Transgenic and gene knockout mice in gastric cancer research

PubMed Central

Jiang, Yannan; Yu, Yingyan

2017-01-01

Mouse models are useful tool for carcinogenic study. They will greatly enrich the understanding of pathogenesis and molecular mechanisms for gastric cancer. However, only few of mice could develop gastric cancer spontaneously. With the development and improvement of gene transfer technology, investigators created a variety of transgenic and knockout/knockin mouse models of gastric cancer, such as INS-GAS mice and gastrin knockout mice. Combined with helicobacter infection and carcinogens treatment, these transgenic/knockout/knockin mice developed precancerous or cancerous lesions, which are proper for gene function study or experimental therapy. Here we review the progression of genetically engineered mouse models on gastric cancer research, and emphasize the effects of chemical carcinogens or infectious factors on carcinogenesis of genetically modified mouse. We also emphasize the histological examination on mouse stomach. We expect to provide researchers with some inspirations on this field. PMID:27713138

Exacerbation of experimental autoimmune encephalomyelitis by passive transfer of IgG antibodies from a multiple sclerosis patient responsive to immunoadsorption.

PubMed

Pedotti, Rosetta; Musio, Silvia; Scabeni, Stefano; Farina, Cinthia; Poliani, Pietro Luigi; Colombo, Emanuela; Costanza, Massimo; Berzi, Angela; Castellucci, Fabrizio; Ciusani, Emilio; Confalonieri, Paolo; Hemmer, Bernhard; Mantegazza, Renato; Antozzi, Carlo

2013-09-15

The pathogenic role of antibodies in multiple sclerosis (MS) is still controversial. We transferred to mice with experimental autoimmune encephalomyelitis (EAE), animal model of MS, IgG antibodies purified from a MS patient presenting a dramatic clinical improvement during relapse after selective IgG removal with immunoadsorption. Passive transfer of patient's IgG exacerbated motor paralysis and increased mouse central nervous system (CNS) inflammation and demyelination. Binding of patient's IgG was demonstrated in mouse CNS, with a diffuse staining of white matter oligodendrocytes. These data support a growing body of evidence that antibodies can play an important role in the pathobiology of MS. Copyright © 2013 Elsevier B.V. All rights reserved.

Strategies and Considerations for Distributing and Recovering Mouse Lines

PubMed Central

Du, Yubin; Xie, Wen; Liu, Chengyu

2012-01-01

As more and more genetically modified mouse lines are being generated, it becomes increasingly common to share animal models among different research institutions. Live mice are routinely transferred between animal facilities. Due to various issues concerning animal welfare, intellectual property rights, colony health status and biohazard, significant paperwork and coordination are required before any animal travel can take place. Shipping fresh or frozen preimplantation embryos, gametes, or reproductive organs can bypass some of the issues associated with live animal transfer, but it requires the receiving facilities to be able to perform delicate and sometimes intricate procedures such as embryo transfer, in vitro fertilization (IVF), or ovary transplantation. Here, we summarize the general requirements for live animal transport and review some of the assisted reproductive technologies (ART) that can be applied to shipping and reviving mouse lines. Intended users of these methods should consult their institution’s responsible official to find out whether each specific method is legal or appropriate in their own animal facilities. PMID:20691859

Aberrant muscle antigen exposure in mice is sufficient to cause myositis in a Treg cell-deficient milieu.

PubMed

Young, Nicholas A; Sharma, Rahul; Friedman, Alexandra K; Kaffenberger, Benjamin H; Bolon, Brad; Jarjour, Wael N

2013-12-01

Myositis is associated with muscle-targeted inflammation and is observed in some Treg cell-deficient mouse models. Because an autoimmune pathogenesis has been strongly implicated, the aim of this study was to investigate the hypothesis that abnormal exposure to muscle antigens, as observed in muscle injury, can induce autoimmune-mediated myositis in susceptible hosts. FoxP3 mutant (scurfy) mice were mated to synaptotagmin VII (Syt VII) mutant mice, which resulted in a new mouse strain that combines impaired membrane resealing with Treg cell deficiency. Lymphocyte preparations from double-mutant mice were adoptively transferred intraperitoneally, with or without purified Treg cells, into recombination-activating gene 1 (RAG-1)-null recipients. Lymph node cells from mice with the FoxP3 mutation were transferred into RAG-1-null mice either 1) intraperitoneally in conjunction with muscle homogenate or purified myosin protein or 2) intramuscularly with or without cotransfer of purified Treg cells. FoxP3-deficient mouse lymph node cells transferred in conjunction with myosin protein or muscle homogenate induced robust skeletal muscle inflammation. The infiltrates consisted predominantly of CD4+ and CD8+ T cells, a limited number of macrophages, and no B cells. Significant inflammation was also seen in similar experiments using lymph node cells from FoxP3/Syt VII double-mutant mice but was absent in experiments using adoptive transfer of FoxP3 mutant mouse cells alone. The cotransfer of Treg cells completely suppressed myositis. These data, derived from a new, reproducible model, demonstrate the critical roles of Treg cell deficiency and aberrant muscle antigen exposure in the priming of autoreactive cells to induce myositis. This mouse system has multifaceted potential for examining the interplay in vivo between tissue injury and autoimmunity. © 2013 The Authors. Arthritis & Rheumatism is published by Wiley Periodicals, Inc. on behalf of the American College of Rheumatology.

Somatic Cell Nuclear Transfer in the Mouse

NASA Astrophysics Data System (ADS)

Kishigami, Satoshi; Wakayama, Teruhiko

Somatic cell nuclear transfer (SCNT) has become a unique and powerful tool for epigenetic reprogramming research and gene manipulation in animals since “Dolly,” the first animal cloned from an adult cell was reported in 1997. Although the success rates of somatic cloning have been inefficient and the mechanism of reprogramming is still largely unknown, this technique has been proven to work in more than 10 mammalian species. Among them, the mouse provides the best model for both basic and applied research of somatic cloning because of its abounding genetic resources, rapid sexual maturity and propagation, minimal requirements for housing, etc. This chapter describes a basic protocol for mouse cloning using cumulus cells, the most popular cell type for NT, in which donor nuclei are directly injected into the oocyte using a piezo-actuated micromanipulator. In particular, we focus on a new, more efficient mouse cloning protocol using trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, which increases both in vitro and in vivo developmental rates from twofold to fivefold. This new method including TSA will be helpful to establish mouse cloning in many laboratories.

Targeting CD6 for the treatment of experimental autoimmune uveitis.

PubMed

Zhang, Lingjun; Li, Yan; Qiu, Wen; Bell, Brent A; Dvorina, Nina; Baldwin, William M; Singer, Nora; Kern, Timothy; Caspi, Rachel R; Fox, David A; Lin, Feng

2018-06-01

CD6 is emerging as a new target for treating many pathological conditions in which T cells are integrally involved, but even the latest data from studies of CD6 gene engineered mice were still contradictory. To address this issue, we studied experimental autoimmune uveitis (EAU), a model of autoimmune uveitis, in wild-type (WT) and CD6 knockout (KO) mice. After EAU induction in WT and CD6 KO mice, we evaluated ocular inflammation and compared retinal antigen-specific T-cell responses using scanning laser ophthalmoscopy, spectral-domain optical coherence tomography, histopathology, and T cell recall assays. Uveitogenic T cells from WT and CD6 KO mice were adoptively transferred into WT naïve mice to confirm the impact of CD6 on T cells. In addition, we immunized CD6 KO mice with recombinant CD6 protein to develop mouse anti-mouse CD6 monoclonal antibodies (mAbs) in which functional antibodies exhibiting cross-reactivity with human CD6 were screened and identified for treatment studies. In CD6 KO mice with EAU, we found significantly decreased retinal inflammation and reduced autoreactive T-cell responses, and confirmed the impaired uveitogenic capacity of T cells from these mice in an adoptive transfer experiment. Notably, one of these cross-reactive mAbs significantly ameliorated retinal inflammation in EAU induced by the adoptive transfer of uveitogenic T cells. Together, these data strongly suggest that CD6 plays a previously unknown, but pivotal role in autoimmune uveitis, and may be a promising new treatment target for this blinding disease. In addition, the newly developed mouse anti-mouse/human CD6 mAbs could be valuable tools for testing CD6-targeted therapies in other mouse models of human diseases. Copyright © 2018 Elsevier Ltd. All rights reserved.

Correction of mouse ornithine transcarbamylase deficiency by gene transfer into the germ line.

PubMed Central

Cavard, C; Grimber, G; Dubois, N; Chasse, J F; Bennoun, M; Minet-Thuriaux, M; Kamoun, P; Briand, P

1988-01-01

The sparse fur with abnormal skin and hair (Spf-ash) mouse is a model for the human X-linked hereditary disorder, ornithine transcarbamylase (OTC) deficiency. In Spf-ash mice, both OTC mRNA and enzyme activity are 5% of control values resulting in hyperammonemia, pronounced orotic aciduria and an abnormal phenotype characterized by growth retardation and sparse fur. Using microinjection, we introduced a construction containing rat OTC cDNA linked to the SV40 early promoter into fertilized eggs of Spf-ash mice. The expression of the transgene resulted in the development of a transgenic mouse whose phenotype and orotic acid excretion are fully normalized. Thus, the possibility of correcting hereditary enzymatic defect by gene transfer of heterologous cDNA coding for the normal enzyme has been demonstrated. Images PMID:3162766

A Novel Form of Compensation in the Tg2576 Amyloid Mouse Model of Alzheimer’s Disease

PubMed Central

Somogyi, Attila; Katonai, Zoltán; Alpár, Alán; Wolf, Ervin

2016-01-01

One century after its first description, pathology of Alzheimer’s disease (AD) is still poorly understood. Amyloid-related dendritic atrophy and membrane alterations of susceptible brain neurons in AD, and in animal models of AD are widely recognized. However, little effort has been made to study the potential effects of combined morphological and membrane alterations on signal transfer and synaptic integration in neurons that build up affected neural networks in AD. In this study spatial reconstructions and electrophysiological measurements of layer II/III pyramidal neurons of the somatosensory cortex from wild-type (WT) and transgenic (TG) human amyloid precursor protein (hAPP) overexpressing Tg2576 mice were used to build faithful segmental cable models of these neurons. Local synaptic activities were simulated in various points of the dendritic arbors and properties of subthreshold dendritic impulse propagation and predictors of synaptic input pattern recognition ability were quantified and compared in modeled WT and TG neurons. Despite the widespread dendritic degeneration and membrane alterations in mutant mouse neurons, surprisingly little, or no change was detected in steady-state and 50 Hz sinusoidal voltage transfers, current transfers, and local and propagation delays of PSPs traveling along dendrites of TG neurons. Synaptic input pattern recognition ability was also predicted to be unaltered in TG neurons in two different soma-dendritic membrane models investigated. Our simulations predict the way how subthreshold dendritic signaling and pattern recognition are preserved in TG neurons: amyloid-related membrane alterations compensate for the pathological effects that dendritic atrophy has on subthreshold dendritic signal transfer and integration in layer II/III somatosensory neurons of this hAPP mouse model for AD. Since neither propagation of single PSPs nor integration of multiple PSPs (pattern recognition) changes in TG neurons, we conclude that AD-related neuronal hyperexcitability cannot be accounted for by altered subthreshold dendritic signaling in these neurons but hyperexcitability is related to changes in active membrane properties and network connectivity. PMID:27378850

A Novel Form of Compensation in the Tg2576 Amyloid Mouse Model of Alzheimer's Disease.

PubMed

Somogyi, Attila; Katonai, Zoltán; Alpár, Alán; Wolf, Ervin

2016-01-01

One century after its first description, pathology of Alzheimer's disease (AD) is still poorly understood. Amyloid-related dendritic atrophy and membrane alterations of susceptible brain neurons in AD, and in animal models of AD are widely recognized. However, little effort has been made to study the potential effects of combined morphological and membrane alterations on signal transfer and synaptic integration in neurons that build up affected neural networks in AD. In this study spatial reconstructions and electrophysiological measurements of layer II/III pyramidal neurons of the somatosensory cortex from wild-type (WT) and transgenic (TG) human amyloid precursor protein (hAPP) overexpressing Tg2576 mice were used to build faithful segmental cable models of these neurons. Local synaptic activities were simulated in various points of the dendritic arbors and properties of subthreshold dendritic impulse propagation and predictors of synaptic input pattern recognition ability were quantified and compared in modeled WT and TG neurons. Despite the widespread dendritic degeneration and membrane alterations in mutant mouse neurons, surprisingly little, or no change was detected in steady-state and 50 Hz sinusoidal voltage transfers, current transfers, and local and propagation delays of PSPs traveling along dendrites of TG neurons. Synaptic input pattern recognition ability was also predicted to be unaltered in TG neurons in two different soma-dendritic membrane models investigated. Our simulations predict the way how subthreshold dendritic signaling and pattern recognition are preserved in TG neurons: amyloid-related membrane alterations compensate for the pathological effects that dendritic atrophy has on subthreshold dendritic signal transfer and integration in layer II/III somatosensory neurons of this hAPP mouse model for AD. Since neither propagation of single PSPs nor integration of multiple PSPs (pattern recognition) changes in TG neurons, we conclude that AD-related neuronal hyperexcitability cannot be accounted for by altered subthreshold dendritic signaling in these neurons but hyperexcitability is related to changes in active membrane properties and network connectivity.

Immunocompetent Mouse Model for Tracking Cancer Progression | NCI Technology Transfer Center | TTC

Cancer.gov

The National Cancer Institute seeks licensees or research collaborators to develop and commercialize transgenic mice having immunocompetent rat growth hormone-firefly Luciferase-enhanced green fluorescent protein.

The Bruton Tyrosine Kinase (BTK) Inhibitor Acalabrutinib Demonstrates Potent On-Target Effects and Efficacy in Two Mouse Models of Chronic Lymphocytic Leukemia.

PubMed

Herman, Sarah E M; Montraveta, Arnau; Niemann, Carsten U; Mora-Jensen, Helena; Gulrajani, Michael; Krantz, Fanny; Mantel, Rose; Smith, Lisa L; McClanahan, Fabienne; Harrington, Bonnie K; Colomer, Dolors; Covey, Todd; Byrd, John C; Izumi, Raquel; Kaptein, Allard; Ulrich, Roger; Johnson, Amy J; Lannutti, Brian J; Wiestner, Adrian; Woyach, Jennifer A

2017-06-01

Purpose: Acalabrutinib (ACP-196) is a novel, potent, and highly selective Bruton tyrosine kinase (BTK) inhibitor, which binds covalently to Cys481 in the ATP-binding pocket of BTK. We sought to evaluate the antitumor effects of acalabrutinib treatment in two established mouse models of chronic lymphocytic leukemia (CLL). Experimental Design: Two distinct mouse models were used, the TCL1 adoptive transfer model where leukemic cells from Eμ-TCL1 transgenic mice are transplanted into C57BL/6 mice, and the human NSG primary CLL xenograft model. Mice received either vehicle or acalabrutinib formulated into the drinking water. Results: Utilizing biochemical assays, we demonstrate that acalabrutinib is a highly selective BTK inhibitor as compared with ibrutinib. In the human CLL NSG xenograft model, treatment with acalabrutinib demonstrated on-target effects, including decreased phosphorylation of PLCγ2, ERK, and significant inhibition of CLL cell proliferation. Furthermore, tumor burden in the spleen of the mice treated with acalabrutinib was significantly decreased compared with vehicle-treated mice. Similarly, in the TCL1 adoptive transfer model, decreased phosphorylation of BTK, PLCγ2, and S6 was observed. Most notably, treatment with acalabrutinib resulted in a significant increase in survival compared with mice receiving vehicle. Conclusions: Treatment with acalabrutinib potently inhibits BTK in vivo , leading to on-target decreases in the activation of key signaling molecules (including BTK, PLCγ2, S6, and ERK). In two complementary mouse models of CLL, acalabrutinib significantly reduced tumor burden and increased survival compared with vehicle treatment. Overall, acalabrutinib showed increased BTK selectivity compared with ibrutinib while demonstrating significant antitumor efficacy in vivo on par with ibrutinib. Clin Cancer Res; 23(11); 2831-41. ©2016 AACR . ©2016 American Association for Cancer Research.

The Bruton’s tyrosine kinase (BTK) inhibitor acalabrutinib demonstrates potent on-target effects and efficacy in two mouse models of chronic lymphocytic leukemia

PubMed Central

Herman, Sarah E. M.; Montraveta, Arnau; Niemann, Carsten U.; Mora-Jensen, Helena; Gulrajani, Michael; Krantz, Fanny; Mantel, Rose; Smith, Lisa L.; McClanahan, Fabienne; Harrington, Bonnie K.; Colomer, Dolors; Covey, Todd; Byrd, John C.; Izumi, Raquel; Kaptein, Allard; Ulrich, Roger; Johnson, Amy J.; Lannutti, Brian J.; Wiestner, Adrian; Woyach, Jennifer A.

2017-01-01

Purpose Acalabrutinib (ACP-196) is a novel, potent, and highly selective BTK inhibitor, which binds covalently to Cys481 in the ATP-binding pocket of BTK. We sought to evaluate the anti-tumor effects of acalabrutinib treatment in two established mouse models of chronic lymphocytic leukemia (CLL). Experimental Design Two distinct mouse models were used, the TCL1 adoptive transfer model where leukemic cells from Eμ-TCL1 transgenic mice are transplanted into C57BL/6 mice, and the human NSG primary CLL xenograft model. Mice received either vehicle or acalabrutinib formulated into the drinking water. Results Utilizing biochemical assays we demonstrate that acalabrutinib is a highly selective BTK inhibitor as compared to ibrutinib. In the human CLL NSG xenograft model, treatment with acalabrutinib demonstrated on-target effects including decreased phosphorylation of PLCγ2, ERK and significant inhibition of CLL cell proliferation. Further, tumor burden in the spleen of the mice treated with acalabrutinib was significantly decreased compared to vehicle treated mice. Similarly, in the TCL1 adoptive transfer model, decreased phosphorylation of BTK, PLCγ2 and S6 was observed. Most notably, treatment with acalabrutinib resulted in a significant increase in survival compared to mice receiving vehicle. Conclusions Treatment with acalabrutinib potently inhibits BTK in vivo, leading to on-target decreases in the activation of key signaling molecules (including BTK, PLCγ2, S6 and ERK). In two complementary mouse models of CLL acalabrutinib significantly reduced tumor burden and increased survival compared to vehicle treatment. Overall, acalabrutinib showed increased BTK selectivity compared to ibrutinib while demonstrating significant anti-tumor efficacy in vivo on par with ibrutinib. PMID:27903679

Defective natural killer cell activity in a mouse model of eczema herpeticum.

PubMed

Kawakami, Yuko; Ando, Tomoaki; Lee, Jong-Rok; Kim, Gisen; Kawakami, Yu; Nakasaki, Tae; Nakasaki, Manando; Matsumoto, Kenji; Choi, Youn Soo; Kawakami, Toshiaki

2017-03-01

Patients with atopic dermatitis (AD) are susceptible to several viruses, including herpes simplex virus (HSV). Some patients experience 1 or more episodes of a severe skin infection caused by HSV termed eczema herpeticum (EH). There are numerous mouse models of AD, but no established model exists for EH. We sought to establish and characterize a mouse model of EH. We infected AD-like skin lesions with HSV1 to induce severe skin lesions in a dermatitis-prone mouse strain of NC/Nga. Gene expression was investigated by using a microarray and quantitative PCR; antibody titers were measured by means of ELISA; and natural killer (NK) cell, cytotoxic T-cell, regulatory T-cell, and follicular helper T-cell populations were evaluated by using flow cytometry. The role of NK cells in HSV1-induced development of severe skin lesions was examined by means of depletion and adoptive transfer. Inoculation of HSV1 induced severe erosive skin lesions in eczematous mice, which had an impaired skin barrier, but milder lesions in small numbers of normal mice. Eczematous mice exhibited lower NK cell activity but similar cytotoxic T-cell activity and humoral immune responses compared with normal mice. The role of NK cells in controlling HSV1-induced skin lesions was demonstrated by experiments depleting or transferring NK cells. A murine model of EH with an impaired skin barrier was established in this study. We demonstrated a critical role of defective NK activities in the development of HSV1-induced severe skin lesions in eczematous mice. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. All rights reserved.

T1, diffusion tensor, and quantitative magnetization transfer imaging of the hippocampus in an Alzheimer's disease mouse model.

PubMed

Whittaker, Heather T; Zhu, Shenghua; Di Curzio, Domenico L; Buist, Richard; Li, Xin-Min; Noy, Suzanna; Wiseman, Frances K; Thiessen, Jonathan D; Martin, Melanie

2018-07-01

Alzheimer's disease (AD) pathology causes microstructural changes in the brain. These changes, if quantified with magnetic resonance imaging (MRI), could be studied for use as an early biomarker for AD. The aim of our study was to determine if T 1 relaxation, diffusion tensor imaging (DTI), and quantitative magnetization transfer imaging (qMTI) metrics could reveal changes within the hippocampus and surrounding white matter structures in ex vivo transgenic mouse brains overexpressing human amyloid precursor protein with the Swedish mutation. Delineation of hippocampal cell layers using DTI color maps allows more detailed analysis of T 1 -weighted imaging, DTI, and qMTI metrics, compared with segmentation of gross anatomy based on relaxation images, and with analysis of DTI or qMTI metrics alone. These alterations are observed in the absence of robust intracellular Aβ accumulation or plaque deposition as revealed by histology. This work demonstrates that multiparametric quantitative MRI methods are useful for characterizing changes within the hippocampal substructures and surrounding white matter tracts of mouse models of AD. Copyright © 2018. Published by Elsevier Inc.

Limitations of gaze transfer: without visual context, eye movements do not to help to coordinate joint action, whereas mouse movements do.

PubMed

Müller, Romy; Helmert, Jens R; Pannasch, Sebastian

2014-10-01

Remote cooperation can be improved by transferring the gaze of one participant to the other. However, based on a partner's gaze, an interpretation of his communicative intention can be difficult. Thus, gaze transfer has been inferior to mouse transfer in remote spatial referencing tasks where locations had to be pointed out explicitly. Given that eye movements serve as an indicator of visual attention, it remains to be investigated whether gaze and mouse transfer differentially affect the coordination of joint action when the situation demands an understanding of the partner's search strategies. In the present study, a gaze or mouse cursor was transferred from a searcher to an assistant in a hierarchical decision task. The assistant could use this cursor to guide his movement of a window which continuously opened up the display parts the searcher needed to find the right solution. In this context, we investigated how the ease of using gaze transfer depended on whether a link could be established between the partner's eye movements and the objects he was looking at. Therefore, in addition to the searcher's cursor, the assistant either saw the positions of these objects or only a grey background. When the objects were visible, performance and the number of spoken words were similar for gaze and mouse transfer. However, without them, gaze transfer resulted in longer solution times and more verbal effort as participants relied more strongly on speech to coordinate the window movement. Moreover, an analysis of the spatio-temporal coupling of the transmitted cursor and the window indicated that when no visual object information was available, assistants confidently followed the searcher's mouse but not his gaze cursor. Once again, the results highlight the importance of carefully considering task characteristics when applying gaze transfer in remote cooperation. Copyright © 2013 Elsevier B.V. All rights reserved.

Rescue of bilirubin-induced neonatal lethality in a mouse model of Crigler-Najjar syndrome type I by AAV9-mediated gene transfer

PubMed Central

Bortolussi, Giulia; Zentilin, Lorena; Baj, Gabriele; Giraudi, Pablo; Bellarosa, Cristina; Giacca, Mauro; Tiribelli, Claudio; Muro, Andrés F.

2012-01-01

Crigler-Najjar type I (CNI) syndrome is a recessively inherited disorder characterized by severe unconjugated hyperbilirubinemia caused by uridine diphosphoglucuronosyltransferase 1A1 (UGT1A1) deficiency. The disease is lethal due to bilirubin-induced neurological damage unless phototherapy is applied from birth. However, treatment becomes less effective during growth, and liver transplantation is required. To investigate the pathophysiology of the disease and therapeutic approaches in mice, we generated a mouse model by introducing a premature stop codon in the UGT1a1 gene, which results in an inactive enzyme. Homozygous mutant mice developed severe jaundice soon after birth and died within 11 d, showing significant cerebellar alterations. To rescue neonatal lethality, newborns were injected with a single dose of adeno-associated viral vector 9 (AAV9) expressing the human UGT1A1. Gene therapy treatment completely rescued all AAV-treated mutant mice, accompanied by lower plasma bilirubin levels and normal brain histology and motor coordination. Our mouse model of CNI reproduces genetic and phenotypic features of the human disease. We have shown, for the first time, the full recovery of the lethal effects of neonatal hyperbilirubinemia. We believe that, besides gene-addition-based therapies, our mice could represent a very useful model to develop and test novel technologies based on gene correction by homologous recombination.—Bortolussi, G., Zentilin, L., Baj, G., Giraudi, P., Bellarosa, C., Giacca, M., Tiribelli, C., Muro, A. F. Rescue of bilirubin-induced neonatal lethality in a mouse model of Crigler-Najjar syndrome type I by AAV9-mediated gene transfer. PMID:22094718

Lessons learned using different mouse models during space radiation-induced lung tumorigenesis experiments

NASA Astrophysics Data System (ADS)

Wang, Jian; Zhang, Xiangming; Wang, Ping; Wang, Xiang; Farris, Alton B.; Wang, Ya

2016-06-01

Unlike terrestrial ionizing radiation, space radiation, especially galactic cosmic rays (GCR), contains high energy charged (HZE) particles with high linear energy transfer (LET). Due to a lack of epidemiologic data for high-LET radiation exposure, it is highly uncertain how high the carcinogenesis risk is for astronauts following exposure to space radiation during space missions. Therefore, using mouse models is necessary to evaluate the risk of space radiation-induced tumorigenesis; however, which mouse model is better for these studies remains uncertain. Since lung tumorigenesis is the leading cause of cancer death among both men and women, and low-LET radiation exposure increases human lung carcinogenesis, evaluating space radiation-induced lung tumorigenesis is critical to enable safe Mars missions. Here, by comparing lung tumorigenesis obtained from different mouse strains, as well as miR-21 in lung tissue/tumors and serum, we believe that wild type mice with a low spontaneous tumorigenesis background are ideal for evaluating the risk of space radiation-induced lung tumorigenesis, and circulating miR-21 from such mice model might be used as a biomarker for predicting the risk.

VEGF-C gene therapy augments postnatal lymphangiogenesis and ameliorates secondary lymphedema

PubMed Central

Yoon, Young-sup; Murayama, Toshinori; Gravereaux, Edwin; Tkebuchava, Tengiz; Silver, Marcy; Curry, Cynthia; Wecker, Andrea; Kirchmair, Rudolf; Hu, Chun Song; Kearney, Marianne; Ashare, Alan; Jackson, David G.; Kubo, Hajime; Isner, Jeffrey M.; Losordo, Douglas W.

2003-01-01

Although lymphedema is a common clinical condition, treatment for this disabling condition remains limited and largely ineffective. Recently, it has been reported that overexpression of VEGF-C correlates with increased lymphatic vessel growth (lymphangiogenesis). However, the effect of VEGF-C–induced lymphangiogenesis on lymphedema has yet to be demonstrated. Here we investigated the impact of local transfer of naked plasmid DNA encoding human VEGF-C (phVEGF-C) on two animal models of lymphedema: one in the rabbit ear and the other in the mouse tail. In a rabbit model, following local phVEGF-C gene transfer, VEGFR-3 expression was significantly increased. This gene transfer led to a decrease in thickness and volume of lymphedema, improvement of lymphatic function demonstrated by serial lymphoscintigraphy, and finally, attenuation of the fibrofatty changes of the skin, the final consequences of lymphedema. The favorable effect of phVEGF-C on lymphedema was reconfirmed in a mouse tail model. Immunohistochemical analysis using lymphatic-specific markers: VEGFR-3, lymphatic endothelial hyaluronan receptor-1, together with the proliferation marker Ki-67 Ab revealed that phVEGF-C transfection potently induced new lymphatic vessel growth. This study, we believe for the first time, documents that gene transfer of phVEGF-C resolves lymphedema through direct augmentation of lymphangiogenesis. This novel therapeutic strategy may merit clinical investigation in patients with lymphedema. PMID:12618526

Single-domain monoclonal antibodies for the treatment of hepatocellular carcinoma | NCI Technology Transfer Center | TTC

Cancer.gov

The National Cancer Institute seeks parties to license human monoclonal antibodies and immunoconjugates and co-develop, evaluate, and/or commercialize large-scale antibody production and hepatocellular carcinoma (HCC) xenograft mouse models.

Development of Gene Therapy for Thalassemia

PubMed Central

Nienhuis, Arthur W.; Persons, Derek A.

2012-01-01

Retroviral vector–mediated gene transfer into hematopoietic stem cells provides a potentially curative therapy for severe β-thalassemia. Lentiviral vectors based on human immunodeficiency virus have been developed for this purpose and have been shown to be effective in curing thalassemia in mouse models. One participant in an ongoing clinical trial has achieved transfusion independence after gene transfer into bone marrow stem cells owing, in part, to a genetically modified, dominant clone. Ongoing efforts are focused on improving the efficiency of lentiviral vector–mediated gene transfer into stem cells so that the curative potential of gene transfer can be consistently achieved. PMID:23125203

ART culture conditions change the probability of mouse embryo gestation through defined cellular and molecular responses.

PubMed

Schwarzer, Caroline; Esteves, Telma Cristina; Araúzo-Bravo, Marcos J; Le Gac, Séverine; Nordhoff, Verena; Schlatt, Stefan; Boiani, Michele

2012-09-01

Do different human ART culture protocols prepare embryos differently for post-implantation development? The type of ART culture protocol results in distinct cellular and molecular phenotypes in vitro at the blastocyst stage as well as subsequently during in vivo development. It has been reported that ART culture medium affects human development as measured by gestation rates and birthweights. However, due to individual variation across ART patients, it is not possible as yet to pinpoint a cause-effect relationship between choice of culture medium and developmental outcome. In a prospective study, 13 human ART culture protocols were compared two at a time against in vivo and in vitro controls. Superovulated mouse oocytes were fertilized in vivo using outbred and inbred mating schemes. Zygotes were cultured in medium or in the oviduct and scored for developmental parameters 96 h later. Blastocysts were either analyzed or transferred into fosters to measure implantation rates and fetal development. In total, 5735 fertilized mouse oocytes, 1732 blastocysts, 605 fetuses and 178 newborns were examined during the course of the study (December 2010-December 2011). Mice of the B6C3F1, C57Bl/6 and CD1 strains were used as oocyte donors, sperm donors and recipients for embryo transfer, respectively. In vivo fertilized B6C3F1 oocytes were allowed to cleave in 13 human ART culture protocols compared with mouse oviduct and optimized mouse medium (KSOM(aa)). Cell lineage composition of resultant blastocysts was analyzed by immunostaining and confocal microscopy (trophectoderm, Cdx2; primitive ectoderm, Nanog; primitive endoderm, Sox17), global gene expression by microarray analysis, and rates of development to midgestation and to term. Mouse zygotes show profound variation in blastocyst (49.9-91.9%) and fetal (15.7-62.0%) development rates across the 13 ART culture protocols tested (R(2)= 0.337). Two opposite protocols, human tubal fluid/multiblast (high fetal rate) and ISM1/ISM2 (low fetal rate), were analyzed in depth using outbred and inbred fertilization schemes. Resultant blastocysts show imbalances of cell lineage composition; culture medium-specific deviation of gene expression (38 genes, ≥ 4-fold) compared with the in vivo pattern; and produce different litter sizes (P ≤ 0.0076) after transfer into fosters. Confounding effects of subfertility, life style and genetic heterogeneity are reduced to a minimum in the mouse model compared with ART patients. This is an animal model study. Mouse embryo responses to human ART media are not transferable 1-to-1 to human development due to structural and physiologic differences between oocytes of the two species. Our data promote awareness that human ART culture media affect embryo development. Effects reported here in the mouse may apply also in human, because no ART medium presently available on the market has been optimized for human embryo development. The mouse embryo assay (MEA), which requires ART media to support at least 80% blastocyst formation, is in need of reform and should be extended to include post-implantation development.

Broad AOX expression in a genetically tractable mouse model does not disturb normal physiology

PubMed Central

Szibor, Marten; Dhandapani, Praveen K.; Dufour, Eric; Holmström, Kira M.; Zhuang, Yuan; Salwig, Isabelle; Wittig, Ilka; Heidler, Juliana; Gizatullina, Zemfira; Fuchs, Helmut; Gailus-Durner, Valérie; de Angelis, Martin Hrabě; Nandania, Jatin; Velagapudi, Vidya; Wietelmann, Astrid; Rustin, Pierre; Gellerich, Frank N.; Braun, Thomas

2017-01-01

ABSTRACT Plants and many lower organisms, but not mammals, express alternative oxidases (AOXs) that branch the mitochondrial respiratory chain, transferring electrons directly from ubiquinol to oxygen without proton pumping. Thus, they maintain electron flow under conditions when the classical respiratory chain is impaired, limiting excess production of oxygen radicals and supporting redox and metabolic homeostasis. AOX from Ciona intestinalis has been used to study and mitigate mitochondrial impairments in mammalian cell lines, Drosophila disease models and, most recently, in the mouse, where multiple lentivector-AOX transgenes conferred substantial expression in specific tissues. Here, we describe a genetically tractable mouse model in which Ciona AOX has been targeted to the Rosa26 locus for ubiquitous expression. The AOXRosa26 mouse exhibited only subtle phenotypic effects on respiratory complex formation, oxygen consumption or the global metabolome, and showed an essentially normal physiology. AOX conferred robust resistance to inhibitors of the respiratory chain in organello; moreover, animals exposed to a systemically applied LD50 dose of cyanide did not succumb. The AOXRosa26 mouse is a useful tool to investigate respiratory control mechanisms and to decipher mitochondrial disease aetiology in vivo. PMID:28067626

[Development of the next generation humanized mouse for drug discovery].

PubMed

Ito, Ryoji

A humanized mouse, which is efficiently engrafted human cells and tissues, is an important tool to mimic human physiology for biomedical researches. Since 2000s, severe combined immunodeficient mouse strains such as NOG, BRG, and NSG mice have been generated. They are great recipients to create humanized mouse models compared to previous other immunodeficient strains due to their multiple dysfunctions of innate and acquired immunity. Especially, the transfer of human hematopoietic stem cells into these immunodeficient mice has been enabled to reconstitute human immune systems, because the mice show high engraftment level of human leukocyte in peripheral blood (～50%), spleen and bone marrow (60～90%) and generate well-differentiated multilineage human immune cells including lymphoid and myeloid lineage cells. Using these mice, several human disease models such as cancer, allergy, graft-versus-host disease (GVHD), and etc. have been established to understand the pathogenic mechanisms of the diseases and to evaluate the efficacy and safety of novel drugs. In this review, I provide an overview of recent advances in the humanized mouse technology, including generation of novel platforms of genetically modified NOG (next generation NOG) mice and some applications of them to create human disease models for drug discovery in preclinical researches.

A Mouse Model for Conditional Secretion of Specific Single-Chain Antibodies Provides Genetic Evidence for Regulation of Cortical Plasticity by a Non-cell Autonomous Homeoprotein Transcription Factor.

PubMed

Bernard, Clémence; Vincent, Clémentine; Testa, Damien; Bertini, Eva; Ribot, Jérôme; Di Nardo, Ariel A; Volovitch, Michel; Prochiantz, Alain

2016-05-01

During postnatal life the cerebral cortex passes through critical periods of plasticity allowing its physiological adaptation to the environment. In the visual cortex, critical period onset and closure are influenced by the non-cell autonomous activity of the Otx2 homeoprotein transcription factor, which regulates the maturation of parvalbumin-expressing inhibitory interneurons (PV cells). In adult mice, the maintenance of a non-plastic adult state requires continuous Otx2 import by PV cells. An important source of extra-cortical Otx2 is the choroid plexus, which secretes Otx2 into the cerebrospinal fluid. Otx2 secretion and internalization requires two small peptidic domains that are part of the DNA-binding domain. Thus, mutating these "transfer" sequences also modifies cell autonomous transcription, precluding this approach to obtain a cell autonomous-only mouse. Here, we develop a mouse model with inducible secretion of an anti-Otx2 single-chain antibody to trap Otx2 in the extracellular milieu. Postnatal secretion of this single-chain antibody by PV cells delays PV maturation and reduces plasticity gene expression. Induced adult expression of this single-chain antibody in cerebrospinal fluid decreases Otx2 internalization by PV cells, strongly induces plasticity gene expression and reopens physiological plasticity. We provide the first mammalian genetic evidence for a signaling mechanism involving intercellular transfer of a homeoprotein transcription factor. Our single-chain antibody mouse model is a valid strategy for extracellular neutralization that could be applied to other homeoproteins and signaling molecules within and beyond the nervous system.

Combinations of chromosome transfer and genome editing for the development of cell/animal models of human disease and humanized animal models.

PubMed

Uno, Narumi; Abe, Satoshi; Oshimura, Mitsuo; Kazuki, Yasuhiro

2018-02-01

Chromosome transfer technology, including chromosome modification, enables the introduction of Mb-sized or multiple genes to desired cells or animals. This technology has allowed innovative developments to be made for models of human disease and humanized animals, including Down syndrome model mice and humanized transchromosomic (Tc) immunoglobulin mice. Genome editing techniques are developing rapidly, and permit modifications such as gene knockout and knockin to be performed in various cell lines and animals. This review summarizes chromosome transfer-related technologies and the combined technologies of chromosome transfer and genome editing mainly for the production of cell/animal models of human disease and humanized animal models. Specifically, these include: (1) chromosome modification with genome editing in Chinese hamster ovary cells and mouse A9 cells for efficient transfer to desired cell types; (2) single-nucleotide polymorphism modification in humanized Tc mice with genome editing; and (3) generation of a disease model of Down syndrome-associated hematopoiesis abnormalities by the transfer of human chromosome 21 to normal human embryonic stem cells and the induction of mutation(s) in the endogenous gene(s) with genome editing. These combinations of chromosome transfer and genome editing open up new avenues for drug development and therapy as well as for basic research.

A mouse model with age-dependent immune response and immune-tolerance for HBV infection.

PubMed

Yi, Xuerui; Yuan, Youcheng; Li, Na; Yi, Lu; Wang, Cuiling; Qi, Ying; Gong, Liang; Liu, Guangze; Kong, Xiangping

2018-02-01

Viral clearance of human HBV infection largely depends on the age of exposure. Thus, a mouse model with age-dependent immune response and immune-tolerance for HBV infection was established. HBVRag1 mice were generated by crossing Rag1 -/- mice with HBV-Tg mice. Following adoptive transfer of splenocytes adult (8-9 weeks old) and young (3 weeks old) HBVRag1 mice were named as HBVRag-ReA and HBVRag-ReY mice respectively. The biochemical parameters that were associated with viral load and immune function, as well as the histological evaluation of the liver tissues between the two mouse models were detected. The immune tolerance of HBVRag-ReY mice that were reconstituted at the early stages of life was evaluated by quantitative hepatitis B core antibody assay, adoptive transfer, and modulation of gut microbiota with the addition of antibiotics. HBVRag-ReA mice indicated apparent hepatocytes damage, clearance of HBsAg and production of HBsAb and HBcAb. HBVRag-ReY mice did not develop ALT elevation, and produced HBcAb and HBsAg. A higher number of hepatic CD8 + T and B cells promoted clearance of HBsAg in HBVRag-ReA mice following 30 days of lymphocyte transfer. In contrast to HBVRag-ReA mice, HBVRag-ReY mice exhibited higher levels of Th1/Th2 cytokines. HBVRag-ReY mice exhibited significantly higher (P < .01, approximately 10-fold) serum quantitative anti-HBc levels than HBV-Tg mice, which might be similar to the phase of immune clearance and immune tolerance in human HBV infection. Furthermore, the age-related tolerance in HBVRag-ReY mice that were sensitive to antibiotic treatment was different from that noted in HBV-Tg mice. GS-9620 could inhibit the production of HBsAg, whereas HBV vaccination could induce sustained seroconversion in HBVRag-ReY mice with low levels of HBsAg. The present study described a mouse model with age-dependent immunity and immune-tolerance for HBV infection in vivo, which may mimic chronic HBV infection in humans. Copyright © 2017 Elsevier Ltd. All rights reserved.

Hemagglutination and graft-versus-host disease in the severe combined immunodeficiency mouse lymphoproliferative disease model.

PubMed Central

Pirruccello, S. J.; Nakamine, H.; Beisel, K. W.; Kleveland, K. L.; Okano, M.; Taguchi, Y.; Davis, J. R.; Mahloch, M. L.; Purtilo, D. T.

1992-01-01

In the course of evaluating the severe combined immunodeficiency mouse-human peripheral blood lymphocyte (SCID-PBL) model of lymphoproliferative disease, we noted hemagglutination occurring in peripheral blood smears of mice with serum human immunoglobulin levels greater than 1.0 mg/ml. The hemagglutinating process was mediated by human anti-mouse red cell antibodies of the IgM class, peaked at five to seven weeks post-transfer of 5 to 7 x 10(7) human PBL and was generally self limiting. However, death resulted in some mice when serum immunoglobulin levels were greater than 3.0 mg/ml. The most severely affected mice had hemagglutination induced congestion of liver, lungs and spleen. Several mice also had lesions consistent with graft-versus-host disease (GVHD) including focal hepatic necrosis and destruction of mouse splenic hematopoietic elements. The lesions associated with hemagglutination and GVHD in SCID-PBL mice are distinct from those associated with EBV-induced lymphoproliferation. Recognition of these pathologic processes are required for a thorough understanding of the SCID-PBL model. Images Figure 1 Figure 3 Figure 4 PMID:1580330

Lessons learned using different mouse models during space radiation-induced lung tumorigenesis experiments.

PubMed

Wang, Jian; Zhang, Xiangming; Wang, Ping; Wang, Xiang; Farris, Alton B; Wang, Ya

2016-06-01

Unlike terrestrial ionizing radiation, space radiation, especially galactic cosmic rays (GCR), contains high energy charged (HZE) particles with high linear energy transfer (LET). Due to a lack of epidemiologic data for high-LET radiation exposure, it is highly uncertain how high the carcinogenesis risk is for astronauts following exposure to space radiation during space missions. Therefore, using mouse models is necessary to evaluate the risk of space radiation-induced tumorigenesis; however, which mouse model is better for these studies remains uncertain. Since lung tumorigenesis is the leading cause of cancer death among both men and women, and low-LET radiation exposure increases human lung carcinogenesis, evaluating space radiation-induced lung tumorigenesis is critical to enable safe Mars missions. Here, by comparing lung tumorigenesis obtained from different mouse strains, as well as miR-21 in lung tissue/tumors and serum, we believe that wild type mice with a low spontaneous tumorigenesis background are ideal for evaluating the risk of space radiation-induced lung tumorigenesis, and circulating miR-21 from such mice model might be used as a biomarker for predicting the risk. Copyright © 2016 The Committee on Space Research (COSPAR). Published by Elsevier Ltd. All rights reserved.

Longitudinal assessment of mouse renal injury using high-resolution anatomic and magnetization transfer MR imaging.

PubMed

Wang, Feng; Jiang, Rosie; Takahashi, Keiko; Gore, John; Harris, Raymond C; Takahashi, Takamune; Quarles, C Chad

2014-11-01

The purpose of this study is to evaluate the utility of high-resolution non-invasive endogenous high-field MRI methods for the longitudinal structural and quantitative assessments of mouse kidney disease using the model of unilateral ureter obstruction (UUO). T1-weighted, T2-weighted and magnetization transfer (MT) imaging protocols were optimized to improve the regional contrast in mouse kidney. Conventional T1 and T2 weighted images were collected in UUO mice on day 0 (~3h), day 1, day 3 and day 6 after injury, on a 7 T small animal MRI system. Cortical and medullary thickness, corticomedullary contrast and Magnetization Transfer Ratio (MTR) were assessed longitudinally. Masson trichrome staining was used to histologically assess changes in tissue microstructure. Over the course of UUO progression there were significant (p<0.05) changes in thickness of cortex and outer medulla, and regional changes in T2 signal intensity and MTR values. Histological changes included tubular cell death, tubular dilation, urine retention, and interstitial fibrosis, assessed by histology. The MRI measures of renal cortical and medullary atrophy, cortical-medullary differentiation and MTR changes provide an endogenous, non-invasive and quantitative evaluation of renal morphology and tissue composition during UUO progression. Copyright © 2014 Elsevier Inc. All rights reserved.

Massively Parallel Sequencing Reveals the Complex Structure of an Irradiated Human Chromosome on a Mouse Background in the Tc1 Model of Down Syndrome

PubMed Central

Clayton, Stephen; Prigmore, Elena; Langley, Elizabeth; Yang, Fengtang; Maguire, Sean; Fu, Beiyuan; Rajan, Diana; Sheppard, Olivia; Scott, Carol; Hauser, Heidi; Stephens, Philip J.; Stebbings, Lucy A.; Ng, Bee Ling; Fitzgerald, Tomas; Quail, Michael A.; Banerjee, Ruby; Rothkamm, Kai; Tybulewicz, Victor L. J.; Fisher, Elizabeth M. C.; Carter, Nigel P.

2013-01-01

Down syndrome (DS) is caused by trisomy of chromosome 21 (Hsa21) and presents a complex phenotype that arises from abnormal dosage of genes on this chromosome. However, the individual dosage-sensitive genes underlying each phenotype remain largely unknown. To help dissect genotype – phenotype correlations in this complex syndrome, the first fully transchromosomic mouse model, the Tc1 mouse, which carries a copy of human chromosome 21 was produced in 2005. The Tc1 strain is trisomic for the majority of genes that cause phenotypes associated with DS, and this freely available mouse strain has become used widely to study DS, the effects of gene dosage abnormalities, and the effect on the basic biology of cells when a mouse carries a freely segregating human chromosome. Tc1 mice were created by a process that included irradiation microcell-mediated chromosome transfer of Hsa21 into recipient mouse embryonic stem cells. Here, the combination of next generation sequencing, array-CGH and fluorescence in situ hybridization technologies has enabled us to identify unsuspected rearrangements of Hsa21 in this mouse model; revealing one deletion, six duplications and more than 25 de novo structural rearrangements. Our study is not only essential for informing functional studies of the Tc1 mouse but also (1) presents for the first time a detailed sequence analysis of the effects of gamma radiation on an entire human chromosome, which gives some mechanistic insight into the effects of radiation damage on DNA, and (2) overcomes specific technical difficulties of assaying a human chromosome on a mouse background where highly conserved sequences may confound the analysis. Sequence data generated in this study is deposited in the ENA database, Study Accession number: ERP000439. PMID:23596509

What We Have Learned from Animal Models of Dry Eye

PubMed Central

Stern, Michael E.; Pflugfelder, Stephen C.

2017-01-01

Animal models have proved valuable to investigate the pathogenesis of dry eye disease, identify therapeutic targets and the efficacy of candidate therapeutics for dry eye. Pharmacological inhibition of the lacrimal functional unit and exposure of the mouse eye to desiccating stress was found to activate innate immune pathways, promote dendritic cell maturation and initiate an adaptive T cell response to ocular surface antigens. Disease relevant mediators and pathways have been identified through use of genetically altered mice, specific inhibitors and adoptive transfer of desiccating stress primed CD4+ T cells to naïve recipients. Findings from mouse models have elucidated the mechanism of action of cyclosporine A and the rationale for developing lifitegrast, the two currently approved therapeutics in the US. PMID:28282318

Production of cloned mice by somatic cell nuclear transfer.

PubMed

Kishigami, Satoshi; Wakayama, Sayaka; Thuan, Nguyen Van; Ohta, Hiroshi; Mizutani, Eiji; Hikichi, Takafusa; Bui, Hong-Thuy; Balbach, Sebastian; Ogura, Atsuo; Boiani, Michele; Wakayama, Teruhiko

2006-01-01

Although it has now been 10 years since the first cloned mammals were generated from somatic cells using nuclear transfer (NT), the success rate for producing live offspring by cloning remains < 5%. Nevertheless, the techniques have potential as important tools for future research in basic biology. We have been able to develop a stable NT method in the mouse, in which donor nuclei are directly injected into the oocyte using a piezo-actuated micromanipulator. Although manipulation of the piezo unit is complex, once mastered it is of great help not only in NT experiments but also in almost all other forms of micromanipulation. In addition to this technique, embryonic stem (ES) cell lines established from somatic cell nuclei by NT can be generated relatively easily from a variety of mouse genotypes and cell types. Such NT-ES cells can be used not only for experimental models of human therapeutic cloning but also as a backup of the donor cell's genome. Our most recent protocols for mouse cloning, as described here, will allow the production of cloned mice in > or = 3 months.

Gene Transfer of Brain-derived Neurotrophic Factor (BDNF) Prevents Neurodegeneration Triggered by FXN Deficiency.

PubMed

Katsu-Jiménez, Yurika; Loría, Frida; Corona, Juan Carlos; Díaz-Nido, Javier

2016-05-01

Friedreich's ataxia is a predominantly neurodegenerative disease caused by recessive mutations that produce a deficiency of frataxin (FXN). Here, we have used a herpesviral amplicon vector carrying a gene encoding for brain-derived neurotrophic factor (BDNF) to drive its overexpression in neuronal cells and test for its effect on FXN-deficient neurons both in culture and in the mouse cerebellum in vivo. Gene transfer of BDNF to primary cultures of mouse neurons prevents the apoptosis which is triggered by the knockdown of FXN gene expression. This neuroprotective effect of BDNF is also observed in vivo in a viral vector-based knockdown mouse cerebellar model. The injection of a lentiviral vector carrying a minigene encoding for a FXN-specific short hairpin ribonucleic acid (shRNA) into the mouse cerebellar cortex triggers a FXN deficit which is accompanied by significant apoptosis of granule neurons as well as loss of calbindin in Purkinje cells. These pathological changes are accompanied by a loss of motor coordination of mice as assayed by the rota-rod test. Coinjection of a herpesviral vector encoding for BDNF efficiently prevents both the development of cerebellar neuropathology and the ataxic phenotype. These data demonstrate the potential therapeutic usefulness of neurotrophins like BDNF to protect FXN-deficient neurons from degeneration.

Adeno-associated virus–targeted disruption of the CFTR gene in cloned ferrets

PubMed Central

Sun, Xingshen; Yan, Ziying; Yi, Yaling; Li, Ziyi; Lei, Diana; Rogers, Christopher S.; Chen, Juan; Zhang, Yulong; Welsh, Michael J.; Leno, Gregory H.; Engelhardt, John F.

2008-01-01

Somatic cell gene targeting combined with nuclear transfer cloning presents tremendous potential for the creation of new, large-animal models of human diseases. Mouse disease models often fail to reproduce human phenotypes, underscoring the need for the generation and study of alternative disease models. Mice deficient for CFTR have been poor models for cystic fibrosis (CF), lacking many aspects of human CF lung disease. In this study, we describe the production of a CFTR gene–deficient model in the domestic ferret using recombinant adeno-associated virus–mediated gene targeting in fibroblasts, followed by nuclear transfer cloning. As part of this approach, we developed a somatic cell rejuvenation protocol using serial nuclear transfer to produce live CFTR-deficient clones from senescent gene-targeted fibroblasts. We transferred 472 reconstructed embryos into 11 recipient jills and obtained 8 healthy male ferret clones heterozygous for a disruption in exon 10 of the CFTR gene. To our knowledge, this study represents the first description of genetically engineered ferrets and describes an approach that may be of substantial utility in modeling not only CF, but also other genetic diseases. PMID:18324338

Mouse Models for Drug Discovery. Can New Tools and Technology Improve Translational Power?

PubMed Central

Zuberi, Aamir; Lutz, Cathleen

2016-01-01

Abstract The use of mouse models in biomedical research and preclinical drug evaluation is on the rise. The advent of new molecular genome-altering technologies such as CRISPR/Cas9 allows for genetic mutations to be introduced into the germ line of a mouse faster and less expensively than previous methods. In addition, the rapid progress in the development and use of somatic transgenesis using viral vectors, as well as manipulations of gene expression with siRNAs and antisense oligonucleotides, allow for even greater exploration into genomics and systems biology. These technological advances come at a time when cost reductions in genome sequencing have led to the identification of pathogenic mutations in patient populations, providing unprecedented opportunities in the use of mice to model human disease. The ease of genetic engineering in mice also offers a potential paradigm shift in resource sharing and the speed by which models are made available in the public domain. Predictively, the knowledge alone that a model can be quickly remade will provide relief to resources encumbered by licensing and Material Transfer Agreements. For decades, mouse strains have provided an exquisite experimental tool to study the pathophysiology of the disease and assess therapeutic options in a genetically defined system. However, a major limitation of the mouse has been the limited genetic diversity associated with common laboratory mice. This has been overcome with the recent development of the Collaborative Cross and Diversity Outbred mice. These strains provide new tools capable of replicating genetic diversity to that approaching the diversity found in human populations. The Collaborative Cross and Diversity Outbred strains thus provide a means to observe and characterize toxicity or efficacy of new therapeutic drugs for a given population. The combination of traditional and contemporary mouse genome editing tools, along with the addition of genetic diversity in new modeling systems, are synergistic and serve to make the mouse a better model for biomedical research, enhancing the potential for preclinical drug discovery and personalized medicine. PMID:28053071

Studying Neutrophil Migration In Vivo Using Adoptive Cell Transfer.

PubMed

Miyabe, Yoshishige; Kim, Nancy D; Miyabe, Chie; Luster, Andrew D

2016-01-01

Adoptive cell transfer experiments can be used to study the roles of cell trafficking molecules on the migratory behavior of specific immune cell populations in vivo. Chemoattractants and their G protein-coupled seven-transmembrane-spanning receptors regulate migration of cells in vivo, and dysregulated expression of chemoattractants and their receptors is implicated in autoimmune and inflammatory diseases. Inflammatory arthritides, such as rheumatoid arthritis (RA), are characterized by the recruitment of inflammatory cells into joints. The K/BxN serum transfer mouse model of inflammatory arthritis shares many similar features with RA. In this autoantibody-induced model of arthritis, neutrophils are the critical immune cells necessary for the development of joint inflammation and damage. We have used adoptive neutrophil transfer to define the contributions of chemoattractant receptors, cytokines, and activation receptors expressed on neutrophils that critically regulate their entry into the inflamed joint. In this review, we describe the procedure of neutrophil adoptive transfer to study the influence of neutrophil-specific receptors or mediators upon the their recruitment into the joint using the K/BxN model of inflammatory arthritis as a model of how adoptive cell transfer studies can be used to study immune cell migration in vivo.

Transplantation of wild-type mouse hematopoietic stem and progenitor cells ameliorates deficits in a mouse model of Friedreich’s ataxia

PubMed Central

Rocca, Celine J.; Goodman, Spencer M.; Dulin, Jennifer N.; Haquang, Joseph H.; Gertsman, Ilya; Blondelle, Jordan; Smith, Janell L. M.; Heyser, Charles J.; Cherqui, Stephanie

2017-01-01

Friedreich’s ataxia (FRDA) is an incurable autosomal recessive neurodegenerative disease caused by reduced expression of the mitochondrial protein frataxin due to an intronic GAA-repeat expansion in the FXN gene. We report the therapeutic efficacy of transplanting wild-type mouse hematopoietic stem and progenitor cells (HSPCs) into the YG8R mouse model of FRDA. In the HSPC-transplanted YG8R mice, development of muscle weakness and locomotor deficits was abrogated as was degeneration of large sensory neurons in the dorsal root ganglia (DRGs) and mitochondrial capacity was improved in brain, skeletal muscle, and heart. Transplanted HSPCs engrafted and then differentiated into microglia in the brain and spinal cord and into macrophages in the DRGs, heart, and muscle of YG8R FRDA mice. We observed the transfer of wild-type frataxin and Cox8 mitochondrial proteins from HSPC-derived microglia/macrophages to FRDA mouse neurons and muscle myocytes in vivo. Our results show the HSPC-mediated phenotypic rescue of FRDA in YG8R mice and suggest that this approach should be investigated further as a strategy for treating FRDA. PMID:29070698

An inducible model of abacterial prostatitis induces antigen specific inflammatory and proliferative changes in the murine prostate

PubMed Central

Haverkamp, Jessica M.; Charbonneau, Bridget; Meyerholz, David K.; Cohen, Michael B.; Snyder, Paul W.; Svensson, Robert U.; Henry, Michael D.; Wang, Hsing- Hui

2011-01-01

Background Prostatitis is a poorly understood disease and increasing evidence suggests inflammation is involved in other prostatic diseases, including prostate cancer. Methods The ability of pre-activated CD8 T cells to induce prostatitis was examined by adoptive transfer into POET-3 mice or POET-3/Luc/Pten−/+ mice. Characterization of the inflammatory response was determined by examining leukocyte infiltration by histological analysis, flow cytometry and by evaluating cytokine and chemokine levels in prostate tissue. The impact of inflammation on the prostate was evaluated by monitoring epithelial cell proliferation over time. Results Initiation of inflammation by ovalbumin specific CD8+ T cells (OT-I cells) resulted in development of acute prostatitis in the anterior, dorsolateral and anterior prostate of POET-3 and POET-3/Luc/Pten−/+ mice. Acute prostatitis was characterized by recruitment of adoptively transferred OT-I cells and importantly, autologous CD4+ and CD8+ T cells, myeloid-derived suppressor cells (MDSC) and regulatory T cells (Treg). In concert with leukocyte infiltration elevated levels of pro-inflammatory cytokines and chemokines were observed. Inflammation also resulted in marked epithelial cell proliferation that was sustained up to 80 days post adoptive-transfer of OT-I cells. Conclusions The POET-3 model represents a novel mouse model to study both acute and chronic prostate inflammation in an antigen-specific system. Further, the POET-3 mouse model can be crossed with other genetic models of disease such as the C57/Luc/Pten−/− model of prostate cancer, allowing the impact of prostatitis on other prostatic diseases to be evaluated. PMID:21656824

Protection against Chlamydia psittaci in mice conferred by Lyt-2+ T cells.

PubMed Central

Buzoni-Gatel, D; Guilloteau, L; Bernard, F; Bernard, S; Chardès, T; Rocca, A

1992-01-01

A murine model was used to study the respective roles of L3T4+ and Lyt-2+ T cells in protection against Chlamydia psittaci. Donor mice were intravenously (i.v.) infected with 1 x 10(5) plaque-forming units (PFU) per mice of live C. psittaci. One month after inoculation, splenic cells from donors were transferred into syngenic recipients (5 x 10(7) cells/mouse). As measured by splenic colonization on Day 6 after i.v. challenge (1 x 10(5) PFU/mouse), transfer with primed (untreated) cells conferred a 3 log protection in this model. In vitro treatment, before transfer, of splenic cells with anti-Lyt-2 monoclonal antibody (mAb) and complement, markedly impaired the protection in comparison with control mice transferred with primed untreated cells, whereas treatment with anti-L3T4 mAb did not reduce the transferred protection. Resistance to a reinfection with C. psittaci was also studied after selective in vivo depletion of L3T4+ and Lyt-2+ T cells. One month after primary infection, mice were treated with anti-L3T4 or anti-Lyt-2 mAb and challenged thereafter (i.v., 1 x 10(5) PFU). The splenic colonization on Day 6 after challenge demonstrated that treatment with anti-Lyt-2 mAb impaired resistance against a subsequent infection with C. psittaci. Treatment with anti-L3T4 mAb in vivo had no effect on protection, as previously described in vitro. The mechanisms by which Lyt-2+ T cells could participate in the elimination of bacteria were discussed. PMID:1427980

Expression of endogenous mouse APP modulates β-amyloid deposition in hAPP-transgenic mice.

PubMed

Steffen, Johannes; Krohn, Markus; Schwitlick, Christina; Brüning, Thomas; Paarmann, Kristin; Pietrzik, Claus U; Biverstål, Henrik; Jansone, Baiba; Langer, Oliver; Pahnke, Jens

2017-06-20

Amyloid-β (Aβ) deposition is one of the hallmarks of the amyloid hypothesis in Alzheimer's disease (AD). Mouse models using APP-transgene overexpression to generate amyloid plaques have shown to model only certain parts of the disease. The extent to which the data from mice can be transferred to man remains controversial. Several studies have shown convincing treatment results in reducing Aβ and enhancing cognition in mice but failed totally in human. One model-dependent factor has so far been almost completely neglected: the endogenous expression of mouse APP and its effects on the transgenic models and the readout for therapeutic approaches.Here, we report that hAPP-transgenic models of amyloidosis devoid of endogenous mouse APP expression (mAPP-knockout / mAPPko) show increased amounts and higher speed of Aβ deposition than controls with mAPP. The number of senile plaques and the level of aggregated hAβ were elevated in mAPPko mice, while the deposition in cortical blood vessels was delayed, indicating an alteration in the general aggregation propensity of hAβ together with endogenous mAβ. Furthermore, the cellular response to Aβ deposition was modulated: mAPPko mice developed a pronounced and age-dependent astrogliosis, while microglial association to amyloid plaques was diminished. The expression of human and murine aggregation-prone proteins with differing amino acid sequences within the same mouse model might not only alter the extent of deposition but also modulate the route of pathogenesis, and thus, decisively influence the study outcome, especially in translational research.

Importing, caring, breeding, genotyping, and phenotyping a genetic mouse in a Chinese university.

PubMed

Kuo, S T; Wu, Q H; Liu, B; Xie, Z L; Wu, X; Shang, S J; Zhang, X Y; Kang, X J; Liu, L N; Zhu, F P; Wang, Y S; Hu, M Q; Xu, H D; Zhou, L; Liu, B; Chai, Z Y; Zhang, Q F; Liu, W; Teng, S S; Wang, C H; Guo, N; Dou, H Q; Zuo, P L; Zheng, L H; Zhang, C X; Zhu, D S; Wang, L; Wang, S R; Zhou, Z

2014-07-01

The genetic manipulation of the laboratory mouse has been well developed and generated more and more mouse lines for biomedical research. To advance our science exploration, it is necessary to share genetically modified mouse lines with collaborators between institutions, even in different countries. The transfer process is complicated. Significant paperwork and coordination are required, concerning animal welfare, intellectual property rights, colony health status, and biohazard. Here, we provide a practical example of importing a transgenic mice line, Dynamin 1 knockout mice, from Yale University in the USA to Perking University in China for studying cell secretion. This example including the length of time that required for paper work, mice quarantine at the receiving institution, and expansion of the mouse line for experiments. The procedure described in this paper for delivery live transgenic mice from USA to China may serve a simple reference for transferring mouse lines between other countries too.

Molecular and cellular events during blastocyst implantation in the receptive uterus: clues from mouse models

PubMed Central

MATSUMOTO, Hiromichi

2017-01-01

The success of implantation is an interactive process between the blastocyst and the uterus. Synchronized development of embryos with uterine differentiation to a receptive state is necessary to complete pregnancy. The period of uterine receptivity for implantation is limited and referred to as the “implantation window”, which is regulated by ovarian steroid hormones. Implantation process is complicated due to the many signaling molecules in the hierarchical mechanisms with the embryo-uterine dialogue. The mouse is widely used in animal research, and is uniquely suited for reproductive studies, i.e., having a large litter size and brief estrous cycles. This review first describes why the mouse is the preferred model for implantation studies, focusing on uterine morphology and physiological traits, and then highlights the knowledge on uterine receptivity and the hormonal regulation of blastocyst implantation in mice. Our recent study revealed that selective proteolysis in the activated blastocyst is associated with the completion of blastocyst implantation after embryo transfer. Furthermore, in the context of blastocyst implantation in the mouse, this review discusses the window of uterine receptivity, hormonal regulation, uterine vascular permeability and angiogenesis, the delayed-implantation mouse model, morphogens, adhesion molecules, crosslinker proteins, extracellular matrix, and matricellular proteins. A better understanding of uterine and blastocyst biology during the peri-implantation period should facilitate further development of reproductive technology. PMID:28638003

A mouse model of mitochondrial complex III dysfunction induced by myxothiazol

DOE Office of Scientific and Technical Information (OSTI.GOV)

Davoudi, Mina; Kallijärvi, Jukka; Marjavaara, Sanna

2014-04-18

Highlights: • Reversible chemical inhibition of complex III in wild type mouse. • Myxothiazol causes decreased complex III activity in mouse liver. • The model is useful for therapeutic trials to improve mitochondrial function. - Abstract: Myxothiazol is a respiratory chain complex III (CIII) inhibitor that binds to the ubiquinol oxidation site Qo of CIII. It blocks electron transfer from ubiquinol to cytochrome b and thus inhibits CIII activity. It has been utilized as a tool in studies of respiratory chain function in in vitro and cell culture models. We developed a mouse model of biochemically induced and reversible CIIImore » inhibition using myxothiazol. We administered myxothiazol intraperitoneally at a dose of 0.56 mg/kg to C57Bl/J6 mice every 24 h and assessed CIII activity, histology, lipid content, supercomplex formation, and gene expression in the livers of the mice. A reversible CIII activity decrease to 50% of control value occurred at 2 h post-injection. At 74 h only minor histological changes in the liver were found, supercomplex formation was preserved and no significant changes in the expression of genes indicating hepatotoxicity or inflammation were found. Thus, myxothiazol-induced CIII inhibition can be induced in mice for four days in a row without overt hepatotoxicity or lethality. This model could be utilized in further studies of respiratory chain function and pharmacological approaches to mitochondrial hepatopathies.« less

A partial structural and functional rescue of a retinitis pigmentosa model with compacted DNA nanoparticles.

PubMed

Cai, Xue; Nash, Zack; Conley, Shannon M; Fliesler, Steven J; Cooper, Mark J; Naash, Muna I

2009-01-01

Previously we have shown that compacted DNA nanoparticles can drive high levels of transgene expression after subretinal injection in the mouse eye. Here we delivered compacted DNA nanoparticles containing a therapeutic gene to the retinas of a mouse model of retinitis pigmentosa. Nanoparticles containing the wild-type retinal degeneration slow (Rds) gene were injected into the subretinal space of rds(+/-) mice on postnatal day 5. Gene expression was sustained for up to four months at levels up to four times higher than in controls injected with saline or naked DNA. The nanoparticles were taken up into virtually all photoreceptors and mediated significant structural and biochemical rescue of the disease without histological or functional evidence of toxicity. Electroretinogram recordings showed that nanoparticle-mediated gene transfer restored cone function to a near-normal level in contrast to transfer of naked plasmid DNA. Rod function was also improved. These findings demonstrate that compacted DNA nanoparticles represent a viable option for development of gene-based interventions for ocular diseases and obviate major barriers commonly encountered with non-viral based therapies.

Effects of embryo culture media do not persist after implantation: a histological study in mice.

PubMed

Hemkemeyer, Sandra A; Schwarzer, Caroline; Boiani, Michele; Ehmcke, Jens; Le Gac, Séverine; Schlatt, Stefan; Nordhoff, Verena

2014-02-01

Is post-implantation embryonic development after blastocyst transfer affected by exposure to different assisted reproduction technology (ART) culture media? Fetal development and placental histology of ART embryos cultured in vitro in different ART media was not impaired compared with embryos grown in vivo. The application of different in vitro culture (IVC) media for human ART has an effect on birthweight of newborns. In the mouse model, differences in blastocyst formation were reported after culture in different ART media. Moreover, abnormalities in the liver and heart have been detected as a result of suboptimal IVC conditions. Fertilized oocytes from inbred and outbred breeding schemes were retrieved and either immediately transferred to foster mothers or incubated in control or human ART culture media up to the blastocyst stage prior to transfer. Placental and fetal anatomy and particularly bone development were evaluated. B6C3F1 female mice were used as oocyte donors after ovulation induction. C57Bl/6 and CD1 males were used for mating and CD1 females as foster mothers for embryo transfer. Fertilized oocytes were recovered from mated females and incubated in sequential human ART media (ISM1/ISM2 and HTF/Multiblast), in control media [KSOM(aa) and Whitten's medium] or grown in utero without IVC (zygote control). As in vivo, control B6C3F1 females were superovulated and left untreated. Fetuses and placentae were isolated by Caesarean section and analysed at 18.5 days post-coitum (dpc) for placenta composition and at 15.5 dpc for body weight, crown-rump length (CRL), fetal organ development, morphological development, total bone length and extent of bone ossification. No major differences in the number of implantation sites or in histological appearance of the placentae were detected. CRL of KSOM(aa) fetuses was higher compared with zygote control and Whitten's medium. Histological analysis of tissue sections revealed no gross morphological differences compared with the in vitro groups or in vivo controls. Furthermore, no changes in skeletal development and degree of ossification were observed. However, fibula and tibia of ISM1/ISM2 fetuses were longer than the respective ones from in vivo fetuses. Findings in the mouse embryo and fetus may not be fully transferable to humans. In addition to skeletal development and placentation, there may be other parameters, e.g. on the molecular level which respond to IVC in ART media. Some comparisons have limited statistical power. Our data suggest that once implantation is achieved, subsequent post-implantation development unfolds normally, resulting in healthy fetuses. With mouse models, we gather information for the safety of human ART culture media. Our mouse study is reassuring for the safety of ART conditions on human embryonic development, given the lack of bold detrimental effects observed in the mouse model. This work was supported by the Deutsche Forschungsgemeinschaft (BO 2540/4-1 and SCHL 394/9-1) and by the Nederlandse Organisatie voor Wetenschappelijk Onderzoek (S.L.G.); Bilateral grant NWO-DFG 63-258. None of the authors has any conflict of interest to declare. Not applicable.

Cellular Specificity of the Blood–CSF Barrier for Albumin Transfer across the Choroid Plexus Epithelium

PubMed Central

Liddelow, Shane A.; Dzięgielewska, Katarzyna M.; Møllgård, Kjeld; Whish, Sophie C.; Noor, Natassya M.; Wheaton, Benjamin J.; Gehwolf, Renate; Wagner, Andrea; Traweger, Andreas; Bauer, Hannelore; Bauer, Hans-Christian; Saunders, Norman R.

2014-01-01

To maintain the precise internal milieu of the mammalian central nervous system, well-controlled transfer of molecules from periphery into brain is required. Recently the soluble and cell-surface albumin-binding glycoprotein SPARC (secreted protein acidic and rich in cysteine) has been implicated in albumin transport into developing brain, however the exact mechanism remains unknown. We postulate that SPARC is a docking site for albumin, mediating its uptake and transfer by choroid plexus epithelial cells from blood into cerebrospinal fluid (CSF). We used in vivo physiological measurements of transfer of endogenous (mouse) and exogenous (human) albumins, in situ Proximity Ligation Assay (in situ PLA), and qRT-PCR experiments to examine the cellular mechanism mediating protein transfer across the blood–CSF interface. We report that at all developmental stages mouse albumin and SPARC gave positive signals with in situ PLAs in plasma, CSF and within individual plexus cells suggesting a possible molecular interaction. In contrast, in situ PLA experiments in brain sections from mice injected with human albumin showed positive signals for human albumin in the vascular compartment that were only rarely identifiable within choroid plexus cells and only at older ages. Concentrations of both endogenous mouse albumin and exogenous (intraperitoneally injected) human albumin were estimated in plasma and CSF and expressed as CSF/plasma concentration ratios. Human albumin was not transferred through the mouse blood–CSF barrier to the same extent as endogenous mouse albumin, confirming results from in situ PLA. During postnatal development Sparc gene expression was higher in early postnatal ages than in the adult and changed in response to altered levels of albumin in blood plasma in a differential and developmentally regulated manner. Here we propose a possible cellular route and mechanism by which albumin is transferred from blood into CSF across a sub-population of specialised choroid plexus epithelial cells. PMID:25211495

Clusterin Facilitates Exchange of Glycosyl Phosphatidylinositol-Linked SPAM1 Between Reproductive Luminal Fluids and Mouse and Human Sperm Membranes1

PubMed Central

Griffiths, Genevieve S.; Galileo, Deni S.; Aravindan, Rolands G.; Martin-DeLeon, Patricia A.

2009-01-01

Glycosyl phosphatidylinositol (GPI)-linked proteins, which are involved in post-testicular maturation of sperm and have a role in fertilization, are acquired on the sperm surface from both vesicular and membrane-free soluble fractions of epididymal luminal fluid (LF) and uterine LF. Herein, we investigate the mechanism of uptake of these proteins from the soluble fraction of LFs using sperm adhesion molecule 1 (SPAM1) as a model. Ultracentrifugation and native Western blot analysis of the soluble fraction revealed that SPAM1 is present in low-molecular-weight (monomeric) and high-molecular-weight (oligomeric) complexes. The latter are incapable of transferring SPAM1 and may serve to produce monomers. Monomers are stabilized by hydrophobic interactions with clusterin (CLU), a lipid carrier that is abundantly expressed in LFs. We show that CLU is involved in the transfer of SPAM1 monomers, whose delivery was decreased by anti-CLU antibody under normal and apolipoprotein-enhanced conditions. Coimmunoprecipitation revealed an intimate association of CLU with SPAM1. Both plasma and recombinant CLU had a dose-related effect on transfer efficiency: high concentrations reduced and low concentrations enhanced delivery of SPAM1 to human and mouse sperm membranes, reflecting physiological states in the epididymal tract. We propose a lipid exchange model (akin to the lipid-poor model for cholesterol efflux) for the delivery of GPI-linked proteins to sperm membranes via CLU. Our investigation defines specific conditions for membrane-free GPI-linked protein transfer in vitro and could lead to technology for improving fertility or treating sperm pathology by the addition of relevant GPI-linked proteins critical for successful fertilization in humans and domestic animals. PMID:19357365

Effect of tumor properties on energy absorption, temperature mapping, and thermal dose in 13.56-MHz radiofrequency hyperthermia.

PubMed

Prasad, Bibin; Kim, Subin; Cho, Woong; Kim, Suzy; Kim, Jung Kyung

2018-05-01

Computational techniques can enhance personalized hyperthermia-treatment planning by calculating tissue energy absorption and temperature distribution. This study determined the effect of tumor properties on energy absorption, temperature mapping, and thermal dose distribution in mild radiofrequency hyperthermia using a mouse xenograft model. We used a capacitive-heating radiofrequency hyperthermia system with an operating frequency of 13.56 MHz for in vivo mouse experiments and performed simulations on a computed tomography mouse model. Additionally, we measured the dielectric properties of the tumors and considered temperature dependence for thermal properties, metabolic heat generation, and perfusion. Our results showed that dielectric property variations were more dominant than thermal properties and other parameters, and that the measured dielectric properties provided improved temperature-mapping results relative to the property values taken from previous study. Furthermore, consideration of temperature dependency in the bio heat-transfer model allowed elucidation of precise thermal-dose calculations. These results suggested that this method might contribute to effective thermoradiotherapy planning in clinics. Copyright © 2018 Elsevier Ltd. All rights reserved.

Reperfusion injury intensifies the adaptive human T cell alloresponse in a human-mouse chimeric artery model.

PubMed

Yi, Tai; Fogal, Birgit; Hao, Zhengrong; Tobiasova, Zuzana; Wang, Chen; Rao, Deepak A; Al-Lamki, Rafia S; Kirkiles-Smith, Nancy C; Kulkarni, Sanjay; Bradley, John R; Bothwell, Alfred L M; Sessa, William C; Tellides, George; Pober, Jordan S

2012-02-01

Perioperative nonimmune injuries to an allograft can decrease graft survival. We have developed a model for studying this process using human materials. Human artery segments were transplanted as infrarenal aortic interposition grafts into an immunodeficient mouse host, allowed to "heal in" for 30 days, and then retransplanted into a second mouse host. To induce a reperfusion injury, the healed-in artery segments were incubated for 3 hours under hypoxic conditions ex vivo before retransplantation. To induce immunologic rejection, the animals receiving the retransplanted artery segment were adoptively transferred with human peripheral blood mononuclear cells or purified T cells from a donor allogeneic to the artery 1 week before surgery. To compare rejection of injured versus healthy tissues, these manipulations were combined. Results were analyzed ex vivo by histology, morphometry, immunohistochemistry, and mRNA quantitation or in vivo by ultrasound. Our results showed that reperfusion injury, which otherwise heals with minimal sequelae, intensifies the degree of allogeneic T cell-mediated injury to human artery segments. We developed a new human-mouse chimeric model demonstrating interactions of reperfusion injury and alloimmunity using human cells and tissues that may be adapted to study other forms of nonimmune injury and other types of adaptive immune responses.

Mouse Models for Drug Discovery. Can New Tools and Technology Improve Translational Power?

PubMed

Zuberi, Aamir; Lutz, Cathleen

2016-12-01

The use of mouse models in biomedical research and preclinical drug evaluation is on the rise. The advent of new molecular genome-altering technologies such as CRISPR/Cas9 allows for genetic mutations to be introduced into the germ line of a mouse faster and less expensively than previous methods. In addition, the rapid progress in the development and use of somatic transgenesis using viral vectors, as well as manipulations of gene expression with siRNAs and antisense oligonucleotides, allow for even greater exploration into genomics and systems biology. These technological advances come at a time when cost reductions in genome sequencing have led to the identification of pathogenic mutations in patient populations, providing unprecedented opportunities in the use of mice to model human disease. The ease of genetic engineering in mice also offers a potential paradigm shift in resource sharing and the speed by which models are made available in the public domain. Predictively, the knowledge alone that a model can be quickly remade will provide relief to resources encumbered by licensing and Material Transfer Agreements. For decades, mouse strains have provided an exquisite experimental tool to study the pathophysiology of the disease and assess therapeutic options in a genetically defined system. However, a major limitation of the mouse has been the limited genetic diversity associated with common laboratory mice. This has been overcome with the recent development of the Collaborative Cross and Diversity Outbred mice. These strains provide new tools capable of replicating genetic diversity to that approaching the diversity found in human populations. The Collaborative Cross and Diversity Outbred strains thus provide a means to observe and characterize toxicity or efficacy of new therapeutic drugs for a given population. The combination of traditional and contemporary mouse genome editing tools, along with the addition of genetic diversity in new modeling systems, are synergistic and serve to make the mouse a better model for biomedical research, enhancing the potential for preclinical drug discovery and personalized medicine. © The Author 2016. Published by Oxford University Press.

T cell transfer model of chronic colitis: concepts, considerations, and tricks of the trade.

PubMed

Ostanin, Dmitry V; Bao, Jianxiong; Koboziev, Iurii; Gray, Laura; Robinson-Jackson, Sherry A; Kosloski-Davidson, Melissa; Price, V Hugh; Grisham, Matthew B

2009-02-01

The inflammatory bowel diseases (Crohn's disease; ulcerative colitis) are idiopathic chronic inflammatory disorders of the intestine and/or colon. A major advancement in our understanding of the pathogenesis of these diseases has been the development of mouse models of chronic gut inflammation. One model that has been instrumental in delineating the immunological mechanisms responsible for the induction as well as regulation of intestinal inflammation is the T cell transfer model of chronic colitis. This paper presents a detailed protocol describing the methods used to induce chronic colitis in mice. Special attention is given to the immunological concepts that explain disease pathogenesis in this model, considerations and potential pitfalls in using this model, and finally different "tricks" that we have learned over the past 12 years that have allowed us to develop a more simplified version of this model of experimental IBD.

Legal Agreements and the Governance of Research Commons: Lessons from Materials Sharing in Mouse Genomics

PubMed Central

Mishra, Amrita

2014-01-01

Abstract Omics research infrastructure such as databases and bio-repositories requires effective governance to support pre-competitive research. Governance includes the use of legal agreements, such as Material Transfer Agreements (MTAs). We analyze the use of such agreements in the mouse research commons, including by two large-scale resource development projects: the International Knockout Mouse Consortium (IKMC) and International Mouse Phenotyping Consortium (IMPC). We combine an analysis of legal agreements and semi-structured interviews with 87 members of the mouse model research community to examine legal agreements in four contexts: (1) between researchers; (2) deposit into repositories; (3) distribution by repositories; and (4) exchanges between repositories, especially those that are consortium members of the IKMC and IMPC. We conclude that legal agreements for the deposit and distribution of research reagents should be kept as simple and standard as possible, especially when minimal enforcement capacity and resources exist. Simple and standardized legal agreements reduce transactional bottlenecks and facilitate the creation of a vibrant and sustainable research commons, supported by repositories and databases. PMID:24552652

NADPH–Cytochrome P450 Oxidoreductase: Roles in Physiology, Pharmacology, and Toxicology

PubMed Central

Ding, Xinxin; Wolf, C. Roland; Porter, Todd D.; Pandey, Amit V.; Zhang, Qing-Yu; Gu, Jun; Finn, Robert D.; Ronseaux, Sebastien; McLaughlin, Lesley A.; Henderson, Colin J.; Zou, Ling; Flück, Christa E.

