequences of forced Survivin expression in melanocytes in vivo. Transgenic (Tg) mouse lines (Dct-Survivin) were generated with melanocyte-specific expression of Survivin, and melanocytes grown from Dct-Survivin mice expressed Survivin. Dct-Survivin melanocytes exhibited decreased susceptibility to UV-induced apoptosis but no difference in proliferative capacity compared to melanocytes derived from non-Tg littermates. Induction of nevi in Dct-Survivin and non-Tg mice by topical application of DMBA did not reveal significant differences in lesion onset (median 10 wks) or density (4 lesions/mouse after 15 wks). Dct-Survivin mice were bred with melanoma-prone MH19/HGF-B6 Tg mice and all progeny expressing either individual, neither, or both (Survivin/HGF) transgenes were UV-treated as neonates and then monitored for 43 wks. Melanocytes in neonatal Survivin+/HGF+ mouse skin were less susceptible to UV-induced apoptosis than those from Survivin−/HGF+ mice. Onset of melanocytic tumors was earlier (median 18 vs. 24 wks, p = .01, log-rank test) and overall tumor density was greater (7.7 vs. 5.2 tumors/mouse, p = .04) in Survivin+/HGF+ compared to Survivin−/HGF+ mice. Strikingly, melanomas arising in Survivin+/HGF+ mice demonstrated a greater tendency for lymph node (35% vs. 0%, p = .04) and lung (53% vs. 22%) metastasis, and lower rates of spontaneous apoptosis, than those in Survivin−/HGF+ mice. These studies demonstrate a role for Survivin in promoting both early and late events of UV-induced melanoma development in vivo. PMID:17545596

  3. Suppressed rate of carcinogenesis and decreases in tumour volume and lung metastasis in CXCL14/BRAK transgenic mice.

    PubMed

    Hata, Ryu-Ichiro; Izukuri, Kazuhito; Kato, Yasumasa; Sasaki, Soichiro; Mukaida, Naofumi; Maehata, Yojiro; Miyamoto, Chihiro; Akasaka, Tetsu; Yang, Xiaoyan; Nagashima, Yoji; Takeda, Kazuyoshi; Kiyono, Tohru; Taniguchi, Masaru

    2015-01-01

    Cancer progression involves carcinogenesis, an increase in tumour size, and metastasis. Here, we investigated the effect of overexpressed CXC chemokine ligand 14 (CXCL14) on these processes by using CXCL14/BRAK (CXCL14) transgenic (Tg) mice. The rate of AOM/DSS-induced colorectal carcinogenesis in these mice was significantly lower compared with that for isogenic wild type C57BL/6 (Wt) mice. When tumour cells were injected into these mice, the size of the tumours that developed and the number of metastatic nodules in the lungs of the animals were always significantly lower in the Tg mice than in the Wt ones. Injection of anti-asialo-GM1 antibodies to the mice before and after injection of tumour cells attenuated the suppressing effects of CXCL14 on the tumor growth and metastasis, suggesting that NK cell activity played an important role during CXCL14-mediated suppression of tumour growth and metastasis. The importance of NK cells on the metastasis was also supported when CXCL14 was expressed in B16 melanoma cells. Further, the survival rates after tumour cell injection were significantly increased for the Tg mice. As these Tg mice showed no obvious abnormality, we propose that CXCL14 to be a promising molecular target for cancer suppression/prevention. PMID:25765541

  4. Age-related changes in brain metabolites and cognitive function in APP/PS1 transgenic mice.

    PubMed

    Chen, Shuang-qing; Cai, Qing; Shen, Yu-ying; Wang, Pei-jun; Teng, Gao-jun; Zhang, Wei; Zang, Feng-chao

    2012-11-01

    Proton magnetic resonance spectroscopy ((1)H-MRS) and the Morris water maze (MWM) have played an important role in Alzheimer's disease (AD) research. The aim of this study was to determine whether (1)H-MRS and the MWM can detect for early AD in APP/PS1 transgenic (tg) mice. (1)H-MRS was performed in 20 tg mice and 15 wild-type mice at 3, 5 and 8 months of age. The concentration of N-acetylaspartate (NAA), glutamate (Glu), myo-inositol (mI), choline (Cho) and creatine (Cr) in the hippocampus were measured, and the NAA/Cr, Glu/Cr, mI/Cr and Cho/Cr ratios were quantified. Additionally, the spatial learning and memory of the mice were evaluated by MWM. The (1)H-MRS revealed that mI levels in tg mice were significantly higher at 3 months of age compared to wt mice, while the NAA and Glu levels in 5- and 8-month-old tg mice were significantly decreased (p<0.05). Additionally, significant cognitive changes only occurred at 8 months of age in APP/PS1 tg mice. These results indicated that metabolic changes preceded overt cognitive dysfunctions in early-stage AD, suggesting that (1)H-MRS is a more sensitive biomarker for assessing early AD. PMID:22828014

  5. Feed-forward regulation of bile acid detoxification by CYP3A4: studies in humanized transgenic mice.

    PubMed

    Stedman, Catherine; Robertson, Graham; Coulter, Sally; Liddle, Christopher

    2004-03-19

    Bile acids are potentially toxic end products of cholesterol metabolism and their concentrations must be tightly regulated. Homeostasis is maintained by both feed-forward regulation and feedback regulation. We used humanized transgenic mice incorporating 13 kb of the 5' regulatory flanking sequence of CYP3A4 linked to a lacZ reporter gene to explore the in vivo relationship between bile acids and physiological adaptive CYP3A gene regulation in acute cholestasis after bile duct ligation (BDL). Male transgenic mice were subjected to BDL or sham surgery prior to sacrifice on days 3, 6, and 10, and others were injected with intraperitoneal lithocholic acid (LCA) or vehicle alone. BDL resulted in marked hepatic activation of the CYP3A4/lacZ transgene in pericentral hepatocytes, with an 80-fold increase in transgene activation by day 10. Individual bile acids were quantified by liquid chromatography/mass spectrometry. Serum 6beta-hydroxylated bile acids were increased following BDL, confirming the physiological relevance of endogenous Cyp3a induction to bile acid detoxification. Although concentrations of conjugated primary bile acids increased after BDL, there was no increase in LCA, a putative PXR ligand, indicating that this cannot be the only endogenous bile acid mediating this protective response. Moreover, in LCA-treated animals, 5-bromo-4-chloro-3-indolyl-beta-d-galactopyranoside staining showed hepatic activation of the CYP3A4 transgene only on the liver capsular surface, and minimal parenchymal induction, despite significant liver injury. This study demonstrates that CYP3A up-regulation is a significant in vivo adaptive response to cholestasis. However, this up-regulation is not dependent on increases in circulating LCA and the role of other bile acids as regulatory molecules requires further exploration. PMID:14681232

  6. Prolonged ethanol administration depletes mitochondrial DNA in MnSOD-overexpressing transgenic mice, but not in their wild type littermates

    SciTech Connect

    Larosche, Isabelle; Choumar, Amal; Fromenty, Bernard; Letteron, Philippe; Abbey-Toby, Adje; Van Remmen, Holly; Epstein, Charles J.; Richardson, Arlan; Feldmann, Gerard; Pessayre, Dominique; Mansouri, Abdellah

    2009-02-01

    Alcohol consumption increases reactive oxygen species formation and lipid peroxidation, whose products can damage mitochondrial DNA (mtDNA) and alter mitochondrial function. A possible role of manganese superoxide dismutase (MnSOD) on these effects has not been investigated. To test whether MnSOD overexpression modulates alcohol-induced mitochondrial alterations, we added ethanol to the drinking water of transgenic MnSOD-overexpressing (TgMnSOD) mice and their wild type (WT) littermates for 7 weeks. In TgMnSOD mice, alcohol administration further increased the activity of MnSOD, but decreased cytosolic glutathione as well as cytosolic glutathione peroxidase activity and peroxisomal catalase activity. Whereas ethanol increased cytochrome P-450 2E1 and mitochondrial ROS generation in both WT and TgMnSOD mice, hepatic iron, lipid peroxidation products and respiratory complex I protein carbonyls were only increased in ethanol-treated TgMnSOD mice but not in WT mice. In ethanol-fed TgMnSOD mice, but not ethanol-fed WT mice, mtDNA was depleted, and mtDNA lesions blocked the progress of polymerases. The iron chelator, DFO prevented hepatic iron accumulation, lipid peroxidation, protein carbonyl formation and mtDNA depletion in alcohol-treated TgMnSOD mice. Alcohol markedly decreased the activities of complexes I, IV and V of the respiratory chain in TgMnSOD, with absent or lesser effects in WT mice. There was no inflammation, apoptosis or necrosis, and steatosis was similar in ethanol-treated WT and TgMnSOD mice. In conclusion, prolonged alcohol administration selectively triggers iron accumulation, lipid peroxidation, respiratory complex I protein carbonylation, mtDNA lesions blocking the progress of polymerases, mtDNA depletion and respiratory complex dysfunction in TgMnSOD mice but not in WT mice.

  7. Expression of a human surfactant protein C mutation associated with interstitial lung disease disrupts lung development in transgenic mice.

    PubMed

    Bridges, James P; Wert, Susan E; Nogee, Lawrence M; Weaver, Timothy E

    2003-12-26

    Surfactant Protein C (SP-C) is a secreted transmembrane protein that is exclusively expressed by alveolar type II epithelial cells of the lung. SP-C associates with surfactant lipids to reduce surface tension within the alveolus, maintaining lung volume at end expiration. Mutations in the gene encoding SP-C (SFTPC) have recently been linked to chronic lung disease in children and adults. The goal of this study was to determine whether a disease-linked mutation in SFTPC causes lung disease in transgenic mice. The SFTPC mutation, designated g.1728 G --> A, results in the deletion of exon4, generating a truncated form of SP-C (SP-C(Deltaexon4)). cDNA encoding SP-C(Deltaexon4) was constitutively expressed in type II epithelial cells of transgenic mice. Viable F0 transgene-positive mice were not generated after two separate rounds of pronuclear injections. Histological analysis of lung tissue harvested from embryonic day 17.5 F0 transgene-positive fetuses revealed that SP-C(Deltaexon4) caused a dose-dependent disruption in branching morphogenesis of the lung associated with epithelial cell cytotoxicity. Transient expression of SP-C(Deltaexon4) in isolated type II epithelial cells or HEK293 cells resulted in incomplete processing of the mutant proprotein, a dose-dependent increase in BiP transcription, trapping of the proprotein in the endoplasmic reticulum, and rapid degradation via a proteasome-dependent pathway. Taken together, these data suggest that the g.1728 G --> A mutation causes misfolding of the SP-C proprotein with subsequent induction of the unfolded protein response and endoplasmic reticulum-associated degradation pathways ultimately resulting in disrupted lung morphogenesis. PMID:14525980

  8. Gene expression profiling of R6/2 transgenic mice with different CAG repeat lengths reveals genes associated with disease onset and progression in Huntington's disease.