2013-01-01

This is a report on a symposium sponsored by the American Society for Pharmacology and Experimental Therapeutics and held at the Experimental Biology 2012 meeting in San Diego, California, on April 25, 2012. The symposium speakers summarized and critically evaluated our current understanding of the physiologic, pharmacological, and toxicological roles of NADPH–cytochrome P450 oxidoreductase (POR), a flavoprotein involved in electron transfer to microsomal cytochromes P450 (P450), cytochrome b5, squalene mono-oxygenase, and heme oxygenase. Considerable insight has been derived from the development and characterization of mouse models with conditional Por deletion in particular tissues or partial suppression of POR expression in all tissues. Additional mouse models with global or conditional hepatic deletion of cytochrome b5 are helping to clarify the P450 isoform- and substrate-specific influences of cytochrome b5 on P450 electron transfer and catalytic function. This symposium also considered studies using siRNA to suppress POR expression in a hepatoma cell–culture model to explore the basis of the hepatic lipidosis phenotype observed in mice with conditional deletion of Por in liver. The symposium concluded with a strong translational perspective, relating the basic science of human POR structure and function to the impacts of POR genetic variation on human drug and steroid metabolism. PMID:23086197

Evaluation of biolistic gene transfer methods in vivo using non-invasive bioluminescent imaging techniques.

PubMed

Xia, Jixiang; Martinez, Angela; Daniell, Henry; Ebert, Steven N

2011-06-02

Gene therapy continues to hold great potential for treating many different types of disease and dysfunction. Safe and efficient techniques for gene transfer and expression in vivo are needed to enable gene therapeutic strategies to be effective in patients. Currently, the most commonly used methods employ replication-defective viral vectors for gene transfer, while physical gene transfer methods such as biolistic-mediated ("gene-gun") delivery to target tissues have not been as extensively explored. In the present study, we evaluated the efficacy of biolistic gene transfer techniques in vivo using non-invasive bioluminescent imaging (BLI) methods. Plasmid DNA carrying the firefly luciferase (LUC) reporter gene under the control of the human Cytomegalovirus (CMV) promoter/enhancer was transfected into mouse skin and liver using biolistic methods. The plasmids were coupled to gold microspheres (1 μm diameter) using different DNA Loading Ratios (DLRs), and "shot" into target tissues using a helium-driven gene gun. The optimal DLR was found to be in the range of 4-10. Bioluminescence was measured using an In Vivo Imaging System (IVIS-50) at various time-points following transfer. Biolistic gene transfer to mouse skin produced peak reporter gene expression one day after transfer. Expression remained detectable through four days, but declined to undetectable levels by six days following gene transfer. Maximum depth of tissue penetration following biolistic transfer to abdominal skin was 200-300 μm. Similarly, biolistic gene transfer to mouse liver in vivo also produced peak early expression followed by a decline over time. In contrast to skin, however, liver expression of the reporter gene was relatively stable 4-8 days post-biolistic gene transfer, and remained detectable for nearly two weeks. The use of bioluminescence imaging techniques enabled efficient evaluation of reporter gene expression in vivo. Our results demonstrate that different tissues show different expression kinetics following gene transfer of the same reporter plasmid to different mouse tissues in vivo. We evaluated superficial (skin) and abdominal organ (liver) targets, and found that reporter gene expression peaked within the first two days post-transfer in each case, but declined most rapidly in the skin (3-4 days) compared to liver (10-14 days). This information is essential for designing effective gene therapy strategies in different target tissues.

Immune Rejection after Pancreatic Islet Cell Transplantation: In Vivo Dual Contrast-enhanced MR Imaging in a Mouse Model

PubMed Central

Wang, Ping; Schuetz, Christian; Ross, Alana; Dai, Guangping; Markmann, James F.

2013-01-01

Purpose: To detect adoptively transferred immune attack in a mouse model of islet cell transplantation by using a long-circulating paramagnetic T1 contrast agent, a protected graft copolymer (PGC) that is covalently linked to gadolinium–diethylenetriaminepentaacetic acid with fluorescein isothiocyanate (Gd-DTPA-F), which accumulates in the sites of inflammation that are characterized by vascular disruption. Materials and Methods: All animal experiments were performed in compliance with institutional guidelines and approved by the subcommittee on research animal care. Six nonobese diabetic severe combined immunodeficiency mice received transplanted human islet cells under the kidney capsule and adoptively transferred 5 × 106 splenocytes from 6-week-old nonobese diabetic mice. These mice also served as control subjects for comparison of pre- and postadoptive transfer MR imaging results. Mice that received phosphate-buffered saline solution only were included as nonadoptive-transfer control subjects (n = 2). In vivo magnetic resonance (MR) imaging was performed before and 17 hours after intravenous injections of PGC-Gd-DTPA-F, followed by histologic examination. Statistical differences were analyzed by means of a paired Student t test and repeated two-way analysis of variance. Results: MR imaging results showed significantly greater accumulation of PGC-Gd-DTPA-F in the graft area after immune attack initiated by adoptive transfer of splenocytes compared with that of the same area before the transfer (T1, 137.2 msec ± 39.3 and 239.5 msec ± 17.6, respectively; P < .001). These results were confirmed at histologic examination, which showed considerable leakage of the contrast agent into the islet cell interstitium. Conclusion: PGC-Gd-DTPA-F–enhanced MR imaging allows for the in vivo assessment of vascular damage of the graft T cell challenge. © RSNA, 2012 Supplemental material: http://radiology.rsna.org/lookup/suppl/doi:10.1148/radiol.12121129/-/DC1 PMID:23264346

Demonstration of neuron-glia transfer of precursors for GABA biosynthesis in a co-culture system of dissociated mouse cerebral cortex.

PubMed

Leke, Renata; Bak, Lasse K; Schousboe, Arne; Waagepetersen, Helle S

2008-12-01

Co-cultures of neurons and astrocytes were prepared from dissociated embryonic mouse cerebral cortex and cultured for 7 days. To investigate if these cultures may serve as a functional model system to study neuron-glia interaction with regard to GABA biosynthesis, the cells were incubated either in media containing [U-(13)C]glutamine (0.1, 0.3 and 0.5 mM) or 1 mM acetate plus 2.5 mM glucose plus 1 mM lactate. In the latter case one of the 3 substrates was uniformly (13)C labeled. Cellular contents and (13)C labeling of glutamate, GABA, aspartate and glutamine were determined in the cells after an incubation period of 2.5 h. The GABA biosynthetic machinery exhibited the expected complexity with regard to metabolic compartmentation and involvement of TCA cycle activity as seen in other culture systems containing GABAergic neurons. Metabolism of acetate clearly demonstrated glial synthesis of glutamine and its transfer to the neuronal compartment. It is concluded that this co-culture system serves as a reliable model in which functional and pharmacological aspects of GABA biosynthesis can be investigated.

Plasma butyrylcholinesterase regulates ghrelin to control aggression

PubMed Central

Chen, Vicky Ping; Gao, Yang; Geng, Liyi; Parks, Robin J.; Pang, Yuan-Ping; Brimijoin, Stephen

2015-01-01

Ongoing mouse studies of a proposed therapy for cocaine abuse based on viral gene transfer of butyrylcholinesterase (BChE) mutated for accelerated cocaine hydrolysis have yielded surprising effects on aggression. Further investigation has linked these effects to a reduction in circulating ghrelin, driven by BChE at levels ∼100-fold above normal. Tests with human BChE showed ready ghrelin hydrolysis at physiologic concentrations, and multiple low-mass molecular dynamics simulations revealed that ghrelin’s first five residues fit sterically and electrostatically into BChE’s active site. Consistent with in vitro results, male BALB/c mice with high plasma BChE after gene transfer exhibited sharply reduced plasma ghrelin. Unexpectedly, such animals fought less, both spontaneously and in a resident/intruder provocation model. One mutant BChE was found to be deficient in ghrelin hydrolysis. BALB/c mice transduced with this variant retained normal plasma ghrelin levels and did not differ from untreated controls in the aggression model. In contrast, C57BL/6 mice with BChE gene deletion exhibited increased ghrelin and fought more readily than wild-type animals. Collectively, these findings indicate that BChE-catalyzed ghrelin hydrolysis influences mouse aggression and social stress, with potential implications for humans. PMID:25646463

Alanine–glyoxylate aminotransferase-deficient mice, a model for primary hyperoxaluria that responds to adenoviral gene transfer

PubMed Central

Salido, Eduardo C.; Li, Xiao M.; Lu, Yang; Wang, Xia; Santana, Alfredo; Roy-Chowdhury, Namita; Torres, Armando; Shapiro, Larry J.; Roy-Chowdhury, Jayanta

2006-01-01

Mutations in the alanine–glyoxylate amino transferase gene (AGXT) are responsible for primary hyperoxaluria type I, a rare disease characterized by excessive hepatic oxalate production that leads to renal failure. We generated a null mutant mouse by targeted mutagenesis of the homologous gene, Agxt, in embryonic stem cells. Mutant mice developed normally, and they exhibited hyperoxaluria and crystalluria. Approximately half of the male mice in mixed genetic background developed calcium oxalate urinary stones. Severe nephrocalcinosis and renal failure developed after enhancement of oxalate production by ethylene glycol administration. Hepatic expression of human AGT1, the protein encoded by AGXT, by adenoviral vector-mediated gene transfer in Agxt−/− mice normalized urinary oxalate excretion and prevented oxalate crystalluria. Subcellular fractionation and immunofluorescence studies revealed that, as in the human liver, the expressed wild-type human AGT1 was predominantly localized in mouse hepatocellular peroxisomes, whereas the most common mutant form of AGT1 (G170R) was localized predominantly in the mitochondria. PMID:17110443

Whole-body irradiation increases the magnitude and persistence of adoptively transferred T cells associated with tumor regression in a mouse model of prostate cancer

PubMed Central

Ward-Kavanagh, Lindsay K.; Zhu, Junjia; Cooper, Timothy K.; Schell, Todd D.

2014-01-01

Adoptive immunotherapy has demonstrated efficacy in a subset of clinical and preclinical studies, but the T cells used for therapy often are rendered rapidly non-functional in tumor-bearing hosts. Recent evidence indicates that prostate cancer can be susceptible to immunotherapy, but most studies using autochthonous tumor models demonstrate only short-lived T-cell responses in the tolerogenic prostate microenvironment. Here, we assessed the efficacy of sublethal whole-body irradiation (WBI) to enhance the magnitude and duration of adoptively transferred CD8+ T cells in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model. We demonstrate that WBI promoted high-level accumulation of granzyme B (GzB)-expressing donor T cells both in lymphoid organs and in the prostate of TRAMP mice. Donor T cells remained responsive to vaccination in irradiated recipients, but a single round of WBI-enhanced adoptive immunotherapy failed to impact significantly the existing disease. Addition of a second round of immunotherapy promoted regression of established disease in half of the treated mice, with no progressions observed. Regression was associated with long-term persistence of effector/memory phenotype CD8+ donor cells. Administration of the second round of adoptive immunotherapy led to reacquisition of GzB expression by persistent T cells from the first transfer. These results indicate that WBI conditioning amplifies tumor-specific T cells in the TRAMP prostate and lymphoid tissue, and suggest that the initial treatment alters the tolerogenic microenvironment to increase antitumor activity by a second wave of donor cells. PMID:24801834

Reducing FLI1 Levels in the MRL/lpr Lupus Mouse Model Impacts T Cell Function by Modulating Glycosphingolipid Metabolism

PubMed Central

Richard, Erin Morris; Thiyagarajan, Thirumagal; Bunni, Marlene A.; Basher, Fahmin; Roddy, Patrick O.; Siskind, Leah J.; Nietert, Paul J.; Nowling, Tamara K.

2013-01-01

Systemic Lupus erythematosus (SLE) is an autoimmune disease caused, in part, by abnormalities in cells of the immune system including B and T cells. Genetically reducing globally the expression of the ETS transcription factor FLI1 by 50% in two lupus mouse models significantly improves disease measures and survival through an unknown mechanism. In this study we analyze the effects of reducing FLI1 in the MRL/lpr lupus prone model on T cell function. We demonstrate that adoptive transfer of MRL/lpr Fli1 +/+ or Fli1 +/- T cells and B cells into Rag1-deficient mice results in significantly decreased serum immunoglobulin levels in animals receiving Fli1 +/- lupus T cells compared to animals receiving Fli1 +/+ lupus T cells regardless of the genotype of co-transferred lupus B cells. Ex vivo analyses of MRL/lpr T cells demonstrated that Fli1 +/- T cells produce significantly less IL-4 during early and late disease and exhibited significantly decreased TCR-specific activation during early disease compared to Fli1 +/+ T cells. Moreover, the Fli1 +/- T cells expressed significantly less neuraminidase 1 (Neu1) message and decreased NEU activity during early disease and significantly decreased levels of glycosphingolipids during late disease compared to Fli1 +/+ T cells. FLI1 dose-dependently activated the Neu1 promoter in mouse and human T cell lines. Together, our results suggest reducing FLI1 in lupus decreases the pathogenicity of T cells by decreasing TCR-specific activation and IL-4 production in part through the modulation of glycosphingolipid metabolism. Reducing the expression of FLI1 or targeting the glycosphingolipid metabolic pathway in lupus may serve as a therapeutic approach to treating lupus. PMID:24040398

Reducing FLI1 levels in the MRL/lpr lupus mouse model impacts T cell function by modulating glycosphingolipid metabolism.

PubMed

Richard, Erin Morris; Thiyagarajan, Thirumagal; Bunni, Marlene A; Basher, Fahmin; Roddy, Patrick O; Siskind, Leah J; Nietert, Paul J; Nowling, Tamara K

2013-01-01

Systemic Lupus erythematosus (SLE) is an autoimmune disease caused, in part, by abnormalities in cells of the immune system including B and T cells. Genetically reducing globally the expression of the ETS transcription factor FLI1 by 50% in two lupus mouse models significantly improves disease measures and survival through an unknown mechanism. In this study we analyze the effects of reducing FLI1 in the MRL/lpr lupus prone model on T cell function. We demonstrate that adoptive transfer of MRL/lpr Fli1(+/+) or Fli1(+/-) T cells and B cells into Rag1-deficient mice results in significantly decreased serum immunoglobulin levels in animals receiving Fli1(+/-) lupus T cells compared to animals receiving Fli1(+/+) lupus T cells regardless of the genotype of co-transferred lupus B cells. Ex vivo analyses of MRL/lpr T cells demonstrated that Fli1(+/-) T cells produce significantly less IL-4 during early and late disease and exhibited significantly decreased TCR-specific activation during early disease compared to Fli1(+/+) T cells. Moreover, the Fli1(+/-) T cells expressed significantly less neuraminidase 1 (Neu1) message and decreased NEU activity during early disease and significantly decreased levels of glycosphingolipids during late disease compared to Fli1(+/+) T cells. FLI1 dose-dependently activated the Neu1 promoter in mouse and human T cell lines. Together, our results suggest reducing FLI1 in lupus decreases the pathogenicity of T cells by decreasing TCR-specific activation and IL-4 production in part through the modulation of glycosphingolipid metabolism. Reducing the expression of FLI1 or targeting the glycosphingolipid metabolic pathway in lupus may serve as a therapeutic approach to treating lupus.

Insights into the pathogenesis of GvHD: what mice can teach us about man.

PubMed

Hülsdünker, J; Zeiser, R

2015-01-01

Acute graft-vs-host disease (GvHD) is a life-threatening complication of allogeneic hematopoietic cell transplantation (allo-HCT). Most of the knowledge about the biology of GvHD is derived from mouse models of this disease and therefore a critical analysis of potential advantages and disadvantages of the murine GvHD models is important to classify and understand the findings made in these models. The central events leading up to GvHD were characterized in three phases which includes the tissue damage-phase, the T cell priming-phase and the effector-phase, when the disease becomes clinically overt. The role of individual cytokines, chemokines, transcription factor or receptors was studied in these models by using gene deficient or transgenic mice in the donor or recipient compartments. Besides, numerous studies have been performed in these models to prevent or treat GvHD. Several recent clinical trials were all based on previously reported findings from the mouse model of GvHD such as the trials on CCR5-blockade, donor statin treatment, vorinostat treatment or adoptive transfer of regulatory T cells for GvHD prevention. The different mouse models for GvHD and graft-vs-leukemia effects are critically reviewed and their impact on current clinical practice is discussed. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Various heterologous cells exhibit interferon induced transfer of viral resistance.

PubMed

Hughes, T K; Blalock, J E; Baron, S

1978-01-01

Previously it was shown that cocultivation of mouse L and human WISH or baby hamster kidney cells in the presence of mouse interferon resulted in decreased viral yield from both cell species. We now show that this phenomenon also occurs when rabbit kidney and human WISH cells, with their corresponding interferons, are cocultivated with human WISH and baby hamster kidney cells, respectively. This finding increases the number of donor cell types to three. The related finding that monkey VERO and chick embryo cells can be recipients of transferred resistance expands the number of heterologous recipient cell species capable of receiving transferred resistence to five. Not all cell types tested have been shown to function in this transfer system. The fact that VERO cells, which do not produce interferon, are capable of receiving transferred resistence is significant because it indicates that the mechanism of transfer does not involve production or interferon by the recipient cells.

Modeling of optical quadrature microscopy for imaging mouse embryos

NASA Astrophysics Data System (ADS)

Warger, William C., II; DiMarzio, Charles A.

2008-02-01

Optical quadrature microscopy (OQM) has been shown to provide the optical path difference through a mouse embryo, and has led to a novel method to count the total number of cells further into development than current non-toxic imaging techniques used in the clinic. The cell counting method has the potential to provide an additional quantitative viability marker for blastocyst transfer during in vitro fertilization. OQM uses a 633 nm laser within a modified Mach-Zehnder interferometer configuration to measure the amplitude and phase of the signal beam that travels through the embryo. Four cameras preceded by multiple beamsplitters record the four interferograms that are used within a reconstruction algorithm to produce an image of the complex electric field amplitude. Here we present a model for the electric field through the primary optical components in the imaging configuration and the reconstruction algorithm to calculate the signal to noise ratio when imaging mouse embryos. The model includes magnitude and phase errors in the individual reference and sample paths, fixed pattern noise, and noise within the laser and detectors. This analysis provides the foundation for determining the imaging limitations of OQM and the basis to optimize the cell counting method in order to introduce additional quantitative viability markers.

Adenovirus-mediated E2-EPF UCP Gene Transfer Prevents Autoamputation in a Mouse Model of Hindlimb Ischemia

PubMed Central

Lim, Jung Hwa; Shin, Hyo Jung; Park, Kyeong-Su; Lee, Chan Hee; Jung, Cho-Rok; Im, Dong-Soo

2012-01-01

E2-EPF ubiquitin carrier protein (UCP) stabilizes hypoxia-inducible factor-1α (HIF-1α) inducing ischemic vascular responses. Here, we investigated the effect of UCP gene transfer on therapeutic angiogenesis. Adenovirus-encoded UCP (Ad-F-UCP) increased the expression of vascular endothelial growth factor (VEGF) and fibroblast growth factor-2 (FGF-2) in cells and mice. Conditioned media from UCP-overexpressing cells promoted proliferation, tubule formation, and invasion of human umbilical-vascular-endothelial cells (HUVECs), and vascularization in chorioallantoic membrane (CAM) assay. Ad-F-UCP increased the vessel density in the Martigel plug assay, and generated copious vessel-like structures in the explanted muscle. The UCP effect on angiogenesis was dependent on VEGF and FGF-2. In mouse hindlimb ischemia model (N = 30/group), autoamputation (limb loss) occurred in 87% and 68% of the mice with saline and Ad encoding β-galactosidase (Ad-LacZ), respectively, whereas only 23% of the mice injected with Ad-F-UCP showed autoamputation after 21 days of treatment. Ad-F-UCP increased protein levels of HIF-1α, platelet-endothelial cell adhesion molecule-1 (PECAM-1), smooth muscle cell actin (SMA) in the ischemic muscle, and augmented blood vessels doubly positive for PECAM-1 and SMA. Consequently, UCP gene transfer prevented muscle degeneration and autoamputation of ischemic limb. The results suggest that E2-EPF UCP may be a target for therapeutic angiogenesis. PMID:22294149

Adenovirus-mediated E2-EPF UCP gene transfer prevents autoamputation in a mouse model of hindlimb ischemia.

PubMed

Lim, Jung Hwa; Shin, Hyo Jung; Park, Kyeong-Su; Lee, Chan Hee; Jung, Cho-Rok; Im, Dong-Soo

2012-04-01

E2-EPF ubiquitin carrier protein (UCP) stabilizes hypoxia-inducible factor-1α (HIF-1α) inducing ischemic vascular responses. Here, we investigated the effect of UCP gene transfer on therapeutic angiogenesis. Adenovirus-encoded UCP (Ad-F-UCP) increased the expression of vascular endothelial growth factor (VEGF) and fibroblast growth factor-2 (FGF-2) in cells and mice. Conditioned media from UCP-overexpressing cells promoted proliferation, tubule formation, and invasion of human umbilical-vascular-endothelial cells (HUVECs), and vascularization in chorioallantoic membrane (CAM) assay. Ad-F-UCP increased the vessel density in the Martigel plug assay, and generated copious vessel-like structures in the explanted muscle. The UCP effect on angiogenesis was dependent on VEGF and FGF-2. In mouse hindlimb ischemia model (N = 30/group), autoamputation (limb loss) occurred in 87% and 68% of the mice with saline and Ad encoding β-galactosidase (Ad-LacZ), respectively, whereas only 23% of the mice injected with Ad-F-UCP showed autoamputation after 21 days of treatment. Ad-F-UCP increased protein levels of HIF-1α, platelet-endothelial cell adhesion molecule-1 (PECAM-1), smooth muscle cell actin (SMA) in the ischemic muscle, and augmented blood vessels doubly positive for PECAM-1 and SMA. Consequently, UCP gene transfer prevented muscle degeneration and autoamputation of ischemic limb. The results suggest that E2-EPF UCP may be a target for therapeutic angiogenesis.

TRANSFER OF DRUG RESISTANCE BETWEEN ENTERIC BACTERIA INDUCED IN THE MOUSE INTESTINE

PubMed Central

Kasuya, Morimasa

1964-01-01

Kasuya, Morimasa (Nagoya University School of Medicine, Nagoya, Japan). Transfer of drug resistance between enteric bacteria induced in the mouse intestine. J. Bacteriol. 88:322–328. 1964.—Transfer of multiple drug resistance in the intestines of germ-free and conventional mice was studied with strains of Shigella, Escherichia, and Klebsiella. The transfer experiment was carried out under antibiotic-free conditions to eliminate the production of drug-resistant bacteria by antibiotics. All resistance factors (chloramphenicol, streptomycin, tetracycline, and sulfathiazole) were transferred with ease in the intestinal tracts of mice, when donors and recipients multiplied freely, and acquired resistance was further transferred to other sensitive enteric bacteria in the intestinal tract. Bacteria to which resistance factors were transferred showed, in most of the experiments, exactly the same level and pattern of resistance as the donors. Based on the above, a hypothesis that the same process may possibly occur in the human intestine is presented. PMID:14203347

Maternal exposure to an environmentally relevant dose of triclocarban results in perinatal exposure and potential alterations in offspring development in the mouse model.

PubMed

Enright, Heather A; Falso, Miranda J S; Malfatti, Michael A; Lao, Victoria; Kuhn, Edward A; Hum, Nicholas; Shi, Yilan; Sales, Ana Paula; Haack, Kurt W; Kulp, Kristen S; Buchholz, Bruce A; Loots, Gabriela G; Bench, Graham; Turteltaub, Kenneth W

2017-01-01

Triclocarban (TCC) is among the top 10 most commonly detected wastewater contaminants in both concentration and frequency. Its presence in water, as well as its propensity to bioaccumulate, has raised numerous questions about potential endocrine and developmental effects. Here, we investigated whether exposure to an environmentally relevant concentration of TCC could result in transfer from mother to offspring in CD-1 mice during gestation and lactation using accelerator mass spectrometry (AMS). 14C-TCC (100 nM) was administered to dams through drinking water up to gestation day 18, or from birth to post-natal day 10. AMS was used to quantify 14C-concentrations in offspring and dams after exposure. We demonstrated that TCC does effectively transfer from mother to offspring, both trans-placentally and via lactation. TCC-related compounds were detected in the tissues of offspring with significantly higher concentrations in the brain, heart and fat. In addition to transfer from mother to offspring, exposed offspring were heavier in weight than unexposed controls demonstrating an 11% and 8.5% increase in body weight for females and males, respectively. Quantitative real-time polymerase chain reaction (qPCR) was used to examine changes in gene expression in liver and adipose tissue in exposed offspring. qPCR suggested alterations in genes involved in lipid metabolism in exposed female offspring, which was consistent with the observed increased fat pad weights and hepatic triglycerides. This study represents the first report to quantify the transfer of an environmentally relevant concentration of TCC from mother to offspring in the mouse model and evaluate bio-distribution after exposure using AMS. Our findings suggest that early-life exposure to TCC may interfere with lipid metabolism and could have implications for human health.

Gaze transfer in remote cooperation: is it always helpful to see what your partner is attending to?

PubMed

Müller, Romy; Helmert, Jens R; Pannasch, Sebastian; Velichkovsky, Boris M

2013-01-01

Establishing common ground in remote cooperation is challenging because nonverbal means of ambiguity resolution are limited. In such settings, information about a partner's gaze can support cooperative performance, but it is not yet clear whether and to what extent the abundance of information reflected in gaze comes at a cost. Specifically, in tasks that mainly rely on spatial referencing, gaze transfer might be distracting and leave the partner uncertain about the meaning of the gaze cursor. To examine this question, we let pairs of participants perform a joint puzzle task. One partner knew the solution and instructed the other partner's actions by (1) gaze, (2) speech, (3) gaze and speech, or (4) mouse and speech. Based on these instructions, the acting partner moved the pieces under conditions of high or low autonomy. Performance was better when using either gaze or mouse transfer compared to speech alone. However, in contrast to the mouse, gaze transfer induced uncertainty, evidenced in delayed responses to the cursor. Also, participants tried to resolve ambiguities by engaging in more verbal effort, formulating more explicit object descriptions and fewer deictic references. Thus, gaze transfer seems to increase uncertainty and ambiguity, thereby complicating grounding in this spatial referencing task. The results highlight the importance of closely examining task characteristics when considering gaze transfer as a means of support.

Altering host resistance to infections through microbial transplantation.

PubMed

Willing, Benjamin P; Vacharaksa, Anjalee; Croxen, Matthew; Thanachayanont, Teerawat; Finlay, B Brett

2011-01-01

Host resistance to bacterial infections is thought to be dictated by host genetic factors. Infections by the natural murine enteric pathogen Citrobacter rodentium (used as a model of human enteropathogenic and enterohaemorrhagic E. coli infections) vary between mice strains, from mild self-resolving colonization in NIH Swiss mice to lethality in C3H/HeJ mice. However, no clear genetic component had been shown to be responsible for the differences observed with C. rodentium infections. Because the intestinal microbiota is important in regulating resistance to infection, and microbial composition is dependent on host genotype, it was tested whether variations in microbial composition between mouse strains contributed to differences in "host" susceptibility by transferring the microbiota of resistant mice to lethally susceptible mice prior to infection. Successful transfer of the microbiota from resistant to susceptible mice resulted in delayed pathogen colonization and mortality. Delayed mortality was associated with increased IL-22 mediated innate defense including antimicrobial peptides Reg3γ and Reg3β, and immunono-neutralization of IL-22 abrogated the beneficial effect of microbiota transfer. Conversely, depletion of the native microbiota in resistant mice by antibiotics and transfer of the susceptible mouse microbiota resulted in reduced innate defenses and greater pathology upon infection. This work demonstrates the importance of the microbiota and how it regulates mucosal immunity, providing an important factor in susceptibility to enteric infection. Transfer of resistance through microbial transplantation (bacteriotherapy) provides additional mechanisms to alter "host" resistance, and a novel means to alter enteric infection and to study host-pathogen interactions.

Sildenafil Citrate Increases Fetal Weight in a Mouse Model of Fetal Growth Restriction with a Normal Vascular Phenotype

PubMed Central

Dilworth, Mark Robert; Andersson, Irene; Renshall, Lewis James; Cowley, Elizabeth; Baker, Philip; Greenwood, Susan; Sibley, Colin Peter; Wareing, Mark

2013-01-01

Fetal growth restriction (FGR) is defined as the inability of a fetus to achieve its genetic growth potential and is associated with a significantly increased risk of morbidity and mortality. Clinically, FGR is diagnosed as a fetus falling below the 5th centile of customised growth charts. Sildenafil citrate (SC, Viagra™), a potent and selective phosphodiesterase-5 inhibitor, corrects ex vivo placental vascular dysfunction in FGR, demonstrating potential as a therapy for this condition. However, many FGR cases present without an abnormal vascular phenotype, as assessed by Doppler measures of uterine/umbilical artery blood flow velocity. Thus, we hypothesized that SC would not increase fetal growth in a mouse model of FGR, the placental-specific Igf2 knockout mouse, which has altered placental exchange capacity but normal placental blood flow. Fetal weights were increased (by 8%) in P0 mice following maternal SC treatment (0.4 mg/ml) via drinking water. There was also a trend towards increased placental weight in treated P0 mice (P = 0.056). Additionally, 75% of the P0 fetal weights were below the 5th centile, the criterion used to define human FGR, of the non-treated WT fetal weights; this was reduced to 51% when dams were treated with SC. Umbilical artery and vein blood flow velocity measures confirmed the lack of an abnormal vascular phenotype in the P0 mouse; and were unaffected by SC treatment. 14C-methylaminoisobutyric acid transfer (measured to assess effects on placental nutrient transporter activity) per g placenta was unaffected by SC, versus untreated, though total transfer was increased, commensurate with the trend towards larger placentas in this group. These data suggest that SC may improve fetal growth even in the absence of an abnormal placental blood flow, potentially affording use in multiple sub-populations of individuals presenting with FGR. PMID:24204949

Adeno-associated virus-mediated gene delivery into the scala media of the normal and deafened adult mouse ear.

PubMed

Kilpatrick, L A; Li, Q; Yang, J; Goddard, J C; Fekete, D M; Lang, H

2011-06-01

Murine models are ideal for studying cochlear gene transfer, as many hearing loss-related mutations have been discovered and mapped within the mouse genome. However, because of the small size and delicate nature, the membranous labyrinth of the mouse is a challenging target for the delivery of viral vectors. To minimize injection trauma, we developed a procedure for the controlled release of adeno-associated viruses (AAVs) into the scala media of adult mice. This procedure poses minimal risk of injury to structures of the cochlea and middle ear, and allows for near-complete preservation of low and middle frequency hearing. In this study, transduction efficiency and cellular specificity of AAV vectors (serotypes 1, 2, 5, 6 and 8) were investigated in normal and drug-deafened ears. Using the cytomegalovirus promoter to drive gene expression, a variety of cell types were transduced successfully, including sensory hair cells and supporting cells, as well as cells in the auditory nerve and spiral ligament. Among all five serotypes, inner hair cells were the most effectively transduced cochlear cell type. All five serotypes of AAV vectors transduced cells of the auditory nerve, though serotype 8 was the most efficient vector for transduction. Our findings indicate that efficient AAV inoculation (via the scala media) can be performed in adult mouse ears, with hearing preservation a realistic goal. The procedure we describe may also have applications for intra-endolymphatic drug delivery in many mouse models of human deafness.

Adeno-associated virus-mediated gene delivery into the scala media of the normal and deafened adult mouse ear

PubMed Central

Kilpatrick, Lauren A.; Li, Qian; Yang, John; Goddard, John C; Fekete, Donna M.; Lang, Hainan

2010-01-01

Murine models are ideal for studying cochlear gene transfer as many hearing loss-related mutations have been discovered and mapped within the mouse genome. However, due to its small size and delicate nature, the membranous labyrinth of the mouse is a challenging target for delivery of viral vectors. To minimize injection trauma, we developed a procedure for the controlled release of adeno-associated viruses (AAV) into the scala media of adult mice. This procedure poses minimal risk of injury to structures of the cochlea and middle ear and allows for near-complete preservation of low and middle frequency hearing. In the present study, transduction efficiency and cellular specificity of AAV vectors (serotypes 1, 2, 5, 6, and 8) were investigated in normal and drug-deafened ears. Using the cytomegalovirus (CMV) promoter to drive gene expression, a variety of cell types were transduced successfully, including sensory hair cells and supporting cells, as well as cells in the auditory nerve and spiral ligament. Among all five serotypes, inner hair cells (IHCs) were the most effectively transduced cochlear cell type. All five serotypes of AAV vectors transduced cells of the auditory nerve, though serotype 8 was the most efficient vector for transduction. Our findings indicate that efficient AAV inoculation (via the scala media) can be performed in adult mouse ears, with hearing preservation a realistic goal. The procedure we describe may also have applications for intra-endolymphatic drug delivery in many mouse models of human deafness. PMID:21209625

Induction of Human Blood Group A Antigen Expression on Mouse Cells, Using Lentiviral Gene Transduction

PubMed Central

Fan, Xiaohu; Lang, Haili; Zhou, Xianpei; Zhang, Li; Yin, Rong; Maciejko, Jessica; Giannitsos, Vasiliki; Motyka, Bruce; Medin, Jeffrey A.; Platt, Jeffrey L.

2010-01-01

Abstract The ABO histo-blood group system is the most important antigen system in transplantation medicine, yet no small animal model of the ABO system exists. To determine the feasibility of developing a murine model, we previously subcloned the human α-1,2-fucosyltransferase (H-transferase, EC 2.4.1.69) cDNA and the human α-1,3-N-acetylgalactosaminyltransferase (A-transferase, EC 2.4.1.40) cDNA into lentiviral vectors to study their ability to induce human histo-blood group A antigen expression on mouse cells. Herein we investigated the optimal conditions for human A and H antigen expression in murine cells. We determined that transduction of a bicistronic lentiviral vector (LvEF1-AH-trs) resulted in the expression of A antigen in a mouse endothelial cell line. We also studied the in vivo utility of this vector to induce human A antigen expression in mouse liver. After intrahepatic injection of LvEF1-AH-trs, A antigen expression was observed on hepatocytes as detected by immunohistochemistry and real-time RT-PCR. In human group A erythrocyte-sensitized mice, A antigen expression in the liver was associated with tissue damage, and deposition of antibody and complement. These results suggest that this gene transfer strategy can be used to simulate the human ABO blood group system in a murine model. This model will facilitate progress in the development of interventions for ABO-incompatible transplantation and transfusion scenarios, which are difficult to develop in clinical or large animal settings. PMID:20163247

Quantitative Metaproteomics and Activity-Based Probe Enrichment Reveals Significant Alterations in Protein Expression from a Mouse Model of Inflammatory Bowel Disease.

PubMed

Mayers, Michael D; Moon, Clara; Stupp, Gregory S; Su, Andrew I; Wolan, Dennis W

2017-02-03

Tandem mass spectrometry based shotgun proteomics of distal gut microbiomes is exceedingly difficult due to the inherent complexity and taxonomic diversity of the samples. We introduce two new methodologies to improve metaproteomic studies of microbiome samples. These methods include the stable isotope labeling in mammals to permit protein quantitation across two mouse cohorts as well as the application of activity-based probes to enrich and analyze both host and microbial proteins with specific functionalities. We used these technologies to study the microbiota from the adoptive T cell transfer mouse model of inflammatory bowel disease (IBD) and compare these samples to an isogenic control, thereby limiting genetic and environmental variables that influence microbiome composition. The data generated highlight quantitative alterations in both host and microbial proteins due to intestinal inflammation and corroborates the observed phylogenetic changes in bacteria that accompany IBD in humans and mouse models. The combination of isotope labeling with shotgun proteomics resulted in the total identification of 4434 protein clusters expressed in the microbial proteomic environment, 276 of which demonstrated differential abundance between control and IBD mice. Notably, application of a novel cysteine-reactive probe uncovered several microbial proteases and hydrolases overrepresented in the IBD mice. Implementation of these methods demonstrated that substantial insights into the identity and dysregulation of host and microbial proteins altered in IBD can be accomplished and can be used in the interrogation of other microbiome-related diseases.

Number and location of mouse mammary tumor virus proviral DNA in mouse DNA of normal tissue and of mammary tumors.

PubMed Central

Groner, B; Hynes, N E

1980-01-01

The Southern DNA filter transfer technique was used to characterize the genomic location of the mouse mammary tumor proviral DNA in different inbred strains of mice. Two of the strains (C3H and CBA) arose from a cross of a Bagg albino (BALB/c) mouse and a DBA mouse. The mouse mammary tumor virus-containing restriction enzyme DNA fragments of these strains had similar patterns, suggesting that the proviruses of these mice are in similar genomic locations. Conversely, the pattern arising from the DNA of the GR mouse, a strain genetically unrelated to the others, appeared different, suggesting that its mouse mammary tumor proviruses are located in different genomic sites. The structure of another gene, that coding for beta-globin, was also compared. The mice strains which we studied can be categorized into two classes, expressing either one or two beta-globin proteins. The macroenvironment of the beta-globin gene appeared similar among the mice strains belonging to one genetic class. Female mice of the C3H strain exogenously transmit mouse mammary tumor virus via the milk, and their offspring have a high incidence of mammary tumor occurrence. DNA isolated from individual mammary tumors taken from C3H mice or from BALB/c mice foster nursed on C3H mothers was analyzed by the DNA filter transfer technique. Additional mouse mammary tumor virus-containing fragments were found in the DNA isolated from each mammary tumor. These proviral sequences were integrated into different genomic sites in each tumor. Images PMID:6245257

The chemokine receptor CX3CR1 coordinates monocyte recruitment and endothelial regeneration after arterial injury.

PubMed

Getzin, Tobias; Krishnasamy, Kashyap; Gamrekelashvili, Jaba; Kapanadze, Tamar; Limbourg, Anne; Häger, Christine; Napp, L Christian; Bauersachs, Johann; Haller, Hermann; Limbourg, Florian P

2018-02-01

Regeneration of arterial endothelium after injury is critical for the maintenance of normal blood flow, cell trafficking, and vascular function. Using mouse models of carotid injury, we show that the transition from a static to a dynamic phase of endothelial regeneration is marked by a strong increase in endothelial proliferation, which is accompanied by induction of the chemokine CX 3 CL1 in endothelial cells near the wound edge, leading to progressive recruitment of Ly6C lo monocytes expressing high levels of the cognate CX 3 CR1 chemokine receptor. In Cx3cr1 -deficient mice recruitment of Ly6C lo monocytes, endothelial proliferation and regeneration of the endothelial monolayer after carotid injury are impaired, which is rescued by acute transfer of normal Ly6C lo monocytes. Furthermore, human non-classical monocytes induce proliferation of endothelial cells in co-culture experiments in a VEGFA-dependent manner, and monocyte transfer following carotid injury promotes endothelial wound closure in a hybrid mouse model in vivo Thus, CX 3 CR1 coordinates recruitment of specific monocyte subsets to sites of endothelial regeneration, which promote endothelial proliferation and arterial regeneration. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

Development of an in vitro cell culture model to study milk to plasma ratios of therapeutic drugs.

PubMed

Athavale, Maithili A; Maitra, Anurupa; Patel, Shahnaz; Bhate, Vijay R; Toddywalla, Villi S

2013-01-01

To create an in vitro cell culture model to predict the M/P (concentration of drug in milk/concentration in maternal plasma) ratios of therapeutic drugs viz. rifampicin, theophylline, paracetamol, and aspirin. An in vitro cell culture model using CIT3 cells (mouse mammary epithelial cells) was created by culturing the cells on transwells. The cells formed an integral monolayer, allowing only transcellular transport as it happens in vivo. Functionality of the cells was confirmed through scanning electron microscopy. Time wise transfer of the study drugs from plasma to milk was studied and compared with actual (in vivo) M/P ratios obtained at reported tmax for the respective drugs. The developed model mimicked two important intrinsic factors of mammary epithelial cells viz. secretory and tight-junction properties and also the passive route of drug transport. The in vitro M/P ratios at reported tmax were 0.23, 0.61, 0.87, and 0.03 respectively, for rifampicin, theophylline, paracetamol, and salicylic acid as compared to 0.29, 0.65, 0.65, and 0.22, respectively, in vitro. Our preliminary effort to develop an in vitro physiological model showed promising results. Transfer rate of the drugs using the developed model compared well with the transfer potential seen in vivo except for salicylic acid, which was transferred in far lower concentration in vitro. The model has a potential to be developed as a non-invasive alternative to the in vitro technique for determining the transfer of therapeutic drugs into breast milk.

Pharmacokinetic Properties of Memantine after a Single Intraperitoneal Administration and Multiple Oral Doses in Euploid Mice and in the Ts65Dn Mouse Model of Down's Syndrome.

PubMed

Victorino, Daniella B; Bederman, Ilya R; Costa, Alberto C S

2017-11-01

Memantine is a drug approved for the treatment of moderate-to-severe Alzheimer's disease (AD), and there is ongoing research on the potential expansion of its clinical applicability. Published data on the pharmacokinetics of memantine in the mouse are still incomplete, particularly for chronic administration regimens and mouse models of specific genetic disorders. Down's syndrome (DS) is a genetic disorder known to affect multiple organs and systems, with the potential to alter significantly drug pharmacokinetics. Here, we describe a simple, efficient and sensitive GC/MS-based procedure for the determination of memantine concentrations in murine blood and tissue samples. We analysed pharmacokinetic properties of memantine, particularly its distribution in blood, brain and liver in the Ts65Dn mouse model of DS and euploid F1 hybrid mice after single intraperitoneal administrations of increasing doses of this drug. We also determined steady-state memantine concentrations in plasma, brain and liver after chronic oral administration of this drug in adult male Ts65Dn mice, euploid littermate controls and nursing or pregnant Ts65Dn mice. Our results revalidated the acute dose of memantine used in previously published work, determined the appropriate amount of memantine to be mixed into mouse chow to achieve steady and pharmacologically relevant plasma and tissue levels of this drug and demonstrated that memantine can be transferred from mother to offspring via maternal milk and placenta. Most of these findings are potentially applicable not only to the study of DS but also to other neurodevelopmental and neurodegenerative disorders. © 2017 The Authors. Basic & Clinical Pharmacology & Toxicology published by John Wiley & Sons Ltd on behalf of Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).

Enterococcus faecalis Sex Pheromone cCF10 Enhances Conjugative Plasmid Transfer In Vivo.

PubMed

Hirt, Helmut; Greenwood-Quaintance, Kerryl E; Karau, Melissa J; Till, Lisa M; Kashyap, Purna C; Patel, Robin; Dunny, Gary M

2018-02-13

Cell-cell communication mediated by peptide pheromones (cCF10 [CF]) is essential for high-frequency plasmid transfer in vitro in Enterococcus faecalis To examine the role of pheromone signaling in vivo , we established either a CF-producing (CF+) recipient or a recipient producing a biologically inactive variant of CF (CF- recipient) in a germfree mouse model 3 days before donor inoculation and determined transfer frequencies of the pheromone-inducible plasmid pCF10. Plasmid transfer was detected in the upper and middle sections of the intestinal tract 5 h after donor inoculation and was highly efficient in the absence of antibiotic selection. The transconjugant/donor ratio reached a maximum level approaching 1 on day 4 in the upper intestinal tract. Plasmid transfer was significantly lower with the CF- recipient. While rescue of the CF- mating defect by coculture with CF+ recipients is easily accomplished in vitro , no extracellular complementation occurred in vivo This suggests that most pheromone signaling in the gut occurs between recipient and donor cells in very close proximity. Plasmid-bearing cells (donors plus transconjugants) steadily increased in the population from 0.1% after donor inoculation to about 10% at the conclusion of the experiments. This suggests a selective advantage of pCF10 carriage distinct from antibiotic resistance or bacteriocin production. Our results demonstrate that pheromone signaling is required for efficient pCF10 transfer in vivo In the absence of CF+ recipients, a low level of transfer to CF- recipients occurred in the gut. This may result from low-level host-mediated induction of the donors in the gastrointestinal (GI) tract, similar to that previously observed in serum. IMPORTANCE Horizontal gene transfer is a major factor in the biology of Enterococcus faecalis , an important nosocomial pathogen. Previous studies showing efficient conjugative plasmid transfer in the gastrointestinal (GI) tracts of experimental animals did not examine how the enterococcal sex pheromone response impacts the efficiency of transfer. Our study demonstrates for the first time pheromone-enhanced, high-frequency plasmid transfer of E. faecalis plasmid pCF10 in a mouse model in the absence of antibiotic or bacteriocin selection. Pheromone production by recipients dramatically increased plasmid transfer in germfree mice colonized initially with recipients, followed by donors. The presence of a coresident community of common gut microbes did not significantly reduce in vivo plasmid transfer between enterococcal donors and recipients. In mice colonized with enterococcal recipients, we detected plasmid transfer in the intestinal tract within 5 h of addition of donors, before transconjugants could be cultured from feces. Surprisingly, pCF10 carriage provided a competitive fitness advantage unrelated to antibiotic resistance or bacteriocin production. Copyright © 2018 Hirt et al.

Adoptive transfer of T regulatory cells inhibits lipopolysaccharide-induced inflammation in fetal brain tissue in a late-pregnancy preterm birth mouse model.

PubMed

Wang, Fan; Xiao, Mi; Chen, Ru-Juan; Lin, Xiao-Jie; Siddiq, Muhammad; Liu, Li

2017-02-01

To evaluate the effect of regulatory T cells (Tregs) on the inflammation resulting from lipopolysaccharide (LPS) challenge in prenatal brain tissue, Tregs isolated from pregnant mice were transferred into model mice, and the expression levels of fork head family transcription factor (Foxp3), interleukin-6 (IL-6), CD68 (a marker of microglia), and toll-like receptor 4 (TLR-4) were assessed in the fetal brain tissue. Foxp3, IL-6, and TLR-4 expression were detected by polymerase chain reaction and Western blot; CD68 expression level was detected using immunochemical analysis. Foxp3, IL-6, TLR-4, and CD68 expressions in fetal brain were significantly induced by maternal LPS administration, and the increased expression levels were markedly reduced by adoptive transfer of Tregs. Maternal LPS exposure significantly induced inflammation in perinatal brain tissue, and Tregs negatively regulated this LPS-induced inflammation. © 2016 International Federation for Cell Biology.

Computational modeling highlights disordered Formin Homology 1 domain's role in profilin-actin transfer.

PubMed

Horan, Brandon G; Zerze, Gül H; Kim, Young C; Vavylonis, Dimitrios; Mittal, Jeetain

2018-05-13

Formins accelerate actin polymerization, assumed to occur through flexible FH1 domain mediated transfer of profilin-actin to the barbed end. To study FH1 properties and address sequence effects including varying length/distributionof profilin-binding proline-rich motifs, we performed allatom simulations of mouse mDia1, mDia2; budding yeast Bni1, Bnr1; fission yeast Cdc12, For3, and Fus1 FH1s. We find FH1 has flexible regions between high propensity polyproline helix regions. A coarse-grained model retaining sequence-specificity, assuming rigid polyproline segments,describes their size. Multiple bound profilins or profilin-actin complexes expand mDia1-FH1, which may be important in cells. Simulations of the barbed end bound to Bni1-FH1-FH2 dimer show the leading FH1 can better transfer profilin or profilin-actin, having decreasing probability with increasing distance from FH2. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Toward Distinguishing Recurrent Tumor From Radiation Necrosis: DWI and MTC in a Gamma Knife–Irradiated Mouse Glioma Model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Perez-Torres, Carlos J.; Engelbach, John A.; Cates, Jeremy

Purpose: Accurate noninvasive diagnosis is vital for effective treatment planning. Presently, standard anatomical magnetic resonance imaging (MRI) is incapable of differentiating recurring tumor from delayed radiation injury, as both lesions are hyperintense in both postcontrast T1- and T2-weighted images. Further studies are therefore necessary to identify an MRI paradigm that can differentially diagnose these pathologies. Mouse glioma and radiation injury models provide a powerful platform for this purpose. Methods and Materials: Two MRI contrasts that are widely used in the clinic were chosen for application to a glioma/radiation-injury model: diffusion weighted imaging, from which the apparent diffusion coefficient (ADC) ismore » obtained, and magnetization transfer contrast, from which the magnetization transfer ratio (MTR) is obtained. These metrics were evaluated longitudinally, first in each lesion type alone–glioma versus irradiation – and then in a combined irradiated glioma model. Results: MTR was found to be consistently decreased in all lesions compared to nonlesion brain tissue (contralateral hemisphere), with limited specificity between lesion types. In contrast, ADC, though less sensitive to the presence of pathology, was increased in radiation injury and decreased in tumors. In the irradiated glioma model, ADC also increased immediately after irradiation, but decreased as the tumor regrew. Conclusions: ADC is a better metric than MTR for differentiating glioma from radiation injury. However, MTR was more sensitive to both tumor and radiation injury than ADC, suggesting a possible role in detecting lesions that do not enhance strongly on T1-weighted images.« less

The Role of Heterotypic DENV-specific CD8+T Lymphocytes in an Immunocompetent Mouse Model of Secondary Dengue Virus Infection.

PubMed

Talarico, Laura B; Batalle, Juan P; Byrne, Alana B; Brahamian, Jorge M; Ferretti, Adrián; García, Ayelén G; Mauri, Aldana; Simonetto, Carla; Hijano, Diego R; Lawrence, Andrea; Acosta, Patricio L; Caballero, Mauricio T; Paredes Rojas, Yésica; Ibañez, Lorena I; Melendi, Guillermina A; Rey, Félix A; Damonte, Elsa B; Harris, Eva; Polack, Fernando P

2017-06-01

Dengue is the most prevalent arthropod-borne viral disease worldwide and is caused by the four dengue virus serotypes (DENV-1-4). Sequential heterologous DENV infections can be associated with severe disease manifestations. Here, we present an immunocompetent mouse model of secondary DENV infection using non mouse-adapted DENV strains to investigate the pathogenesis of severe dengue disease. C57BL/6 mice infected sequentially with DENV-1 (strain Puerto Rico/94) and DENV-2 (strain Tonga/74) developed low platelet counts, internal hemorrhages, and increase of liver enzymes. Cross-reactive CD8 + T lymphocytes were found to be necessary and sufficient for signs of severe disease by adoptively transferring of DENV-1-immune CD8 + T lymphocytes before DENV-2 challenge. Disease signs were associated with production of tumor necrosis factor (TNF)-α and elevated cytotoxicity displayed by heterotypic anti-DENV-1 CD8 + T lymphocytes. These findings highlight the critical role of heterotypic anti-DENV CD8 + T lymphocytes in manifestations of severe dengue disease. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Mouse homologues of human hereditary disease.

PubMed Central

Searle, A G; Edwards, J H; Hall, J G

1994-01-01

Details are given of 214 loci known to be associated with human hereditary disease, which have been mapped on both human and mouse chromosomes. Forty two of these have pathological variants in both species; in general the mouse variants are similar in their effects to the corresponding human ones, but exceptions include the Dmd/DMD and Hprt/HPRT mutations which cause little, if any, harm in mice. Possible reasons for phenotypic differences are discussed. In most pathological variants the gene product seems to be absent or greatly reduced in both species. The extensive data on conserved segments between human and mouse chromosomes are used to predict locations in the mouse of over 50 loci of medical interest which are mapped so far only on human chromosomes. In about 80% of these a fairly confident prediction can be made. Some likely homologies between mapped mouse loci and unmapped human ones are also given. Sixty six human and mouse proto-oncogene and growth factor gene homologies are also listed; those of confirmed location are all in known conserved segments. A survey of 18 mapped human disease loci and chromosome regions in which the manifestation or severity of pathological effects is thought to be the result of genomic imprinting shows that most of the homologous regions in the mouse are also associated with imprinting, especially those with homologues on human chromosomes 11p and 15q. Useful methods of accelerating the production of mouse models of human hereditary disease include (1) use of a supermutagen, such as ethylnitrosourea (ENU), (2) targeted mutagenesis involving ES cells, and (3) use of gene transfer techniques, with production of 'knockout mutations'. PMID:8151633

Mutated-leptin gene transfer induces increases in body weight by electroporation and hydrodynamics-based gene delivery in mice.

PubMed

Xiang, Lan; Murai, Atsushi; Muramatsu, Tatsuo

2005-12-01

To investigate whether in vivo gene transfer causes leptin-antagonistic effects on food intake, animal body weight and fat tissue weight, the R128Q mutated-leptin gene, an R to Q substitution at position 128 of mouse leptin, was transferred into mouse liver and leg muscle by electroporation and hydrodynamics-based gene delivery. Mutated-leptin gene transfer by electroporation caused significant increases in body weight at 5 days and after (5.4% increase relative to control; p<0.05). Hydrodynamics-based gene delivery of the mutated-leptin gene also caused an increase in body weight (3.0% increase relative to control; p<0.05). Mutated-leptin gene transfer by electroporation significantly increased the tissue weight of epididymal white fat and neuropeptide Y mRNA expression in the hypothalamus compared with those of the control group 3 weeks after gene transfer (p<0.05). These results suggest that mutated-leptin gene transfer successfully produced leptin-antagonistic effects by modulating the central regulator of energy homeostasis. Also, the extent of leptin-antagonistic effects by electroporation was much higher than hydrodynamics-based gene delivery, with at least single gene transfer.

Archiving and Distributing Mouse Lines by Sperm Cryopreservation, IVF, and Embryo Transfer

PubMed Central

Takahashi, Hideko; Liu, Chengyu

2012-01-01

The number of genetically modified mouse lines has been increasing exponentially in the past few decades. In order to safeguard them from accidental loss and genetic drifting, to reduce animal housing cost, and to efficiently distribute them around the world, it is important to cryopreserve these valuable genetic resources. Preimplantation-stage embryos from thousands of mouse lines have been cryopreserved during the past two to three decades. Although reliable, this method requires several hundreds of embryos, which demands a sizable breeding colony, to safely preserve each line. This requirement imposes significant delay and financial burden for the archiving effort. Sperm cryopreservation is now emerging as the leading method for storing and distributing mouse lines, largely due to the recent finding that addition of a reducing agent, monothioglycerol, into the cryoprotectant can significantly increase the in vitro fertilization (IVF) rate in many mouse strains, including the most widely used C57BL/6 strain. This method is quick, inexpensive, and requires only two breeding age male mice, but it still remains tricky and strain-dependent. A small change in experimental conditions can lead to significant variations in the outcome. In this chapter, we describe in detail our sperm cryopreservation, IVF, and oviduct transfer procedures for storing and reviving genetically modified mouse lines. PMID:20691860

[Development of a hepatitis B virus carrier transgenic mice model].

PubMed

Caner, Müge; Arat, Sezen; Bircan, Rifat

2008-01-01

The studies for the development of transgenic mice models which provide important profits for the studies concerning immunopathogenesis of hepatitis B virus (HBV) infections are in progress since 20 years. For this purpose different lineages bearing whole HBV genome or selected viral genes have been developed and their usage in clarifying the HBV replication and pathogenesis mechanisms have been emphasized. The aim of this study was to develop and breed a HBV carrier mice model. In the study the full HBV genome has been transferred to mouse embryos by microinjection procedure. Following transgenic manipulation, the HBV carriers among the daughter mice have been detected by molecular methods in which HBV-DNA replication and expression have been shown. The manipulations for transgene transfers have been performed in TUBITAK Marmara Research Center Transgene Laboratory, Gebze, Istanbul. The HBV-DNA carrier mice have been demonstrated by polymerase chain reaction (PCR) using the DNA samples obtained from tail tissues and also by dot-blot hybridization of the mice sera. Integrated HBV-DNA has been detected by applying in-situ hybridization to the liver tissue sections. HBV-DNA expression has been shown by reverse transcriptase PCR method with total RNA molecules that have been isolated from the liver tissues of the HBV-DNA carrier mice. HBsAg has been detected in the liver by immunohistochemical method, and HBsAg and HBeAg have additionally been demonstrated by ELISA. HBV genome, expression of the genome and the expression products have been determined in approximately 10% of the mice of which HBV-DNA have been transferred. By inbreeding heterozygote carrier mice, homozygote HBV transgenic mice line have been obtained. These HBV transgenic mice are the first lineages developed in our country. It is hopefully thought that this HBV carrier transgenic mouse model may contribute to the studies on the pathogenesis of HBV infections which are important health problems in the world as well as in Turkey.

Adoptive cell transfer of resistance to Mycobacterium leprae infections in mice.

PubMed Central

Lowe, C; Brett, S J; Rees, R J

1985-01-01

Cells were transferred from mice intradermally vaccinated with killed Mycobacterium leprae to sublethally irradiated recipients. Unseparated cells from lymph nodes or spleens of M. leprae vaccinated mice were found to cause significant inhibition of the growth of a subsequent M. leprae challenge in mouse footpads for up to 26 weeks after vaccination. Vaccination with live BCG and cells transferred from BCG-vaccinated mice caused no significant inhibition of M. leprae growth in mouse footpads. Cell separation into fractions containing predominantly B and T lymphocytes showed that the inhibition of growth was due to M. leprae-sensitized T lymphocytes. M. leprae vaccinated mice were also skin tested with soluble M. leprae antigen and showed maximum delayed hypersensitivity responses 4 weeks after vaccination. PMID:3876183

Propagating Humanized BLT Mice for the Study of Human Immunology and Immunotherapy.

PubMed

Smith, Drake J; Lin, Levina J; Moon, Heesung; Pham, Alexander T; Wang, Xi; Liu, Siyuan; Ji, Sunjong; Rezek, Valerie; Shimizu, Saki; Ruiz, Marlene; Lam, Jennifer; Janzen, Deanna M; Memarzadeh, Sanaz; Kohn, Donald B; Zack, Jerome A; Kitchen, Scott G; An, Dong Sung; Yang, Lili

2016-12-15

The humanized bone marrow-liver-thymus (BLT) mouse model harbors a nearly complete human immune system, therefore providing a powerful tool to study human immunology and immunotherapy. However, its application is greatly limited by the restricted supply of human CD34 + hematopoietic stem cells and fetal thymus tissues that are needed to generate these mice. The restriction is especially significant for the study of human immune systems with special genetic traits, such as certain human leukocyte antigen (HLA) haplotypes or monogene deficiencies. To circumvent this critical limitation, we have developed a method to quickly propagate established BLT mice. Through secondary transfer of bone marrow cells and human thymus implants from BLT mice into NSG (NOD/SCID/IL-2Rγ -/- ) recipient mice, we were able to expand one primary BLT mouse into a colony of 4-5 proBLT (propagated BLT) mice in 6-8 weeks. These proBLT mice reconstituted human immune cells, including T cells, at levels comparable to those of their primary BLT donor mouse. They also faithfully inherited the human immune cell genetic traits from their donor BLT mouse, such as the HLA-A2 haplotype that is of special interest for studying HLA-A2-restricted human T cell immunotherapies. Moreover, an EGFP reporter gene engineered into the human immune system was stably passed from BLT to proBLT mice, making proBLT mice suitable for studying human immune cell gene therapy. This method provides an opportunity to overcome a critical hurdle to utilizing the BLT humanized mouse model and enables its more widespread use as a valuable preclinical research tool.

A Mouse Model of Visual Perceptual Learning Reveals Alterations in Neuronal Coding and Dendritic Spine Density in the Visual Cortex.

PubMed

Wang, Yan; Wu, Wei; Zhang, Xian; Hu, Xu; Li, Yue; Lou, Shihao; Ma, Xiao; An, Xu; Liu, Hui; Peng, Jing; Ma, Danyi; Zhou, Yifeng; Yang, Yupeng

2016-01-01

Visual perceptual learning (VPL) can improve spatial vision in normally sighted and visually impaired individuals. Although previous studies of humans and large animals have explored the neural basis of VPL, elucidation of the underlying cellular and molecular mechanisms remains a challenge. Owing to the advantages of molecular genetic and optogenetic manipulations, the mouse is a promising model for providing a mechanistic understanding of VPL. Here, we thoroughly evaluated the effects and properties of VPL on spatial vision in C57BL/6J mice using a two-alternative, forced-choice visual water task. Briefly, the mice underwent prolonged training at near the individual threshold of contrast or spatial frequency (SF) for pattern discrimination or visual detection for 35 consecutive days. Following training, the contrast-threshold trained mice showed an 87% improvement in contrast sensitivity (CS) and a 55% gain in visual acuity (VA). Similarly, the SF-threshold trained mice exhibited comparable and long-lasting improvements in VA and significant gains in CS over a wide range of SFs. Furthermore, learning largely transferred across eyes and stimulus orientations. Interestingly, learning could transfer from a pattern discrimination task to a visual detection task, but not vice versa. We validated that this VPL fully restored VA in adult amblyopic mice and old mice. Taken together, these data indicate that mice, as a species, exhibit reliable VPL. Intrinsic signal optical imaging revealed that mice with perceptual training had higher cut-off SFs in primary visual cortex (V1) than those without perceptual training. Moreover, perceptual training induced an increase in the dendritic spine density in layer 2/3 pyramidal neurons of V1. These results indicated functional and structural alterations in V1 during VPL. Overall, our VPL mouse model will provide a platform for investigating the neurobiological basis of VPL.

Sex-reversed somatic cell cloning in the mouse.

PubMed

Inoue, Kimiko; Ogonuki, Narumi; Mekada, Kazuyuki; Yoshiki, Atsushi; Sado, Takashi; Ogura, Atsuo

2009-10-01

Somatic cell nuclear transfer has many potential applications in the fields of basic and applied sciences. However, it has a disadvantage that can never be overcome technically-the inflexibility of the sex of the offspring. Here, we report an accidental birth of a female mouse following nuclear transfer using an immature Sertoli cell. We produced a batch of 27 clones in a nuclear transfer experiment using Sertoli cells collected from neonatal male mice. Among them, one pup was female. This "male-derived female" clone grew into a normal adult and produced offspring by natural mating with a littermate. Chromosomal analysis revealed that the female clone had a 39,X karyotype, indicating that the Y chromosome had been deleted in the donor cell or at some early step during nuclear transfer. This finding suggests the possibility of resuming sexual reproduction after a single male is cloned, which should be especially useful for reviving extinct or endangered species.

Ultrastructural Morphology of Sperm from Human Globozoospermia

PubMed Central

Ricci, Giuseppe; Andolfi, Laura; Luppi, Stefania; Boscolo, Rita; Zweyer, Marina; Trevisan, Elisa

2015-01-01

Globozoospermia is a rare disorder characterized by the presence of sperm with round head, lacking acrosome. Coiling tail around the nucleus has been reported since early human studies, but no specific significance has conferred it. By contrast, studies on animal models suggest that coiling tail around the nucleus could represent a crucial step of defective spermatogenesis, resulting in round-headed sperm. No observations, so far, support the transfer of this hypothesis to human globozoospermia. The purpose of this work was to compare ultrastructural morphology of human and mouse model globozoospermic sperm. Sperm have been investigated by using scanning and transmission electron microscopy. The images that we obtained show significant similarities to those described in GOPC knockout mice, an animal model of globozoospermia. By using this model as reference, we were able to identify the probable steps of the tail coiling process in human globozoospermia. Although we have no evidence that there is the same pathophysiology in man and knocked-out mouse, the similarities between these ultrastructural observations in human and those in the experimental model are very suggestive. This is the first demonstration of the existence of relevant morphological homologies between the tail coiling in animal model and human globozoospermia. PMID:26436098

Coenzyme Q10 instilled as eye drops on the cornea reaches the retina and protects retinal layers from apoptosis in a mouse model of kainate-induced retinal damage.

PubMed

Lulli, Matteo; Witort, Ewa; Papucci, Laura; Torre, Eugenio; Schipani, Christian; Bergamini, Christian; Dal Monte, Massimo; Capaccioli, Sergio

2012-12-17

To evaluate if coenzyme Q10 (CoQ10) can protect retinal ganglion cells (RGCs) from apoptosis and, when instilled as eye drops on the cornea, if it can reach the retina and exert its antiapoptotic activity in this area in a mouse model of kainate (KA)-induced retinal damage. Rat primary or cultured RGCs were subjected to glutamate (50 μM) or chemical hypoxia (Antimycin A, 200 μM) or serum withdrawal (FBS, 0.5%) in the presence or absence of CoQ10 (10 μM). Cell viability was evaluated by light microscopy and fluorescence-activated cell sorting analyses. Apoptosis was evaluated by caspase 3/7 activity and mitochondrion depolarization tetramethylrhodamine ethyl ester analysis. CoQ10 transfer to the retina following its instillation as eye drops on the cornea was quantified by HPLC. Retinal protection by CoQ10 (10 μM) eye drops instilled on the cornea was then evaluated in a mouse model of KA-induced excitotoxic retinal cell apoptosis by cleaved caspase 3 immunohistofluorescence, caspase 3/7 activity assays, and quantification of inhibition of RGC loss. CoQ10 significantly increased viable cells by preventing RGC apoptosis. Furthermore, when topically applied as eye drops to the cornea, it reached the retina, thus substantially increasing local CoQ10 concentration and protecting retinal layers from apoptosis. The ability of CoQ10 eye drops to protect retinal cells from apoptosis in the mouse model of KA-induced retinal damage suggests that topical CoQ10 may be evaluated in designing therapies for treating apoptosis-driven retinopathies.

Preimplantation death of xenomitochondrial mouse embryo harbouring bovine mitochondria

PubMed Central

Kawahara, Manabu; Koyama, Shiori; Iimura, Satomi; Yamazaki, Wataru; Tanaka, Aiko; Kohri, Nanami; Sasaki, Keisuke; Takahashi, Masashi

2015-01-01

Mitochondria, cellular organelles playing essential roles in eukaryotic cell metabolism, are thought to have evolved from bacteria. The organization of mtDNA is remarkably uniform across species, reflecting its vital and conserved role in oxidative phosphorylation (OXPHOS). Our objectives were to evaluate the compatibility of xenogeneic mitochondria in the development of preimplantation embryos in mammals. Mouse embryos harbouring bovine mitochondria (mtB-M embryos) were prepared by the cell-fusion technique employing the haemagglutinating virus of Japan (HVJ). The mtB-M embryos showed developmental delay at embryonic days (E) 3.5 after insemination. Furthermore, none of the mtB-M embryos could implant into the maternal uterus after embryo transfer, whereas control mouse embryos into which mitochondria from another mouse had been transferred developed as well as did non-manipulated embryos. When we performed quantitative PCR (qPCR) of mouse and bovine ND5, we found that the mtB-M embryos contained 8.3% of bovine mitochondria at the blastocyst stage. Thus, contamination with mitochondria from another species induces embryonic lethality prior to implantation into the maternal uterus. The heteroplasmic state of these xenogeneic mitochondria could have detrimental effects on preimplantation development, leading to preservation of species-specific mitochondrial integrity in mammals. PMID:26416548

A modified culture method significantly improves the development of mouse somatic cell nuclear transfer embryos.

PubMed

Dai, Xiangpeng; Hao, Jie; Zhou, Qi

2009-08-01

Many strategies have been established to improve the efficiency of somatic cell nuclear transfer (SCNT), but relatively few focused on improving culture conditions. The effect of different culture media on preimplantation development of mouse nuclear transfer embryos was investigated. A modified sequential media method, named D media (M16/KSOM and CZB-EG/KSOM), was successfully established that significantly improves SCNT embryo development. Our result demonstrated that while lacking any adverse effect on in vivo fertilized embryos, the D media dramatically improves the blastocyst development of SCNT embryos compared with other commonly used media, including KSOM, M16, CZB, and alphaMEM. Specifically, the rate of blastocyst formation was 62.3% for D1 (M16/KSOM) versus 10-30% for the other media. An analysis of media components indicated that removing EDTA and glutamine from the media can be beneficial for early SCNT embryo development. Our results suggest that in vitro culture environment plays an important role in somatic cell reprogramming, and D media represent the most efficient culture method reported to date to support mouse SCNT early embryo development in vitro.

Routine habitat change: a source of unrecognized transient alteration of intestinal microbiota in laboratory mice.

PubMed

Ma, Betty W; Bokulich, Nicholas A; Castillo, Patricia A; Kananurak, Anchasa; Underwood, Mark A; Mills, David A; Bevins, Charles L

2012-01-01

The mammalian intestine harbors a vast, complex and dynamic microbial population, which has profound effects on host nutrition, intestinal function and immune response, as well as influence on physiology outside of the alimentary tract. Imbalance in the composition of the dense colonizing bacterial population can increase susceptibility to various acute and chronic diseases. Valuable insights on the association of the microbiota with disease critically depend on investigation of mouse models. Like in humans, the microbial community in the mouse intestine is relatively stable and resilient, yet can be influenced by environmental factors. An often-overlooked variable in research is basic animal husbandry, which can potentially alter mouse physiology and experimental outcomes. This study examined the effects of common husbandry practices, including food and bedding alterations, as well as facility and cage changes, on the gut microbiota over a short time course of five days using three culture-independent techniques, quantitative PCR, terminal restriction fragment length polymorphism (TRFLP) and next generation sequencing (NGS). This study detected a substantial transient alteration in microbiota after the common practice of a short cross-campus facility transfer, but found no comparable alterations in microbiota within 5 days of switches in common laboratory food or bedding, or following an isolated cage change in mice acclimated to their housing facility. Our results highlight the importance of an acclimation period following even simple transfer of mice between campus facilities, and highlights that occult changes in microbiota should be considered when imposing husbandry variables on laboratory animals.

Impeding Xist expression from the active X chromosome improves mouse somatic cell nuclear transfer.

PubMed

Inoue, Kimiko; Kohda, Takashi; Sugimoto, Michihiko; Sado, Takashi; Ogonuki, Narumi; Matoba, Shogo; Shiura, Hirosuke; Ikeda, Rieko; Mochida, Keiji; Fujii, Takashi; Sawai, Ken; Otte, Arie P; Tian, X Cindy; Yang, Xiangzhong; Ishino, Fumitoshi; Abe, Kuniya; Ogura, Atsuo

2010-10-22

Cloning mammals by means of somatic cell nuclear transfer (SCNT) is highly inefficient because of erroneous reprogramming of the donor genome. Reprogramming errors appear to arise randomly, but the nature of nonrandom, SCNT-specific errors remains elusive. We found that Xist, a noncoding RNA that inactivates one of the two X chromosomes in females, was ectopically expressed from the active X (Xa) chromosome in cloned mouse embryos of both sexes. Deletion of Xist on Xa showed normal global gene expression and resulted in about an eight- to ninefold increase in cloning efficiency. We also identified an Xist-independent mechanism that specifically down-regulated a subset of X-linked genes through somatic-type repressive histone blocks. Thus, we have identified nonrandom reprogramming errors in mouse cloning that can be altered to improve the efficiency of SCNT methods.

Increased Engraftment of Human Short Term Repopulating Hematopoietic Cells in NOD/SCID/IL2rγnull Mice by Lentiviral Expression of NUP98-HOXA10HD

PubMed Central

Zhao, Huifen; Humphries, Keith; Persons, Derek A.

2016-01-01

Techniques to expand human hematopoietic stem cells ex-vivo could be beneficial to the fields of clinical hematopoietic stem cell transplantation and gene therapy targeted at hematopoietic stem cells. NUP98-HOXA10HD is a relatively newly discovered fusion gene that in mouse transplant experiments has been shown to increase numbers of hematopoietic stem cells. We evaluated whether this fusion gene could be used to expand engrafting human primitive CD34+ cells in an immunodeficient mouse model. Gene transfer was achieved using a lentiviral based vector. The engraftment of mobilized peripheral blood human CD34+ cells grown in culture for one week after gene transfer was evaluated 3–4 months after transplant and found to be 2–3 fold higher in the NUP98-HOXA10HD groups as compared to controls. These data suggest an expansive effect at least at the short term human repopulating cell level. Further evaluation in long term repopulating models and investment in a NUP98-HOXA10HD protein seems worthy of consideration. Additionally, the results here provide strong impetus to utilize NUP98-HOXA10HD as a tool to search for underlying genes and pathways involved in hematopoietic stem cell expansion that can be enhanced and have an even more potent expansive effect. PMID:26761813

Maternal exposure to an environmentally relevant dose of triclocarban results in perinatal exposure and potential alterations in offspring development in the mouse model

DOE PAGES

Enright, Heather A.; Falso, Miranda J. S.; Malfatti, Michael A.; ...

2017-08-09

Triclocarban (TCC) is among the top 10 most commonly detected wastewater contaminants in both concentration and frequency. Its presence in water, as well as its propensity to bioaccumulate, has raised numerous questions about potential endocrine and developmental effects. Here in this paper, we investigated whether exposure to an environmentally relevant concentration of TCC could result in transfer from mother to offspring in CD-1 mice during gestation and lactation using accelerator mass spectrometry (AMS). 14C-TCC (100 nM) was administered to dams through drinking water up to gestation day 18, or from birth to post-natal day 10. AMS was used to quantifymore » 14C-concentrations in offspring and dams after exposure. We demonstrated that TCC does effectively transfer from mother to offspring, both trans-placentally and via lactation. TCC-related compounds were detected in the tissues of offspring with significantly higher concentrations in the brain, heart and fat. In addition to transfer from mother to offspring, exposed offspring were heavier in weight than unexposed controls demonstrating an 11% and 8.5% increase in body weight for females and males, respectively. Quantitative real-time polymerase chain reaction (qPCR) was used to examine changes in gene expression in liver and adipose tissue in exposed offspring. qPCR suggested alterations in genes involved in lipid metabolism in exposed female offspring, which was consistent with the observed increased fat pad weights and hepatic triglycerides. This study represents the first report to quantify the transfer of an environmentally relevant concentration of TCC from mother to offspring in the mouse model and evaluate bio-distribution after exposure using AMS. Our findings suggest that early-life exposure to TCC may interfere with lipid metabolism and could have implications for human health.« less

Maternal exposure to an environmentally relevant dose of triclocarban results in perinatal exposure and potential alterations in offspring development in the mouse model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Enright, Heather A.; Falso, Miranda J. S.; Malfatti, Michael A.

Triclocarban (TCC) is among the top 10 most commonly detected wastewater contaminants in both concentration and frequency. Its presence in water, as well as its propensity to bioaccumulate, has raised numerous questions about potential endocrine and developmental effects. Here in this paper, we investigated whether exposure to an environmentally relevant concentration of TCC could result in transfer from mother to offspring in CD-1 mice during gestation and lactation using accelerator mass spectrometry (AMS). 14C-TCC (100 nM) was administered to dams through drinking water up to gestation day 18, or from birth to post-natal day 10. AMS was used to quantifymore » 14C-concentrations in offspring and dams after exposure. We demonstrated that TCC does effectively transfer from mother to offspring, both trans-placentally and via lactation. TCC-related compounds were detected in the tissues of offspring with significantly higher concentrations in the brain, heart and fat. In addition to transfer from mother to offspring, exposed offspring were heavier in weight than unexposed controls demonstrating an 11% and 8.5% increase in body weight for females and males, respectively. Quantitative real-time polymerase chain reaction (qPCR) was used to examine changes in gene expression in liver and adipose tissue in exposed offspring. qPCR suggested alterations in genes involved in lipid metabolism in exposed female offspring, which was consistent with the observed increased fat pad weights and hepatic triglycerides. This study represents the first report to quantify the transfer of an environmentally relevant concentration of TCC from mother to offspring in the mouse model and evaluate bio-distribution after exposure using AMS. Our findings suggest that early-life exposure to TCC may interfere with lipid metabolism and could have implications for human health.« less

Non-Targeted Effects Models Predict Significantly Higher Mars Mission Cancer Risk than Targeted Effects Models

DOE PAGES

Cucinotta, Francis A.; Cacao, Eliedonna

2017-05-12

Cancer risk is an important concern for galactic cosmic ray (GCR) exposures, which consist of a wide-energy range of protons, heavy ions and secondary radiation produced in shielding and tissues. Relative biological effectiveness (RBE) factors for surrogate cancer endpoints in cell culture models and tumor induction in mice vary considerable, including significant variations for different tissues and mouse strains. Many studies suggest non-targeted effects (NTE) occur for low doses of high linear energy transfer (LET) radiation, leading to deviation from the linear dose response model used in radiation protection. Using the mouse Harderian gland tumor experiment, the only extensive data-setmore » for dose response modelling with a variety of particle types (>4), for the first-time a particle track structure model of tumor prevalence is used to investigate the effects of NTEs in predictions of chronic GCR exposure risk. The NTE model led to a predicted risk 2-fold higher compared to a targeted effects model. The scarcity of data with animal models for tissues that dominate human radiation cancer risk, including lung, colon, breast, liver, and stomach, suggest that studies of NTEs in other tissues are urgently needed prior to long-term space missions outside the protection of the Earth’s geomagnetic sphere.« less

Non-Targeted Effects Models Predict Significantly Higher Mars Mission Cancer Risk than Targeted Effects Models

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cucinotta, Francis A.; Cacao, Eliedonna

Cancer risk is an important concern for galactic cosmic ray (GCR) exposures, which consist of a wide-energy range of protons, heavy ions and secondary radiation produced in shielding and tissues. Relative biological effectiveness (RBE) factors for surrogate cancer endpoints in cell culture models and tumor induction in mice vary considerable, including significant variations for different tissues and mouse strains. Many studies suggest non-targeted effects (NTE) occur for low doses of high linear energy transfer (LET) radiation, leading to deviation from the linear dose response model used in radiation protection. Using the mouse Harderian gland tumor experiment, the only extensive data-setmore » for dose response modelling with a variety of particle types (>4), for the first-time a particle track structure model of tumor prevalence is used to investigate the effects of NTEs in predictions of chronic GCR exposure risk. The NTE model led to a predicted risk 2-fold higher compared to a targeted effects model. The scarcity of data with animal models for tissues that dominate human radiation cancer risk, including lung, colon, breast, liver, and stomach, suggest that studies of NTEs in other tissues are urgently needed prior to long-term space missions outside the protection of the Earth’s geomagnetic sphere.« less

Lack of passive transfer of renal tubulointerstitial disease by serum or monoclonal antibody specific for renal tubular antigens in the mouse.

PubMed

Evans, B D; Dilwith, R L; Balaban, S L; Rudofsky, U H

1988-01-01

Mice immunized with rabbit renal basement membranes form autoantibodies to their kidney glomerular and tubular basement membranes (GBM/TBM). Development of renal tubular disease (RTD) consists of deposition of autoantibodies along the GBM/TBM with the inter- and intratubular accumulation of lymphocytes and macrophages and destruction of the TBM. Transfer of this disease in mice with either serum or monoclonal antibodies, however, has been difficult to demonstrate and, therefore, attempts were made to confirm a report that RTD is passively transferred by anti-TBM autoantibodies. Using the revised protocol in this later report, we found that 12 weeks after transfer autoantibodies were deposited along the GBM and/or TBM of the recipients, yet RTD was not observed. Although qualitative and quantitative characteristics of the antibody may play a role in the pathogenesis in the murine model of RTD, we could not obtain evidence to support and confirm this study.

Laser microbeam irradiation and renucleation of mouse eggs. Final progress report, July 1, 1979-December 31, 1983

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, T.P.

1983-01-01

We have combined laser microbeam irradiation of mouse egg nuclei with fusion to donor cell nuclei in order to develop a new procedure for transferring nuclei into mammalian eggs. We have been using virus-treated cells injected into the perivitelline for fusion with egg cells. Binucleate cells inside the zona pellucida were often produced indicating nuclear transfer between cells had occurred. To prevent the formation of such abortive polyploidy, host nuclei were inactivated with a laser microbeam. The subsequent cleavage of the microirradiated eggs has been studied.

Dependence and independence of survival parameters on linear energy transfer in cells and tissues

PubMed Central

Ando, Koichi; Goodhead, Dudley T.

2016-01-01

Carbon-ion radiotherapy has been used to treat more than 9000 cancer patients in the world since 1994. Spreading of the Bragg peak is necessary for carbon-ion radiotherapy, and is designed based on the linear–quadratic model that is commonly used for photon therapy. Our recent analysis using in vitro cell kills and in vivo mouse tissue reaction indicates that radiation quality affects mainly the alpha terms, but much less the beta terms, which raises the question of whether this is true in other biological systems. Survival parameters alpha and beta for 45 in vitro mammalian cell lines were obtained by colony formation after irradiation with carbon ions, fast neutrons and X-rays. Relationships between survival parameters and linear energy transfer (LET) below 100 keV/μm were obtained for 4 mammalian cell lines. Mouse skin reaction and tumor growth delay were measured after fractionated irradiation. The Fe-plot provided survival parameters of the tissue reactions. A clear separation between X-rays and high-LET radiation was observed for alpha values, but not for beta values. Alpha values/terms increased with increasing LET in any cells and tissues studied, while beta did not show a systematic change. We have found a puzzle or contradiction in common interpretations of the linear-quadratic model that causes us to question whether the model is appropriate for interpreting biological effectiveness of high-LET radiation up to 500 keV/μm, probably because of inconsistency in the concept of damage interaction. A repair saturation model proposed here was good enough to fit cell kill efficiency by radiation of wide-ranged LET. A model incorporating damage complexity and repair saturation would be suitable for heavy-ion radiotherapy. PMID:27380803

Significant improvement of mouse cloning technique by treatment with trichostatin A after somatic nuclear transfer.

PubMed

Kishigami, Satoshi; Mizutani, Eiji; Ohta, Hiroshi; Hikichi, Takafusa; Thuan, Nguyen Van; Wakayama, Sayaka; Bui, Hong-Thuy; Wakayama, Teruhiko

2006-02-03

The low success rate of animal cloning by somatic cell nuclear transfer (SCNT) is believed to be associated with epigenetic errors including abnormal DNA hypermethylation. Recently, we elucidated by using round spermatids that, after nuclear transfer, treatment of zygotes with trichostatin A (TSA), an inhibitor of histone deacetylase, can remarkably reduce abnormal DNA hypermethylation depending on the origins of transferred nuclei and their genomic regions [S. Kishigami, N. Van Thuan, T. Hikichi, H. Ohta, S. Wakayama. E. Mizutani, T. Wakayama, Epigenetic abnormalities of the mouse paternal zygotic genome associated with microinsemination of round spermatids, Dev. Biol. (2005) in press]. Here, we found that 5-50 nM TSA-treatment for 10 h following oocyte activation resulted in more efficient in vitro development of somatic cloned embryos to the blastocyst stage from 2- to 5-fold depending on the donor cells including tail tip cells, spleen cells, neural stem cells, and cumulus cells. This TSA-treatment also led to more than 5-fold increase in success rate of mouse cloning from cumulus cells without obvious abnormality but failed to improve ES cloning success. Further, we succeeded in establishment of nuclear transfer-embryonic stem (NT-ES) cells from TSA-treated cloned blastocyst at a rate three times higher than those from untreated cloned blastocysts. Thus, our data indicate that TSA-treatment after SCNT in mice can dramatically improve the practical application of current cloning techniques.

Assignment of Etfdh, Etfb, and Etfa to chromosomes 3, 7, and 13: The mouse homologs of genes respondible for glutaric acidemia type II in human

DOE Office of Scientific and Technical Information (OSTI.GOV)

White, R.A.; Dowler, L.L.; Angeloni, S.V.

Electron transfer flavoprotein (composed of {alpha} and {beta} subunits) is an obligatory electron acceptor for several dehydrogenases and is located in the mitochondrial matrix. Electrons accepted by electron transfer flavo-protein (ETF) are transferred to the main mitochondrial respiratory chain by the way of ETF dehydrogenase (ETFDH). In humans, deficiency of ETF or ETFDH leads to glutaric acidemia type II, an inherited metabolic disorder that can be fatal in its neonatal form and is characterized by severe hypoketotic hypoglycemia and acidosis. We used cDNA probes for the Etfdh, Etfb, and Etfa genes to determine localization of these mouse genes to chromosomesmore » 3, 7, and 13. 18 refs., 3 figs.« less

Retroviral gene transfer of an antisense construct against membrane type 1 matrix metalloproteinase reduces the invasiveness of rheumatoid arthritis synovial fibroblasts.

PubMed

Rutkauskaite, Edita; Volkmer, Dagmar; Shigeyama, Yukio; Schedel, Jörg; Pap, Geza; Müller-Ladner, Ulf; Meinecke, Ingmar; Alexander, Dorothea; Gay, Renate E; Drynda, Susanne; Neumann, Wolfram; Michel, Beat A; Aicher, Wilhelm K; Gay, Steffen; Pap, Thomas

2005-07-01

Membrane type 1 matrix metalloproteinase (MT1-MMP) is expressed prominently in rheumatoid arthritis synovial fibroblasts (RASFs), but the specific contribution of MT1-MMP to fibroblast-mediated destruction of articular cartilage is incompletely understood. This study used gene transfer of an antisense expression construct to assess the effects of MT1-MMP inhibition on the invasiveness of RASFs. Retroviral gene transfer of a pLXIN vector-based antisense RNA expression construct (MT1-MMPalphaS) to MT1-MMP was used to stably transduce RASFs. Levels of MT1-MMP RNA and protein were determined by quantitative polymerase chain reaction, Western blotting, and immunocytochemistry in MT1-MMPalphaS-transduced RASFs as well as in control cells, with monitoring for 60 days. The effects of MT1-MMPalphaS on the invasiveness of RASFs were analyzed in the SCID mouse co-implantation model of RA. MT1-MMPalphaS-transduced RASFs produced high levels of antisense RNA that exceeded endogenous levels of MT1-MMP messenger RNA by 15-fold and resulted in a down-regulation of MT1-MMP at the protein level. Inhibition of MT1-MMP production was maintained for 60 days and significantly reduced the invasiveness of RASFs in the SCID mouse model. Whereas prominent invasion into cartilage by non-transduced and mock-transduced RASFs was observed (mean invasion scores 3.0 and 3.1, respectively), MT1-MMPalphaS-transduced cells showed only moderate invasiveness (mean invasion score 1.8; P < 0.05). The data demonstrate that an antisense RNA expression construct against MT1-MMP can be generated and expressed in RASFs for at least 60 days. Inhibition of MT1-MMP significantly reduces the cartilage degradation by RASFs.

Sub-lethal radiation enhances anti-tumor immunotherapy in a transgenic mouse model of pancreatic cancer

PubMed Central

Cao, Zhu Alexander; Daniel, Dylan; Hanahan, Douglas

2002-01-01

Background It is not uncommon to observe circulating tumor antigen-specific T lymphocytes in cancer patients despite a lack of significant infiltration and destruction of their tumors. Thus, an important goal for tumor immunotherapy is to identify ways to modulate in vivo anti-tumor immunity to achieve clinical efficacy. We investigate this proposition in a spontaneous mouse tumor model, Rip1-Tag2. Methods Experimental therapies were carried out in two distinctive trial designs, intended to either intervene in the explosive growth of small tumors, or regress bulky end-stage tumors. Rip1-Tag2 mice received a single transfer of splenocytes from Tag-specific, CD4+ T cell receptor transgenic mice, a single sub-lethal radiation, or a combination therapy in which the lymphocyte transfer was preceded by the sub-lethal radiation. Tumor burden, the extent of lymphocyte infiltration into solid tumors and host survival were used to assess the efficacy of these therapeutic approaches. Results In either intervention or regression, the transfer of Tag-specific T cells alone did not result in significant lymphocyte infiltration into solid tumors, not did it affect tumor growth or host survival. In contrast, the combination therapy resulted in significant reduction in tumor burden, increase in lymphocyte infiltration into solid tumors, and extension of survival. Conclusions The results indicate that certain types of solid tumors may be intrinsically resistant to infiltration and destruction by tumor-specific T lymphocytes. Our data suggest that such resistance can be disrupted by sub-lethal radiation. The combinatorial approach presented here merits consideration in the design of clinical trials aimed to achieve T cell-mediated anti-tumor immunity. PMID:12019035

A Novel Mouse Model of Endometriosis Mimics Human Phenotype and Reveals Insights into the Inflammatory Contribution of Shed Endometrium

PubMed Central

Greaves, Erin; Cousins, Fiona L.; Murray, Alison; Esnal-Zufiaurre, Arantza; Fassbender, Amelie; Horne, Andrew W.; Saunders, Philippa T.K.

2015-01-01

Endometriosis is an estrogen-dependent inflammatory disorder characterized by the presence of endometrial tissue outside the uterine cavity. Patients experience chronic pelvic pain and infertility, with the most likely origin of the tissue deposits (lesions) being endometrial fragments shed at menses. Menstruation is an inflammatory process associated with a dramatic increase in inflammatory mediators and tissue-resident immune cells. In the present study, we developed and validated a mouse model of endometriosis using syngeneic menstrual endometrial tissue introduced into the peritoneum of immunocompetent mice. We demonstrate the establishment of endometriotic lesions that exhibit similarities to those recovered from patients undergoing laparoscopy. Specifically, in both cases, lesions had epithelial (cytokeratin+) and stromal (vimentin/CD10+) cell compartments with a well-developed vasculature (CD31+ endothelial cells). Expression of estrogen receptor β was increased in lesions compared with the peritoneum or eutopic endometrium. By performing experiments using mice with green fluorescent protein–labeled macrophages (MacGreen) in reciprocal transfers with wild-type mice, we obtained evidence that macrophages present in the peritoneum and in menses endometrium can contribute to the inflammatory microenvironment of the lesions. In summary, we developed a mouse model of endometriosis that exhibits similarities to human peritoneal lesions with respect to estrogen receptor expression, inflammation, and macrophage infiltration, providing an opportunity for further studies and the possible identification of novel therapies for this perplexing disorder. PMID:24910298

Factors affecting the electrofusion of mouse and ferret oocytes with ferret somatic cells.

PubMed

Li, Ziyi; Sun, Xingshen; Chen, Juan; Leno, Gregory H; Engelhardt, John F

2005-09-01

The domestic ferret, Mustela putorius furos, holds great promise as a genetic model for human lung disease, provided that key technologies for somatic cell nuclear transfer (SCNT) are developed. In this report, we extend our understanding of SCNT in this species by defining conditions for efficient cell fusion by electrical pulse. Two experimental systems were employed in this study. First, in vivo-matured mouse oocytes and ferret somatic cells were used to establish general parameters for fusion. One fibroblast, or cumulus cell, was agglutinated to nucleate, zona pellucida-free, mouse oocytes, and subjected to an electrical pulse. Similar electrical pulse conditions were also tested with 1 or 2 somatic cells inserted into the perivitelline space (PVS) of intact mouse oocytes. The fusion rate for a single fibroblast with a zona-free oocyte was 80.2%, significantly higher (P < 0.05) than that observed for 1, or 2, fibroblasts placed in the PVS (52.0% and 63.8%, respectively). The fusion rate (44.1%) following insertion of two cumulus cells was significantly higher (P < 0.05) than that following insertion of one cumulus cell (25.1%). Second, in vitro-matured ferret oocytes were enucleated, and one to three fibroblasts or cumulus cells were inserted into the PVS. Zona pellucida-free ferret oocytes were fragile and excluded from the study. The fusion rates with two or three fibroblasts were 71.4% and 76.8%, respectively; significantly higher (P < 0.05) than that for one fibroblast (48.6%). This cell number-dependent difference in fusion efficiency was also observed with cumulus cells. Fusion-derived (ferret-ferret) NT embryos cleaved, formed blastocysts in vitro, and underwent early-stage fetal development following embryo transfer. The rate of development was cell type-independent, in contrast to the cell type-dependent differences observed in fusion efficiency. In conclusion, fibroblasts fused more efficiently than cumulus cells and the efficiency of single cell fusions was improved when two or more cells were inserted into the PVS. These studies define conditions for efficient cell fusion with ferret oocytes and should facilitate SCNT and the development of genetically defined animal models in this species.

Impaired receptivity and decidualization in DHEA-induced PCOS mice.

PubMed

Li, Shu-Yun; Song, Zhuo; Song, Min-Jie; Qin, Jia-Wen; Zhao, Meng-Long; Yang, Zeng-Ming

2016-12-07

Polycystic ovary syndrome (PCOS), a complex endocrine disorder, is a leading cause of female infertility. An obvious reason for infertility in PCOS women is anovulation. However, success rate with high quality embryos selected by assisted reproduction techniques in PCOS patients still remain low with a high rate of early clinical pregnancy loss, suggesting a problem in uterine receptivity. Using a dehydroepiandrosterone-induced mouse model of PCOS, some potential causes of decreased fertility in PCOS patients were explored. In our study, ovulation problem also causes sterility in PCOS mice. After blastocysts from normal mice are transferred into uterine lumen of pseudopregnant PCOS mice, the rate of embryo implantation was reduced. In PCOS mouse uteri, the implantation-related genes are also dysregulated. Additionally, artificial decidualization is severely impaired in PCOS mice. The serum estrogen level is significantly higher in PCOS mice than vehicle control. The high level of estrogen and potentially impaired LIF-STAT3 pathway may lead to embryo implantation failure in PCOS mice. Although there are many studies about effects of PCOS on endometrium, both embryo transfer and artificial decidualization are applied to exclude the effects from ovulation and embryos in our study.

Impaired receptivity and decidualization in DHEA-induced PCOS mice

PubMed Central

Li, Shu-Yun; Song, Zhuo; Song, Min-Jie; Qin, Jia-Wen; Zhao, Meng-Long; Yang, Zeng-Ming

2016-01-01

Polycystic ovary syndrome (PCOS), a complex endocrine disorder, is a leading cause of female infertility. An obvious reason for infertility in PCOS women is anovulation. However, success rate with high quality embryos selected by assisted reproduction techniques in PCOS patients still remain low with a high rate of early clinical pregnancy loss, suggesting a problem in uterine receptivity. Using a dehydroepiandrosterone-induced mouse model of PCOS, some potential causes of decreased fertility in PCOS patients were explored. In our study, ovulation problem also causes sterility in PCOS mice. After blastocysts from normal mice are transferred into uterine lumen of pseudopregnant PCOS mice, the rate of embryo implantation was reduced. In PCOS mouse uteri, the implantation-related genes are also dysregulated. Additionally, artificial decidualization is severely impaired in PCOS mice. The serum estrogen level is significantly higher in PCOS mice than vehicle control. The high level of estrogen and potentially impaired LIF-STAT3 pathway may lead to embryo implantation failure in PCOS mice. Although there are many studies about effects of PCOS on endometrium, both embryo transfer and artificial decidualization are applied to exclude the effects from ovulation and embryos in our study. PMID:27924832

SU-F-T-668: Irradiating Mouse Brain with a Clinical Linear Accelerator

DOE Office of Scientific and Technical Information (OSTI.GOV)

Perez-Torres, C

Purpose: To design and construct a “mouse jig” device that would allow for irradiation of the mouse brain with a clinical Varian 6 MeV Linear Accelerator. This device must serve as a head immobilizer, gaseous anesthesia delivery, and radiation bolus concurrently. Methods: The mouse jig was machined out of nylon given that it is inexpensive, easy to machine, and has similar electron density to water. A cylindrical opening with diameter of 16 mm and 40 mm depth was drilled into a nylon block sized 56×56×50 mm (width, length, depth). Additional slots were included in the block for ear bars andmore » a tooth bar to serve as a three-point immobilization device as well as for anesthesia delivery and scavenging. For ease of access when loading the mouse into the holder, there is a removable piece at the top of the block that is 15 mm in depth. This serves a dual purpose, as with the proper extra shielding, the mouse jig could be used with lower linear energy transfer photons with this piece removed. A baseplate was then constructed with five square slots where the mouse jig can securely be inserted plus additional slots that would allow the baseplate to be mounted on a standard lock bar in the treatment couch. This maximizes the reproducibility of placement between imaging and treatment and between treatment sessions. Results: CT imaging and radiation treatment planning was performed that showed acceptable coverage and uniformity of radiation dose in the mouse brain while sparing the throat and eyes. Conclusion: We have designed and manufactured a device that fulfills our criteria allowing us to selectively irradiate the mouse brain with a clinical linear accelerator. This setup will be used for generating mouse models of radiation-induced brain injury.« less

Different modes of herpes simplex virus type 1 spread in brain and skin tissues.

PubMed

Tsalenchuck, Yael; Tzur, Tomer; Steiner, Israel; Panet, Amos

2014-02-01

Herpes simplex virus type 1 (HSV-1) initially infects the skin and subsequently spreads to the nervous system. To investigate and compare HSV-1 mode of propagation in the two clinically relevant tissues, we have established ex vivo infection models, using native tissues of mouse and human skin, as well as mouse brain, maintained in organ cultures. HSV-1, which is naturally restricted to the human, infects and spreads in the mouse and human skin tissues in a similar fashion, thus validating the mouse model. The spread of HSV-1 in the skin was concentric to form typical plaques of limited size, predominantly of cytopathic cells. By contrast, HSV-1 spread in the brain tissue was directed along specific neuronal networks with no apparent cytopathic effect. Two additional differences were noted following infection of the skin and brain tissues. First, only a negligible amount of extracellular progeny virus was produced of the infected brain tissues, while substantial quantity of infectious progeny virus was released to the media of the infected skin. Second, antibodies against HSV-1, added following the infection, effectively restricted viral spread in the skin but have no effect on viral spread in the brain tissue. Taken together, these results reveal that HSV-1 spread within the brain tissue mostly by direct transfer from cell to cell, while in the skin the progeny extracellular virus predominates, thus facilitating the infection to new individuals.

Sterol transfer between cyclodextrin and membranes: similar but not identical mechanism to NPC2-mediated cholesterol transfer.

PubMed

McCauliff, Leslie A; Xu, Zhi; Storch, Judith

2011-08-30

Niemann--Pick C disease is an inherited disorder in which cholesterol and other lipids accumulate in the late endosomal/lysosomal compartment. Recently, cyclodextrins (CD) have been shown to reduce symptoms and extend lifespan in animal models of the disease. In the present studies we examined the mechanism of sterol transport by CD using in vitro model systems and fluorescence spectroscopy and NPC2-deficient fibroblasts. We demonstrate that cholesterol transport from the lysosomal cholesterol-binding protein NPC2 to CD occurs via aqueous diffusional transfer and is very slow; the rate-limiting step appears to be dissociation of cholesterol from NPC2, suggesting that specific interactions between NPC2 and CD do not occur. In contrast, the transfer rate of the fluorescent cholesterol analogue dehydroergosterol (DHE) from CD to phospholipid membranes is very rapid and is directly proportional to the acceptor membrane concentration, as is DHE transfer from membranes to CD. Moreover, CD dramatically increases the rate of sterol transfer between membranes, with rates that can approach those mediated by NPC2. The results suggest that sterol transfer from CD to membranes occurs by a collisional transfer mechanism involving direct interaction of CD with membranes, similar to that shown previously for NPC2. For CD, however, absolute rates are slower compared to NPC2 for a given concentration, and the lysosomal phospholipid lysobisphosphatidic acid (LBPA) does not stimulate rates of sterol transfer between membranes and CD. As expected from the apparent absence of interaction between CD and NPC2, the addition of CD to NPC2-deficient fibroblasts rapidly rescued the cholesterol accumulation phenotype. Thus, the recent observations of CD efficacy in mouse models of NPC disease are likely the result of CD enhancement of cholesterol transport between membranes, with rapid sterol transfer occurring during CD--membrane interactions.

GAP JUNCTION COMMUNICATON IN A TRANSFECTED HUMAN CELL LINE: ACTION OF MELATONIN AND MAGNETIC FIELDS

EPA Science Inventory

GAP JUNCTION COMMUNICTION IN TRANSFECTED HUMAN CELL LINE: ACTION OF MELATONIN AND MAGNETIC FIELDS.

OBJECTIVE: We previously showed that functional gap junction communication (GJC), as monitored by dye transfer (DT), could be enhanced in mouse C3H 10T112 cells and in mouse...

Inhibition of class IIb histone deacetylase significantly improves cloning efficiency in mice.

PubMed

Ono, Tetsuo; Li, Chong; Mizutani, Eiji; Terashita, Yukari; Yamagata, Kazuo; Wakayama, Teruhiko

2010-12-01

Since the first mouse clone was produced by somatic cell nuclear transfer, the success rate of cloning in mice has been extremely low. Some histone deacetylase inhibitors, such as trichostatin A and scriptaid, have improved the full-term development of mouse clones significantly, but the mechanisms allowing for this are unclear. Here, we found that two other specific inhibitors, suberoylanilide hydroxamic acid and oxamflatin, could also reduce the rate of apoptosis in blastocysts, improve the full-term development of cloned mice, and increase establishment of nuclear transfer-generated embryonic stem cell lines significantly without leading to obvious abnormalities. However, another inhibitor, valproic acid, could not improve cloning efficiency. Suberoylanilide hydroxamic acid, oxamflatin, trichostatin A, and scriptaid are inhibitors for classes I and IIa/b histone deacetylase, whereas valproic acid is an inhibitor for classes I and IIa, suggesting that inhibiting class IIb histone deacetylase is an important step for reprogramming mouse cloning efficiency.

Diabetes Alters Mechanical Properties and Collagen Fiber Re-Alignment in Multiple Mouse Tendons

PubMed Central

Connizzo, Brianne K.; Bhatt, Pankti R.; Liechty, Kenneth W.; Soslowsky, Louis J.

2014-01-01

Tendons function to transfer load from muscle to bone through their complex composition and hierarchical structure, consisting mainly of type I collagen. Recent evidence suggests that type II diabetes may cause alterations in collagen structure, such as irregular fibril morphology and density, which could play a role in the mechanical function of tendons. Using the db/db mouse model of type II diabetes, the diabetic skin was found to have impaired biomechanical properties when compared to the non-diabetic group. The purpose of this study was to assess the effect of diabetes on biomechanics, collagen fiber re-alignment, and biochemistry in three functionally different tendons (Achilles, supraspinatus, patellar) using the db/db mouse model. Results showed that cross-sectional area and stiffness, but not modulus, were significantly reduced in all three tendons. However, the tendon response to load (transition strain, collagen fiber re-alignment) occurred earlier in the mechanical test, contrary to expectations. In addition, the patellar tendon had an altered response to diabetes when compared to the other two tendons, with no changes in fiber realignment and decreased collagen content at the midsubstance of the tendon. Overall, type II diabetes alters tendon mechanical properties and the dynamic response to load. PMID:24833253

A mouse model of fragile X syndrome exhibits heightened arousal and/or emotion following errors or reversal of contingencies.

PubMed

Moon, J; Ota, K T; Driscoll, L L; Levitsky, D A; Strupp, B J

2008-07-01

This study was designed to further assess cognitive and affective functioning in a mouse model of Fragile X syndrome (FXS), the Fmr1(tm1Cgr) or Fmr1 "knockout" (KO) mouse. Male KO mice and wild-type littermate controls were tested on learning set and reversal learning tasks. The KO mice were not impaired in associative learning, transfer of learning, or reversal learning, based on measures of learning rate. Analyses of videotapes of the reversal learning task revealed that both groups of mice exhibited higher levels of activity and wall-climbing during the initial sessions of the task than during the final sessions, a pattern also seen for trials following an error relative to those following a correct response. Notably, the increase in both behavioral measures seen early in the task was significantly more pronounced for the KO mice than for controls, as was the error-induced increase in activity level. This pattern of effects suggests that the KO mice reacted more strongly than controls to the reversal of contingencies and pronounced drop in reinforcement rate, and to errors in general. This pattern of effects is consistent with the heightened emotional reactivity frequently described for humans with FXS. (c) 2008 Wiley Periodicals, Inc.

Outstanding animal studies in allergy II. From atopic barrier and microbiome to allergen-specific immunotherapy.

PubMed

Jensen-Jarolim, Erika; Pali-Schöll, Isabella; Roth-Walter, Franziska

2017-06-01

Animal studies published within the past 18 months were assessed, focusing on innate and specific immunomodulation, providing knowledge of high translational relevance for human atopic and allergic diseases. Allergic companion animals represent alternative models, but most studies were done in mice. Atopic dermatitis mouse models were refined by the utilization of cytokines like IL-23 and relevant skin allergens or enzymes. A novel IL-6 reporter mouse allows biomonitoring of inflammation. Both skin pH and the (transferable) microflora have a pivotal role in modulating the skin barrier. The microflora of the gastrointestinal mucosa maintains tolerance to dietary compounds and can be disturbed by antiacid drugs. A key mouse study evidenced that dust from Amish households, but not from Hutterites protected mice against asthma. In studies on subcutaneous and sublingual allergen-specific immunotherapy, much focus was given on delivery and adjuvants, using poly-lacto-co-glycolic particles, CpGs, probiotics or Vitamin D3. The epicutaneous and intralymphatic routes showed promising results in mice and horses in terms of prophylactic and therapeutic allergy treatment. In atopic dermatitis, food allergies and asthma, environmental factors, together with the resident microflora and barrier status, decide on sensitization versus tolerance. Also allergen-specific immunotherapy operates with immunomodulatory principles.

Outstanding animal studies in allergy II. From atopic barrier and microbiome to allergen-specific immunotherapy

PubMed Central

Jensen-Jarolim, Erika; Pali-Schöll, Isabella; Roth-Walter, Franziska

2017-01-01

Purpose of review Animal studies published within the past 18 months were assessed, focusing on innate and specific immunomodulation, providing knowledge of high translational relevance for human atopic and allergic diseases. Recent findings Allergic companion animals represent alternative models, but most studies were done in mice. Atopic dermatitis mouse models were refined by the utilization of cytokines like IL-23 and relevant skin allergens or enzymes. A novel IL-6 reporter mouse allows biomonitoring of inflammation. Both skin pH and the (transferable) microflora have a pivotal role in modulating the skin barrier. The microflora of the gastrointestinal mucosa maintains tolerance to dietary compounds and can be disturbed by antiacid drugs. A key mouse study evidenced that dust from Amish households, but not from Hutterites protected mice against asthma. In studies on subcutaneous and sublingual allergen-specific immunotherapy, much focus was given on delivery and adjuvants, using poly-lacto-co-glycolic particles, CpGs, probiotics or Vitamin D3. The epicutaneous and intralymphatic routes showed promising results in mice and horses in terms of prophylactic and therapeutic allergy treatment. Summary In atopic dermatitis, food allergies and asthma, environmental factors, together with the resident microflora and barrier status, decide on sensitization versus tolerance. Also allergen-specific immunotherapy operates with immunomodulatory principles. PMID:28375932

Therapeutic cloning in the mouse

PubMed Central

Mombaerts, Peter

2003-01-01

Nuclear transfer technology can be applied to produce autologous differentiated cells for therapeutic purposes, a concept termed therapeutic cloning. Countless articles have been published on the ethics and politics of human therapeutic cloning, reflecting the high expectations from this new opportunity for rejuvenation of the aging or diseased body. Yet the research literature on therapeutic cloning, strictly speaking, is comprised of only four articles, all in the mouse. The efficiency of derivation of embryonic stem cell lines via nuclear transfer is remarkably consistent among these reports. However, the efficiency is so low that, in its present form, the concept is unlikely to become widespread in clinical practice. PMID:12949262

Engraftment of Human Pluripotent Stem Cell-derived Progenitors in the Inner Ear of Prenatal Mice.

PubMed

Takeda, Hiroki; Hosoya, Makoto; Fujioka, Masato; Saegusa, Chika; Saeki, Tsubasa; Miwa, Toru; Okano, Hideyuki; Minoda, Ryosei

2018-01-31

There is, at present, no curative treatment for genetic hearing loss. We have previously reported that transuterine gene transfer of wild type CONNEXIN30 (CX30) genes into otocysts in CX30-deleted mice could restore hearing. Cell transplantation therapy might be another therapeutic option, although it is still unknown whether stem cell-derived progenitor cells could migrate into mouse otocysts. Here, we show successful cell transplantation of progenitors of outer sulcus cell-like cells derived from human-derived induced pluripotent stem cells into mouse otocysts on embryonic day 11.5. The delivered cells engrafted more frequently in the non-sensory region in the inner ear of CX30-deleted mice than in wild type mice and survived for up to 1 week after transplantation. Some of the engrafted cells expressed CX30 proteins in the non-sensory region. This is the first report that demonstrates successful engraftment of exogenous cells in prenatal developing otocysts in mice. Future studies using this mouse otocystic injection model in vivo will provide further clues for developing treatment modalities for congenital hearing loss in humans.

Intestinal bacteria modify lymphoma incidence and latency by affecting systemic inflammatory state, oxidative stress, and leucocyte genotoxicity

PubMed Central

Yamamoto, Mitsuko L.; Maier, Irene; Dang, Angeline Tilly; Berry, David; Liu, Jared; Ruegger, Paul M.; Yang, Jiue-in; Soto, Phillip A.; Presley, Laura L.; Reliene, Ramune; Westbrook, Aya M.; Wei, Bo; Loy, Alexander; Chang, Christopher; Braun, Jonathan; Borneman, James; Schiestl, Robert H.

2013-01-01

Ataxia-telangiectasia (A-T) is a genetic disorder associated with high incidence of B cell lymphoma. Using an A-T mouse model, we compared lymphoma incidence in several isogenic mouse colonies harboring different bacterial communities, finding that intestinal microbiota are a major contributor to disease penetrance and latency, lifespan, molecular oxidative stress and systemic leucocyte genotoxicity. High throughput sequence analysis of rRNA genes identified mucosa-associated bacterial phylotypes that were colony-specific. Lactobacillus johnsonii, which was deficient in the more cancer-prone mouse colony, was causally tested for its capacity to confer reduced genotoxicity when restored by short-term oral transfer. This intervention decreased systemic genotoxicity, a response associated with reduced basal leucocytes and the cytokine-mediated inflammatory state, and mechanistically linked to the host cell biology of systemic genotoxicity. Our results suggest that intestinal microbiota are a potentially modifiable trait for translational intervention in individuals at risk for B cell lymphoma, or for other diseases that are driven by genotoxicity or the molecular response to oxidative stress. PMID:23860718

Immune modulation using transdermal photodynamic therapy

NASA Astrophysics Data System (ADS)

Levy, Julia G.; Chowdhary, R. K.; Ratkay, Leslie G.; Waterfield, Douglas; Obochi, Modestus; Leong, Simon; Hunt, David W. C.; Chan, Agnes H.

1995-01-01

The photosensitizer benzoporphyrin derivative monoacid ring A (VerteporfinR or BPD) has maximum absorption characteristics (690 nm) and biodistribution characteristics which permit activation of the drug in capillaries of the skin without causing skin photosensitivity (transdermal PDT). This permits targeting of cells in the circulation for selective ablation. Since BPD has been shown to accumulate preferentially in activated lymphocytes and monocytes, studies have been undertaken to determine the effect of transdermal PDT on murine models for rheumatoid arthritis (the MRL/lpr adjuvant enhanced model) and multiple sclerosis (the experimental allergic encephalomyelitis (EAE) model in PL mice). Localized transdermal PDT with BPD was found to be completely successful in preventing the development of adjuvant enhanced arthritis in the MRL/lpr mouse as well as improving the underlying arthritic condition of these animals. In the EAE model, in which an adoptive transfer system was used, it was found that transdermal PDT of recipients was effective in preventing EAE if treatments were implemented up to 24 hours after cell transfer but was not effective if given later, indicating the requirement for circulating T cells for effective treatment.

Routine Habitat Change: A Source of Unrecognized Transient Alteration of Intestinal Microbiota in Laboratory Mice

PubMed Central

Ma, Betty W.; Bokulich, Nicholas A.; Castillo, Patricia A.; Kananurak, Anchasa; Underwood, Mark A.; Mills, David A.; Bevins, Charles L.

2012-01-01

The mammalian intestine harbors a vast, complex and dynamic microbial population, which has profound effects on host nutrition, intestinal function and immune response, as well as influence on physiology outside of the alimentary tract. Imbalance in the composition of the dense colonizing bacterial population can increase susceptibility to various acute and chronic diseases. Valuable insights on the association of the microbiota with disease critically depend on investigation of mouse models. Like in humans, the microbial community in the mouse intestine is relatively stable and resilient, yet can be influenced by environmental factors. An often-overlooked variable in research is basic animal husbandry, which can potentially alter mouse physiology and experimental outcomes. This study examined the effects of common husbandry practices, including food and bedding alterations, as well as facility and cage changes, on the gut microbiota over a short time course of five days using three culture-independent techniques, quantitative PCR, terminal restriction fragment length polymorphism (TRFLP) and next generation sequencing (NGS). This study detected a substantial transient alteration in microbiota after the common practice of a short cross-campus facility transfer, but found no comparable alterations in microbiota within 5 days of switches in common laboratory food or bedding, or following an isolated cage change in mice acclimated to their housing facility. Our results highlight the importance of an acclimation period following even simple transfer of mice between campus facilities, and highlights that occult changes in microbiota should be considered when imposing husbandry variables on laboratory animals. PMID:23082164

Targeting Fibroblast Activation Protein in Tumor Stroma with Chimeric Antigen Receptor T Cells Can Inhibit Tumor Growth and Augment Host Immunity Without Severe Toxicity

PubMed Central

Wang, Liang-Chuan S; Lo, Albert; Scholler, John; Sun, Jing; Majumdar, Rajrupa S; Kapoor, Veena; Antzis, Michael; Cotner, Cody E.; Johnson, Laura A; Durham, Amy C; Solomides, Charalambos C.; June, Carl H; Puré, Ellen; Albelda, Steven M

2013-01-01

The majority of chimeric antigen receptor (CAR) T cell research has focused on attacking cancer cells. Here we show that targeting the tumor-promoting, non-transformed stromal cells using CAR T cells may offer several advantages. We developed a retroviral CAR construct specific for the mouse fibroblast activation protein (FAP), comprising a single chain Fv FAP (mAb 73.3) with the CD8α hinge and transmembrane regions, and the human CD3ζ and 4-1BB activation domains. The transduced muFAP-CAR mouse T cells secreted IFNγ and killed FAP-expressing 3T3 target cells specifically. Adoptively transferred 73.3-FAP-CAR mouse T cells selectively reduced FAPhi stromal cells and inhibited the growth of multiple types of subcutaneously transplanted tumors in wild-type, but not FAP-null immune-competent syngeneic mice. The antitumor effects could be augmented by multiple injections of the CAR T cells, by using CAR T cells with a deficiency in diacylglycerol kinase, or by combination with a vaccine. A major mechanism of action of the muFAP-CAR T cells was the augmentation of the endogenous CD8+ T cell antitumor responses. Off-tumor toxicity in our models was minimal following muFAP-CAR T cell therapy. In summary, inhibiting tumor growth by targeting tumor stroma with adoptively transferred CAR T cells directed to FAP can be safe and effective suggesting that further clinical development of anti-human FAP-CAR is warranted. PMID:24778279

Detection of Subclinical Arthritis in Mice by a Thrombin Receptor-Derived Imaging Agent.

PubMed

Friedman, Beth; Whitney, Michael A; Savariar, Elamprakash N; Caneda, Christa; Steinbach, Paul; Xiong, Qing; Hingorani, Dina V; Crisp, Jessica; Adams, Stephen R; Kenner, Michael; Lippert, Csilla N; Nguyen, Quyen T; Guma, Monica; Tsien, Roger Y; Corr, Maripat

2018-01-01

Functional imaging of synovitis could improve both early detection of rheumatoid arthritis (RA) and long-term outcomes. Given the intersection of inflammation with coagulation protease activation, this study was undertaken to examine coagulation protease activities in arthritic mice with a dual-fluorescence ratiometric activatable cell-penetrating peptide (RACPP) that has a linker, norleucine (Nle)-TPRSFL, with a cleavage site for thrombin. K/BxN-transgenic mice with chronic arthritis and mice with day 1 passive serum-transfer arthritis were imaged in vivo for Cy5:Cy7 emission ratiometric fluorescence from proteolytic cleavage and activation of RACPP NleTPRSFL . Joint thickness in mice with serum-transfer arthritis was measured from days 0 to 10. The cleavage-evoked release of Cy5-tagged tissue-adhesive fragments enabled microscopic correlation with immunohistochemistry for inflammatory markers. Thrombin dependence of ratiometric fluorescence was tested by ex vivo application of RACPP NleTPRSFL and argatroban to cryosections obtained from mouse hind paws on day 1 of serum-transfer arthritis. In chronic arthritis, RACPP NleTPRSFL fluorescence ratios of Cy5:Cy7 emission were significantly higher in diseased swollen ankles of K/BxN-transgenic mice than in normal mouse ankles. A high ratio of RACPP NleTPRSFL fluorescence in mouse ankles and toes on day 1 of serum-transfer arthritis correlated with subsequent joint swelling. Foci of high ratiometric fluorescence localized to inflammation, as demarcated by immune reactivity for citrullinated histones, macrophages, mast cells, and neutrophils, in soft tissue on day 1 of serum-transfer arthritis. Ex vivo application of RACPP NleTPRSFL to cryosections obtained from mice on day 1 of serum-transfer arthritis produced ratiometric fluorescence that was inhibited by argatroban. RACPP NleTPRSFL activation detects established experimental arthritis, and the detection of inflammation by RACPP NleTPRSFL on day 1 of serum-transfer arthritis correlates with disease progression. © 2017, American College of Rheumatology.

Comparative biology of cystic fibrosis animal models.

PubMed

Fisher, John T; Zhang, Yulong; Engelhardt, John F

2011-01-01

Animal models of human diseases are critical for dissecting mechanisms of pathophysiology and developing therapies. In the context of cystic fibrosis (CF), mouse models have been the dominant species by which to study CF disease processes in vivo for the past two decades. Although much has been learned through these CF mouse models, limitations in the ability of this species to recapitulate spontaneous lung disease and several other organ abnormalities seen in CF humans have created a need for additional species on which to study CF. To this end, pig and ferret CF models have been generated by somatic cell nuclear transfer and are currently being characterized. These new larger animal models have phenotypes that appear to closely resemble human CF disease seen in newborns, and efforts to characterize their adult phenotypes are ongoing. This chapter will review current knowledge about comparative lung cell biology and cystic fibrosis transmembrane conductance regulator (CFTR) biology among mice, pigs, and ferrets that has implications for CF disease modeling in these species. We will focus on methods used to compare the biology and function of CFTR between these species and their relevance to phenotypes seen in the animal models. These cross-species comparisons and the development of both the pig and the ferret CF models may help elucidate pathophysiologic mechanisms of CF lung disease and lead to new therapeutic approaches.

Calcitriol Treatment Ameliorates Inflammation and Blistering in Mouse Models of Epidermolysis Bullosa Acquisita.

PubMed

Tukaj, Stefan; Bieber, Katja; Witte, Mareike; Ghorbanalipoor, Saeedeh; Schmidt, Enno; Zillikens, Detlef; Ludwig, Ralf J; Kasperkiewicz, Michael

2018-02-01

A link between hypovitaminosis D and development of autoimmune bullous disorders has been suggested recently, but this association has not been elaborated experimentally. Here, the role of vitamin D was investigated in epidermolysis bullosa acquisita (EBA), an anti-type VII collagen autoantibody-induced blistering skin disease. Oral administration of the hormonally active vitamin D metabolite calcitriol ameliorated clinical disease severity and dermal neutrophil infiltration in both an antibody transfer- and immunization-induced EBA mouse model. Mechanistically, calcitriol hindered immune effector cell activation as evidenced by increased L-selectin expression on Gr-1 + cells in calcitriol-treated mice with antibody transfer-induced EBA, as well as suppressed in vitro immune complex-induced reactive oxygen species production in calcitriol-treated murine neutrophils. Additionally, calcitriol administration was associated with an increase of regulatory T (CD4 + FoxP3 + ) and B (CD19 + IL10 + ) cells as well as reduction of pro-inflammatory T helper 17 (CD4 + IL-17 + ) cells in mice with immunization-induced EBA. In line, levels of circulating anti-type VII collagen autoantibodies were lower in mice that received calcitriol compared to solvent-treated animals. Together with the observed state of hypovitaminosis D in most cases of an analyzed EBA patient cohort, the results of this study support the use of vitamin D derivatives or analogs for patients with EBA and related diseases. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Impairment of extramitochondrial oxidative phosphorylation in mouse rod outer segments by blue light irradiation.

PubMed

Calzia, Daniela; Panfoli, Isabella; Heinig, Nora; Schumann, Ulrike; Ader, Marius; Traverso, Carlo Enrico; Funk, Richard H W; Roehlecke, Cora

2016-06-01

Exposure to short wavelength light causes increased reactive oxygen intermediates production in the outer retina, particularly in the rod Outer Segments (OS). Consistently, the OS were shown to conduct aerobic ATP production through the ectopic expression of the electron transfer chain complexes I-IV and F1Fo-ATP synthase. These facts prompted us to verify if the oxidative phosphorylation in the OS is implied in the oxidative damage of the blue-light (BL) treated OS, in an organotypic model of mouse retina. Whole mouse eyeball cultures were treated with short wavelength BL (peak at 405 nm, output power 1 mW/cm(2)) for 6 h. Immunogold transmission electron microscopy confirmed the expression of Complex I and F1Fo-ATP synthase in the OS. In situ histochemical assays on unfixed sections showed impairment of respiratory Complexes I and II after BL exposure, both in the OS and IS, utilized as a control. Basal O2 consumption and ATP synthesis were impaired in the OS purified from blue-light irradiated eyeball cultures. Electron transfer capacity between Complex I and II as well as activity of Complexes I and II was decreased in blue-light irradiated purified OS. The severe malfunctioning of the OS aerobic respiratory capacity after 6 h BL treatment may be the consequence of a self-induced damage. BL exposure would cause an initial over-functioning of both the phototransduction and respiratory chain, with reactive oxygen species production. In a self-renewal vicious cycle, membrane and protein oxidative damage, proton leakage and uncoupling, would impair redox chains, perpetuating the damage and causing hypo-metabolism with eventual apoptosis of the rod. Data may shed new light on the rod-driven retinopathies such as Age Related Macular Degeneration, of which blue-light irradiated retina represents a model. Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

A novel mouse model of endometriosis mimics human phenotype and reveals insights into the inflammatory contribution of shed endometrium.

PubMed

Greaves, Erin; Cousins, Fiona L; Murray, Alison; Esnal-Zufiaurre, Arantza; Fassbender, Amelie; Horne, Andrew W; Saunders, Philippa T K

2014-07-01

Endometriosis is an estrogen-dependent inflammatory disorder characterized by the presence of endometrial tissue outside the uterine cavity. Patients experience chronic pelvic pain and infertility, with the most likely origin of the tissue deposits (lesions) being endometrial fragments shed at menses. Menstruation is an inflammatory process associated with a dramatic increase in inflammatory mediators and tissue-resident immune cells. In the present study, we developed and validated a mouse model of endometriosis using syngeneic menstrual endometrial tissue introduced into the peritoneum of immunocompetent mice. We demonstrate the establishment of endometriotic lesions that exhibit similarities to those recovered from patients undergoing laparoscopy. Specifically, in both cases, lesions had epithelial (cytokeratin(+)) and stromal (vimentin/CD10(+)) cell compartments with a well-developed vasculature (CD31(+) endothelial cells). Expression of estrogen receptor β was increased in lesions compared with the peritoneum or eutopic endometrium. By performing experiments using mice with green fluorescent protein-labeled macrophages (MacGreen) in reciprocal transfers with wild-type mice, we obtained evidence that macrophages present in the peritoneum and in menses endometrium can contribute to the inflammatory microenvironment of the lesions. In summary, we developed a mouse model of endometriosis that exhibits similarities to human peritoneal lesions with respect to estrogen receptor expression, inflammation, and macrophage infiltration, providing an opportunity for further studies and the possible identification of novel therapies for this perplexing disorder. Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

A fully implantable pacemaker for the mouse: from battery to wireless power.

PubMed

Laughner, Jacob I; Marrus, Scott B; Zellmer, Erik R; Weinheimer, Carla J; MacEwan, Matthew R; Cui, Sophia X; Nerbonne, Jeanne M; Efimov, Igor R

2013-01-01

Animal models have become a popular platform for the investigation of the molecular and systemic mechanisms of pathological cardiovascular physiology. Chronic pacing studies with implantable pacemakers in large animals have led to useful models of heart failure and atrial fibrillation. Unfortunately, molecular and genetic studies in these large animal models are often prohibitively expensive or not available. Conversely, the mouse is an excellent species for studying molecular mechanisms of cardiovascular disease through genetic engineering. However, the large size of available pacemakers does not lend itself to chronic pacing in mice. Here, we present the design for a novel, fully implantable wireless-powered pacemaker for mice capable of long-term (>30 days) pacing. This design is compared to a traditional battery-powered pacemaker to demonstrate critical advantages achieved through wireless inductive power transfer and control. Battery-powered and wireless-powered pacemakers were fabricated from standard electronic components in our laboratory. Mice (n = 24) were implanted with endocardial, battery-powered devices (n = 14) and epicardial, wireless-powered devices (n = 10). Wireless-powered devices were associated with reduced implant mortality and more reliable device function compared to battery-powered devices. Eight of 14 (57.1%) mice implanted with battery-powered pacemakers died following device implantation compared to 1 of 10 (10%) mice implanted with wireless-powered pacemakers. Moreover, device function was achieved for 30 days with the wireless-powered device compared to 6 days with the battery-powered device. The wireless-powered pacemaker system presented herein will allow electrophysiology studies in numerous genetically engineered mouse models as well as rapid pacing-induced heart failure and atrial arrhythmia in mice.

Recent advancements in cloning by somatic cell nuclear transfer.

PubMed

Ogura, Atsuo; Inoue, Kimiko; Wakayama, Teruhiko

2013-01-05

Somatic cell nuclear transfer (SCNT) cloning is the sole reproductive engineering technology that endows the somatic cell genome with totipotency. Since the first report on the birth of a cloned sheep from adult somatic cells in 1997, many technical improvements in SCNT have been made by using different epigenetic approaches, including enhancement of the levels of histone acetylation in the chromatin of the reconstructed embryos. Although it will take a considerable time before we fully understand the nature of genomic programming and totipotency, we may expect that somatic cell cloning technology will soon become broadly applicable to practical purposes, including medicine, pharmaceutical manufacturing and agriculture. Here we review recent progress in somatic cell cloning, with a special emphasis on epigenetic studies using the laboratory mouse as a model.

Recent advancements in cloning by somatic cell nuclear transfer

PubMed Central

Ogura, Atsuo; Inoue, Kimiko; Wakayama, Teruhiko

2013-01-01

Somatic cell nuclear transfer (SCNT) cloning is the sole reproductive engineering technology that endows the somatic cell genome with totipotency. Since the first report on the birth of a cloned sheep from adult somatic cells in 1997, many technical improvements in SCNT have been made by using different epigenetic approaches, including enhancement of the levels of histone acetylation in the chromatin of the reconstructed embryos. Although it will take a considerable time before we fully understand the nature of genomic programming and totipotency, we may expect that somatic cell cloning technology will soon become broadly applicable to practical purposes, including medicine, pharmaceutical manufacturing and agriculture. Here we review recent progress in somatic cell cloning, with a special emphasis on epigenetic studies using the laboratory mouse as a model. PMID:23166393

In vivo and in vitro gene transfer to mammalian somatic cells by particle bombardment.

PubMed Central

Yang, N S; Burkholder, J; Roberts, B; Martinell, B; McCabe, D

1990-01-01

Chimeric chloramphenicol acetyltransferase and beta-galactosidase marker genes were coated onto fine gold particles and used to bombard a variety of mammalian tissues and cells. Transient expression of the genes was obtained in liver, skin, and muscle tissues of rat and mouse bombarded in vivo. Similar results were obtained with freshly isolated ductal segments of rat and human mammary glands and primary cultures derived from these explants. Gene transfer and transient expression were also observed in eight human cell culture lines, including cells of epithelial, endothelial, fibroblast, and lymphocyte origin. Using CHO and MCF-7 cell cultures as models, we obtained stable gene transfer at frequencies of 1.7 x 10(-3) and 6 x 10(-4), respectively. The particle bombardment technology thus provides a useful means to transfer foreign genes into a variety of mammalian somatic cell systems. The method is applicable to tissues in vivo as well as to isolated cells in culture and has proven effective with all cell or tissue types tested thus far. This technology may therefore prove to be applicable in various aspects of gene therapy. Images PMID:2175906

Mammalian diversity: gametes, embryos and reproduction.

PubMed

Behringer, Richard R; Eakin, Guy S; Renfree, Marilyn B

2006-01-01

The class Mammalia is composed of approximately 4800 extant species. These mammalian species are divided into three subclasses that include the monotremes, marsupials and eutherians. Monotremes are remarkable because these mammals are born from eggs laid outside of the mother's body. Marsupial mammals have relatively short gestation periods and give birth to highly altricial young that continue a significant amount of 'fetal' development after birth, supported by a highly sophisticated lactation. Less than 10% of mammalian species are monotremes or marsupials, so the great majority of mammals are grouped into the subclass Eutheria, including mouse and human. Mammals exhibit great variety in morphology, physiology and reproduction. In the present article, we highlight some of this remarkable diversity relative to the mouse, one of the most widely used mammalian model organisms, and human. This diversity creates challenges and opportunities for gamete and embryo collection, culture and transfer technologies.

T cell–derived interleukin (IL)-21 promotes brain injury following stroke in mice

PubMed Central

Clarkson, Benjamin D.S.; Ling, Changying; Shi, Yejie; Harris, Melissa G.; Rayasam, Aditya; Sun, Dandan; Salamat, M. Shahriar; Kuchroo, Vijay; Lambris, John D.; Sandor, Matyas

2014-01-01

T lymphocytes are key contributors to the acute phase of cerebral ischemia reperfusion injury, but the relevant T cell–derived mediators of tissue injury remain unknown. Using a mouse model of transient focal brain ischemia, we report that IL-21 is highly up-regulated in the injured mouse brain after cerebral ischemia. IL-21–deficient mice have smaller infarcts, improved neurological function, and reduced lymphocyte accumulation in the brain within 24 h of reperfusion. Intracellular cytokine staining and adoptive transfer experiments revealed that brain-infiltrating CD4+ T cells are the predominant IL-21 source. Mice treated with decoy IL-21 receptor Fc fusion protein are protected from reperfusion injury. In postmortem human brain tissue, IL-21 localized to perivascular CD4+ T cells in the area surrounding acute stroke lesions, suggesting that IL-21–mediated brain injury may be relevant to human stroke. PMID:24616379

Cancer immunotherapy and immunological memory.

PubMed

Murata, Kenji; Tsukahara, Tomohide; Torigoe, Toshihiko

2016-01-01

Human immunological memory is the key distinguishing hallmark of the adaptive immune system and plays an important role in the prevention of morbidity and the severity of infection. The differentiation system of T cell memory has been clarified using mouse models. However, the human T cell memory system has great diversity induced by natural antigens derived from many pathogens and tumor cells throughout life, and profoundly differs from the mouse memory system constructed using artificial antigens and transgenic T cells. We believe that only human studies can elucidate the human immune system. The importance of immunological memory in cancer immunotherapy has been pointed out, and the trafficking properties and long-lasting anti-tumor capacity of memory T cells play a crucial role in the control of malignant tumors. Adoptive cell transfer of less differentiated T cells has consistently demonstrated superior anti-tumor capacity relative to more differentiated T cells. Therefore, a human T cell population with the characteristics of stem cell memory is thought to be attractive for peptide vaccination and adoptive cell transfer. A novel human memory T cell population that we have identified is closer to the naive state than previous memory T cells in the T cell differentiation lineage, and has the characteristics of stem-like chemoresistance. Here we introduce this novel population and describe the fundamentals of immunological memory in cancer immunotherapy.

Mouse strain and injection site are crucial for detecting linked suppression in transplant recipients by trans-vivo DTH assay.

PubMed

Burlingham, W J; Jankowska-Gan, E

2007-02-01

Chemokine-driven accumulation of lymphocytes, mononuclear and polymorphonuclear proinflammatory cells in antigenic tissue sites is a key feature of several types of T-cell-dependent autoimmunity and transplant rejection pathology. It is now clear that the immune system expends considerable energy to control this process, exemplified by the sequential layers of regulatory cell input, both innate and adaptive, designed to prevent a classical Type IV or 'delayed-type' hypersensitivity (DTH) reaction from occurring in the visual field of the eye. Yet, despite an abundance of in vitro assays currently available to the human T-cell immunologist, none of them adequately models the human DTH response and its various control features. The theme of this article is that it is relatively easy to model the effector side of the human DTH response with xenogeneic adoptive transfer models. However, we show that in order to detect inhibition of a recall DTH in response to colocalized donor antigen (linked suppression)--a characteristic feature of peripheral tolerance to an organ transplant--both the challenge site and the immunocompetence of the mouse adoptive host are critical factors limiting the sensitivity of the trans-vivo DTH test.

Recapitulating X-Linked Juvenile Retinoschisis in Mouse Model by Knock-In Patient-Specific Novel Mutation.

PubMed

Chen, Ding; Xu, Tao; Tu, Mengjun; Xu, Jinlin; Zhou, Chenchen; Cheng, Lulu; Yang, Ruqing; Yang, Tanchu; Zheng, Weiwei; He, Xiubin; Deng, Ruzhi; Ge, Xianglian; Li, Jin; Song, Zongming; Zhao, Junzhao; Gu, Feng

2017-01-01

X-linked juvenile retinoschisis (XLRS) is a retinal disease caused by mutations in the gene encoding retinoschisin (RS1), which leads to a significant proportion of visual impairment and blindness. To develop personalized genome editing based gene therapy, knock-in animal disease models that have the exact mutation identified in the patients is extremely crucial, and that the way which genome editing in knock-in animals could be easily transferred to the patients. Here we recruited a family diagnosed with XLRS and identified the causative mutation ( RS1 , p.Y65X), then a knock-in mouse model harboring this disease-causative mutation was generated via TALEN (transcription activator-like effector nucleases). We found that the b-wave amplitude of the ERG of the RS1 -KI mice was significantly decreased. Moreover, we observed that the structure of retina in RS1 -KI mice has become disordered, including the disarray of inner nuclear layer and outer nuclear layer, chaos of outer plexiform layer, decreased inner segments of photoreceptor and the loss of outer segments. The novel knock-in mice ( RS1 -KI) harboring patient-specific mutation will be valuable for development of treatment via genome editing mediated gene correction.

Estimation of placental and lactational transfer and tissue distribution of atrazine and its main metabolites in rodent dams, fetuses, and neonates with physiologically based pharmacokinetic modeling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Zhoumeng; Interdisciplinary Toxicology Program, University of Georgia, Athens, GA 30602; Fisher, Jeffrey W.

Atrazine (ATR) is a widely used chlorotriazine herbicide, a ubiquitous environmental contaminant, and a potential developmental toxicant. To quantitatively evaluate placental/lactational transfer and fetal/neonatal tissue dosimetry of ATR and its major metabolites, physiologically based pharmacokinetic models were developed for rat dams, fetuses and neonates. These models were calibrated using pharmacokinetic data from rat dams repeatedly exposed (oral gavage; 5 mg/kg) to ATR followed by model evaluation against other available rat data. Model simulations corresponded well to the majority of available experimental data and suggest that: (1) the fetus is exposed to both ATR and its major metabolite didealkylatrazine (DACT) atmore » levels similar to maternal plasma levels, (2) the neonate is exposed mostly to DACT at levels two-thirds lower than maternal plasma or fetal levels, while lactational exposure to ATR is minimal, and (3) gestational carryover of DACT greatly affects its neonatal dosimetry up until mid-lactation. To test the model's cross-species extrapolation capability, a pharmacokinetic study was conducted with pregnant C57BL/6 mice exposed (oral gavage; 5 mg/kg) to ATR from gestational day 12 to 18. By using mouse-specific parameters, the model predictions fitted well with the measured data, including placental ATR/DACT levels. However, fetal concentrations of DACT were overestimated by the model (10-fold). This overestimation suggests that only around 10% of the DACT that reaches the fetus is tissue-bound. These rodent models could be used in fetal/neonatal tissue dosimetry predictions to help design/interpret early life toxicity/pharmacokinetic studies with ATR and as a foundation for scaling to humans. - Highlights: • We developed PBPK models for atrazine in rat dams, fetuses, and neonates. • We conducted pharmacokinetic (PK) study with atrazine in pregnant mice. • Model predictions were in good agreement with experimental rat and mouse PK data. • The fetus is exposed to atrazine/its main metabolite at levels similar to the dam. • The nursing neonate is exposed primarily to atrazine's main metabolite DACT.« less

Effects of nuclear transfer procedures on ES cell cloning efficiency in the mouse.

PubMed

Yabuuchi, Akiko; Yasuda, Yoshiko; Kato, Yoko; Tsunoda, Yukio

2004-04-01

Enucleated oocytes receiving mouse embryonic stem (ES) cells develop into fertile young. The developmental potential to young is low, however, and the rate of postnatal death is high. We examined the effect of various nuclear transfer procedures on the in vitro and in vivo developmental potential of nuclear-transferred oocytes. The potential of oocytes receiving ES cells at M phase to develop into blastocysts after fusion by Sendai virus was high compared with that after direct injection (67% vs. 30%). The developmental potential of oocytes receiving ES cells at the M phase is higher than that of oocytes receiving ES cells at the G(1) phase (30-67% vs. 2-5%). Developmental ability to live young was low in all groups (0-4%). Different activation protocols affected the potential to develop into blastocysts to a different extent (27-62%), but did not affect the potential to develop into live young (0-3%). The present study demonstrated that the various conditions examined did not affect the potential of nuclear-transferred oocytes receiving ES cells to develop into live young or the incidence of postnatal death.

Hydrodynamic liver gene transfer mechanism involves transient sinusoidal blood stasis and massive hepatocyte endocytic vesicles.

PubMed

Crespo, A; Peydró, A; Dasí, F; Benet, M; Calvete, J J; Revert, F; Aliño, S F

2005-06-01

The present study contributes to clarify the mechanism underlying the high efficacy of hepatocyte gene transfer mediated by hydrodynamic injection. Gene transfer experiments were performed employing the hAAT gene, and the efficacy and differential identification in mouse plasma of human transgene versus mouse gene was assessed by ELISA and proteomic procedures, respectively. By applying different experimental strategies such as cumulative dose-response efficacy, hemodynamic changes reflected by venous pressures, intravital microscopy, and morphological changes established by transmission electron microscopy, we found that: (a) cumulative multiple doses of transgene by hydrodynamic injection are efficient and well tolerated, resulting in therapeutic plasma levels of hAAT; (b) hydrodynamic injection mediates a transient inversion of intrahepatic blood flow, with circulatory stasis for a few minutes mainly in pericentral vein sinusoids; (c) transmission electron microscopy shows hydrodynamic injection to promote massive megafluid endocytic vesicles among hepatocytes around the central vein but not in hepatocytes around the periportal vein. We suggest that the mechanism of hydrodynamic liver gene transfer involves transient inversion of intrahepatic flow, sinusoidal blood stasis, and massive fluid endocytic vesicles in pericentral vein hepatocytes.

Heat loss regulation: role of appendages and torso in the deer mouse and the white rabbit.

PubMed

Conley, K E; Porter, W P

1985-01-01

Thermal conductance was subdivided into the component conductances of the appendages and torso using a heat transfer analysis for the deer mouse, Peromyscus maniculatus, and the white rabbit, Oryctolagus cuniculus. Our analysis was based on laboratory measurements of skin temperature and respiratory gas exchange made between air temperatures of 8 and 34 degrees C for the deer mouse, and from published data for the white rabbit. Two series conductances to heat transfer for each appendage and torso were evaluated: internal (hin), for blood flow and tissue conduction to the skin surface, and external (hex), for heat loss from the skin surface to the environment. These two series conductances were represented in a single, total conductance (htot). The limit to htot was set by hex and was reached by the torso htot of both animals. The increase in torso htot observed with air temperature for the mouse suggests that a pilomotor change in fur depth occurred. A control of htot below the limit set by hex was achieved by the hin of each appendage. Elevation of mouse thermal conductance (C) resulted from increases in feet, tail, and torso htot. In contrast, the rabbit showed no change in torso htot between 5 and 30 degrees C and ear htot exclusively increased C over these air temperatures. We suggest that the hyperthermia reported for the rabbit at 35 degrees C resulted from C reaching the physical limit set by torso and near hex. Thus the ear alone adjusted rabbit C, whereas the feet, tail, and the torso contributed to the adjustment of mouse C.

Tumor-homing peptides as tools for targeted delivery of payloads to the placenta

PubMed Central

King, Anna; Ndifon, Cornelia; Lui, Sylvia; Widdows, Kate; Kotamraju, Venkata R.; Agemy, Lilach; Teesalu, Tambet; Glazier, Jocelyn D.; Cellesi, Francesco; Tirelli, Nicola; Aplin, John D.; Ruoslahti, Erkki; Harris, Lynda K.

2016-01-01

The availability of therapeutics to treat pregnancy complications is severely lacking mainly because of the risk of causing harm to the fetus. As enhancement of placental growth and function can alleviate maternal symptoms and improve fetal growth in animal models, we have developed a method for targeted delivery of payloads to the placenta. We show that the tumor-homing peptide sequences CGKRK and iRGD bind selectively to the placental surface of humans and mice and do not interfere with normal development. Peptide-coated nanoparticles intravenously injected into pregnant mice accumulated within the mouse placenta, whereas control nanoparticles exhibited reduced binding and/or fetal transfer. We used targeted liposomes to efficiently deliver cargoes of carboxyfluorescein and insulin-like growth factor 2 to the mouse placenta; the latter significantly increased mean placental weight when administered to healthy animals and significantly improved fetal weight distribution in a well-characterized model of fetal growth restriction. These data provide proof of principle for targeted delivery of drugs to the placenta and provide a novel platform for the development of placenta-specific therapeutics. PMID:27386551

HSV-mediated gene transfer of vascular endothelial growth factor to dorsal root ganglia prevents diabetic neuropathy

PubMed Central

Chattopadhyay, M; Krisky, D; Wolfe, D; Glorioso, JC; Mata, M; Fink, DJ

2005-01-01

We examined the utility of herpes simplex virus (HSV) vector-mediated gene transfer of vascular endothelial growth factor (VEGF) in a mouse model of diabetic neuropathy. A replication-incompetent HSV vector with VEGF under the control of the HSV ICP0 promoter (vector T0VEGF) was constructed. T0VEGF expressed and released VEGF from primary dorsal root ganglion (DRG) neurons in vitro, and following subcutaneous inoculation in the foot, expressed VEGF in DRG and nerve in vivo. At 2 weeks after induction of diabetes, subcutaneous inoculation of T0VEGF prevented the reduction in sensory nerve amplitude characteristic of diabetic neuropathy measured 4 weeks later, preserved autonomic function measured by pilocarpine-induced sweating, and prevented the loss of nerve fibers in the skin and reduction of neuropeptide calcitonin gene-related peptide and substance P in DRG neurons of the diabetic mice. HSV-mediated transfer of VEGF to DRG may prove useful in treatment of diabetic neuropathy. PMID:15843809

Chromosomal integration of adenoviral vector DNA in vivo.

PubMed

Stephen, Sam Laurel; Montini, Eugenio; Sivanandam, Vijayshankar Ganesh; Al-Dhalimy, Muhseen; Kestler, Hans A; Finegold, Milton; Grompe, Markus; Kochanek, Stefan

2010-10-01

So far there has been no report of any clinical or preclinical evidence for chromosomal vector integration following adenovirus (Ad) vector-mediated gene transfer in vivo. We used liver gene transfer with high-capacity Ad vectors in the FAH(Deltaexon5) mouse model to analyze homologous and heterologous recombination events between vector and chromosomal DNA. Intravenous injection of Ad vectors either expressing a fumarylacetoacetate hydrolase (FAH) cDNA or carrying part of the FAH genomic locus resulted in liver nodules of FAH-expressing hepatocytes, demonstrating chromosomal vector integration. Analysis of junctions between vector and chromosomal DNA following heterologous recombination indicated integration of the vector genome through its termini. Heterologous recombination occurred with a median frequency of 6.72 x 10(-5) per transduced hepatocyte, while homologous recombination occurred more rarely with a median frequency of 3.88 x 10(-7). This study has established quantitative and qualitative data on recombination of adenoviral vector DNA with genomic DNA in vivo, contributing to a risk-benefit assessment of the biosafety of Ad vector-mediated gene transfer.

AAV-dominant negative tumor necrosis factor (DN-TNF) gene transfer to the striatum does not rescue medium spiny neurons in the YAC128 mouse model of Huntington's disease.

PubMed

Alto, Laura Taylor; Chen, Xi; Ruhn, Kelly A; Treviño, Isaac; Tansey, Malú G

2014-01-01

CNS inflammation is a hallmark of neurodegenerative disease, and recent studies suggest that the inflammatory response may contribute to neuronal demise. In particular, increased tumor necrosis factor (TNF) signaling is implicated in the pathology of both Parkinson's disease (PD) and Alzheimer's disease (AD). We have previously shown that localized gene delivery of dominant negative TNF to the degenerating brain region can limit pathology in animal models of PD and AD. TNF is upregulated in Huntington's disease (HD), like in PD and AD, but it is unknown whether TNF signaling contributes to neuronal degeneration in HD. We used in vivo gene delivery to test whether selective reduction of soluble TNF signaling could attenuate medium spiny neuron (MSN) degeneration in the YAC128 transgenic (TG) mouse model of Huntington's disease (HD). AAV vectors encoding cDNA for dominant-negative tumor necrosis factor (DN-TNF) or GFP (control) were injected into the striatum of young adult wild type WT and YAC128 TG mice and achieved 30-50% target coverage. Expression of dominant negative TNF protein was confirmed immunohistologically and biochemically and was maintained as mice aged to one year, but declined significantly over time. However, the extent of striatal DN-TNF gene transfer achieved in our studies was not sufficient to achieve robust effects on neuroinflammation, rescue degenerating MSNs or improve motor function in treated mice. Our findings suggest that alternative drug delivery strategies should be explored to determine whether greater target coverage by DN-TNF protein might afford some level of neuroprotection against HD-like pathology and/or that soluble TNF signaling may not be the primary driver of striatal neuroinflammation and MSN loss in YAC128 TG mice.

High-level resistance to gentamicin: genetic transfer between Enterococcus faecalis isolated from food of animal origin and human microbiota.

PubMed

Sparo, M; Urbizu, L; Solana, M V; Pourcel, G; Delpech, G; Confalonieri, A; Ceci, M; Sánchez Bruni, S F

2012-02-01

  To investigate the in vivo gene transfer of high-level gentamicin resistance (HLRG) from Enterococcus faecalis isolated from the food of animal origin to a human isolate, using a mouse model of intestinally colonized human microbiota.   In vitro study: The presence of plasmids involved in HLRG coding was investigated. After the conjugation experiment, the recipient strain, Ent. faecalis JH2-SS, acquired a plasmid responsible for HLRG [minimal inhibitory concentration (MIC) >800 μg ml(-1) ], in a similar position to the donor cells. In vivo study: Seven BALB/c mice were dosed with ceftriaxone (400 mg kg(-1) ) and then inoculated with a dilution of 1/100 of human faeces (HFc). After 72 h, Ent. faecalis JH2-SS (recipient) was inoculated and then, after a further 72 h, the animals were given Ent. faecalis CS19, isolated from the food of animal origin, involved in HLRG (donor). The presence of transconjugant strains in HFc was subsequently recorded on a daily basis until the end of the experiment. The clonal relationship between Ent. faecalis and Escherichia coli in faeces was assessed by RAPD-PCR. Both the in vitro and in vivo studies showed that the receptor strain acquired a plasmid responsible for HLRG (MICs >800 μg ml(-1) ), which migrated with a similar relative mobility value. Transconjugant strains were detected from 24 h after the donor strain inoculation and persisted until the end of the experiment.   The in vivo gene transfer of HLRG from Ent. faecalis strains, isolated from the food of animal origin, to human microbiota has been demonstrated in a mouse model.   The complexity found on the therapeutic responses of invasive infectious diseases caused by Ent. faecalis facilitates the assessment of food of animal origin as a resistant pathogen reservoir. In addition, this study may contribute to the understanding of antimicrobials' resistance gene transfer between Ent. faecalis strains from food and human GI tract. © 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.

At the bench: adoptive cell therapy for melanoma.

PubMed

Urba, Walter J

2014-06-01

The cellular and molecular principles that furnish the foundation for ACT of melanoma and their implications for further clinical research are reviewed. The parallel advances in basic immunology, preclinical animal studies, and clinical trials over the last two decades have been integrated successfully with improvements in technology to produce an effective ACT strategy for patients with melanoma. From the initial observation that tumors could be treated effectively by the transfer of immune cells to current strategies using preconditioning with myeloablative therapy before adoptive transfer of native or genetically altered T cells, the role of preclinical animal models is discussed. The importance of the pmel transgenic mouse model in the determination of the mechanisms of lymphodepletion, the ongoing work to identify the optimal T cells for adoptive immunotherapy, and the early impact of the emerging discipline of synthetic biology are highlighted. The clinical consequences of the research described herein are reviewed in the companion manuscript. © 2014 Society for Leukocyte Biology.

Widespread and highly persistent gene transfer to the CNS by retrovirus vector in utero: implication for gene therapy to Krabbe disease.

PubMed

Shen, Jin-Song; Meng, Xing-Li; Yokoo, Takashi; Sakurai, Ken; Watabe, Kazuhiko; Ohashi, Toya; Eto, Yoshikatsu

2005-05-01

Brain-directed prenatal gene therapy may benefit some lysosomal storage diseases that affect the central nervous system (CNS) before birth. Our previous study showed that intrauterine introduction of recombinant adenoviruses into cerebral ventricles results in efficient gene transfer to the CNS in the mouse. However, transgene expression decreased with time due to the non-integrative property of adenoviral vectors. In this study, in order to obtain permanent gene transduction, we investigated the feasibility of retrovirus-mediated in utero gene transduction. Concentrated retrovirus encoding the LacZ gene was injected into the cerebral ventricles of the embryos of normal and twitcher mice (a murine model of Krabbe disease) at embryonic day 12. The distribution and maintenance of the transgene expression in the recipient brain were analyzed histochemically, biochemically and by the quantitative polymerase chain reaction method pre- and postnatally. Efficient and highly persistent gene transduction to the brain was achieved both in normal and the twitcher mouse. Transduced neurons, astrocytes and oligodendrocytes were distributed throughout the brain. The transduced LacZ gene, its transcript and protein expression in the brain were maintained for 14 months without decrement. In addition, gene transduction to multiple tissues other than the brain was also detected at low levels. This study suggests that brain-directed in utero gene transfer using retrovirus vector may be beneficial to the treatment of lysosomal storage diseases with severe brain damage early in life, such as Krabbe disease. Copyright (c) 2005 John Wiley & Sons, Ltd.

Enhancing Autophagy with Drugs or Lung-directed Gene Therapy Reverses the Pathological Effects of Respiratory Epithelial Cell Proteinopathy.

PubMed

Hidvegi, Tunda; Stolz, Donna B; Alcorn, John F; Yousem, Samuel A; Wang, Jieru; Leme, Adriana S; Houghton, A McGarry; Hale, Pamela; Ewing, Michael; Cai, Houming; Garchar, Evelyn Akpadock; Pastore, Nunzia; Annunziata, Patrizia; Kaminski, Naftali; Pilewski, Joseph; Shapiro, Steven D; Pak, Stephen C; Silverman, Gary A; Brunetti-Pierri, Nicola; Perlmutter, David H

2015-12-11

Recent studies have shown that autophagy mitigates the pathological effects of proteinopathies in the liver, heart, and skeletal muscle but this has not been investigated for proteinopathies that affect the lung. This may be due at least in part to the lack of an animal model robust enough for spontaneous pathological effects from proteinopathies even though several rare proteinopathies, surfactant protein A and C deficiencies, cause severe pulmonary fibrosis. In this report we show that the PiZ mouse, transgenic for the common misfolded variant α1-antitrypsin Z, is a model of respiratory epithelial cell proteinopathy with spontaneous pulmonary fibrosis. Intracellular accumulation of misfolded α1-antitrypsin Z in respiratory epithelial cells of the PiZ model resulted in activation of autophagy, leukocyte infiltration, and spontaneous pulmonary fibrosis severe enough to elicit functional restrictive deficits. Treatment with autophagy enhancer drugs or lung-directed gene transfer of TFEB, a master transcriptional activator of the autophagolysosomal system, reversed these proteotoxic consequences. We conclude that this mouse is an excellent model of respiratory epithelial proteinopathy with spontaneous pulmonary fibrosis and that autophagy is an important endogenous proteostasis mechanism and an attractive target for therapy. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Transfer in SDS of biotinylated proteins from acrylamide gels to an avidin-coated membrane filter.

PubMed

Karlin, Arthur; Wang, Chaojian; Li, Jing; Xu, Qiang

2004-06-01

Avidin was covalently linked to aldehyde-derivatized polyethersulfone membrane filters. These filters were used in Western blot analysis of proteins reacted with biotinylation reagents and electrophoresed in sodium dodecyl sulfate (SDS) on polyacrylamide gels. Electrophoretic transfer from the gels to these filters was in 0.1% SDS, in which the covalently bound avidin retained its biotin-binding capacity. We compared Western blots on avidin-coated membrane filters of biotinylated and nonbiotinylated forms of mouse immunoglobulin G (IgG), mouse IgG heavy chain, muscle-type acetylcholine receptor alpha subunit, and fused alpha and beta subunits of receptor. Biotinylated proteins were captured with high specificity compared to their nonbiotinylated counterparts and sensitively detected on the avidin-coated membranes.

A novel homologous model for gene therapy of dwarfism by non-viral transfer of the mouse growth hormone gene into immunocompetent dwarf mice.

PubMed

Cecchi, Claudia R; Higuti, Eliza; Oliveira, Nelio A J; Lima, Eliana R; Jakobsen, Maria; Dagnaes-Hansen, Frederick; Gissel, Hanne; Aagaard, Lars; Jensen, Thomas G; Jorge, Alexander A L; Bartolini, Paolo; Peroni, Cibele N

2014-02-01

The possibilities for non-viral GH gene therapy are studied in immunocompetent dwarf mice (lit/lit). As expression vector we used a plasmid previously employed in immunodeficient dwarf mice (pUBI-hGH-gDNA) by replacing the human GH gene with the genomic sequence of mouse-GH DNA (pUBI-mGH-gDNA). HEK-293 human cells transfected with pUBI-mGH-gDNA produced 3.0 µg mGH/10(6) cells/day compared to 3.7 µg hGH/10(6) cells/day for pUBIhGH- gDNA transfected cells. The weight of lit/lit mice treated with the same two plasmids (50 µg DNA/mouse) by electrotransfer into the quadriceps muscle was followed for 3 months. The weight increase up to 15 days for mGH, hGH and saline treated mice were 0.130, 0.112 and 0.027 g/mouse/day. Most sera from hGH-treated mice contained anti-hGH antibodies already on day 15, with the highest titers on day 45, while no significant anti-mGH antibodies were observed in mGH-treated mice. At the end of 3 months, the weight increase for mGH-treated mice was 34.3%, while the nose-to-tail and femur lengths increased 9.5% and 24.3%. Mouse-GH and hGH circulating levels were 4-5 ng/mL 15 days after treatment, versus control levels of ~0.7 ng GH/mL (P<0.001). In mGH-treated mice, mIGF-I determined on days 15, 45 and 94 were 1.5- to 3-fold higher than the control and 1.2- to 1.6-fold higher than hGH-treated mice. The described homologous model represents an important progress forming the basis for preclinical testing of non-viral gene therapy for GH deficiency.

Discovery and Targeted Proteomics on Cutaneous Biopsies Infected by Borrelia to Investigate Lyme Disease*

PubMed Central

Schnell, Gilles; Boeuf, Amandine; Westermann, Benoît; Jaulhac, Benoît; Lipsker, Dan; Carapito, Christine; Boulanger, Nathalie; Ehret-Sabatier, Laurence

2015-01-01

Lyme disease is the most important vector-borne disease in the Northern hemisphere and represents a major public health challenge with insufficient means of reliable diagnosis. Skin is rarely investigated in proteomics but constitutes in the case of Lyme disease the key interface where the pathogens can enter, persist, and multiply. Therefore, we investigated proteomics on skin samples to detect Borrelia proteins directly in cutaneous biopsies in a robust and specific way. We first set up a discovery gel prefractionation-LC-MS/MS approach on a murine model infected by Borrelia burgdorferi sensu stricto that allowed the identification of 25 Borrelia proteins among more than 1300 mouse proteins. Then we developed a targeted gel prefractionation-LC-selected reaction monitoring (SRM) assay to detect 9/33 Borrelia proteins/peptides in mouse skin tissue samples using heavy labeled synthetic peptides. We successfully transferred this assay from the mouse model to human skin biopsies (naturally infected by Borrelia), and we were able to detect two Borrelia proteins: OspC and flagellin. Considering the extreme variability of OspC, we developed an extended SRM assay to target a large set of variants. This assay afforded the detection of nine peptides belonging to either OspC or flagellin in human skin biopsies. We further shortened the sample preparation and showed that Borrelia is detectable in mouse and human skin biopsies by directly using a liquid digestion followed by LC-SRM analysis without any prefractionation. This study thus shows that a targeted SRM approach is a promising tool for the early direct diagnosis of Lyme disease with high sensitivity (<10 fmol of OspC/mg of human skin biopsy). PMID:25713121

Prevention of Asthma Exacerbation in a Mouse Model by Simultaneous Inhibition of NF-κB and STAT6 Activation Using a Chimeric Decoy Strategy.

PubMed

Miyake, Tetsuo; Miyake, Takashi; Sakaguchi, Makoto; Nankai, Hirokazu; Nakazawa, Takahiro; Morishita, Ryuichi

2018-03-02

Transactivation of inflammatory and immune mediators in asthma is tightly regulated by nuclear factor κB (NF-κB) and signal transducer and activator of transcription 6 (STAT6). Therefore, we investigated the efficacy of simultaneous inhibition of NF-κB and STAT6 using a chimeric decoy strategy to prevent asthma exacerbation. The effects of decoy oligodeoxynucleotides were evaluated using an ovalbumin-induced mouse asthma model. Ovalbumin-sensitized mice received intratracheal administration of decoy oligodeoxynucleotides 3 days before ovalbumin challenge. Fluorescent-dye-labeled decoy oligodeoxynucleotides could be detected in lymphocytes and macrophages in the lung, and activation of NF-κB and STAT6 was inhibited by chimeric decoy oligodeoxynucleotide transfer. Consequently, treatment with chimeric or NF-κB decoy oligodeoxynucleotides protected against methacholine-induced airway hyperresponsiveness, whereas the effect of chimeric decoy oligodeoxynucleotides was significantly greater than that of NF-κB decoy oligodeoxynucleotides. Treatment with chimeric decoy oligodeoxynucleotides suppressed airway inflammation through inhibition of overexpression of interleukin-4 (IL-4), IL-5, and IL-13 and inflammatory infiltrates. Histamine levels in the lung were reduced via suppression of mast cell accumulation. A significant reduction in mucin secretion was observed due to suppression of MUC5AC gene expression. Interestingly, the inhibitory effects on IL-5, IL-13, and histamine secretion were achieved by transfer of chimeric decoy oligodeoxynucleotides only. This novel therapeutic approach could be useful to treat patients with various types of asthma. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Bone marrow endothelial progenitors augment atherosclerotic plaque regression in a mouse model of plasma lipid lowering

PubMed Central

Yao, Longbiao; Heuser-Baker, Janet; Herlea-Pana, Oana; Iida, Ryuji; Wang, Qilong; Zou, Ming-Hui; Barlic-Dicen, Jana

2012-01-01

The major event initiating atherosclerosis is hypercholesterolemia-induced disruption of vascular endothelium integrity. In settings of endothelial damage, endothelial progenitor cells (EPCs) are mobilized from bone marrow into circulation and home to sites of vascular injury where they aid endothelial regeneration. Given the beneficial effects of EPCs in vascular repair, we hypothesized that these cells play a pivotal role in atherosclerosis regression. We tested our hypothesis in the atherosclerosis-prone mouse model in which hypercholesterolemia, one of the main factors affecting EPC homeostasis, is reversible (Reversa mice). In these mice normalization of plasma lipids decreased atherosclerotic burden; however, plaque regression was incomplete. To explore whether endothelial progenitors contribute to atherosclerosis regression, bone marrow EPCs from a transgenic strain expressing green fluorescent protein under the control of endothelial cell-specific Tie2 promoter (Tie2-GFP+) were isolated. These cells were then adoptively transferred into atheroregressing Reversa recipients where they augmented plaque regression induced by reversal of hypercholesterolemia. Advanced plaque regression correlated with engraftment of Tie2-GFP+ EPCs into endothelium and resulted in an increase in atheroprotective nitric oxide and improved vascular relaxation. Similarly augmented plaque regression was also detected in regressing Reversa mice treated with the stem cell mobilizer AMD3100 which also mobilizes EPCs to peripheral blood. We conclude that correction of hypercholesterolemia in Reversa mice leads to partial plaque regression that can be augmented by AMD3100 treatment or by adoptive transfer of EPCs. This suggests that direct cell therapy or indirect progenitor cell mobilization therapy may be used in combination with statins to treat atherosclerosis. PMID:23081735

Chronic CCl4 intoxication causes liver and bone damage similar to the human pathology of hepatic osteodystrophy: a mouse model to analyse the liver-bone axis.