    PubMed

    Tang, Bin; Seredenina, Tamara; Coppola, Giovanni; Kuhn, Alexandre; Geschwind, Daniel H; Luthi-Carter, Ruth; Thomas, Elizabeth A

    2011-06-01

    R6/2 transgenic mice with expanded CAG repeats (>300) have a surprisingly prolonged disease progression and longer lifespan than prototypical parent R6/2 mice (carrying 150 CAGs); however, the mechanism of this phenotype amelioration is unknown. We compared gene expression profiles in the striatum of R6/2 transgenic mice carrying ~300 CAG repeats (R6/2(Q300) transgenic mice) to those carrying ~150 CAG repeats (R6/2(Q150) transgenic mice) and littermate wildtype controls in order to identify genes that may play determinant roles in the time course of phenotypic expression in these mice. Of the top genes showing concordant expression changes in the striatum of both R6/2 lines, 85% were decreased in expression, while discordant expression changes were observed mostly for genes upregulated in R6/2(Q300) transgenic mice. Upregulated genes in the R6/2(Q300) mice were associated with the ubiquitin ligase complex, cell adhesion, protein folding, and establishment of protein localization. We qPCR-validated increases in expression of genes related to the latter category, including Lrsam1, Erp29, Nasp, Tap1, Rab9b, and Pfdn5 in R6/2(Q300) mice, changes that were not observed in R6/2 mice with shorter CAG repeats, even in late stages (i.e., 12 weeks of age). We further tested Lrsam1 and Erp29, the two genes showing the greatest upregulation in R6/2(Q300) transgenic mice, for potential neuroprotective effects in primary striatal cultures overexpressing a mutated human huntingtin (htt) fragment. Overexpression of Lrsam1 prevented the loss of NeuN-positive cell bodies in htt171-82Q cultures, concomitant with a reduction of nuclear htt aggregates. Erp29 showed no significant effects in this model. This is consistent with the distinct pattern of htt inclusion localization observed in R6/2(Q300) transgenic mice, in which smaller cytoplasmic inclusions represent the major form of insoluble htt in the cell, as opposed to large nuclear inclusions observed in R6/2(Q150) transgenic mice

  9. Lipid-Based Diets Improve Muscarinic Neurotransmission in the Hippocampus of Transgenic APPswe/PS1dE9 Mice.

    PubMed

    Janickova, Helena; Rudajev, Vladimir; Dolejsi, Eva; Koivisto, Hennariikka; Jakubik, Jan; Tanila, Heikki; El-Fakahany, Esam E; Dolezal, Vladimir

    2015-01-01

    Transgenic APPswe/PS1dE9 mice modeling Alzheimer's disease demonstrate ongoing accumulation of β-amyloid fragments resulting in formation of amyloid plaques that starts at the age of 4-5 months. Buildup of β-amyloid fragments is accompanied by impairment of muscarinic transmission that becomes detectable at this age, well before the appearance of cognitive deficits that manifest around the age of 12 months. We have recently demonstrated that long-term feeding of trangenic mice with specific isocaloric fish oil-based diets improves specific behavioral parameters. Now we report on the influence of short-term feeding (3 weeks) of three isocaloric diets supplemented with Fortasyn (containing fish oil and ingredients supporting membrane renewal), the plant sterol stigmasterol together with fish oil, and stigmasterol alone on markers of cholinergic neurotransmission in the hippocampus of 5-month-old transgenic mice and their wild-type littermates. Transgenic mice fed normal diet demostrated increase in ChAT activity and attenuation of carbachol-stimulated GTP-γ(35)S binding compared to wild-type mice. None of the tested diets compared to control diet influenced the activities of ChAT, AChE, BuChE, muscarinic receptor density or carbachol-stimulated GTP-γ(35)S binding in wild-type mice. In contrast, all experimental diets increased the potency of carbachol in stimulating GTP-γ(35)S binding in trangenic mice to the level found in wild-type animals. Only the Fortasyn diet increased markers of cholinergic synapses in transgenic mice. Our data demonstrate that even short-term feeding of transgenic mice with chow containing specific lipid-based dietary supplements can influence markers of cholinergic synapses and rectify impaired muscarinic signal transduction that develops in transgenic mice. PMID:26502816

  10. Transgenic mice over-expressing ET-1 in the endothelial cells develop systemic hypertension with altered vascular reactivity.

    PubMed

    Leung, Justin Wai-Chung; Wong, Wing Tak; Koon, Hon Wai; Mo, Fong Ming; Tam, Sidney; Huang, Yu; Vanhoutte, Paul M; Chung, Stephen Sum Man; Chung, Sookja Kim

    2011-01-01

    Endothelin-1 (ET-1) is a potent vasoconstrictor involved in the regulation of vascular tone and implicated in hypertension. However, the role of small blood vessels endothelial ET-1 in hypertension remains unclear. The present study investigated the effect of chronic over-expression of endothelial ET-1 on arterial blood pressure and vascular reactivity using transgenic mice approach. Transgenic mice (TET-1) with endothelial ET-1 over-expression showed increased in ET-1 level in the endothelial cells of small pulmonary blood vessels. Although TET-1 mice appeared normal, they developed mild hypertension which was normalized by the ET(A) receptor (BQ123) but not by ET(B) receptor (BQ788) antagonist. Tail-cuff measurements showed a significant elevation of systolic and mean blood pressure in conscious TET-1 mice. The mice also exhibited left ventricular hypertrophy and left axis deviation in electrocardiogram, suggesting an increased peripheral resistance. The ionic concentrations in the urine and serum were normal in 8-week old TET-1 mice, indicating that the systemic hypertension was independent of renal function, although, higher serum urea levels suggested the occurrence of kidney dysfunction. The vascular reactivity of the aorta and the mesenteric artery was altered in the TET-1 mice indicating that chronic endothelial ET-1 up-regulation leads to vascular tone imbalance in both conduit and resistance arteries. These findings provide evidence for the role of spatial expression of ET-1 in the endothelium contributing to mild hypertension was mediated by ET(A) receptors. The results also suggest that chronic endothelial ET-1 over-expression affects both cardiac and vascular functions, which, at least in part, causes blood pressure elevation. PMID:22096514

  11. Liver proteomic response to hypertriglyceridemia in human-apolipoprotein C-III transgenic mice at cellular and mitochondrial compartment levels

    PubMed Central

    2014-01-01

    Background Hypertriglyceridemia (HTG) is defined as a triglyceride (TG) plasma level exceeding 150 mg/dl and is tightly associated with atherosclerosis, metabolic syndrome, obesity, diabetes and acute pancreatitis. The present study was undertaken to investigate the mitochondrial, sub-mitochondrial and cellular proteomic impact of hypertriglyceridemia in the hepatocytes of hypertriglyceridemic transgenic mice (overexpressing the human apolipoproteinC-III). Methods Quantitative proteomics (2D-DIGE) analysis was carried out on both “low-expressor” (LE) and “high-expressor” (HE) mice, respectively exhibiting moderate and severe HTG, to characterize the effect of the TG plasma level on the proteomic response. Results The mitoproteome analysis has revealed a large-scale phenomenon in transgenic mice, i.e. a general down-regulation of matricial proteins and up-regulation of inner membrane proteins. These data also demonstrate that the magnitude of proteomic changes strongly depends on the TG plasma level. Our different analyses indicate that, in HE mice, the capacity of several metabolic pathways is altered to promote the availability of acetyl-CoA, glycerol-3-phosphate, ATP and NADPH for TG de novo biosynthesis. The up-regulation of several cytosolic ROS detoxifying enzymes has also been observed, suggesting that the cytoplasm of HTG mice is subjected to oxidative stress. Moreover, our results suggest that iron over-accumulation takes place in the cytosol of HE mice hepatocytes and may contribute to enhance oxidative stress and to promote cellular proliferation. Conclusions These results indicate that the metabolic response to HTG in human apolipoprotein C-III overexpressing mice may support a high TG production rate and that the cytosol of hepatocytes is subjected to an important oxidative stress, probably as a result of FFA over-accumulation, iron overload and enhanced activity of some ROS-producing catabolic enzymes. PMID:25047818

  12. Redox Proteomic Profiling of Specifically Carbonylated Proteins in the Serum of Triple Transgenic Alzheimer’s Disease Mice

    PubMed Central

    Shen, Liming; Chen, Youjiao; Yang, Aochu; Chen, Cheng; Liao, Liping; Li, Shuiming; Ying, Ming; Tian, Jing; Liu, Qiong; Ni, Jiazuan

    2016-01-01

    Oxidative stress is a key event in the onset and progression of neurodegenerative diseases, including Alzheimer’s disease (AD). To investigate the role of oxidative stress in AD and to search for potential biomarkers in peripheral blood, serums were collected in this study from the 3-, 6-, and 12-month-old triple transgenic AD mice (3×Tg-AD mice) and the age- and sex-matched non-transgenic (non-Tg) littermates. The serum oxidized proteins were quantified by slot-blot analysis and enzyme-linked immunosorbent assay (ELISA) to investigate the total levels of serum protein carbonyl groups. Western blotting, in conjunction with two-dimensional gel electrophoresis (2D-Oxyblot), was employed to identify and quantify the specifically-carbonylated proteins in the serum of 3×Tg-AD mice. The results showed that the levels of serum protein carbonyls were increased in the three month old 3×Tg-AD mice compared with the non-Tg control mice, whereas no significant differences were observed in the six and 12 months old AD mice, suggesting that oxidative stress is an early event in AD progression. With the application of 2D-Oxyblot analysis, (immunoglobin) Ig gamma-2B chain C region (IGH-3), Ig lambda-2 chain C region (IGLC2), Ig kappa chain C region (IGKC), and Ig kappa chain V-V region HP R16.7 were identified as significantly oxidized proteins compared with the control. Among them IGH-3 and IGKC were validated via immunoprecipitation and Western blot analysis. Identification of oxidized proteins in the serums of 3×Tg-AD mice can not only reveal potential roles of those proteins in the pathogenesis of AD but also provide potential biomarkers of AD at the early stage. PMID:27077851

  13. Pharmacologic blockade of 12/15-lipoxygenase ameliorates memory deficits, Aβ and tau neuropathology in the triple-transgenic mice.