PubMed

Nussler, Andreas K; Wildemann, Britt; Freude, Thomas; Litzka, Christian; Soldo, Petra; Friess, Helmut; Hammad, Seddik; Hengstler, Jan G; Braun, Karl F; Trak-Smayra, Viviane; Godoy, Patricio; Ehnert, Sabrina

2014-04-01

Patients with chronic liver diseases frequently exhibit decreased bone mineral densities (BMD), which is defined as hepatic osteodystrophy (HOD). HOD is a multifactorial disease whose regulatory mechanisms are barely understood. Thus, an early diagnosis and therapy is hardly possible. Therefore, the aim of our study consisted in characterizing a mouse model reflecting the human pathomechanism. Serum samples were collected from patients with chronic liver diseases and 12-week old C57Bl6/N mice after 6-week treatment with carbon tetrachloride (CCl4). Repetitive injections of CCl4 induced liver damage in mice, resembling liver fibrosis in patients, as assessed by serum analysis and histological staining. Although CCl4 did not affect primary osteoblast cultures, μCT analysis revealed significantly decreased BMD, bone volume, trabecular number and thickness in CCl4-treated mice. In both HOD patients and CCl4-treated mice, an altered vitamin D metabolism with decreased CYP27A1, CYP2R1, vitamin D-binding protein GC and increased 7-dehydrocholesterol reductase hepatic gene expression, results in decreased 25-OH vitamin D serum levels. Moreover, both groups exhibit excessively high active transforming growth factor-beta (TGF-β) serum levels, inhibiting osteoblast function in vitro. Summarizing, our mouse model presents possible mediators of HOD, e.g. altered vitamin D metabolism and increased active TGF-β. Liver damage and significant changes in bone structure and mineralization are already visible by μCT analysis after 6 weeks of CCl4 treatment. This fast response and easy transferability makes it an ideal model to investigate specific gene functions in HOD.

A Fully Implantable Pacemaker for the Mouse: From Battery to Wireless Power

PubMed Central

Zellmer, Erik R.; Weinheimer, Carla J.; MacEwan, Matthew R.; Cui, Sophia X.; Nerbonne, Jeanne M.; Efimov, Igor R.

2013-01-01

Animal models have become a popular platform for the investigation of the molecular and systemic mechanisms of pathological cardiovascular physiology. Chronic pacing studies with implantable pacemakers in large animals have led to useful models of heart failure and atrial fibrillation. Unfortunately, molecular and genetic studies in these large animal models are often prohibitively expensive or not available. Conversely, the mouse is an excellent species for studying molecular mechanisms of cardiovascular disease through genetic engineering. However, the large size of available pacemakers does not lend itself to chronic pacing in mice. Here, we present the design for a novel, fully implantable wireless-powered pacemaker for mice capable of long-term (>30 days) pacing. This design is compared to a traditional battery-powered pacemaker to demonstrate critical advantages achieved through wireless inductive power transfer and control. Battery-powered and wireless-powered pacemakers were fabricated from standard electronic components in our laboratory. Mice (n = 24) were implanted with endocardial, battery-powered devices (n = 14) and epicardial, wireless-powered devices (n = 10). Wireless-powered devices were associated with reduced implant mortality and more reliable device function compared to battery-powered devices. Eight of 14 (57.1%) mice implanted with battery-powered pacemakers died following device implantation compared to 1 of 10 (10%) mice implanted with wireless-powered pacemakers. Moreover, device function was achieved for 30 days with the wireless-powered device compared to 6 days with the battery-powered device. The wireless-powered pacemaker system presented herein will allow electrophysiology studies in numerous genetically engineered mouse models as well as rapid pacing-induced heart failure and atrial arrhythmia in mice. PMID:24194832

A Mouse Model of Harlequin Ichthyosis Delineates a Key Role for Abca12 in Lipid Homeostasis

PubMed Central

Smyth, Ian; Mukhamedova, Nigora; Meikle, Peter J.; Ellis, Sarah; Slattery, Keith; Collinge, Janelle E.; de Graaf, Carolyn A.; Bahlo, Melanie; Sviridov, Dmitri

2008-01-01

Harlequin Ichthyosis (HI) is a severe and often lethal hyperkeratotic skin disease caused by mutations in the ABCA12 transport protein. In keratinocytes, ABCA12 is thought to regulate the transfer of lipids into small intracellular trafficking vesicles known as lamellar bodies. However, the nature and scope of this regulation remains unclear. As part of an original recessive mouse ENU mutagenesis screen, we have identified and characterised an animal model of HI and showed that it displays many of the hallmarks of the disease including hyperkeratosis, loss of barrier function, and defects in lipid homeostasis. We have used this model to follow disease progression in utero and present evidence that loss of Abca12 function leads to premature differentiation of basal keratinocytes. A comprehensive analysis of lipid levels in mutant epidermis demonstrated profound defects in lipid homeostasis, illustrating for the first time the extent to which Abca12 plays a pivotal role in maintaining lipid balance in the skin. To further investigate the scope of Abca12's activity, we have utilised cells from the mutant mouse to ascribe direct transport functions to the protein and, in doing so, we demonstrate activities independent of its role in lamellar body function. These cells have severely impaired lipid efflux leading to intracellular accumulation of neutral lipids. Furthermore, we identify Abca12 as a mediator of Abca1-regulated cellular cholesterol efflux, a finding that may have significant implications for other diseases of lipid metabolism and homeostasis, including atherosclerosis. PMID:18802465

Finding mouse models of human lymphomas and leukemia's using the Jackson laboratory mouse tumor biology database.

PubMed

Begley, Dale A; Sundberg, John P; Krupke, Debra M; Neuhauser, Steven B; Bult, Carol J; Eppig, Janan T; Morse, Herbert C; Ward, Jerrold M

2015-12-01

Many mouse models have been created to study hematopoietic cancer types. There are over thirty hematopoietic tumor types and subtypes, both human and mouse, with various origins, characteristics and clinical prognoses. Determining the specific type of hematopoietic lesion produced in a mouse model and identifying mouse models that correspond to the human subtypes of these lesions has been a continuing challenge for the scientific community. The Mouse Tumor Biology Database (MTB; http://tumor.informatics.jax.org) is designed to facilitate use of mouse models of human cancer by providing detailed histopathologic and molecular information on lymphoma subtypes, including expertly annotated, on line, whole slide scans, and providing a repository for storing information on and querying these data for specific lymphoma models. Copyright © 2015 Elsevier Inc. All rights reserved.

Mouse models rarely mimic the transcriptome of human neurodegenerative diseases: A systematic bioinformatics-based critique of preclinical models.

PubMed

Burns, Terry C; Li, Matthew D; Mehta, Swapnil; Awad, Ahmed J; Morgan, Alexander A

2015-07-15

Translational research for neurodegenerative disease depends intimately upon animal models. Unfortunately, promising therapies developed using mouse models mostly fail in clinical trials, highlighting uncertainty about how well mouse models mimic human neurodegenerative disease at the molecular level. We compared the transcriptional signature of neurodegeneration in mouse models of Alzheimer׳s disease (AD), Parkinson׳s disease (PD), Huntington׳s disease (HD) and amyotrophic lateral sclerosis (ALS) to human disease. In contrast to aging, which demonstrated a conserved transcriptome between humans and mice, only 3 of 19 animal models showed significant enrichment for gene sets comprising the most dysregulated up- and down-regulated human genes. Spearman׳s correlation analysis revealed even healthy human aging to be more closely related to human neurodegeneration than any mouse model of AD, PD, ALS or HD. Remarkably, mouse models frequently upregulated stress response genes that were consistently downregulated in human diseases. Among potential alternate models of neurodegeneration, mouse prion disease outperformed all other disease-specific models. Even among the best available animal models, conserved differences between mouse and human transcriptomes were found across multiple animal model versus human disease comparisons, surprisingly, even including aging. Relative to mouse models, mouse disease signatures demonstrated consistent trends toward preserved mitochondrial function protein catabolism, DNA repair responses, and chromatin maintenance. These findings suggest a more complex and multifactorial pathophysiology in human neurodegeneration than is captured through standard animal models, and suggest that even among conserved physiological processes such as aging, mice are less prone to exhibit neurodegeneration-like changes. This work may help explain the poor track record of mouse-based translational therapies for neurodegeneration and provides a path forward to critically evaluate and improve animal models of human disease. Copyright © 2015 Elsevier B.V. All rights reserved.

Senescence of nickel-transformed cells by an X chromosome: Possible epigenetic control

DOE Office of Scientific and Technical Information (OSTI.GOV)

Klein, C.B.; Xin Wei Wang; Bhamra, R.K.

1991-02-15

Transfer of a normal Chinese hamster X chromosome (carried in a mouse A9 donor cell line) to a nickel-transformed Chinese hamster cell line with an Xq chromosome deletion resulted in senescense of these previously immortal cells. At early passages of the A9/CX donor cells, the hamster X chromosome was highly active, inducing senescence in 100% of the colonies obtained after its transfer into the nickel-transformed cells. However, senescence was reduced to 50% when Chinese hamster X chromosomes were transferred from later passage A9 cells. Full senescing activity of the intact hamster X chromosome was restored by treatment of the donormore » mouse cells with 5-azacytidine, which induced demethylation of DMA. These results suggest that a senescence gene or genes, which may be located on the Chinese hamster X chromosome, can be regulated by DNA methylation, and that escape from senescence and possibly loss of tumor suppressor gene activity can occur by epigenetic mechanisms.« less

Induction of cardiomyocyte-like cells in infarct hearts by gene transfer of Gata4, Mef2c, and Tbx5.

PubMed

Inagawa, Kohei; Miyamoto, Kazutaka; Yamakawa, Hiroyuki; Muraoka, Naoto; Sadahiro, Taketaro; Umei, Tomohiko; Wada, Rie; Katsumata, Yoshinori; Kaneda, Ruri; Nakade, Koji; Kurihara, Chitose; Obata, Yuichi; Miyake, Koichi; Fukuda, Keiichi; Ieda, Masaki

2012-10-12

After myocardial infarction (MI), massive cell death in the myocardium initiates fibrosis and scar formation, leading to heart failure. We recently found that a combination of 3 cardiac transcription factors, Gata4, Mef2c, and Tbx5 (GMT), reprograms fibroblasts directly into functional cardiomyocytes in vitro. To investigate whether viral gene transfer of GMT into infarcted hearts induces cardiomyocyte generation. Coronary artery ligation was used to generate MI in the mouse. In vitro transduction of GMT retrovirus converted cardiac fibroblasts from the infarct region into cardiomyocyte-like cells with cardiac-specific gene expression and sarcomeric structures. Injection of the green fluorescent protein (GFP) retrovirus into mouse hearts, immediately after MI, infected only proliferating noncardiomyocytes, mainly fibroblasts, in the infarct region. The GFP expression diminished after 2 weeks in immunocompetent mice but remained stable for 3 months in immunosuppressed mice, in which cardiac induction did not occur. In contrast, injection of GMT retrovirus into α-myosin heavy chain (αMHC)-GFP transgenic mouse hearts induced the expression of αMHC-GFP, a marker of cardiomyocytes, in 3% of virus-infected cells after 1 week. A pooled GMT injection into the immunosuppressed mouse hearts induced cardiac marker expression in retrovirus-infected cells within 2 weeks, although few cells showed striated muscle structures. To transduce GMT efficiently in vivo, we generated a polycistronic retrovirus expressing GMT separated by 2A "self-cleaving" peptides (3F2A). The 3F2A-induced cardiomyocyte-like cells in fibrotic tissue expressed sarcomeric α-actinin and cardiac troponin T and had clear cross striations. Quantitative RT-PCR also demonstrated that FACS-sorted 3F2A-transduced cells expressed cardiac-specific genes. GMT gene transfer induced cardiomyocyte-like cells in infarcted hearts.

Antiangiogenic agents can increase lymphocyte infiltration into tumor and enhance the effectiveness of adoptive immunotherapy of cancer.

PubMed

Shrimali, Rajeev K; Yu, Zhiya; Theoret, Marc R; Chinnasamy, Dhanalakshmi; Restifo, Nicholas P; Rosenberg, Steven A

2010-08-01

Adoptive cell transfer (ACT)-based immunotherapies can mediate objective cancer regression in animal models and in up to 70% of patients with metastatic melanoma; however, it remains unclear whether the tumor vasculature impedes the egress of tumor-specific T cells, thus hindering this immunotherapy. Disruption of the proangiogenic interaction of vascular endothelial growth factor (VEGF) with its receptor (VEGFR-2) has been reported to "normalize" tumor vasculature, enhancing the efficacy of chemotherapeutic agents by increasing their delivery to the tumor intersitium. We thus sought to determine whether disrupting VEGF/VEGFR-2 signaling could enhance the effectiveness of ACT in a murine cancer model. The administration of an antibody against mouse VEGF synergized with ACT to enhance inhibition of established, vascularized, B16 melanoma (P = 0.009) and improve survival (P = 0.003). Additive effects of an antibody against VEGFR-2 in conjunction with ACT were seen in this model (P = 0.013). Anti-VEGF, but not anti-VEGFR-2, antibody significantly increased infiltration of transferred cells into the tumor. Thus, normalization of tumor vasculature through disruption of the VEGF/VEGFR-2 axis can increase extravasation of adoptively transferred T cells into the tumor and improve ACT-based immunotherapy. These studies provide a rationale for the exploration of combining antiangiogenic agents with ACT for the treatment of patients with cancer.

Arthritis imaging using a near-infrared fluorescence folate-targeted probe

PubMed Central

Chen, Wei-Tsung; Mahmood, Umar; Weissleder, Ralph; Tung, Ching-Hsuan

2005-01-01

A recently developed near-infrared fluorescence-labeled folate probe (NIR2-folate) was tested for in vivo imaging of arthritis using a lipopolysaccharide intra-articular injection model and a KRN transgenic mice serum induction mouse model. In the lipopolysaccharide injection model, the fluorescence signal intensity of NIR2-folate (n = 12) and of free NIR2 (n = 5) was compared between lipopolysaccharide-treated and control joints. The fluorescence signal intensity of the NIR2-folate probe at the inflammatory joints was found to be significantly higher than the control normal joints (up to 2.3-fold, P < 0.001). The NIR2-free dye injection group showed a persistent lower enhancement ratio than the NIR2-folate probe injection group. Excessive folic acid was also given to demonstrate a competitive effect with the NIR2-folate. In the KRN serum transfer model (n = 4), NIR2-folate was applied at different time points after serum transfer, and the inflamed joints could be detected as early as 30 hours after arthritogenic antibody transfer (1.8-fold increase in signal intensity). Fluorescence microscopy, histology, and immunohistochemistry validated the optical imaging results. We conclude that in vivo arthritis detection was feasible using a folate-targeted near-infrared fluorescence probe. This receptor-targeted imaging method may facilitate improved arthritis diagnosis and early assessment of the disease progress by providing an in vivo characterization of active macrophage status in inflammatory joint diseases. PMID:15743478

IL-12 is required for differentiation of pathogenic CD8+ T cell effectors that cause myocarditis

PubMed Central

Grabie, Nir; Delfs, Michael W.; Westrich, Jason R.; Love, Victoria A.; Stavrakis, George; Ahmad, Ferhaan; Seidman, Christine E.; Seidman, Jonathan G.; Lichtman, Andrew H.

2003-01-01

Cardiac antigen–specific CD8+ T cells are involved in the autoimmune component of human myocarditis. Here, we studied the differentiation and migration of pathogenic CD8+ T cell effector cells in a new mouse model of autoimmune myocarditis. A transgenic mouse line was derived that expresses cardiac myocyte restricted membrane-bound ovalbumin (CMy-mOva). The endogenous adaptive immune system of CMy-mOva mice displays tolerance to ovalbumin. Adoptive transfer of naive CD8+ T cells from the ovalbumin-specific T cell receptor–transgenic (TCR-transgenic) OT-I strain induces myocarditis in CMy-mOva mice only after subsequent inoculation with ovalbumin-expressing vesicular stomatitis virus (VSV-Ova). OT-I effector T cells derived in vitro in the presence or absence of IL-12 were adoptively transferred into CMy-mOva mice, and the consequences were compared. Although IL-12 was not required for the generation of cytolytic and IFN-γ–producing effector T cells, only effectors primed in the presence of IL-12 infiltrated CMy-mOva hearts in significant numbers, causing lethal myocarditis. Furthermore, analysis of OT-I effectors collected from a mediastinal draining lymph node indicated that only effectors primed in vitro in the presence of IL-12 proliferated in vivo. These data demonstrate the importance of IL-12 in the differentiation of pathogenic CD8+ T cells that can cause myocarditis. PMID:12618521

A unique mouse strain that develops spontaneous, iodine-accelerated, pathogenic antibodies to the human thyrotrophin receptor 1

PubMed Central

Rapoport, Basil; Aliesky, Holly A.; Banuelos, Bianca; Chen, Chun-Rong; McLachlan, Sandra M.

2015-01-01

Antibodies that stimulate the thyrotropin receptor (TSHR), the cause of Graves’ hyperthyroidism, only develop in humans. TSHR antibodies can be induced in mice by immunization but studying pathogenesis and therapeutic intervention requires a model without immunization. Spontaneous, iodine-accelerated, thyroid autoimmunity develops in NOD.H2h4 mice associated with thyroglobulin and thyroid-peroxidase, but not TSHR, antibodies. We hypothesized that transferring the human (h)TSHR A-subunit to NOD.H2h4 mice would result in loss of tolerance to this protein. BALB/c hTSHR A-subunit mice were bred to NOD.H2h4 mice and transgenic offspring were repeatedly backcrossed to NOD.H2h4 mice. All offspring developed antibodies to thyroglobulin and thyroid-peroxidase. However, only TSHR-transgenic NOD.H2h4 mice (TSHR/NOD.H2h4) developed pathogenic TSHR antibodies as detected using clinical Graves’ disease assays. As in humans, TSHR/NOD.H2h4 females were more prone than males to developing pathogenic TSHR antibodies. Fortunately, in view of the confounding effect of excess thyroid hormone on immune responses, spontaneously arising pathogenic (h)TSHR antibodies cross-react poorly with the mouse TSHR and do not cause thyrotoxicosis. In summary, the TSHR/NOD.H2h4 mouse strain develops spontaneous, iodine-accelerated, pathogenic TSHR antibodies in females, providing a unique model to investigate disease pathogenesis and test novel TSHR-antigen specific immunotherapies aimed at curing Graves’ disease in humans. PMID:25825442

Lamellipodin-Deficient Mice: A Model of Rectal Carcinoma

PubMed Central

Miller, Cassandra L.; Muthupalani, Sureshkumar; Shen, Zeli; Drees, Frauke; Ge, Zhongming; Feng, Yan; Chen, Xiaowei; Gong, Guanyu; Nagar, Karan K.; Wang, Timothy C.; Gertler, Frank B.; Fox, James G.

2016-01-01

During a survey of clinical rectal prolapse (RP) cases in the mouse population at MIT animal research facilities, a high incidence of RP in the lamellipodin knock-out strain, C57BL/6-Raph1tm1Fbg (Lpd-/-) was documented. Upon further investigation, the Lpd-/- colony was found to be infected with multiple endemic enterohepatic Helicobacter species (EHS). Lpd-/- mice, a transgenic mouse strain produced at MIT, have not previously shown a distinct immune phenotype and are not highly susceptible to other opportunistic infections. Predominantly male Lpd-/- mice with RP exhibited lesions consistent with invasive rectal carcinoma concomitant to clinically evident RP. Multiple inflammatory cytokines, CD11b+Gr1+ myeloid-derived suppressor cell (MDSC) populations, and epithelial cells positive for a DNA damage biomarker, H2AX, were elevated in affected tissue, supporting their role in the neoplastic process. An evaluation of Lpd-/- mice with RP compared to EHS-infected, but clinically normal (CN) Lpd-/- animals indicated that all of these mice exhibit some degree of lower bowel inflammation; however, mice with prolapses had significantly higher degree of focal lesions at the colo-rectal junction. When Helicobacter spp. infections were eliminated in Lpd-/- mice by embryo transfer rederivation, the disease phenotype was abrogated, implicating EHS as a contributing factor in the development of rectal carcinoma. Here we describe lesions in Lpd-/- male mice consistent with a focal inflammation-induced neoplastic transformation and propose this strain as a mouse model of rectal carcinoma. PMID:27045955

System parameters for erythropoiesis control model: Comparison of normal values in human and mouse model

NASA Technical Reports Server (NTRS)

1979-01-01

The computer model for erythropoietic control was adapted to the mouse system by altering system parameters originally given for the human to those which more realistically represent the mouse. Parameter values were obtained from a variety of literature sources. Using the mouse model, the mouse was studied as a potential experimental model for spaceflight. Simulation studies of dehydration and hypoxia were performed. A comparison of system parameters for the mouse and human models is presented. Aside from the obvious differences expected in fluid volumes, blood flows and metabolic rates, larger differences were observed in the following: erythrocyte life span, erythropoietin half-life, and normal arterial pO2.

A novel surgical approach for intratracheal administration of bioactive agents in a fetal mouse model.

PubMed

Carlon, Marianne S; Toelen, Jaan; da Cunha, Marina Mori; Vidović, Dragana; Van der Perren, Anke; Mayer, Steffi; Sbragia, Lourenço; Nuyts, Johan; Himmelreich, Uwe; Debyser, Zeger; Deprest, Jan

2012-10-31

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and acquired diseases. Apart from congenital or inherited abnormalities with the requirement for long-term expression of the delivered gene, several non-inherited perinatal conditions, where short-term gene expression or pharmacological intervention is sufficient to achieve therapeutic effects, are considered as potential future indications for this kind of approach. Candidate diseases for the application of short-term prenatal therapy could be the transient neonatal deficiency of surfactant protein B causing neonatal respiratory distress syndrome(1,2) or hyperoxic injuries of the neonatal lung(3). Candidate diseases for permanent therapeutic correction are Cystic Fibrosis (CF)(4), genetic variants of surfactant deficiencies(5) and α1-antitrypsin deficiency(6). Generally, an important advantage of prenatal gene therapy is the ability to start therapeutic intervention early in development, at or even prior to clinical manifestations in the patient, thus preventing irreparable damage to the individual. In addition, fetal organs have an increased cell proliferation rate as compared to adult organs, which could allow a more efficient gene or stem cell transfer into the fetus. Furthermore, in utero gene delivery is performed when the individual's immune system is not completely mature. Therefore, transplantation of heterologous cells or supplementation of a non-functional or absent protein with a correct version should not cause immune sensitization to the cell, vector or transgene product, which has recently been proven to be the case with both cellular and genetic therapies(7). In the present study, we investigated the potential to directly target the fetal trachea in a mouse model. This procedure is in use in larger animal models such as rabbits and sheep(8), and even in a clinical setting(9), but has to date not been performed before in a mouse model. When studying the potential of fetal gene therapy for genetic diseases such as CF, the mouse model is very useful as a first proof-of-concept because of the wide availability of different transgenic mouse strains, the well documented embryogenesis and fetal development, less stringent ethical regulations, short gestation and the large litter size. Different access routes have been described to target the fetal rodent lung, including intra-amniotic injection(10-12), (ultrasound-guided) intrapulmonary injection(13,14) and intravenous administration into the yolk sac vessels(15,16) or umbilical vein(17). Our novel surgical procedure enables researchers to inject the agent of choice directly into the fetal mouse trachea which allows for a more efficient delivery to the airways than existing techniques(18).

Transgenic Parasites Stably Expressing Full-Length Plasmodium falciparum Circumsporozoite Protein as a Model for Vaccine Down-Selection in Mice Using Sterile Protection as an Endpoint

PubMed Central

Porter, Michael D.; Nicki, Jennifer; Pool, Christopher D.; DeBot, Margot; Illam, Ratish M.; Brando, Clara; Bozick, Brooke; De La Vega, Patricia; Angra, Divya; Spaccapelo, Roberta; Crisanti, Andrea; Murphy, Jittawadee R.; Bennett, Jason W.; Schwenk, Robert J.; Ockenhouse, Christian F.

2013-01-01

Circumsporozoite protein (CSP) of Plasmodium falciparum is a protective human malaria vaccine candidate. There is an urgent need for models that can rapidly down-select novel CSP-based vaccine candidates. In the present study, the mouse-mosquito transmission cycle of a transgenic Plasmodium berghei malaria parasite stably expressing a functional full-length P. falciparum CSP was optimized to consistently produce infective sporozoites for protection studies. A minimal sporozoite challenge dose was established, and protection was defined as the absence of blood-stage parasites 14 days after intravenous challenge. The specificity of protection was confirmed by vaccinating mice with multiple CSP constructs of differing lengths and compositions. Constructs that induced high NANP repeat-specific antibody titers in enzyme-linked immunosorbent assays were protective, and the degree of protection was dependent on the antigen dose. There was a positive correlation between antibody avidity and protection. The antibodies in the protected mice recognized the native CSP on the parasites and showed sporozoite invasion inhibitory activity. Passive transfer of anti-CSP antibodies into naive mice also induced protection. Thus, we have demonstrated the utility of a mouse efficacy model to down-select human CSP-based vaccine formulations. PMID:23536694

Human mammary microenvironment better regulates the biology of human breast cancer in humanized mouse model.

PubMed

Zheng, Ming-Jie; Wang, Jue; Xu, Lu; Zha, Xiao-Ming; Zhao, Yi; Ling, Li-Jun; Wang, Shui

2015-02-01

During the past decades, many efforts have been made in mimicking the clinical progress of human cancer in mouse models. Previously, we developed a human breast tissue-derived (HB) mouse model. Theoretically, it may mimic the interactions between "species-specific" mammary microenvironment of human origin and human breast cancer cells. However, detailed evidences are absent. The present study (in vivo, cellular, and molecular experiments) was designed to explore the regulatory role of human mammary microenvironment in the progress of human breast cancer cells. Subcutaneous (SUB), mammary fat pad (MFP), and HB mouse models were developed for in vivo comparisons. Then, the orthotopic tumor masses from three different mouse models were collected for primary culture. Finally, the biology of primary cultured human breast cancer cells was compared by cellular and molecular experiments. Results of in vivo mouse models indicated that human breast cancer cells grew better in human mammary microenvironment. Cellular and molecular experiments confirmed that primary cultured human breast cancer cells from HB mouse model showed a better proliferative and anti-apoptotic biology than those from SUB to MFP mouse models. Meanwhile, primary cultured human breast cancer cells from HB mouse model also obtained the migratory and invasive biology for "species-specific" tissue metastasis to human tissues. Comprehensive analyses suggest that "species-specific" mammary microenvironment of human origin better regulates the biology of human breast cancer cells in our humanized mouse model of breast cancer, which is more consistent with the clinical progress of human breast cancer.

In vivo quantitative magnetization transfer imaging correlates with histology during de- and remyelination in cuprizone-treated mice.

PubMed

Turati, Laura; Moscatelli, Marco; Mastropietro, Alfonso; Dowell, Nicholas G; Zucca, Ileana; Erbetta, Alessandra; Cordiglieri, Chiara; Brenna, Greta; Bianchi, Beatrice; Mantegazza, Renato; Cercignani, Mara; Baggi, Fulvio; Minati, Ludovico

2015-03-01

The pool size ratio measured by quantitative magnetization transfer MRI is hypothesized to closely reflect myelin density, but their relationship has so far been confirmed mostly in ex vivo conditions. We investigate the correspondence between this parameter measured in vivo at 7.0 T, with Black Gold II staining for myelin fibres, and with myelin basic protein and beta-tubulin immunofluorescence in a hybrid longitudinal study of C57BL/6 and SJL/J mice treated with cuprizone, a neurotoxicant causing relatively selective myelin loss followed by spontaneous remyelination upon treatment suspension. Our results confirm that pool size ratio measurements correlate with myelin content, with the correlation coefficient depending on strain and staining method, and demonstrate the in vivo applicability of this MRI technique to experimental mouse models of multiple sclerosis. Copyright © 2015 John Wiley & Sons, Ltd.

Potentiation of antimicrobial photodynamic inactivation mediated by a cationic fullerene by added iodide: in vitro and in vivo studies

PubMed Central

Zhang, Yunsong; Dai, Tianhong; Wang, Min; Vecchio, Daniela; Chiang, Long Y; Hamblin, Michael R

2016-01-01

Background Antimicrobial photodynamic inactivation with fullerenes bearing cationic charges may overcome resistant microbes. Methods & results We synthesized C60-fullerene (LC16) bearing decaquaternary chain and deca-tertiary-amino groups that facilitates electron-transfer reactions via the photoexcited fullerene. Addition of the harmless salt, potassium iodide (10 mM) potentiated the ultraviolet A (UVA) or white light-mediated killing of Gram-negative bacteria Acinetobacter baumannii, Gram-positive methicillin-resistant Staphylococcus aureus and fungal yeast Candida albicans by 1–2+ logs. Mouse model infected with bioluminescent Acinetobacter baumannii gave increased loss of bioluminescence when iodide (10 mM) was combined with LC16 and UVA/white light. Conclusion The mechanism may involve photoinduced electron reduction of 1(C60>)* or 3(C60>)* by iodide producing I· or I2 followed by subsequent intermolecular electron-transfer events of (C60>)−· to produce reactive radicals. PMID:25723093

Genetically engineered T cells to target EGFRvIII expressing glioblastoma.

PubMed

Bullain, Szofia S; Sahin, Ayguen; Szentirmai, Oszkar; Sanchez, Carlos; Lin, Ning; Baratta, Elizabeth; Waterman, Peter; Weissleder, Ralph; Mulligan, Richard C; Carter, Bob S

2009-09-01

Glioblastoma remains a significant therapeutic challenge, warranting further investigation of novel therapies. We describe an immunotherapeutic strategy to treat glioblastoma based on adoptive transfer of genetically modified T-lymphocytes (T cells) redirected to kill EGFRvIII expressing gliomas. We constructed a chimeric immune receptor (CIR) specific to EGFRvIII, (MR1-zeta). After in vitro selection and expansion, MR1-zeta genetically modified primary human T-cells specifically recognized EGFRvIII-positive tumor cells as demonstrated by IFN-gamma secretion and efficient tumor lysis compared to control CIRs defective in EGFRvIII binding (MRB-zeta) or signaling (MR1-delzeta). MR1-zeta expressing T cells also inhibited EGFRvIII-positive tumor growth in vivo in a xenografted mouse model. Successful targeting of EGFRvIII-positive tumors via adoptive transfer of genetically modified T cells may represent a new immunotherapy strategy with great potential for clinical applications.

Tumor-derived exosomes promote tumor self-seeding in hepatocellular carcinoma by transferring miRNA-25-5p to enhance cell motility.

PubMed

Liu, Hao; Chen, Wei; Zhi, Xiao; Chen, En-Jiang; Wei, Tao; Zhang, Jian; Shen, Jian; Hu, Li-Qiang; Zhao, Bin; Feng, Xin-Hua; Bai, Xue-Li; Liang, Ting-Bo

2018-05-22

Tumor self-seeding occurs when circulating malignant cells reinfiltrate the original tumor. The process may breed more aggressive tumor cells, which may contribute to cancer progression. In this study, we observed tumor self-seeding in mouse xenograft models of hepatocellular carcinoma (HCC) for the first time. We confirmed that circulating tumor cell uptake of tumor-derived exosomes, which are increasingly recognized as key instigators of cancer progression by facilitating cell-cell communication, promoted tumor self-seeding by enhancing the invasive and migration capability of recipient HCC cells. Horizontal transfer of exosomal microRNA-25-5p to anoikis-resistant HCC cells significantly enhanced their migratory and invasive abilities, whereas inhibiting microRNA-25-5p alleviated these effects. Our experiments delineate an exosome-based novel pathway employed by functional microRNA from the original tumor cells that can influence the biological fate of circulating tumor cells.

Production of cloned mice and ES cells from adult somatic cells by nuclear transfer: how to improve cloning efficiency?

PubMed

Wakayama, Teruhiko

2007-02-01

Although it has now been 10 years since the first cloned mammals were generated from somatic cells using nuclear transfer (NT), most cloned embryos usually undergo developmental arrest prior to or soon after implantation, and the success rate for producing live offspring by cloning remains below 5%. The low success rate is believed to be associated with epigenetic errors, including abnormal DNA hypermethylation, but the mechanism of "reprogramming" is unclear. We have been able to develop a stable NT method in the mouse in which donor nuclei are directly injected into the oocyte using a piezo-actuated micromanipulator. Especially in the mouse, only a few laboratories can make clones from adult somatic cells, and cloned mice are never successfully produced from most mouse strains. However, this technique promises to be an important tool for future research in basic biology. For example, NT can be used to generate embryonic stem (NT-ES) cell lines from a patient's own somatic cells. We have shown that NT-ES cells are equivalent to ES cells derived from fertilized embryos and that they can be generated relatively easily from a variety of mouse genotypes and cell types of both sexes, even though it may be more difficult to generate clones directly. In general, NT-ES cell techniques are expected to be applied to regenerative medicine; however, this technique can also be applied to the preservation of genetic resources of mouse strain instead of embryos, oocytes and spermatozoa. This review describes how to improve cloning efficiency and NT-ES cell establishment and further applications.

The Mouse Tumor Biology Database: A Comprehensive Resource for Mouse Models of Human Cancer.

PubMed

Krupke, Debra M; Begley, Dale A; Sundberg, John P; Richardson, Joel E; Neuhauser, Steven B; Bult, Carol J

2017-11-01

Research using laboratory mice has led to fundamental insights into the molecular genetic processes that govern cancer initiation, progression, and treatment response. Although thousands of scientific articles have been published about mouse models of human cancer, collating information and data for a specific model is hampered by the fact that many authors do not adhere to existing annotation standards when describing models. The interpretation of experimental results in mouse models can also be confounded when researchers do not factor in the effect of genetic background on tumor biology. The Mouse Tumor Biology (MTB) database is an expertly curated, comprehensive compendium of mouse models of human cancer. Through the enforcement of nomenclature and related annotation standards, MTB supports aggregation of data about a cancer model from diverse sources and assessment of how genetic background of a mouse strain influences the biological properties of a specific tumor type and model utility. Cancer Res; 77(21); e67-70. ©2017 AACR . ©2017 American Association for Cancer Research.

Inflammatory arthritis increases mouse osteoclast precursors with myeloid suppressor function

PubMed Central

Charles, Julia F.; Hsu, Lih-Yun; Niemi, Erene C.; Weiss, Arthur; Aliprantis, Antonios O.; Nakamura, Mary C.

2012-01-01

Increased osteoclastic bone resorption leads to periarticular erosions and systemic osteoporosis in RA patients. Although a great deal is known about how osteoclasts differentiate from precursors and resorb bone, the identity of an osteoclast precursor (OCP) population in vivo and its regulatory role in RA remains elusive. Here, we report the identification of a CD11b–/loLy6Chi BM population with OCP activity in vitro and in vivo. These cells, which can be distinguished from previously characterized precursors in the myeloid lineage, display features of both M1 and M2 monocytes and expand in inflammatory arthritis models. Surprisingly, in one mouse model of RA (adoptive transfer of SKG arthritis), cotransfer of OCP with SKG CD4+ T cells diminished inflammatory arthritis. Similar to monocytic myeloid-derived suppressor cells (M-MDSCs), OCPs suppressed CD4+ and CD8+ T cell proliferation in vitro through the production of NO. This study identifies a BM myeloid precursor population with osteoclastic and T cell–suppressive activity that is expanded in inflammatory arthritis. Therapeutic strategies that prevent the development of OCPs into mature bone-resorbing cells could simultaneously prevent bone resorption and generate an antiinflammatory milieu in the RA joint. PMID:23114597

Foamy Virus Vector-mediated Gene Correction of a Mouse Model of Wiskott–Aldrich Syndrome

PubMed Central

Uchiyama, Toru; Adriani, Marsilio; Jagadeesh, G Jayashree; Paine, Adam; Candotti, Fabio

2012-01-01

The Wiskott–Aldrich syndrome (WAS) is an X-linked disorder characterized by eczema, thrombocytopenia and immunodeficiency. Hematopoietic cell transplantation can cure the disease and gene therapy is being tested as an alternative treatment option. In this study, we assessed the use of foamy virus (FV) vectors as a gene transfer system for WAS, using a Was knockout (KO) mouse model. Preliminary experiments using FV vectors expressing the green fluorescent protein under the transcriptional control of the endogenous WAS promoter or a ubiquitously acting chromatin opening element allowed us to define transduction conditions resulting in high (>40%) and long-term in-vivo marking of blood cells after transplantation. In following experiments, Was KO mice were treated with FV vectors containing the human WAS complementary DNA (cDNA). Transplanted animals expressed the WAS protein (WASp) in T and B lymphocytes, as well as platelets and showed restoration of both T-cell receptor-mediated responses and B-cell migration. We also observed recovery of platelet adhesion and podosome formation in dendritic cells (DCs) of treated mice. These data demonstrate that FV vectors can be effective for hematopoietic stem cell (HSC)-directed gene correction of WAS. PMID:22215016

Imaging the accumulation and suppression of tau pathology using multiparametric MRI

PubMed Central

Holmes, Holly E.; Colgan, Niall; Ismail, Ozama; Ma, Da; Powell, Nick M.; O'Callaghan, James M.; Harrison, Ian F.; Johnson, Ross A.; Murray, Tracey K.; Ahmed, Zeshan; Heggenes, Morton; Fisher, Alice; Cardoso, M.J.; Modat, Marc; Walker-Samuel, Simon; Fisher, Elizabeth M.C.; Ourselin, Sebastien; O'Neill, Michael J.; Wells, Jack A.; Collins, Emily C.; Lythgoe, Mark F.

2016-01-01

Mouse models of Alzheimer's disease have served as valuable tools for investigating pathogenic mechanisms relating to neurodegeneration, including tau-mediated and neurofibrillary tangle pathology—a major hallmark of the disease. In this work, we have used multiparametric magnetic resonance imaging (MRI) in a longitudinal study of neurodegeneration in the rTg4510 mouse model of tauopathy, a subset of which were treated with doxycycline at different time points to suppress the tau transgene. Using this paradigm, we investigated the sensitivity of multiparametric MRI to both the accumulation and suppression of pathologic tau. Tau-related atrophy was discernible from 5.5 months within the cortex and hippocampus. We observed markedly less atrophy in the treated rTg4510 mice, which was enhanced after doxycycline intervention from 3.5 months. We also observed differences in amide proton transfer, cerebral blood flow, and diffusion tensor imaging parameters in the rTg4510 mice, which were significantly less altered after doxycycline treatment. We propose that these non-invasive MRI techniques offer insight into pathologic mechanisms underpinning Alzheimer's disease that may be important when evaluating emerging therapeutics targeting one of more of these processes. PMID:26923415

Adult Mouse DRG Explant and Dissociated Cell Models to Investigate Neuroplasticity and Responses to Environmental Insults Including Viral Infection.

PubMed

Fornaro, Michele; Sharthiya, Harsh; Tiwari, Vaibhav

2018-03-09

This protocol describes an ex vivo model of mouse-derived dorsal root ganglia (DRG) explant and in vitro DRG-derived co-culture of dissociated sensory neurons and glial satellite cells. These are useful and versatile models to investigate a variety of biological responses associated with physiological and pathological conditions of the peripheral nervous system (PNS) ranging from neuron-glial interaction, neuroplasticity, neuroinflammation, and viral infection. The usage of DRG explant is scientifically advantageous compared to simplistic single cells models for multiple reasons. For instance, as an organotypic culture, the DRG explant allows ex vivo transfer of an entire neuronal network including the extracellular microenvironment that play a significant role in all the neuronal and glial functions. Further, DRG explants can also be maintained ex vivo for several days and the culture conditions can be perturbed as desired. In addition, the harvested DRG can be further dissociated into an in vitro co-culture of primary sensory neurons and satellite glial cells to investigate neuronal-glial interaction, neuritogenesis, axonal cone interaction with the extracellular microenvironment, and more general, any aspect associated with the neuronal metabolism. Therefore, the DRG-explant system offers a great deal of flexibility to study a wide array of events related to biological, physiological, and pathological conditions in a cost-effective manner.

Divergent Roles of Interferon-γ and Innate Lymphoid Cells in Innate and Adaptive Immune Cell-Mediated Intestinal Inflammation.

PubMed

Brasseit, Jennifer; Kwong Chung, Cheong K C; Noti, Mario; Zysset, Daniel; Hoheisel-Dickgreber, Nina; Genitsch, Vera; Corazza, Nadia; Mueller, Christoph

2018-01-01

Aberrant interferon gamma (IFNγ) expression is associated with the pathogenesis of numerous autoimmune- and inflammatory disorders, including inflammatory bowel diseases (IBD). However, the requirement of IFNγ for the pathogenesis of chronic intestinal inflammation remains controversial. The aim of this study was thus to investigate the role of IFNγ in experimental mouse models of innate and adaptive immune cell-mediated intestinal inflammation using genetically and microbiota-stabilized hosts. While we find that IFNγ drives acute intestinal inflammation in the anti-CD40 colitis model in an innate lymphoid cell (ILC)-dependent manner, IFNγ secreted by both transferred CD4 T cells and/or cells of the lymphopenic Rag1 -/- recipient mice was dispensable for CD4 T cell-mediated colitis. In the absence of IFNγ, intestinal inflammation in CD4 T cell recipient mice was associated with enhanced IL17 responses; consequently, targeting IL17 signaling in IFNγ-deficient mice reduced T cell-mediated colitis. Intriguingly, in contrast to the anti-CD40 model of colitis, depletion of ILC in the Rag1 -/- recipients of colitogenic CD4 T cells did not prevent induction of colonic inflammation. Together, our findings demonstrate that IFNγ represents an essential, or a redundant, pro-inflammatory cytokine for the induction of intestinal inflammation, depending on the experimental mouse model used and on the nature of the critical disease inducing immune cell populations involved.

Effect of the timing of the first cleavage on the developmental potential of nuclear-transferred mouse oocytes receiving embryonic stem cells.

PubMed

Kobayashi, T; Kato, Y; Tsunoda, Y

2004-09-01

The present study examined whether the timing of the first cleavage has an effect on the in vitro and in vivo developmental potential of nuclear-transferred mouse oocytes receiving embryonic stem cells. First, the timing of the first cleavage and the developmental potential of nuclear-transferred oocytes were examined every hour from 12 to 24 h after the start of culture and compared with in vitro-fertilized oocytes. The developmental potential of in vitro-fertilized oocytes decreased gradually according to the time required for cleavage (84% (32/38) for 15 h to 50% (1/2) for 20 h), but intermediate-cleaved (15-16 h) nuclear-transferred oocytes had a higher potential to develop into blastocysts (55% (17/31) to 67% (45/67) versus 0-43% (6/14)]. Second the nuclear-transferred oocytes were divided into three groups according to the timing of the first cleavage; each group was cultured to blastocysts in vitro, and then transferred to recipients. The potential of intermediate-cleaved oocytes (15-16 h) to develop into blastocysts was significantly higher than fast-cleaved (before 15 h) and slow-cleaved (after 16 h) oocytes (65, 46, and 37%). The proportion of fetuses on Day 10.5 of pregnancy was highest in the intermediate-cleaved group (4 versus 2 and 1%, respectively) and a full-term fetus was obtained from this group. The present study demonstrated that the timing of the first cleavage could be used to determine the potential of nuclear-transferred oocytes with embryonic stem cells to develop to the blastocyst stage in vitro, but not to determine post-implantation viability after transfer to recipients.

HSP70: therapeutic potential in acute and chronic cardiac disease settings.

PubMed

Bernardo, Bianca C; Weeks, Kate L; Patterson, Natalie L; McMullen, Julie R

2016-12-01

Heat shock proteins are a family of proteins that are produced by cells in response to exposure to stressful conditions. The best studied heat shock protein is HSP70, which is known to act as a molecular chaperone to maintain cellular homeostasis and inhibit protein aggregation in response to stress. While early animal studies suggested that increasing HSP70 in the heart (using a transgenic, gene transfer or pharmacological approach) provided cardiac protection against acute cardiac stress, recent studies have found no benefit of increasing HSP70 in mouse models of chronic cardiac stress. As HSP70 has been considered a potential therapeutic target, it is important to comprehensively assess HSP70 therapies in preclinical models of acute and chronic cardiac disease.

In Vivo Fluorescence Resonance Energy Transfer Imaging for Targeted Anti-Cancer Drug Delivery Kinetics

NASA Astrophysics Data System (ADS)

Webb, Kevin; Gaind, Vaibhav; Tsai, Hsiaorho; Bentz, Brian; Chelvam, Venkatesh; Low, Philip

2012-02-01

We describe an approach for the evaluation of targeted anti-cancer drug delivery in vivo. The method emulates the drug release and activation process through acceptor release from a targeted donor-acceptor pair that exhibits fluorescence resonance energy transfer (FRET). In this case, folate targeting of the cancer cells is used - 40 % of all human cancers, including ovarian, lung, breast, kidney, brain and colon cancer, over-express folate receptors. We demonstrate the reconstruction of the spatially-dependent FRET parameters in a mouse model and in tissue phantoms. The FRET parameterization is incorporated into a source for a diffusion equation model for photon transport in tissue, in a variant of optical diffusion tomography (ODT) called FRET-ODT. In addition to the spatially-dependent tissue parameters in the diffusion model (absorption and diffusion coefficients), the FRET parameters (donor-acceptor distance and yield) are imaged as a function of position. Modulated light measurements are made with various laser excitation positions and a gated camera. More generally, our method provides a new vehicle for studying disease at the molecular level by imaging FRET parameters in deep tissue, and allows the nanometer FRET ruler to be utilized in deep tissue.

Complementation of DNA repair defect in xeroderma pigmentosum cells of group C by the transfer of human chromosome 5

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kaur, G.P.; Athwal, R.S.

1993-01-01

Complementation of DNA excision repair defect in xeroderma pigmentosum cells of group C (XP-C) has been achieved by the transfer of human chromosome 5. Individual human chromosomes tagged with a selectable marker were transferred to XP-C cells by microcell fusion from mouse-human hybrid cell lines each bearing a single different human chromosome. Analysis of the chromosome transfer clones revealed that introduction of chromosome 5 into XP-C cells corrected the DNA repair defect as well as UV-sensitive phenotypes, while chromosomes 2, 6, 7, 9, 13, 15, 17, and 21 failed to complement. The introduced chromosome 5 in complemented UV[sup r] clonesmore » was distinguished from the parental XP-C chromosomes by polymorphism for dinucleotide (CA)[sub n] repeats at two loci, D5S117 and D5S209. In addition, an intact marked chromosome 5 was rescued into mouse cells from a complemented UV[sup r] clone by microcell fusion. Five subclones of a complemented clone that had lost the marked chromosome 5 exhibited UV-sensitive and repair-deficient phenotypes identical to parental XP-C cells. Concordant loss of the transferred chromosome and reappearance of XP-C phenotype further confirmed the presence of a DNA repair gene on human chromosome 5. 38 refs., 7 figs., 1 tab.« less

Worm Proteins of Schistosoma mansoni Reduce the Severity of Experimental Chronic Colitis in Mice by Suppressing Colonic Proinflammatory Immune Responses

PubMed Central

Heylen, Marthe; Ruyssers, Nathalie E.; De Man, Joris G.; Timmermans, Jean-Pierre; Pelckmans, Paul A.; Moreels, Tom G.; De Winter, Benedicte Y.

2014-01-01

Although helminthic therapy as a possible new option to treat inflammatory bowel disease is a well-established concept by now, the search for immunomodulatory helminth-derived compounds and their mechanisms of action is still ongoing. We investigated the therapeutic potential and the underlying immunological mechanisms of Schistosoma mansoni soluble worm proteins (SmSWP) in an adoptive T cell transfer mouse model of chronic colitis. Both a curative and a preventive treatment protocol were included in this study. The curative administration of SmSWP (started when colitis was established), resulted in a significant improvement of the clinical disease score, colonoscopy, macroscopic and microscopic inflammation score, colon length and myeloperoxidase activity. The therapeutic potential of the preventive SmSWP treatment (started before colitis was established), was less pronounced compared with the curative SmSWP treatment but still resulted in an improved clinical disease score, body weight loss, colon length and microscopic inflammation score. Both the curative and preventive SmSWP treatment downregulated the mRNA expression of the proinflammatory cytokines IFN-γ and IL-17A and upregulated the mRNA expression of the anti-inflammatory cytokine IL-4 in the colon at the end of the experiment. This colonic immunomodulatory effect of SmSWP could not be confirmed at the protein level. Moreover, the effect of SmSWP appeared to be a local colonic phenomenon, since the flow cytometric T cell characterization of the mesenteric lymph nodes and the cytokine measurements in the serum did not reveal any effect of SmSWP treatment. In conclusion, SmSWP treatment reduced the severity of colitis in the adoptive transfer mouse model via the suppression of proinflammatory cytokines and the induction of an anti-inflammatory response in the colon. PMID:25313594

Clarithromycin expands CD11b+Gr-1+ cells via the STAT3/Bv8 axis to ameliorate lethal endotoxic shock and post-influenza bacterial pneumonia

PubMed Central

Fujii, Hideki; Yagi, Kazuma; Suzuki, Shoji; Hegab, Ahmed E.; Tasaka, Sadatomo; Nakamoto, Nobuhiro; Iwata, Satoshi; Honda, Kenya; Kanai, Takanori; Hasegawa, Naoki; Betsuyaku, Tomoko

2018-01-01

Macrolides are used to treat various inflammatory diseases owing to their immunomodulatory properties; however, little is known about their precise mechanism of action. In this study, we investigated the functional significance of the expansion of myeloid-derived suppressor cell (MDSC)-like CD11b+Gr-1+ cells in response to the macrolide antibiotic clarithromycin (CAM) in mouse models of shock and post-influenza pneumococcal pneumonia as well as in humans. Intraperitoneal administration of CAM markedly expanded splenic and lung CD11b+Gr-1+ cell populations in naïve mice. Notably, CAM pretreatment enhanced survival in a mouse model of lipopolysaccharide (LPS)-induced shock. In addition, adoptive transfer of CAM-treated CD11b+Gr-1+ cells protected mice against LPS-induced lethality via increased IL-10 expression. CAM also improved survival in post-influenza, CAM-resistant pneumococcal pneumonia, with improved lung pathology as well as decreased interferon (IFN)-γ and increased IL-10 levels. Adoptive transfer of CAM-treated CD11b+Gr-1+ cells protected mice from post-influenza pneumococcal pneumonia. Further analysis revealed that the CAM-induced CD11b+Gr-1+ cell expansion was dependent on STAT3-mediated Bv8 production and may be facilitated by the presence of gut commensal microbiota. Lastly, an analysis of peripheral blood obtained from healthy volunteers following oral CAM administration showed a trend toward the expansion of human MDSC-like cells (Lineage−HLA-DR−CD11b+CD33+) with increased arginase 1 mRNA expression. Thus, CAM promoted the expansion of a unique population of immunosuppressive CD11b+Gr-1+ cells essential for the immunomodulatory properties of macrolides. PMID:29621339

Capsaicin-sensitive sensory nerves exert complex regulatory functions in the serum-transfer mouse model of autoimmune arthritis.

PubMed

Borbély, Éva; Botz, Bálint; Bölcskei, Kata; Kenyér, Tibor; Kereskai, László; Kiss, Tamás; Szolcsányi, János; Pintér, Erika; Csepregi, Janka Zsófia; Mócsai, Attila; Helyes, Zsuzsanna

2015-03-01

The K/BxN serum-transfer arthritis is a widely-used translational mouse model of rheumatoid arthritis, in which the immunological components have thoroughly been investigated. In contrast, little is known about the role of sensory neural factors and the complexity of neuro-immune interactions. Therefore, we analyzed the involvement of capsaicin-sensitive peptidergic sensory nerves in autoantibody-induced arthritis with integrative methodology. Arthritogenic K/BxN or control serum was injected to non-pretreated mice or resiniferatoxin (RTX)-pretreated animals where capsaicin-sensitive nerves were inactivated. Edema, touch sensitivity, noxious heat threshold, joint function, body weight and clinical arthritis severity scores were determined repeatedly throughout two weeks. Micro-CT and in vivo optical imaging to determine matrix-metalloproteinase (MMP) and neutrophil-derived myeloperoxidase (MPO) activities, semiquantitative histopathological scoring and radioimmunoassay to measure somatostatin in the joint homogenates were also performed. In RTX-pretreated mice, the autoantibody-induced joint swelling, arthritis severity score, MMP and MPO activities, as well as histopathological alterations were significantly greater compared to non-pretreated animals. Self-control quantification of the bone mass revealed decreased values in intact female mice, but significantly greater arthritis-induced pathological bone formation after RTX-pretreatment. In contrast, mechanical hyperalgesia from day 10 was smaller after inactivating capsaicin-sensitive afferents. Although thermal hyperalgesia did not develop, noxious heat threshold was significantly higher following RTX pretreatment. Somatostatin-like immunoreactivity elevated in the tibiotarsal joints in non-pretreated, which was significantly less in RTX-pretreated mice. Although capsaicin-sensitive sensory nerves mediate mechanical hyperalgesia in the later phase of autoantibody-induced chronic arthritis, they play important anti-inflammatory roles at least partially through somatostatin release. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Capsaicin-sensitive sensory nerves exert complex regulatory functions in the serum-transfer mouse model of autoimmune arthritis

PubMed Central

Borbély, Éva; Botz, Bálint; Bölcskei, Kata; Kenyér, Tibor; Kereskai, László; Kiss, Tamás; Szolcsányi, János; Pintér, Erika; Csepregi, Janka Zsófia; Mócsai, Attila; Helyes, Zsuzsanna

2015-01-01

Objective The K/BxN serum-transfer arthritis is a widely-used translational mouse model of rheumatoid arthritis, in which the immunological components have thoroughly been investigated. In contrast, little is known about the role of sensory neural factors and the complexity of neuro–immune interactions. Therefore, we analyzed the involvement of capsaicin-sensitive peptidergic sensory nerves in autoantibody-induced arthritis with integrative methodology. Methods Arthritogenic K/BxN or control serum was injected to non-pretreated mice or resiniferatoxin (RTX)-pretreated animals where capsaicin-sensitive nerves were inactivated. Edema, touch sensitivity, noxious heat threshold, joint function, body weight and clinical arthritis severity scores were determined repeatedly throughout two weeks. Micro-CT and in vivo optical imaging to determine matrix-metalloproteinase (MMP) and neutrophil-derived myeloperoxidase (MPO) activities, semiquantitative histopathological scoring and radioimmunoassay to measure somatostatin in the joint homogenates were also performed. Results In RTX-pretreated mice, the autoantibody-induced joint swelling, arthritis severity score, MMP and MPO activities, as well as histopathological alterations were significantly greater compared to non-pretreated animals. Self-control quantification of the bone mass revealed decreased values in intact female mice, but significantly greater arthritis-induced pathological bone formation after RTX-pretreatment. In contrast, mechanical hyperalgesia from day 10 was smaller after inactivating capsaicin-sensitive afferents. Although thermal hyperalgesia did not develop, noxious heat threshold was significantly higher following RTX pretreatment. Somatostatin-like immunoreactivity elevated in the tibiotarsal joints in non-pretreated, which was significantly less in RTX-pretreated mice. Conclusions Although capsaicin-sensitive sensory nerves mediate mechanical hyperalgesia in the later phase of autoantibody-induced chronic arthritis, they play important anti-inflammatory roles at least partially through somatostatin release. PMID:25524130

Nanoamplifiers synthesized from gadolinium and gold nanocomposites for magnetic resonance imaging

NASA Astrophysics Data System (ADS)

Tian, Xiumei; Shao, Yuanzhi; He, Haoqiang; Liu, Huan; Shen, Yingying; Huang, Wenlin; Li, Li

2013-03-01

We have synthesized an efficient and highly sensitive nanoamplifier composed of gadolinium-doped silica nanoparticles and gold nanoparticles (AuNPs). Magnetic resonance imaging (MRI) in vitro and in vivo assays revealed enhancement of signal sensitivity, which may be explained by electron transfer between water and gadolinium-doped nanoparticles, apparent in the presence of gold. In vitro and in vivo evaluation demonstrated nanoamplifier incurred minimal cytotoxicity and immunotoxicity, increased stability, and gradual excretion patterns. Tumor targeted properties were preliminarily determined when the nanoamplifier was injected into mouse models of colon cancer liver metastasis. Furthermore, although AuNPs departed from the nanoamplifiers in specific mice tissues, optical and magnetic resonance imaging was efficient, especially in metastatic tumors. These assays validate our nanoamplifier as an effective MRI signal enhancer with sensitive cancer diagnosis potential.We have synthesized an efficient and highly sensitive nanoamplifier composed of gadolinium-doped silica nanoparticles and gold nanoparticles (AuNPs). Magnetic resonance imaging (MRI) in vitro and in vivo assays revealed enhancement of signal sensitivity, which may be explained by electron transfer between water and gadolinium-doped nanoparticles, apparent in the presence of gold. In vitro and in vivo evaluation demonstrated nanoamplifier incurred minimal cytotoxicity and immunotoxicity, increased stability, and gradual excretion patterns. Tumor targeted properties were preliminarily determined when the nanoamplifier was injected into mouse models of colon cancer liver metastasis. Furthermore, although AuNPs departed from the nanoamplifiers in specific mice tissues, optical and magnetic resonance imaging was efficient, especially in metastatic tumors. These assays validate our nanoamplifier as an effective MRI signal enhancer with sensitive cancer diagnosis potential. Electronic supplementary information (ESI) available: Protocols for the characterization, immunotoxicity and pharmacokinetics analyses. Additional supporting figures. See DOI: 10.1039/c3nr00170a

Interluekin-12 enhances the function and anti-tumor activity in murine and human CD8+ T cells

PubMed Central

Rubinstein, Mark P.; Su, Ee Wern; Suriano, Samantha; Cloud, Colleen A.; Andrijauskaite, Kristina; Kesarwani, Pravin; Schwartz, Kristina M.; Williams, Katelyn; Johnson, C. Bryce; Li, Mingli; Scurti, Gina M.; Salem, Mohamed L.; Paulos, Chrystal M.; Garrett-Mayer, Elizabeth; Mehrotra, Shikhar; Cole, David J.

2016-01-01

Mouse CD8+ T cells conditioned with Interleukin (IL)-12 ex vivo mediate the potent regression of established melanoma when transferred into lymphodepleted mice. However, the quantitative and qualitative changes induced by IL-12 in the responding mouse CD8+ T cells have not been well defined. Moreover, the mechanisms by which IL-12-conditioning impacts human CD8+ T cells, and how such cells might be expanded prior to infusion into patients is not known. We found that ex vivo IL-12-conditioning of mouse CD8+ T cells led to a 10- to 100-fold increase in persistence and anti-tumor efficacy upon adoptive transfer into lymphodepleted mice. The enhancing effect of IL-12 was associated with maintenance of functional avidity. Importantly, in the context of ongoing ACT clinical trials, human CD8+ T cells genetically modified with a tyrosinase-specific T-cell receptor exhibited significantly enhanced functional activity when conditioned with IL-12 as indicated by heightened granzyme B expression and elevated peptide-specific CD107a degranulation. This effect was sustainable despite the 20 days of in vitro cellular expansion required to expand cells over 1,000-fold allowing adequate cell numbers for administration to cancer patients. Overall, these findings support the efficacy and feasibility of ex vivo IL-12-conditioning of TCR-modified human CD8+ T cells for adoptive transfer and cancer therapy. PMID:25676709

Drug discovery in prostate cancer mouse models.

PubMed

Valkenburg, Kenneth C; Pienta, Kenneth J

2015-01-01

The mouse is an important, though imperfect, organism with which to model human disease and to discover and test novel drugs in a preclinical setting. Many experimental strategies have been used to discover new biological and molecular targets in the mouse, with the hopes of translating these discoveries into novel drugs to treat prostate cancer in humans. Modeling prostate cancer in the mouse, however, has been challenging, and often drugs that work in mice have failed in human trials. The authors discuss the similarities and differences between mice and men; the types of mouse models that exist to model prostate cancer; practical questions one must ask when using a mouse as a model; and potential reasons that drugs do not often translate to humans. They also discuss the current value in using mouse models for drug discovery to treat prostate cancer and what needs are still unmet in field. With proper planning and following practical guidelines by the researcher, the mouse is a powerful experimental tool. The field lacks genetically engineered metastatic models, and xenograft models do not allow for the study of the immune system during the metastatic process. There remain several important limitations to discovering and testing novel drugs in mice for eventual human use, but these can often be overcome. Overall, mouse modeling is an essential part of prostate cancer research and drug discovery. Emerging technologies and better and ever-increasing forms of communication are moving the field in a hopeful direction.

Orthology for comparative genomics in the mouse genome database.

PubMed

Dolan, Mary E; Baldarelli, Richard M; Bello, Susan M; Ni, Li; McAndrews, Monica S; Bult, Carol J; Kadin, James A; Richardson, Joel E; Ringwald, Martin; Eppig, Janan T; Blake, Judith A

2015-08-01

The mouse genome database (MGD) is the model organism database component of the mouse genome informatics system at The Jackson Laboratory. MGD is the international data resource for the laboratory mouse and facilitates the use of mice in the study of human health and disease. Since its beginnings, MGD has included comparative genomics data with a particular focus on human-mouse orthology, an essential component of the use of mouse as a model organism. Over the past 25 years, novel algorithms and addition of orthologs from other model organisms have enriched comparative genomics in MGD data, extending the use of orthology data to support the laboratory mouse as a model of human biology. Here, we describe current comparative data in MGD and review the history and refinement of orthology representation in this resource.

Molecular Biology: Conference on Genetic Engineering Techniques (2nd) Held in London (United Kingdom) on 20-21 November 1986.

DTIC Science & Technology

1987-05-27

system in Chinese t-PA to be a serine protease of 327 amino ovary hamster cells. Precise yields from acids in length. The protein appears, high-level...ham- ster or mouse cell line, allowing the differentiation of human and hamster or ________ mouse clones by hybridization with total human DNA or...appropriate lo- functional protein when transferred into cation downstream of a strong promoter in baby hamster kidney (BHK) cells or rat place of one or

Fluorescent Labeling and Biodistribution of Latex Nanoparticles Formed by Surfactant-Free RAFT Emulsion Polymerization.

PubMed

Poon, Cheuk Ka; Tang, Owen; Chen, Xin-Ming; Kim, Byung; Hartlieb, Matthias; Pollock, Carol A; Hawkett, Brian S; Perrier, Sébastien

2017-10-01

The authors report the preparation of a novel range of functional polyacrylamide stabilized polystyrene nanoparticles, obtained by surfactant-free reversible addition-fragmentation chain transfer (RAFT) emulsion polymerization, their fluorescent tagging, cellular uptake, and biodistribution. The authors show the versatility of the RAFT emulsion process for the design of functional nanoparticles of well-defined size that can be used as drug delivery vectors. Functionalization with a fluorescent tag offers a useful visualization tool for tracing, localization, and clearance studies of these carriers in biological models. The studies are carried out by labeling the sterically stabilized latex particles chemically with rhodamine B. The fluorescent particles are incubated in a healthy human renal proximal tubular cell line model, and intravenously injected into a mouse model. Cellular localization and biodistribution of these particles on the biological models are explored. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Genetically Engineered Mouse Models for Studying Inflammatory Bowel Disease

PubMed Central

Mizoguchi, Atsushi; Takeuchi, Takahito; Himuro, Hidetomo; Okada, Toshiyuki; Mizoguchi, Emiko

2015-01-01

Inflammatory bowel disease (IBD) is a chronic intestinal inflammatory condition that is mediated by very complex mechanisms controlled by genetic, immune, and environmental factors. More than 74 kinds of genetically engineered mouse strains have been established since 1993 for studying IBD. Although mouse models cannot fully reflect human IBD, they have provided significant contributions for not only understanding the mechanism, but also developing new therapeutic means for IBD. Indeed, 20 kinds of genetically engineered mouse models carry the susceptibility genes identified in human IBD, and the functions of some other IBD susceptibility genes have also been dissected out using mouse models. Cutting-edge technologies such as cell-specific and inducible knockout systems, which were recently employed to mouse IBD models, have further enhanced the ability of investigators to provide important and unexpected rationales for developing new therapeutic strategies for IBD. In this review article, we briefly introduce 74 kinds of genetically engineered mouse models that spontaneously develop intestinal inflammation. PMID:26387641

Altered social behavior and neuronal development in mice lacking the Uba6-Use1 ubiquitin transfer system

PubMed Central

Lee, Peter C. W.; Dodart, Jean-Cosme; Aron, Liviu; Finley, Lydia W.; Bronson, Roderick T.; Haigis, Marcia C.; Yankner, Bruce A.; Harper, J. Wade

2013-01-01

The Uba6 (E1)-Use1 (E2) ubiquitin transfer cascade is a poorly understood alternative arm of the ubiquitin proteasome system (UPS) required for mouse embryonic development, independent of the canonical Uba1-E2-E3 pathway. Loss of neuronal Uba6 during embryonic development results in altered patterning of neurons in the hippocampus and the amygdala, decreased dendritic spine density, and numerous behavioral disorders. The levels of the E3 ubiquitin ligase Ube3a (E6-AP) and Shank3, both linked with dendritic spine function, are elevated in the amygdala of Uba6-deficient mice, while levels of the Ube3a substrate Arc are reduced. Uba6 and Use1 promote proteasomal turnover of Ube3a in mouse embryo fibroblasts (MEFs) and catalyze Ube3a ubiquitylation in vitro. These activities occur in parallel with an independent pathway involving Uba1-UbcH7, but in a spatially distinct manner in MEFs. These data reveal an unanticipated role for Uba6 in neuronal development, spine architecture, mouse behavior, and turnover of Ube3a. PMID:23499007

Assessment of the developmental totipotency of neural cells in the cerebral cortex of mouse embryo by nuclear transfer

PubMed Central

Yamazaki, Yukiko; Makino, Hatsune; Hamaguchi-Hamada, Kayoko; Hamada, Shun; Sugino, Hidehiko; Kawase, Eihachiro; Miyata, Takaki; Ogawa, Masaharu; Yanagimachi, Ryuzo; Yagi, Takeshi

2001-01-01

When neural cells were collected from the entire cerebral cortex of developing mouse fetuses (15.5–17.5 days postcoitum) and their nuclei were transferred into enucleated oocytes, 5.5% of the reconstructed oocytes developed into normal offspring. This success rate was the highest among all previous mouse cloning experiments that used somatic cells. Forty-four percent of live embryos at 10.5 days postcoitum were morphologically normal when premature and early-postmitotic neural cells from the ventricular side of the cortex were used. In contrast, the majority (95%) of embryos were morphologically abnormal (including structural abnormalities in the neural tube) when postmitotic-differentiated neurons from the pial side of the cortex were used for cloning. Whereas 4.3% of embryos cloned with ventricular-side cells developed into healthy offspring, only 0.5% of those cloned with differentiated neurons in the pial side did so. These facts seem to suggest that the nuclei of neural cells in advanced stages of differentiation had lost their developmental totipotency. The underlying mechanism for this developmental limitation could be somatic DNA rearrangements in differentiating neural cells. PMID:11698647

Genome-wide expression profiling of five mouse models identifies similarities and differences with human psoriasis.

PubMed

Swindell, William R; Johnston, Andrew; Carbajal, Steve; Han, Gangwen; Wohn, Christian; Lu, Jun; Xing, Xianying; Nair, Rajan P; Voorhees, John J; Elder, James T; Wang, Xiao-Jing; Sano, Shigetoshi; Prens, Errol P; DiGiovanni, John; Pittelkow, Mark R; Ward, Nicole L; Gudjonsson, Johann E

2011-04-04

Development of a suitable mouse model would facilitate the investigation of pathomechanisms underlying human psoriasis and would also assist in development of therapeutic treatments. However, while many psoriasis mouse models have been proposed, no single model recapitulates all features of the human disease, and standardized validation criteria for psoriasis mouse models have not been widely applied. In this study, whole-genome transcriptional profiling is used to compare gene expression patterns manifested by human psoriatic skin lesions with those that occur in five psoriasis mouse models (K5-Tie2, imiquimod, K14-AREG, K5-Stat3C and K5-TGFbeta1). While the cutaneous gene expression profiles associated with each mouse phenotype exhibited statistically significant similarity to the expression profile of psoriasis in humans, each model displayed distinctive sets of similarities and differences in comparison to human psoriasis. For all five models, correspondence to the human disease was strong with respect to genes involved in epidermal development and keratinization. Immune and inflammation-associated gene expression, in contrast, was more variable between models as compared to the human disease. These findings support the value of all five models as research tools, each with identifiable areas of convergence to and divergence from the human disease. Additionally, the approach used in this paper provides an objective and quantitative method for evaluation of proposed mouse models of psoriasis, which can be strategically applied in future studies to score strengths of mouse phenotypes relative to specific aspects of human psoriasis.

Sex reversal in the mouse (Mus musculus) is caused by a recurrent nonreciprocal crossover involving the x and an aberrant y chromosome.

PubMed

Singh, L; Jones, K W

1982-02-01

Satellite DNA (Bkm) from the W sex-determining chromosome of snakes, which is related to sequences on the mouse Y chromosome, has been used to analyze the DNA and chromosomes of sex-reversed (Sxr) XXSxr male mice. Such mice exhibit a male-specific Southern blot Bkm hybridization pattern, consistent with the presence of Y-chromosome DNA. In situ hybridization of Bkm to chromosomes of XXSxr mice shows an aberrant concentration of related sequences on the distal terminus of a large mouse chromosome. The XYSxr carrier male, however, shows a pair of small chromosomes, which are presumed to be aberrant Y derivatives. Meiosis in the XYSxr mouse involves transfer of chromatin rich in Bkm-related DNA from the Y-Y1 complex to the X distal terminus. We suggest that this event is responsible for the transmission of the Sxr trait.

Near-infrared laser irradiation improves the development of mouse pre-implantation embryos.

PubMed

Yokoo, Masaki; Mori, Miho

2017-05-27

The aim of the present study was to assess the effects of near-infrared laser irradiation on the in vitro development of mouse embryos. Female ICR mice were superovulated with pregnant mare serum gonadotropin and human chorionic gonadotropin (hCG), and mated with male mice. Two-cell stage embryos were collected 40 h after administering hCG and cultured in M16 medium. Two-cell embryos (0 h after culture), 8-cell embryos (approx. 30 h after culture), morula (approx. 48 h after culture), and blastocysts (approx. 73 h after culture) were irradiated at 904 nm for 60 s. These embryos were cultured in a time-lapse monitoring system and the timing of blastocyst hatching was evaluated. Some of the irradiated blastocysts were transferred to the uterine horns of pseudopregnant recipients immediately after irradiation. Pregnancy rates, and offspring growth and fertility, were evaluated. Near-infrared laser irradiation increased the speed of in vitro mouse embryo development. In irradiated blastocysts, hatching was faster than in control (non-irradiated) blastocysts (18.4 vs. 28.2 h, P < 0.05). When 195 irradiated blastocysts were transferred to 18 pseudopregnant mice, all became pregnant and 92 (47.2%) normal-looking pups were born alive. When 182 control blastocysts were transferred to 17 pseudopregnant mice, 14 (82.4%) became pregnant and 54 (29.7%) normal-looking pups were born alive. The growth trajectories (up to 5 weeks) of offspring from irradiated blastocysts were similar to those from control blastocysts. Second generation offspring from transplanted animals were all fertile. These results indicate that near-infrared laser irradiation improves the quality of mouse embryo development in vitro, and increases the live birth rate without affecting the normality of the offspring. Thus, the near-infrared laser method may enhance the quality of embryos and contribute to improvements in reproductive technologies in mammals. Copyright © 2017 Elsevier Inc. All rights reserved.

Periodontitis induced by Porphyromonas gingivalis drives periodontal microbiota dysbiosis and insulin resistance via an impaired adaptive immune response

PubMed Central

Blasco-Baque, Vincent; Garidou, Lucile; Pomié, Céline; Escoula, Quentin; Loubieres, Pascale; Le Gall-David, Sandrine; Lemaitre, Mathieu; Nicolas, Simon; Klopp, Pascale; Waget, Aurélie; Azalbert, Vincent; Colom, André; Bonnaure-Mallet, Martine; Kemoun, Philippe; Serino, Matteo; Burcelin, Rémy

2017-01-01

Objective To identify a causal mechanism responsible for the enhancement of insulin resistance and hyperglycaemia following periodontitis in mice fed a fat-enriched diet. Design We set-up a unique animal model of periodontitis in C57Bl/6 female mice by infecting the periodontal tissue with specific and alive pathogens like Porphyromonas gingivalis (Pg), Fusobacterium nucleatum and Prevotella intermedia. The mice were then fed with a diabetogenic/non-obesogenic fat-enriched diet for up to 3 months. Alveolar bone loss, periodontal microbiota dysbiosis and features of glucose metabolism were quantified. Eventually, adoptive transfer of cervical (regional) and systemic immune cells was performed to demonstrate the causal role of the cervical immune system. Results Periodontitis induced a periodontal microbiota dysbiosis without mainly affecting gut microbiota. The disease concomitantly impacted on the regional and systemic immune response impairing glucose metabolism. The transfer of cervical lymph-node cells from infected mice to naive recipients guarded against periodontitis-aggravated metabolic disease. A treatment with inactivated Pg prior to the periodontal infection induced specific antibodies against Pg and protected the mouse from periodontitis-induced dysmetabolism. Finally, a 1-month subcutaneous chronic infusion of low rates of lipopolysaccharides from Pg mimicked the impact of periodontitis on immune and metabolic parameters. Conclusions We identified that insulin resistance in the high-fat fed mouse is enhanced by pathogen-induced periodontitis. This is caused by an adaptive immune response specifically directed against pathogens and associated with a periodontal dysbiosis. PMID:26838600

Potentiating Tissue-Resident Type 2 Innate Lymphoid Cells by IL-33 to Prevent Renal Ischemia-Reperfusion Injury.

PubMed

Cao, Qi; Wang, Yiping; Niu, Zhiguo; Wang, Chengshi; Wang, Ruifeng; Zhang, Zhiqiang; Chen, Titi; Wang, Xin Maggie; Li, Qing; Lee, Vincent W S; Huang, Qingsong; Tan, Jing; Guo, Minghao; Wang, Yuan Min; Zheng, Guoping; Yu, Di; Alexander, Stephen I; Wang, Hui; Harris, David C H

2018-03-01

The IL-33-type 2 innate lymphoid cell (ILC2) axis has an important role in tissue homeostasis, inflammation, and wound healing. However, the relative importance of this innate immune pathway for immunotherapy against inflammation and tissue damage remains unclear. Here, we show that treatment with recombinant mouse IL-33 prevented renal structural and functional injury and reduced mortality in mice subjected to ischemia-reperfusion injury (IRI). Compared with control-treated IRI mice, IL-33-treated IRI mice had increased levels of IL-4 and IL-13 in serum and kidney and more ILC2, regulatory T cells (Tregs), and anti-inflammatory (M2) macrophages. Depletion of ILC2, but not Tregs, substantially abolished the protective effect of IL-33 on renal IRI. Adoptive transfer of ex vivo -expanded ILC2 prevented renal injury in mice subjected to IRI. This protective effect associated with induction of M2 macrophages in kidney and required ILC2 production of amphiregulin. Treatment of mice with IL-33 or ILC2 after IRI was also renoprotective. Furthermore, in a humanized mouse model of renal IRI, treatment with human IL-33 or transfer of ex vivo -expanded human ILC2 ameliorated renal IRI. This study has uncovered a major protective role of the IL-33-ILC2 axis in renal IRI that could be potentiated as a therapeutic strategy. Copyright © 2018 by the American Society of Nephrology.

Mesenchymal stem cells alleviate oxidative stress-induced mitochondrial dysfunction in the airways.

PubMed

Li, Xiang; Michaeloudes, Charalambos; Zhang, Yuelin; Wiegman, Coen H; Adcock, Ian M; Lian, Qizhou; Mak, Judith C W; Bhavsar, Pankaj K; Chung, Kian Fan

2018-05-01

Oxidative stress-induced mitochondrial dysfunction can contribute to inflammation and remodeling in patients with chronic obstructive pulmonary disease (COPD). Mesenchymal stem cells protect against lung damage in animal models of COPD. It is unknown whether these effects occur through attenuating mitochondrial dysfunction in airway cells. We sought to examine the effect of induced pluripotent stem cell-derived mesenchymal stem cells (iPSC-MSCs) on oxidative stress-induce mitochondrial dysfunction in human airway smooth muscle cells (ASMCs) in vitro and in mouse lungs in vivo. ASMCs were cocultured with iPSC-MSCs in the presence of cigarette smoke medium (CSM), and mitochondrial reactive oxygen species (ROS) levels, mitochondrial membrane potential (ΔΨm), and apoptosis were measured. Conditioned medium from iPSC-MSCs and transwell cocultures were used to detect any paracrine effects. The effect of systemic injection of iPSC-MSCs on airway inflammation and hyperresponsiveness in ozone-exposed mice was also investigated. Coculture of iPSC-MSCs with ASMCs attenuated CSM-induced mitochondrial ROS, apoptosis, and ΔΨm loss in ASMCs. iPSC-MSC-conditioned medium or transwell cocultures with iPSC-MSCs reduced CSM-induced mitochondrial ROS but not ΔΨm or apoptosis in ASMCs. Mitochondrial transfer from iPSC-MSCs to ASMCs was observed after direct coculture and was enhanced by CSM. iPSC-MSCs attenuated ozone-induced mitochondrial dysfunction, airway hyperresponsiveness, and inflammation in mouse lungs. iPSC-MSCs offered protection against oxidative stress-induced mitochondrial dysfunction in human ASMCs and in mouse lungs while reducing airway inflammation and hyperresponsiveness. These effects are, at least in part, dependent on cell-cell contact, which allows for mitochondrial transfer, and paracrine regulation. Therefore iPSC-MSCs show promise as a therapy for oxidative stress-dependent lung diseases, such as COPD. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

77 FR 26660 - Guidelines for the Transfer of Excess Computers or Other Technical Equipment Pursuant to Section...

Federal Register 2010, 2011, 2012, 2013, 2014

2012-05-07

....usda.gov . SUPPLEMENTARY INFORMATION: A. Background A proposed rule was published in the Federal.... Computers or other technical equipment means central processing units, laptops, desktops, computer mouses...

Optimizing mouse models of neurodegenerative disorders: are therapeutics in sight?

PubMed

Lutz, Cathleen M; Osborne, Melissa A

2013-01-01

The genomic and biologic conservation between mice and humans, along with our increasing ability to manipulate the mouse genome, places the mouse as a premier model for deciphering disease mechanisms and testing potential new therapies. Despite these advantages, mouse models of neurodegenerative disease are sometimes difficult to generate and can present challenges that must be carefully addressed when used for preclinical studies. For those models that do exist, the standardization and optimization of the models is a critical step in ensuring success in both basic research and preclinical use. This review looks back on the history of model development for neurodegenerative diseases and highlights the key strategies that have been learned in order to improve the design, development and use of mouse models in the study of neurodegenerative disease.

Live embryo imaging to follow cell cycle and chromosomes stability after nuclear transfer.

PubMed

Balbach, Sebastian T; Boiani, Michele

2015-01-01

Nuclear transfer (NT) into mouse oocytes yields a transcriptionally and functionally heterogeneous population of cloned embryos. Most studies of NT embryos consider only embryos at predefined key stages (e.g., morula or blastocyst), that is, after the bulk of reprogramming has taken place. These retrospective approaches are of limited use to elucidate mechanisms of reprogramming and to predict developmental success. Observing cloned embryo development using live embryo cinematography has the potential to reveal otherwise undetectable embryo features. However, light exposure necessary for live cell cinematography is highly toxic to cloned embryos. Here we describe a protocol for combined bright-field and fluorescence live-cell imaging of histone H2b-GFP expressing mouse embryos, to record cell divisions up to the blastocyst stage. This protocol, which can be adapted to observe other reporters such as Oct4-GFP or Nanog-GFP, allowed us to quantitatively analyze cleavage kinetics of cloned embryos.