    PubMed

    Chu, J; Li, J-G; Giannopoulos, P F; Blass, B E; Childers, W; Abou-Gharbia, M; Praticò, D

    2015-11-01

    The 12/15-lipoxygenase (12/15LO) enzyme is widely distributed within the central nervous system. Previous work showed that this protein is upregulated in Alzheimer's disease (AD), and plays an active role in the development of brain amyloidosis in amyloid beta (Aβ)-precursor protein transgenic mice (Tg2576). In the present paper, we studied the effect of its pharmacologic inhibition on the AD-like phenotype of a mouse model with plaques and tangles, the triple-transgenic mice. Compared with mice receiving placebo, the group treated with PD146176, a specific 12/15LO inhibitor, manifested a significant improvement of their memory deficits. The same animals had a significant reduction in Aβ levels and deposition, which was secondary to a decrease in the β-secretase pathway. In addition, while total tau-soluble levels were unchanged for both groups, PD146176-treated mice had a significant reduction in its phosphorylation state and insoluble fraction, which specifically associated with decrease in stress-activated protein kinase/c-Jun N-terminal kinase activity. In vitro study showed that the effect on tau and Aβ were independent from each other. These data establish a functional role for 12/15LO in the pathogenesis of the full spectrum of the AD-like phenotype and represent the successful completion of the initial step for the preclinical development of 12/15LO inhibitors as novel therapeutic agents for AD. PMID:25560760

  14. Hla-DR2-restricted responses to proteolipid protein 95-116 peptide cause autoimmune encephalitis in transgenic mice.

    PubMed

    Kawamura, K; Yamamura, T; Yokoyama, K; Chui, D H; Fukui, Y; Sasazuki, T; Inoko, H; David, C S; Tabira, T

    2000-04-01

    In multiple sclerosis (MS) patients who carry the Class II major histocompatibility (MHC) type HLA-DR2, T cells specific for amino acids 95-116 in the proteolipid protein (PLP) are activated and clonally expanded. However, it remains unclear whether these autoreactive T cells play a pathogenic role or, rather, protect against the central nervous system (CNS) damage. We have addressed this issue, using mice transgenic for the human MHC class II region carrying the HLA-DR2 (DRB1* 1502) haplotype. After stimulating cultured lymph node cells repeatedly with PLP95-116, we generated 2 HLA-DR2-restricted, PLP95-116-specific T-cell lines (TCLs) from the transgenic mice immunized with this portion of PLP. The TCLs were CD4+ and produced T-helper 1 (Th1) cytokines in response to the peptide. These TCLs were adoptively transferred into RAG-2/2 mice expressing HLA-DR2 (DRG1* 1502) molecules. Mice receiving 1 of the TCLs developed a neurological disorder manifested ataxic movement without apparent paresis on day 3, 4, or 5 after cell transfer. Histological examination revealed inflammatory foci primarily restricted to the cerebrum and cerebellum, in association with scattered demyelinating lesions in the deep cerebral cortex. These results support a pathogenic role for PLP95-116-specific T cells in HLA-DR2+ MS patients, and shed light on the possible correlation between autoimmune target epitope and disease phenotype in human CNS autoimmune diseases. PMID:10841661

  15. Morphological, Histochemical, Immunohistochemical, and Ultrastructural Characterization of Tumors and Dysplastic and Non-Neoplastic Lesions Arising in BK Virus/tat Transgenic Mice

    PubMed Central

    Altavilla, Giuseppe; Trabanelli, Cecilia; Merlin, Michela; Caputo, Antonella; Lanfredi, Massimo; Barbanti-Brodano, Giuseppe; Corallini, Alfredo

    1999-01-01

    To study the role in AIDS pathogenesis of the human immunodeficiency virus type 1 (HIV-1) Tat protein, a transactivator of viral and cellular genes, we generated transgenic mice with a recombinant DNA containing BK virus (BKV) early region and the HIV-1 tat gene, directed by its own promoter-enhancer. DNA hybridization revealed that the transgene is stably maintained in all organs of transgenic mice as a tandem insertion in a number of copies ranging from 5 to 20 per cell. In addition, tat and BKV RNA were expressed in all tissues. Transgenic mice developed three types of lesions: 1) tumors, 2) hyperplastic and dysplastic lesions, and 3) non-neoplastic lesions. Tumors of different histotypes, such as lymphomas, adenocarcinomas of skin glands, leiomyosarcomas, skin squamous cell carcinomas, hepatomas, hepatocarcinomas, and cavernous liver hemangiomas, developed in 29% of transgenic animals. The majority of tumors were malignant, invasive, and producing metastases. Conversely, tumors of only two histotypes (lymphomas and adenocarcinomas of skin glands) appeared in control mice. Hyperplastic and dysplastic lesions were more frequent in transgenic than in control mice and involved the skin or its adnexes, the liver and the rectum, indicating multiple targets for the activity of the transgene. Pyelonephritis, frequently complicated with hydronephrosis, inflammatory eye lesions, and amyloid depositions represented the most frequent non-neoplastic lesions detected in transgenic mice. Many of the pathological findings observed in this animal model are comparable to similar lesions appearing in AIDS patients, suggesting a relevant role for Tat in the pathogenesis of such lesions during the course of AIDS. PMID:10233861

  16. Targeting the cannabinoid CB2 receptor to attenuate the progression of motor deficits in LRRK2-transgenic mice.

    PubMed

    Palomo-Garo, Cristina; Gómez-Gálvez, Yolanda; García, Concepción; Fernández-Ruiz, Javier

    2016-08-01

    Most of cases of Parkinson's disease (PD) have a sporadic origin, with their causes mostly unknown, although overexposure to some environmental factors has been found to occur in some cases. Other forms of parkinsonism are the consequence of dominant or recessive mutations in specific genes, e.g. α-synuclein, parkin and, more recently, leucine-rich repeat kinase 2 (LRRK2), whose G2019S mutation represents the most prevalent form of late-onset, autosomal dominant familial PD. A transgenic mouse model expressing the G2019S mutation of LRRK2 is already available and apparently may represent a valuable experimental model for investigating PD pathogenesis and novel treatments. We designed a long-term study with these animals aimed at: (i) elucidating the changes experienced by the endocannabinoid signaling system in the basal ganglia during the progression of the disease in these mice, paying emphasis in the CB2 receptor, which has emerged as a promising target in PD, and (ii) evaluating the potential of compounds selectively activating this CB2 receptor, as disease-modifying agents in these mice. Our results unequivocally demonstrate that LRRK2 transgenic mice develop motor impairment consisting of small anomalies in rotarod performance (presumably reflecting a deficit in motor coordination and dystonia) and a strong deficiency in the hanging-wire test (reflecting muscle weakness), rather than hypokinesia which was difficult to be demonstrated in the actimeter. These behavioral responses occurred in absence of any evidence of reactive gliosis and neuronal losses, as well as synaptic deterioration in the basal ganglia, except an apparent impairment in autophagy reflected by elevated LAMP-1 immunolabelling in the striatum and substantia nigra. Furthermore, there were no changes in the status of the CB2 receptor, as well as in other elements of the endocannabinoid signaling, in the basal ganglia, but, paradoxically, the selective activation of this receptor partially

  17. Transgenic expression of interleukin 6 in the central nervous system regulates brain metallothionein-I and -III expression in mice.

    PubMed

    Hernández, J; Molinero, A; Campbell, I L; Hidalgo, J

    1997-08-01

    The metallothionein (MT) gene family consists of several members (MT-I-IV) that are tightly regulated during development. MT-I and MT-II are expressed in many tissues, including the brain, whereas MT-III is expressed mainly in the central nervous system. However, the physiological roles of these isoforms in the brain and their regulation are poorly characterized. In this report, we have studied the putative role of IL-6 in the regulation of brain MT. The present results demonstrated that transgenic mice expressing IL-6 under the regulatory control of the glial fibrillary acidic protein gene promoter (GFAP-IL6 mice), and which develop chronic progressive neurodegenerative disease, show significantly increased MT-I + II protein levels in specific brain areas. Thus, the MT-I + II levels of 1- and 3-month-old GFAP-IL6 mice (G16 and/or G36 lines) were not altered in hippocampus but they were elevated in the cerebellum (highest induction), medulla plus pons, hypothalamus and remaining brain (lowest induction). The effect of the transgenic expression of IL-6 was more dramatic for MT-I + II protein than for MT-I mRNA levels, with the latter only marginally elevated in the G16 line at 3 months but not at 6 months of age where there was a tendency to decreased levels. Brain MT-I mRNA levels also tended to decrease in the higher expressor G36 line in 3-month-old mice despite the strongly elevated MT-I + II protein levels at this age. Therefore, in addition to increasing MT gene transcription, these results suggest a post-transcriptional effect of IL-6 or of a IL-6-dependent factor, in this chronic situation. The up-regulated brain MT-I + II protein levels in the GFAP-IL6 mice was comparable to the expression of the acute-phase response gene EB22/5, suggesting that these MT isoforms could be considered acute-phase response proteins in the brain. Brain MT-III mRNA levels followed a somewhat similar pattern that those of MT-I mRNA but the decreasing effect of IL-6 transgene

  18. Fully functional HLA B27-restricted CD4+ as well as CD8+ T cell responses in TCR transgenic mice.

    PubMed

    Roddis, Matthew; Carter, Robert W; Sun, Mei-Yi; Weissensteiner, Thomas; McMichael, Andrew J; Bowness, Paul; Bodmer, Helen C

    2004-01-01

    The strong association of HLA B27 with spondyloarthropathies contrasts strikingly with most autoimmune diseases, which are HLA class II associated and thought to be mediated by CD4+ T lymphocytes. By introducing a human-derived HLA B27-restricted TCR into HLA B27 transgenic mice, we have obtained a functional TCR transgenic model, GRb, dependent on HLA B27 for response. Surprisingly, HLA B27 supported CD4+ as well as CD8+ T cell responses in vivo and in vitro. Further, HLA B27-restricted CD4+ T cells were capable of differentiation into a range of Th1 and Th2 T cell subsets with normal patterns of cytokine expression. The transgenic T cells were also able to enhance clearance of recombinant vaccinia virus containing influenza nucleoprotein in vivo. This is the first description of a human HLA class I-restricted TCR transgenic line. The existence of CD4+ MHC class I-restricted T cells has significant implications for immune regulation in autoimmunity and, in particular, in HLA B27-associated arthritis. We believe that this model provides a novel system for the study of unusual T cell behavior in vivo. PMID:14688321

  19. Two primate-specific small non-protein-coding RNAs in transgenic mice: neuronal expression, subcellular localization and binding partners

    PubMed Central

    Khanam, Tasneem; Rozhdestvensky, Timofey S.; Bundman, Marsha; Galiveti, Chenna R.; Handel, Sergej; Sukonina, Valentina; Jordan, Ursula; Brosius, Jürgen; Skryabin, Boris V.