Applications and Limitations of Mouse Models for Understanding Human Atherosclerosis

PubMed Central

von Scheidt, Moritz; Zhao, Yuqi; Kurt, Zeyneb; Pan, Calvin; Zeng, Lingyao; Yang, Xia; Schunkert, Heribert; Lusis, Aldons J.

2017-01-01

Most of the biological understanding of mechanisms underlying coronary artery disease (CAD) derives from studies of mouse models. The identification of multiple CAD loci and strong candidate genes in large human genome-wide association studies (GWAS) presented an opportunity to examine the relevance of mouse models for the human disease. We comprehensively reviewed the mouse literature, including 827 literature-derived genes, and compared it to human data. First, we observed striking concordance of risk factors for atherosclerosis in mice and humans. Second, there was highly significant overlap of mouse genes with human genes identified by GWAS. In particular, of the 46 genes with strong association signals in CAD-GWAS that were studied in mouse models all but one exhibited consistent effects on atherosclerosis-related phenotypes. Third, we compared 178 CAD-associated pathways derived from human GWAS with 263 from mouse studies and observed that over 50% were consistent between both species. PMID:27916529

Cyclophosphamide augments antitumor immunity: studies in an autochthonous prostate cancer model.

PubMed

Wada, Satoshi; Yoshimura, Kiyoshi; Hipkiss, Edward L; Harris, Tim J; Yen, Hung-Rong; Goldberg, Monica V; Grosso, Joseph F; Getnet, Derese; Demarzo, Angelo M; Netto, George J; Anders, Robert; Pardoll, Drew M; Drake, Charles G

2009-05-15

To study the immune response to prostate cancer, we developed an autochthonous animal model based on the transgenic adenocarcinoma of the mouse prostate (TRAMP) mouse in which spontaneously developing tumors express influenza hemagglutinin as a unique, tumor-associated antigen. Our prior studies in these animals showed immunologic tolerance to hemagglutinin, mirroring the clinical situation in patients with cancer who are generally nonresponsive to their disease. We used this physiologically relevant animal model to assess the immunomodulatory effects of cyclophosphamide when administered in combination with an allogeneic, cell-based granulocyte-macrophage colony-stimulating factor-secreting cancer immunotherapy. Through adoptive transfer of prostate/prostate cancer-specific CD8 T cells as well as through studies of the endogenous T-cell repertoire, we found that cyclophosphamide induced a marked augmentation of the antitumor immune response. This effect was strongly dependent on both the dose and the timing of cyclophosphamide administration. Mechanistic studies showed that immune augmentation by cyclophosphamide was associated with a transient depletion of regulatory T cells in the tumor draining lymph nodes but not in the peripheral circulation. Interestingly, we also noted effects on dendritic cell phenotype; low-dose cyclophosphamide was associated with increased expression of dendritic cell maturation markers. Taken together, these data clarify the dose, timing, and mechanism of action by which immunomodulatory cyclophosphamide can be translated to a clinical setting in a combinatorial cancer treatment strategy.

Genetically engineered mouse models for studying inflammatory bowel disease.

PubMed

Mizoguchi, Atsushi; Takeuchi, Takahito; Himuro, Hidetomo; Okada, Toshiyuki; Mizoguchi, Emiko

2016-01-01

Inflammatory bowel disease (IBD) is a chronic intestinal inflammatory condition that is mediated by very complex mechanisms controlled by genetic, immune, and environmental factors. More than 74 kinds of genetically engineered mouse strains have been established since 1993 for studying IBD. Although mouse models cannot fully reflect human IBD, they have provided significant contributions for not only understanding the mechanism, but also developing new therapeutic means for IBD. Indeed, 20 kinds of genetically engineered mouse models carry the susceptibility genes identified in human IBD, and the functions of some other IBD susceptibility genes have also been dissected out using mouse models. Cutting-edge technologies such as cell-specific and inducible knockout systems, which were recently employed to mouse IBD models, have further enhanced the ability of investigators to provide important and unexpected rationales for developing new therapeutic strategies for IBD. In this review article, we briefly introduce 74 kinds of genetically engineered mouse models that spontaneously develop intestinal inflammation. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Mouse Tumor Biology (MTB): a database of mouse models for human cancer.

PubMed

Bult, Carol J; Krupke, Debra M; Begley, Dale A; Richardson, Joel E; Neuhauser, Steven B; Sundberg, John P; Eppig, Janan T

2015-01-01

The Mouse Tumor Biology (MTB; http://tumor.informatics.jax.org) database is a unique online compendium of mouse models for human cancer. MTB provides online access to expertly curated information on diverse mouse models for human cancer and interfaces for searching and visualizing data associated with these models. The information in MTB is designed to facilitate the selection of strains for cancer research and is a platform for mining data on tumor development and patterns of metastases. MTB curators acquire data through manual curation of peer-reviewed scientific literature and from direct submissions by researchers. Data in MTB are also obtained from other bioinformatics resources including PathBase, the Gene Expression Omnibus and ArrayExpress. Recent enhancements to MTB improve the association between mouse models and human genes commonly mutated in a variety of cancers as identified in large-scale cancer genomics studies, provide new interfaces for exploring regions of the mouse genome associated with cancer phenotypes and incorporate data and information related to Patient-Derived Xenograft models of human cancers. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

The latest animal models of ovarian cancer for novel drug discovery.

PubMed

Magnotti, Elizabeth; Marasco, Wayne A

2018-03-01

Epithelial ovarian cancer is a heterogeneous disease classified into five subtypes, each with a different molecular profile. Most cases of ovarian cancer are diagnosed after metastasis of the primary tumor and are resistant to traditional platinum-based chemotherapeutics. Mouse models of ovarian cancer have been utilized to discern ovarian cancer tumorigenesis and the tumor's response to therapeutics. Areas covered: The authors provide a review of mouse models currently employed to understand ovarian cancer. This article focuses on advances in the development of orthotopic and patient-derived tumor xenograft (PDX) mouse models of ovarian cancer and discusses current humanized mouse models of ovarian cancer. Expert opinion: The authors suggest that humanized mouse models of ovarian cancer will provide new insight into the role of the human immune system in combating and augmenting ovarian cancer and aid in the development of novel therapeutics. Development of humanized mouse models will take advantage of the NSG and NSG-SGM3 strains of mice as well as new strains that are actively being derived.

How Genetically Engineered Mouse Tumor Models Provide Insights Into Human Cancers

PubMed Central

Politi, Katerina; Pao, William

2011-01-01

Genetically engineered mouse models (GEMMs) of human cancer were first created nearly 30 years ago. These early transgenic models demonstrated that mouse cells could be transformed in vivo by expression of an oncogene. A new field emerged, dedicated to generating and using mouse models of human cancer to address a wide variety of questions in cancer biology. The aim of this review is to highlight the contributions of mouse models to the diagnosis and treatment of human cancers. Because of the breadth of the topic, we have selected representative examples of how GEMMs are clinically relevant rather than provided an exhaustive list of experiments. Today, as detailed here, sophisticated mouse models are being created to study many aspects of cancer biology, including but not limited to mechanisms of sensitivity and resistance to drug treatment, oncogene cooperation, early detection, and metastasis. Alternatives to GEMMs, such as chemically induced or spontaneous tumor models, are not discussed in this review. PMID:21263096

Integrating model behavior, optimization, and sensitivity/uncertainty analysis: overview and application of the MOUSE software toolbox

USDA-ARS?s Scientific Manuscript database

This paper provides an overview of the Model Optimization, Uncertainty, and SEnsitivity Analysis (MOUSE) software application, an open-source, Java-based toolbox of visual and numerical analysis components for the evaluation of environmental models. MOUSE is based on the OPTAS model calibration syst...

Defects in Mitochondrial Dynamics and Metabolomic Signatures of Evolving Energetic Stress in Mouse Models of Familial Alzheimer's Disease

PubMed Central

Trushina, Eugenia; Nemutlu, Emirhan; Zhang, Song; Christensen, Trace; Camp, Jon; Mesa, Janny; Siddiqui, Ammar; Tamura, Yasushi; Sesaki, Hiromi; Wengenack, Thomas M.; Dzeja, Petras P.; Poduslo, Joseph F.

2012-01-01

Background The identification of early mechanisms underlying Alzheimer's Disease (AD) and associated biomarkers could advance development of new therapies and improve monitoring and predicting of AD progression. Mitochondrial dysfunction has been suggested to underlie AD pathophysiology, however, no comprehensive study exists that evaluates the effect of different familial AD (FAD) mutations on mitochondrial function, dynamics, and brain energetics. Methods and Findings We characterized early mitochondrial dysfunction and metabolomic signatures of energetic stress in three commonly used transgenic mouse models of FAD. Assessment of mitochondrial motility, distribution, dynamics, morphology, and metabolomic profiling revealed the specific effect of each FAD mutation on the development of mitochondrial stress and dysfunction. Inhibition of mitochondrial trafficking was characteristic for embryonic neurons from mice expressing mutant human presenilin 1, PS1(M146L) and the double mutation of human amyloid precursor protein APP(Tg2576) and PS1(M146L) contributing to the increased susceptibility of neurons to excitotoxic cell death. Significant changes in mitochondrial morphology were detected in APP and APP/PS1 mice. All three FAD models demonstrated a loss of the integrity of synaptic mitochondria and energy production. Metabolomic profiling revealed mutation-specific changes in the levels of metabolites reflecting altered energy metabolism and mitochondrial dysfunction in brains of FAD mice. Metabolic biomarkers adequately reflected gender differences similar to that reported for AD patients and correlated well with the biomarkers currently used for diagnosis in humans. Conclusions Mutation-specific alterations in mitochondrial dynamics, morphology and function in FAD mice occurred prior to the onset of memory and neurological phenotype and before the formation of amyloid deposits. Metabolomic signatures of mitochondrial stress and altered energy metabolism indicated alterations in nucleotide, Krebs cycle, energy transfer, carbohydrate, neurotransmitter, and amino acid metabolic pathways. Mitochondrial dysfunction, therefore, is an underlying event in AD progression, and FAD mouse models provide valuable tools to study early molecular mechanisms implicated in AD. PMID:22393443

The VP40 protein of Marburg virus exhibits impaired budding and increased sensitivity to human tetherin following mouse adaptation.

PubMed

Feagins, Alicia R; Basler, Christopher F

2014-12-01

The Marburg virus VP40 protein is a viral matrix protein that spontaneously buds from cells. It also functions as an interferon (IFN) signaling antagonist by targeting Janus kinase 1 (JAK1). A previous study demonstrated that the VP40 protein of the Ravn strain of Marburg virus (Ravn virus [RAVV]) failed to block IFN signaling in mouse cells, whereas the mouse-adapted RAVV (maRAVV) VP40 acquired the ability to inhibit IFN responses in mouse cells. The increased IFN antagonist function of maRAVV VP40 mapped to residues 57 and 165, which were mutated during the mouse adaptation process. In the present study, we demonstrate that maRAVV VP40 lost the capacity to efficiently bud from human cell lines, despite the fact that both parental and maRAVV VP40s bud efficiently from mouse cell lines. The impaired budding in human cells corresponds with the appearance of protrusions on the surface of maRAVV VP40-expressing Huh7 cells and with an increased sensitivity of maRAVV VP40 to restriction by human tetherin but not mouse tetherin. However, transfer of the human tetherin cytoplasmic tail to mouse tetherin restored restriction of maRAVV VP40. Residues 57 and 165 were demonstrated to contribute to the failure of maRAVV VP40 to bud from human cells, and residue 57 was demonstrated to alter VP40 oligomerization, as assessed by coprecipitation assay, and to determine sensitivity to human tetherin. This suggests that RAVV VP40 acquired, during adaptation to mice, changes in its oligomerization potential that enhanced IFN antagonist function. However, this new capacity impaired RAVV VP40 budding from human cells. Filoviruses, which include Marburg viruses and Ebola viruses, are zoonotic pathogens that cause severe disease in humans and nonhuman primates but do not cause similar disease in wild-type laboratory strains of mice unless first adapted to these animals. Although mouse adaptation has been used as a method to develop small animal models of pathogenesis, the molecular determinants associated with filovirus mouse adaptation are poorly understood. Our study demonstrates how genetic changes that accrued during mouse adaptation of the Ravn strain of Marburg virus have impacted the budding function of the viral VP40 matrix protein. Strikingly, we find impairment of mouse-adapted VP40 budding function in human but not mouse cell lines, and we correlate the impairment with an increased sensitivity of VP40 to restriction by human but not mouse tetherin and with changes in VP40 oligomerization. These data suggest that there are functional costs associated with filovirus adaptation to new hosts and implicate tetherin as a filovirus host restriction factor. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Disrupting the male germ line to find infertility and contraception targets.

PubMed

Archambeault, Denise R; Matzuk, Martin M

2014-05-01

Genetically-manipulated mouse models have become indispensible for broadening our understanding of genes and pathways related to male germ cell development. Until suitable in vitro systems for studying spermatogenesis are perfected, in vivo models will remain the gold standard for inquiry into testicular function. Here, we discuss exciting advances that are allowing researchers faster, easier, and more customizable access to their mouse models of interest. Specifically, the trans-NIH Knockout Mouse Project (KOMP) is working to generate knockout mouse models of every gene in the mouse genome. The related Knockout Mouse Phenotyping Program (KOMP2) is performing systematic phenotypic analysis of this genome-wide collection of knockout mice, including fertility screening. Together, these programs will not only uncover new genes involved in male germ cell development but also provide the research community with the mouse models necessary for further investigations. In addition to KOMP/KOMP2, another promising development in the field of mouse models is the advent of CRISPR (clustered regularly interspaced short palindromic repeat)-Cas technology. Utilizing 20 nucleotide guide sequences, CRISPR/Cas has the potential to introduce sequence-specific insertions, deletions, and point mutations to produce null, conditional, activated, or reporter-tagged alleles. CRISPR/Cas can also successfully target multiple genes in a single experimental step, forgoing the multiple generations of breeding traditionally required to produce mouse models with deletions, insertions, or mutations in multiple genes. In addition, CRISPR/Cas can be used to create mouse models carrying variants identical to those identified in infertile human patients, providing the opportunity to explore the effects of such mutations in an in vivo system. Both the KOMP/KOMP2 projects and the CRISPR/Cas system provide powerful, accessible genetic approaches to the study of male germ cell development in the mouse. A more complete understanding of male germ cell biology is critical for the identification of novel targets for potential non-hormonal contraceptive intervention. Copyright © 2014. Published by Elsevier Masson SAS.

Implausibility of the vibrational theory of olfaction

DOE PAGES

Block, Eric; Ertem, Mehmed Z.; Jang, Seogjoo; ...

2015-04-21

The vibrational theory of olfaction assumes that electron transfer occurs across odorants at the active sites of odorant receptors (ORs), serving as a sensitive measure of odorant vibrational frequencies, ultimately leading to olfactory perception. A previous study reported that human subjects differentiated hydrogen/deuterium isotopomers (isomers with isotopic atoms) of the musk compound cyclopentadecanone as evidence supporting the theory. Here, we find no evidence for such differentiation at the molecular level. In fact, we find that the human musk-recognizing receptor, OR5AN1, identified using a heterologous OR expression system and robustly responding to cyclopentadecanone and muscone, fails to distinguish isotopomers of thesemore » compounds in vitro. Furthermore, the mouse (methylthio)methanethiol (MTMT)-recognizing receptor, MOR244-3, and other selected human and mouse ORs, responded similarly to normal, deuterated, and ¹³C isotopomers of their respective ligands, paralleling our results with the musk receptor OR5AN1. These findings suggest that the proposed vibration theory does not apply to the human musk receptor OR5AN1, mouse thiol receptor MOR244-3, or other ORs examined. Also, contrary to the vibration theory predictions, muscone-d₃₀ lacks the 1,380-1,550 cm⁻¹ IR bands claimed to be essential for musk odor. Furthermore, our theoretical analysis shows that the proposed electron transfer mechanism of the vibrational frequencies of odorants could be easily suppressed by quantum effects of non-odorant molecular vibrational modes. As a result, these and other concerns about electron transfer at ORs, together with our extensive experimental data, argue against the plausibility of the vibration theory.« less

Experimental embryology of mammals at the Jastrzebiec Institute of Genetics and Animal Breeding.

PubMed

Karasiewicz, Jolanta; Andrzej-Modlinski, Jacek

2008-01-01

Our Department of Experimental Embryology originated from The Laboratory of Embryo Biotechnology, which was organized and directed by Dr. Maria Czlonkowska until her premature death in 1991. Proving successful international transfer of frozen equine embryos and generation of an embryonic sheep-goat chimaera surviving ten years were outstanding achievements of her term. In the 1990s, we produced advanced fetuses of mice after reconstructing enucleated oocytes with embryonic stem (ES) cells, as well as mice originating entirely from ES cells by substitution of the inner cell mass with ES cells. Attempts at obtaining ES cells in sheep resulted in the establishment of embryo-derived epithelioid cell lines from Polish Heatherhead and Polish Merino breeds, producing overt chimaeras upon blastocyst injection. Successful re-cloning was achieved from 8-cell rabbit embryos, and healthy animals were born from the third generation of cloned embryos. Recently mice were born after transfer of 8-cell embryonic nuclei into selectively enucleated zygotes, and mouse blastocysts were produced from selectively enucleated germinal vesicle oocytes surrounded by follicular cells, upon their reconstruction with 2-cell nuclei and subsequent activation. Embryonic-somatic chimaeras were born after transfer of foetal fibroblasts into 8-cell embryos (mouse) and into morulae and blastocysts (sheep). We also regularly perform the following applications: in vitro production of bovine embryos from slaughterhouse oocytes or those recovered by ovum pick up; cryopreservation of oocytes and embryos (freezing: mouse, rabbit, sheep, goat; vitrification: rabbit, cow); and banking of somatic cells from endangered wild mammalian species (mainly Cervidae).

Implausibility of the vibrational theory of olfaction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Block, Eric; Ertem, Mehmed Z.; Jang, Seogjoo

The vibrational theory of olfaction assumes that electron transfer occurs across odorants at the active sites of odorant receptors (ORs), serving as a sensitive measure of odorant vibrational frequencies, ultimately leading to olfactory perception. A previous study reported that human subjects differentiated hydrogen/deuterium isotopomers (isomers with isotopic atoms) of the musk compound cyclopentadecanone as evidence supporting the theory. Here, we find no evidence for such differentiation at the molecular level. In fact, we find that the human musk-recognizing receptor, OR5AN1, identified using a heterologous OR expression system and robustly responding to cyclopentadecanone and muscone, fails to distinguish isotopomers of thesemore » compounds in vitro. Furthermore, the mouse (methylthio)methanethiol (MTMT)-recognizing receptor, MOR244-3, and other selected human and mouse ORs, responded similarly to normal, deuterated, and ¹³C isotopomers of their respective ligands, paralleling our results with the musk receptor OR5AN1. These findings suggest that the proposed vibration theory does not apply to the human musk receptor OR5AN1, mouse thiol receptor MOR244-3, or other ORs examined. Also, contrary to the vibration theory predictions, muscone-d₃₀ lacks the 1,380-1,550 cm⁻¹ IR bands claimed to be essential for musk odor. Furthermore, our theoretical analysis shows that the proposed electron transfer mechanism of the vibrational frequencies of odorants could be easily suppressed by quantum effects of non-odorant molecular vibrational modes. As a result, these and other concerns about electron transfer at ORs, together with our extensive experimental data, argue against the plausibility of the vibration theory.« less

Genome-Wide Expression Profiling of Five Mouse Models Identifies Similarities and Differences with Human Psoriasis

PubMed Central

Swindell, William R.; Johnston, Andrew; Carbajal, Steve; Han, Gangwen; Wohn, Christian; Lu, Jun; Xing, Xianying; Nair, Rajan P.; Voorhees, John J.; Elder, James T.; Wang, Xiao-Jing; Sano, Shigetoshi; Prens, Errol P.; DiGiovanni, John; Pittelkow, Mark R.; Ward, Nicole L.; Gudjonsson, Johann E.

2011-01-01

Development of a suitable mouse model would facilitate the investigation of pathomechanisms underlying human psoriasis and would also assist in development of therapeutic treatments. However, while many psoriasis mouse models have been proposed, no single model recapitulates all features of the human disease, and standardized validation criteria for psoriasis mouse models have not been widely applied. In this study, whole-genome transcriptional profiling is used to compare gene expression patterns manifested by human psoriatic skin lesions with those that occur in five psoriasis mouse models (K5-Tie2, imiquimod, K14-AREG, K5-Stat3C and K5-TGFbeta1). While the cutaneous gene expression profiles associated with each mouse phenotype exhibited statistically significant similarity to the expression profile of psoriasis in humans, each model displayed distinctive sets of similarities and differences in comparison to human psoriasis. For all five models, correspondence to the human disease was strong with respect to genes involved in epidermal development and keratinization. Immune and inflammation-associated gene expression, in contrast, was more variable between models as compared to the human disease. These findings support the value of all five models as research tools, each with identifiable areas of convergence to and divergence from the human disease. Additionally, the approach used in this paper provides an objective and quantitative method for evaluation of proposed mouse models of psoriasis, which can be strategically applied in future studies to score strengths of mouse phenotypes relative to specific aspects of human psoriasis. PMID:21483750

RNA Sequencing Identifies Novel Translational Biomarkers of Kidney Fibrosis

PubMed Central

Craciun, Florin L.; Bijol, Vanesa; Ajay, Amrendra K.; Rao, Poornima; Kumar, Ramya K.; Hutchinson, John; Hofmann, Oliver; Joshi, Nikita; Luyendyk, James P.; Kusebauch, Ulrike; Moss, Christopher L.; Srivastava, Anand; Himmelfarb, Jonathan; Waikar, Sushrut S.; Moritz, Robert L.

2016-01-01

CKD is the gradual, asymptomatic loss of kidney function, but current tests only identify CKD when significant loss has already happened. Several potential biomarkers of CKD have been reported, but none have been approved for preclinical or clinical use. Using RNA sequencing in a mouse model of folic acid-induced nephropathy, we identified ten genes that track kidney fibrosis development, the common pathologic finding in patients with CKD. The gene expression of all ten candidates was confirmed to be significantly higher (approximately ten- to 150-fold) in three well established, mechanistically distinct mouse models of kidney fibrosis than in models of nonfibrotic AKI. Protein expression of these genes was also high in the folic acid model and in patients with biopsy-proven kidney fibrosis. mRNA expression of the ten genes increased with increasing severity of kidney fibrosis, decreased in response to therapeutic intervention, and increased only modestly (approximately two- to five-fold) with liver fibrosis in mice and humans, demonstrating specificity for kidney fibrosis. Using targeted selected reaction monitoring mass spectrometry, we detected three of the ten candidates in human urine: cadherin 11 (CDH11), macrophage mannose receptor C1 (MRC1), and phospholipid transfer protein (PLTP). Furthermore, urinary levels of each of these three proteins distinguished patients with CKD (n=53) from healthy individuals (n=53; P<0.05). In summary, we report the identification of urinary CDH11, MRC1, and PLTP as novel noninvasive biomarkers of CKD. PMID:26449608

Breast Cancer Redox Heterogeneity Detectable with Chemical Exchange Satruation Transfer (CEST) MRI

PubMed Central

Cai, Kejia; Xu, He N.; Singh, Anup; Moon, Lily; Haris, Mohammad; Reddy, Ravinder; Li, Lin

2014-01-01

Purpose Tissue redox state is an important mediator of various biological processes in health and diseases such as cancer. Previously, we discovered that the mitochondrial redox state of ex vivo tissues detected by redox scanning (an optical imaging method) revealed interesting tumor redox state heterogeneity that could differentiate tumor aggressiveness. Because the noninvasive chemical exchange saturation transfer (CEST) MRI can probe the proton transfer and generate contrasts from endogenous metabolites, we aim to investigate if the in vivo CEST contrast is sensitive to proton transfer of the redox reactions so as to reveal the tissue redox states in breast cancer animal models. Procedures CEST MRI has been employed to characterize tumor metabolic heterogeneity and correlated with the redox states measured by the redox scanning in two human breast cancer mouse xenograft models, MDA-MB-231 and MCF-7. The possible biological mechanism on the correlation between the two imaging modalities was further investigated by phantom studies where the reductants and the oxidants of the representative redox reactions were measured. Results The CEST contrast is found linearly correlated with NADH concentration and the NADH redox ratio with high statistical significance, where NADH is the reduced form of nicotinamide adenine dinucleotide. The phantom studies showed that the reductants of the redox reactions have more CEST contrast than the corresponding oxidants, indicating that higher CEST effect corresponds to the more reduced redox state. Conclusions This preliminary study suggests that CEST MRI, once calibrated, might provide a novel noninvasive imaging surrogate for the tissue redox state and a possible diagnostic biomarker for breast cancer in the clinic. PMID:24811957

Breast cancer redox heterogeneity detectable with chemical exchange saturation transfer (CEST) MRI.

PubMed

Cai, Kejia; Xu, He N; Singh, Anup; Moon, Lily; Haris, Mohammad; Reddy, Ravinder; Li, Lin Z

2014-10-01

Tissue redox state is an important mediator of various biological processes in health and diseases such as cancer. Previously, we discovered that the mitochondrial redox state of ex vivo tissues detected by redox scanning (an optical imaging method) revealed interesting tumor redox state heterogeneity that could differentiate tumor aggressiveness. Because the noninvasive chemical exchange saturation transfer (CEST) MRI can probe the proton transfer and generate contrasts from endogenous metabolites, we aim to investigate if the in vivo CEST contrast is sensitive to proton transfer of the redox reactions so as to reveal the tissue redox states in breast cancer animal models. CEST MRI has been employed to characterize tumor metabolic heterogeneity and correlated with the redox states measured by the redox scanning in two human breast cancer mouse xenograft models, MDA-MB-231 and MCF-7. The possible biological mechanism on the correlation between the two imaging modalities was further investigated by phantom studies where the reductants and the oxidants of the representative redox reactions were measured. The CEST contrast is found linearly correlated with NADH concentration and the NADH redox ratio with high statistical significance, where NADH is the reduced form of nicotinamide adenine dinucleotide. The phantom studies showed that the reductants of the redox reactions have more CEST contrast than the corresponding oxidants, indicating that higher CEST effect corresponds to the more reduced redox state. This preliminary study suggests that CEST MRI, once calibrated, might provide a novel noninvasive imaging surrogate for the tissue redox state and a possible diagnostic biomarker for breast cancer in the clinic.

Near-infrared laser irradiation improves the development of mouse pre-implantation embryos

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yokoo, Masaki; Mori, Miho

The aim of the present study was to assess the effects of near-infrared laser irradiation on the in vitro development of mouse embryos. Female ICR mice were superovulated with pregnant mare serum gonadotropin and human chorionic gonadotropin (hCG), and mated with male mice. Two-cell stage embryos were collected 40 h after administering hCG and cultured in M16 medium. Two-cell embryos (0 h after culture), 8-cell embryos (approx. 30 h after culture), morula (approx. 48 h after culture), and blastocysts (approx. 73 h after culture) were irradiated at 904 nm for 60 s. These embryos were cultured in a time-lapse monitoring system and the timing ofmore » blastocyst hatching was evaluated. Some of the irradiated blastocysts were transferred to the uterine horns of pseudopregnant recipients immediately after irradiation. Pregnancy rates, and offspring growth and fertility, were evaluated. Near-infrared laser irradiation increased the speed of in vitro mouse embryo development. In irradiated blastocysts, hatching was faster than in control (non-irradiated) blastocysts (18.4 vs. 28.2 h, P < 0.05). When 195 irradiated blastocysts were transferred to 18 pseudopregnant mice, all became pregnant and 92 (47.2%) normal-looking pups were born alive. When 182 control blastocysts were transferred to 17 pseudopregnant mice, 14 (82.4%) became pregnant and 54 (29.7%) normal-looking pups were born alive. The growth trajectories (up to 5 weeks) of offspring from irradiated blastocysts were similar to those from control blastocysts. Second generation offspring from transplanted animals were all fertile. These results indicate that near-infrared laser irradiation improves the quality of mouse embryo development in vitro, and increases the live birth rate without affecting the normality of the offspring. Thus, the near-infrared laser method may enhance the quality of embryos and contribute to improvements in reproductive technologies in mammals. - Highlights: • Irradiation of blastocysts with a near-infrared laser improves embryo development. • Irradiation of blastocysts increases the live birth rate after embryo transfer. • Irradiation of blastocysts did not affect the normality of the pups. • Near-infrared laser irradiation may be useful to enhance the quality of embryos. • This study may contribute to improvements in reproductive technologies in mammals.« less

Lentivirus-meditated frataxin gene delivery reverses genome instability in Friedreich ataxia patient and mouse model fibroblasts.

PubMed

Khonsari, H; Schneider, M; Al-Mahdawi, S; Chianea, Y G; Themis, M; Parris, C; Pook, M A; Themis, M

2016-12-01

Friedreich ataxia (FRDA) is a progressive neurodegenerative disease caused by deficiency of frataxin protein, with the primary sites of pathology being the large sensory neurons of the dorsal root ganglia and the cerebellum. FRDA is also often accompanied by severe cardiomyopathy and diabetes mellitus. Frataxin is important in mitochondrial iron-sulfur cluster (ISC) biogenesis and low-frataxin expression is due to a GAA repeat expansion in intron 1 of the FXN gene. FRDA cells are genomically unstable, with increased levels of reactive oxygen species and sensitivity to oxidative stress. Here we report the identification of elevated levels of DNA double strand breaks (DSBs) in FRDA patient and YG8sR FRDA mouse model fibroblasts compared to normal fibroblasts. Using lentivirus FXN gene delivery to FRDA patient and YG8sR cells, we obtained long-term overexpression of FXN mRNA and frataxin protein levels with reduced DSB levels towards normal. Furthermore, γ-irradiation of FRDA patient and YG8sR cells revealed impaired DSB repair that was recovered on FXN gene transfer. This suggests that frataxin may be involved in DSB repair, either directly by an unknown mechanism, or indirectly via ISC biogenesis for DNA repair enzymes, which may be essential for the prevention of neurodegeneration.

Lentiviral gene transfer regenerates hematopoietic stem cells in a mouse model for Mpl-deficient aplastic anemia.

PubMed

Heckl, Dirk; Wicke, Daniel C; Brugman, Martijn H; Meyer, Johann; Schambach, Axel; Büsche, Guntram; Ballmaier, Matthias; Baum, Christopher; Modlich, Ute

2011-04-07

Thpo/Mpl signaling plays an important role in the maintenance of hematopoietic stem cells (HSCs) in addition to its role in megakaryopoiesis. Patients with inactivating mutations in Mpl develop thrombocytopenia and aplastic anemia because of progressive loss of HSCs. Yet, it is unknown whether this loss of HSCs is an irreversible process. In this study, we used the Mpl knockout (Mpl(-/-)) mouse model and expressed Mpl from newly developed lentiviral vectors specifically in the physiologic Mpl target populations, namely, HSCs and megakaryocytes. After validating lineage-specific expression in vivo using lentiviral eGFP reporter vectors, we performed bone marrow transplantation of transduced Mpl(-/-) bone marrow cells into Mpl(-/-) mice. We show that restoration of Mpl expression from transcriptionally targeted vectors prevents lethal adverse reactions of ectopic Mpl expression, replenishes the HSC pool, restores stem cell properties, and corrects platelet production. In some mice, megakaryocyte counts were atypically high, accompanied by bone neo-formation and marrow fibrosis. Gene-corrected Mpl(-/-) cells had increased long-term repopulating potential, with a marked increase in lineage(-)Sca1(+)cKit(+) cells and early progenitor populations in reconstituted mice. Transcriptome analysis of lineage(-)Sca1(+)cKit(+) cells in Mpl-corrected mice showed functional adjustment of genes involved in HSC self-renewal.

[Effect of topical application of a recombinant adenovirus carrying promyelocytic leukemia gene in a psoriasis-like mouse model].

PubMed

Wang, Qiongyu; Zhang, Aijun; Ma, Huiqun; Wang, Shijie; Ma, Yunyun; Zou, Xingwei; Li, Ruilian

2013-03-01

To investigate the effects of topical treatment with adenovirus-mediated promyelocytic leukemia gene (PML) gene in a psoriasis-like mouse model. The effect of adenovirus-mediated PML gene on the granular layer of mouse tail scale epidermis and epithelial mitosis were observed on longitudinal histological sections prepared from the tail skin and vaginal epithelium of the mice. Adenovirus-mediated PML gene significantly inhibited mitosis of mouse vaginal epithelial cells and promoted the formation of granular layer in mouse tail scale epidermis. The therapeutic effect of PML gene in the psoriasis-like mouse model may be associated with increased granular cells and suppressed epidemic cell proliferation.

On the high frequency transfer of mechanical stimuli from the surface of the head to the macular neuroepithelium of the mouse.

PubMed

Jones, Timothy A; Lee, Choongheon; Gaines, G Christopher; Grant, J W Wally

2015-04-01

Vestibular macular sensors are activated by a shearing motion between the otoconial membrane and underlying receptor epithelium. Shearing motion and sensory activation in response to an externally induced head motion do not occur instantaneously. The mechanically reactive elastic and inertial properties of the intervening tissue introduce temporal constraints on the transfer of the stimulus to sensors. Treating the otoconial sensory apparatus as an overdamped second-order mechanical system, we measured the governing long time constant (Τ(L)) for stimulus transfer from the head surface to epithelium. This provided the basis to estimate the corresponding upper cutoff for the frequency response curve for mouse otoconial organs. A velocity step excitation was used as the forcing function. Hypothetically, the onset of the mechanical response to a step excitation follows an exponential rise having the form Vel(shear) = U(1-e(-t/TL)), where U is the applied shearing velocity step amplitude. The response time of the otoconial apparatus was estimated based on the activation threshold of macular neural responses to step stimuli having durations between 0.1 and 2.0 ms. Twenty adult C57BL/6 J mice were evaluated. Animals were anesthetized. The head was secured to a shaker platform using a non-invasive head clip or implanted skull screws. The shaker was driven to produce a theoretical forcing step velocity excitation at the otoconial organ. Vestibular sensory evoked potentials (VsEPs) were recorded to measure the threshold for macular neural activation. The duration of the applied step motion was reduced systematically from 2 to 0.1 ms and response threshold determined for each duration (nine durations). Hypothetically, the threshold of activation will increase according to the decrease in velocity transfer occurring at shorter step durations. The relationship between neural threshold and stimulus step duration was characterized. Activation threshold increased exponentially as velocity step duration decreased below 1.0 ms. The time constants associated with the exponential curve were Τ(L) = 0.50 ms for the head clip coupling and T(L) = 0.79 ms for skull screw preparation. These corresponded to upper -3 dB frequency cutoff points of approximately 318 and 201 Hz, respectively. T(L) ranged from 224 to 379 across individual animals using the head clip coupling. The findings were consistent with a second-order mass-spring mechanical system. Threshold data were also fitted to underdamped models post hoc. The underdamped fits suggested natural resonance frequencies on the order of 278 to 448 Hz as well as the idea that macular systems in mammals are less damped than generally acknowledged. Although estimated indirectly, it is argued that these time constants reflect largely if not entirely the mechanics of transfer to the sensory apparatus. The estimated governing time constant of 0.50 ms for composite data predicts high frequency cutoffs of at least 318 Hz for the intact otoconial apparatus of the mouse.

Inferring the Impact of Regulatory Mechanisms that Underpin CD8+ T Cell Control of B16 Tumor Growth In vivo Using Mechanistic Models and Simulation.

PubMed

Klinke, David J; Wang, Qing

2016-01-01

A major barrier for broadening the efficacy of immunotherapies for cancer is identifying key mechanisms that limit the efficacy of tumor infiltrating lymphocytes. Yet, identifying these mechanisms using human samples and mouse models for cancer remains a challenge. While interactions between cancer and the immune system are dynamic and non-linear, identifying the relative roles that biological components play in regulating anti-tumor immunity commonly relies on human intuition alone, which can be limited by cognitive biases. To assist natural intuition, modeling and simulation play an emerging role in identifying therapeutic mechanisms. To illustrate the approach, we developed a multi-scale mechanistic model to describe the control of tumor growth by a primary response of CD8+ T cells against defined tumor antigens using the B16 C57Bl/6 mouse model for malignant melanoma. The mechanistic model was calibrated to data obtained following adenovirus-based immunization and validated to data obtained following adoptive transfer of transgenic CD8+ T cells. More importantly, we use simulation to test whether the postulated network topology, that is the modeled biological components and their associated interactions, is sufficient to capture the observed anti-tumor immune response. Given the available data, the simulation results also provided a statistical basis for quantifying the relative importance of different mechanisms that underpin CD8+ T cell control of B16F10 growth. By identifying conditions where the postulated network topology is incomplete, we illustrate how this approach can be used as part of an iterative design-build-test cycle to expand the predictive power of the model.

X‐linked retinoschisis: an update

PubMed Central

Sikkink, Stephen K; Biswas, Susmito; Parry, Neil R A; Stanga, Paulo E; Trump, Dorothy

2007-01-01

X‐linked retinoschisis is the leading cause of macular degeneration in males and leads to splitting within the inner retinal layers leading to visual deterioration. Many missense and protein truncating mutations have now been identified in the causative retinoschisis gene (RS1) which encodes a 224 amino acid secreting retinal protein, retinoschisin. Retinoschisin octamerises is implicated in cell–cell interactions and cell adhesion perhaps by interacting with β2 laminin. Mutations cause loss of retinoschisin function by one of the three mechanisms: by interfering with protein secretion, by preventing its octamerisation or by reducing function in the secreted octamerised protein. The development of retinoschisis mouse models have provided a model system that closely resembles the human disease. Recent reports of RS1 gene transfer to these models and the sustained restoration of some retinal function and morphology suggest gene replacement may be a possible future therapy for patients. PMID:17172462

Generation of transgenic mouse model using PTTG as an oncogene.

PubMed

Kakar, Sham S; Kakar, Cohin

2015-01-01

The close physiological similarity between the mouse and human has provided tools to understanding the biological function of particular genes in vivo by introduction or deletion of a gene of interest. Using a mouse as a model has provided a wealth of resources, knowledge, and technology, helping scientists to understand the biological functions, translocation, trafficking, and interaction of a candidate gene with other intracellular molecules, transcriptional regulation, posttranslational modification, and discovery of novel signaling pathways for a particular gene. Most importantly, the generation of the mouse model for a specific human disease has provided a powerful tool to understand the etiology of a disease and discovery of novel therapeutics. This chapter describes in detail the step-by-step generation of the transgenic mouse model, which can be helpful in guiding new investigators in developing successful models. For practical purposes, we will describe the generation of a mouse model using pituitary tumor transforming gene (PTTG) as the candidate gene of interest.

Establishing a learned-helplessness effect paradigm in C57BL/6 mice: behavioural evidence for emotional, motivational and cognitive effects of aversive uncontrollability per se.

PubMed

Pryce, Christopher R; Azzinnari, Damiano; Sigrist, Hannes; Gschwind, Tilo; Lesch, Klaus-Peter; Seifritz, Erich

2012-01-01

Uncontrollability of major life events has been proposed to be central to depression onset and maintenance. The learned helplessness (LH) effect describes a deficit in terminating controllable aversive stimuli in individuals that experienced aversive stimuli as uncontrollable relative to individuals that experienced the same stimuli as controllable. The LH effect translates across species and therefore can provide an objective-valid readout in animal models of depression. Paradigms for a robust LH effect are established and currently applied in rat but there are few reports of prior and current study of the LH effect in mouse. This includes the C57BL/6 mouse, typically the strain of choice for application of molecular-genetic tools in pre-clinical depression research. The aims of this study were to develop a robust paradigm for the LH effect in BL/6 mice, provide evidence for underlying psychological processes, and study the effect of a depression-relevant genotype on the LH effect. The apparatus used for in/escapable electro-shock exposure and escape test was a two-way shuttle arena with continuous automated measurement of locomotion, compartment transfers, e-shock escapes, vertical activity and freezing. Brother-pairs of BL/6 mice were allocated to either escapable e-shocks (ES) or inescapable e-shocks (IS), with escape latencies of the ES brother used as e-shock durations for the IS brother. The standard two-way shuttle paradigm was modified: the central gate was replaced by a raised divider and e-shock escape required transfer to the distal part of the safe compartment. These refinements yielded reduced superstitious, pre-adaptive e-shock transfers in IS mice and thereby increased the LH effect. To obtain a robust LH effect in all brother pairs, pre-screening for minor between-brother ES differences was necessary and did not confound the LH effect. IS mice developed reduced motor responses to e-shock, consistent with a motivational deficit, and absence of a learning curve for escapes at escape test, consistent with a cognitive deficit. When a tone CS was used to predict e-shock, IS mice exhibited increased reactivity to the CS, consistent with hyper-emotionality. There was no ES-IS difference in pain sensitivity. Mice heterozygous knockout for the 5-HTT gene exhibited an increased LH effect relative to wildtype mice. This mouse model will allow for the detailed molecular study of the aetiology, psychology, neurobiology and neuropharmacology of uncontrollability of aversive stimuli, a potential major aetiological factor and state marker in depression. This article is part of a Special Issue entitled 'Anxiety and Depression'. Copyright © 2011 Elsevier Ltd. All rights reserved.

A fully human chimeric antigen receptor with potent activity against cancer cells but reduced risk for off-tumor toxicity

PubMed Central

Song, De-Gang; Ye, Qunrui; Poussin, Mathilde; Liu, Lin; Figini, Mariangela; Powell, Daniel J.

2015-01-01

Chimeric antigen receptors (CARs) can redirect T cells against antigen-expressing tumors in an HLA-independent manner. To date, various CARs have been constructed using mouse single chain antibody variable fragments (scFvs) of high affinity that are immunogenic in humans and have the potential to mediate “on-target” toxicity. Here, we developed and evaluated a fully human CAR comprised of the human C4 folate receptor-alpha (αFR)-specific scFv coupled to intracellular T cell signaling domains. Human T cells transduced to express the C4 CAR specifically secreted proinflammatory cytokine and exerted cytolytic functions when cultured with αFR-expressing tumors in vitro. Adoptive transfer of C4 CAR T cells mediated the regression of large, established human ovarian cancer in a xenogeneic mouse model. Relative to a murine MOv19 scFv-based αFR CAR, C4 CAR T cells mediated comparable cytotoxic tumor activity in vitro and in vivo but had lower affinity for αFR protein and exhibited reduced recognition of normal cells expressing low levels of αFR. Thus, T cells expressing a fully human CAR of intermediate affinity can efficiently kill antigen-expressing tumors in vitro and in vivo and may overcome issues of transgene immunogenicity and “on-target off-tumor” toxicity that plague trials utilizing CARs containing mouse-derived, high affinity scFvs. PMID:26101914

Development, glycolytic activity, and viability of preimplantation mouse embryos subjected to different periods of glucose starvation.

PubMed

Leppens-Luisier, G; Sakkas, D

1997-03-01

After compaction, the preimplantation mouse embryo switches to a glucose-based metabolism, whereas for the 2- to 4-cell stage embryo, glucose can be inhibitory. In this study, we investigated the adaptability of preimplantation embryos to different periods of glucose starvation by culturing in vitro fertilized (IVF) and in vivo-fertilized 1-cell OF1 mouse embryos. Blastocysts obtained from exposure to glucose starvation for different periods of time were examined for the number of cells in the trophectoderm and inner cell mass, and for glycolytic activity and viability. A high percentage of blastocysts was obtained when 1-cell embryos fertilized in vitro or in vivo were cultured in M16 until the 2-cell stage, were transferred to M16 without glucose (M16-G) until the 4- or 8-cell stage, and then were transferred to fresh M16-G. When in vivo-fertilized 1-cell embryos were cultured to the 2-cell stage and then left in M16, less than 5% formed blastocysts compared to 26% of those transferred into M16-G. Blastocysts obtained when in vivo-fertilized 1-cell embryos were left in M16-G after the 2-cell stage, however, showed a significantly elevated glycolytic activity compared to those transferred to fresh M16 or M16-G medium at the 4- or 8-cell stage. Interestingly, even though these embryos displayed elevated glycolytic activity, they did not exhibit differences in the numbers of inner cell mass and trophectoderm cells or in viability compared to embryos cultured according to other protocols. Blastocysts from all cultured protocols had a significantly lower total cell number and a lower trophectoderm, but not inner cell mass, cell number compared to blastocysts developed in vivo. This study documents the metabolic adaptability of the preimplantation embryo by highlighting its ability to proceed with development and retain viability when challenged with glucose starvation at different periods.

Chromosome-wise dissection of the genome of the extremely big mouse line DU6i.

PubMed

Bevova, Marianna R; Aulchenko, Yurii S; Aksu, Soner; Renne, Ulla; Brockmann, Gudrun A

2006-01-01

The extreme high-body-weight-selected mouse line DU6i is a polygenic model for growth research, harboring many small-effect QTL. We dissected the genome of this line into 19 autosomes and the Y chromosome by the construction of a new panel of chromosome substitution strains (CSS). The DU6i chromosomes were transferred to a DBA/2 mice genetic background by marker-assisted recurrent backcrossing. Mitochondria and the X chromosome were of DBA/2 origin in the backcross. During the construction of these novel strains, >4000 animals were generated, phenotyped, and genotyped. Using these data, we studied the genetic control of variation in body weight and weight gain at 21, 42, and 63 days. The unique data set facilitated the analysis of chromosomal interaction with sex and parent-of-origin effects. All analyzed chromosomes affected body weight and weight gain either directly or in interaction with sex or parent of origin. The effects were age specific, with some chromosomes showing opposite effects at different stages of development.

Phagocytic cells contribute to the antibody-mediated elimination of pulmonary-infected SARS coronavirus.

PubMed

Yasui, Fumihiko; Kohara, Michinori; Kitabatake, Masahiro; Nishiwaki, Tetsu; Fujii, Hideki; Tateno, Chise; Yoneda, Misako; Morita, Kouichi; Matsushima, Kouji; Koyasu, Shigeo; Kai, Chieko

2014-04-01

While the 2002-2003 outbreak of severe acute respiratory syndrome (SARS) resulted in 774 deaths, patients who were affected with mild pulmonary symptoms successfully recovered. The objective of the present work was to identify, using SARS coronavirus (SARS-CoV) mouse infection models, immune factors responsible for clearing of the virus. The elimination of pulmonary SARS-CoV infection required the activation of B cells by CD4(+) T cells. Furthermore, passive immunization (post-infection) with homologous (murine) anti-SARS-CoV antiserum showed greater elimination efficacy against SARS-CoV than that with heterologous (rabbit) antiserum, despite the use of equivalent titers of neutralizing antibodies. This distinction was mediated by mouse phagocytic cells (monocyte-derived infiltrating macrophages and partially alveolar macrophages, but not neutrophils), as demonstrated both by adoptive transfer from donors and by immunological depletion of selected cell types. These results indicate that the cooperation of anti-SARS-CoV antibodies and phagocytic cells plays an important role in the elimination of SARS-CoV. Copyright © 2014 Elsevier Inc. All rights reserved.

[Establishment and identification of mouse lymphoma cell line EL4 expressing red fluorescent protein].

PubMed

Li, Yan-Jie; Cao, Jiang; Chen, Chong; Wang, Dong-Yang; Zeng, Ling-Yu; Pan, Xiu-Ying; Xu, Kai-Lin

2010-02-01

This study was purposed to construct a lentiviral vector encoding red fluorescent protein (DsRed) and transfect DsRed into EL4 cells for establishing mouse leukemia/lymphoma model expressing DsRed. The bicistronic SIN lentiviral transfer plasmid containing the genes encoding neo and internal ribosomal entry site-red fluorescent protein (IRES-DsRed) was constructed. Human embryonic kidney 293FT cells were co-transfected with the three plasmids by liposome method. The viral particles were collected and used to transfect EL4 cells, then the cells were selected by G418. The results showed that the plasmid pXZ208-neo-IRES-DsRed was constructed successfully, and the viral titer reached to 10(6) U/ml. EL4 cells were transfected by the viral solution efficiently. The transfected EL4 cells expressing DsRed survived in the final concentration 600 microg/ml of G418. The expression of DsRed in the transfected EL4 cells was demonstrated by fluorescence microscopy and flow cytometry. In conclusion, the EL4/DsRed cell line was established successfully.

Receptor-mediated transfer of pSV2CAT DNA to mouse liver cells using asialofetuin-labeled liposomes.

PubMed

Hara, T; Aramaki, Y; Takada, S; Koike, K; Tsuchiya, S

1995-12-01

Asialofetuin-labeled liposomes (AF-liposomes) were developed as a nonviral vector having high transfection activity for receptor-mediated gene transfer to hepatocytes by systemic administration. Initially, the majority of pSV2CAT, a chloramphenicol acetyltransferase (CAT) gene expression plasmid, was associated with AF-liposomes (AF-liposome-pSV2CAT), and they were injected into the portal vein of an adult mouse. Significantly high CAT activity was observed in the liver. The CAT activity in the liver was further increased two-fold by using AF-liposomes completely encapsulating pSV2CAT. Nonlabeled control liposomes, on the other hand, showed lower CAT activity in the liver than in the spleen or lung. The level of CAT mRNA reflected the CAT activity obtained by each liposome preparation in each tissue. Immunohistochemical staining showed that CAT was produced in a large number of parenchymal cells localizing in the periportal area. The plasmid encapsulated in the internal aqueous layer of the liposomes was effectively protected from environmental degradation. Thus, by administration into the blood circulation, AF-liposomes would be successfully incorporated into hepatocytes through receptor-mediated endocytosis, and the encapsulated plasmid would be transferred to the intracellular pathway.

What Is the Predictive Value of Animal Models for Vaccine Efficacy in Humans? Reevaluating the Potential of Mouse Models for the Human Immune System.

PubMed

Jameson, Stephen C; Masopust, David

2018-04-02

Much of what we understand about immunology, including the response to vaccines, come from studies in mice because they provide many practical advantages compared with research in higher mammals and humans. Nevertheless, modalities for preventing or treating disease do not always translate from mouse to humans, which has led to increasing scrutiny of the continued merits of mouse research. Here, we summarize the pros and cons of current laboratory mouse models for immunology research and discuss whether overreliance on nonphysiological, ultra-hygienic animal husbandry approaches has limited the ultimate translation potential of mouse-derived data to humans. Alternative approaches are discussed that may extend the use of the mouse model for preclinical studies. Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kishigami, Satoshi; Mizutani, Eiji; Ohta, Hiroshi

The low success rate of animal cloning by somatic cell nuclear transfer (SCNT) is believed to be associated with epigenetic errors including abnormal DNA hypermethylation. Recently, we elucidated by using round spermatids that, after nuclear transfer, treatment of zygotes with trichostatin A (TSA), an inhibitor of histone deacetylase, can remarkably reduce abnormal DNA hypermethylation depending on the origins of transferred nuclei and their genomic regions [S. Kishigami, N. Van Thuan, T. Hikichi, H. Ohta, S. Wakayama. E. Mizutani, T. Wakayama, Epigenetic abnormalities of the mouse paternal zygotic genome associated with microinsemination of round spermatids, Dev. Biol. (2005) in press]. Here,more » we found that 5-50 nM TSA-treatment for 10 h following oocyte activation resulted in more efficient in vitro development of somatic cloned embryos to the blastocyst stage from 2- to 5-fold depending on the donor cells including tail tip cells, spleen cells, neural stem cells, and cumulus cells. This TSA-treatment also led to more than 5-fold increase in success rate of mouse cloning from cumulus cells without obvious abnormality but failed to improve ES cloning success. Further, we succeeded in establishment of nuclear transfer-embryonic stem (NT-ES) cells from TSA-treated cloned blastocyst at a rate three times higher than those from untreated cloned blastocysts. Thus, our data indicate that TSA-treatment after SCNT in mice can dramatically improve the practical application of current cloning techniques.« less

Low-frequency ultrasound increases non-viral gene transfer to the mouse lung.

PubMed

Xenariou, Stefania; Liang, Hai-Dong; Griesenbach, Uta; Zhu, Jie; Farley, Raymond; Somerton, Lucinda; Singh, Charanjit; Jeffery, Peter K; Scheule, Ronald K; Cheng, Seng H; Geddes, Duncan M; Blomley, Martin; Alton, Eric W F W

2010-01-01

The aim of the study was to assess if low-frequency ultrasound (US), in the range of 30-35 kHz, increases non-viral gene transfer to the mouse lung. US is greatly attenuated in the lung due to large energy losses at the air/tissue interfaces. The advantages of low-frequency US, compared with high-frequency US are: (i) increased cavitation (responsible for the formation of transient pores in the cell membrane) and (ii) reduced energy losses during lung penetration. Cationic lipid GL67/plasmid DNA (pDNA), polyethylenimine (PEI)/pDNA and naked pDNA were delivered via intranasal instillation and the animals were then exposed to US (sonoporation) at 0.07 or 0.1 MPa for 10 min. Under these conditions, US did not enhance GL67 or PEI-mediated transfection. It did, however, increase naked pDNA gene transfer by approximately 4 folds. Importantly, this was achieved in the absence of microbubbles, which are crucial for the commonly used high-frequency (1 MHz) sonoporation but may not be able to withstand nebulization in a clinically relevant setup. Lung hemorrhage was also assessed and shown to increase with US pressure in a dose-dependent manner. We have thus, established that low-frequency US can enhance lung gene transfer with naked pDNA and this enhancement is more effective than the previously reported 1 MHz US.

Zika Virus Trafficking and Interactions in the Human Male Reproductive Tract.

PubMed

da Silva, Lucia Regina Cangussu

2018-05-11

Sexual transmission of Zika virus (ZIKV) is a matter of great concern. Infectious viral particles can be shed in semen for as long as six months after infection and can be transferred to male and female sexual partners during unprotected sexual intercourse. The virus can be found inside spermatozoa and could be directly transferred to the oocyte during fertilization. Sexual transmission of ZIKV can contribute to the rise in number of infected individuals in endemic areas as well as in countries where the mosquito vector does not thrive. There is also the possibility, as has been demonstrated in mouse models, that the vaginal deposition of ZIKV particles present in semen could lead to congenital syndrome. In this paper, we review the current literature to understand ZIKV trafficking from the bloodstream to the human male reproductive tract and viral interactions with host cells in interstitial spaces, tubule walls, annexed glands and semen. We hope to highlight gaps to be filled by future research and potential routes for vaccine and antiviral development.

DUSP3 genetic deletion confers M2-like−macrophage-dependent tolerance to septic shock

PubMed Central

Singh, Pratibha; Dejager, Lien; Amand, Mathieu; Theatre, Emilie; Vandereyken, Maud; Zurashvilli, Tinatin; Singh, Maneesh; Mack, Matthias; Timmermans, Steven; Musumeci, Lucia; Dejardin, Emmanuel; Mustelin, Tomas; Van Ginderachter, Jo A.; Moutschen, Michel; Oury, Cécile; Libert, Claude; Rahmouni, Souad

2015-01-01

DUSP3 is a small dual-specificity protein phosphatase with an unknown physiological function. We report that DUSP3 is strongly expressed in human and mouse monocytes and macrophages and that its deficiency in mice promotes tolerance to lipopolysaccharide (LPS)-induced endotoxin shock and to polymicrobial septic shock following cecal ligation and puncture. By using adoptive transfer experiments, we demonstrate that resistance to endotoxin is macrophage-dependent and transferable and that this protection is associated with a striking increase of M2-like macrophages in DUSP3−/− mice in both the LPS and cecal ligation and puncture models. We show that the altered response of DUSP3−/− mice to sepsis is reflected in decreased TNF production and impaired ERK1/2 activation. Our results demonstrate that DUSP3 plays a key and non-redundant role as a regulator of innate immune responses by mechanisms involving the control of ERK1/2 activation, TNF secretion and macrophage polarization. PMID:25876765

Maternal-fetal transfer of selenium in the mouse

PubMed Central

Burk, Raymond F.; Olson, Gary E.; Hill, Kristina E.; Winfrey, Virginia P.; Motley, Amy K.; Kurokawa, Suguru

2013-01-01

Selenoprotein P (Sepp1) is taken up by receptor-mediated endocytosis for its selenium. The other extracellular selenoprotein, glutathione peroxidase-3 (Gpx3), has not been shown to transport selenium. Mice with genetic alterations of Sepp1, the Sepp1 receptors apolipoprotein E receptor-2 (apoER2) and megalin, and Gpx3 were used to investigate maternal-fetal selenium transfer. Immunocytochemistry (ICC) showed receptor-independent uptake of Sepp1 and Gpx3 in the same vesicles of d-13 visceral yolk sac cells, suggesting uptake by pinocytosis. ICC also showed apoER2-mediated uptake of maternal Sepp1 in the d-18 placenta. Thus, two selenoprotein-dependent maternal-fetal selenium transfer mechanisms were identified. Selenium was quantified in d-18 fetuses with the mechanisms disrupted. Maternal Sepp1 deletion, which lowers maternal whole-body selenium, decreased fetal selenium under selenium-adequate conditions but deletion of fetal apoER2 did not. Fetal apoER2 deletion did decrease fetal selenium, by 51%, under selenium-deficient conditions, verifying function of the placental Sepp1-apoER2 mechanism. Maternal Gpx3 deletion decreased fetal selenium, by 13%, but only under selenium-deficient conditions. These findings indicate that the selenoprotein uptake mechanisms ensure selenium transfer to the fetus under selenium-deficient conditions. The failure of their disruptions (apoER2 deletion, Gpx3 deletion) to affect fetal selenium under selenium-adequate conditions indicates the existence of an additional maternal-fetal selenium transfer mechanism.—Burk, R. F., Olson, G. E., Hill, K. E., Winfrey, V. P., Motley, A. K., and Kurokawa, S. Maternal-fetal transfer of selenium in the mouse. PMID:23651543

Intercellular Transfer of a Soluble Viral Superantigen

PubMed Central

Reilly, Melissa; Mix, Denise; Reilly, Andrew A.; Yang Ye, Xiang; Winslow, Gary M.

2000-01-01

Mouse mammary tumor virus (MMTV) superantigens (vSAgs) can undergo intercellular transfer in vivo and in vitro such that a vSAg can be presented to T cells by major histocompatibility complex (MHC) class II proteins on antigen-presenting cells (APCs) that do not express the superantigen. This process may allow T-cell activation to occur prior to viral infection. Consistent with these findings, vSAg produced by Chinese hamster ovary (CHO) cells was readily transferred to class II IE and IA (H-2k and H-2d) proteins on a B-cell lymphoma or mouse splenocytes. Fixed class II-expressing acceptor cells were used to demonstrate that the vSAg, but not the class II proteins, underwent intercellular transfer, indicating that vSAg binding to class II MHC could occur directly at the cell surface. Intercellular transfer also occurred efficiently to splenocytes from endogenous retrovirus-free mice, indicating that other proviral proteins were not involved. Presentation of vSAg7 produced by a class II-negative, furin protease-deficient CHO variant (FD11) was unsuccessful, indicating that proteolytic processing was a requisite event and that proteolytic activity could not be provided by an endoprotease on the acceptor APC. Furthermore, vSAg presentation was effected using cell-free supernatant from class II-negative, vSAg-positive cells, indicating that a soluble molecule, most likely produced by proteolytic processing, was sufficient to stimulate T cells. Because the membrane-proximal endoproteolytic cleavage site in the vSAg (residues 68 to 71) was not necessary for intercellular transfer, the data support the notion that the carboxy-terminal endoproteolytic cleavage product is an active vSAg moiety. PMID:10954523

Tick-Borne Encephalitis Virus Vaccine-Induced Human Antibodies Mediate Negligible Enhancement of Zika Virus Infection InVitro and in a Mouse Model.

PubMed

Duehr, James; Lee, Silviana; Singh, Gursewak; Foster, Gregory A; Krysztof, David; Stramer, Susan L; Bermúdez González, Maria C; Menichetti, Eva; Geretschläger, Robert; Gabriel, Christian; Simon, Viviana; Lim, Jean K; Krammer, Florian

2018-01-01

Recent reports in the scientific literature have suggested that anti-dengue virus (DENV) and anti-West Nile virus (WNV) immunity exacerbates Zika virus (ZIKV) pathogenesis in vitro and in vivo in mouse models. Large populations of immune individuals exist for a related flavivirus (tick-borne encephalitis virus [TBEV]), due to large-scale vaccination campaigns and endemic circulation throughout most of northern Europe and the southern Russian Federation. As a result, the question of whether anti-TBEV immunity can affect Zika virus pathogenesis is a pertinent one. For this study, we obtained 50 serum samples from individuals vaccinated with the TBEV vaccine FSME-IMMUN (Central European/Neudörfl strain) and evaluated their enhancement capacity in vitro using K562 human myeloid cells expressing CD32 and in vivo using a mouse model of ZIKV pathogenesis. Among the 50 TBEV vaccinee samples evaluated, 29 had detectable reactivity against ZIKV envelope (E) protein by enzyme-linked immunosorbent assay (ELISA), and 36 showed enhancement of ZIKV infection in vitro . A pool of the most highly reacting and enhanced samples resulted in no significant change in the morbidity/mortality of ZIKV disease in immunocompromised Stat2 -/- mice. Our results suggest that humoral immunity against TBEV is unlikely to enhance Zika virus pathogenesis in humans. No clinical reports indicating that TBEV vaccinees experiencing enhanced ZIKV disease have been published so far, and though the epidemiological data are sparse, our findings suggest that there is little reason for concern. This study also displays a clear relationship between the phylogenetic distance between two flaviviruses and their capacity for pathogenic enhancement. IMPORTANCE The relationship between serial infections of two different serotypes of dengue virus and more severe disease courses is well-documented in the literature, driven by so-called antibody-dependent enhancement (ADE). Recently, studies have shown the possibility of ADE in cells exposed to anti-DENV human plasma and then infected with ZIKV and also in mouse models of ZIKV pathogenesis after passive transfer of anti-DENV human plasma. In this study, we evaluated the extent to which this phenomenon occurs using sera from individuals immunized against tick-borne encephalitis virus (TBEV). This is highly relevant, since large proportions of the European population are vaccinated against TBEV or otherwise seropositive.

Late-onset cone photoreceptor degeneration induced by R172W mutation in Rds and partial rescue by gene supplementation.

PubMed

Conley, Shannon; Nour, May; Fliesler, Steven J; Naash, Muna I

2007-12-01

R172W is a common mutation in the human retinal degeneration slow (RDS) gene, associated with a late-onset dominant macular dystrophy. In this study, the authors characterized a mouse model that closely mimics the human phenotype and tested the feasibility of gene supplementation as a disease treatment strategy. Transgenic mouse lines carrying the R172W mutation were generated. The retinal phenotype associated with this mutation in a low-expresser line (L-R172W) was examined, both structurally (histology with correlative immunohistochemistry) and functionally (electroretinography). By examining animals over time and with various rds genetic backgrounds, the authors evaluated the dominance of the defect. To assess the efficacy of gene transfer therapy as a treatment for this defect, a previously characterized transgenic line expressing the normal mouse peripherin/Rds (NMP) was crossed with a higher-expresser Rds line harboring the R172W mutation (H-R172W). Functional, structural, and biochemical analyses were used to assess rescue of the retinal disease phenotype. In the wild-type (WT) background, L-R172W mice exhibited late-onset (12-month) dominant cone degeneration without any apparent effect on rods. The degeneration was slightly accelerated (9 months) in the rds(+/-) background. L-R172W retinas did not form outer segments in the absence of endogenous Rds. With use of the H-R172W line on an rds(+/-) background for proof-of-principle genetic supplementation studies, the NMP transgene product rescued rod and cone functional defects and supported outer segment integrity up to 3 months of age, but the rescue effect did not persist in older (11-month) animals. The R172W mutation leads to dominant cone degeneration in the mouse model, regardless of the expression level of the transgene. In contrast, effects of the mutation on rods are dose dependent, underscoring the usefulness of the L-R172W line as a faithful model of the human phenotype. This model may prove helpful in future studies on the mechanisms of cone degeneration and for elucidating the different roles of Rds in rods and cones. This study provides evidence that Rds genetic supplementation can be used to partially rescue visual function. Although this strategy is capable of rescuing haploinsufficiency, it does not rescue the long-term degeneration associated with a gain-of-function mutation.

Mouse Models in Bone Marrow Transplantation and Adoptive Cellular Therapy

PubMed Central

Arber, Caroline; Brenner, Malcolm K.; Reddy, Pavan

2014-01-01

Mouse models of transplantation have been indispensable to the development of bone marrow transplantation (BMT). Their role in the generation of basic science knowledge is invaluable and is subject to discussion below. However, this article focuses on the direct role and relevance of mouse models towards the clinical development and advances in BMT and adoptive T-cell therapy for human diseases. The authors aim to present a thoughtful perspective on the pros and cons of mouse models while noting that despite imperfections these models are obligatory for the development of science-based medicine. PMID:24216170

Hybrid light transport model based bioluminescence tomography reconstruction for early gastric cancer detection

NASA Astrophysics Data System (ADS)

Chen, Xueli; Liang, Jimin; Hu, Hao; Qu, Xiaochao; Yang, Defu; Chen, Duofang; Zhu, Shouping; Tian, Jie

2012-03-01

Gastric cancer is the second cause of cancer-related death in the world, and it remains difficult to cure because it has been in late-stage once that is found. Early gastric cancer detection becomes an effective approach to decrease the gastric cancer mortality. Bioluminescence tomography (BLT) has been applied to detect early liver cancer and prostate cancer metastasis. However, the gastric cancer commonly originates from the gastric mucosa and grows outwards. The bioluminescent light will pass through a non-scattering region constructed by gastric pouch when it transports in tissues. Thus, the current BLT reconstruction algorithms based on the approximation model of radiative transfer equation are not optimal to handle this problem. To address the gastric cancer specific problem, this paper presents a novel reconstruction algorithm that uses a hybrid light transport model to describe the bioluminescent light propagation in tissues. The radiosity theory integrated with the diffusion equation to form the hybrid light transport model is utilized to describe light propagation in the non-scattering region. After the finite element discretization, the hybrid light transport model is converted into a minimization problem which fuses an l1 norm based regularization term to reveal the sparsity of bioluminescent source distribution. The performance of the reconstruction algorithm is first demonstrated with a digital mouse based simulation with the reconstruction error less than 1mm. An in situ gastric cancer-bearing nude mouse based experiment is then conducted. The primary result reveals the ability of the novel BLT reconstruction algorithm in early gastric cancer detection.

Centralized mouse repositories.

PubMed

Donahue, Leah Rae; Hrabe de Angelis, Martin; Hagn, Michael; Franklin, Craig; Lloyd, K C Kent; Magnuson, Terry; McKerlie, Colin; Nakagata, Naomi; Obata, Yuichi; Read, Stuart; Wurst, Wolfgang; Hörlein, Andreas; Davisson, Muriel T

2012-10-01

Because the mouse is used so widely for biomedical research and the number of mouse models being generated is increasing rapidly, centralized repositories are essential if the valuable mouse strains and models that have been developed are to be securely preserved and fully exploited. Ensuring the ongoing availability of these mouse strains preserves the investment made in creating and characterizing them and creates a global resource of enormous value. The establishment of centralized mouse repositories around the world for distributing and archiving these resources has provided critical access to and preservation of these strains. This article describes the common and specialized activities provided by major mouse repositories around the world.

Centralized Mouse Repositories

PubMed Central

Donahue, Leah Rae; de Angelis, Martin Hrabe; Hagn, Michael; Franklin, Craig; Lloyd, K. C. Kent; Magnuson, Terry; McKerlie, Colin; Nakagata, Naomi; Obata, Yuichi; Read, Stuart; Wurst, Wolfgang; Hörlein, Andreas; Davisson, Muriel T.

2013-01-01

Because the mouse is used so widely for biomedical research and the number of mouse models being generated is increasing rapidly, centralized repositories are essential if the valuable mouse strains and models that have been developed are to be securely preserved and fully exploited. Ensuring the ongoing availability of these mouse strains preserves the investment made in creating and characterizing them and creates a global resource of enormous value. The establishment of centralized mouse repositories around the world for distributing and archiving these resources has provided critical access to and preservation of these strains. This article describes the common and specialized activities provided by major mouse repositories around the world. PMID:22945696

ALS patients' regulatory T lymphocytes are dysfunctional, and correlate with disease progression rate and severity.

PubMed

Beers, David R; Zhao, Weihua; Wang, Jinghong; Zhang, Xiujun; Wen, Shixiang; Neal, Dan; Thonhoff, Jason R; Alsuliman, Abdullah S; Shpall, Elizabeth J; Rezvani, Katy; Appel, Stanley H

2017-03-09

Neuroinflammation is a pathological hallmark of ALS in both transgenic rodent models and patients, and is characterized by proinflammatory T lymphocytes and activated macrophages/microglia. In ALS mouse models, decreased regulatory T lymphocytes (Tregs) exacerbate the neuroinflammatory process, leading to accelerated motoneuron death and shortened survival; passive transfer of Tregs suppresses the neuroinflammation and prolongs survival. Treg numbers and FOXP3 expression are also decreased in rapidly progressing ALS patients. A key question is whether the marked neuroinflammation in ALS can be attributed to the impaired suppressive function of ALS Tregs in addition to their decreased numbers. To address this question, T lymphocyte proliferation assays were performed. Compared with control Tregs, ALS Tregs were less effective in suppressing responder T lymphocyte proliferation. Although both slowly and rapidly progressing ALS patients had dysfunctional Tregs, the greater the clinically assessed disease burden or the more rapidly progressing the patient, the greater the Treg dysfunction. Epigenetically, the percentage methylation of the Treg-specific demethylated region was greater in ALS Tregs. After in vitro expansion, ALS Tregs regained suppressive abilities to the levels of control Tregs, suggesting that autologous passive transfer of expanded Tregs might offer a novel cellular therapy to slow disease progression.

Mouse cloning and somatic cell reprogramming using electrofused blastomeres.

PubMed

Riaz, Amjad; Zhao, Xiaoyang; Dai, Xiangpeng; Li, Wei; Liu, Lei; Wan, Haifeng; Yu, Yang; Wang, Liu; Zhou, Qi

2011-05-01

Mouse cloning from fertilized eggs can assist development of approaches for the production of "genetically tailored" human embryonic stem (ES) cell lines that are not constrained by the limitations of oocyte availability. However, to date only zygotes have been successfully used as recipients of nuclei from terminally differentiated somatic cell donors leading to ES cell lines. In fertility clinics, embryos of advanced embryonic stages are usually stored for future use, but their ability to support the derivation of ES cell lines via somatic nuclear transfer has not yet been proved. Here, we report that two-cell stage electrofused mouse embryos, arrested in mitosis, can support developmental reprogramming of nuclei from donor cells ranging from blastomeres to somatic cells. Live, full-term cloned pups from embryonic donors, as well as pluripotent ES cell lines from embryonic or somatic donors, were successfully generated from these reconstructed embryos. Advanced stage pre-implantation embryos were unable to develop normally to term after electrofusion and transfer of a somatic cell nucleus, indicating that discarded pre-implantation human embryos could be an important resource for research that minimizes the ethical concerns for human therapeutic cloning. Our approach provides an attractive and practical alternative to therapeutic cloning using donated oocytes for the generation of patient-specific human ES cell lines.

Interpretation of magnetization transfer from inhomogeneously broadened lines (ihMT) in tissues as a dipolar order effect within motion restricted molecules

NASA Astrophysics Data System (ADS)

Varma, G.; Girard, O. M.; Prevost, V. H.; Grant, A. K.; Duhamel, G.; Alsop, D. C.

2015-11-01

Comparison of off-resonance saturation with single and dual frequency irradiation indicates a contribution of inhomogeneously broadened lines to magnetization transfer in tissues. This inhomogeneous magnetization transfer (ihMT) phenomenon can be exploited to produce images that highlight tissues containing myelin, in vivo. Here, a model for ihMT is described that includes dipolar order effects from magnetization associated with motion-restricted macromolecules. In this model, equal irradiation at positive and negative frequency offsets eliminates dipolar order and achieves greater saturation than irradiation at a single offset frequency using the same power. Fitting of mouse and human volunteer brain data at different irradiation powers and offset frequencies was performed to assess the relevance of the model and approximate tissue parameters. A key parameter in determining ihMT signal was found to be the relaxation time T1D associated with the dipolar order reservoir and the fraction f of the semi-solid, bound magnetization that possessed a nonzero T1D. Indeed, better fits of myelinated tissue were achieved when assuming f ≠ 1. From such fits, estimated T1Ds of mice in the white matter, (34 ± 14)ms, were much longer than in muscle, T1D = (1 ± 1)ms and the average f from white matter volunteer data was 2.2 times greater than that in grey matter. The combination of f and longer T1Ds was primarily responsible for the much higher ihMT in myelinated tissues, and provided explanation for the species variation. This dipolar order ihMT model should help guide future research, pulse sequence optimization, and clinical applications.

Molecular Indicators of Stress-Induced Neuroinflammation in a Mouse Model Simulating Features of Post-Traumatic Stress Disorder (Open Access)

DTIC Science & Technology

2017-05-23

OPEN ORIGINAL ARTICLE Molecular indicators of stress-induced neuroinflammation in a mouse model simulating features of post -traumatic stress disorder... post -traumatic stress disorder (PTSD). The model involved exposure of an intruder (male C57BL/6) mouse to a resident aggressor (male SJL) mouse for 5...revealed that neurogenesis and synaptic plasticity pathways were activated during the early responses but were inhibited after the later post -trauma

Social transfer of alcohol withdrawal-induced hyperalgesia in female prairie voles.

PubMed

Walcott, Andre T; Smith, Monique L; Loftis, Jennifer M; Ryabinin, Andrey E

2018-03-27

The expression of pain serves as a way for animals to communicate potential dangers to nearby conspecifics. Recent research demonstrated that mice undergoing alcohol or morphine withdrawal, or inflammation, could socially communicate their hyperalgesia to nearby mice. However, it is unknown whether such social transfer of hyperalgesia can be observed in other species of rodents. Therefore, the present study investigated if the social transfer of hyperalgesia occurs in the highly social prairie vole (Microtus ochrogaster). We observe that adult female prairie voles undergoing withdrawal from voluntary two-bottle choice alcohol drinking display an increase in nociception. This alcohol withdrawal-induced hypersensitiity is socially transferred to female siblings within the same cage and female strangers housed in separate cages within the same room. These experiments reveal that the social transfer of pain phenomenon is not specific to inbred mouse strains and that prairie voles display alcohol withdrawal and social transfer-induced hyperalgesia.

Reproduction of mouse-pup ultrasonic vocalizations by nanocrystalline silicon thermoacoustic emitter

NASA Astrophysics Data System (ADS)

Kihara, Takashi; Harada, Toshihiro; Kato, Masahiro; Nakano, Kiyoshi; Murakami, Osamu; Kikusui, Takefumi; Koshida, Nobuyoshi

2006-01-01

As one of the functional properties of ultrasound generator based on efficient thermal transfer at the nanocrystalline silicon (nc-Si) layer surface, its potential as an ultrasonic simulator of vocalization signals is demonstrated by using the acoustic data of mouse-pup calls. The device composed of a surface-heating thin-film electrode, an nc-Si layer, and a single-crystalline silicon (c-Si) wafer, exhibits an almost completely flat frequency response over a wide range without any mechanical surface vibration systems. It is shown that the fabricated emitter can reproduce digitally recorded ultrasonic mouse-pups vocalizations very accurately in terms of the call duration, frequency dispersion, and sound pressure level. The thermoacoustic nc-Si device provides a powerful physical means for the understanding of ultrasonic communication mechanisms in various living animals.

Genetic characterization and improved genotyping of the dysferlin-deficient mouse strain Dysf (tm1Kcam).

PubMed

Wiktorowicz, Tatiana; Kinter, Jochen; Kobuke, Kazuhiro; Campbell, Kevin P; Sinnreich, Michael

2015-01-01

Mouse models of dysferlinopathies are valuable tools with which to investigate the pathomechanisms underlying these diseases and to test novel therapeutic strategies. One such mouse model is the Dysf (tm1Kcam) strain, which was generated using a targeting vector to replace a 12-kb region of the dysferlin gene and which features a progressive muscular dystrophy. A prerequisite for successful animal studies using genetic mouse models is an accurate genotyping protocol. Unfortunately, the lack of robustness of currently available genotyping protocols for the Dysf (tm1Kcam) mouse has prevented efficient colony management. Initial attempts to improve the genotyping protocol based on the published genomic structure failed. These difficulties led us to analyze the targeted locus of the dysferlin gene of the Dysf (tm1Kcam) mouse in greater detail. In this study we resequenced and analyzed the targeted locus of the Dysf (tm1Kcam) mouse and developed a novel PCR protocol for genotyping. We found that instead of a deletion, the dysferlin locus in the Dysf (tm1Kcam) mouse carries a targeted insertion. This genetic characterization enabled us to establish a reliable method for genotyping of the Dysf (tm1Kcam) mouse, and thus has made efficient colony management possible. Our work will make the Dysf (tm1Kcam) mouse model more attractive for animal studies of dysferlinopathies.

Mouse models of neurodegenerative diseases: criteria and general methodology.

PubMed

Janus, Christopher; Welzl, Hans

2010-01-01

The major symptom of Alzheimer's disease is rapidly progressing dementia, coinciding with the formation of amyloid and tau deposits in the central nervous system, and neuronal death. At present familial cases of dementias provide the most promising foundation for modelling neurodegeneration. We describe the mnemonic and other major behavioral symptoms of tauopathies, briefly outline the genetics underlying familiar cases and discuss the arising implications for modelling the disease in mostly transgenic mouse lines. We then depict to what degree the most recent mouse models replicate pathological and cognitive characteristics observed in patients.There is no universally valid behavioral test battery to evaluate mouse models. The selection of individual tests depends on the behavioral and/or memory system in focus, the type of a model and how well it replicates the pathology of a disease and the amount of control over the genetic background of the mouse model. However it is possible to provide guidelines and criteria for modelling the neurodegeneration, setting up the experiments and choosing relevant tests. One should not adopt a "one (trans)gene, one disease" interpretation, but should try to understand how the mouse genome copes with the protein expression of the transgene in question. Further, it is not possible to recommend some mouse models over others since each model is valuable within its own constraints, and the way experiments are performed often reflects the idiosyncratic reality of specific laboratories. Our purpose is to improve bridging molecular and behavioural approaches in translational research.

Soiled-bedding Sentinel Detection of Murine Norovirus 4

PubMed Central

Manuel, Christopher A; Hsu, Charlie C; Riley, Lela K; Livingston, Robert S

2008-01-01

According to serologic surveys, murine norovirus (MNV) is the most prevalent viral pathogen infecting mice used in biomedical research. However, the use of sentinel mice to detect MNV-infected mouse populations has not been evaluated thoroughly. To this end, an experimental method of soiled bedding transfer was created to mimic a quarterly sentinel monitoring program. Soiled bedding (15 or 30 cm3) from ICR mice experimentally infected with MNV4 was transferred weekly to cages of pair-housed 4-wk-old ICR mice. After 12 wk, both mice in 80% (4 of 5) of cages receiving either 15 or 30 cm3 of soiled bedding were seropositive for MNV and were shedding virus in feces. To evaluate the stability of MNV RNA in mouse feces, fecal pellets from MNV-infected sentinel mice were stored at room temperature for as long as 14 d. After storage, all fecal samples tested positive for MNV by RT-PCR. To determine whether fecal samples could be pooled for MNV detection, 1 MNV-positive fecal pellet was combined with either 9 or 19 MNV-negative fecal pellets. All pooled fecal samples were positive for MNV by RT-PCR at both dilutions. These data indicate that although MNV-infected mouse populations can be detected by exposing sentinel mice to MNV-contaminated bedding, detection failures can occur. In addition, there was high agreement in the MNV infection status of cohoused sentinel mice. These data also demonstrate that MNV is readily detectable in pooled fecal samples and in mouse feces stored at room temperature for 2 wk. PMID:18459710

Noise in a Laboratory Animal Facility from the Human and Mouse Perspectives

PubMed Central

Reynolds, Randall P; Kinard, Will L; Degraff, Jesse J; Leverage, Ned; Norton, John N

2010-01-01

The current study was performed to understand the level of sound produced by ventilated racks, animal transfer stations, and construction equipment that mice in ventilated cages hear relative to what humans would hear in the same environment. Although the ventilated rack and animal transfer station both produced sound pressure levels above the ambient level within the human hearing range, the sound pressure levels within the mouse hearing range did not increase above ambient noise from either noise source. When various types of construction equipment were used 3 ft from the ventilated rack, the sound pressure level within the mouse hearing range was increased but to a lesser degree for each implement than were the sound pressure levels within the human hearing range. At more distant locations within the animal facility, sound pressure levels from the large jackhammer within the mouse hearing range decreased much more rapidly than did those in the human hearing range, indicating that less of the sound is perceived by mice than by humans. The relatively high proportion of low-frequency sound produced by the shot blaster, used without the metal shot that it normally uses to clean concrete, increased the sound pressure level above the ambient level for humans but did not increase sound pressure levels above ambient noise for mice at locations greater than 3 ft from inside of the cage, where sound was measured. This study demonstrates that sound clearly audible to humans in the animal facility may be perceived to a lesser degree or not at all by mice, because of the frequency content of the sound. PMID:20858361

Direct comparison of the pharmacodynamics of four antifungal drugs in a mouse model of disseminated candidiasis using microbiological assays of serum drug concentrations.