    2007-01-01

    In a rare occasion a single chromosomal locus was targeted twice by independent Alu-related retroposon insertions, and in both cases supported neuronal expression of the respective inserted genes encoding small non-protein coding RNAs (npcRNAs): BC200 RNA in anthropoid primates and G22 RNA in the Lorisoidea branch of prosimians. To avoid primate experimentation, we generated transgenic mice to study neuronal expression and protein binding partners for BC200 and G22 npcRNAs. The BC200 gene, with sufficient upstream flanking sequences, is expressed in transgenic mouse brain areas comparable to those in human brain, and G22 gene, with upstream flanks, has a similar expression pattern. However, when all upstream regions of the G22 gene were removed, expression was completely abolished, despite the presence of intact internal RNA polymerase III promoter elements. Transgenic BC200 RNA is transported into neuronal dendrites as it is in human brain. G22 RNA, almost twice as large as BC200 RNA, has a similar subcellular localization. Both transgenically expressed npcRNAs formed RNP complexes with poly(A) binding protein and the heterodimer SRP9/14, as does BC200 RNA in human. These observations strongly support the possibility that the independently exapted npcRNAs have similar functions, perhaps in translational regulation of dendritic protein biosynthesis in neurons of the respective primates. PMID:17175535

  20. Xanthohumol Prevents Atherosclerosis by Reducing Arterial Cholesterol Content via CETP and Apolipoprotein E in CETP-Transgenic Mice

    PubMed Central

    Segawa, Shuichi; Ozaki, Moeko; Kobayashi, Naoyuki; Shigyo, Tatsuro; Chiba, Hitoshi

    2012-01-01

    Background Xanthohumol is expected to be a potent anti-atherosclerotic agent due to its inhibition of cholesteryl ester transfer protein (CETP). In this study, we hypothesized that xanthohumol prevents atherosclerosis in vivo and used CETP-transgenic mice (CETP-Tg mice) to evaluate xanthohumol as a functional agent. Methodology/Principal Findings Two strains of mice, CETP-Tg and C57BL/6N (wild-type), were fed a high cholesterol diet with or without 0.05% (w/w) xanthohumol ad libitum for 18 weeks. In CETP-Tg mice, xanthohumol significantly decreased accumulated cholesterol in the aortic arch and increased HDL cholesterol (HDL-C) when compared to the control group (without xanthohumol). Xanthohumol had no significant effect in wild-type mice. CETP activity was significantly decreased after xanthohumol addition in CETP-Tg mice compared with the control group and it inversely correlated with HDL-C (%) (P<0.05). Furthermore, apolipoprotein E (apoE) was enriched in serum and the HDL-fraction in CETP-Tg mice after xanthohumol addition, suggesting that xanthohumol ameliorates reverse cholesterol transport via apoE-rich HDL resulting from CETP inhibition. Conclusions Our results suggest xanthohumol prevents cholesterol accumulation in atherogenic regions by HDL-C metabolism via CETP inhibition leading to apoE enhancement. PMID:23166663

  1. Spontaneous generation of rapidly transmissible prions in transgenic mice expressing wild-type bank vole prion protein.

    PubMed

    Watts, Joel C; Giles, Kurt; Stöhr, Jan; Oehler, Abby; Bhardwaj, Sumita; Grillo, Sunny K; Patel, Smita; DeArmond, Stephen J; Prusiner, Stanley B

    2012-02-28

    Currently, there are no animal models of the most common human prion disorder, sporadic Creutzfeldt-Jakob disease (CJD), in which prions are formed spontaneously from wild-type (WT) prion protein (PrP). Interestingly, bank voles (BV) exhibit an unprecedented promiscuity for diverse prion isolates, arguing that bank vole PrP (BVPrP) may be inherently prone to adopting misfolded conformations. Therefore, we constructed transgenic (Tg) mice expressing WT BVPrP. Tg(BVPrP) mice developed spontaneous CNS dysfunction between 108 and 340 d of age and recapitulated the hallmarks of prion disease, including spongiform degeneration, pronounced astrogliosis, and deposition of alternatively folded PrP in the brain. Brain homogenates of ill Tg(BVPrP) mice transmitted disease to Tg(BVPrP) mice in ∼35 d, to Tg mice overexpressing mouse PrP in under 100 d, and to WT mice in ∼185 d. Our studies demonstrate experimentally that WT PrP can spontaneously form infectious prions in vivo. Thus, Tg(BVPrP) mice may be useful for studying the spontaneous formation of prions, and thus may provide insight into the etiology of sporadic CJD. PMID:22331873

  2. Extended survival of misfolded G85R SOD1-linked ALS mice by transgenic expression of chaperone Hsp110

    PubMed Central

    Nagy, Maria; Fenton, Wayne A.; Li, Di; Furtak, Krystyna; Horwich, Arthur L.

    2016-01-01

    Recent studies have indicated that mammalian cells contain a cytosolic protein disaggregation machinery comprised of Hsc70, DnaJ homologs, and Hsp110 proteins, the last of which acts to accelerate a rate-limiting step of nucleotide exchange of Hsc70. We tested the ability of transgenic overexpression of a Thy1 promoter-driven human Hsp110 protein, HspA4L (Apg1), in neuronal cells of a transgenic G85R SOD1YFP ALS mouse strain to improve survival. Notably, G85R is a mutant version of Cu/Zn superoxide dismutase 1 (SOD1) that is unable to reach native form and that is prone to aggregation, with prominent YFP-fluorescent aggregates observed in the motor neurons of the transgenic mice as early as 1 mo of age. The several-fold overexpression of Hsp110 in motor neurons of these mice was associated with an increased median survival from ∼5.5 to 7.5 mo and increased maximum survival from 6.5 to 12 mo. Improvement of survival was also observed for a G93A mutant SOD1 ALS strain. We conclude that neurodegeneration associated with cytosolic misfolding and aggregation can be ameliorated by overexpression of Hsp110, likely enhancing the function of a cytosolic disaggregation machinery. PMID:27114530

  3. Transgenic mouse milk expressing human bile salt-stimulated lipase improves the survival and growth status of premature mice.

    PubMed

    Wang, Yuanyuan; Sheng, Zheya; Wang, Yuhang; Li, Qinghe; Gao, Yu; Wang, Yuhui; Dai, Yunping; Liu, George; Zhao, Yaofeng; Li, Ning

    2015-03-01

    The lactating human mammary gland and the pancreas both produce bile salt-stimulated lipase (BSSL), a lipolytic enzyme acting on a wide range of substrates, including triglyceride, cholesterol esters, and fat-soluble vitamins esters. Breast milk BSSL has a particularly important role in the digestion of milk fat by newborn infants. We report the generation of transgenic mice that harbored a human BSSL gene controlled by a mammary gland-specific promoter. BSSL levels in transgenic mouse milk were raised to 376.8 μg/ml, corresponding to an activity of 9.15 U/ml. Premature wild-type neonates nursed by transgenic dams exhibited significantly higher survival rate than did the control neonates nursed by wild dams (95 vs. 83.3 % and, P < 0.05). They also showed 43.8 % greater body weight gain and 33.3 % lesser fecal crude fat levels than did the controls. This study provides significant evidence that increased levels of BSSL in milk may reduce mortality and improve the growth and fat absorption in premature mice during neonatal development. PMID:25385005

  4. The targeted overexpression of a Claudin mutant in the epidermis of transgenic mice elicits striking epidermal and hair follicle abnormalities.

    PubMed

    Troy, Tammy-Claire; Turksen, Kursad

    2007-06-01

    Skin is one of the largest organs of the body, and is formed during development through a highly orchestrated process involving mesenchymal-epithelial interactions, cell commitment, and terminal differentiation. It protects against microorganism invasion and UV irradiation, inhibits water loss, regulates body temperature, and is an important part of the immune system. Using transgenic mouse technology, we have demonstrated that Claudin (Cldn)-containing tight junctions (TJs) are intricately involved in cell signaling during epidermal differentiation and that an epidermal suprabasal overexpression of Cldn6 results in a perturbed epidermal terminal differentiation program with distinct phenotypic abnormalities. To delineate the role of the Cldn cytoplasmic tail domain in epidermal differentiation, we engineered transgenic mice targeting the overexpression of a Cldn6 cytoplasmic tail-truncation mutant in the epidermis. Transgenic mice were characterized by a lethal barrier dysfunction in addition to the existence of hyperproliferative squamous invaginations/cysts replacing hair follicles. Immunohistochemical analysis revealed an epidermal cytoplasmic accumulation of Cldn6, Cldn11, Cldn12, and Cldn18, downregulation of Cldn1 and aberrant expression of various classical markers of epidermal differentiation; namely the basal keratins as well as K1, involucrin, loricrin, and filaggrin. Collectively these studies suggest an important role for Cldns in epidermal/hair follicle differentiation programs likely involving cross talk to signaling pathways (e.g., Notch) directing cell fate selection and differentiation. PMID:17914196

  5. Regional brain cytochrome oxidase activity in beta-amyloid precursor protein transgenic mice with the Swedish mutation.

    PubMed

    Strazielle, C; Sturchler-Pierrat, C; Staufenbiel, M; Lalonde, R

    2003-01-01

    Cytochrome oxidase activity was examined in a transgenic mouse model of Alzheimer's disease with overexpression of the 751 amino acid isoform of beta-amyloid precursor protein with the Swedish mutation under control of the murine thy-1 promoter. The neuritic plaques, abundantly localized in the hippocampus and anterior neocortical areas, showed a core devoid of enzymatic activity surrounded by higher cytochrome oxidase activity at the sites of the dystrophic neurites and activated glial cells. Quantitative measures, taken only in the healthy-appearing regional areas without neuritic plaques, were higher in numerous limbic and non-limbic regions of transgenic mice in comparison with controls. Enzymatic activity was higher in the dentate gyrus and CA2-CA3 region of the hippocampus, the anterior cingulate and primary visual cortex, two olfactory structures, the ventral part of the neostriatum, the parafascicularis nucleus of the thalamus, and the subthalamic nucleus. Brainstem regions anatomically related with altered forebrain regions were more heavily labeled as well, including the substantia nigra, the periaqueductal gray, the superior colliculus, the medial raphe, the locus coeruleus and the adjacent parabrachial nucleus, as well as the pontine nuclei, red nucleus, and trigeminal motor nucleus. Functional brain organization is discussed in the context of Alzheimer's disease. Although hypometabolism is generally observed in this pathology, the increased cytochrome oxidase activity obtained in these transgenic mice can be the result of a functional compensation on the surviving neurons, or of an early mitochondrial alteration related to increased oxidative damage. PMID:12732258

  6. Transgenic overexpression of the alpha-synuclein interacting protein synphilin-1 leads to behavioral and neuropathological alterations in mice.