PubMed

Maki, Katsuyuki; Holmes, Ann R; Watabe, Etsuko; Iguchi, Yumi; Matsumoto, Satoru; Ikeda, Fumiaki; Tawara, Shuichi; Mutoh, Seitaro

2007-01-01

The aim of this study was to compare the pharmacodynamics of the azole antifungal drugs fluconazole, itraconazole and ketoconazole, and the polyene antifungal amphotericin B, in a mouse model of disseminated Candida albicans infection. In order to directly compare effective serum concentrations of these antifungals, drug concentrations were assayed microbiologically by measuring inhibition of C. albicans mycelial growth (mMIC) in a mouse serum-based assay (serum antifungal titer). Efficacy in the mouse infection model was determined using an organ-based (kidney burden) endpoint. For all four drugs, the serum antifungal titers, 8 hr after administration of single doses of drugs at a range of drug concentrations, correlated closely with C. albicans kidney fungal burden in the mouse model. The results showed that determining serum antifungal titer may be used to accurately represent kidney fungal burden in a mouse model of disseminated candidiasis and allowed direct comparison of the pharmacodynamics of differing classes of antifungal drugs.

Mouse Models as Tools to Identify Genetic Pathways for Retinal Degeneration, as Exemplified by Leber's Congenital Amaurosis.

PubMed

Chang, Bo

2016-01-01

Leber's congenital amaurosis (LCA) is an inherited retinal degenerative disease characterized by severe loss of vision in the first year of life. In addition to early vision loss, a variety of other eye-related abnormalities including roving eye movements, deep-set eyes, and sensitivity to bright light also occur with this disease. Many animal models of LCA are available and the study them has led to a better understanding of the pathology of the disease, and has led to the development of therapeutic strategies aimed at curing or slowing down LCA. Mouse models, with their well-developed genetics and similarity to human physiology and anatomy, serve as powerful tools with which to investigate the etiology of human LCA. Such mice provide reproducible, experimental systems for elucidating pathways of normal development, function, designing strategies and testing compounds for translational research and gene-based therapies aimed at delaying the diseases progression. In this chapter, I describe tools used in the discovery and evaluation of mouse models of LCA including a Phoenix Image-Guided Optical Coherence Tomography (OCT) and a Diagnosys Espion Visual Electrophysiology System. Three mouse models are described, the rd3 mouse model for LCA12 and LCA1, the rd12 mouse model for LCA2, and the rd16 mouse model for LCA10.

Differences in Pathogenesis for Salmonella enterica serovar Typhimurium in the Mouse Versus the Swine Model Identify Bacterial Gene Products Required for Systemic but not Gastrointestinal Disease

USDA-ARS?s Scientific Manuscript database

Over the last several decades, the mouse model of Typhoid fever has been an extremely productive model to investigate Salmonella enterica serovar Typhimurium pathogenesis. The mouse is the paradigm for investigating systemic disease due to infection by Salmonella; however, the swine model of gastro...

Weaker control of the electrical properties of cerebellar granule cells by tonically active GABAA receptors in the Ts65Dn mouse model of Down’s syndrome

PubMed Central

2013-01-01

Background Down’s syndrome (DS) is caused by triplication of all or part of human chromosome 21 and is characterized by a decrease in the overall size of the brain. One of the brain regions most affected is the cerebellum, in which the number of granule cells (GCs) is markedly decreased. GCs process sensory information entering the cerebellum via mossy fibres and pass it on to Purkinje cells and inhibitory interneurons. How GCs transform incoming signals depends on their input–output relationship, which is adjusted by tonically active GABAA receptor channels. Results We report that in the Ts65Dn mouse model of DS, in which cerebellar volume and GC number are decreased as in DS, the tonic GABAA receptor current in GCs is smaller than in wild-type mice and is less effective in moderating input resistance and raising the minimum current required for action potential firing. We also find that tonically active GABAA receptors curb the height and broaden the width of action potentials in wild-type GCs but not in Ts65Dn GCs. Single-cell real-time quantitative PCR reveals that these electrical differences are accompanied by decreased expression of the gene encoding the GABAA receptor β3 subunit but not genes coding for some of the other GABAA receptor subunits expressed in GCs (α1, α6, β2 and δ). Conclusions Weaker moderation of excitability and action potential waveform in GCs of the Ts65Dn mouse by tonically active GABAA receptors is likely to contribute to atypical transfer of information through the cerebellum. Similar changes may occur in DS. PMID:23870245

A unified model of the excitability of mouse sensory and motor axons.

PubMed

Makker, Preet G S; Matamala, José Manuel; Park, Susanna B; Lees, Justin G; Kiernan, Matthew C; Burke, David; Moalem-Taylor, Gila; Howells, James

2018-06-19

Non-invasive nerve excitability techniques have provided valuable insight into the understanding of neurological disorders. The widespread use of mice in translational research on peripheral nerve disorders and by pharmaceutical companies during drug development requires valid and reliable models that can be compared to humans. This study established a novel experimental protocol that enables comparative assessment of the excitability properties of motor and sensory axons at the same site in mouse caudal nerve, compared the mouse data to data for motor and sensory axons in human median nerve at the wrist, and constructed a mathematical model of the excitability of mouse axons. In a separate study, ischaemia was employed as an experimental manoeuvre to test the translational utility of this preparation. The patterns of mouse sensory and motor excitability were qualitatively similar to human studies under normal and ischaemic conditions. The most conspicuous differences between mouse and human studies were observed in the recovery cycle and the response to hyperpolarization. Modelling showed that an increase in temperature in mouse axons could account for most of the differences in the recovery cycle. The modelling also suggested a larger hyperpolarization-activated conductance in mouse axons. The kinetics of this conductance appeared to be much slower raising the possibility that an additional or different hyperpolarization-activated cyclic-nucleotide gated (HCN) channel isoform underlies the accommodation to hyperpolarization in mouse axons. Given a possible difference in HCN isoforms, caution should be exercised in extrapolating from studies of mouse motor and sensory axons to human nerve disorders. This article is protected by copyright. All rights reserved.

Rapamycin improves sociability in the BTBR T(+)Itpr3(tf)/J mouse model of autism spectrum disorders.

PubMed

Burket, Jessica A; Benson, Andrew D; Tang, Amy H; Deutsch, Stephen I

2014-01-01

Overactivation of the mammalian target of rapamycin (mTOR) has been implicated in the pathogenesis of syndromic forms of autism spectrum disorders (ASDs), such as tuberous sclerosis complex, neurofibromatosis 1, and fragile X syndrome. Administration of mTORC1 (mTOR complex 1) inhibitors (e.g. rapamycin) in syndromic mouse models of ASDs improved behavior, cognition, and neuropathology. However, since only a minority of ASDs are due to the effects of single genes (∼10%), there is a need to explore inhibition of mTOR activity in mouse models that may be more relevant to the majority of nonsyndromic presentations, such as the genetically inbred BTBR T(+)Itpr3(tf)/J (BTBR) mouse model of ASDs. BTBR mice have social impairment and exhibit increased stereotypic behavior. In prior work, d-cycloserine, a partial glycineB site agonist that targets the N-methyl-d-aspartate (NMDA) receptor, was shown to improve sociability in both Balb/c and BTBR mouse models of ASDs. Importantly, NMDA receptor activation regulates mTOR signaling activity. The current study investigated the ability of rapamycin (10mg/kg, i.p.×four days), an mTORC1 inhibitor, to improve sociability and stereotypic behavior in BTBR mice. Using a standard paradigm to assess mouse social behavior, rapamycin improved several measures of sociability in the BTBR mouse, suggesting that mTOR overactivation represents a therapeutic target that mediates or contributes to impaired sociability in the BTBR mouse model of ASDs. Interestingly, there was no effect of rapamycin on stereotypic behaviors in this mouse model. Copyright © 2013 Elsevier Inc. All rights reserved.

Reward-Related Behavioral Paradigms for Addiction Research in the Mouse: Performance of Common Inbred Strains

PubMed Central

Feyder, Michael; Brigman, Jonathan L.; Crombag, Hans S.; Saksida, Lisa M.; Bussey, Timothy J.; Holmes, Andrew

2011-01-01

The mouse has emerged as a uniquely valuable species for studying the molecular and genetic basis of complex behaviors and modeling neuropsychiatric disease states. While valid and reliable preclinical assays for reward-related behaviors are critical to understanding addiction-related processes, and various behavioral procedures have been developed and characterized in rats and primates, there have been relatively few studies using operant-based addiction-relevant behavioral paradigms in the mouse. Here we describe the performance of the C57BL/6J inbred mouse strain on three major reward-related paradigms, and replicate the same procedures in two other commonly used inbred strains (DBA/2J, BALB/cJ). We examined Pavlovian-instrumental transfer (PIT) by measuring the ability of an auditory cue associated with food reward to promote an instrumental (lever press) response. In a separate experiment, we assessed the acquisition and extinction of a simple stimulus-reward instrumental behavior on a touchscreen-based task. Reinstatement of this behavior was then examined following either continuous exposure to cues (conditioned reinforcers, CRs) associated with reward, brief reward and CR exposure, or brief reward exposure followed by continuous CR exposure. The third paradigm examined sensitivity of an instrumental (lever press) response to devaluation of food reward (a probe for outcome insensitive, habitual behavior) by repeated pairing with malaise. Results showed that C57BL/6J mice displayed robust PIT, as well as clear extinction and reinstatement, but were insensitive to reinforcer devaluation. DBA/2J mice showed good PIT and (rewarded) reinstatement, but were slow to extinguish and did not show reinforcer devaluation or significant CR-reinstatement. BALB/cJ mice also displayed good PIT, extinction and reinstatement, and retained instrumental responding following devaluation, but, unlike the other strains, demonstrated reduced Pavlovian approach behavior (food magazine head entries). Overall, these assays provide robust paradigms for future studies using the mouse to elucidate the neural, molecular and genetic factors underpinning reward-related behaviors relevant to addiction research. PMID:21249214

Reward-related behavioral paradigms for addiction research in the mouse: performance of common inbred strains.

PubMed

Lederle, Lauren; Weber, Susanna; Wright, Tara; Feyder, Michael; Brigman, Jonathan L; Crombag, Hans S; Saksida, Lisa M; Bussey, Timothy J; Holmes, Andrew

2011-01-10

The mouse has emerged as a uniquely valuable species for studying the molecular and genetic basis of complex behaviors and modeling neuropsychiatric disease states. While valid and reliable preclinical assays for reward-related behaviors are critical to understanding addiction-related processes, and various behavioral procedures have been developed and characterized in rats and primates, there have been relatively few studies using operant-based addiction-relevant behavioral paradigms in the mouse. Here we describe the performance of the C57BL/6J inbred mouse strain on three major reward-related paradigms, and replicate the same procedures in two other commonly used inbred strains (DBA/2J, BALB/cJ). We examined Pavlovian-instrumental transfer (PIT) by measuring the ability of an auditory cue associated with food reward to promote an instrumental (lever press) response. In a separate experiment, we assessed the acquisition and extinction of a simple stimulus-reward instrumental behavior on a touch screen based task. Reinstatement of this behavior was then examined following either continuous exposure to cues (conditioned reinforcers, CRs) associated with reward, brief reward and CR exposure, or brief reward exposure followed by continuous CR exposure. The third paradigm examined sensitivity of an instrumental (lever press) response to devaluation of food reward (a probe for outcome insensitive, habitual behavior) by repeated pairing with malaise. Results showed that C57BL/6J mice displayed robust PIT, as well as clear extinction and reinstatement, but were insensitive to reinforcer devaluation. DBA/2J mice showed good PIT and (rewarded) reinstatement, but were slow to extinguish and did not show reinforcer devaluation or significant CR-reinstatement. BALB/cJ mice also displayed good PIT, extinction and reinstatement, and retained instrumental responding following devaluation, but, unlike the other strains, demonstrated reduced Pavlovian approach behavior (food magazine head entries). Overall, these assays provide robust paradigms for future studies using the mouse to elucidate the neural, molecular and genetic factors underpinning reward-related behaviors relevant to addiction research.

In Vivo Hyperthermic Stress Model: An Easy Tool to Study the Effects of Oxidative Stress on Neuronal Tau Functionality in Mouse Brain.

PubMed

Chauderlier, Alban; Delattre, Lucie; Buée, Luc; Galas, Marie-Christine

2017-01-01

Oxidative damage is an early event in neurodegenerative disorders such as Alzheimer disease. To increase oxidative stress in AD-related mouse models is essential to study early mechanisms involved in the physiopathology of these diseases. In this chapter, we describe an experimental mouse model of transient and acute hyperthermic stress to induce in vivo an increase of oxidative stress in the brain of any kind of wild-type or transgenic mouse.

α-Synuclein propagates from mouse brain to grafted dopaminergic neurons and seeds aggregation in cultured human cells

PubMed Central

Hansen, Christian; Angot, Elodie; Bergström, Ann-Louise; Steiner, Jennifer A.; Pieri, Laura; Paul, Gesine; Outeiro, Tiago F.; Melki, Ronald; Kallunki, Pekka; Fog, Karina; Li, Jia-Yi; Brundin, Patrik

2011-01-01

Post-mortem analyses of brains from patients with Parkinson disease who received fetal mesencephalic transplants show that α-synuclein–containing (α-syn–containing) Lewy bodies gradually appear in grafted neurons. Here, we explored whether intercellular transfer of α-syn from host to graft, followed by seeding of α-syn aggregation in recipient neurons, can contribute to this phenomenon. We assessed α-syn cell-to-cell transfer using microscopy, flow cytometry, and high-content screening in several coculture model systems. Coculturing cells engineered to express either GFP– or DsRed-tagged α-syn resulted in a gradual increase in double-labeled cells. Importantly, α-syn–GFP derived from 1 neuroblastoma cell line localized to red fluorescent aggregates in other cells expressing DsRed–α-syn, suggesting a seeding effect of transmitted α-syn. Extracellular α-syn was taken up by cells through endocytosis and interacted with intracellular α-syn. Next, following intracortical injection of recombinant α-syn in rats, we found neuronal uptake was attenuated by coinjection of an endocytosis inhibitor. Finally, we demonstrated in vivo transfer of α-syn between host cells and grafted dopaminergic neurons in mice overexpressing human α-syn. In summary, intercellularly transferred α-syn interacts with cytoplasmic α-syn and can propagate α-syn pathology. These results suggest that α-syn propagation is a key element in the progression of Parkinson disease pathology. PMID:21245577

Real-time PCR to determine transgene copy number and to quantitate the biolocalization of adoptively transferred cells from EGFP-transgenic mice.

PubMed

Joshi, Molishree; Keith Pittman, H; Haisch, Carl; Verbanac, Kathryn

2008-09-01

Quantitative real-time PCR (qPCR) is a sensitive technique for the detection and quantitation of specific DNA sequences. Here we describe a Taqman qPCR assay for quantification of tissue-localized, adoptively transferred enhanced green fluorescent protein (EGFP)-transgenic cells. A standard curve constructed from serial dilutions of a plasmid containing the EGFP transgene was (i) highly reproducible, (ii) detected as few as two copies, and (iii) was included in each qPCR assay. qPCR analysis of genomic DNA was used to determine transgene copy number in several mouse strains. Fluorescent microscopy of tissue sections showed that adoptively transferred vascular endothelial cells (VEC) from EGFP-transgenic mice specifically localized to tissue with metastatic tumors in syngeneic recipients. VEC microscopic enumeration of liver metastases strongly correlated with qPCR analysis of identical sections (Pearson correlation 0.81). EGFP was undetectable in tissue from control mice by qPCR. In another study using intra-tumor EGFP-VEC delivery to subcutaneous tumors, manual cell count and qPCR analysis of alternating sections also strongly correlated (Pearson correlation 0.82). Confocal microscopy of the subcutaneous tumor sections determined that visual fluorescent signals were frequently tissue artifacts. This qPCR methodology offers specific, objective, and rapid quantitation, uncomplicated by tissue autofluorescence, and should be readily transferable to other in vivo models to quantitate the biolocalization of transplanted cells.

Implementation of a manual for working with wobbler mice and criteria for discontinuation of the experiment.

PubMed

Ott, Bastian; Dahlke, Carolin; Meller, Karl; Napirei, Markus; Schmitt-John, Thomas; Brand-Saberi, Beate; Theiss, Carsten; Saberi, Darius

2015-07-01

Mouse breeding is of importance to a whole range of medical and biological research. There are many known mouse models for motor neuron diseases. However, it must be kept in mind that especially mouse models for amyotrophic lateral sclerosis develop severe symptoms causing intense stress. This article is designed to summarize conscientious work with the wobbler mouse, a model for the sporadic form of amyotrophic lateral sclerosis. This mouse model is characterized by a degeneration of α-motor-neurons leading to head tremor, loss of body weight and rapidly progressive paralysis. Although this mouse model has been known since 1956, there are no guidelines for breeding wobbler mice. Due to the lack of such guidelines the present study tries to close this gap and implements a manual for further studies. It includes the whole workflow in regard to wobbler mice from breeding and animal care taking, genotyping and phenotype analysis, but also gives some examples for the use of various neuronal tissues for histological investigation. Beside the progress in research a second aim should always be the enhancement of mouse welfare and reduction of stress for the laboratory animals. Copyright © 2015 Elsevier GmbH. All rights reserved.

Cross-reactive mouse monoclonal antibodies raised against the hemagglutinin of A/Shanghai/1/2013 (H7N9) protect against novel H7 virus isolates in the mouse model.

PubMed

Stadlbauer, Daniel; Amanat, Fatima; Strohmeier, Shirin; Nachbagauer, Raffael; Krammer, Florian

2018-06-20

Influenza viruses remain a major global public health risk. In addition to seasonal influenza viruses, epizootic influenza A H7 subtype viruses of both the Asian and North American lineage are of concern due to their pandemic potential. In China, the simultaneous occurrence of H7N9 zoonotic episodes and seasonal influenza virus epidemics could potentially lead to novel reassortant viruses with the ability to efficiently spread among humans. Recently, the H7N9 virus has evolved into two new lineages, the Pearl River Delta and the Yangtze River Delta clade. This development has also resulted in viruses with a polybasic cleavage site in the hemagglutinin that are highly pathogenic in avian species and have caused human infections. In addition, an outbreak of a highly pathogenic H7N8 strain was reported in the US state of Indiana in 2016. Furthermore, an H7N2 feline virus strain caused an outbreak in cats in an animal shelter in New York City in 2016, resulting in one human zoonotic event. In this study, mouse monoclonal antibodies previously raised against the hemagglutinin of the A/Shanghai/1/2013 (H7N9) virus were tested for their (cross-) reactivity to these novel H7 viruses. Moreover, the functionality of these antibodies was assessed in vitro in hemagglutination inhibition and microneutralization assays. The therapeutic and prophylactic efficacy of the broadly reactive antibodies against novel H7 viruses was determined in vivo in mouse passive transfer-viral challenge experiments. Our results provide data about the conservation of critical H7 epitopes and could inform the selection of pre-pandemic H7 vaccine strains.

Psychological Stress on Female Mice Diminishes the Developmental Potential of Oocytes: A Study Using the Predatory Stress Model

PubMed Central

Liu, Yu-Xiang; Cheng, Ya-Nan; Miao, Yi-Long; Wei, De-Li; Zhao, Li-Hua; Luo, Ming-Jiu; Tan, Jing-He

2012-01-01

Although the predatory stress experimental protocol is considered more psychological than the restraint protocol, it has rarely been used to study the effect of psychological stress on reproduction. Few studies exist on the direct effect of psychological stress to a female on developmental competence of her oocytes, and the direct effect of predatory maternal stress on oocytes has not been reported. In this study, a predatory stress system was first established for mice with cats as predators. Beginning 24 h after injection of equine chorionic gonadotropin, female mice were subjected to predatory stress for 24 h. Evaluation of mouse responses showed that the predatory stress system that we established increased anxiety-like behaviors and plasma cortisol concentrations significantly and continuously while not affecting food and water intake of the mice. In vitro experiments showed that whereas oocyte maturation and Sr2+ activation or fertilization were unaffected by maternal predatory stress, rate of blastocyst formation and number of cells per blastocyst decreased significantly in stressed mice compared to non-stressed controls. In vivo embryo development indicated that both the number of blastocysts recovered per donor mouse and the average number of young per recipient after embryo transfer of blastocysts with similar cell counts were significantly lower in stressed than in unstressed donor mice. It is concluded that the predatory stress system we established was both effective and durative to induce mouse stress responses. Furthermore, predatory stress applied during the oocyte pre-maturation stage significantly impaired oocyte developmental potential while exerting no measurable impact on nuclear maturation, suggesting that cytoplasmic maturation of mouse oocytes was more vulnerable to maternal stress than nuclear maturation. PMID:23118931

PreImplantation Factor (PIF) correlates with early mammalian embryo development-bovine and murine models

PubMed Central

2011-01-01

Background PreImplantation Factor (PIF), a novel peptide secreted by viable embryos is essential for pregnancy: PIF modulates local immunity, promotes decidual pro-adhesion molecules and enhances trophoblast invasion. To determine the role of PIF in post-fertilization embryo development, we measured the peptide's concentration in the culture medium and tested endogenous PIF's potential trophic effects and direct interaction with the embryo. Methods Determine PIF levels in culture medium of multiple mouse and single bovine embryos cultured up to the blastocyst stage using PIF-ELISA. Examine the inhibitory effects of anti-PIF-monoclonal antibody (mAb) added to medium on cultured mouse embryos development. Test FITC-PIF uptake by cultured bovine blastocysts using fluorescent microscopy. Results PIF levels in mouse embryo culture medium significantly increased from the morula to the blastocyst stage (ANOVA, P = 0.01). In contrast, atretic embryos medium was similar to the medium only control. Detectable - though low - PIF levels were secreted already by 2-cell stage mouse embryos. In single bovine IVF-derived embryos, PIF levels in medium at day 3 of culture were higher than non-cleaving embryos (control) (P = 0.01) and at day 7 were higher than day 3 (P = 0.03). In non-cleaving embryos culture medium was similar to medium alone (control). Anti-PIF-mAb added to mouse embryo cultures lowered blastocyst formation rate 3-fold in a dose-dependent manner (2-way contingency table, multiple groups, X2; P = 0.01) as compared with non-specific mouse mAb, and medium alone, control. FITC-PIF was taken-up by cultured bovine blastocysts, but not by scrambled FITC-PIF (control). Conclusions PIF is an early embryo viability marker that has a direct supportive role on embryo development in culture. PIF-ELISA use to assess IVF embryo quality prior to transfer is warranted. Overall, our data supports PIF's endogenous self sustaining role in embryo development and the utility of PIF- ELISA to detect viable embryos in a non-invasive manner. PMID:21569635

Rational Design of Mouse Models for Cancer Research.

PubMed

Landgraf, Marietta; McGovern, Jacqui A; Friedl, Peter; Hutmacher, Dietmar W

2018-03-01

The laboratory mouse is widely considered as a valid and affordable model organism to study human disease. Attempts to improve the relevance of murine models for the investigation of human pathologies led to the development of various genetically engineered, xenograft and humanized mouse models. Nevertheless, most preclinical studies in mice suffer from insufficient predictive value when compared with cancer biology and therapy response of human patients. We propose an innovative strategy to improve the predictive power of preclinical cancer models. Combining (i) genomic, tissue engineering and regenerative medicine approaches for rational design of mouse models with (ii) rapid prototyping and computational benchmarking against human clinical data will enable fast and nonbiased validation of newly generated models. Copyright © 2017 Elsevier Ltd. All rights reserved.

Cytoplasmic asters are required for progression past the first cell cycle in cloned mouse embryos.

PubMed

Miki, Hiromi; Inoue, Kimiko; Ogonuki, Narumi; Mochida, Keiji; Nagashima, Hiroshi; Baba, Tadashi; Ogura, Atsuo

2004-12-01

Unlike the oocytes of most other animal species, unfertilized murine oocytes contain cytoplasmic asters, which act as microtubule-organizing centers following fertilization. This study examined the role of asters during the first cell cycle of mouse nuclear transfer (NT) embryos. NT was performed by intracytoplasmic injection of cumulus cells. Cytoplasmic asters were localized by staining with an anti-alpha-tubulin antibody. Enucleation of MII oocytes caused no significant change in the number of cytoplasmic asters. The number of asters decreased after transfer of the donor nuclei into these enucleated oocytes, probably because some of the asters participated in the formation of the spindle that anchors the donor chromosomes. The cytoplasmic asters became undetectable within 2 h of oocyte activation, irrespective of the presence or absence of the donor chromosomes. After the standard NT protocol, a spindle-like structure persisted between the pseudopronuclei of these oocytes throughout the pronuclear stage. The asters reappeared shortly before the first mitosis and formed the mitotic spindle. When the donor nucleus was transferred into preactivated oocytes (delayed NT) that were devoid of free asters, the microtubules and microfilaments were distributed irregularly in the ooplasm and formed dense bundles within the cytoplasm. Thereafter, all of the delayed NT oocytes underwent fragmentation and arrested development. Treatment of these delayed NT oocytes with Taxol, which is a microtubule-assembling agent, resulted in the formation of several aster-like structures and reduced fragmentation. Some Taxol-treated oocytes completed the first cell cycle and developed further. This study demonstrates that cytoplasmic asters play a crucial role during the first cell cycle of murine NT embryos. Therefore, in mouse NT, the use of MII oocytes as recipients is essential, not only for chromatin reprogramming as previously reported, but also for normal cytoskeletal organization in reconstructed oocytes.

Review of DoD Malaria Research Programs,

DTIC Science & Technology

1992-05-01

the irraliated sporozoite vaccine. Work in the mouse model system and then extrapolate to human malarias. Study naturally acquired immune ...recombinant vaccines. Work simultaneously in the mouse model system and with human malarias. 3. Identify targets and mechanisms of protective immunity not...multivalent vaccines that attack these same targets. 3. Working again in the mouse model, non- human primate model, andI human systems we

The use of carboxymethylcellulose gel to increase non-viral gene transfer in mouse airways

PubMed Central

Griesenbach, Uta; Meng, Cuixiang; Farley, Raymond; Wasowicz, Marguerite; Munkonge, Felix M; Chan, Mario; Stoneham, Charlotte; Sumner-Jones, Stephanie; Pringle, Ian A.; Gill, Deborah R.; Hyde, Stephen C.; Stevenson, Barbara; Holder, Emma; Ban, Hiroshi; Hasegawa, Mamoru; Cheng, Seng H; Scheule, Ronald K; Sinn, Patrick L; McCray, Paul B; Alton, Eric WFW

2014-01-01

We have assessed whether viscoelastic gels known to inhibit mucociliary clearance can increase lipid-mediated gene transfer. Methylcellulose or carboxymethylcellulose (0.25 to 1.5%) were mixed with complexes of the cationic lipid GL67A and plasmids encoding luciferase and perfused onto the nasal epithelium of mice. Survival after perfusion with 1% CMC or1% MC was 90 and 100%, respectively. In contrast 1.5% CMC was uniformly lethal likely due to the viscous solution blocking the airways. Perfusion with 0.5% CMC containing lipid/DNA complexes reproducibly increased gene expression by approximately 3-fold (n= 16, p<0.05). Given this benefit, likely related to increased duration of contact, we also assessed the effect of prolonging contact time of the liposome/DNA complexes by delivering our standard 80 μg DNA dose over either approximately 22 or 60 min of perfusion. This independently increased gene transfer by 6-fold (n=8, p<0.05) and could be further enhanced by the addition of 0.5% CMC, leading to an overall 25-fold enhancement (n=8, p<0.001) in gene expression. As a result of these interventions CFTR transgene mRNA transgene levels were increased several logs above background. Interestingly, this did not lead to correction of the ion transport defects in the nasal epithelium of cystic fibrosis mice nor for immunohistochemical quantification of CFTR expression. To assess if 0.5% CMC also increased gene transfer in the mouse lung, we used whole body nebulisation chambers. CMC was nebulised for 1 hr immediately before, or simultaneously with GL67A/pCIKLux. The former did not increase gene transfer, whereas co-administration significantly increased gene transfer by 4-fold (p<0.0001, n=18). This study suggests that contact time of non-viral gene transfer agents is a key factor for gene delivery, and suggests two methods which may be translatable for use in man. PMID:20022367

Functional imaging of the angiogenic switch in a transgenic mouse model of human breast cancer by dynamic contrast enhanced magnetic resonance imaging.

PubMed

Consolino, Lorena; Longo, Dario Livio; Dastrù, Walter; Cutrin, Juan Carlos; Dettori, Daniela; Lanzardo, Stefania; Oliviero, Salvatore; Cavallo, Federica; Aime, Silvio

2016-07-15

Tumour progression depends on several sequential events that include the microenvironment remodelling processes and the switch to the angiogenic phenotype, leading to new blood vessels recruitment. Non-invasive imaging techniques allow the monitoring of functional alterations in tumour vascularity and cellularity. The aim of this work was to detect functional changes in vascularisation and cellularity through Dynamic Contrast Enhanced (DCE) and Diffusion Weighted (DW) Magnetic Resonance Imaging (MRI) modalities during breast cancer initiation and progression of a transgenic mouse model (BALB-neuT mice). Histological examination showed that BALB-neuT mammary glands undergo a slow neoplastic progression from simple hyperplasia to invasive carcinoma, still preserving normal parts of mammary glands. DCE-MRI results highlighted marked functional changes in terms of vessel permeability (K(trans) , volume transfer constant) and vascularisation (vp , vascular volume fraction) in BALB-neuT hyperplastic mammary glands if compared to BALB/c ones. When breast tissue progressed from simple to atypical hyperplasia, a strong increase in DCE-MRI biomarkers was observed in BALB-neuT in comparison to BALB/c mice (K(trans)  = 5.3 ± 0.7E-4 and 3.1 ± 0.5E-4; vp  = 7.4 ± 0.8E-2 and 4.7 ± 0.6E-2 for BALB-neuT and BALB/c, respectively) that remained constant during the successive steps of the neoplastic transformation. Consistent with DCE-MRI observations, microvessel counting revealed a significant increase in tumour vessels. Our study showed that DCE-MRI estimates can accurately detect the angiogenic switch at early step of breast cancer carcinogenesis. These results support the view that this imaging approach is an excellent tool to characterize microvasculature changes, despite only small portions of the mammary glands developed neoplastic lesions in a transgenic mouse model. © 2016 UICC.

An Inducible Transgenic Mouse Model for Immune Mediated Hepatitis Showing Clearance of Antigen Expressing Hepatocytes by CD8+ T Cells

PubMed Central

Cebula, Marcin; Ochel, Aaron; Hillebrand, Upneet; Pils, Marina C.; Schirmbeck, Reinhold; Hauser, Hansjörg; Wirth, Dagmar

2013-01-01

The liver has the ability to prime immune responses against neo antigens provided upon infections. However, T cell immunity in liver is uniquely modulated by the complex tolerogenic property of this organ that has to also cope with foreign agents such as endotoxins or food antigens. In this respect, the nature of intrahepatic T cell responses remains to be fully characterized. To gain deeper insight into the mechanisms that regulate the CD8+ T cell responses in the liver, we established a novel OVA_X_CreERT2 mouse model. Upon tamoxifen administration OVA antigen expression is observed in a fraction of hepatocytes, resulting in a mosaic expression pattern. To elucidate the cross-talk of CD8+ T cells with antigen-expressing hepatocytes, we adoptively transferred Kb/OVA257-264-specific OT-I T cells to OVA_X_CreERT2 mice or generated triple transgenic OVA_X CreERT2_X_OT-I mice. OT-I T cells become activated in OVA_X_CreERT2 mice and induce an acute and transient hepatitis accompanied by liver damage. In OVA_X_CreERT2_X_OT-I mice, OVA induction triggers an OT-I T cell mediated, fulminant hepatitis resulting in 50% mortality. Surviving mice manifest a long lasting hepatitis, and recover after 9 weeks. In these experimental settings, recovery from hepatitis correlates with a complete loss of OVA expression indicating efficient clearance of the antigen-expressing hepatocytes. Moreover, a relapse of hepatitis can be induced upon re-induction of cured OVA_X_CreERT2_X_OT-I mice indicating absence of tolerogenic mechanisms. This pathogen-free, conditional mouse model has the advantage of tamoxifen inducible tissue specific antigen expression that reflects the heterogeneity of viral antigen expression and enables the study of intrahepatic immune responses to both de novo and persistent antigen. It allows following the course of intrahepatic immune responses: initiation, the acute phase and antigen clearance. PMID:23869228

Metabolic coupling of glutathione between mouse and quail cardiac myocytes and its protective role against oxidative stress.

PubMed

Nakamura, T Y; Yamamoto, I; Kanno, Y; Shiba, Y; Goshima, K

1994-05-01

Cultured quail myocytes were much more resistant to H2O2 toxicity than cultured mouse myocytes. The intracellular concentration of glutathione ([GSH]i) and the activity of gamma-glutamylcysteine synthetase (gamma-GCS) in quail heart cells were about five and three times higher, respectively, than in mouse heart cells, although catalase and glutathione peroxidase (GSHpx) activity was similar in both. Preloading of gamma-glutamylcysteine monoethyl ester (gamma-GCE), a membrane-permeating GSH precursor, increased the H2O2 resistance of cultured mouse myocytes. These observations suggest that the high [GSH]i and the high activity of gamma-GCS in quail myocytes are responsible for their high resistance to H2O2. Both H2O2 sensitivity and [GSH]i of mosaic sheets composed of equal amounts of mouse and quail myocytes approximated those of sheets composed entirely of quail myocytes. From these observations, it is hypothesized that GSH was transferred from quail myocytes to mouse myocytes, probably through gap junctions between them, and that quail myocytes resynthesized GSH by a feedback mechanism, thus maintaining their intracellular GSH levels. When the fluorescent dye lucifer yellow was injected into a beating quail myocyte in a mosaic sheet, it spread to neighboring mouse myocytes but not to neighboring L cells (a cell line derived from mouse connective tissue). These observations indicate that existence of gap junctions in the region of cell contact between mouse and quail myocytes but not between quail myocytes and L cells. When quail myocytes preloaded with [3H]gamma-GCE were cocultured with mouse myocytes and L cells, the radioactivity was transmitted to neighboring mouse myocytes but not L cells. These observations show that GSH and/or its precursors can be transmitted from quail myocytes to mouse myocytes through gap junctions and that this can protect mouse myocytes from H2O2 toxicity. Mouse myocyte sheets composed of 10(4) cells or more showed higher resistance to H2O2 toxicity than single isolated mouse myocytes. Metabolic coupling of GSH between myocytes may contribute at least in part to this high resistance of the cell sheets.

Treatment of d-galactose induced mouse aging with Lycium barbarum polysaccharides and its mechanism study.

PubMed

Tang, Tao; He, Bixiu

2013-01-01

We evaluated the effects of Lycium barbarum polysaccharides LBP) on D-galactose aging model mouse, and explored its possible mechanism. Kunming mice were randomly divided into the control group, the model group, the high-dose LBP group, and the low-dose LBP group. Except the control group, D-galactose was used for modelling. The drug was administrated when modelling. Mouse behavioural, learning and memory changes were observed, and the contents of lipid peroxidation (LPO), lipofuscin (LF) and monoamine oxidase B (MAO-B) in mouse brain tissue and the weight of immune organs were measured after 6 weeks. Compared with the control group, mouse weight gain in the model group reduced significantly. Compared with model group, after mice drank LBP, the times of electric shock was less than aging mice (in which, the high-dose LBP group, P<0.05), and electric shock incubation period was longer (P<0.01). On Day 45 after modelling and drug administration, the contents of LPO, LF and MAO-B in mouse brain tissue in the model group increased significantly, while those in the drug administration groups decreased significantly. The thymus index in the aging model group decreased significantly; the thymus index and the spleen index in the high-dose LBP group and the low-dose LBP group rebounded significantly (P<0.01). We concluded that LBP has an anti-aging effect on D-galactose induced aging model mouse, and its mechanism may be related with the alleviation of glucose metabolism disorder and the resistance of the generation of lipid peroxide and other substances, which damage cell membrane lipid.

Adoptive transfer of ex vivo expanded Vγ9Vδ2 T cells in combination with zoledronic acid inhibits cancer growth and limits osteolysis in a murine model of osteolytic breast cancer.

PubMed

Zysk, Aneta; DeNichilo, Mark O; Panagopoulos, Vasilios; Zinonos, Irene; Liapis, Vasilios; Hay, Shelley; Ingman, Wendy; Ponomarev, Vladimir; Atkins, Gerald; Findlay, David; Zannettino, Andrew; Evdokiou, Andreas

2017-02-01

Bone metastases occur in over 75% of patients with advanced breast cancer and are responsible for high levels of morbidity and mortality. In this study, ex vivo expanded cytotoxic Vγ9Vδ2 T cells isolated from human peripheral blood were tested for their anti-cancer efficacy in combination with zoledronic acid (ZOL), using a mouse model of osteolytic breast cancer. In vitro, expanded Vγ9Vδ2 T cells were cytotoxic against a panel of human breast cancer cell lines, and ZOL pre-treatment further sensitised breast cancer cells to killing by Vγ9Vδ2 T cells. Vγ9Vδ2 T cells adoptively transferred into NOD/SCID mice localised to osteolytic breast cancer lesions in the bone, and multiple infusions of Vγ9Vδ2 T cells reduced tumour growth in the bone. ZOL pre-treatment potentiated the anti-cancer efficacy of Vγ9Vδ2 T cells, with mice showing further reductions in tumour burden. Mice treated with the combination also had reduced tumour burden of secondary pulmonary metastases, and decreased bone degradation. Our data suggests that adoptive transfer of Vγ9Vδ2 T cell in combination with ZOL may prove an effective immunotherapeutic approach for the treatment of breast cancer bone metastases. Copyright © 2016. Published by Elsevier Ireland Ltd.

Plasmodium berghei infection ameliorates atopic dermatitis-like skin lesions in NC/Nga mice.

PubMed

Kishi, C; Amano, H; Suzue, K; Ishikawa, O

2014-10-01

Atopic diseases are more prevalent in industrialized countries than in developing countries. In addition, significant differences in the prevalence of allergic diseases are observed between rural and urban areas within the same country. This difference in prevalence has been attributed to what is called the 'hygiene hypothesis'. Although parasitic infections are known to protect against allergic reactions, the mechanism is still unknown. The aim of this study was to investigate whether or not malarial infections can inhibit atopic dermatitis (AD)-like skin lesions in a mouse model of AD. We used NC/Nga mice which are a model for AD. The NC/Nga mice were intraperitoneally infected with 1 × 10(5) Plasmoduim berghei (Pb) XAT-infected erythrocytes. Malarial infections ameliorated AD-like skin lesions in the NC/Nga mice. This improvement was blocked by the administration of anti-asialo GM1 antibodies, which are anti-natural killer (NK) cells. Additionally, adoptive transfer of NK cells markedly improved AD-like skin lesions in conventional NC/Nga mice; these suggest that the novel protective mechanism associated with malaria parasitic infections is at least, in part, dependent on NK cells. We have experimentally demonstrated for the first time that malarial infections ameliorated AD-like skin lesions in a mouse model of AD. Our study could explain in part the mechanism of the 'hygiene hypothesis', which states that parasitic infections can inhibit the development of allergic diseases. © 2014 The Authors. Allergy Published by John Wiley & Sons Ltd.

21 CFR 884.6160 - Assisted reproduction labware.

Code of Federal Regulations, 2011 CFR

2011-04-01

... equipment or supplies intended to prepare, store, manipulate, or transfer human gametes or embryos for in..., dishes, plates, and other vessels that come into physical contact with gametes, embryos or tissue culture media. (b)Classification. Class II (special controls) (mouse embryo assay information, endotoxin testing...

21 CFR 884.6160 - Assisted reproduction labware.

Code of Federal Regulations, 2012 CFR

2012-04-01

... equipment or supplies intended to prepare, store, manipulate, or transfer human gametes or embryos for in..., dishes, plates, and other vessels that come into physical contact with gametes, embryos or tissue culture media. (b)Classification. Class II (special controls) (mouse embryo assay information, endotoxin testing...

21 CFR 884.6160 - Assisted reproduction labware.

Code of Federal Regulations, 2014 CFR

2014-04-01

... equipment or supplies intended to prepare, store, manipulate, or transfer human gametes or embryos for in..., dishes, plates, and other vessels that come into physical contact with gametes, embryos or tissue culture media. (b)Classification. Class II (special controls) (mouse embryo assay information, endotoxin testing...

21 CFR 884.6160 - Assisted reproduction labware.

Code of Federal Regulations, 2013 CFR

2013-04-01

... equipment or supplies intended to prepare, store, manipulate, or transfer human gametes or embryos for in..., dishes, plates, and other vessels that come into physical contact with gametes, embryos or tissue culture media. (b)Classification. Class II (special controls) (mouse embryo assay information, endotoxin testing...

Fatal autoimmune hepatitis induced by concurrent loss of naturally arising regulatory T cells and PD-1-mediated signaling.

PubMed

Kido, Masahiro; Watanabe, Norihiko; Okazaki, Taku; Akamatsu, Takuji; Tanaka, Junya; Saga, Kazuyuki; Nishio, Akiyoshi; Honjo, Tasuku; Chiba, Tsutomu

2008-10-01

Because of the lack of animal models developing spontaneous autoimmune hepatitis (AIH), the molecular mechanisms involved in the development of AIH are still unclear. This study aims to examine the regulatory roles of naturally arising CD4(+)CD25(+) regulatory T (Treg) cells and programmed cell death 1 (PD-1)-mediated signaling in the development of AIH. To induce a concurrent loss of Treg cells and PD-1-mediated signaling, neonatal thymectomy (NTx), which severely reduces the number of Treg cells, was performed on PD-1(-/-) mice. After the NTx, we performed histologic examination, assessed autoantibody production and infiltrating cells in the liver, and conducted adoptive transfer experiments. In contrast to NTx mice and PD-1(-/-) mice, NTx-PD-1(-/-) mice produced antinuclear antibodies and developed fatal hepatitis characterized by a CD4(+) and CD8(+) T-cell infiltration invading the parenchyma with massive lobular necrosis. Induction of AIH in NTx-PD-1(-/-) mice was suppressed by transfer of Treg cells, even derived from PD-1(-/-) mice. Transfer of total but not CD4(+) T-cell-depleted splenocytes from NTx-PD-1(-/-) mice into RAG2(-/-) mice induced the development of severe hepatitis. In contrast, the transfer of CD8(+) T-cell-depleted splenocytes triggered only mononuclear infiltrates without massive necrosis of the parenchyma. NTx-PD-1(-/-) mice are the first mouse model of spontaneous fatal AIH. The concurrent loss of Treg cells and PD-1-mediated signaling can induce the development of fatal AIH. Autoreactive CD4(+) T cells are essential for induction of AIH, whereas CD8(+) T cells play an important role in progression to fatal hepatic damage.

Differential Regulatory Role of Pituitary Adenylate Cyclase–Activating Polypeptide in the Serum-Transfer Arthritis Model

PubMed Central

Botz, Bálint; Bölcskei, Kata; Kereskai, László; Kovács, Miklós; Németh, Tamás; Szigeti, Krisztián; Horváth, Ildikó; Máthé, Domokos; Kovács, Noémi; Hashimoto, Hitoshi; Reglődi, Dóra; Szolcsányi, János; Pintér, Erika; Mócsai, Attila; Helyes, Zsuzsanna

2014-01-01

Objective Pituitary adenylate cyclase–activating polypeptide (PACAP) expressed in capsaicin-sensitive sensory neurons and immune cells has divergent functions in inflammatory and pain processes. This study was undertaken to investigate the involvement of PACAP in a mouse model of rheumatoid arthritis. Methods Arthritis was induced in PACAP−/− and wild-type (PACAP+/+) mice by K/BxN serum transfer. General features of the disease were investigated by semiquantitative scoring, plethysmometry, and histopathologic analysis. Mechano- and thermonociceptive thresholds and motor functions were also evaluated. Metabolic activity was assessed by positron emission tomography. Bone morphology was measured by in vivo micro–computed tomography, myeloperoxidase activity and superoxide production by bioluminescence imaging with luminol and lucigenin, respectively, and vascular permeability by fluorescent indocyanine green dye study. Results PACAP+/+ mice developed notable joint swelling, reduced grasping ability, and mechanical (but not thermal) hyperalgesia after K/BxN serum transfer. In PACAP−/− mice clinical scores and edema were significantly reduced, and mechanical hyperalgesia and motor impairment were absent, throughout the 2-week period of observation. Metabolic activity and superoxide production increased in the tibiotarsal joints of wild-type mice but were significantly lower in PACAP−/− animals. Myeloperoxidase activity in the ankle joints of PACAP−/− mice was significantly reduced in the early phase of arthritis, but increased in the late phase. Synovial hyperplasia was also significantly increased, and progressive bone spur formation was observed in PACAP-deficient mice only. Conclusion In PACAP-deficient mice with serum-transfer arthritis, joint swelling, vascular leakage, hyperalgesia, and early inflammatory cell accumulation are reduced; in the later phase of the disease, immune cell function and bone neoformation are increased. Elucidation of the underlying pathways of PACAP activity may open promising new avenues for development of therapy in inflammatory arthritis. PMID:25048575

Mutagenicity testing with transgenic mice. Part II: Comparison with the mouse spot test

PubMed Central

Wahnschaffe, Ulrich; Bitsch, Annette; Kielhorn, Janet; Mangelsdorf, Inge

2005-01-01

The mouse spot test, an in vivo mutation assay, has been used to assess a number of chemicals. It is at present the only in vivo mammalian test system capable of detecting somatic gene mutations according to OECD guidelines (OECD guideline 484). It is however rather insensitive, animal consuming and expensive type of test. More recently several assays using transgenic animals have been developed. From data in the literature, the present study compares the results of in vivo testing of over twenty chemicals using the mouse spot test and compares them with results from the two transgenic mouse models with the best data base available, the lacI model (commercially available as the Big Blue® mouse), and the lacZ model (commercially available as the Muta™ Mouse). There was agreement in the results from the majority of substances. No differences were found in the predictability of the transgenic animal assays and the mouse spot test for carcinogenicity. However, from the limited data available, it seems that the transgenic mouse assay has several advantages over the mouse spot test and may be a suitable test system replacing the mouse spot test for detection of gene but not chromosome mutations in vivo. PMID:15676065

The Mouse Genome Database (MGD): facilitating mouse as a model for human biology and disease.

PubMed

Eppig, Janan T; Blake, Judith A; Bult, Carol J; Kadin, James A; Richardson, Joel E

2015-01-01

The Mouse Genome Database (MGD, http://www.informatics.jax.org) serves the international biomedical research community as the central resource for integrated genomic, genetic and biological data on the laboratory mouse. To facilitate use of mouse as a model in translational studies, MGD maintains a core of high-quality curated data and integrates experimentally and computationally generated data sets. MGD maintains a unified catalog of genes and genome features, including functional RNAs, QTL and phenotypic loci. MGD curates and provides functional and phenotype annotations for mouse genes using the Gene Ontology and Mammalian Phenotype Ontology. MGD integrates phenotype data and associates mouse genotypes to human diseases, providing critical mouse-human relationships and access to repositories holding mouse models. MGD is the authoritative source of nomenclature for genes, genome features, alleles and strains following guidelines of the International Committee on Standardized Genetic Nomenclature for Mice. A new addition to MGD, the Human-Mouse: Disease Connection, allows users to explore gene-phenotype-disease relationships between human and mouse. MGD has also updated search paradigms for phenotypic allele attributes, incorporated incidental mutation data, added a module for display and exploration of genes and microRNA interactions and adopted the JBrowse genome browser. MGD resources are freely available to the scientific community. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

Genetically engineered mouse models of craniopharyngioma: an opportunity for therapy development and understanding of tumor biology

PubMed Central

Martinez‐Barbera, Juan Pedro

2017-01-01

Abstract Adamantinomatous craniopharyngioma (ACP) is the commonest tumor of the sellar region in childhood. Two genetically engineered mouse models have been developed and are giving valuable insights into ACP biology. These models have identified novel pathways activated in tumors, revealed an important function of paracrine signalling and extended conventional theories about the role of organ‐specific stem cells in tumorigenesis. In this review, we summarize these mouse models, what has been learnt, their limitations and open questions for future research. We then discussed how these mouse models may be used to test novel therapeutics against potentially targetable pathways recently identified in human ACP. PMID:28414891

17-β Estradiol reduces atherosclerosis without exacerbating lupus in ovariectomized systemic lupus erythematosus-susceptible LDLr(-/-) mice.

PubMed

Shelton, K A; Cline, J M; Cann, J A

2013-04-01

To test the hypothesis that estrogen treatment in a radiation chimera mouse model of systemic lupus erythematosus (SLE) and atherosclerosis will increase SLE-associated atherosclerosis by increasing autoantibody production and inflammation. We used a radiation chimera mouse model in which bone marrow from the polygenic B6.Sle1.2.3 model of SLE was transferred to the low density lipoprotein receptor knock out (LDLr(-/-)) model of atherosclerosis on a C57BL/6 background (Sle/LDLr(-/-)). Ovariectomized chimeric mice were treated for 10 weeks with either 5.6 μg/day of 17β-estradiol or placebo; outcomes included atherosclerosis plaque size, anti-dsDNA autoantibody production and renal pathology. Mean atherosclerosis plaque size was 67.4 ± 7.6% smaller in the estrogen treated group (p < 0.0001). Estrogen treated Sle/LDLr(-/-) mice had no significant difference in serum cholesterol concentration, lipoprotein distribution, anti-dsDNA autoantibody concentration, antibody isotype concentration and renal histopathology score compared to placebo. However, they had significantly lower mean urine protein to urine creatinine ratio (UP:UC). There was no correlation between atherosclerosis lesion size and either the renal histology score or UP:UC ratio in Sle/LDLr(-/-) mice. These results indicate that 17β-estradiol is atheroprotective within the context of murine SLE independent of changes in serum cholesterol concentration, autoantibody concentration, or renal pathology. The SLE phenotype in Sle/LDLr(-/-) mice is not exacerbated by exogenous 17β-estradiol administration, and the reduced UP:UC ratio suggests a protective effect against lupus nephritis. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

17-β Estradiol reduces atherosclerosis without exacerbating lupus in ovariectomized systemic lupus erythematosus-susceptible LDLr−/− mice

PubMed Central

Shelton, KA; Cline, JM; Cann, JA

2013-01-01

Objective To test the hypothesis that estrogen treatment in a radiation chimera mouse model of systemic lupus erythematosus (SLE) and atherosclerosis will increase SLE-associated atherosclerosis by increasing autoantibody production and inflammation. Methods We used a radiation chimera mouse model in which bone marrow from the polygenic B6.Sle1.2.3 model of SLE was transferred to the low density lipoprotein receptor knock out (LDLr−/−) model of atherosclerosis on a C57BL/6 background (Sle/LDLr−/−). Ovariectomized chimeric mice were treated for 10 weeks with either 5.6 ug/day of 17β-estradiol or placebo; outcomes included atherosclerosis plaque size, anti-dsDNA autoantibody production and renal pathology. Results Mean atherosclerosis plaque size was 67.4 ± 7.6% smaller in the estrogen treated group (p<0.0001). Estrogen treated Sle/LDLr−/− mice had no significant difference in serum cholesterol concentration, lipoprotein distribution, anti-dsDNA autoantibody concentration, antibody isotype concentration and renal histopathology score compared to placebo. However, they had significantly lower mean urine protein to urine creatinine ratio (UP:UC). There was no correlation between atherosclerosis lesion size and either the renal histology score or UP:UC ratio in Sle/LDLr−/− mice. Conclusion These results indicate that 17β-estradiol is atheroprotective within the context of murine SLE independent of changes in serum cholesterol concentration, autoantibody concentration, or renal pathology. The SLE phenotype in Sle/LDLr−/− mice is not exacerbated by exogenous 17β-estradiol administration, and the reduced UP:UC ratio suggests a protective effect against lupus nephritis. PMID:23395521

3D visualization and quantification of bone and teeth mineralization for the study of osteo/dentinogenesis in mice models

NASA Astrophysics Data System (ADS)

Marchadier, A.; Vidal, C.; Ordureau, S.; Lédée, R.; Léger, C.; Young, M.; Goldberg, M.

2011-03-01

Research on bone and teeth mineralization in animal models is critical for understanding human pathologies. Genetically modified mice represent highly valuable models for the study of osteo/dentinogenesis defects and osteoporosis. Current investigations on mice dental and skeletal phenotype use destructive and time consuming methods such as histology and scanning microscopy. Micro-CT imaging is quicker and provides high resolution qualitative phenotypic description. However reliable quantification of mineralization processes in mouse bone and teeth are still lacking. We have established novel CT imaging-based software for accurate qualitative and quantitative analysis of mouse mandibular bone and molars. Data were obtained from mandibles of mice lacking the Fibromodulin gene which is involved in mineralization processes. Mandibles were imaged with a micro-CT originally devoted to industrial applications (Viscom, X8060 NDT). 3D advanced visualization was performed using the VoxBox software (UsefulProgress) with ray casting algorithms. Comparison between control and defective mice mandibles was made by applying the same transfer function for each 3D data, thus allowing to detect shape, colour and density discrepencies. The 2D images of transverse slices of mandible and teeth were similar and even more accurate than those obtained with scanning electron microscopy. Image processing of the molars allowed the 3D reconstruction of the pulp chamber, providing a unique tool for the quantitative evaluation of dentinogenesis. This new method is highly powerful for the study of oro-facial mineralizations defects in mice models, complementary and even competitive to current histological and scanning microscopy appoaches.

Melanocortin MC1 receptor in human genetics and model systems

PubMed Central

Beaumont, Kimberley A.; Wong, Shu S.; Ainger, Stephen A.; Liu, Yan Yan; Patel, Mira P.; Millhauser, Glenn L.; Smith, Jennifer J.; Alewood, Paul F.; Leonard, J. Helen; Sturm, Richard A.

2011-01-01

The melanocortin MC1 receptor is a G -protein coupled receptor expressed in melanocytes of the skin and hair and is known for its key role in regulation of human pigmentation. Melanocortin MC1 receptor activation after ultraviolet radiation exposure results in a switch from the red/yellow pheomelanin to the brown/black eumelanin pigment synthesis within cutaneous melanocytes; this pigment is then transferred to the surrounding keratinocytes of the skin. The increase in melanin maturation and uptake results in tanning of the skin, providing a physical protection of skin cells from ultraviolet radiation induced DNA damage. Melanocortin MC 1 receptor polymorphism is widespread within the Caucasian population and some variant alleles are associated with red hair colour, fair skin, poor tanning and increased risk of skin cancer. Here we will discuss the use of mouse coat colour models, human genetic association studies, and in vitro cell culture studies to determine the complex functions of the melanocortin MC1 receptor and the molecular mechanisms underlying the association between melanocortin MC1 receptor variant alleles and the red hair colour phenotype. Recent research indicates that melanocortin MC1 receptor has many non-pigmentary functions, and that the increased risk of skin cancer conferred by melanocortin MC1 receptor variant alleles is to some extent independent of pigmentation phenotypes. The use of new transgenic mouse models, the study of novel melanocortin MC1 receptor response genes and the use of more advanced human skin models such as 3D skin reconstruction may provide key elements in understanding the pharmacogenetics of human melanocortin MC1 receptor polymorphism . PMID:21199646

Efficacy of a Trivalent Hand, Foot, and Mouth Disease Vaccine against Enterovirus 71 and Coxsackieviruses A16 and A6 in Mice.

PubMed

Caine, Elizabeth A; Fuchs, Jeremy; Das, Subash C; Partidos, Charalambos D; Osorio, Jorge E

2015-11-17

Hand, foot, and mouth disease (HFMD) has recently emerged as a major public health concern across the Asian-Pacific region. Enterovirus 71 (EV71) and Coxsackievirus A16 (CVA16) are the primary causative agents of HFMD, but other members of the Enterovirus A species, including Coxsackievirus A6 (CVA6), can cause disease. The lack of small animal models for these viruses have hampered the development of a licensed HFMD vaccine or antivirals. We have previously reported on the development of a mouse model for EV71 and demonstrated the protective efficacy of an inactivated EV71 vaccine candidate. Here, mouse-adapted strains of CVA16 and CVA6 were produced by sequential passage of the viruses through mice deficient in interferon (IFN) α/β (A129) and α/β and γ (AG129) receptors. Adapted viruses were capable of infecting 3 week-old A129 (CVA6) and 12 week-old AG129 (CVA16) mice. Accordingly, these models were used in active and passive immunization studies to test the efficacy of a trivalent vaccine candidate containing inactivated EV71, CVA16, and CVA6. Full protection from lethal challenge against EV71 and CVA16 was observed in trivalent vaccinated groups. In contrast, monovalent vaccinated groups with non-homologous challenges failed to cross protect. Protection from CVA6 challenge was accomplished through a passive transfer study involving serum raised against the trivalent vaccine. These animal models will be useful for future studies on HFMD related pathogenesis and the efficacy of vaccine candidates.

The In Vitro Response of Tissue Stem Cells to Irradiation With Different Linear Energy Transfers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nagle, Peter W.; Hosper, Nynke A.; Department of Radiation Oncology, University of Groningen, University Medical Center Groningen, Groningen

Purpose: A reduction in the dose, irradiated volume, and sensitivity of, in particular, normal tissue stem cells is needed to advance radiation therapy. This could be obtained with the use of particles for radiation therapy. However, the radiation response of normal tissue stem cells is still an enigma. Therefore, in the present study, we developed a model to investigate the in vitro response of stem cells to particle irradiation. Methods and Materials: We used the immortalized human salivary gland (HSG) cell line resembling salivary gland (SG) cells to translate the radiation response in 2-dimensional (2D) to 3-dimensional (3D) conditions. This responsemore » was subsequently translated to the response of SG stem cells (SGSCs). Dispersed single cells were irradiated with photons or carbon ions at different linear energy transfers (LETs; 48.76 ± 2.16, 149.9 ± 10.8, and 189 ± 15 keV/μm). Subsequently, 2D or 3D clonogenicity was determined by counting the colonies or secondary stem cell-derived spheres in Matrigel. γH2AX immunostaining was used to assess DNA double strand break repair. Results: The 2D response of HSG cells showed a similar increase in dose response to increasing higher LET irradiation as other cell lines. The 3D response of HSG cells to increasing LET irradiation was reduced compared with the 2D response. Finally, the response of mouse SGSCs to photons was similar to the 3D response of HSG cells. The response to higher LET irradiation was reduced in the stem cells. Conclusions: Mouse SGSC radiosensitivity seems reduced at higher LET radiation compared with transformed HSG cells. The developed model to assess the radiation response of SGSCs offers novel possibilities to study the radiation response of normal tissue in vitro.« less

Interspecies chimera between primate embryonic stem cells and mouse embryos: Monkey ESCs engraft into mouse embryos, but not post-implantation fetuses

PubMed Central

Simerly, Calvin; McFarland, Dave; Castro, Carlos; Lin, Chih-Cheng; Redinger, Carrie; Jacoby, Ethan; Mich-Basso, Jocelyn; Orwig, Kyle; Mills, Parker; Ahrens, Eric; Navara, Chris; Schatten, Gerald

2016-01-01

Unequivocal evidence for pluripotency in which embryonic stem cells contribute to chimeric offspring has yet to be demonstrated in human or nonhuman primates (NHPs). Here, rhesus and baboons ESCs were investigated in interspecific mouse chimera generated by aggregation or blastocyst injection. Aggregation chimera produced mouse blastocysts with GFP-nhpESCs at the inner cell mass (ICM), and embryo transfers (ETs) generated dimly-fluorescencing abnormal fetuses. Direct injection of GFP-nhpESCs into blastocysts produced normal non-GFP-fluorescencing fetuses. Injected chimera showed >70% loss of GFP-nhpESCs after 21 h culture. Outgrowths of all chimeric blastocysts established distinct but separate mouse- and NHP-ESC colonies. Extensive endogenous autofluorescence compromised anti-GFP detection and PCR analysis did not detect nhpESCs in fetuses. NhpESCs localize to the ICM in chimera and generate pregnancies. Because primate ESCs do not engraft post-implantation, and also because endogenous autofluorescence results in misleading positive signals, interspecific chimera assays for pluripotency with primate stem cells is unreliable with the currently available ESCs. Testing primate ESCs reprogrammed into even more naïve states in these inter-specific chimera assays will be an important future endeavor. PMID:21543277

Electroporation of Postimplantation Mouse Embryos In Utero.

PubMed

Huang, Cheng-Chiu; Carcagno, Abel

2018-02-01

Gene transfer by electroporation is possible in mouse fetuses within the uterus. As described in this protocol, the pregnant female is anesthetized, the abdominal cavity is opened, and the uterus with the fetuses is exteriorized. A solution of plasmid DNA is injected through the uterine wall directly into the fetus, typically into a cavity like the brain ventricle, guided by fiber optic illumination. Electrodes are positioned on the uterus around the region of the fetus that was injected, and electrical pulses are delivered. The uterus is returned to the abdominal cavity, the body wall is sutured closed, and the female is allowed to recover. The manipulated fetuses can then be collected and analyzed at various times after the electroporation. This method allows experimental access to later-stage developing mouse embryos. © 2018 Cold Spring Harbor Laboratory Press.

Modulation of the malignant phenotype with the urokinase-type plasminogen activator and the type I plasminogen activator inhibitor.

PubMed

Sordat, B; Reiter, L; Cajot, J F

1990-12-02

Gene transfer techniques were utilized to evaluate the role of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor type 1 (PAI-1) in enhancing or preventing the expression of the invasive malignant phenotype, respectively. Mouse L-cell transfectants expressing human uPA or human PAI-1 as well as mouse B16 transfectants expressing mouse uPA or human PAI-1 were generated. These transfectants were tested using a variety of experimental methods including smooth muscle cell matrix solubilization in vitro, lung colony formation in vivo and co-cultures of antagonist-expressing cells in vitro. Results from these studies provide direct evidence for an enhancing role of uPA in malignant invasion and experimental metastasis and for a modulatory role of PAI-1 in tumor cell-mediated breakdown of extracellular matrices.

Mouse neuroblastoma cell based model and the effect of epileptic events on calcium oscillations and neural spikes

NASA Astrophysics Data System (ADS)

Kim, Suhwan; Baek, Juyeong; Jung, Unsang; Lee, Sangwon; Jung, Woonggyu; Kim, Jeehyun; Kang, Shinwon

2013-05-01

Recently, Mouse neuroblastoma cells are considered as an attractive model for the study of human neurological and prion diseases, and intensively used as a model system in different areas. Among those areas, differentiation of neuro2a (N2A) cells, receptor mediated ion current, and glutamate induced physiological response are actively investigated. The reason for the interest to mouse neuroblastoma N2A cells is that they have a fast growing rate than other cells in neural origin with a few another advantages. This study evaluated the calcium oscillations and neural spikes recording of mouse neuroblastoma N2A cells in an epileptic condition. Based on our observation of neural spikes in mouse N2A cell with our proposed imaging modality, we report that mouse neuroblastoma N2A cells can be an important model related to epileptic activity studies. It is concluded that the mouse neuroblastoma N2A cells produce the epileptic spikes in vitro in the same way as produced by the neurons or the astrocytes. This evidence advocates the increased and strong level of neurotransmitters release by enhancement in free calcium using the 4-aminopyridine which causes the mouse neuroblastoma N2A cells to produce the epileptic spikes and calcium oscillation.

Prolongation of the survival of breast cancer-bearing mice immunized with GM-CSF-secreting syngeneic/allogeneic fibroblasts transfected with a cDNA expression library from breast cancer cells.

PubMed

Kim, Tae S; Jung, Mi Y; Cho, Daeho; Cohen, Edward P

2006-10-30

Breast cancer cells, like other types of neoplastic cells, form weakly immunogenic tumor-associated antigens. The antigenic properties of the tumor-associated antigens can be enhanced if they are expressed by highly immunogenic cells. In this study, a cancer vaccine was prepared by transfer of a cDNA expression library from SB5b breast carcinoma into mouse fibroblast cells of C3H/He mouse origin (H-2(k)), that had been previously modified to secrete GM-CSF and to express allogeneic class I-determinants (H-2(b)). The transfected syngeneic/allogeneic fibroblasts secreting GM-CSF were used as a vaccine in C3H/He mice. Robust cell-mediated immunity toward the breast cancer cells was generated in mice immunized with the cDNA-based vaccine. The immunity, mediated predominantly by CD8(+) T lymphocytes, was directed toward the breast cancer cells, but not against either of two other non-cross-reactive neoplasms of C3H/He mice. The immunity was sufficient to prolong the survival of mice with established breast cancer. Among other advantages, preparation of the vaccine by cDNA-transfer into a fibroblast cell line enabled the recipient cells to be modified in advance of DNA-transfer to augment their immunogenic properties. As the transferred DNA is replicated as the transfected cells divide, the vaccine could be prepared from microgram quantities of tumor tissue.

Dynamics of circulating gamma delta T cell activity in an immunocompetent mouse model of high-grade glioma

USDA-ARS?s Scientific Manuscript database

Human gamma delta T cells are potent effectors against glioma cell lines in vitro and in human/mouse xenograft models of glioblastoma, however, this effect has not been investigated in an immunocompetent mouse model. In this report, we established GL261 intracranial gliomas in syngeneic WT C57BL/6 m...

Lack of species-specific difference in pulmonary function when using mouse versus human plasma in a mouse model of hemorrhagic shock.

PubMed

Peng, Zhanglong; Pati, Shibani; Fontaine, Magali J; Hall, Kelly; Herrera, Anthony V; Kozar, Rosemary A

2016-11-01

Clinical studies have demonstrated that the early and empiric use of plasma improves survival after hemorrhagic shock. We have demonstrated in rodent models of hemorrhagic shock that resuscitation with plasma is protective to the lungs compared with lactated Ringer's solution. As our long-term objective is to determine the molecular mechanisms that modulate plasma's protective effects in injured bleeding patients, we have used human plasma in a mouse model of hemorrhagic shock. The goal of the current experiments is to determine if there are significant adverse effects on lung injury when using human versus mouse plasma in an established murine model of hemorrhagic shock and laparotomy. Mice underwent laparotomy and 90 minutes of hemorrhagic shock to a mean arterial pressure (MAP) of 35 ± 5 mm Hg followed by resuscitation at 1× shed blood using either mouse fresh frozen plasma (FFP), human FFP, or human lyophilized plasma. Mean arterial pressure was recorded during shock and for the first 30 minutes of resuscitation. After 3 hours, animals were killed, and lungs collected for analysis. There was a significant increase in early MAP when mouse FFP was used to resuscitate animals compared with human FFP or human lyophilized plasma. However, despite these differences, analysis of the mouse lungs revealed no significant differences in pulmonary histopathology, lung permeability, or lung edema between all three plasma groups. Analysis of neutrophil infiltration in the lungs revealed that mouse FFP decreased neutrophil influx as measured by neutrophil staining; however, myeloperoxidase immunostaining revealed no significant differences in between groups. The study of human plasma in a mouse model of hemorrhagic shock is feasible but does reveal some differences compared with mouse plasma-based resuscitation in physiologic measures such as MAP postresuscitation. Measures of end organ function such as lung injury appear to be comparable in this acute model of hemorrhagic shock and resuscitation.

In vivo quantitative bioluminescence tomography using heterogeneous and homogeneous mouse models.

PubMed

Liu, Junting; Wang, Yabin; Qu, Xiaochao; Li, Xiangsi; Ma, Xiaopeng; Han, Runqiang; Hu, Zhenhua; Chen, Xueli; Sun, Dongdong; Zhang, Rongqing; Chen, Duofang; Chen, Dan; Chen, Xiaoyuan; Liang, Jimin; Cao, Feng; Tian, Jie

2010-06-07

Bioluminescence tomography (BLT) is a new optical molecular imaging modality, which can monitor both physiological and pathological processes by using bioluminescent light-emitting probes in small living animal. Especially, this technology possesses great potential in drug development, early detection, and therapy monitoring in preclinical settings. In the present study, we developed a dual modality BLT prototype system with Micro-computed tomography (MicroCT) registration approach, and improved the quantitative reconstruction algorithm based on adaptive hp finite element method (hp-FEM). Detailed comparisons of source reconstruction between the heterogeneous and homogeneous mouse models were performed. The models include mice with implanted luminescence source and tumor-bearing mice with firefly luciferase report gene. Our data suggest that the reconstruction based on heterogeneous mouse model is more accurate in localization and quantification than the homogeneous mouse model with appropriate optical parameters and that BLT allows super-early tumor detection in vivo based on tomographic reconstruction of heterogeneous mouse model signal.

Of Mice and Men: Comparative Analysis of Neuro-Inflammatory Mechanisms in Human and Mouse Using Cause-and-Effect Models.

PubMed

Kodamullil, Alpha Tom; Iyappan, Anandhi; Karki, Reagon; Madan, Sumit; Younesi, Erfan; Hofmann-Apitius, Martin

2017-01-01

Perturbance in inflammatory pathways have been identified as one of the major factors which leads to neurodegenerative diseases (NDD). Owing to the limited access of human brain tissues and the immense complexity of the brain, animal models, specifically mouse models, play a key role in advancing the NDD field. However, many of these mouse models fail to reproduce the clinical manifestations and end points of the disease. NDD drugs, which passed the efficacy test in mice, were repeatedly not successful in clinical trials. There are numerous studies which are supporting and opposing the applicability of mouse models in neuroinflammation and NDD. In this paper, we assessed to what extend a mouse can mimic the cellular and molecular interactions in humans at a mechanism level. Based on our mechanistic modeling approach, we investigate the failure of a neuroinflammation targeted drug in the late phases of clinical trials based on the comparative analyses between the two species.

NCI Mouse Repository | FNLCR Staging

Cancer.gov

The NCI Mouse Repository is an NCI-funded resource for mouse cancer models and associated strains. The repository makes strains available to all members of the scientific community (academic, non-profit, and commercial). NCI Mouse Repository strains

Cloning Mice.

PubMed

Ogura, Atsuo

2017-08-01

Viable and fertile mice can be generated by somatic nuclear transfer into enucleated oocytes, presumably because the transplanted somatic cell genome becomes reprogrammed by factors in the oocyte. The first somatic cloned offspring of mice were obtained by directly injecting donor nuclei into recipient enucleated oocytes. When this method is used (the so-called Honolulu method of somatic cell nuclear transfer [SCNT]), the donor nuclei readily and completely condense within the enucleated metaphase II-arrested oocytes, which contain high levels of M-phase-promoting factor (MPF). It is believed that the condensation of the donor chromosomes promotes complete reprogramming of the donor genome within the mouse oocytes. Another key to the success of mouse cloning is the use of blunt micropipettes attached to a piezo impact-driving micromanipulation device. This system saves a significant amount of time during the micromanipulation of oocytes and thus minimizes the loss of oocyte viability in vitro. For example, a group of 20 oocytes can be enucleated within 10 min by an experienced operator. This protocol is composed of seven parts: (1) preparing micropipettes, (2) setting up the enucleation and injection micropipettes, (3) collecting and enucleating oocytes, (4) preparing nucleus donor cells, (5) injecting donor nuclei, (6) activating embryos and culturing, and (7) transferring cloned embryos. © 2017 Cold Spring Harbor Laboratory Press.

Effects of Multimodal Analgesia on the Success of Mouse Embryo Transfer Surgery

PubMed Central

Parker, John M.; Austin, Jamie; Wilkerson, James; Carbone, Larry

2011-01-01

Multimodal analgesia is promoted as the best practice pain management for invasive animal research procedures. Universal acceptance and incorporation of multimodal analgesia requires assessing potential effects on study outcome. The focus of this study was to assess effects on embryo survival after multimodal analgesia comprising an opioid and nonsteroidal antiinflammatory drug (NSAID) compared with opioid-only analgesia during embryo transfer procedures in transgenic mouse production. Mice were assigned to receive either carprofen (5 mg/kg) with buprenorphine (0.1 mg/kg; CB) or vehicle with buprenorphine (0.1 mg/kg; VB) in a prospective, double-blinded placebo controlled clinical trial. Data were analyzed in surgical sets of 1 to 3 female mice receiving embryos chimeric for a shared targeted embryonic stem-cell clone and host blastocyst cells. A total of 99 surgical sets were analyzed, comprising 199 Crl:CD1 female mice and their 996 offspring. Neither yield (pups weaned per embryo implanted in the surgical set) nor birth rate (average number of pups weaned per dam in the set) differed significantly between the CB and VB conditions. Multimodal opioid–NSAID analgesia appears to have no significant positive or negative effect on the success of producing novel lines of transgenic mice by blastocyst transfer. PMID:21838973

Vector delivery technique affects gene transfer in the cornea in vivo.

PubMed

Mohan, Rajiv R; Sharma, Ajay; Cebulko, Tyler C; Tandon, Ashish

2010-11-27

This study tested whether controlled drying of the cornea increases vector absorption in mouse and rabbit corneas in vivo and human cornea ex vivo, and studied the effects of corneal drying on gene transfer, structure and inflammatory reaction in the mouse cornea in vivo. Female C57 black mice and New Zealand White rabbits were used for in vivo studies. Donor human corneas were used for ex vivo experiments. A hair dryer was used for drying the corneas after removing corneal epithelium by gentle scraping. The corneas received no, once, twice, thrice, or five times warm air for 10 s with a 5 s interval after each 10 s hair dryer application. Thereafter, balanced salt solution (BSS) was topically applied immediately on the cornea for 2 min using a custom-cloning cylinder. The absorbed BSS was quantified using Hamilton microsyringes. The adeno-associated virus 8 (AAV8) vector (1.1×10(8) genomic copies/µl) expressing marker gene was used to study the effect of corneal drying on gene transfer. Animals were sacrificed on day 14 and gene expression was analyzed using commercial staining kit. Morphological changes and infiltration of inflammatory cells were examined with H & E staining and immunocytochemistry. Mice, rabbit or human corneas subjected to no or 10 s drying showed 6%-8% BSS absorption whereas 20, 30, or 50 s corneal drying showed significantly high 14%-19% (p<0.001), 21%-22% (p<0.001), and 25%-27% (p<0.001) BSS absorption, respectively. The AAV8 application on mouse cornea after 50 s drying showed significantly higher transgene delivery (p<0.05) in vivo with mild-to-moderate changes in corneal morphology. The 30 s of drying also showed significantly (p<0.05) high transgene delivery in mouse stroma in vivo without jeopardizing corneal morphology whereas 10 or 20 s drying showed moderate degree of gene transfer with no altered corneal morphology. Corneas that underwent 50 s drying showed high CD11b-positive cells (p<0.01) compared to control corneas whereas 20 or 30 s air-dried corneas showed insignificant CD11b-positive cells compared to control corneas. Controlled corneal drying with hair dryer increases vector absorption significantly. The dispensing of efficacious AAV serotype into cornea with optimized minimally invasive topical application technique could provide high and targeted expression of therapeutic genes in the stroma in vivo without causing significant side effects.

Importance of murine study design for testing toxicity of retroviral vectors in support of phase I trials.

PubMed

Will, Elke; Bailey, Jeff; Schuesler, Todd; Modlich, Ute; Balcik, Brenden; Burzynski, Ben; Witte, David; Layh-Schmitt, Gerlinde; Rudolph, Cornelia; Schlegelberger, Brigitte; von Kalle, Christof; Baum, Christopher; Sorrentino, Brian P; Wagner, Lars M; Kelly, Patrick; Reeves, Lilith; Williams, David A

2007-04-01

Although retroviral vectors are one of the most widely used vehicles for gene transfer, there is no uniformly accepted pre-clinical model defined to assess their safety, in particular their risk related to insertional mutagenesis. In the murine pre-clinical study presented here, 40 test and 10 control mice were transplanted with ex vivo manipulated bone marrow cells to assess the long-term effects of the transduction of hematopoietic cells with the retroviral vector MSCV-MGMT(P140K)wc. Test mice had significant gene marking 8-12 months post-transplantation with an average of 0.93 vector copies per cell and 41.5% of peripheral blood cells expressing the transgene MGMT(P140K), thus confirming persistent vector expression. Unexpectedly, six test mice developed malignant lymphoma. No vector was detected in the tumor cells of five animals with malignancies, indicating that the malignancies were not caused by insertional mutagenesis or MGMT(P140K) expression. Mice from a concurrent study with a different transgene also revealed additional cases of vector-negative lymphomas of host origin. We conclude that the background tumor formation in this mouse model complicates safety determination of retroviral vectors and propose an improved study design that we predict will increase the relevance and accuracy of interpretation of pre-clinical mouse studies.

Nanoparticle-mediated rhodopsin cDNA but not intron-containing DNA delivery causes transgene silencing in a rhodopsin knockout model.

PubMed

Zheng, Min; Mitra, Rajendra N; Filonov, Nazar A; Han, Zongchao

2016-03-01

Previously, we compared the efficacy of nanoparticle (NP)-mediated intron-containing rhodopsin (sgRho) vs. intronless cDNA in ameliorating retinal disease phenotypes in a rhodopsin knockout (RKO) mouse model of retinitis pigmentosa. We showed that NP-mediated sgRho delivery achieved long-term expression and phenotypic improvement in RKO mice, but not NP housing cDNA. However, the protein level of the NP-sgRho construct was only 5-10% of wild-type at 8 mo postinjection. To have a better understanding of the reduced levels of long-term expression of the vectors, in the present study, we evaluated the epigenetic changes of subretinal delivering NP-cDNA vs. NP-sgRho in the RKO mouse eyes. Following the administration, DNA methylation and histone status of specific regions (bacteria plasmid backbone, promoter, rhodopsin gene, and scaffold/matrix attachment region) of the vectors were evaluated at various time points. We documented that epigenetic transgene silencing occurred in vector-mediated gene transfer, which were caused by the plasmid backbone and the cDNA of the transgene, but not the intron-containing transgene. No toxicity or inflammation was found in the treated eyes. Our results suggest that cDNA of the rhodopsin transgene and bacteria backbone interfered with the host defense mechanism of DNA methylation-mediated transgene silencing through heterochromatin-associated modifications. © FASEB.

Non-invasive near-infrared fluorescence imaging of the neutrophil response in a mouse model of transient cerebral ischaemia.

PubMed

Vaas, Markus; Enzmann, Gaby; Perinat, Therese; Siler, Ulrich; Reichenbach, Janine; Licha, Kai; Kipar, Anja; Rudin, Markus; Engelhardt, Britta; Klohs, Jan

2017-08-01

Near-infrared fluorescence (NIRF) imaging enables non-invasive monitoring of molecular and cellular processes in live animals. Here we demonstrate the suitability of NIRF imaging to investigate the neutrophil response in the brain after transient middle cerebral artery occlusion (tMCAO). We established procedures for ex vivo fluorescent labelling of neutrophils without affecting their activation status. Adoptive transfer of labelled neutrophils in C57BL/6 mice before surgery resulted in higher fluorescence intensities over the ischaemic hemisphere in tMCAO mice with NIRF imaging when compared with controls, corroborated by ex vivo detection of labelled neutrophils using fluorescence microscopy. NIRF imaging showed that neutrophils started to accumulate immediately after tMCAO, peaking at 18 h, and were still visible until 48 h after reperfusion. Our data revealed accumulation of neutrophils also in extracranial tissue, indicating damage in the external carotid artery territory in the tMCAO model. Antibody-mediated inhibition of α4-integrins did reduce fluorescence signals at 18 and 24, but not at 48 h after reperfusion, compared with control treatment animals. Antibody treatment reduced cerebral lesion volumes by 19%. In conclusion, the non-invasive nature of NIRF imaging allows studying the dynamics of neutrophil recruitment and its modulation by targeted interventions in the mouse brain after transient experimental cerebral ischaemia.