    PubMed

    Nuber, Silke; Franck, Thomas; Wolburg, Hartwig; Schumann, Ulrike; Casadei, Nicolas; Fischer, Kristina; Calaminus, Carsten; Pichler, Bernd J; Chanarat, Sittinan; Teismann, Peter; Schulz, Jörg B; Luft, Andreas R; Tomiuk, Jürgen; Wilbertz, Johannes; Bornemann, Antje; Krüger, Rejko; Riess, Olaf

    2010-02-01

    Synphilin-1 has been identified as an interacting protein of alpha-synuclein, Parkin, and LRRK2, proteins which are mutated in familial forms of Parkinson disease (PD). Subsequently, synphilin-1 has also been shown to be an intrinsic component of Lewy bodies in sporadic PD. In order to elucidate the role of synphilin-1 in the pathogenesis of PD, we generated transgenic mice overexpressing wild-type and mutant (R621C) synphilin-1 driven by a mouse prion protein promoter. Transgenic expression of both wild-type and the R621C variant synphilin-1 resulted in increased dopamine levels of the nigrostriatal system in 3-month-old mice. Furthermore, we found pathological ubiquitin-positive inclusions in cerebellar sections and dark-cell degeneration of Purkinje cells. Both transgenic mouse lines showed significant reduction of motor skill learning and motor performance. These findings suggest a pathological role of overexpressed synphilin-1 in vivo and will help to further elucidate the mechanisms of protein aggregation and neuronal cell death. PMID:19760259

  7. Extended survival of misfolded G85R SOD1-linked ALS mice by transgenic expression of chaperone Hsp110.

    PubMed

    Nagy, Maria; Fenton, Wayne A; Li, Di; Furtak, Krystyna; Horwich, Arthur L

    2016-05-10

    Recent studies have indicated that mammalian cells contain a cytosolic protein disaggregation machinery comprised of Hsc70, DnaJ homologs, and Hsp110 proteins, the last of which acts to accelerate a rate-limiting step of nucleotide exchange of Hsc70. We tested the ability of transgenic overexpression of a Thy1 promoter-driven human Hsp110 protein, HspA4L (Apg1), in neuronal cells of a transgenic G85R SOD1YFP ALS mouse strain to improve survival. Notably, G85R is a mutant version of Cu/Zn superoxide dismutase 1 (SOD1) that is unable to reach native form and that is prone to aggregation, with prominent YFP-fluorescent aggregates observed in the motor neurons of the transgenic mice as early as 1 mo of age. The several-fold overexpression of Hsp110 in motor neurons of these mice was associated with an increased median survival from ∼5.5 to 7.5 mo and increased maximum survival from 6.5 to 12 mo. Improvement of survival was also observed for a G93A mutant SOD1 ALS strain. We conclude that neurodegeneration associated with cytosolic misfolding and aggregation can be ameliorated by overexpression of Hsp110, likely enhancing the function of a cytosolic disaggregation machinery. PMID:27114530

  8. Testis hormone-sensitive lipase expression in spermatids is governed by a short promoter in transgenic mice.

    PubMed

    Blaise, R; Guillaudeux, T; Tavernier, G; Daegelen, D; Evrard, B; Mairal, A; Holm, C; Jégou, B; Langin, D

    2001-02-16

    A testicular form of hormone-sensitive lipase (HSL(tes)), a triacylglycerol lipase, and cholesterol esterase, is expressed in male germ cells. Northern blot analysis showed HSL(tes) mRNA expression in early spermatids. Immunolocalization of the protein in human and rodent seminiferous tubules indicated that the highest level of expression occurred in elongated spermatids. We have previously shown that 0.5 kilobase pairs of the human HSL(tes) promoter directs testis-specific expression of a chloramphenicol acetyltransferase reporter gene in transgenic mice and determined regions binding nuclear proteins expressed in testis but not in liver (Blaise, R., Grober, J., Rouet, P., Tavernier, G., Daegelen, D., and Langin, D. (1999) J. Biol. Chem. 274, 9327-9334). Mutation of a SRY/Sox-binding site in one of the regions did not impair in vivo testis-specific expression of the reporter gene. Further transgenic analyses established that 95 base pairs upstream of the transcription start site were sufficient for correct testis expression. In gel retardation assays using early spermatid nuclear extracts, a germ cell-specific DNA-protein interaction was mapped between -46 and -29 base pairs. The DNA binding nuclear protein showed properties of zinc finger transcription factors. Mutation of the region abolished reporter gene activity in transgenic mice, showing that it is necessary for testis expression of HSL(tes). PMID:11076952

  9. Efficient generation of transgenic mice by lentivirus-mediated modification of spermatozoa.

    PubMed

    Chandrashekran, Anil; Sarkar, Rupa; Thrasher, Adrian; Fraser, Scott E; Dibb, Nicholas; Casimir, Colin; Winston, Robert; Readhead, Carol

    2014-02-01

    Transgenic technologies conventionally rely on the oocyte as a substrate for genetic modification. Owing to their accessibility, however, male germ cells, including mature sperm, have material advantages for use in transgenesis. Here we have exploited lentiviruses to generate transgenic animals via the male germline. When pseudotyped lentiviral vectors encoding green fluorescent protein (GFP) were incubated with mouse spermatozoa, these sperm were highly successful in producing transgenics. Lentivirally transduced mouse spermatozoa were used in in vitro fertilization (IVF) studies, and when followed by embryo transfer, ≥ 42% of founders were found to be transgenic for GFP. Inverse PCR strategy for integration site analysis demonstrated integration of at least 1 or 2 copies of GFP in the transgenics, mapping to different chromosomes. GFP expression was detected in a wide range of murine tissues, including testis and the transgene was stably transmitted to a third generation of transgenic animals. This relatively simple, yet highly efficient, technique for generating transgenic animals by transducing spermatozoa with lentiviral vectors in vitro is a powerful tool for the study of fertilization/preimplantation development, vertical viral gene transmission, gene function and regulation, and epigenetic inheritance. PMID:24297703

  10. Postnatal male germ-cell expression of cre recombinase in Tex101-iCre transgenic mice.

    PubMed

    Lei, Zhenmin; Lin, Jing; Li, Xian; Li, Shengqiang; Zhou, Huaxin; Araki, Yoshihiko; Lan, Zi-Jian

    2010-12-01

    We have generated a transgenic mouse line that expresses improved Cre recombinase (iCre) under the control of the testis-expressed gene 101 (Tex101) promoter. This transgenic mouse line was named Tex101-iCre. Using the floxed ROSA reporter mice, we found that robust Cre recombinase activity was detected in postnatal testes with weak or no activity in other tissues. Within the testis, Cre recombinase was active in spermatogenic cells as early as the prospermatogonia stage at day 1 after birth. In 30- and 60-day-old mice, positive Cre recombinase activity was detected not only in prospermatogonia but also in spermatogenic cells at later stages of spermatogenesis. There was little or no Cre activity in interstitial cells. Breeding wild-type females with homozygous floxed fibroblast growth factor receptor 2 (Fgfr2) males carrying the Tex101-iCre transgene did not produce any progeny with the floxed Fgfr2 allele. All the progeny inherited a recombined Fgfr2 allele, indicating that complete deletion of the floxed Fgfr2 allele by Tex101-iCre can be achieved in the male germline. Furthermore, FGFR2 protein was not detected in spermatocytes and spermatids of adult Fgfr2(fl/fl) ;Tex101-iCre mice. Taken together, our results suggest that the Tex101-iCre mouse line allows the inactivation of a floxed gene in spermatogenic cells in adult mice, which will facilitate the functional characterization of genes in normal spermatogenesis and male fertility. PMID:20853429

  11. Skeletal phenotype of growing transgenic mice that express a function-perturbing form of beta1 integrin in osteoblasts

    NASA Technical Reports Server (NTRS)

    Globus, R. K.; Amblard, D.; Nishimura, Y.; Iwaniec, U. T.; Kim, J-B; Almeida, E. A. C.; Damsky, C. D.; Wronski, T. J.; van der Meulen, M. C. H.

    2005-01-01

    Skeletal modeling entails the deposition of large amounts of extracellular matrix (ECM) to form structures tailored to withstand increasing mechanical loads during rapid growth. Specific ECM molecules bind to integrin receptors on the cell surface, thereby triggering a cascade of signaling events that affect critical cell functions. To evaluate the role of integrins during skeletal growth, transgenic mice were engineered to express a function-perturbing fragment of beta1 integrin consisting of the transmembrane domain and cytoplasmic tail under the control of the osteocalcin promoter (TG mice). Thus, transgene expression was targeted to mature cells of the osteoblast lineage, and herein we show that cultured cells resembling osteocytes from 90-day-old TG mice display impaired adhesion to collagen I, a ligand for beta1 integrin. To determine the influence of beta1 integrin on bones that are responsible for providing structural support during periods of rapid growth, we examined the phenotype of the appendicular skeleton in TG mice compared to wild type (WT) mice. According to radiographs, bones from mice of both genotypes between 14 and 90 days of age appeared similar in gross structure and density, although proximal tibiae from 35-90 days old TG mice were less curved than those of WT mice (72-92% TG/WT). Although there were only mild and transient differences in absolute bone mass and strength, once normalized to body mass, the tibial dry mass (79.1% TG/WT females), ash mass (78.5% TG/WT females), and femoral strength in torsion (71.6% TG/WT females) were reduced in TG mice compared to WT mice at 90 days of age. Similar effects of genotype on bone mass and curvature were observed in 1-year-old retired breeders, indicating that these phenotypic differences between TG and WT mice were stable well into adulthood. Effects of genotype on histomorphometric indices of cancellous bone turnover were minimal and evident only transiently during growth, but when present they

  12. Transgenic complementation of MeCP2 deficiency: phenotypic rescue of Mecp2-null mice by isoform-specific transgenes

    PubMed Central

    Kerr, Bredford; Soto C, Jessica; Saez, Mauricio; Abrams, Alexander; Walz, Katherina; Young, Juan I

    2012-01-01

    Rett syndrome (RTT) is a disorder that affects patients' ability to communicate, move and behave. RTT patients are characterized by impaired language, stereotypic behaviors, frequent seizures, ataxia and sleep disturbances, with the onset of symptoms occurring after a period of seemingly normal development. RTT is caused by mutations in methyl-CpG binding protein 2 (MECP2), an X-chromosome gene encoding for MeCP2, a protein that regulates gene expression. MECP2 generates two alternative splice variants encoding two protein isoforms that differ only in the N-terminus. Although no functional differences have been identified for these splice variants, it has been suggested that the RTT phenotype may occur in the presence of a functional MeCP2-e2 protein. This suggests that the two isoforms might be functionally distinct. Supporting this notion, the two variants show regional and age-related differences in transcript abundance. Here, we show that transgenic expression of either the MeCP2-e1 or MeCP2-e2 splice variant results in prevention of development of RTT-like phenotypic manifestations in a mouse model lacking Mecp2. Our results indicate that the two MeCP2 splice variants can substitute for each other and fulfill the basic functions of MeCP2 in the mouse brain. PMID:21829232