Non-invasive near-infrared fluorescence imaging of the neutrophil response in a mouse model of transient cerebral ischaemia

PubMed Central

Vaas, Markus; Enzmann, Gaby; Perinat, Therese; Siler, Ulrich; Reichenbach, Janine; Licha, Kai; Kipar, Anja; Rudin, Markus; Engelhardt, Britta

2016-01-01

Near-infrared fluorescence (NIRF) imaging enables non-invasive monitoring of molecular and cellular processes in live animals. Here we demonstrate the suitability of NIRF imaging to investigate the neutrophil response in the brain after transient middle cerebral artery occlusion (tMCAO). We established procedures for ex vivo fluorescent labelling of neutrophils without affecting their activation status. Adoptive transfer of labelled neutrophils in C57BL/6 mice before surgery resulted in higher fluorescence intensities over the ischaemic hemisphere in tMCAO mice with NIRF imaging when compared with controls, corroborated by ex vivo detection of labelled neutrophils using fluorescence microscopy. NIRF imaging showed that neutrophils started to accumulate immediately after tMCAO, peaking at 18 h, and were still visible until 48 h after reperfusion. Our data revealed accumulation of neutrophils also in extracranial tissue, indicating damage in the external carotid artery territory in the tMCAO model. Antibody-mediated inhibition of α4-integrins did reduce fluorescence signals at 18 and 24, but not at 48 h after reperfusion, compared with control treatment animals. Antibody treatment reduced cerebral lesion volumes by 19%. In conclusion, the non-invasive nature of NIRF imaging allows studying the dynamics of neutrophil recruitment and its modulation by targeted interventions in the mouse brain after transient experimental cerebral ischaemia. PMID:27789786

A Novel Chimeric Antigen Receptor Against Prostate Stem Cell Antigen Mediates Tumor Destruction in a Humanized Mouse Model of Pancreatic Cancer

PubMed Central

Lagisetty, Kiran H.; Tran, Eric; Zheng, Zhili; Gattinoni, Luca; Yu, Zhiya; Burns, William R.; Miermont, Anne M.; Teper, Yaroslav; Rudloff, Udo; Restifo, Nicholas P.; Feldman, Steven A.; Rosenberg, Steven A.; Morgan, Richard A.

2014-01-01

Abstract Despite advances in the understanding of its molecular pathophysiology, pancreatic cancer remains largely incurable, highlighting the need for novel therapies. We developed a chimeric antigen receptor (CAR) specific for prostate stem cell antigen (PSCA), a glycoprotein that is overexpressed in pancreatic cancer starting at early stages of malignant transformation. To optimize the CAR design, we used antigen-recognition domains derived from mouse or human antibodies, and intracellular signaling domains containing one or two T cell costimulatory elements, in addition to CD3zeta. Comparing multiple constructs established that the CAR based on human monoclonal antibody Ha1-4.117 had the greatest reactivity in vitro. To further analyze this CAR, we developed a human pancreatic cancer xenograft model and adoptively transferred CAR-engineered T cells into animals with established tumors. CAR-engineered human lymphocytes induced significant antitumor activity, and unlike what has been described for other CARs, a second-generation CAR (containing CD28 cosignaling domain) induced a more potent antitumor effect than a third-generation CAR (containing CD28 and 41BB cosignaling domains). While our results provide evidence to support PSCA as a target antigen for CAR-based immunotherapy of pancreatic cancer, the expression of PSCA on selected normal tissues could be a source of limiting toxicity. PMID:24694017

Production of cloned mice from somatic cells, ES cells, and frozen bodies.

PubMed

Wakayama, Sayaka; Mizutani, Eiji; Wakayama, Teruhiko

2010-01-01

Somatic cell nuclear transfer (SCNT) has become a unique and powerful tool for epigenetic reprogramming research and gene manipulation in animals. Although the success rates of somatic cloning have been inefficient and the mechanism of reprogramming is still largely unknown, therefore, the nuclear transfer (NT) method has been thought of as a "black box approach" and inadequate to determine the detail of how genomic reprogramming occurs. However, only the NT approach can reveal dynamic and global modifications in the epigenome without using genetic modification, as well as can create live animals. At present, this is the only technique available for the preservation and propagation of valuable genetic resources from mutant mice that are infertile or too old, or recovered from carcasses, without the use of germ cells. This chapter describes a basic protocol for mouse cloning and embryonic stem (ES) cell establishment from cloned embryo using a piezo-actuated micromanipulator. This technique will greatly help not only in mouse cloning but also in other forms of micromanipulation such as intracytoplasmic sperm injection (ICSI) into oocytes or ES cell injection into blastocysts. In addition, we describe a new, more efficient mouse cloning protocol using histone deacetylase inhibitor (HDACi), which increases the success rates of cloned mice or establish rate of ES cells to fivefold. Copyright (c) 2010 Elsevier Inc. All rights reserved.

Characterisation of mouse epididymosomes reveals a complex profile of microRNAs and a potential mechanism for modification of the sperm epigenome

PubMed Central

Reilly, Jackson N.; McLaughlin, Eileen A.; Stanger, Simone J.; Anderson, Amanda L.; Hutcheon, Kate; Church, Kiralee; Mihalas, Bettina P.; Tyagi, Sonika; Holt, Janet E.; Eamens, Andrew L.; Nixon, Brett

2016-01-01

Recent evidence has shown that the sperm epigenome is vulnerable to dynamic modifications arising from a variety of paternal environment exposures and that this legacy can serve as an important determinant of intergenerational inheritance. It has been postulated that such exchange is communicated to maturing spermatozoa via the transfer of small non-protein-coding RNAs (sRNAs) in a mechanism mediated by epididymosomes; small membrane bound vesicles released by the soma of the male reproductive tract (epididymis). Here we confirm that mouse epididymosomes encapsulate an impressive cargo of >350 microRNAs (miRNAs), a developmentally important sRNA class, the majority (~60%) of which are also represented by the miRNA signature of spermatozoa. This includes >50 miRNAs that were found exclusively in epididymal sperm and epididymosomes, but not in the surrounding soma. We also documented substantial changes in the epididymosome miRNA cargo, including significant fold changes in almost half of the miRNAs along the length of the epididymis. Finally, we provide the first direct evidence for the transfer of several prominent miRNA species between mouse epididymosomes and spermatozoa to afford novel insight into a mechanism of intercellular communication by which the sRNA payload of sperm can be selectively modified during their post-testicular maturation. PMID:27549865

An extended Kalman filter for mouse tracking.

PubMed

Choi, Hongjun; Kim, Mingi; Lee, Onseok

2018-05-19

Animal tracking is an important tool for observing behavior, which is useful in various research areas. Animal specimens can be tracked using dynamic models and observation models that require several types of data. Tracking mouse has several barriers due to the physical characteristics of the mouse, their unpredictable movement, and cluttered environments. Therefore, we propose a reliable method that uses a detection stage and a tracking stage to successfully track mouse. The detection stage detects the surface area of the mouse skin, and the tracking stage implements an extended Kalman filter to estimate the state variables of a nonlinear model. The changes in the overall shape of the mouse are tracked using an oval-shaped tracking model to estimate the parameters for the ellipse. An experiment is conducted to demonstrate the performance of the proposed tracking algorithm using six video images showing various types of movement, and the ground truth values for synthetic images are compared to the values generated by the tracking algorithm. A conventional manual tracking method is also applied to compare across eight experimenters. Furthermore, the effectiveness of the proposed tracking method is also demonstrated by applying the tracking algorithm with actual images of mouse. Graphical abstract.

Mouse Models for Down Syndrome-Associated Developmental Cognitive Disabilities

PubMed Central

Liu, Chunhong; Belichenko, Pavel V.; Zhang, Li; Fu, Dawei; Kleschevnikov, Alexander M.; Baldini, Antonio; Antonarakis, Stylianos E.; Mobley, William C.; Yu, Y. Eugene

2011-01-01

Down syndrome (DS) is mainly caused by the presence of an extra copy of human chromosome 21 (Hsa21) and is a leading genetic cause for developmental cognitive disabilities in humans. The mouse is a premier model organism for DS because the regions on Hsa21 are syntenically conserved with three regions in the mouse genome, which are located on mouse chromosome 10 (Mmu10), Mmu16 and Mmu17. With the advance of chromosomal manipulation technologies, new mouse mutants have been generated to mimic DS at both the genotypic and phenotypic levels. Further mouse-based molecular genetic studies in the future may lead to the unraveling of the mechanisms underlying DS-associated developmental cognitive disabilities, which would lay the groundwork for developing effective treatments for this phenotypic manifestation. In this review, we will discuss recent progress and future challenges in modeling DS-associated developmental cognitive disability in mice with an emphasis on hippocampus-related phenotypes. PMID:21865664

Adoptively transferred TRAIL+ T cells suppress GVHD and augment antitumor activity

PubMed Central

Ghosh, Arnab; Dogan, Yildirim; Moroz, Maxim; Holland, Amanda M.; Yim, Nury L.; Rao, Uttam K.; Young, Lauren F.; Tannenbaum, Daniel; Masih, Durva; Velardi, Enrico; Tsai, Jennifer J.; Jenq, Robert R.; Penack, Olaf; Hanash, Alan M.; Smith, Odette M.; Piersanti, Kelly; Lezcano, Cecilia; Murphy, George F.; Liu, Chen; Palomba, M. Lia; Sauer, Martin G.; Sadelain, Michel; Ponomarev, Vladimir; van den Brink, Marcel R.M.

2013-01-01

Current strategies to suppress graft-versus-host disease (GVHD) also compromise graft-versus-tumor (GVT) responses. Furthermore, most experimental strategies to separate GVHD and GVT responses merely spare GVT function without actually enhancing it. We have previously shown that endogenously expressed TNF-related apoptosis-inducing ligand (TRAIL) is required for optimal GVT activity against certain malignancies in recipients of allogeneic hematopoietic stem cell transplantation (allo-HSCT). In order to model a donor-derived cellular therapy, we genetically engineered T cells to overexpress TRAIL and adoptively transferred donor-type unsorted TRAIL+ T cells into mouse models of allo-HSCT. We found that murine TRAIL+ T cells induced apoptosis of alloreactive T cells, thereby reducing GVHD in a DR5-dependent manner. Furthermore, murine TRAIL+ T cells mediated enhanced in vitro and in vivo antilymphoma GVT response. Moreover, human TRAIL+ T cells mediated enhanced in vitro cytotoxicity against both human leukemia cell lines and against freshly isolated chronic lymphocytic leukemia (CLL) cells. Finally, as a model of off-the-shelf, donor-unrestricted antitumor cellular therapy, in vitro–generated TRAIL+ precursor T cells from third-party donors also mediated enhanced GVT response in the absence of GVHD. These data indicate that TRAIL-overexpressing donor T cells could potentially enhance the curative potential of allo-HSCT by increasing GVT response and suppressing GVHD. PMID:23676461

Methods in Molecular Biology Mouse Genetics: Methods and Protocols | Center for Cancer Research

Cancer.gov

Mouse Genetics: Methods and Protocols provides selected mouse genetic techniques and their application in modeling varieties of human diseases. The chapters are mainly focused on the generation of different transgenic mice to accomplish the manipulation of genes of interest, tracing cell lineages, and modeling human diseases.

Use of mouse models to study the mechanisms and consequences of RBC clearance

PubMed Central

Hod, E. A.; Arinsburg, S. A.; Francis, R. O.; Hendrickson, J. E.; Zimring, J. C.; Spitalnik, S. L.

2013-01-01

Mice provide tractable animal models for studying the pathophysiology of various human disorders. This review discusses the use of mouse models for understanding red-blood-cell (RBC) clearance. These models provide important insights into the pathophysiology of various clinically relevant entities, such as autoimmune haemolytic anaemia, haemolytic transfusion reactions, other complications of RBC transfusions and immunomodulation by Rh immune globulin therapy. Mouse models of both antibody- and non-antibody-mediated RBC clearance are reviewed. Approaches for exploring unanswered questions in transfusion medicine using these models are also discussed. PMID:20345515

Generation Of A Mouse Model For Schwannomatosis

DTIC Science & Technology

2010-09-01

TITLE: Generation of a Mouse Model for Schwannomatosis PRINCIPAL INVESTIGATOR: Long-Sheng Chang, Ph.D. CONTRACTING ORGANIZATION: The...Annual 3. DATES COVERED (From - To) 1 Sep 2009 - 31 Aug 2010 4. TITLE AND SUBTITLE Generation of a Mouse Model for Schwannomatosis 5a. CONTRACT...hypothesis involving inactivation of both the INI1/SNF5 and NF2 tumor suppressor genes in the formation of schwannomatosis -associated tumors. To

Butyrylcholinesterase gene transfer in obese mice prevents postdieting body weight rebound by suppressing ghrelin signaling.

PubMed

Chen, Vicky Ping; Gao, Yang; Geng, Liyi; Brimijoin, Stephen

2017-10-10

The worldwide prevalence of obesity is increasing at an alarming rate but treatment options remain limited. Despite initial success, weight loss by calorie restriction (CR) often fails because of rebound weight gain. Postdieting hyperphagia along with altered hypothalamic neuro-architecture appears to be one direct cause of this undesirable outcome. In response to calorie deficiency the circulating levels of the appetite-promoting hormone, acyl-ghrelin, rise sharply. We hypothesize that proper modulation of acyl-ghrelin and its receptor's sensitivity will favorably impact energy intake and reprogram the body weight set point. Here we applied viral gene transfer of the acyl-ghrelin hydrolyzing enzyme, butyrylcholinesterase (BChE), in a mouse model of diet-induced obesity. Our results confirmed that BChE overexpression decreased circulating acyl-ghrelin levels, suppressed CR-provoked ghrelin signaling, and restored central ghrelin sensitivity. In addition to maintaining healthy body weights, BChE treated mice had modest postdieting food intake and showed normal glucose homeostasis. Spontaneous activity and energy expenditure did not differ significantly between treated and untreated mice after body weight rebound, suggesting that BChE gene transfer did not alter energy expenditure in the long term. These findings indicate that combining BChE treatment with CR could be an effective approach in treating human obesity and aiding lifelong weight management.

Mouse Genome Database: From sequence to phenotypes and disease models

PubMed Central

Richardson, Joel E.; Kadin, James A.; Smith, Cynthia L.; Blake, Judith A.; Bult, Carol J.

2015-01-01

Summary The Mouse Genome Database (MGD, www.informatics.jax.org) is the international scientific database for genetic, genomic, and biological data on the laboratory mouse to support the research requirements of the biomedical community. To accomplish this goal, MGD provides broad data coverage, serves as the authoritative standard for mouse nomenclature for genes, mutants, and strains, and curates and integrates many types of data from literature and electronic sources. Among the key data sets MGD supports are: the complete catalog of mouse genes and genome features, comparative homology data for mouse and vertebrate genes, the authoritative set of Gene Ontology (GO) annotations for mouse gene functions, a comprehensive catalog of mouse mutations and their phenotypes, and a curated compendium of mouse models of human diseases. Here, we describe the data acquisition process, specifics about MGD's key data areas, methods to access and query MGD data, and outreach and user help facilities. genesis 53:458–473, 2015. © 2015 The Authors. Genesis Published by Wiley Periodicals, Inc. PMID:26150326

Hydroxyurea therapy requires HbF induction for clinical benefit in a sickle cell mouse model

PubMed Central

Lebensburger, Jeffrey D.; Pestina, Tamara I.; Ware, Russell E.; Boyd, Kelli L.; Persons, Derek A.

2010-01-01

Hydroxyurea has proven clinical efficacy in patients with sickle cell disease. Potential mechanisms for the beneficial effects include fetal hemoglobin induction and the reduction of cell adhesive properties, inflammation and hypercoagulability. Using a murine model of sickle cell disease in which fetal hemoglobin induction does not occur, we evaluated whether hydroxyurea administration would still yield improvements in hematologic parameters and reduce end-organ damage. Animals given a maximally tolerated dose of hydroxyurea that resulted in significant reductions in the neutrophil and platelet counts showed no improvement in hemolytic anemia and end-organ damage compared to control mice. In contrast, animals having high levels of fetal hemoglobin due to gene transfer with a γ-globin lentiviral vector showed correction of anemia and organ damage. These data suggest that induction of fetal hemoglobin by hydroxyurea is an essential mechanism for its clinical benefits. PMID:20378564

Effective Gene Therapy of Mice with Congenital Erythropoietic Porphyria Is Facilitated by a Survival Advantage of Corrected Erythroid Cells

PubMed Central

Robert-Richard, Elodie; Moreau-Gaudry, François; Lalanne, Magalie; Lamrissi-Garcia, Isabelle; Cario-André, Muriel; Guyonnet-Dupérat, Véronique; Taine, Laurence; Ged, Cécile; de Verneuil, Hubert

2008-01-01

Achieving long-term expression of a therapeutic gene in a given hematopoietic lineage remains an important goal of gene therapy. Congenital erythropoietic porphyria (CEP) is a severe autosomal-recessive disorder characterized by a deficiency in uroporphyrinogen III synthase (UROS), the fourth enzyme of the heme biosynthetic pathway. We used a recently obtained murine model to check the feasibility of gene therapy in this disease. Lentivirus-mediated transfer of the human UROS cDNA into hematopoietic stem cells (HSCs) from Urosmut248 mice resulted in a complete and long-term enzymatic, metabolic, and phenotypic correction of the disease, favored by a survival advantage of corrected red blood cells. These results demonstrate that the cure of this mouse model of CEP at a moderate transduction level supports the proof of concept of a gene therapy in this disease by transplantation of genetically modified hematopoietic stem cells. PMID:18179890

Overcoming preexisting humoral immunity to AAV using capsid decoys.

PubMed

Mingozzi, Federico; Anguela, Xavier M; Pavani, Giulia; Chen, Yifeng; Davidson, Robert J; Hui, Daniel J; Yazicioglu, Mustafa; Elkouby, Liron; Hinderer, Christian J; Faella, Armida; Howard, Carolann; Tai, Alex; Podsakoff, Gregory M; Zhou, Shangzhen; Basner-Tschakarjan, Etiena; Wright, John Fraser; High, Katherine A

2013-07-17

Adeno-associated virus (AAV) vectors delivered through the systemic circulation successfully transduce various target tissues in animal models. However, similar attempts in humans have been hampered by the high prevalence of neutralizing antibodies to AAV, which completely block vector transduction. We show in both mouse and nonhuman primate models that addition of empty capsid to the final vector formulation can, in a dose-dependent manner, adsorb these antibodies, even at high titers, thus overcoming their inhibitory effect. To further enhance the safety of the approach, we mutated the receptor binding site of AAV2 to generate an empty capsid mutant that can adsorb antibodies but cannot enter a target cell. Our work suggests that optimizing the ratio of full/empty capsids in the final formulation of vector, based on a patient's anti-AAV titers, will maximize the efficacy of gene transfer after systemic vector delivery.

Local Inflammation Exacerbates the Severity of Staphylococcus aureus Skin Infection

PubMed Central

Montgomery, Christopher P.; Daniels, Melvin D.; Zhao, Fan; Spellberg, Brad; Chong, Anita S.; Daum, Robert S.

2013-01-01

Staphylococcus aureus is the leading cause of skin infections. In a mouse model of S. aureus skin infection, we found that lesion size did not correlate with bacterial burden. Athymic nude mice had smaller skin lesions that contained lower levels of myeloperoxidase, IL-17A, and CXCL1, compared with wild type mice, although there was no difference in bacterial burden. T cell deficiency did not explain the difference in lesion size, because TCR βδ (-/-) mice did not have smaller lesions, and adoptive transfer of congenic T cells into athymic nude mice prior to infection did not alter lesion size. The differences observed were specific to the skin, because mortality in a pneumonia model was not different between wild type and athymic nude mice. Thus, the clinical severity of S. aureus skin infection is driven by the inflammatory response to the bacteria, rather than bacterial burden, in a T cell independent manner. PMID:23861974

Overcoming Preexisting Humoral Immunity to AAV Using Capsid Decoys

PubMed Central

Anguela, Xavier M.; Pavani, Giulia; Chen, Yifeng; Davidson, Robert J.; Hui, Daniel J.; Yazicioglu, Mustafa; Elkouby, Liron; Hinderer, Christian J.; Faella, Armida; Howard, Carolann; Tai, Alex; Podsakoff, Gregory M.; Zhou, Shangzhen; Basner-Tschakarjan, Etiena; Wright, John Fraser

2014-01-01

Adeno-associated virus (AAV) vectors delivered through the systemic circulation successfully transduce various target tissues in animal models. However, similar attempts in humans have been hampered by the high prevalence of neutralizing antibodies to AAV, which completely block vector transduction. We show in both mouse and nonhuman primate models that addition of empty capsid to the final vector formulation can, in a dose-dependent manner, adsorb these antibodies, even at high titers, thus overcoming their inhibitory effect. To further enhance the safety of the approach, we mutated the receptor binding site of AAV2 to generate an empty capsid mutant that can adsorb antibodies but cannot enter a target cell. Our work suggests that optimizing the ratio of full/empty capsids in the final formulation of vector, based on a patient's anti-AAV titers, will maximize the efficacy of gene transfer after systemic vector delivery. PMID:23863832

Establishing a laboratory animal model from a transgenic animal: RasH2 mice as a model for carcinogenicity studies in regulatory science.

PubMed

Urano, K; Tamaoki, N; Nomura, T

2012-01-01

Transgenic animal models have been used in small numbers in gene function studies in vivo for a period of time, but more recently, the use of a single transgenic animal model has been approved as a second species, 6-month alternative (to the routine 2-year, 2-animal model) used in short-term carcinogenicity studies for generating regulatory application data of new drugs. This article addresses many of the issues associated with the creation and use of one of these transgenic models, the rasH2 mouse, for regulatory science. The discussion includes strategies for mass producing mice with the same stable phenotype, including constructing the transgene, choosing a founder mouse, and controlling both the transgene and background genes; strategies for developing the model for regulatory science, including measurements of carcinogen susceptibility, stability of a large-scale production system, and monitoring for uniform carcinogenicity responses; and finally, efficient use of the transgenic animal model on study. Approximately 20% of mouse carcinogenicity studies for new drug applications in the United States currently use transgenic models, typically the rasH2 mouse. The rasH2 mouse could contribute to animal welfare by reducing the numbers of animals used as well as reducing the cost of carcinogenicity studies. A better understanding of the advantages and disadvantages of the transgenic rasH2 mouse will result in greater and more efficient use of this animal model in the future.

NCI Mouse Repository | Frederick National Laboratory for Cancer Research

Cancer.gov

The NCI Mouse Repository is an NCI-funded resource for mouse cancer models and associated strains. The repository makes strains available to all members of the scientific community (academic, non-profit, and commercial). NCI Mouse Repository strains

Cage Change Influences Serum Corticosterone and Anxiety-Like Behaviors in the Mouse

PubMed Central

Rasmussen, Skye; Miller, Melinda M.; Filipski, Sarah B.; Tolwani, Ravi J.

2011-01-01

Environmental variables and husbandry practices can influence physiology and alter behavior in mice. Our study evaluated the effects of cage change on serum corticosterone levels and anxiety-like behaviors in C57BL/6 male mice. We examined the effects of 3 different methods of performing cage transfer and of transferring mice to a clean or a dirty familiar cage microenvironment. The 3 different handling methods were forceps transfer, gentle transfer with gloved hands, and a passive transfer technique that did not involve active handling. Active handling methods and transfer to both clean and dirty cage microenvironments significantly increased serum corticosterone 15 min after cage change; however, at 60 min after cage change, levels were comparable to those of unmanipulated mice. Although the effects were transient, cage change altered anxiety-like behaviors in the open field when behavioral testing was performed on the same day. These results demonstrate that the timing of cage change can influence behavioral results, an effect that is an important consideration for rodent behavioral studies. PMID:21838975

Immunosuppressive Effect of B7-H4 Pathway in a Murine Systemic Lupus Erythematosus Model.

PubMed

Xiao, Ze Xiu; Zheng, Xu; Hu, Li; Wang, Julie; Olsen, Nancy; Zheng, Song Guo

2017-01-01

B7-H4, one of the co-stimulatory molecules of the B7 family, has been shown to play an important role in negatively regulating the adaptive immune response by inhibiting the proliferation, activation, and cytokine production of T cells. In this study, we investigate the role of B7-H4 in development of systemic lupus erythematosus (SLE). We investigated a murine model of SLE using transfer of bone marrow-derived dendritic cells (BMDCs) that were incubated with activated syngeneic lymphocyte-derived DNA. The recipient mouse produced anti-ds-DNA antibodies as well as displayed splenomegaly and lymphadenopathy as shown by significantly increased weights, and the kidneys showed lupus-like pathological changes include urine protein and glomerulonephritis with hyperplasia in glomeruli and increased mesangial cells and vasculitis with perivascular cell infiltration, glomerular deposition of IgG and complement C3. We showed that B7-H4 deficiency in BMDCs could cause greater production of anti-ds-DNA antibodies in transferred mice, and the lymph tissue swelling and the kidney lesions were also exacerbated with B7-H4 deficiency. Treatment with a B7-H4 antagonist antibody also aggravated the lupus model. Conversely, B7-H4 Ig alleviated the lupus manifestations. Therefore, we conclude that B7-H4 is a negative check point for the development of SLE in this murine model. These results suggest that this approach may have a clinical potential in treating human SLE.

Immunosuppressive Effect of B7-H4 Pathway in a Murine Systemic Lupus Erythematosus Model

PubMed Central

Xiao, Ze Xiu; Zheng, Xu; Hu, Li; Wang, Julie; Olsen, Nancy; Zheng, Song Guo

2017-01-01

B7-H4, one of the co-stimulatory molecules of the B7 family, has been shown to play an important role in negatively regulating the adaptive immune response by inhibiting the proliferation, activation, and cytokine production of T cells. In this study, we investigate the role of B7-H4 in development of systemic lupus erythematosus (SLE). We investigated a murine model of SLE using transfer of bone marrow-derived dendritic cells (BMDCs) that were incubated with activated syngeneic lymphocyte-derived DNA. The recipient mouse produced anti-ds-DNA antibodies as well as displayed splenomegaly and lymphadenopathy as shown by significantly increased weights, and the kidneys showed lupus-like pathological changes include urine protein and glomerulonephritis with hyperplasia in glomeruli and increased mesangial cells and vasculitis with perivascular cell infiltration, glomerular deposition of IgG and complement C3. We showed that B7-H4 deficiency in BMDCs could cause greater production of anti-ds-DNA antibodies in transferred mice, and the lymph tissue swelling and the kidney lesions were also exacerbated with B7-H4 deficiency. Treatment with a B7-H4 antagonist antibody also aggravated the lupus model. Conversely, B7-H4 Ig alleviated the lupus manifestations. Therefore, we conclude that B7-H4 is a negative check point for the development of SLE in this murine model. These results suggest that this approach may have a clinical potential in treating human SLE. PMID:29321778

Screening and identification of proteins mediating senna induced gastrointestinal motility enhancement in mouse colon

PubMed Central

Wang, Xin; Zhong, Yue-Xia; Lan, Mei; Zhang, Zong-You; Shi, Yong-Quan; Lu, Ju; Ding, Jie; Wu, Kai-Cun; Jin, Jian-Ping; Pan, Bo-Rong; Fan, Dai Min

2002-01-01

AIM: To isolate the proteins involved in pharmacologic action of senna extract (SE) from mouse gastrointestinal tract and to explore the molecular mechanism of gastrointestinal motility change induced by SE. METHODS: SE was administrated to mice by different routes. Gastrointestinal motility of mice was observed using cathartic, gastrointestinal propellant movement experiments and X-ray analysis. Mouse model for gastrointestinal motility enhancement was established through continuous gastric administration of SE at progressively increased dose. At 3 h and week 3, 4, 6 and 10, morphological changes of gastrointestinal tissues were found under light microscope. Ultrastructural changes of intestinal and colonic tissues at week 6 were observed under transmission electron microscope. The colonic proteomic changes in model mice were examined by two-dimension polyacrylamide gel electrophoresis with immobilized pH gradient isoelectric focusing to screen the differentially expressed proteins, and their molecular masses and isoelectric points were determined. Two N-terminal sequences of the samples were also determined by mass spectrometry. RESULTS: SE (0.3 g) caused diarrhea after gastric administration in 1-6 h and enhanced gastrointestinal propellant (65.1% ± 7.5%; 45.8% ± 14.6%,P < 0.01) in mice, but intramuscular and hypodermic injection had no cathartic effect. X-ray analysis of gastrointestinal motility demonstrated that gastric administration of SE enhanced gastric evacuation and gastrointestinal transferring function. At 3 h and week 3 and 4 after gastric administration of SE, light microscopic examination revealed no apparent change in gastrointestinal mucosal tissues, but transmission electron microscopic examination revealed inflammatory changes in whole layer of intestinal and colonic wall. Twenty differential proteins were detected in the colonic tissues of the model mice by two-dimensional electrophoresis, and the N-terminal amino acid sequences of two proteins were determined. CONCLUSION: SE causes diarrhea and enhances gastrointestinal motility through digestive tract administration. Long-term gastric administration of SE induces inflammatory changes and cell damage in the whole gastrointestinal tract. The differential proteins screened from the colonic tissues of the model mice might mediate the enhancing effect of SE on gastrointestinal motility. PMID:11833095

An efficient and robust MRI-guided radiotherapy planning approach for targeting abdominal organs and tumours in the mouse

PubMed Central

Bird, Luke; Tullis, Iain D. C.; Newman, Robert G.; Corroyer-Dulmont, Aurelien; Falzone, Nadia; Azad, Abul; Vallis, Katherine A.; Sansom, Owen J.; Muschel, Ruth J.; Vojnovic, Borivoj; Hill, Mark A.; Fokas, Emmanouil; Smart, Sean C.

2017-01-01

Introduction Preclinical CT-guided radiotherapy platforms are increasingly used but the CT images are characterized by poor soft tissue contrast. The aim of this study was to develop a robust and accurate method of MRI-guided radiotherapy (MR-IGRT) delivery to abdominal targets in the mouse. Methods A multimodality cradle was developed for providing subject immobilisation and its performance was evaluated. Whilst CT was still used for dose calculations, target identification was based on MRI. Each step of the radiotherapy planning procedure was validated initially in vitro using BANG gel dosimeters. Subsequently, MR-IGRT of normal adrenal glands with a size-matched collimated beam was performed. Additionally, the SK-N-SH neuroblastoma xenograft model and the transgenic KPC model of pancreatic ductal adenocarcinoma were used to demonstrate the applicability of our methods for the accurate delivery of radiation to CT-invisible abdominal tumours. Results The BANG gel phantoms demonstrated a targeting efficiency error of 0.56 ± 0.18 mm. The in vivo stability tests of body motion during MR-IGRT and the associated cradle transfer showed that the residual body movements are within this MR-IGRT targeting error. Accurate MR-IGRT of the normal adrenal glands with a size-matched collimated beam was confirmed by γH2AX staining. Regression in tumour volume was observed almost immediately post MR-IGRT in the neuroblastoma model, further demonstrating accuracy of x-ray delivery. Finally, MR-IGRT in the KPC model facilitated precise contouring and comparison of different treatment plans and radiotherapy dose distributions not only to the intra-abdominal tumour but also to the organs at risk. Conclusion This is, to our knowledge, the first study to demonstrate preclinical MR-IGRT in intra-abdominal organs. The proposed MR-IGRT method presents a state-of-the-art solution to enabling robust, accurate and efficient targeting of extracranial organs in the mouse and can operate with a sufficiently high throughput to allow fractionated treatments to be given. PMID:28453537

Temperature mapping and thermal dose calculation in combined radiation therapy and 13.56 MHz radiofrequency hyperthermia for tumor treatment

NASA Astrophysics Data System (ADS)

Kim, Jung Kyung; Prasad, Bibin; Kim, Suzy

2017-02-01

To evaluate the synergistic effect of radiotherapy and radiofrequency hyperthermia therapy in the treatment of lung and liver cancers, we studied the mechanism of heat absorption and transfer in the tumor using electro-thermal simulation and high-resolution temperature mapping techniques. A realistic tumor-induced mouse anatomy, which was reconstructed and segmented from computed tomography images, was used to determine the thermal distribution in tumors during radiofrequency (RF) heating at 13.56 MHz. An RF electrode was used as a heat source, and computations were performed with the aid of the multiphysics simulation platform Sim4Life. Experiments were carried out on a tumor-mimicking agar phantom and a mouse tumor model to obtain a spatiotemporal temperature map and thermal dose distribution. A high temperature increase was achieved in the tumor from both the computation and measurement, which elucidated that there was selective high-energy absorption in tumor tissue compared to the normal surrounding tissues. The study allows for effective treatment planning for combined radiation and hyperthermia therapy based on the high-resolution temperature mapping and high-precision thermal dose calculation.

C5a Receptor (CD88) Blockade Protects against MPO-ANCA GN

PubMed Central

Xiao, Hong; Dairaghi, Daniel J.; Powers, Jay P.; Ertl, Linda S.; Baumgart, Trageen; Wang, Yu; Seitz, Lisa C.; Penfold, Mark E.T.; Gan, Lin; Hu, Peiqi; Lu, Bao; Gerard, Norma P.; Gerard, Craig; Schall, Thomas J.; Jaen, Juan C.; Falk, Ronald J.

2014-01-01

Necrotizing and crescentic GN (NCGN) with a paucity of glomerular immunoglobulin deposits is associated with ANCA. The most common ANCA target antigens are myeloperoxidase (MPO) and proteinase 3. In a manner that requires activation of the alternative complement pathway, passive transfer of antibodies to mouse MPO (anti-MPO) induces a mouse model of ANCA NCGN that closely mimics human disease. Here, we confirm the importance of C5aR/CD88 in the mediation of anti-MPO–induced NCGN and report that C6 is not required. We further demonstrate that deficiency of C5a-like receptor (C5L2) has the reverse effect of C5aR/CD88 deficiency and results in more severe disease, indicating that C5aR/CD88 engagement enhances inflammation and C5L2 engagement suppresses inflammation. Oral administration of CCX168, a small molecule antagonist of human C5aR/CD88, ameliorated anti-MPO–induced NCGN in mice expressing human C5aR/CD88. These observations suggest that blockade of C5aR/CD88 might have therapeutic benefit in patients with ANCA-associated vasculitis and GN. PMID:24179165

C5a receptor (CD88) blockade protects against MPO-ANCA GN.

PubMed

Xiao, Hong; Dairaghi, Daniel J; Powers, Jay P; Ertl, Linda S; Baumgart, Trageen; Wang, Yu; Seitz, Lisa C; Penfold, Mark E T; Gan, Lin; Hu, Peiqi; Lu, Bao; Gerard, Norma P; Gerard, Craig; Schall, Thomas J; Jaen, Juan C; Falk, Ronald J; Jennette, J Charles

2014-02-01

Necrotizing and crescentic GN (NCGN) with a paucity of glomerular immunoglobulin deposits is associated with ANCA. The most common ANCA target antigens are myeloperoxidase (MPO) and proteinase 3. In a manner that requires activation of the alternative complement pathway, passive transfer of antibodies to mouse MPO (anti-MPO) induces a mouse model of ANCA NCGN that closely mimics human disease. Here, we confirm the importance of C5aR/CD88 in the mediation of anti-MPO-induced NCGN and report that C6 is not required. We further demonstrate that deficiency of C5a-like receptor (C5L2) has the reverse effect of C5aR/CD88 deficiency and results in more severe disease, indicating that C5aR/CD88 engagement enhances inflammation and C5L2 engagement suppresses inflammation. Oral administration of CCX168, a small molecule antagonist of human C5aR/CD88, ameliorated anti-MPO-induced NCGN in mice expressing human C5aR/CD88. These observations suggest that blockade of C5aR/CD88 might have therapeutic benefit in patients with ANCA-associated vasculitis and GN.

GaN-based micro-LED arrays on flexible substrates for optical cochlear implants

NASA Astrophysics Data System (ADS)

Goßler, Christian; Bierbrauer, Colin; Moser, Rüdiger; Kunzer, Michael; Holc, Katarzyna; Pletschen, Wilfried; Köhler, Klaus; Wagner, Joachim; Schwaerzle, Michael; Ruther, Patrick; Paul, Oliver; Neef, Jakob; Keppeler, Daniel; Hoch, Gerhard; Moser, Tobias; Schwarz, Ulrich T.

2014-05-01

Currently available cochlear implants are based on electrical stimulation of the spiral ganglion neurons. Optical stimulation with arrays of micro-sized light-emitting diodes (µLEDs) promises to increase the number of distinguishable frequencies. Here, the development of a flexible GaN-based micro-LED array as an optical cochlear implant is reported for application in a mouse model. The fabrication of 15 µm thin and highly flexible devices is enabled by a laser-based layer transfer process of the GaN-LEDs from sapphire to a polyimide-on-silicon carrier wafer. The fabricated 50 × 50 µm2 LEDs are contacted via conducting paths on both p- and n-sides of the LEDs. Up to three separate channels could be addressed. The probes, composed of a linear array of the said µLEDs bonded to the flexible polyimide substrate, are peeled off the carrier wafer and attached to flexible printed circuit boards. Probes with four µLEDs and a width of 230 µm are successfully implanted in the mouse cochlea both in vitro and in vivo. The LEDs emit 60 µW at 1 mA after peel-off, corresponding to a radiant emittance of 6 mW mm-2.

Luciferase-tagged wild-type and tropism-deficient mouse cytomegaloviruses reveal early dynamics of host colonization following peripheral challenge.

PubMed

Farrell, Helen; Oliveira, Martha; Macdonald, Kate; Yunis, Joseph; Mach, Michael; Bruce, Kimberley; Stevenson, Philip; Cardin, Rhonda; Davis-Poynter, Nicholas

2016-12-01

Cytomegaloviruses (CMVs) establish persistent, systemic infections and cause disease by maternal-foetal transfer, suggesting that their dissemination is a key target for antiviral intervention. Late clinical presentation has meant that human CMV (HCMV) dissemination is not well understood. Murine CMV (MCMV) provides a tractable model. Whole mouse imaging of virus-expressed luciferase has proved a useful way to track systemic infections. MCMV, in which the abundant lytic gene M78 was luciferase-tagged via a self-cleaving peptide (M78-LUC), allowed serial, unbiased imaging of systemic and peripheral infection without significant virus attenuation. Ex vivo luciferase imaging showed greater sensitivity than plaque assay, and revealed both well-known infection sites (the lungs, lymph nodes, salivary glands, liver, spleen and pancreas) and less explored sites (the bone marrow and upper respiratory tract). We applied luciferase imaging to tracking MCMV lacking M33, a chemokine receptor conserved in HCMV and a proposed anti-viral drug target. M33-deficient M78-LUC colonized normally in peripheral sites and local draining lymph nodes but spread poorly to the salivary gland, suggesting a defect in vascular transport consistent with properties of a chemokine receptor.

Therapeutic effects of autologous lymphocytes activated with trastuzumab for xenograft mouse models of human breast cancer.

PubMed

Nakagawa, Shinichiro; Matsuoka, Yusuke; Ichihara, Hideaki; Yoshida, Hitoji; Yoshida, Kenshi; Ueoka, Ryuichi

2013-01-01

Trastuzumab (TTZ) is molecular targeted drug used for metastatic breast cancer patients overexpressing human epidermal growth factor receptor 2 (HER2). Therapeutic effects of lymphocytes activated with TTZ (TTZ-LAK) using xenograft mouse models of human breast cancer (MDA-MB-453) cells were examined in vivo. Remarkable reduction of tumor volume in a xenograft mouse models intravenously treated with TTZ-LAK cells after the subcutaneously inoculated of MDA-MB-453 cells was verified in vivo. The migration of TTZ-LAK cells in tumor of mouse models subcutaneously inoculated MDA-MB-453 cells was observed on the basis of histological analysis using immunostaining with CD-3. Induction of apoptosis in tumor of xenograft mice treated with TTZ-LAK cells was observed in micrographs using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) method. It was noteworthy that the therapeutic effects of TTZ-LAK cells along with apoptosis were obtained for xenograft mouse models of human breast tumor in vivo.

Generation of a neuro-specific microarray reveals novel differentially expressed noncoding RNAs in mouse models for neurodegenerative diseases.

PubMed

Gstir, Ronald; Schafferer, Simon; Scheideler, Marcel; Misslinger, Matthias; Griehl, Matthias; Daschil, Nina; Humpel, Christian; Obermair, Gerald J; Schmuckermair, Claudia; Striessnig, Joerg; Flucher, Bernhard E; Hüttenhofer, Alexander

2014-12-01

We have generated a novel, neuro-specific ncRNA microarray, covering 1472 ncRNA species, to investigate their expression in different mouse models for central nervous system diseases. Thereby, we analyzed ncRNA expression in two mouse models with impaired calcium channel activity, implicated in Epilepsy or Parkinson's disease, respectively, as well as in a mouse model mimicking pathophysiological aspects of Alzheimer's disease. We identified well over a hundred differentially expressed ncRNAs, either from known classes of ncRNAs, such as miRNAs or snoRNAs or which represented entirely novel ncRNA species. Several differentially expressed ncRNAs in the calcium channel mouse models were assigned as miRNAs and target genes involved in calcium signaling, thus suggesting feedback regulation of miRNAs by calcium signaling. In the Alzheimer mouse model, we identified two snoRNAs, whose expression was deregulated prior to amyloid plaque formation. Interestingly, the presence of snoRNAs could be detected in cerebral spine fluid samples in humans, thus potentially serving as early diagnostic markers for Alzheimer's disease. In addition to known ncRNAs species, we also identified 63 differentially expressed, entirely novel ncRNA candidates, located in intronic or intergenic regions of the mouse genome, genomic locations, which previously have been shown to harbor the majority of functional ncRNAs. © 2014 Gstir et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

PDX-1 Is a Therapeutic Target for Pancreatic Cancer, Insulinoma and Islet Neoplasia Using a Novel RNA Interference Platform

PubMed Central

Liu, Shi-He; Rao, Donald D.; Nemunaitis, John; Senzer, Neil; Zhou, Guisheng; Dawson, David; Gingras, Marie-Claude; Wang, Zhaohui; Gibbs, Richard; Norman, Michael; Templeton, Nancy S.; DeMayo, Francesco J.; O'Malley, Bert; Sanchez, Robbi; Fisher, William E.; Brunicardi, F. Charles

2012-01-01

Pancreatic and duodenal homeobox-1 (PDX-1) is a transcription factor that regulates insulin expression and islet maintenance in the adult pancreas. Our recent studies demonstrate that PDX-1 is an oncogene for pancreatic cancer and is overexpressed in pancreatic cancer. The purpose of this study was to demonstrate that PDX-1 is a therapeutic target for both hormonal symptoms and tumor volume in mouse models of pancreatic cancer, insulinoma and islet neoplasia. Immunohistochemistry of human pancreatic and islet neoplasia specimens revealed marked PDX-1 overexpression, suggesting PDX-1 as a “drugable” target within these diseases. To do so, a novel RNA interference effector platform, bifunctional shRNAPDX-1, was developed and studied in mouse and human cell lines as well as in mouse models of pancreatic cancer, insulinoma and islet neoplasia. Systemic delivery of bi-shRNAhumanPDX-1 lipoplexes resulted in marked reduction of tumor volume and improved survival in a human pancreatic cancer xenograft mouse model. bi-shRNAmousePDX-1 lipoplexes prevented death from hyperinsulinemia and hypoglycemia in an insulinoma mouse model. shRNAmousePDX-1 lipoplexes reversed hyperinsulinemia and hypoglycemia in an immune-competent mouse model of islet neoplasia. PDX-1 was overexpressed in pancreatic neuroendocrine tumors and nesidioblastosis. These data demonstrate that PDX-1 RNAi therapy controls hormonal symptoms and tumor volume in mouse models of pancreatic cancer, insulinoma and islet neoplasia, therefore, PDX-1 is a potential therapeutic target for these pancreatic diseases. PMID:22905092

Development and Characterization of a Mouse Model for Marburg Hemorrhagic Fever

DTIC Science & Technology

2009-07-01

Microbiology. All Rights Reserved. Development and Characterization of a Mouse Model for Marburg Hemorrhagic Fever Kelly L. Warfield,* Steven B...mouse model has hampered an understanding of the pathogenesis and immunity of Marburg hemorrhagic fever (MHF), the disease caused by marburgvirus (MARV...cause severe hemorrhagic fevers in humans and non- human primates (27). The incubation time is estimated to be 3 to 21 days, with human case fatality

Producing a Mouse Model to Explore the Linkages Between Tocopherol Biology and Prostate Cancer

DTIC Science & Technology

2005-07-01

Edwards, Prostate cancer and supplementation with alpha-tocopherol and beta -carotene: incidence and mortality in a controlled trial. J Natl Cancer ...1-0153 TITLE: Producing a Mouse Model to Explore the Linkages Between Tocopherol Biology and Prostate Cancer ...TITLE AND SUBTITLE Producing a Mouse Model to Explore the Linkages Between Tocopherol 5a. CONTRACT NUMBER Biology and Prostate Cancer 5b. GRANT

Synergistic Action of FOXP3 and TSC1 Pathways During Tumor Progression

DTIC Science & Technology

2015-10-01

invasive carcinoma and, ultimately, metastatic disease [1-3]. Mouse models of PIN (mPIN) generated by a single- mutant gene in prostate do not progress...downstream target) is sufficient to significantly reduce the initiation of prostate cancer in the Pten conditional knockout mouse model [19-21...the possibility that these two genetic hits cooperate to promote tumor progression, and mouse models show that this cooperation accelerates

Assessing the potential for AAV vector genotoxicity in a murine model

PubMed Central

Li, Hojun; Malani, Nirav; Hamilton, Shari R.; Schlachterman, Alexander; Bussadori, Giulio; Edmonson, Shyrie E.; Shah, Rachel; Arruda, Valder R.; Mingozzi, Federico; Fraser Wright, J.; Bushman, Frederic D.

2011-01-01

Gene transfer using adeno-associated virus (AAV) vectors has great potential for treating human disease. Recently, questions have arisen about the safety of AAV vectors, specifically, whether integration of vector DNA in transduced cell genomes promotes tumor formation. This study addresses these questions with high-dose liver-directed AAV-mediated gene transfer in the adult mouse as a model (80 AAV-injected mice and 52 controls). After 18 months of follow-up, AAV-injected mice did not show a significantly higher rate of hepatocellular carcinoma compared with controls. Tumors in mice treated with AAV vectors did not have significantly different amounts of vector DNA compared with adjacent normal tissue. A novel high-throughput method for identifying AAV vector integration sites was developed and used to clone 1029 integrants. Integration patterns in tumor tissue and adjacent normal tissue were similar to each other, showing preferences for active genes, cytosine-phosphate-guanosine islands, and guanosine/cysteine-rich regions. Gene expression data showed that genes near integration sites did not show significant changes in expression patterns compared with genes more distal to integration sites. No integration events were identified as causing increased oncogene expression. Thus, we did not find evidence that AAV vectors cause insertional activation of oncogenes and subsequent tumor formation. PMID:21106988

ALS patients’ regulatory T lymphocytes are dysfunctional, and correlate with disease progression rate and severity

PubMed Central

Beers, David R.; Zhao, Weihua; Wang, Jinghong; Zhang, Xiujun; Wen, Shixiang; Neal, Dan; Thonhoff, Jason R.; Alsuliman, Abdullah S.; Shpall, Elizabeth J.; Rezvani, Katy

2017-01-01

Neuroinflammation is a pathological hallmark of ALS in both transgenic rodent models and patients, and is characterized by proinflammatory T lymphocytes and activated macrophages/microglia. In ALS mouse models, decreased regulatory T lymphocytes (Tregs) exacerbate the neuroinflammatory process, leading to accelerated motoneuron death and shortened survival; passive transfer of Tregs suppresses the neuroinflammation and prolongs survival. Treg numbers and FOXP3 expression are also decreased in rapidly progressing ALS patients. A key question is whether the marked neuroinflammation in ALS can be attributed to the impaired suppressive function of ALS Tregs in addition to their decreased numbers. To address this question, T lymphocyte proliferation assays were performed. Compared with control Tregs, ALS Tregs were less effective in suppressing responder T lymphocyte proliferation. Although both slowly and rapidly progressing ALS patients had dysfunctional Tregs, the greater the clinically assessed disease burden or the more rapidly progressing the patient, the greater the Treg dysfunction. Epigenetically, the percentage methylation of the Treg-specific demethylated region was greater in ALS Tregs. After in vitro expansion, ALS Tregs regained suppressive abilities to the levels of control Tregs, suggesting that autologous passive transfer of expanded Tregs might offer a novel cellular therapy to slow disease progression. PMID:28289705

Designing Mouse Behavioral Tasks Relevant to Autistic-Like Behaviors

ERIC Educational Resources Information Center

Crawley, Jacqueline N.

2004-01-01

The importance of genetic factors in autism has prompted the development of mutant mouse models to advance our understanding of biological mechanisms underlying autistic behaviors. Mouse models of human neuropsychiatric diseases are designed to optimize (1) face validity, i.e., resemblance to the human symptoms; (2) construct validity, i.e.,…

Behavioral phenotypes of genetic mouse models of autism

PubMed Central

Kazdoba, T. M.; Leach, P. T.; Crawley, J. N.

2016-01-01

More than a hundred de novo single gene mutations and copy-number variants have been implicated in autism, each occurring in a small subset of cases. Mutant mouse models with syntenic mutations offer research tools to gain an understanding of the role of each gene in modulating biological and behavioral phenotypes relevant to autism. Knockout, knockin and transgenic mice incorporating risk gene mutations detected in autism spectrum disorder and comorbid neurodevelopmental disorders are now widely available. At present, autism spectrum disorder is diagnosed solely by behavioral criteria. We developed a constellation of mouse behavioral assays designed to maximize face validity to the types of social deficits and repetitive behaviors that are central to an autism diagnosis. Mouse behavioral assays for associated symptoms of autism, which include cognitive inflexibility, anxiety, hyperactivity, and unusual reactivity to sensory stimuli, are frequently included in the phenotypic analyses. Over the past 10 years, we and many other laboratories around the world have employed these and additional behavioral tests to phenotype a large number of mutant mouse models of autism. In this review, we highlight mouse models with mutations in genes that have been identified as risk genes for autism, which work through synaptic mechanisms and through the mTOR signaling pathway. Robust, replicated autism-relevant behavioral outcomes in a genetic mouse model lend credence to a causal role for specific gene contributions and downstream biological mechanisms in the etiology of autism. PMID:26403076

Defining the role of polyamines in colon carcinogenesis using mouse models

PubMed Central

Ignatenko, Natalia A.; Gerner, Eugene W.; Besselsen, David G.

2011-01-01

Genetics and diet are both considered important risk determinants for colorectal cancer, a leading cause of death in the US and worldwide. Genetically engineered mouse (GEM) models have made a significant contribution to the characterization of colorectal cancer risk factors. Reliable, reproducible, and clinically relevant animal models help in the identification of the molecular events associated with disease progression and in the development of effictive treatment strategies. This review is focused on the use of mouse models for studying the role of polyamines in colon carcinogenesis. We describe how the available mouse models of colon cancer such as the multiple intestinal neoplasia (Min) mice and knockout genetic models facilitate understanding of the role of polyamines in colon carcinogenesis and help in the development of a rational strategy for colon cancer chemoprevention. PMID:21712957

CSE-MECC two-dimensional capillary electrophoresis analysis of proteins in the mouse tumor cell (AtT-20) homogenate

PubMed Central

Chen, Xingguo; Fazal, Md. Abul; Dovichi, Norman J.

2007-01-01

Two-dimensional capillary electrophoresis was used for the separation of proteins and biogenic amines from the mouse AtT-20 cell line. The first-dimension capillary contained a TRIS-CHES-SDS-dextran buffer to perform capillary sieving electrophoresis, which is based on molecular weight of proteins. The second-dimension capillary contained a TRIS-CHES-SDS buffer for micel1ar electrokinetic capillary chromatography. After a 61 seconds preliminary separation, fractions from the first-dimension capillary were successively transferred to the second-dimension capillary, where they further separated by MECC. The two-dimensional separation required 60 minutes. PMID:17637850

Selective plane illumination microscopy (SPIM) with time-domain fluorescence lifetime imaging microscopy (FLIM) for volumetric measurement of cleared mouse brain samples

NASA Astrophysics Data System (ADS)

Funane, Tsukasa; Hou, Steven S.; Zoltowska, Katarzyna Marta; van Veluw, Susanne J.; Berezovska, Oksana; Kumar, Anand T. N.; Bacskai, Brian J.

2018-05-01

We have developed an imaging technique which combines selective plane illumination microscopy with time-domain fluorescence lifetime imaging microscopy (SPIM-FLIM) for three-dimensional volumetric imaging of cleared mouse brains with micro- to mesoscopic resolution. The main features of the microscope include a wavelength-adjustable pulsed laser source (Ti:sapphire) (near-infrared) laser, a BiBO frequency-doubling photonic crystal, a liquid chamber, an electrically focus-tunable lens, a cuvette based sample holder, and an air (dry) objective lens. The performance of the system was evaluated with a lifetime reference dye and micro-bead phantom measurements. Intensity and lifetime maps of three-dimensional human embryonic kidney (HEK) cell culture samples and cleared mouse brain samples expressing green fluorescent protein (GFP) (donor only) and green and red fluorescent protein [positive Förster (fluorescence) resonance energy transfer] were acquired. The results show that the SPIM-FLIM system can be used for sample sizes ranging from single cells to whole mouse organs and can serve as a powerful tool for medical and biological research.

Host Genotype and Gut Microbiome Modulate Insulin Secretion and Diet-Induced Metabolic Phenotypes.

PubMed

Kreznar, Julia H; Keller, Mark P; Traeger, Lindsay L; Rabaglia, Mary E; Schueler, Kathryn L; Stapleton, Donald S; Zhao, Wen; Vivas, Eugenio I; Yandell, Brian S; Broman, Aimee Teo; Hagenbuch, Bruno; Attie, Alan D; Rey, Federico E

2017-02-14

Genetic variation drives phenotypic diversity and influences the predisposition to metabolic disease. Here, we characterize the metabolic phenotypes of eight genetically distinct inbred mouse strains in response to a high-fat/high-sucrose diet. We found significant variation in diabetes-related phenotypes and gut microbiota composition among the different mouse strains in response to the dietary challenge and identified taxa associated with these traits. Follow-up microbiota transplant experiments showed that altering the composition of the gut microbiota modifies strain-specific susceptibility to diet-induced metabolic disease. Animals harboring microbial communities with enhanced capacity for processing dietary sugars and for generating hydrophobic bile acids showed increased susceptibility to metabolic disease. Notably, differences in glucose-stimulated insulin secretion between different mouse strains were partially recapitulated via gut microbiota transfer. Our results suggest that the gut microbiome contributes to the genetic and phenotypic diversity observed among mouse strains and provide a link between the gut microbiome and insulin secretion. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Targeting Cancer Protein Profiles with Split-Enzyme Reporter Fragments to Achieve Chemical Resolution for Molecular Imaging

DTIC Science & Technology

2013-08-01

We next tested the utility of the construct to accumulate in tumors expressing EGFR using an orthotopic mouse model for brain tumors. Glioma cells...filament tumor marker, identified implanted cells within the orthotopic mouse model which were of human origin, i.e. Gli36Δ5 cells, and demonstrated that...forward into in vivo animal tumor model studies. • In vivo imaging of EGFR targeted-complex in orthotopic mouse model of brain tumor. • Ex vivo validation

The plasma membrane-associated NADH oxidase (ECTO-NOX) of mouse skin responds to blue light

NASA Technical Reports Server (NTRS)

Morre, D. James; Morre, Dorothy M.

2003-01-01

NADH oxidases of the external plasma membrane surface (ECTO-NOX proteins) are characterized by oscillations in activity with a regular period length of 24 min. Explants of mouse skin exhibit the oscillatory activity as estimated from the decrease in A(340) suggesting that individual ECTO-NOX molecules must somehow be induced to function synchronously. Transfer of explants of mouse skin from darkness to blue light (495 nm, 2 min, 50 micromol m(-1) s(-1)) resulted in initiation of a new activity maximum (entrainment) with a midpoint 36 min after light exposure followed by maxima every 24 min thereafter. Addition of melatonin resulted in a new maximum 24 min after melatonin addition. The findings suggest that the ECTO-NOX proteins play a central role in the entrainment of the biological clock both by light and by melatonin.

Properties of the branched-chain 2-hydroxy acid/2-oxo acid shuttle in mouse spermatozoa.

PubMed

Coronel, C E; Gallina, F G; Gerez de Burgos, N M; Burgos, C; Blanco, A

1986-05-01

Operation of the branched-chain 2-hydroxy acid/2-oxo acid shuttle for the transfer of reducing equivalents in mitochondria of mouse spermatozoa was studied in vitro in reconstituted systems. Results show that the branched-chain 2-oxo acids within the mitochondria are offered several metabolic pathways. (a) Decarboxylation: mouse sperm mitochondria possess high branched-chain 2-oxo acid decarboxylase activity. (b) Recycling to the cytosol by using a transport system which can be inhibited by alpha-cyano-3-hydroxycinnamate and pH 6.8. (c) Transamination to the corresponding amino acids: experiments presented indicate that leucine formed from 4-methyl-2-oxopentanoate may pass to the external phase, re-initiating the cycle. These two last possibilities would allow autocatalytic operation of the shuttle. The branched-chain 2-hydroxy acids apparently do not utilize the monocarboxylate carrier to penetrate the mitochondria.

Convection Enhanced Delivery of Recombinant Adeno-associated Virus into the Mouse Brain.

PubMed

Nash, Kevin R; Gordon, Marcia N

2016-01-01

Recombinant adeno-associated virus (rAAV) has become an extremely useful tool for the study of gene over expression or knockdown in the central nervous system of experimental animals. One disadvantage of intracranial injections of rAAV vectors into the brain parenchyma has been restricted distribution to relatively small volumes of the brain. Convection enhanced delivery (CED) is a method for delivery of clinically relevant amounts of therapeutic agents to large areas of the brain in a direct intracranial injection procedure. CED uses bulk flow to increase the hydrostatic pressure and thus improve volume distribution. The CED method has shown robust gene transfer and increased distribution within the CNS and can be successfully used for different serotypes of rAAV for increased transduction of the mouse CNS. This chapter details the surgical injection of rAAV by CED into a mouse brain.

Genetically engineered mouse models of melanoma.

PubMed

Pérez-Guijarro, Eva; Day, Chi-Ping; Merlino, Glenn; Zaidi, M Raza

2017-06-01

Melanoma is a complex disease that exhibits highly heterogeneous etiological, histopathological, and genetic features, as well as therapeutic responses. Genetically engineered mouse (GEM) models provide powerful tools to unravel the molecular mechanisms critical for melanoma development and drug resistance. Here, we expound briefly the basis of the mouse modeling design, the available technology for genetic engineering, and the aspects influencing the use of GEMs to model melanoma. Furthermore, we describe in detail the currently available GEM models of melanoma. Cancer 2017;123:2089-103. © 2017 American Cancer Society. © 2017 American Cancer Society.

CRISPR-Mediated Knockout of Cybb in NSG Mice Establishes a Model of Chronic Granulomatous Disease for Human Stem-Cell Gene Therapy Transplants.

PubMed

Sweeney, Colin L; Choi, Uimook; Liu, Chengyu; Koontz, Sherry; Ha, Seung-Kwon; Malech, Harry L

2017-07-01

Chronic granulomatous disease (CGD) is characterized by defects in the production of microbicidal reactive oxygen species (ROS) by phagocytes. Testing of gene and cell therapies for the treatment of CGD in human hematopoietic cells requires preclinical transplant models. The use of the lymphocyte-deficient NOD.Cg-Prkdc scid Il2rg tm1Wjl/ SzJ (NSG) mouse strain for human hematopoietic cell xenografts to test CGD therapies is complicated by the presence of functional mouse granulocytes capable of producing ROS for subsequent bacterial and fungal killing. To establish a phagocyte-defective mouse model of X-linked CGD (X-CGD) in NSG mice, clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 was utilized for targeted knockout of mouse Cybb on the X-chromosome by microinjection of NSG mouse zygotes with Cas9 mRNA and CRISPR single-guide RNA targeting Cybb exon 1 or exon 3. This resulted in a high incidence of indel formation at the CRISPR target site, with all mice exhibiting deletions in at least one Cybb allele based on sequence analysis of tail snip DNA. A female mouse heterozygous for a 235-bp deletion in Cybb exon 1 was bred to an NSG male to establish the X-CGD NSG mouse strain, NSG.Cybb[KO]. Resulting male offspring with the 235 bp deletion were found to be defective for production of ROS by neutrophils and other phagocytes, and demonstrated increased susceptibility to spontaneous bacterial and fungal infections with granulomatous inflammation. The establishment of the phagocyte-defective NSG.Cybb[KO] mouse model enables the in vivo assessment of gene and cell therapy strategies for treating CGD in human hematopoietic cell transplants without obfuscation by functional mouse phagocytes, and may also be useful for modeling other phagocyte disorders in humanized NSG mouse xenografts.

Genetically engineered mouse models of craniopharyngioma: an opportunity for therapy development and understanding of tumor biology.

PubMed

Apps, John Richard; Martinez-Barbera, Juan Pedro

2017-05-01

Adamantinomatous craniopharyngioma (ACP) is the commonest tumor of the sellar region in childhood. Two genetically engineered mouse models have been developed and are giving valuable insights into ACP biology. These models have identified novel pathways activated in tumors, revealed an important function of paracrine signalling and extended conventional theories about the role of organ-specific stem cells in tumorigenesis. In this review, we summarize these mouse models, what has been learnt, their limitations and open questions for future research. We then discussed how these mouse models may be used to test novel therapeutics against potentially targetable pathways recently identified in human ACP. © 2017 The Authors. Brain Pathology published by John Wiley & Sons Ltd on behalf of International Society of Neuropathology.

Late-Onset Cone Photoreceptor Degeneration Induced by R172W Mutation in Rds and Partial Rescue by Gene Supplementation

PubMed Central

Conley, Shannon; Nour, May; Fliesler, Steven J.; Naash, Muna I.

2008-01-01

Purpose R172W is a common mutation in the human retinal degeneration slow (RDS) gene, associated with a late-onset dominant macular dystrophy. In this study, the authors characterized a mouse model that closely mimics the human phenotype and tested the feasibility of gene supplementation as a disease treatment strategy. Methods Transgenic mouse lines carrying the R172W mutation were generated. The retinal phenotype associated with this mutation in a low-expresser line (L-R172W) was examined, both structurally (histology with correlative immunohistochemistry) and functionally (electroretinography). By examining animals over time and with various rds genetic backgrounds, the authors evaluated the dominance of the defect. To assess the efficacy of gene transfer therapy as a treatment for this defect, a previously characterized transgenic line expressing the normal mouse peripherin/Rds (NMP) was crossed with a higher-expresser Rds line harboring the R172W mutation (H-R172W). Functional, structural, and biochemical analyses were used to assess rescue of the retinal disease phenotype. Results In the wild-type (WT) background, L-R172W mice exhibited late-onset (12-month) dominant cone degeneration without any apparent effect on rods. The degeneration was slightly accelerated (9 months) in the rds+/− background. L-R172W retinas did not form outer segments in the absence of endogenous Rds. With use of the H-R172W line on an rds+/− background for proof-of-principle genetic supplementation studies, the NMP transgene product rescued rod and cone functional defects and supported outer segment integrity up to 3 months of age, but the rescue effect did not persist in older (11-month) animals. Conclusions The R172W mutation leads to dominant cone degeneration in the mouse model, regardless of the expression level of the transgene. In contrast, effects of the mutation on rods are dose dependent, underscoring the usefulness of the L-R172W line as a faithful model of the human phenotype. This model may prove helpful in future studies on the mechanisms of cone degeneration and for elucidating the different roles of Rds in rods and cones. This study provides evidence that Rds genetic supplementation can be used to partially rescue visual function. Although this strategy is capable of rescuing haplo-insufficiency, it does not rescue the long-term degeneration associated with a gain-of-function mutation. PMID:18055786

Nlrp3-dependent IL-1β inhibits CD103+ dendritic cell differentiation in the gut.

PubMed

Mak'Anyengo, Rachel; Duewell, Peter; Reichl, Cornelia; Hörth, Christine; Lehr, Hans-Anton; Fischer, Sandra; Clavel, Thomas; Denk, Gerald; Hohenester, Simon; Kobold, Sebastian; Endres, Stefan; Schnurr, Max; Bauer, Christian

2018-03-08

Inflammatory bowel disease (IBD) is associated with enhanced levels of the IL-1 family cytokines IL-1β and IL-18, which are activated by the Nlrp3 inflammasome. Here, we investigated the role of inflammasome-driven cytokine release on T cell polarization and DC differentiation in steady state and T cell transfer colitis. In vitro and in vivo data showed that IL-1β induces Th17 polarization and increases GM‑CSF production by T cells. Reduced IL-1β levels in Nlrp3-/- mice correlated with enhanced FLT3L levels and increased frequency of tolerogenic CD103+ DC. In the T cell transfer colitis model, Nlrp3 deficiency resulted in lower IL‑1β levels, reduced Th17 immunity, and less severe colitis. Unaltered IL-18 levels in both mouse strains pointed toward Nlrp3-independent processing. Importantly, cohousing revealed that the gut microbiome had no impact on the observed Nlrp3-/- phenotype. This study demonstrates that NLRP3 acts as a molecular switch of intestinal homeostasis by shifting local immune cells toward an inflammatory phenotype via IL-1β.

Reactive microglia drive tau pathology and contribute to the spreading of pathological tau in the brain

PubMed Central

Maphis, Nicole; Xu, Guixiang; Kokiko-Cochran, Olga N.; Jiang, Shanya; Cardona, Astrid; Ransohoff, Richard M.; Lamb, Bruce T.

2015-01-01

Pathological aggregation of tau is a hallmark of Alzheimer’s disease and related tauopathies. We have previously shown that the deficiency of the microglial fractalkine receptor (CX3CR1) led to the acceleration of tau pathology and memory impairment in an hTau mouse model of tauopathy. Here, we show that microglia drive tau pathology in a cell-autonomous manner. First, tau hyperphosphorylation and aggregation occur as early as 2 months of age in hTauCx3cr1−/− mice. Second, CD45+ microglial activation correlates with the spatial memory deficit and spread of tau pathology in the anatomically connected regions of the hippocampus. Third, adoptive transfer of purified microglia derived from hTauCx3cr1−/− mice induces tau hyperphosphorylation within the brains of non-transgenic recipient mice. Finally, inclusion of interleukin 1 receptor antagonist (Kineret®) in the adoptive transfer inoculum significantly reduces microglia-induced tau pathology. Together, our results suggest that reactive microglia are sufficient to drive tau pathology and correlate with the spread of pathological tau in the brain. PMID:25833819

Compensatory Limb Use and Behavioral Assessment of Motor Skill Learning Following Sensorimotor Cortex Injury in a Mouse Model of Ischemic Stroke

PubMed Central

Kerr, Abigail L.; Tennant, Kelly A.

2014-01-01

Mouse models have become increasingly popular in the field of behavioral neuroscience, and specifically in studies of experimental stroke. As models advance, it is important to develop sensitive behavioral measures specific to the mouse. The present protocol describes a skilled motor task for use in mouse models of stroke. The Pasta Matrix Reaching Task functions as a versatile and sensitive behavioral assay that permits experimenters to collect accurate outcome data and manipulate limb use to mimic human clinical phenomena including compensatory strategies (i.e., learned non-use) and focused rehabilitative training. When combined with neuroanatomical tools, this task also permits researchers to explore the mechanisms that support behavioral recovery of function (or lack thereof) following stroke. The task is both simple and affordable to set up and conduct, offering a variety of training and testing options for numerous research questions concerning functional outcome following injury. Though the task has been applied to mouse models of stroke, it may also be beneficial in studies of functional outcome in other upper extremity injury models. PMID:25045916

Protective immunity to Japanese encephalitis virus associated with anti-NS1 antibodies in a mouse model.

PubMed

Li, Yize; Counor, Dorian; Lu, Peng; Duong, Veasna; Yu, Yongxin; Deubel, Vincent

2012-07-24

Japanese encephalitis virus (JEV) is a major mosquito-borne pathogen that causes viral encephalitis throughout Asia. Vaccination with an inactive JEV particle or attenuated virus is an efficient preventative measure for controlling infection. Flavivirus NS1 protein is a glycoprotein secreted during viral replication that plays multiple roles in the viral life cycle and pathogenesis. Utilizing JEV NS1 as an antigen in viral vectors induces a limited protective immune response against infection. Previous studies using E. coli-expressed JEV NS1 to immunize mice induced protection against lethal challenge; however, the protection mechanism through cellular and humoral immune responses was not described. JEV NS1 was expressed in and purified from Drosophila S2 cells in a native glycosylated multimeric form, which induced T-cell and antibody responses in immunized C3H/HeN mice. Mice vaccinated with 1 μg NS1 with or without water-in-oil adjuvant were partially protected against viral challenge and higher protection was observed in mice with higher antibody titers. IgG1 was preferentially elicited by an adjuvanted NS1 protein, whereas a larger load of IFN-γ was produced in splenocytes from mice immunized with aqueous NS1. Mice that passively received anti-NS1 mouse polyclonal immune sera were protected, and this phenomenon was dose-dependent, whereas protection was low or delayed after the passive transfer of anti-NS1 MAbs. The purified NS1 subunit induced protective immunity in relation with anti-NS1 IgG1 antibodies. NS1 protein efficiently stimulated Th1-cell proliferation and IFN-γ production. Protection against lethal challenge was elicited by passive transfer of anti-NS1 antisera, suggesting that anti-NS1 antibodies play a substantial role in anti-viral immunity.

The Effects of Di-(2-ethylhexyl)-phthalate Exposure on Fertilization and Embryonic Development In Vitro and Testicular Genomic Mutation In Vivo

PubMed Central

Gu, Yi-Hua; Liu, Miao; Xu, Yan; Yuan, Yao; Sun, Fei; Zhang, Hui-Qin; Shi, Hui-Juan

2012-01-01

The present study was undertaken to determine the reproductive hazards of Di-(2-ethylhexyl)-phthalate (DEHP) on mouse spermatozoa and embryos in vitro and genomic changes in vivo. Direct low-level DEHP exposure (1 μg/ml) on spermatozoa and embryos was investigated by in vitro fertilization (IVF) process, culture of preimplanted embryos in DEHP-supplemented medium and embryo transfer to achieve full term development. Big Blue® transgenic mouse model was employed to evaluate the mutagenesis of testicular genome with in vivo exposure concentration of DEHP (500 mg/kg/day). Generally, DEHP-treated spermatozoa (1 μg/ml, 30 min) presented reduced fertilization ability (P<0.05) and the resultant embryos had decreased developmental potential compared to DMSO controls (P<0.05). Meanwhile, the transferred 2-cell stage embryos derived from treated spermatozoa also exhibited decreased birth rate than that of control (P<0.05). When fertilized oocytes or 2-cell stage embryos were recovered by in vivo fertilization (without treatment) and then exposed to DEHP, the subsequent development proceed to blastocysts was different, fertilized oocytes were significantly affected (P<0.05) whereas developmental progression of 2-cell stage embryos was similar to controls (P>0.05). Testes of the Big Blue® transgenic mice treated with DEHP for 4 weeks indicated an approximately 3-fold increase in genomic DNA mutation frequency compared with controls (P<0.05). These findings unveiled the hazardous effects of direct low-level exposure of DEHP on spermatozoa's fertilization ability as well as embryonic development, and proved that in vivo DEHP exposure posed mutagenic risks in the reproductive organ – at least in testes, are of great concern to human male reproductive health. PMID:23226291

Periodontitis induced by Porphyromonas gingivalis drives periodontal microbiota dysbiosis and insulin resistance via an impaired adaptive immune response.

PubMed

Blasco-Baque, Vincent; Garidou, Lucile; Pomié, Céline; Escoula, Quentin; Loubieres, Pascale; Le Gall-David, Sandrine; Lemaitre, Mathieu; Nicolas, Simon; Klopp, Pascale; Waget, Aurélie; Azalbert, Vincent; Colom, André; Bonnaure-Mallet, Martine; Kemoun, Philippe; Serino, Matteo; Burcelin, Rémy

2017-05-01

To identify a causal mechanism responsible for the enhancement of insulin resistance and hyperglycaemia following periodontitis in mice fed a fat-enriched diet. We set-up a unique animal model of periodontitis in C57Bl/6 female mice by infecting the periodontal tissue with specific and alive pathogens like Porphyromonas gingivalis ( Pg ), Fusobacterium nucleatum and Prevotella intermedia . The mice were then fed with a diabetogenic/non-obesogenic fat-enriched diet for up to 3 months. Alveolar bone loss, periodontal microbiota dysbiosis and features of glucose metabolism were quantified. Eventually, adoptive transfer of cervical (regional) and systemic immune cells was performed to demonstrate the causal role of the cervical immune system. Periodontitis induced a periodontal microbiota dysbiosis without mainly affecting gut microbiota. The disease concomitantly impacted on the regional and systemic immune response impairing glucose metabolism. The transfer of cervical lymph-node cells from infected mice to naive recipients guarded against periodontitis-aggravated metabolic disease. A treatment with inactivated Pg prior to the periodontal infection induced specific antibodies against Pg and protected the mouse from periodontitis-induced dysmetabolism. Finally, a 1-month subcutaneous chronic infusion of low rates of lipopolysaccharides from Pg mimicked the impact of periodontitis on immune and metabolic parameters. We identified that insulin resistance in the high-fat fed mouse is enhanced by pathogen-induced periodontitis. This is caused by an adaptive immune response specifically directed against pathogens and associated with a periodontal dysbiosis. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

The effects of Di-(2-ethylhexyl)-phthalate exposure on fertilization and embryonic development in vitro and testicular genomic mutation in vivo.

PubMed

Huang, Xue-Feng; Li, Yan; Gu, Yi-Hua; Liu, Miao; Xu, Yan; Yuan, Yao; Sun, Fei; Zhang, Hui-Qin; Shi, Hui-Juan

2012-01-01

The present study was undertaken to determine the reproductive hazards of Di-(2-ethylhexyl)-phthalate (DEHP) on mouse spermatozoa and embryos in vitro and genomic changes in vivo. Direct low-level DEHP exposure (1 μg/ml) on spermatozoa and embryos was investigated by in vitro fertilization (IVF) process, culture of preimplanted embryos in DEHP-supplemented medium and embryo transfer to achieve full term development. Big Blue® transgenic mouse model was employed to evaluate the mutagenesis of testicular genome with in vivo exposure concentration of DEHP (500 mg/kg/day). Generally, DEHP-treated spermatozoa (1 μg/ml, 30 min) presented reduced fertilization ability (P<0.05) and the resultant embryos had decreased developmental potential compared to DMSO controls (P<0.05). Meanwhile, the transferred 2-cell stage embryos derived from treated spermatozoa also exhibited decreased birth rate than that of control (P<0.05). When fertilized oocytes or 2-cell stage embryos were recovered by in vivo fertilization (without treatment) and then exposed to DEHP, the subsequent development proceed to blastocysts was different, fertilized oocytes were significantly affected (P<0.05) whereas developmental progression of 2-cell stage embryos was similar to controls (P>0.05). Testes of the Big Blue® transgenic mice treated with DEHP for 4 weeks indicated an approximately 3-fold increase in genomic DNA mutation frequency compared with controls (P<0.05). These findings unveiled the hazardous effects of direct low-level exposure of DEHP on spermatozoa's fertilization ability as well as embryonic development, and proved that in vivo DEHP exposure posed mutagenic risks in the reproductive organ - at least in testes, are of great concern to human male reproductive health.

Recombinant adeno-associated virus serotype 6 (rAAV2/6)-mediated gene transfer to nociceptive neurons through different routes of delivery

PubMed Central

Towne, Chris; Pertin, Marie; Beggah, Ahmed T; Aebischer, Patrick; Decosterd, Isabelle

2009-01-01

Background Gene transfer to nociceptive neurons of the dorsal root ganglia (DRG) is a promising approach to dissect mechanisms of pain in rodents and is a potential therapeutic strategy for the treatment of persistent pain disorders such as neuropathic pain. A number of studies have demonstrated transduction of DRG neurons using herpes simplex virus, adenovirus and more recently, adeno-associated virus (AAV). Recombinant AAV are currently the gene transfer vehicles of choice for the nervous system and have several advantages over other vectors, including stable and safe gene expression. We have explored the capacity of recombinant AAV serotype 6 (rAAV2/6) to deliver genes to DRG neurons and characterized the transduction of nociceptors through five different routes of administration in mice. Results Direct injection of rAAV2/6 expressing green fluorescent protein (eGFP) into the sciatic nerve resulted in transduction of up to 30% eGFP-positive cells of L4 DRG neurons in a dose dependant manner. More than 90% of transduced cells were small and medium sized neurons (< 700 μm2), predominantly colocalized with markers of nociceptive neurons, and had eGFP-positive central terminal fibers in the superficial lamina of the spinal cord dorsal horn. The efficiency and profile of transduction was independent of mouse genetic background. Intrathecal administration of rAAV2/6 gave the highest level of transduction (≈ 60%) and had a similar size profile and colocalization with nociceptive neurons. Intrathecal administration also transduced DRG neurons at cervical and thoracic levels and resulted in comparable levels of transduction in a mouse model for neuropathic pain. Subcutaneous and intramuscular delivery resulted in low levels of transduction in the L4 DRG. Likewise, delivery via tail vein injection resulted in relatively few eGFP-positive cells within the DRG, however, this transduction was observed at all vertebral levels and corresponded to large non-nociceptive cell types. Conclusion We have found that rAAV2/6 is an efficient vector to deliver transgenes to nociceptive neurons in mice. Furthermore, the characterization of the transduction profile may facilitate gene transfer studies to dissect mechanisms behind neuropathic pain. PMID:19737386

A comparison of some organizational characteristics of the mouse central retina and the human macula.

PubMed

Volland, Stefanie; Esteve-Rudd, Julian; Hoo, Juyea; Yee, Claudine; Williams, David S

2015-01-01

Mouse models have greatly assisted our understanding of retinal degenerations. However, the mouse retina does not have a macula, leading to the question of whether the mouse is a relevant model for macular degeneration. In the present study, a quantitative comparison between the organization of the central mouse retina and the human macula was made, focusing on some structural characteristics that have been suggested to be important in predisposing the macula to stresses leading to degeneration: photoreceptor density, phagocytic load on the RPE, and the relative thinness of Bruch's membrane. Light and electron microscopy measurements from retinas of two strains of mice, together with published data on human retinas, were used for calculations and subsequent comparisons. As in the human retina, the central region of the mouse retina possesses a higher photoreceptor cell density and a thinner Bruch's membrane than in the periphery; however, the magnitudes of these periphery to center gradients are larger in the human. Of potentially greater relevance is the actual photoreceptor cell density, which is much greater in the mouse central retina than in the human macula, underlying a higher phagocytic load for the mouse RPE. Moreover, at eccentricities that correspond to the peripheral half of the human macula, the rod to cone ratio is similar between mouse and human. Hence, with respect to photoreceptor density and phagocytic load of the RPE, the central mouse retina models at least the more peripheral part of the macula, where macular degeneration is often first evident.