  13. Transgenic complementation of MeCP2 deficiency: phenotypic rescue of Mecp2-null mice by isoform-specific transgenes.

    PubMed

    Kerr, Bredford; Soto C, Jessica; Saez, Mauricio; Abrams, Alexander; Walz, Katherina; Young, Juan I

    2012-01-01

    Rett syndrome (RTT) is a disorder that affects patients' ability to communicate, move and behave. RTT patients are characterized by impaired language, stereotypic behaviors, frequent seizures, ataxia and sleep disturbances, with the onset of symptoms occurring after a period of seemingly normal development. RTT is caused by mutations in methyl-CpG binding protein 2 (MECP2), an X-chromosome gene encoding for MeCP2, a protein that regulates gene expression. MECP2 generates two alternative splice variants encoding two protein isoforms that differ only in the N-terminus. Although no functional differences have been identified for these splice variants, it has been suggested that the RTT phenotype may occur in the presence of a functional MeCP2-e2 protein. This suggests that the two isoforms might be functionally distinct. Supporting this notion, the two variants show regional and age-related differences in transcript abundance. Here, we show that transgenic expression of either the MeCP2-e1 or MeCP2-e2 splice variant results in prevention of development of RTT-like phenotypic manifestations in a mouse model lacking Mecp2. Our results indicate that the two MeCP2 splice variants can substitute for each other and fulfill the basic functions of MeCP2 in the mouse brain. PMID:21829232

  14. Allergic inflammatory response to short ragweed allergenic extract in HLA-DQ transgenic mice lacking CD4 gene.

    PubMed

    Chapoval, Svetlana P; Iijima, Koji; Marietta, Eric V; Smart, Michele K; Chapoval, Andrei I; Andrews, Amy G; David, Chella S

    2002-01-15

    To investigate the role of HLA-DQ molecules and/or CD4(+) T cells in the pathogenesis of allergic asthma, we generated HLA-DQ6 and HLA-DQ8 transgenic mice lacking endogenous class II (Abeta(null)) and CD4 genes and challenged them intranasally with short ragweed allergenic extract (SRW). We found that DQ6/CD4(null) mice developed a strong eosinophilic infiltration into the bronchoalveolar lavage and lung tissue, while DQ8/CD4(null) mice were normal. However, neither cytokines nor eosinophil peroxidase in the bronchoalveolar lavage of DQ6/CD4(null) mice was found. In addition, the airway reactivity to methacholine was elevated moderately in DQ6/CD4(null) mice compared with the high response in DQ/CD4(+) counterparts and was only partially augmented by CD4(+) T cell transfer. The DQ6/CD4(null) mice showed Th1/Th2-type cytokines and SRW-specific Abs in the immune sera in contrast to a direct Th2 response observed in DQ6/CD4(+) mice. The proliferative response of spleen mononuclear cells and peribronchial lymph node cells demonstrated that the response to SRW in DQ6/CD4(null) mice was mediated by HLA-DQ-restricted CD4(-)CD8(-)NK1.1(-) T cells. FACS analysis of PBMC and spleen mononuclear cells demonstrated an expansion of double-negative (DN) CD4(-)CD8(-)TCRalphabeta(+) T cells in SRW-treated DQ6/CD4(null) mice. These cells produced IL-4, IL-5, IL-13, and IFN-gamma when stimulated with immobilized anti-CD3. IL-5 ELISPOT assay revealed that DN T cells were the cellular origin of IL-5 in allergen-challenged DQ6/CD4(null) mice. Our study shows a role for HLA-DQ-restricted CD4(+) and DN T cells in the allergic response. PMID:11777987

  15. Differential susceptibity of transgenic mice lacking one or both apolipoprotein alleles to folate and vitamin E deprivation.

    PubMed

    Shea, Thomas B; Ortiz, Daniela; Rogers, Eugene

    2004-06-01

    The E4 allele of apolipoprotein E (ApoE) is associated with neurodegeneration in part due to increased oxidative stress. Transgenic mice lacking ApoE (-/-) represent a model for the consequences of deficiencies in ApoE function. Dietary deficiency in folate and vitamin E has previously been shown to potentiate the impact of ApoE deficiency; ApoE-/- mice deprived of folate and vitamin E for 1 month demonstrated increased oxidative damage in brain tissue and impaired cognitive performance as compared to ApoE+/+ mice. Since individuals homozygous for E4 can demonstrate more increased risk for neurodegeneration and an earlier age of onset than individuals heterozygous for E4, we tested the impact of folate and vitamin E deprivation on ApoE+/- mice. Thiobarbituric acid-reactive substances in brain tissue of ApoE+/- were significantly increased compared to ApoE+/+ mice, but this increase was less than that observed in ApoE-/- mice. By contrast, livers of ApoE+/- and -/- mice displayed an identical increase over that of +/+ mice. ApoE-/- mice, but not +/- or +/+ mice, exhibited impaired cognitive performance in maze trials when deprived of folate and vitamin E. These findings support the notion that homozygous deficiency of ApoE function can be more severe than heterozygous deficiency. They further suggest that the impact of partial deficiency in ApoE function may present a latent risk that may manifest only when compounded by other factors such as dietary deficiency. PMID:15201481

  16. Differential susceptibility of transgenic mice expressing human surfactant protein B genetic variants to Pseudomonas aeruginosa induced pneumonia.

    PubMed

    Ge, Lin; Liu, Xinyu; Chen, Rimei; Xu, Yongan; Zuo, Yi Y; Cooney, Robert N; Wang, Guirong

    2016-01-01

    Surfactant protein B (SP-B) is essential for lung function. Previous studies have indicated that a SP-B 1580C/T polymorphism (SNP rs1130866) was associated with lung diseases including pneumonia. The SNP causes an altered N-linked glycosylation modification at Asn129 of proSP-B, e.g. the C allele with this glycosylation site but not in the T allele. This study aimed to generate humanized SP-B transgenic mice carrying either SP-B C or T allele without a mouse SP-B background and then examine functional susceptibility to bacterial pneumonia in vivo. A total of 18 transgenic mouse founders were generated by the DNA microinjection method. These founders were back-crossed with SP-B KO mice to eliminate mouse SP-B background. Four founder lines expressing similar SP-B levels to human lung were chosen for further investigation. After intratracheal infection with 50 μl of Pseudomonas aeruginosa solution (1 × 10(6) CFU/mouse) or saline in SP-B-C, SP-B-T mice the mice were sacrificed 24 h post-infection and tissues were harvested. Analysis of surfactant activity revealed differential susceptibility between SP-B-C and SP-B-T mice to bacterial infection, e.g. higher minimum surface tension in infected SP-B-C versus infected SP-B-T mice. These results demonstrate for the first time that human SP-B C allele is more susceptible to bacterial pneumonia than SP-B T allele in vivo. PMID:26620227

  17. Differential expression of cholesteryl ester transfer protein in the liver and plasma of fasted and fed transgenic mice.

    PubMed

    MacLean, P S; Vadlamudi, S; Hao, E; Barakat, H A

    2000-06-01

    Because cholesteryl ester transfer protein (CETP) is considered a potential target in the treatment of atherosclerosis, several reports have focused on the regulation of this enzyme, and there is evidence that insulin may be a regulatory factor. The present study examines the differential expression of the human CETP gene between physiologic conditions that are accompanied by low (fasted) and high (fed) insulin levels. CETP expression was examined in plasma and tissues of transgenic mice expressing the human CETP minigene after 12 hours of fasting (n = 20) or ad libitum feeding (n = 20) with normal mouse chow. Plasma cholesteryl ester transfer activity (CETA) was 20% higher in fed than in fasted mice, reflecting higher levels of CETP (P < 0.05). This observation was accompanied by higher liver mRNA in fed mice (100%, P < 0.05), as determined by ribonuclease protection assays, as well as by higher CETA (23%, P < 0.05) and CETP mass (29%, P < 0.05) in the particulate fraction of liver homogenates. These parameters of liver CETP expression correlated well with each other, as well as with plasma CETA. CETP in the liver particulate fraction was found as a doublet (approximately 70 and 65 kDa), which resolved to a single band (approximately 60 kDa) upon deglycosylation. No differences in CETP expression were observed in pooled adipose tissue samples from fed and fasted mice. Insulin and glucose were not related to any plasma or tissue parameter of CETP expression. In summary, the concerted, differential expression of CETP in the liver of fed and fasted transgenic mice appears to contribute to higher plasma CETP levels in fed mice, but the precise role of insulin and glucose in regulating CETP expression under fasted and fed conditions needs to be defined. PMID:11002127

  18. Generation of an ABCG2{sup GFPn-puro} transgenic line - A tool to study ABCG2 expression in mice

    SciTech Connect

    Orford, Michael; Mean, Richard; Lapathitis, George; Genethliou, Nicholas; Panayiotou, Elena; Panayi, Helen; Malas, Stavros

    2009-06-26

    The ATP-binding cassette (ABC) transporter 2 (ABCG2) is expressed by stem cells in many organs and in stem cells of solid tumors. These cells are isolated based on the side population (SP) phenotype, a Hoechst 3342 dye efflux property believed to be conferred by ABCG2. Because of the limitations of this approach we generated transgenic mice that express Nuclear GFP (GFPn) coupled to the Puromycin-resistance gene, under the control of ABCG2 promoter/enhancer sequences. We show that ABCG2 is expressed in neural progenitors of the developing forebrain and spinal cord and in embryonic and adult endothelial cells of the brain. Using the neurosphere assay, we isolated tripotent ABCG2-expressing neural stem cells from embryonic mouse brain. This transgenic line is a powerful tool for studying the expression of ABCG2 in many tissues and for performing functional studies in different experimental settings.

  19. Cardiac and skeletal myopathy in beta myosin heavy-chain simian virus 40 tsA58 transgenic mice.

    PubMed Central

    De Leon, J R; Federoff, H J; Dickson, D W; Vikstrom, K L; Fishman, G I

    1994-01-01

    The mechanisms regulating cardiac muscle differentiation and development are incompletely understood. To examine the relationships between cardiocyte proliferation and differentiation, we tested the ability of a fragment from the rat beta myosin heavy-chain (MHC beta) gene to correctly target expression of a thermolabile simian virus 40 large tumor antigen allele (tsA58) in the developing mouse. Transgene expression in the heart was observed as early as 10 days postconception and was developmentally regulated in parallel with the endogenous MHC beta gene. Expression was also detected in developing skeletal muscle, although at low levels. Despite the temperature sensitivity of the mutant large tumor antigen protein, a subset of transgenic mice in several lineages developed marked cardiac and skeletal myopathies. Images Fig. 2 Fig. 3 Fig. 4 PMID:8290557

  20. Cell-type-specific and hypoxia-inducible expression of the human erythropoietin gene in transgenic mice.

    PubMed Central

    Semenza, G L; Koury, S T; Nejfelt, M K; Gearhart, J D; Antonarakis, S E

    1991-01-01

    Synthesis of erythropoietin, the primary humoral regulator of erythropoiesis, in liver and kidney is inducible by anemia or hypoxia. Analysis of human erythropoietin gene expression in transgenic mice revealed that sequences located 6-14 kilobases 5' to the gene direct expression to the kidney, whereas sequences within the immediate 3'-flanking region control hepatocyte-specific expression. Human erythropoietin transcription initiation sites were differentially utilized in liver and kidney. Inducible transgene expression was precisely targeted to peritubular interstitial cells in the renal cortex that synthesize endogenous mouse erythropoietin. These studies demonstrate that multiple erythropoietin gene regulatory elements control cell-type-specific expression and inducibility by a fundamental physiologic stimulus, hypoxia. Images PMID:1924331

  1. Expression of Autoactivated Stromelysin-1 in Mammary Glands of Transgenic Mice Leads to a Reactive Stroma During Early Development

    SciTech Connect

    Thomasset, N.; Lochter, A.; Sympson, C.J.; Lund, L.R.; Williams, D.R.; Behrendtsen, O.; Werb, Z.; Bissell, M.J.

    1998-04-24

    Extracellular matrix and extracellular matrix-degrading matrix metalloproteinases play a key role in interactions between the epithelium and the mesenchyme during mammary gland development and disease. In patients with breast cancer, the mammary mesenchyme undergoes a stromal reaction, the etiology of which is unknown. We previously showed that targeting of an autoactivating mutant of the matrix metalloproteinase stromelysin-1 to mammary epithelia of transgenic mice resulted in reduced mammary function during pregnancy and development of preneoplastic and neoplastic lesions. Here we examine the cascade of alterations before breast tumor formation in the mammary gland stroma once the expression of the stromelysin-1 transgene commences. Beginning in postpubertal virgin animals, low levels of transgene expression in mammary epithelia led to increased expression of endogenous stromelysin-1 in stromal fibroblasts and up-regulation of other matrix metalloproteinases, without basement membrane disruption. These changes were accompanied by the progressive development of a compensatory reactive stroma, characterized by increased collagen content and vascularization in glands from virgin mice. This remodeling of the gland affected epithelial-mesenchymal communication as indicated by inappropriate expression of tenascin-C starting by day 6 of pregnancy. This, together with increased transgene expression, led to basement membrane disruption starting by day 15 of pregnancy. We propose that the highly reactive stroma provides a prelude to breast epithelial tumors observed in these animals. Epithelial development depends on an exquisite series of inductive and instructive interactions between the differentiating epithelium and the mesenchymal (stromal) compartment. The epithelium, which consists of luminal and myoepithelial cells, is separated from the stroma by a basement membrane (BM), which plays a central role in mammary gland homeostasis and gene expression. In vivo, stromal

  2. Progressive paralysis associated with diffuse astrocyte anaplasia in delta 202 mice homozygous for a transgene encoding the SV40 T antigen.

    PubMed

    López-Revilla, Rubén; Soto-Zárate, Carlos; Ridaura, Cecilia; Chávez-Dueñas, Lucía; Paul, Dieter

    2004-03-01

    A convenient transgenic astrocytoma model in delta202 mice, homozygous for a construct encoding the early region of the SV40 virus genome, is described. In the offspring of crosses between delta202 mice heterozygous for the transgene nearly 60% were transgenic; one third of these developed progressive paralysis starting in the hindlimbs at approximately 35 days of age and died at 90 +/- 30 days of age. In affected mice proliferating-non-neuronal cells immunostained with antibodies to the GFAP, an astrocyte marker, whose number increased with age were found in the white matter of the brain, cerebellum and spinal cord, and progressive degeneration and necrosis of spinal motoneurons was observed that-may explain the paralysis. The early onset and reproducible time course of the neurological disease suggest that homozygous delta202 mice, whose proliferating astrocytes appear to damage spinal motoneurons, are a useful model to study astrocyte differentiation, function and tumorigenesis. PMID:15068170

  3. Cyclin D2 Overexpression in Transgenic Mice Induces Thymic and Epidermal Hyperplasia whereas Cyclin D3 Expression Results Only in Epidermal Hyperplasia

    PubMed Central

    Rodriguez-Puebla, Marcelo L.; LaCava, Margaret; Miliani de Marval, Paula L.; Jorcano, Jose L.; Richie, Ellen R.; Conti, Claudio J.

    2000-01-01

    In a previous report, we described the effects of cyclin D1 expression in epithelial tissues of transgenic mice. To study the involvement of D-type cyclins (D1, D2, and D3) in epithelial growth and differentiation and their putative role as oncogenes in skin, transgenic mice were developed which carry cyclin D2 or D3 genes driven by a keratin 5 promoter. As expected, both transgenic lines showed expression of these proteins in most of the squamous tissues analyzed. Epidermal proliferation increased in transgenic animals and basal cell hyperplasia was observed. All of the animals also had a minor thickening of the epidermis. The pattern of expression of keratin 1 and keratin 5 indicated that epidermal differentiation was not affected. Transgenic K5D2 mice developed mild thymic hyperplasia that reversed at 4 months of age. On the other hand, high expression of cyclin D3 in the thymus did not produce hyperplasia. This model provides in vivo evidence of the action of cyclin D2 and cyclin D3 as mediators of proliferation in squamous epithelial cells. A direct comparison among the three D-type cyclin transgenic mice suggests that cyclin D1 and cyclin D2 have similar roles in epithelial thymus cells. However, overexpression of each D-type cyclin produces a distinct phenotype in thymic epithelial cells. PMID:10980142

  4. Glucagon gene 5'-flanking sequences direct expression of simian virus 40 large T antigen to the intestine, producing carcinoma of the large bowel in transgenic mice.

    PubMed

    Lee, Y C; Asa, S L; Drucker, D J

    1992-05-25

    Glucagon and the glucagon-like peptides play important roles in the regulation of glucose homeostasis. Previous studies have demonstrated that approximately 1300 base pairs of rat glucagon gene 5'-flanking sequences direct transgene expression to the pancreas and brain, but not to the intestine, of transgenic mice. These observations suggested that different tissue-specific enhancer elements mediate activation of glucagon gene transcription in the pancreas and intestine. We have now generated mice that express SV40 large T antigen under the control of approximately 2000 base pairs of glucagon gene 5'-flanking sequences. Transgene expression was observed in the brain and pancreas in association with the development of pancreatic endocrine tumors. In contrast to the mice described previously, we also detected transgene expression throughout the gastrointestinal tract in endocrine cells of the stomach and small and large intestine. Focal areas of enteroendocrine cell hyperplasia in the large bowel invariably progressed to invasive and metastasizing plurihormonal endocrine carcinoma, which was clinically and pathologically evident by 4 weeks of age. In contrast, transgene expression in the small bowel and stomach was not associated with progression to either hyperplasia or carcinoma. The results of these studies provide functional evidence for the existence of an upstream cis-acting regulatory domain that directs glucagon gene transcription to the endocrine cells of the intestine in transgenic mice. PMID:1587847

  5. Body composition and grip strength are improved in transgenic sickle mice fed a high-protein diet.

    PubMed

    Capers, Patrice L; Hyacinth, Hyacinth I; Cue, Shayla; Chappa, Prasanthi; Vikulina, Tatyana; Roser-Page, Susanne; Weitzmann, M Neale; Archer, David R; Newman, Gale W; Quarshie, Alexander; Stiles, Jonathan K; Hibbert, Jacqueline M

    2015-01-01

    Key pathophysiology of sickle cell anaemia includes compensatory erythropoiesis, vascular injury and chronic inflammation, which divert amino acids from tissue deposition for growth/weight gain and muscle formation. We hypothesised that sickle mice maintained on an isoenergetic diet with a high percentage of energy derived from protein (35 %), as opposed to a standard diet with 20 % of energy derived from protein, would improve body composition, bone mass and grip strength. Male Berkeley transgenic sickle mice (S; n 8-12) were fed either 20 % (S20) or 35 % (S35) diets for 3 months. Grip strength (BIOSEB meter) and body composition (dual-energy X-ray absorptiometry scan) were measured. After 3 months, control mice had the highest bone mineral density (BMD) and bone mineral content (BMC) (P < 0·005). S35 mice had the largest increase in grip strength. A two-way ANOVA of change in grip strength (P = 0·043) attributed this difference to genotype (P = 0·025) and a trend in type of diet (P = 0·067). l-Arginine (l-Arg) supplementation of the 20 % diet was explored, as a possible mechanism for improvement obtained with the 35 % diet. Townes transgenic sickle mice (TS; n 6-9) received 0·8, 1·6, 3·2 or 6·4 % l-Arg based on the same protocol and outcome measures used for the S mice. TS mice fed 1·6 % l-Arg for 3 months (TS1.6) had the highest weight gain, BMD, BMC and lean body mass compared with other groups. TS3.2 mice showed significantly more improvement in grip strength than TS0·8 and TS1.6 mice (P < 0·05). In conclusion, the high-protein diet improved body composition and grip strength. Outcomes observed with TS1.6 and TS3.2 mice, respectively, confirm the hypothesis and reveal l-Arg as part of the mechanism. PMID:26090102

  6. Blockade of NF-κB using IκBκ dominant negative mice ameliorates cardiac hypertrophy in myotrophin overexpressed transgenic mice

    PubMed Central

    Young, David; Popovic, Zoran B.; Jones, W. Keith; Gupta, Sudhiranjan

    2008-01-01

    NF-κB is a ubiquitous transcription factor that regulates various kinds of genes including inflammatory molecules, macrophage infiltration factors, cell adhesion molecules, etc., in various disease processes including cardiac hypertrophy and heart failure (HF). Previously, we have demonstrated that activation of NF-κB was required in myotrophin induced cardiac hypertrophy, in spontaneously hypertensive rats (SHR) and in dilated cardiomyopathy (DCM) human hearts. Moreover, our recent study using the myotrophin overexpressed transgenic mouse (Myo-Tg) model showed that shRNA-mediated knock down of NF-κB significantly attenuated cardiac mass associated with improved cardiac function. Although, it has been shown that NF-κB is substantially involved in cardiovascular remodeling, it is not clear whether the continuous blockade of NF-κB is effective in cardiovascular remodeling. To address this question, we took a genetic approach using IκBα triple mutant mice (3M) bred with Myo-Tg mice (a progressive hypertrophy/HF model). The double transgenic mice (Myo-3M) displayed an attenuated cardiac hypertrophy (9.8 ± 0.62 vs 5.4 ± 0.34, p<0.001) and improved cardiac function associated with significant inhibition of the NF-κB signaling cascade, hypertrophy marker gene expression, inflammatory and macrophage gene expression at 24 weeks of age compared to Myo-Tg mice. NF-κB-targeted gene array profiling displayed several important genes were significantly down regulated in Myo-3M mice compared to Myo-Tg mice. Furthermore, Myo-3M did not show any changes of apoptotic gene expression indicating that complete inhibition of NF-κB activation reduces further pro-inflammatory reactions without affecting susceptibility to apoptosis. Therefore, development of therapeutic strategies targeting NF-κB may provide an effective approach to prevent adverse cardiac pathophysiological consequences. PMID:18620706

  7. Enhanced aggressiveness of benzopyrene-induced squamous carcinomas in transgenic mice overexpressing the proprotein convertase PACE4 (PCSK6).

    PubMed

    Bassi, Daniel E; Cenna, Jonathan; Zhang, Jirong; Cukierman, Edna; Klein-Szanto, Andres J

    2015-10-01

    PACE4 (PCSK6) is a proprotein convertase (PC) capable of processing numerous substrates involved in tumor growth, invasion, and metastasis. Because of the human relevancy of the tobacco-associated carcinogen benzo[a]pyrene (B(a)P) we investigated whether transgenic mice in which this PC is targeted to the epidermis (K5-PACE4) may be more susceptible to B(a)P complete carcinogenesis than wild type (WT) mice. In an in vitro experiment, using cell lines derived from skin tumors obtained after B(a)P treatment, we observed that PACE4 overexpression and activity accounts for an increased proliferation rate, exaggerated sensitivity to the PC inhibitor CMK, and interference with IGF-1R autophosphorylation. Squamous cell carcinomas, obtained from K5-PACE4 mice subjected to complete chemical carcinogenesis, were characterized by a 50% increase in cell proliferation, when compared with similar tumors from WT mice. In addition, tumors from K5-PACE4 mice showed deeper invasion into the underlying dermis. Thus, mice overexpressing PACE4 exhibited tumors of increased growth rate and invasive potential when exposed to the human carcinogen B(a)P, further supporting the significance of PCs in tumor growth and progression. PMID:24845697

  8. Inhibition of the HER2 pathway by n-3 polyunsaturated fatty acids prevents breast cancer in fat-1 transgenic mice

    PubMed Central

    Zou, Zuquan; Bellenger, Sandrine; Massey, Karen A.; Nicolaou, Anna; Geissler, Audrey; Bidu, Célia; Bonnotte, Bernard; Pierre, Anne-Sophie; Minville-Walz, Mélaine; Rialland, Michaël; Seubert, John; Kang, Jing X.; Lagrost, Laurent; Narce, Michel; Bellenger, Jérôme

    2013-01-01

    Overexpression of the tyrosine kinase receptor, ErbB2/HER2/Neu, occurs in 25–30% of invasive breast cancer (BC) with poor patient prognosis. Due to confounding factors, inconsistencies still remain regarding the protective effects of n-3 polyunsaturated fatty acids (PUFAs) on BC. We therefore evaluated whether fat-1 transgenic mice, endogenously synthesizing n-3 PUFAs from n-6 PUFAs, were protected against BC development, and we then aimed to study in vivo a mechanism potentially involved in such protection. E0771 BC cells were implanted into fat-1 and wild-type (WT) mice. After tumorigenesis examination, we analyzed the expression of proteins involved in the HER2 signaling pathway and lipidomic analyses were performed in tumor tissues and plasma. Our results showed that tumors totally disappeared by day 15 in fat-1 mice but continued to grow in WT mice. This prevention can be related in part to significant repression of the HER2/β-catenin signaling pathway and formation of significant levels of n-3 PUFA-derived bioactive mediators (particularly 15-hydroxyeicosapentaenoic acid, 17-hydroxydocosahexaenoic acid, and prostaglandin E3) in the tumors of fat-1 mice compared with WT mice. All together these data demonstrate an anti-BC effect of n-3 PUFAs through, at least in part, HER2 signaling pathway downregulation, and highlight the importance of gene-diet interactions in BC. PMID:24052576

  9. AP2-NR4A3 transgenic mice display reduced serum epinephrine because of increased catecholamine catabolism in adipose tissue.

    PubMed

    Walton, R Grace; Zhu, Xiaolin; Tian, Ling; Heywood, Elizabeth B; Liu, Jian; Hill, Helliner S; Liu, Jiarong; Bruemmer, Dennis; Yang, Qinglin; Fu, Yuchang; Garvey, W Timothy

    2016-07-01

    The NR4A orphan nuclear receptors function as early response genes to numerous stimuli. Our laboratory has previously demonstrated that overexpression of NR4A3 (NOR-1, MINOR) in 3T3-L1 adipocytes enhances insulin-stimulated glucose uptake. To assess the in vivo effect of NR4A3 on adipocytes, we generated transgenic mice with NR4A3 overexpression driven by the adipocyte fatty acid-binding protein (AP2) promoter (AP2-NR4A3 mice). We hypothesized that AP2-NR4A3 mice would display enhanced glucose tolerance and insulin sensitivity. However, AP2-NR4A3 mice exhibit metabolic impairment, including increased fasting glucose and insulin, impaired glucose tolerance, insulin resistance, decreased serum free fatty acids, and increased low-density lipoprotein-cholesterol. AP2-NR4A3 mice also display a significant reduction in serum epinephrine due to increased expression of catecholamine-catabolizing enzymes in adipose tissue, including monoamine oxidase-A. Furthermore, enhanced expression of monoamine oxidase-A is due to direct transcriptional activation by NR4A3. Finally, AP2-NR4A3 mice display cardiac and behavioral alterations consistent with chronically low circulating epinephrine levels. In conclusion, overexpression of NR4A3 in adipocytes produces a complex phenotype characterized by impaired glucose metabolism and low serum catecholamines due to enhanced degradation by adipose tissue. PMID:27166283

  10. Influence of fatty acid-free diet on mammary tumor development and growth rate in HER-2/Neu transgenic mice.

    PubMed

    Rossini, Anna; Zanobbio, Laura; Sfondrini, Lucia; Cavalleri, Adalberto; Secreto, Giorgio; Morelli, Daniele; Palazzo, Marco; Sommariva, Michele; Tagliabue, Elda; Rumio, Cristiano; Balsari, Andrea

    2013-01-01

    Numerous investigations have found a relationship between higher risk of cancer and increased intake of fats, while results of clinical studies of fat reduction and breast cancer recurrence have been mixed. A diet completely free of fats cannot be easily administered to humans, but experimental studies in mice can be done to determine whether this extreme condition influences tumor development. Here, we examined the effects of a FA-free diet on mammary tumor development and growth rate in female FVB-neu proto-oncogene transgenic mice that develop spontaneous multifocal mammary tumors after a long latency period. Mice were fed a fatty acid-free diet beginning at 112, 35, and 30 days of age. In all these experiments, tumor appearance was delayed, tumor incidence was reduced and the mean number of palpable mammary tumors per mouse was lower, as compared to standard diet-fed mice. By contrast, tumor growth rate was unaffected in mice fed the fatty acid-free diet. Plasma of mice fed the fatty acid-free diet revealed significantly higher contents of oleic, palmitoleic and 20:3ω9 acids and lower contents of linoleic and palmitic acids. In conclusion, these findings indicate that a FA-free diet reduces tumor incidence and latency but not tumor growth rate, suggesting that a reduction in dietary FAs in humans may have a protective effect on tumorigenesis but not on tumors once they appear. PMID:22689438

  11. Transgenic mice with inactive alleles for procollagen N-proteinase (ADAMTS-2) develop fragile skin and male sterility.

    PubMed Central

    Li, S W; Arita, M; Fertala, A; Bao, Y; Kopen, G C; Långsjö, T K; Hyttinen, M M; Helminen, H J; Prockop, D J

    2001-01-01

    Transgenic mice were prepared with inactive alleles for procollagen N-proteinase (ADAMTS-2; where ADAMTS stands for a disintegrin and metalloproteinase with thrombospondin repeats). Homozygous mice were grossly normal at birth, but after 1-2 months they developed thin skin that tore after gentle handling. Although the gene was inactivated, a large fraction of the N-propeptides of type I procollagen in skin and the N-propeptides of type II procollagen in cartilage were cleaved. Therefore the results suggested the tissues contained one or more additional enzymes that slowly process the proteins. Electron microscopy did not reveal any defects in the morphology of collagen fibrils in newborn mice. However, in two-month-old mice, the collagen fibrils in skin were seen as bizarre curls in cross-section and the mean diameters of the fibrils were approx. half of the controls. Although a portion of the N-propeptides of type II procollagen in cartilage were not cleaved, no defects in the morphology of the fibrils were seen by electron microscopy or by polarized-light microscopy. Female homozygous mice were fertile, but male mice were sterile with a marked decrease in testicular sperm. Therefore the results indicated that ADAMTS-2 plays an essential role in the maturation of spermatogonia. PMID:11284712

  12. Running Exercise Reduces Myelinated Fiber Loss in the Dentate Gyrus of the Hippocampus in APP/PS1 Transgenic Mice.

    PubMed

    Chao, Fenglei; Zhang, Lei; Luo, Yanmin; Xiao, Qian; Lv, Fulin; He, Qi; Zhou, Chunni; Zhang, Yi; Jiang, Lin; Jiang, Rong; Gu, Hengwei; Tang, Yong

    2015-01-01

    To investigate the effect of running exercise on myelinated fibers in the dentate gyrus (DG) of the hippocampus during Alzheimer's disease (AD), 6-month-old male APP/PS1 transgenic mice were randomly assigned to control or running groups. The running group mice were subjected to a running protocol for four months. The behaviors of the mice from both group mice were then assessed using the Morris water maze, and the total volume of the DG and the related quantitative parameters with characteristics of the myelinated nerve fiber and the myelin sheath in the DG were investigated using unbiased stereological techniques and electron microscopy. Learning and spatial memory performances were both significantly increased in the running group compared with the control group. There was no significant difference in the gratio of the myelinated axons between the two groups. However, the DG volume, the myelinated fiber length and volume in the DG, and the myelin sheath volume and thickness in the DG were all significantly increased in the running group mice compared with the control group mice. These results indicated that running exercise was able to prevent DG atrophy and delay the progression of the myelinated fiber loss and the demyelination of the myelin sheaths in the DG in an AD mouse model, which may underlie the running-induced improvement in learning and spatial memory. Taken together, these results demonstrated that running exercise could delay the progression of AD. PMID:25817255