A Genetically Engineered Mouse Model of Neuroblastoma Driven by Mutated ALK and MYCN

DTIC Science & Technology

2014-09-01

AWARD NUMBER: W81XWH-13-1-0220 TITLE: A Genetically Engineered Mouse Model of Neuroblastoma ...CONTRACT NUMBER A Genetically Engineered Mouse Model of Neuroblastoma Driven by Mutated ALK and MYCN 5b. GRANT NUMBER W81XWH-13-1-0220 5c...common ALK mutations in neuroblastoma , F1174L and R1275Q. We have determined that in tumors cells expressing mutated ALK, different downstream

Developing Novel Therapeutic Approaches in Small Cell Lung Carcinoma Using Genetically Engineered Mouse Models and Human Circulating Tumor Cells

DTIC Science & Technology

2014-10-01

AD_________________ Award Number: W81XWH-13-1-0325 TITLE: Developing Novel Therapeutic Approaches in Small Cell Lung Carcinoma Using ...Genetically Engineered Mouse Models and Human Circulating Tumor Cells PRINCIPAL INVESTIGATOR: Jeffrey Engelman MD PhD CONTRACTING ORGANIZATION ...Novel Therapeutic Approaches in Small Cell Lung 5a. CONTRACT NUMBER W81XWH-13-1-0325 Carcinoma Using Genetically Engineered Mouse Models and 5b

Behavioural phenotyping assays for mouse models of autism

PubMed Central

Silverman, Jill L.; Yang, Mu; Lord, Catherine; Crawley, Jacqueline N.

2011-01-01

Autism is a heterogeneous neurodevelopmental disorder of unknown aetiology that affects 1 in 100–150 individuals. Diagnosis is based on three categories of behavioural criteria: abnormal social interactions, communication deficits and repetitive behaviours. Strong evidence for a genetic basis has prompted the development of mouse models with targeted mutations in candidate genes for autism. As the diagnostic criteria for autism are behavioural, phenotyping these mouse models requires behavioural assays with high relevance to each category of the diagnostic symptoms. Behavioural neuroscientists are generating a comprehensive set of assays for social interaction, communication and repetitive behaviours to test hypotheses about the causes of austism. Robust phenotypes in mouse models hold great promise as translational tools for discovering effective treatments for components of autism spectrum disorders. PMID:20559336

Mucosal BCG Vaccination Induces Protective Lung-Resident Memory T Cell Populations against Tuberculosis.

PubMed

Perdomo, Carolina; Zedler, Ulrike; Kühl, Anja A; Lozza, Laura; Saikali, Philippe; Sander, Leif E; Vogelzang, Alexis; Kaufmann, Stefan H E; Kupz, Andreas

2016-11-22

Mycobacterium bovis Bacille Calmette-Guérin (BCG) is the only licensed vaccine against tuberculosis (TB), yet its moderate efficacy against pulmonary TB calls for improved vaccination strategies. Mucosal BCG vaccination generates superior protection against TB in animal models; however, the mechanisms of protection remain elusive. Tissue-resident memory T (T RM ) cells have been implicated in protective immune responses against viral infections, but the role of T RM cells following mycobacterial infection is unknown. Using a mouse model of TB, we compared protection and lung cellular infiltrates of parenteral and mucosal BCG vaccination. Adoptive transfer and gene expression analyses of lung airway cells were performed to determine the protective capacities and phenotypes of different memory T cell subsets. In comparison to subcutaneous vaccination, intratracheal and intranasal BCG vaccination generated T effector memory and T RM cells in the lung, as defined by surface marker phenotype. Adoptive mucosal transfer of these airway-resident memory T cells into naive mice mediated protection against TB. Whereas airway-resident memory CD4 + T cells displayed a mixture of effector and regulatory phenotype, airway-resident memory CD8 + T cells displayed prototypical T RM features. Our data demonstrate a key role for mucosal vaccination-induced airway-resident T cells in the host defense against pulmonary TB. These results have direct implications for the design of refined vaccination strategies. BCG remains the only licensed vaccine against TB. Parenterally administered BCG has variable efficacy against pulmonary TB, and thus, improved prevention strategies and a more refined understanding of correlates of vaccine protection are required. Induction of memory T cells has been shown to be essential for protective TB vaccines. Mimicking the natural infection route by mucosal vaccination has been known to generate superior protection against TB in animal models; however, the mechanisms of protection have remained elusive. Here we performed an in-depth analysis to dissect the immunological mechanisms associated with superior mucosal protection in the mouse model of TB. We found that mucosal, and not subcutaneous, BCG vaccination generates lung-resident memory T cell populations that confer protection against pulmonary TB. We establish a comprehensive phenotypic characterization of these populations, providing a framework for future vaccine development. Copyright © 2016 Perdomo et al.

Mouse cumulus-denuded oocytes restore developmental capacity completely when matured with optimal supplementation of cysteamine, cystine, and cumulus cells.

PubMed

Zhou, Ping; Wu, Yan-Guang; Wei, De-Li; Li, Qing; Wang, Gang; Zhang, Jie; Luo, Ming-Jiu; Tan, Jing-He

2010-04-01

Our objectives were to study how cysteamine, cystine, and cumulus cells (CCs), as well as oocytes interact to increase oocyte intracellular glutathione (GSH) and thereby to establish an efficient in vitro maturation system for cumulus-denuded oocytes (DOs). Using M16 that contained no thiol as maturation medium, we showed that when supplemented alone, neither cystine nor cysteamine promoted GSH synthesis of mouse DOs, but they did when used together. Although goat CCs required either cysteamine or cystine to promote GSH synthesis, mouse CCs required both. In the presence of cystine, goat CCs produced cysteine but mouse CCs did not. Cysteamine reduced cystine to cysteine in cell-free M16. When TCM-199 that contained 83 microM cystine was used as maturation medium, supplementation with cysteamine alone had no effect, but supplementation with 100 microM cysteamine and 200 microM cystine increased blastulation of DOs matured with CC coculture to a level as high as achieved in cumulus-surrounded oocytes (COCs). Similar numbers of young were produced after two-cell embryos from mouse COCs or CC-cocultured DOs matured with optimal thiol supplementation were transferred to pseudopregnant recipients. It is concluded that 1) mouse CCs can use neither cysteamine nor cystine to promote GSH synthesis, but goat CCs can use either one; 2) goat CCs promote mouse oocyte GSH synthesis by reducing cystine to cysteine, but how they use cysteamine requires further investigation; and 3) mouse DOs can use neither cystine nor cysteamine for GSH synthesis, but they restore developmental capacity completely when matured in the presence of optimum supplementation of cysteamine, cystine, and CCs.

A developmentally plastic adult mouse kidney cell line spontaneously generates multiple adult kidney structures

DOE Office of Scientific and Technical Information (OSTI.GOV)

Webb, Carol F., E-mail: carol-webb@omrf.org; Immunobiology and Cancer Research, Oklahoma Medical Research Foundation, Oklahoma City, OK; Department of Microbiology and Immunology, University of Oklahoma Health Sciences Center, Oklahoma City, OK

Despite exciting new possibilities for regenerative therapy posed by the ability to induce pluripotent stem cells, recapitulation of three-dimensional kidneys for repair or replacement has not been possible. ARID3a-deficient mouse tissues generated multipotent, developmentally plastic cells. Therefore, we assessed the adult mouse ARID3a−/− kidney cell line, KKPS5, which expresses renal progenitor surface markers as an alternative cell source for modeling kidney development. Remarkably, these cells spontaneously developed into multicellular nephron-like structures in vitro, and engrafted into immunocompromised medaka mesonephros, where they formed mouse nephron structures. These data implicate KKPS5 cells as a new model system for studying kidney development. - Highlights:more » • An ARID3a-deficient mouse kidney cell line expresses multiple progenitor markers. • This cell line spontaneously forms multiple nephron-like structures in vitro. • This cell line formed mouse kidney structures in immunocompromised medaka fish kidneys. • Our data identify a novel model system for studying kidney development.« less

Mouse phenotyping.

PubMed

Fuchs, Helmut; Gailus-Durner, Valérie; Adler, Thure; Aguilar-Pimentel, Juan Antonio; Becker, Lore; Calzada-Wack, Julia; Da Silva-Buttkus, Patricia; Neff, Frauke; Götz, Alexander; Hans, Wolfgang; Hölter, Sabine M; Horsch, Marion; Kastenmüller, Gabi; Kemter, Elisabeth; Lengger, Christoph; Maier, Holger; Matloka, Mikolaj; Möller, Gabriele; Naton, Beatrix; Prehn, Cornelia; Puk, Oliver; Rácz, Ildikó; Rathkolb, Birgit; Römisch-Margl, Werner; Rozman, Jan; Wang-Sattler, Rui; Schrewe, Anja; Stöger, Claudia; Tost, Monica; Adamski, Jerzy; Aigner, Bernhard; Beckers, Johannes; Behrendt, Heidrun; Busch, Dirk H; Esposito, Irene; Graw, Jochen; Illig, Thomas; Ivandic, Boris; Klingenspor, Martin; Klopstock, Thomas; Kremmer, Elisabeth; Mempel, Martin; Neschen, Susanne; Ollert, Markus; Schulz, Holger; Suhre, Karsten; Wolf, Eckhard; Wurst, Wolfgang; Zimmer, Andreas; Hrabě de Angelis, Martin

2011-02-01

Model organisms like the mouse are important tools to learn more about gene function in man. Within the last 20 years many mutant mouse lines have been generated by different methods such as ENU mutagenesis, constitutive and conditional knock-out approaches, knock-down, introduction of human genes, and knock-in techniques, thus creating models which mimic human conditions. Due to pleiotropic effects, one gene may have different functions in different organ systems or time points during development. Therefore mutant mouse lines have to be phenotyped comprehensively in a highly standardized manner to enable the detection of phenotypes which might otherwise remain hidden. The German Mouse Clinic (GMC) has been established at the Helmholtz Zentrum München as a phenotyping platform with open access to the scientific community (www.mousclinic.de; [1]). The GMC is a member of the EUMODIC consortium which created the European standard workflow EMPReSSslim for the systemic phenotyping of mouse models (http://www.eumodic.org/[2]). Copyright © 2010 Elsevier Inc. All rights reserved.

Gene transfer and gene mapping in mammalian cells in culture.

PubMed

Shows, T B; Sakaguchi, A Y

1980-01-01

The ability to transfer mammalian genes parasexually has opened new possibilities for gene mapping and fine structure mapping and offers great potential for contributing to several aspects of mammalian biology, including gene expression and genetic engineering. The DNA transferred has ranged from whole genomes to single genes and smaller segments of DNA. The transfer of whole genomes by cell fusion forms cell hybrids, which has promoted the extensive mapping of human and mouse genes. Transfer, by cell fusion, of rearranged chromosomes has contributed significantly to determining close linkage and the assignment of genes to specific chromosomal regions. Transfer of single chromosomes has been achieved utilizing microcells fused to recipient cells. Metaphase chromosomes have been isolated and used to transfer single-to-multigenic DNA segments. DNA-mediated gene transfer, simulating bacterial transformation, has achieved transfer of single-copy genes. By utilizing DNA cleaved with restriction endonucleases, gene transfer is being empolyed as a bioassay for the purification of genes. Gene mapping and the fate of transferred genes can be examined now at the molecular level using sequence-specific probles. Recently, single genes have been cloned into eucaryotic and procaryotic vectors for transfer into mammalian cells. Moreover, recombinant libraries in which entire mammalian genomes are represented collectively are a rich new source of transferable genes. Methodology for transferring mammalian genetic information and applications for mapping mammalian genes is presented and prospects for the future discussed.

Interspecies chimera between primate embryonic stem cells and mouse embryos: monkey ESCs engraft into mouse embryos, but not post-implantation fetuses.

PubMed

Simerly, Calvin; McFarland, Dave; Castro, Carlos; Lin, Chih-Cheng; Redinger, Carrie; Jacoby, Ethan; Mich-Basso, Jocelyn; Orwig, Kyle; Mills, Parker; Ahrens, Eric; Navara, Chris; Schatten, Gerald

2011-07-01

Unequivocal evidence for pluripotency in which embryonic stem cells contribute to chimeric offspring has yet to be demonstrated in human or nonhuman primates (NHPs). Here, rhesus and baboons ESCs were investigated in interspecific mouse chimera generated by aggregation or blastocyst injection. Aggregation chimera produced mouse blastocysts with GFP-nhpESCs at the inner cell mass (ICM), and embryo transfers (ETs) generated dimly-fluorescencing abnormal fetuses. Direct injection of GFP-nhpESCs into blastocysts produced normal non-GFP-fluorescencing fetuses. Injected chimera showed >70% loss of GFP-nhpESCs after 21 h culture. Outgrowths of all chimeric blastocysts established distinct but separate mouse- and NHP-ESC colonies. Extensive endogenous autofluorescence compromised anti-GFP detection and PCR analysis did not detect nhpESCs in fetuses. NhpESCs localize to the ICM in chimera and generate pregnancies. Because primate ESCs do not engraft post-implantation, and also because endogenous autofluorescence results in misleading positive signals, interspecific chimera assays for pluripotency with primate stem cells is unreliable with the currently available ESCs. Testing primate ESCs reprogrammed into even more naïve states in these inter-specific chimera assays will be an important future endeavor. Copyright © 2011 Elsevier B.V. All rights reserved.

Human androgen deficiency: insights gained from androgen receptor knockout mouse models

PubMed Central

Rana, Kesha; Davey, Rachel A; Zajac, Jeffrey D

2014-01-01

The mechanism of androgen action is complex. Recently, significant advances have been made into our understanding of how androgens act via the androgen receptor (AR) through the use of genetically modified mouse models. A number of global and tissue-specific AR knockout (ARKO) models have been generated using the Cre-loxP system which allows tissue- and/or cell-specific deletion. These ARKO models have examined a number of sites of androgen action including the cardiovascular system, the immune and hemopoetic system, bone, muscle, adipose tissue, the prostate and the brain. This review focuses on the insights that have been gained into human androgen deficiency through the use of ARKO mouse models at each of these sites of action, and highlights the strengths and limitations of these Cre-loxP mouse models that should be considered to ensure accurate interpretation of the phenotype. PMID:24480924

Modelling clinical systemic lupus erythematosus: similarities, differences and success stories

PubMed Central

Celhar, Teja

2017-01-01

Abstract Mouse models of SLE have been indispensable tools to study disease pathogenesis, to identify genetic susceptibility loci and targets for drug development, and for preclinical testing of novel therapeutics. Recent insights into immunological mechanisms of disease progression have boosted a revival in SLE drug development. Despite promising results in mouse studies, many novel drugs have failed to meet clinical end points. This is probably because of the complexity of the disease, which is driven by polygenic predisposition and diverse environmental factors, resulting in a heterogeneous clinical presentation. Each mouse model recapitulates limited aspects of lupus, especially in terms of the mechanism underlying disease progression. The main mouse models have been fairly successful for the evaluation of broad-acting immunosuppressants. However, the advent of targeted therapeutics calls for a selection of the most appropriate model(s) for testing and, ultimately, identification of patients who will be most likely to respond. PMID:28013204

Mouse Models of Gastric Cancer

PubMed Central

Hayakawa, Yoku; Fox, James G.; Gonda, Tamas; Worthley, Daniel L.; Muthupalani, Sureshkumar; Wang, Timothy C.

2013-01-01

Animal models have greatly enriched our understanding of the molecular mechanisms of numerous types of cancers. Gastric cancer is one of the most common cancers worldwide, with a poor prognosis and high incidence of drug-resistance. However, most inbred strains of mice have proven resistant to gastric carcinogenesis. To establish useful models which mimic human gastric cancer phenotypes, investigators have utilized animals infected with Helicobacter species and treated with carcinogens. In addition, by exploiting genetic engineering, a variety of transgenic and knockout mouse models of gastric cancer have emerged, such as INS-GAS mice and TFF1 knockout mice. Investigators have used the combination of carcinogens and gene alteration to accelerate gastric cancer development, but rarely do mouse models show an aggressive and metastatic gastric cancer phenotype that could be relevant to preclinical studies, which may require more specific targeting of gastric progenitor cells. Here, we review current gastric carcinogenesis mouse models and provide our future perspectives on this field. PMID:24216700

Characterization of skin abnormalities in a mouse model of osteogenesis imperfecta using high resolution magnetic resonance imaging and Fourier transform infrared imaging spectroscopy.

PubMed

Canuto, H C; Fishbein, K W; Huang, A; Doty, S B; Herbert, R A; Peckham, J; Pleshko, N; Spencer, R G

2012-01-01

Evaluation of the skin phenotype in osteogenesis imperfecta (OI) typically involves biochemical measurements, such as histologic or biochemical assessment of the collagen produced from biopsy-derived dermal fibroblasts. As an alternative, the current study utilized non-invasive magnetic resonance imaging (MRI) microscopy and optical spectroscopy to define biophysical characteristics of skin in an animal model of OI. MRI of skin harvested from control, homozygous oim/oim and heterozygous oim/+ mice demonstrated several differences in anatomic and biophysical properties. Fourier transform infrared imaging spectroscopy (FT-IRIS) was used to interpret observed MRI signal characteristics in terms of chemical composition. Differences between wild-type and OI mouse skin included the appearance of a collagen-depleted lower dermal layer containing prominent hair follicles in the oim/oim mice, accounting for 55% of skin thickness in these. The MRI magnetization transfer rate was lower by 50% in this layer as compared to the upper dermis, consistent with lower collagen content. The MRI transverse relaxation time, T2, was greater by 30% in the dermis of the oim/oim mice compared to controls, consistent with a more highly hydrated collagen network. Similarly, an FT-IRIS-defined measure of collagen integrity was 30% lower in the oim/oim mice. We conclude that characterization of phenotypic differences between the skin of OI and wild-type mice by MRI and FT-IRIS is feasible, and that these techniques provide powerful complementary approaches for the analysis of the skin phenotype in animal models of disease. Copyright © 2011 John Wiley & Sons, Ltd.

Transplantation of Hepatocyte Growth Factor-Modified Dental Pulp Stem Cells Prevents Bone Loss in the Early Phase of Ovariectomy-Induced Osteoporosis.

PubMed

Kong, Fanxuan; Shi, Xuefeng; Xiao, Fengjun; Yang, Yuefeng; Zhang, Xiaoyan; Wang, Li-Sheng; Wu, Chu-Tse; Wang, Hua

2018-02-01

Investigations based on mesenchymal stem cells (MSCs) for osteoporosis have attracted attention recently. MSCs can be derived from various tissues, such as bone marrow, adipose, umbilical cord, placenta, and dental pulp. Among these, dental pulp-derived MSCs (DPSCs) and hepatocyte growth factor (HGF)-modified DPSCs (DPSCs-HGF) highly express osteogenic-related genes and have stronger osteogenic differentiation capacities. DPSCs have more benefits in treating osteoporosis. The purpose of this study was to investigate the roles of HGF gene-modified DPSCs in bone regeneration using a mouse model of ovariectomy (OVX)-induced bone loss. The HGF and luciferase genes were transferred into human DPSCs using recombinant adenovirus. These transduced cells were assayed for distribution or bone regeneration assay by transplantation into an OVX-induced osteoporosis model. By using bioluminogenic imaging, it was determined that some DPSCs could survive for >1 month in vivo. The DPSCs were mainly distributed to the lung in the early stage and to the liver in the late stage of OVX osteoporosis after administration, but they were scarcely distributed to the bone. The homing efficiency of DPSCs is higher when administrated in the early stage of a mouse OVX model. Micro-computed tomography indicated that DPSCs-Null or DPSCs-HGF transplantation significantly reduces OVX-induced bone loss in the trabecular bone of the distal femur metaphysis, and DPSCs-HGF show a stronger capacity to reduce bone loss. The data suggest that systemic infusion of DPSCs-HGF is a potential therapeutic approach for OVX-induced bone loss, which might be mediated by paracrine mechanisms.

Phenotyping male infertility in the mouse: how to get the most out of a 'non-performer'.

PubMed

Borg, Claire L; Wolski, Katja M; Gibbs, Gerard M; O'Bryan, Moira K

2010-01-01

Functional male gametes are produced through complex processes that take place within the testis, epididymis and female reproductive tract. A breakdown at any of these phases can result in male infertility. The production of mutant mouse models often yields an unexpected male infertility phenotype. It is with this in mind that the current review has been written. The review aims to act as a guide to the 'non-reproductive biologist' to facilitate a systematic analysis of sterile or subfertile mice and to assist in extracting the maximum amount of information from each model. This is a review of the original literature on defects in the processes that take a mouse spermatogonial stem cell through to a fully functional spermatozoon, which result in male infertility. Based on literature searches and personal experience, we have outlined a step-by-step strategy for the analysis of an infertile male mouse line. A wide range of methods can be used to define the phenotype of an infertile male mouse. These methods range from histological methods such as electron microscopy and immunohistochemistry, to hormone analyses and methods to assess sperm maturation status and functional competence. With the increased rate of genetically modified mouse production, the generation of mouse models with unexpected male infertility is increasing. This manuscript will help to ensure that the maximum amount of information is obtained from each mouse model and, by extension, will facilitate the knowledge of both normal fertility processes and the causes of human infertility.

Time evolution of interhemispheric coupling in a model of focal neocortical epilepsy in mice

NASA Astrophysics Data System (ADS)

Vallone, F.; Vannini, E.; Cintio, A.; Caleo, M.; Di Garbo, A.

2016-09-01

Epilepsy is characterized by substantial network rearrangements leading to spontaneous seizures and little is known on how an epileptogenic focus impacts on neural activity in the contralateral hemisphere. Here, we used a model of unilateral epilepsy induced by injection of the synaptic blocker tetanus neurotoxin (TeNT) in the mouse primary visual cortex (V1). Local field potential (LFP) signals were simultaneously recorded from both hemispheres of each mouse in acute phase (peak of toxin action) and chronic condition (completion of TeNT effects). To characterize the neural electrical activities the corresponding LFP signals were analyzed with several methods of time series analysis. For the epileptic mice, the spectral analysis showed that TeNT determines a power redistribution among the different neurophysiological bands in both acute and chronic phases. Using linear and nonlinear interdependence measures in both time and frequency domains, it was found in the acute phase that TeNT injection promotes a reduction of the interhemispheric coupling for high frequencies (12 -30 Hz) and small time lag (<20 ms), whereas an increase of the coupling is present for low frequencies (0.5 -4 Hz) and long time lag (>40 ms). On the other hand, the chronic period is characterized by a partial or complete recovery of the interhemispheric interdependence level. Granger causality test and symbolic transfer entropy indicate a greater driving influence of the TeNT-injected side on activity in the contralateral hemisphere in the chronic phase. Lastly, based on experimental observations, we built a computational model of LFPs to investigate the role of the ipsilateral inhibition and exicitatory interhemispheric connections in the dampening of the interhemispheric coupling. The time evolution of the interhemispheric coupling in such a relevant model of epilepsy has been addressed here.

Time evolution of interhemispheric coupling in a model of focal neocortical epilepsy in mice.

PubMed

Vallone, F; Vannini, E; Cintio, A; Caleo, M; Di Garbo, A

2016-09-01

Epilepsy is characterized by substantial network rearrangements leading to spontaneous seizures and little is known on how an epileptogenic focus impacts on neural activity in the contralateral hemisphere. Here, we used a model of unilateral epilepsy induced by injection of the synaptic blocker tetanus neurotoxin (TeNT) in the mouse primary visual cortex (V1). Local field potential (LFP) signals were simultaneously recorded from both hemispheres of each mouse in acute phase (peak of toxin action) and chronic condition (completion of TeNT effects). To characterize the neural electrical activities the corresponding LFP signals were analyzed with several methods of time series analysis. For the epileptic mice, the spectral analysis showed that TeNT determines a power redistribution among the different neurophysiological bands in both acute and chronic phases. Using linear and nonlinear interdependence measures in both time and frequency domains, it was found in the acute phase that TeNT injection promotes a reduction of the interhemispheric coupling for high frequencies (12-30 Hz) and small time lag (<20 ms), whereas an increase of the coupling is present for low frequencies (0.5-4 Hz) and long time lag (>40 ms). On the other hand, the chronic period is characterized by a partial or complete recovery of the interhemispheric interdependence level. Granger causality test and symbolic transfer entropy indicate a greater driving influence of the TeNT-injected side on activity in the contralateral hemisphere in the chronic phase. Lastly, based on experimental observations, we built a computational model of LFPs to investigate the role of the ipsilateral inhibition and exicitatory interhemispheric connections in the dampening of the interhemispheric coupling. The time evolution of the interhemispheric coupling in such a relevant model of epilepsy has been addressed here.

Obeticholic acid raises LDL-cholesterol and reduces HDL-cholesterol in the Diet-Induced NASH (DIN) hamster model.

PubMed

Briand, François; Brousseau, Emmanuel; Quinsat, Marjolaine; Burcelin, Rémy; Sulpice, Thierry

2018-01-05

The use of rat and mouse models limits the translation to humans for developing novel drugs targeting nonalcoholic steatohepatitis (NASH). Obeticholic acid (OCA) illustrates this limitation since its dyslipidemic effect in humans cannot be observed in these rodents. Conversely, Golden Syrian hamsters have a lipoprotein metabolism mimicking human dyslipidemia since it does express the cholesteryl ester transfer protein (CETP). We therefore developed a Diet-Induced NASH (DIN) hamster model and evaluated the impact of OCA. Compared with chow fed controls, hamsters fed for 20 weeks with a free-choice (FC) diet, developed obesity, insulin resistance, dyslipidemia and NASH (microvesicular steatosis, inflammation, hepatocyte ballooning and perisinusoidal to bridging fibrosis). After 20 weeks of diet, FC fed hamsters were treated without or with obeticholic acid (15mg/kg/day) for 5 weeks. Although a non-significant trend towards higher dietary caloric intake was observed, OCA significantly lowered body weight after 5 weeks of treatment. OCA significantly increased CETP activity and LDL-C levels by 20% and 27%, and reduced HDL-C levels by 20%. OCA blunted hepatic gene expression of Cyp7a1 and Cyp8b1 and reduced fecal bile acids mass excretion by 64% (P < 0.05). Hamsters treated with OCA showed a trend towards higher scavenger receptor Class B type I (SR-BI) and lower LDL-receptor hepatic protein expression. OCA reduced NAS score for inflammation (P < 0.01) and total NAS score, although not significantly. Compared to mouse and rat models, the DIN hamster replicates benefits and side effects of OCA as observed in humans, and should be useful for evaluating novel drugs targeting NASH. Copyright © 2017 Elsevier B.V. All rights reserved.

Use of electroporation for high-molecular-weight DNA-mediated gene transfer.

PubMed

Jastreboff, M M; Ito, E; Bertino, J R; Narayanan, R

1987-08-01

Electroporation was used to introduce high-molecular-weight DNA into murine hematopoietic cells and NIH3T3 cells. CCRF-CEM cells were stably transfected with SV2NEO plasmid and the genomic DNA from G-418-resistant clones (greater than 65 kb) was introduced into mouse bone marrow and NIH3T3 cells by electroporation. NEO sequences and expression were detected in the hematopoietic tissues of lethally irradiated mice, with 24% of individual spleen colonies expressing NEO. The frequency of genomic DNA transfer into NIH3T3 cells was 0.25 X 10(-3). Electroporation thus offers a powerful mode of gene transfer not only of cloned genes but also of high-molecular-weight DNA into cells.

[Thymus dependent immunity against Tyzzer's disease in the mouse].

PubMed

Fujiwara, K; Machii, K; Nakayama, M; Tamura, T; Ueda, K

1977-01-01

Nude (nu/nu) mice fail to resist to challenge infection of Tyzzer's disease after pretreatment with formalin-killed organisms that was effective for protecting heterozygous haired (nu/ł) mice from challenge. Resistance was induced nu/nu mice after the transfer of spleen cells from immunized nu/ł and concomitant formalin vaccine treatment.

Genetic tracing of the gustatory and trigeminal neural pathways originating from T1R3-expressing taste receptor cells and solitary chemoreceptor cells.

PubMed

Ohmoto, Makoto; Matsumoto, Ichiro; Yasuoka, Akihito; Yoshihara, Yoshihiro; Abe, Keiko

2008-08-01

We established transgenic mouse lines expressing a transneuronal tracer, wheat germ agglutinin (WGA), under the control of mouse T1R3 gene promoter/enhancer. In the taste buds, WGA transgene was faithfully expressed in T1R3-positive sweet/umami taste receptor cells. WGA protein was transferred not laterally to the synapse-bearing, sour-responsive type III cells in the taste buds but directly to a subset of neurons in the geniculate and nodose/petrosal ganglia, and further conveyed to a rostro-central region of the nucleus of solitary tract. In addition, WGA was expressed in solitary chemoreceptor cells in the nasal epithelium and transferred along the trigeminal sensory pathway to the brainstem neurons. The solitary chemoreceptor cells endogenously expressed T1R3 together with bitter taste receptors T2Rs. This result shows an exceptional signature of receptor expression. Thus, the t1r3-WGA transgenic mice revealed the sweet/umami gustatory pathways from taste receptor cells and the trigeminal neural pathway from solitary chemoreceptor cells.

Behavioral phenotypes of genetic mouse models of autism.

PubMed

Kazdoba, T M; Leach, P T; Crawley, J N

2016-01-01

More than a hundred de novo single gene mutations and copy-number variants have been implicated in autism, each occurring in a small subset of cases. Mutant mouse models with syntenic mutations offer research tools to gain an understanding of the role of each gene in modulating biological and behavioral phenotypes relevant to autism. Knockout, knockin and transgenic mice incorporating risk gene mutations detected in autism spectrum disorder and comorbid neurodevelopmental disorders are now widely available. At present, autism spectrum disorder is diagnosed solely by behavioral criteria. We developed a constellation of mouse behavioral assays designed to maximize face validity to the types of social deficits and repetitive behaviors that are central to an autism diagnosis. Mouse behavioral assays for associated symptoms of autism, which include cognitive inflexibility, anxiety, hyperactivity, and unusual reactivity to sensory stimuli, are frequently included in the phenotypic analyses. Over the past 10 years, we and many other laboratories around the world have employed these and additional behavioral tests to phenotype a large number of mutant mouse models of autism. In this review, we highlight mouse models with mutations in genes that have been identified as risk genes for autism, which work through synaptic mechanisms and through the mTOR signaling pathway. Robust, replicated autism-relevant behavioral outcomes in a genetic mouse model lend credence to a causal role for specific gene contributions and downstream biological mechanisms in the etiology of autism. © 2015 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

Using Genetic Mouse Models to Gain Insight into Glaucoma: Past Results and Future Possibilities

PubMed Central

Fernandes, Kimberly A.; Harder, Jeffrey M.; Williams, Pete A.; Rausch, Rebecca L.; Kiernan, Amy E.; Nair, K. Saidas; Anderson, Michael G.; John, Simon W.; Howell, Gareth R.; Libby, Richard T.

2015-01-01

While all forms of glaucoma are characterized by a specific pattern of retinal ganglion cell death, they are clinically divided into several distinct subclasses, including normal tension glaucoma, primary open angle glaucoma, congenital glaucoma, and secondary glaucoma. For each type of glaucoma there are likely numerous molecular pathways that control susceptibility to the disease. Given this complexity, a single animal model will never precisely model all aspects of all the different types of human glaucoma. Therefore, multiple animal models have been utilized to study glaucoma but more are needed. Because of the powerful genetic tools available to use in the laboratory mouse, it has proven to be a highly useful mammalian system for studying the pathophysiology of human disease. The similarity between human and mouse eyes coupled with the ability to use a combination of advanced cell biological and genetic tools in mice have led to a large increase in the number of studies using mice to model specific glaucoma phenotypes. Over the last decade, numerous new mouse models and genetic tools have emerged, providing important insight into the cell biology and genetics of glaucoma. In this review, we describe available mouse genetic models that can be used to study glaucoma-relevant disease/pathobiology. Furthermore, we discuss how these models have been used to gain insights into ocular hypertension (a major risk factor for glaucoma) and glaucomatous retinal ganglion cell death. Finally, the potential for developing new mouse models and using advanced genetic tools and resources for studying glaucoma are discussed. PMID:26116903

Neurocognitive endophenotypes in CGG KI and Fmr1 KO mouse models of Fragile X-Associated disorders: an analysis of the state of the field

PubMed Central

Hunsaker, Michael R.

2013-01-01

It has become increasingly important that the field of behavioral genetics identifies not only the gross behavioral phenotypes associated with a given mutation, but also the behavioral endophenotypes that scale with the dosage of the particular mutation being studied. Over the past few years, studies evaluating the effects of the polymorphic CGG trinucleotide repeat on the FMR1 gene underlying Fragile X-Associated Disorders have reported preliminary evidence for a behavioral endophenotype in human Fragile X Premutation carrier populations as well as the CGG knock-in (KI) mouse model. More recently, the behavioral experiments used to test the CGG KI mouse model have been extended to the Fmr1 knock-out (KO) mouse model. When combined, these data provide compelling evidence for a clear neurocognitive endophenotype in the mouse models of Fragile X-Associated Disorders such that behavioral deficits scale predictably with genetic dosage. Similarly, it appears that the CGG KI mouse effectively models the histopathology in Fragile X-Associated Disorders across CGG repeats well into the full mutation range, resulting in a reliable histopathological endophenotype. These endophenotypes may influence future research directions into treatment strategies for not only Fragile X Syndrome, but also the Fragile X Premutation and Fragile X-Associated Tremor/Ataxia Syndrome (FXTAS). PMID:24627796

Long non-coding RNA expression patterns in lung tissues of chronic cigarette smoke induced COPD mouse model.

PubMed

Zhang, Haiyun; Sun, Dejun; Li, Defu; Zheng, Zeguang; Xu, Jingyi; Liang, Xue; Zhang, Chenting; Wang, Sheng; Wang, Jian; Lu, Wenju

2018-05-15

Long non-coding RNAs (lncRNAs) have critical regulatory roles in protein-coding gene expression. Aberrant expression profiles of lncRNAs have been observed in various human diseases. In this study, we investigated transcriptome profiles in lung tissues of chronic cigarette smoke (CS)-induced COPD mouse model. We found that 109 lncRNAs and 260 mRNAs were significantly differential expressed in lungs of chronic CS-induced COPD mouse model compared with control animals. GO and KEGG analyses indicated that differentially expressed lncRNAs associated protein-coding genes were mainly involved in protein processing of endoplasmic reticulum pathway, and taurine and hypotaurine metabolism pathway. The combination of high throughput data analysis and the results of qRT-PCR validation in lungs of chronic CS-induced COPD mouse model, 16HBE cells with CSE treatment and PBMC from patients with COPD revealed that NR_102714 and its associated protein-coding gene UCHL1 might be involved in the development of COPD both in mouse and human. In conclusion, our study demonstrated that aberrant expression profiles of lncRNAs and mRNAs existed in lungs of chronic CS-induced COPD mouse model. From animal models perspective, these results might provide further clues to investigate biological functions of lncRNAs and their potential target protein-coding genes in the pathogenesis of COPD.

Modeling bladder cancer in mice: opportunities and challenges

PubMed Central

Kobayashi, Takashi; Owczarek, Tomasz B.; McKiernan, James M.; Abate-Shen, Cory

2015-01-01

The prognosis and treatment of bladder cancer have hardly improved in the last 20 years. Bladder cancer remains a debilitating and often fatal disease, and among the most costly cancers to treat. The generation of informative mouse models has the potential to improve our understanding of bladder cancer progression, as well as impact its diagnosis and treatment. However, relatively few mouse models of bladder cancer have been described and particularly few that develop invasive cancer phenotypes. This review focuses on opportunities for improving the landscape of mouse models of bladder cancer. PMID:25533675

Development and function of human innate immune cells in a humanized mouse model.

PubMed

Rongvaux, Anthony; Willinger, Tim; Martinek, Jan; Strowig, Till; Gearty, Sofia V; Teichmann, Lino L; Saito, Yasuyuki; Marches, Florentina; Halene, Stephanie; Palucka, A Karolina; Manz, Markus G; Flavell, Richard A

2014-04-01

Mice repopulated with human hematopoietic cells are a powerful tool for the study of human hematopoiesis and immune function in vivo. However, existing humanized mouse models cannot support development of human innate immune cells, including myeloid cells and natural killer (NK) cells. Here we describe two mouse strains called MITRG and MISTRG, in which human versions of four genes encoding cytokines important for innate immune cell development are knocked into their respective mouse loci. The human cytokines support the development and function of monocytes, macrophages and NK cells derived from human fetal liver or adult CD34(+) progenitor cells injected into the mice. Human macrophages infiltrated a human tumor xenograft in MITRG and MISTRG mice in a manner resembling that observed in tumors obtained from human patients. This humanized mouse model may be used to model the human immune system in scenarios of health and pathology, and may enable evaluation of therapeutic candidates in an in vivo setting relevant to human physiology.

Development and function of human innate immune cells in a humanized mouse model

PubMed Central

Rongvaux, Anthony; Willinger, Tim; Martinek, Jan; Strowig, Till; Gearty, Sofia V.; Teichmann, Lino L.; Saito, Yasuyuki; Marches, Florentina; Halene, Stephanie; Palucka, A. Karolina; Manz, Markus G.; Flavell, Richard A.

2014-01-01

Mice repopulated with human hematopoietic cells are a powerful tool for the study of human hematopoiesis and immune function in vivo. However, existing humanized mouse models are unable to support development of human innate immune cells, including myeloid cells and NK cells. Here we describe a mouse strain, called MI(S)TRG, in which human versions of four genes encoding cytokines important for innate immune cell development are knocked in to their respective mouse loci. The human cytokines support the development and function of monocytes/macrophages and natural killer cells derived from human fetal liver or adult CD34+ progenitor cells injected into the mice. Human macrophages infiltrated a human tumor xenograft in MI(S)TRG mice in a manner resembling that observed in tumors obtained from human patients. This humanized mouse model may be used to model the human immune system in scenarios of health and pathology, and may enable evaluation of therapeutic candidates in an in vivo setting relevant to human physiology. PMID:24633240

Effect of electroacupuncture on brain-derived neurotrophic factor mRNA expression in mouse hippocampus following cerebral ischemia-reperfusion injury.

PubMed

Zhao, Jianxin; Xu, Huazhou; Tian, Yuanxiang; Hu, Manxiang; Xiao, Hongling

2013-04-01

This work aims to observe the effects of electroacupuncture on brain-derived neurotrophic factor (BDNF) mRNA expression in mouse hippocampus following cerebral ischemia-reperfusion injury. The models of mouse cerebral ischemia-reperfusion injury were established. A total of 96 healthy mice were randomly assigned into 4 groups, namely, the sham surgery, model, model + electroacupuncture, and mode + hydergine groups. Mice in the model + electroacupuncture group were treated through electroacupuncture at the Shenshu (BL 23), Geshu (BL 17), and Baihui (GV 20) acupoints. Mice in the model+hydergine group were intragastrically administered with hydergine (0.77 mg/kg(-1) x day(-1)). The levels of BDNF mRNA expressions in the hippocampus were ana lyzed through a semi-quantitative reverse transcription-polymerase chain reaction assay on days 1 and 7 after the surgeries. BDNF mRNA expressions in the mouse hippocampus of the model group on days 1 and 7 after the surgery were higher than those of the sham surgery group (both P < 0.01). On days 1 and 7 of the electroacupuncture treatment, BDNF mRNA expression in the mouse hippocampus of the model + electroacupuncture group was significantly elevated compared with the model group (both P < 0.01) or the model + hydergine group (both P < 0.01). On days 1 and 7 of the hydergine treatment, BDNF mRNA expression in the mouse hippocampus of the model + hydergine group tended to increase compared with the model group; however, statistical significance was not achieved (both P > 0.05). Electroacupuncture treatment enhances endogenous BDNF expression, which may improve the survival environment for intracerebral neurons and inhibit the apoptosis of hippocampal cells.

Intramyocardial Injection of siRNAs Can Efficiently Establish Myocardial Tissue-Specific Renalase Knockdown Mouse Model.

PubMed

Huang, Kun; Liu, Ju; Zhang, Hui; Wang, Jiliang; Li, Huili

2016-01-01

Ischaemia/reperfusion (I/R) injury will cause additional death of cardiomyocytes in ischaemic heart disease. Recent studies revealed that renalase was involved in the I/R injury. So, the myocardial tissue-specific knockdown mouse models were needed for the investigations of renalase. To establish the mouse models, intramyocardial injection of siRNAs targeting renalase was performed in mice. The wild distribution and high transfection efficiency of the siRNAs were approved. And the renalase expression was efficiently suppressed in myocardial tissue. Compared with the high cost, time consumption, and genetic compensation risk of the Cre/loxP technology, RNA interference (RNAi) technology is much cheaper and less time-consuming. Among the RNAi technologies, injection of siRNAs is safer than virus. And considering the properties of the I/R injury mouse models, the efficiency and durability of injection with siRNAs are acceptable for the studies. Altogether, intramyocardial injection of siRNAs targeting renalase is an economical, safe, and efficient method to establish myocardial tissue-specific renalase knockdown mouse models.

Improvements and Limitations of Humanized Mouse Models for HIV Research: NIH/NIAID "Meet the Experts" 2015 Workshop Summary.

PubMed

Akkina, Ramesh; Allam, Atef; Balazs, Alejandro B; Blankson, Joel N; Burnett, John C; Casares, Sofia; Garcia, J Victor; Hasenkrug, Kim J; Kashanchi, Fatah; Kitchen, Scott G; Klein, Florian; Kumar, Priti; Luster, Andrew D; Poluektova, Larisa Y; Rao, Mangala; Sanders-Beer, Brigitte E; Shultz, Leonard D; Zack, Jerome A

2016-02-01

The number of humanized mouse models for the human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) and other infectious diseases has expanded rapidly over the past 8 years. Highly immunodeficient mouse strains, such as NOD/SCID/gamma chain(null) (NSG, NOG), support better human hematopoietic cell engraftment. Another improvement is the derivation of highly immunodeficient mice, transgenic with human leukocyte antigens (HLAs) and cytokines that supported development of HLA-restricted human T cells and heightened human myeloid cell engraftment. Humanized mice are also used to study the HIV reservoir using new imaging techniques. Despite these advances, there are still limitations in HIV immune responses and deficits in lymphoid structures in these models in addition to xenogeneic graft-versus-host responses. To understand and disseminate the improvements and limitations of humanized mouse models to the scientific community, the NIH sponsored and convened a meeting on April 15, 2015 to discuss the state of knowledge concerning these questions and best practices for selecting a humanized mouse model for a particular scientific investigation. This report summarizes the findings of the NIH meeting.

Oral recombinant human or mouse lactoferrin reduces Mycobacterium tuberculosis TDM induced granulomatous lung pathology.

PubMed

Hwang, Shen-An; Kruzel, Marian L; Actor, Jeffrey K

2017-02-01

Trehalose 6'6-dimycolate (TDM) is the most abundant glycolipid on the cell wall of Mycobacterium tuberculosis (MTB). TDM is capable of inducing granulomatous pathology in mouse models that resembles those induced by MTB infection. Using the acute TDM model, this work investigates the effect of recombinant human and mouse lactoferrin to reduce granulomatous pathology. C57BL/6 mice were injected intravenously with TDM at a dose of 25 μg·mouse -1 . At day 4 and 6, recombinant human or mouse lactoferrin (1 mg·(100 μL) -1 ·mouse -1 ) were delivered by gavage. At day 7 after TDM injection, mice were evaluated for lung pathology, cytokine production, and leukocyte populations. Mice given human or mouse lactoferrin had reduced production of IL-12p40 in their lungs. Mouse lactoferrin increased IL-6 and KC (CXCL1) in lung tissue. Increased numbers of macrophages were observed in TDM-injected mice given human or mouse lactoferrin. Granulomatous pathology, composed of mainly migrated leukocytes, was visually reduced in mice that received human or mouse lactoferrin. Quantitation of granulomatous pathology demonstrated a significant decrease in mice given human or mouse lactoferrin compared with TDM control mice. This report is the first to directly compare the immune modulatory effects of both heterologous recombinant human and homologous mouse lactoferrin on the development of TDM-induced granulomas.

Mouse neuroblastoma cell-based model and the effect of epileptic events on calcium oscillations and neural spikes

NASA Astrophysics Data System (ADS)

Kim, Suhwan; Jung, Unsang; Baek, Juyoung; Lee, Sangwon; Jung, Woonggyu; Kim, Jeehyun; Kang, Shinwon

2013-01-01

Recently, mouse neuroblastoma cells have been considered as an attractive model for the study of human neurological and prion diseases, and they have been intensively used as a model system in different areas. For example, the differentiation of neuro2a (N2A) cells, receptor-mediated ion current, and glutamate-induced physiological responses have been actively investigated with these cells. These mouse neuroblastoma N2A cells are of interest because they grow faster than other cells of neural origin and have a number of other advantages. The calcium oscillations and neural spikes of mouse neuroblastoma N2A cells in epileptic conditions are evaluated. Based on our observations of neural spikes in these cells with our proposed imaging modality, we reported that they can be an important model in epileptic activity studies. We concluded that mouse neuroblastoma N2A cells produce epileptic spikes in vitro in the same way as those produced by neurons or astrocytes. This evidence suggests that increased levels of neurotransmitter release due to the enhancement of free calcium from 4-aminopyridine causes the mouse neuroblastoma N2A cells to produce epileptic spikes and calcium oscillations.

High-fertility phenotypes: two outbred mouse models exhibit substantially different molecular and physiological strategies warranting improved fertility.

PubMed

Langhammer, Martina; Michaelis, Marten; Hoeflich, Andreas; Sobczak, Alexander; Schoen, Jennifer; Weitzel, Joachim M

2014-01-01

Animal models are valuable tools in fertility research. Worldwide, there are more than 400 transgenic or knockout mouse models available showing a reproductive phenotype; almost all of them exhibit an infertile or at least subfertile phenotype. By contrast, animal models revealing an improved fertility phenotype are barely described. This article summarizes data on two outbred mouse models exhibiting a 'high-fertility' phenotype. These mouse lines were generated via selection over a time period of more than 40 years and 161 generations. During this selection period, the number of offspring per litter and the total birth weight of the entire litter nearly doubled. Concomitantly with the increased fertility phenotype, several endocrine parameters (e.g. serum testosterone concentrations in male animals), physiological parameters (e.g. body weight, accelerated puberty, and life expectancy), and behavioral parameters (e.g. behavior in an open field and endurance fitness on a treadmill) were altered. We demonstrate that the two independently bred high-fertility mouse lines warranted their improved fertility phenotype using different molecular and physiological strategies. The fertility lines display female- as well as male-specific characteristics. These genetically heterogeneous mouse models provide new insights into molecular and cellular mechanisms that enhance fertility. In view of decreasing fertility in men, these models will therefore be a precious information source for human reproductive medicine. Translated abstract A German translation of abstract is freely available at http://www.reproduction-online.org/content/147/4/427/suppl/DC1.

Ultrasonic vocalizations: a tool for behavioural phenotyping of mouse models of neurodevelopmental disorders

PubMed Central

Scattoni, Maria Luisa; Crawley, Jacqueline; Ricceri, Laura

2009-01-01

In neonatal mice ultrasonic vocalizations have been studied both as an early communicative behavior of the pup-mother dyad and as a sign of an aversive affective state. Adult mice of both sexes produce complex ultrasonic vocalization patterns in different experimental/social contexts. All these vocalizations are becoming an increasingly valuable assay for behavioral phenotyping throughout the mouse life-span and alterations of the ultrasound patterns have been reported in several mouse models of neurodevelopmental disorders. Here we also show that the modulation of vocalizations by maternal cues (maternal potentiation paradigm) – originally identified and investigated in rats - can be measured in C57Bl/6 mouse pups with appropriate modifications of the rat protocol and can likely be applied to mouse behavioral phenotyping. In addition we suggest that a detailed qualitative evaluation of neonatal calls together with analysis of adult mouse vocalization patterns in both sexes in social settings, may lead to a greater understanding of the communication value of vocalizations in mice. Importantly, both neonatal and adult USV altered patterns can be determined during the behavioural phenotyping of mouse models of human neurodevelopmental and neuropsychiatric disorders, starting from those in which deficits in communication are a primary symptom. PMID:18771687

A Comparison of Some Organizational Characteristics of the Mouse Central Retina and the Human Macula

PubMed Central

Hoo, Juyea; Yee, Claudine; Williams, David S.

2015-01-01

Mouse models have greatly assisted our understanding of retinal degenerations. However, the mouse retina does not have a macula, leading to the question of whether the mouse is a relevant model for macular degeneration. In the present study, a quantitative comparison between the organization of the central mouse retina and the human macula was made, focusing on some structural characteristics that have been suggested to be important in predisposing the macula to stresses leading to degeneration: photoreceptor density, phagocytic load on the RPE, and the relative thinness of Bruch’s membrane. Light and electron microscopy measurements from retinas of two strains of mice, together with published data on human retinas, were used for calculations and subsequent comparisons. As in the human retina, the central region of the mouse retina possesses a higher photoreceptor cell density and a thinner Bruch’s membrane than in the periphery; however, the magnitudes of these periphery to center gradients are larger in the human. Of potentially greater relevance is the actual photoreceptor cell density, which is much greater in the mouse central retina than in the human macula, underlying a higher phagocytic load for the mouse RPE. Moreover, at eccentricities that correspond to the peripheral half of the human macula, the rod to cone ratio is similar between mouse and human. Hence, with respect to photoreceptor density and phagocytic load of the RPE, the central mouse retina models at least the more peripheral part of the macula, where macular degeneration is often first evident. PMID:25923208

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ding, Ying; Adachi, Hiroaki, E-mail: hadachi-ns@umin.org; Department of Neurology, University of Occupational and Environmental Health School of Medicine, 1-1 Iseigaoka, Yahata-nishi-ku, Kitakyushu 807-8555

Spinal and bulbar muscular atrophy (SBMA) is an inherited motor neuron disease caused by the expansion of a polyglutamine (polyQ)-encoding tract within the androgen receptor (AR) gene. The pathologic features of SBMA are motor neuron loss in the spinal cord and brainstem and diffuse nuclear accumulation and nuclear inclusions of mutant AR in residual motor neurons and certain visceral organs. Hepatocyte growth factor (HGF) is a polypeptide growth factor which has neuroprotective properties. To investigate whether HGF overexpression can affect disease progression in a mouse model of SBMA, we crossed SBMA transgenic model mice expressing an AR gene with anmore » expanded CAG repeat with mice overexpressing HGF. Here, we report that high expression of HGF induces Akt phosphorylation and modestly ameliorated motor symptoms in an SBMA transgenic mouse model treated with or without castration. These findings suggest that HGF overexpression can provide a potential therapeutic avenue as a combination therapy with disease-modifying therapies in SBMA. - Highlights: • HGF overexpression ameliorates the motor phenotypes of the SBMA mouse model. • HGF overexpression induces Akt phosphorylation in the SBMA mouse model. • This is the first report of combination therapy in a mouse model of polyQ diseases.« less

Defining the optimal animal model for translational research using gene set enrichment analysis.

PubMed

Weidner, Christopher; Steinfath, Matthias; Opitz, Elisa; Oelgeschläger, Michael; Schönfelder, Gilbert

2016-08-01

The mouse is the main model organism used to study the functions of human genes because most biological processes in the mouse are highly conserved in humans. Recent reports that compared identical transcriptomic datasets of human inflammatory diseases with datasets from mouse models using traditional gene-to-gene comparison techniques resulted in contradictory conclusions regarding the relevance of animal models for translational research. To reduce susceptibility to biased interpretation, all genes of interest for the biological question under investigation should be considered. Thus, standardized approaches for systematic data analysis are needed. We analyzed the same datasets using gene set enrichment analysis focusing on pathways assigned to inflammatory processes in either humans or mice. The analyses revealed a moderate overlap between all human and mouse datasets, with average positive and negative predictive values of 48 and 57% significant correlations. Subgroups of the septic mouse models (i.e., Staphylococcus aureus injection) correlated very well with most human studies. These findings support the applicability of targeted strategies to identify the optimal animal model and protocol to improve the success of translational research. © 2016 The Authors. Published under the terms of the CC BY 4.0 license.

AAV-PHP.B-Mediated Global-Scale Expression in the Mouse Nervous System Enables GBA1 Gene Therapy for Wide Protection from Synucleinopathy.

PubMed

Morabito, Giuseppe; Giannelli, Serena G; Ordazzo, Gabriele; Bido, Simone; Castoldi, Valerio; Indrigo, Marzia; Cabassi, Tommaso; Cattaneo, Stefano; Luoni, Mirko; Cancellieri, Cinzia; Sessa, Alessandro; Bacigaluppi, Marco; Taverna, Stefano; Leocani, Letizia; Lanciego, José L; Broccoli, Vania

2017-12-06

The lack of technology for direct global-scale targeting of the adult mouse nervous system has hindered research on brain processing and dysfunctions. Currently, gene transfer is normally achieved by intraparenchymal viral injections, but these injections target a restricted brain area. Herein, we demonstrated that intravenous delivery of adeno-associated virus (AAV)-PHP.B viral particles permeated and diffused throughout the neural parenchyma, targeting both the central and the peripheral nervous system in a global pattern. We then established multiple procedures of viral transduction to control gene expression or inactivate gene function exclusively in the adult nervous system and assessed the underlying behavioral effects. Building on these results, we established an effective gene therapy strategy to counteract the widespread accumulation of α-synuclein deposits throughout the forebrain in a mouse model of synucleinopathy. Transduction of A53T-SCNA transgenic mice with AAV-PHP.B-GBA1 restored physiological levels of the enzyme, reduced α-synuclein pathology, and produced significant behavioral recovery. Finally, we provided evidence that AAV-PHP.B brain penetration does not lead to evident dysfunctions in blood-brain barrier integrity or permeability. Altogether, the AAV-PHP.B viral platform enables non-invasive, widespread, and long-lasting global neural expression of therapeutic genes, such as GBA1, providing an invaluable approach to treat neurodegenerative diseases with diffuse brain pathology such as synucleinopathies. Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

Birth of cloned mice from vaginal smear cells after somatic cell nuclear transfer.

PubMed

Kuwayama, Hiroki; Tanabe, Yoshiaki; Wakayama, Teruhiko; Kishigami, Satoshi

2017-05-01

Less invasive methods for donor cell collection will facilitate reproduction of wild animals using somatic-cell nuclear transfer. Stages of the estrous cycle in mice have long been studies using somatic cells that can be collected from vaginal walls using cotton tipped swabs in a relatively non-invasive manner. In this study, we examined the feasibility of these cells as sources of nuclei for somatic-cell cloning using nuclear transfer. Estrous cycles generally comprise proestrus, estrus, metestrus, and diestrus stages. In the present experiments, more than 60% of cells were nucleated in vaginal smears from all but the estrus stage. However, after somatic-cell nuclear transfer of cells from proestrus, metestrus, and diestrus stages, 66%, 50%, and 72% of cloned embryos developed to the morula/blastocyst, and cloned female mouse birth rates after embryo transfer were 1.5%, 0.3%, and 1%, respectively. These results show that noninvasively collected vaginal smears contain somatic cells that can be used to clone female mice. Copyright © 2017. Published by Elsevier Inc.

Gene Transfer by Guanidinium-Cholesterol Cationic Lipids into Airway Epithelial Cells in vitro and in vivo

NASA Astrophysics Data System (ADS)

Oudrhiri, Noufissa; Vigneron, Jean-Pierre; Peuchmaur, Michel; Leclerc, Tony; Lehn, Jean-Marie; Lehn, Pierre

1997-03-01

Synthetic vectors represent an attractive alternative approach to viral vectors for gene transfer, in particular into airway epithelial cells for lung-directed gene therapy for cystic fibrosis. Having recently found that guanidinium-cholesterol cationic lipids are efficient reagents for gene transfer into mammalian cell lines in vitro, we have investigated their use for gene delivery into primary airway epithelial cells in vitro and in vivo. The results obtained indicate that the lipid bis (guanidinium)-tren-cholesterol (BGTC) can be used to transfer a reporter gene into primary human airway epithelial cells in culture. Furthermore, liposomes composed of BGTC and dioleoyl phosphatidylethanolamine (DOPE) are efficient for gene delivery to the mouse airway epithelium in vivo. Transfected cells were detected both in the surface epithelium and in submucosal glands. In addition, the transfection efficiency of BGTC/DOPE liposomes in vivo was quantitatively assessed by using the luciferase reporter gene system.

A Mouse Model to Investigate Postmenopausal Biology as an Etiology of Ovarian Cancer Risk

DTIC Science & Technology

2006-11-01

Wv mice and genetic alterations such as p53, pten, or p27kip1, which are found in human ovarian cancer. 2. Body: Research Progress In the first year...press (Yang et al., Am. J. Pathology 2007). To collaborate with the mouse model study, we have also examined human ovaries obtained from prophylactic...results in the coming years. Xu, Xiangxi, Ph.D. 8 3. Key Research Accomplishments (1) Further verify the relevance of the Wv mouse model to human

The Oncogenic Role of RhoGAPs in Basal-Like Breast Cancer

DTIC Science & Technology

2015-02-01

cell lines, and mouse models . c) In vivo tumorigenesis and metastasis assays. Milestones: Identify whether ArhGAP11A and RacGAP1 can promote tumor growth...also upregulated in basal (C3(I)-Tag) but not luminal (MMTV-Neu) genetically- engineered mouse models (Fig. 1B). At the protein level, RacGAP1 was...hypothesis that these RhoGAPs are indeed playing an oncogenic role in these cells. Human Tumors Mouse Model Tumors Normal Luminal A Basal-like Normal

A Physiologically Based Kinetic Model of Rat and Mouse Gestation: Disposition of a Weak Acid

EPA Science Inventory

A physiologically based toxicokinetic model of gestation in the rat mouse has been developed. The model is superimposed on the normal growth curve for nonpregnant females. It describes the entire gestation period including organogenesis. The model consists of uterus, mammary tiss...

Priceless GEMMs: genetically engineered mouse models for colorectal cancer drug development.

PubMed

Roper, Jatin; Hung, Kenneth E

2012-08-01

To establish effective drug development for colorectal cancer (CRC), preclinical models that are robust surrogates for human disease are crucial. Mouse models are an attractive platform because of their relatively low cost, short life span, and ease of use. There are two main categories of mouse CRC models: xenografts derived from implantation of CRC cells or tumors in immunodeficient mice; and genetically engineered mouse models (GEMMs) derived from modification of human cancer predisposition genes, resulting in spontaneous tumor formation. Here, we review xenografts and GEMMs and focus on their potential application in translational research. Furthermore, we describe newer GEMMs for sporadic CRC that are particularly suitable for drug testing. Finally, we discuss recent advances in small-animal imaging, such as optical colonoscopy, which allow in vivo assessment of tumors. With the increasing sophistication of GEMMs, our preclinical armamentarium provides new hope for the ongoing war against CRC. Copyright © 2012. Published by Elsevier Ltd.

A Genetically Engineered Mouse Model of Sporadic Colorectal Cancer.

PubMed

Betzler, Alexander M; Kochall, Susan; Blickensdörfer, Linda; Garcia, Sebastian A; Thepkaysone, May-Linn; Nanduri, Lahiri K; Muders, Michael H; Weitz, Jürgen; Reissfelder, Christoph; Schölch, Sebastian

2017-07-06

Despite the advantages of easy applicability and cost-effectiveness, colorectal cancer mouse models based on tumor cell injection have severe limitations and do not accurately simulate tumor biology and tumor cell dissemination. Genetically engineered mouse models have been introduced to overcome these limitations; however, such models are technically demanding, especially in large organs such as the colon in which only a single tumor is desired. As a result, an immunocompetent, genetically engineered mouse model of colorectal cancer was developed which develops highly uniform tumors and can be used for tumor biology studies as well as therapeutic trials. Tumor development is initiated by surgical, segmental infection of the distal colon with adeno-cre virus in compound conditionally mutant mice. The tumors can be easily detected and monitored via colonoscopy. We here describe the surgical technique of segmental adeno-cre infection of the colon, the surveillance of the tumor via high-resolution colonoscopy and present the resulting colorectal tumors.

Absence of Prenatal Forebrain Defects in the Dp(16)1Yey/+ Mouse Model of Down Syndrome

PubMed Central

Goodliffe, Joseph W.; Olmos-Serrano, Jose Luis; Aziz, Nadine M.; Pennings, Jeroen L.A.; Guedj, Faycal; Bianchi, Diana W.

2016-01-01

Studies in humans with Down syndrome (DS) show that alterations in fetal brain development are followed by postnatal deficits in neuronal numbers, synaptic plasticity, and cognitive and motor function. This same progression is replicated in several mouse models of DS. Dp(16)1Yey/+ (hereafter called Dp16) is a recently developed mouse model of DS in which the entire region of mouse chromosome 16 that is homologous to human chromosome 21 has been triplicated. As such, Dp16 mice may more closely reproduce neurodevelopmental changes occurring in humans with DS. Here, we present the first comprehensive cellular and behavioral study of the Dp16 forebrain from embryonic to adult stages. Unexpectedly, our results demonstrate that Dp16 mice do not have prenatal brain defects previously reported in human fetal neocortex and in the developing forebrains of other mouse models, including microcephaly, reduced neurogenesis, and abnormal cell proliferation. Nevertheless, we found impairments in postnatal developmental milestones, fewer inhibitory forebrain neurons, and deficits in motor and cognitive performance in Dp16 mice. Therefore, although this new model does not express prenatal morphological phenotypes associated with DS, abnormalities in the postnatal period appear sufficient to produce significant cognitive deficits in Dp16. SIGNIFICANCE STATEMENT Down syndrome (DS) leads to intellectual disability. Several mouse models have increased our understanding of the neuropathology of DS and are currently being used to test therapeutic strategies. A new mouse model that contains an expanded number of DS-related genes, known as Dp(16)1Yey/+ (Dp16), has been generated recently. We sought to determine whether the extended triplication creates a better phenocopy of DS-related brain pathologies. We measured embryonic development, forebrain maturation, and perinatal/adult behavior and revealed an absence of prenatal phenotypes in Dp16 fetal brain, but specific cellular and behavioral deficits after the first 2 postnatal weeks. These results uncover important differences in prenatal phenotype between Dp16 animals and humans with DS and other DS mouse models. PMID:26961948

Neurobehavioral characterization of APP23 transgenic mice with the SHIRPA primary screen.

PubMed

Lalonde, R; Dumont, M; Staufenbiel, M; Strazielle, C

2005-02-10

The SHIRPA primary screen comprises 40 measures covering various reflexes and basic sensorimotor functions. This multi-test battery was used to compare non-transgenic controls with APP23 transgenic mice, expressing the 751 isoform of human beta-amyloid precursor protein and characterized by amyloid deposits in parenchyma and vessel walls. The APP23 mice were distinguishable from controls by pathological limb reflexes, myoclonic jumping, seizure activity, and tail malformation. In addition, this mouse model of Alzheimer's disease was also marked by a crooked swimming trajectory. APP23 mice were also of lighter weight and were less inclined to stay immobile during a transfer arousal test. Despite the neurologic signs, APP23 transgenic mice were not deficient in stationary beam, coat-hanger, and rotorod tests, indicating intact motor coordination abilities.

Use of a highly sensitive strand-specific quantitative PCR to identify abortive replication in the mouse model of respiratory syncytial virus disease

PubMed Central

2010-01-01

Background The BALB/c mouse is commonly used to study RSV infection and disease. However, despite the many advantages of this well-characterised model, the inoculum is large, viral replication is restricted and only a very small amount of virus can be recovered from infected animals. A key question in this model is the fate of the administered virus. Is replication really being measured or is the model measuring the survival of the virus over time? To answer these questions we developed a highly sensitive strand-specific quantitative PCR (QPCR) able to accurately quantify the amount of RSV replication in the BALB/c mouse lung, allowing characterisation of RSV negative and positive strand RNA dynamics. Results In the mouse lung, no increase in RSV genome was seen above the background of the original inoculum whilst only a limited transient increase (< 1 log) in positive strand, replicative intermediate (RI) RNA occurred. This RNA did however persist at detectable levels for 59 days post infection. As expected, ribavirin therapy reduced levels of infectious virus and RI RNA in the mouse lung. However, whilst Palivizumab therapy was also able to reduce levels of infectious virus, it failed to prevent production of intracellular RI RNA. A comparison of RSV RNA kinetics in human (A549) and mouse (KLN205) cell lines demonstrated that RSV replication was also severely delayed and impaired in vitro in the mouse cells. Conclusions This is the first time that such a sensitive strand-specific QPCR technique has been to the RSV mouse system. We have accurately quantified the restricted and abortive nature of RSV replication in the mouse. Further in vitro studies in human and mouse cells suggest this restricted replication is due at least in part to species-specific host cell-viral interactions. PMID:20860795

Establishment of mouse neuron and microglial cell co-cultured models and its action mechanism.

PubMed

Zhang, Bo; Yang, Yunfeng; Tang, Jun; Tao, Yihao; Jiang, Bing; Chen, Zhi; Feng, Hua; Yang, Liming; Zhu, Gang

2017-06-27

The objective of this study is to establish a co-culture model of mouse neurons and microglial cells, and to analyze the mechanism of action of oxygen glucose deprivation (OGD) and transient oxygen glucose deprivation (tOGD) preconditioning cell models. Mouse primary neurons and BV2 microglial cells were successfully cultured, and the OGD and tOGD models were also established. In the co-culture of mouse primary neurons and microglial cells, the cell number of tOGD mouse neurons and microglial cells was larger than the OGD cell number, observed by a microscope. CCK-8 assay result showed that at 1h after treatment, the OD value in the control group is lower compared to all the other three groups (P < 0.05). The treatment group exhibited the highest OD value among the four groups. The results observed at 5h were consistent with the results at 1 h. Flow cytometry results showed that at 1h after treatment the apoptosis percentages is higher in the control group compared to other three groups (P < 0.05). Mouse brain tissues were collected and primary neurons cells were cultured. In the meantime mouse BV2 microglia cells were cultured. Two types of cells were co-cultured, and OGD and tOGD cell models were established. There were four groups in the experiment: control group (OGD), treatment group (tOGD+OGD), placebo group (tOGD+OGD+saline) and minocycline intervention group (tOGD+OGD+minocycline). CCK-8 kit was used to detect cell viability and flow cytometry was used to detect apoptosis. In this study, mouse primary neurons and microglial cells were co-cultured. The OGD and tOGD models were established successfully. tOGD was able to effectively protect neurons and microglial cells from damage, and inhibit the apoptosis caused by oxygen glucose deprivation.

Inhibition of nuclear factor of activated T cells (NFAT) c3 activation attenuates acute lung injury and pulmonary edema in murine models of sepsis

PubMed Central

Karpurapu, Manjula; Lee, Yong Gyu; Qian, Ziqing; Wen, Jin; Ballinger, Megan N.; Rusu, Luiza; Chung, Sangwoon; Deng, Jing; Qian, Feng; Reader, Brenda F.; Nirujogi, Teja Srinivas; Park, Gye Young; Pei, Dehua; Christman, John W.

2018-01-01

Specific therapies targeting cellular and molecular events of sepsis induced Acute Lung Injury (ALI) pathogenesis are lacking. We have reported a pivotal role for Nuclear Factors of Activated T cells (NFATc3) in regulating macrophage phenotype during sepsis induced ALI and subsequent studies demonstrate that NFATc3 transcriptionally regulates macrophage CCR2 and TNFα gene expression. Mouse pulmonary microvascular endothelial cell monolayer maintained a tighter barrier function when co-cultured with LPS stimulated NFATc3 deficient macrophages whereas wild type macrophages caused leaky monolayer barrier. More importantly, NFATc3 deficient mice showed decreased neutrophilic lung inflammation, improved alveolar capillary barrier function, arterial oxygen saturation and survival benefit in lethal CLP sepsis mouse models. In addition, survival of wild type mice subjected to the lethal CLP sepsis was not improved with broad-spectrum antibiotics, whereas the survival of NFATc3 deficient mice was improved to 40–60% when treated with imipenem. Passive adoptive transfer of NFATc3 deficient macrophages conferred protection against LPS induced ALI in wild type mice. Furthermore, CP9-ZIZIT, a highly potent, cell-permeable peptide inhibitor of Calcineurin inhibited NFATc3 activation. CP9-ZIZIT effectively reduced sepsis induced inflammatory cytokines and pulmonary edema in mice. Thus, this study demonstrates that inhibition of NFATc3 activation by CP9-ZIZIT provides a potential therapeutic option for attenuating sepsis induced ALI/pulmonary edema. PMID:29535830

Modulation of GSK-3 provides cellular and functional neuroprotection in the rd10 mouse model of retinitis pigmentosa.

PubMed

Sánchez-Cruz, Alonso; Villarejo-Zori, Beatriz; Marchena, Miguel; Zaldivar-Díez, Josefa; Palomo, Valle; Gil, Carmen; Lizasoain, Ignacio; de la Villa, Pedro; Martínez, Ana; de la Rosa, Enrique J; Hernández-Sánchez, Catalina

2018-04-16

Retinitis pigmentosa (RP) is a group of hereditary retinal neurodegenerative conditions characterized by primary dysfunction and death of photoreceptor cells, resulting in visual loss and, eventually, blindness. To date, no effective therapies have been transferred to clinic. Given the diverse genetic etiology of RP, targeting common cellular and molecular retinal alterations has emerged as a potential therapeutic strategy. Using the Pde6b rd10/rd10 mouse model of RP, we investigated the effects of daily intraperitoneal administration of VP3.15, a small-molecule heterocyclic GSK-3 inhibitor. Gene expression was analyzed by quantitative PCR and protein expression and phosphorylation by Western blot. Photoreceptor preservation was evaluated by histological analysis and visual function was assessed by electroretinography. In rd10 retinas, increased expression of pro-inflammatory markers and reactive gliosis coincided with the early stages of retinal degeneration. Compared with wild-type controls, GSK-3β expression (mRNA and protein) remained unchanged during the retinal degeneration period. However, levels of GSK-3β Ser9 and its regulator Akt Ser473 were increased in rd10 versus wild-type retinas. In vivo administration of VP3.15 reduced photoreceptor cell loss and preserved visual function. This neuroprotective effect was accompanied by a decrease in the expression of neuroinflammatory markers. These results provide proof of concept of the therapeutic potential of VP3.15 for the treatment of retinal neurodegenerative conditions in general, and RP in particular.

Arginase-1 deficiency.

PubMed

Sin, Yuan Yan; Baron, Garrett; Schulze, Andreas; Funk, Colin D

2015-12-01

Arginase-1 (ARG1) deficiency is a rare autosomal recessive disorder that affects the liver-based urea cycle, leading to impaired ureagenesis. This genetic disorder is caused by 40+ mutations found fairly uniformly spread throughout the ARG1 gene, resulting in partial or complete loss of enzyme function, which catalyzes the hydrolysis of arginine to ornithine and urea. ARG1-deficient patients exhibit hyperargininemia with spastic paraparesis, progressive neurological and intellectual impairment, persistent growth retardation, and infrequent episodes of hyperammonemia, a clinical pattern that differs strikingly from other urea cycle disorders. This review briefly highlights the current understanding of the etiology and pathophysiology of ARG1 deficiency derived from clinical case reports and therapeutic strategies stretching over several decades and reports on several exciting new developments regarding the pathophysiology of the disorder using ARG1 global and inducible knockout mouse models. Gene transfer studies in these mice are revealing potential therapeutic options that can be exploited in the future. However, caution is advised in extrapolating results since the lethal disease phenotype in mice is much more severe than in humans indicating that the mouse models may not precisely recapitulate human disease etiology. Finally, some of the functions and implications of ARG1 in non-urea cycle activities are considered. Lingering questions and future areas to be addressed relating to the clinical manifestations of ARG1 deficiency in liver and brain are also presented. Hopefully, this review will spark invigorated research efforts that lead to treatments with better clinical outcomes.

Overexpression of Heparin-Binding Epidermal Growth Factor-Like Growth Factor Mediates Liver Fibrosis in Transgenic Mice.

PubMed

Guo, Yongze; Ding, Qian; Chen, Lei; Ji, Chenguang; Hao, Huiyao; Wang, Jia; Qi, Wei; Xie, Xiaoli; Ma, Junji; Li, Aidi; Jiang, Xiaoyu; Li, Xiaotian; Jiang, Huiqing

2017-08-01

The role of heparin-binding epidermal growth factor-like growth factor (HB-EGF) in liver fibrosis is not clear and is sometimes even contradictory. To clarify this role, a HB-EGF transgenic (Tg) mouse model was, for the first time, used to evaluate the functions of HB-EGF in liver fibrosis. For the in vivo study, carbon tetrachloride injection and bile duct ligation treatment were used to induce liver fibrosis in HB-EGF Tg mice and wild-type (WT) mice, respectively. Primary hepatic satellite cells (HSCs) were isolated from HB-EGF Tg and WT mice for the in vitro study. Compared with the WT mice, HB-EGF Tg mice were shown to develop more severe liver fibrosis when treated with carbon tetrachloride or bile duct ligation, with increased matrix metalloproteinases 13 activity and enhanced expression of fibrogenic genes including α-smooth muscle actin and collagen I. HB-EGF gene transfer led to an increase in proliferation and a decrease in apoptosis in primary HSCs. The ERK signaling pathway was more highly activated in primary HSCs from HB-EGF Tg mice than in those from WT mice. Our investigation confirmed the profibrotic effect of HB-EGF on the liver using a Tg mouse model. This result may contribute to the elucidation of HB-EGF as a therapeutic target in liver fibrosis. Copyright © 2017 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.

Escherichia coli msbB gene as a virulence factor and a therapeutic target.

PubMed

Somerville, J E; Cassiano, L; Darveau, R P

1999-12-01

A mutation in the msbB gene of Escherichia coli results in the synthesis of E. coli lipopolysaccharide (LPS) that lacks the myristic acid moiety of lipid A. Although such mutant E. coli cells and their purified LPS have a greatly reduced ability to stimulate human immune cells, a minor reduction in the mouse inflammatory response is observed. When the msbB mutation is transferred into a clinical isolate of E. coli, there is a significant loss in virulence, as assessed by lethality in BALB/c mice. When a cloned msbB gene is provided to functionally complement the msbB mutant, virulence returns, providing direct evidence that the msbB gene product is an important virulence factor in a murine model of E. coli pathogenicity. In the genetic background of the clinical E. coli isolate, the msbB mutation also results in filamentation of the cells at 37 degrees C but not at 30 degrees C, a reduction in the level of the K1 capsule, an increase in the level of complement C3 deposition, and an increase in both opsonic and nonopsonic phagocytosis of the msbB mutant, phenotypes that can help to explain the loss in virulence. The demonstration that the inhibition of msbB gene function reduces the virulence of E. coli in a mouse infection model warrants further investigation of the msbB gene product as a novel target for antibiotic therapy.

Actinic keratosis modelling in mice: A translational study

PubMed Central

Vandenberghe, Isabelle; Cartron, Valérie; Cèbe, Patrick; Blanchet, Jean-Christophe; Sibaud, Vincent; Guilbaud, Nicolas; Audoly, Laurent; Lamant, Laurence; Kruczynski, Anna

2017-01-01

Background Actinic keratoses (AK) are pre-malignant cutaneous lesions caused by prolonged exposure to ultraviolet radiation. As AKs lesions are generally accepted to be the initial lesions in a disease continuum that progresses to squamous cell carcinoma (SCC), AK lesions have to be treated. They are also the second most common reason for visits to the dermatologist. Several treatments are available but their efficacy still needs to be improved. The UV-B-induced KA lesion mouse model is used in preclinical studies to assess the efficacy of novel molecules, even though it is often more representative of advanced AK or SCC. Objectives Here we report on a translational study, comparing the various stages of AK development in humans and in the UV-B irradiated mouse model, as well as the optimization of photograph acquisition of AK lesions on mouse skin. Methods Human and mouse skin lesions were analysed by histology and immunohistochemistry. Mouse lesions were also assessed using a digital dermatoscope. Results An histological and phenotypic analysis, including p53, Ki67 and CD3 expression detection, performed on human and mouse AK lesions, shows that overall AK modelling in mice is relevant in the clinical situation. Some differences are observed, such as disorganization of keratinocytes of the basal layer and a number of atypical nuclei which are more numerous in human AK, whereas much more pronounced acanthosis is observed in skin lesion in mice. Thanks to this translational study, we are able to select appropriate experimental conditions for establishing either early or advanced stage AK or an SCC model. Furthermore, we optimized photograph acquisition of AK lesions on mouse skin by using a digital dermatoscope which is also used in clinics and allows reproducible photograph acquisition for further reliable assessment of mouse lesions. Use of this camera is illustrated through a pharmacological study assessing the activity of CARAC®. Conclusion These data demonstrate that this mouse model of UV-B-induced skin lesions is predictive for the identification of novel therapeutic treatments for both early and advanced stages of the disease. PMID:28662116

Taltirelin alleviates fatigue-like behavior in mouse models of cancer-related fatigue.

PubMed

Dougherty, John P; Wolff, Brian S; Cullen, Mary J; Saligan, Leorey N; Gershengorn, Marvin C

2017-10-01

Fatigue affects most cancer patients and has numerous potential causes, including cancer itself and cancer treatment. Cancer-related fatigue (CRF) is not relieved by rest, can decrease quality of life, and has no FDA-approved therapy. Thyrotropin-releasing hormone (TRH) has been proposed as a potential novel treatment for CRF, but its efficacy against CRF remains largely untested. Thus, we tested the TRH analog, taltirelin (TAL), in mouse models of CRF. To model fatigue, we used a mouse model of chemotherapy, a mouse model of radiation therapy, and mice bearing colon 26 carcinoma tumors. We used the treadmill fatigue test to assess fatigue-like behavior after treatment with TAL. Additionally, we used wild-type and TRH receptor knockout mice to determine which TRH receptor was necessary for the actions of TAL. Tumor-bearing mice displayed muscle wasting and all models caused fatigue-like behavior, with mice running a shorter distance in the treadmill fatigue test than controls. TAL reversed fatigue-like behavior in all three models and the mouse TRH 1 receptor was necessary for the effects of TAL. These data suggest that TAL may be useful in alleviating fatigue in all cancer patients and provide further support for evaluating TAL as a potential therapy for CRF in humans. Published by Elsevier Ltd.

Rhein Induces Oxidative Stress and Apoptosis in Mouse Blastocysts and Has Immunotoxic Effects during Embryonic Development.

PubMed

Huang, Chien-Hsun; Chan, Wen-Hsiung

2017-09-20

Rhein, a glucoside chemical compound found in a traditional Chinese medicine derived from the roots of rhubarb, induces cell apoptosis and is considered to have high potential as an antitumor drug. Several previous studies showed that rhein can inhibit cell proliferation and trigger mitochondria-related or endoplasmic reticulum (ER) stress-dependent apoptotic processes. However, the side effects of rhein on pre- and post-implantation embryonic development remain unclear. Here, we show that rhein has cytotoxic effects on blastocyst-stage mouse embryos and induces oxidative stress and immunotoxicity in mouse fetuses. Blastocysts incubated with 5-20 μM rhein showed significant cell apoptosis, as well as decreases in their inner cell mass cell numbers and total cell numbers. An in vitro development assay showed that rhein affected the developmental potentials of both pre- and post-implantation embryos. Incubation of blastocysts with 5-20 μM rhein was associated with increased resorption of post-implantation embryos and decreased fetal weight in an embryo transfer assay. Importantly, in an in vivo model, intravenous injection of dams with rhein (1, 3, and 5 mg/kg body weight/day) for four days resulted in apoptosis of blastocyst-stage embryos, early embryonic developmental injury, and decreased fetal weight. Intravenous injection of dams with 5 mg/kg body weight/day rhein significantly increased the total reactive oxygen species (ROS) content of fetuses and the transcription levels of antioxidant proteins in fetal livers. Additional work showed that rhein induced apoptosis through ROS generation, and that prevention of apoptotic processes effectively rescued the rhein-induced injury effects on embryonic development. Finally, the transcription levels of the innate-immunity related genes, CXCL1 , IL-1 β and IL-8 , were down-regulated in the fetuses of dams that received intravenous injections of rhein. These results collectively show that rhein has the potential to induce embryonic cytotoxicity and induce oxidative stress and immunotoxicity during the development of mouse embryos.

BAMOS: A recording application for BAsso MOuse scale of locomotion in experimental models of spinal cord injury.

PubMed

Gómez, Alberto; Nieto-Díaz, Manuel; Del Águila, Ángela; Arias, Enrique

2018-05-01

Transparency in science is increasingly a hot topic. Scientists are required to show not only results but also evidence of how they have achieved these results. In experimental studies of spinal cord injury, there are a number of standardized tests, such as the Basso-Beattie-Bresnahan locomotor rating scale for rats and Basso Mouse Scale for mice, which researchers use to study the pathophysiology of spinal cord injury and to evaluate the effects of experimental therapies. Although the standardized data from the Basso-Beattie-Bresnahan locomotor rating scale and the Basso Mouse Scale are particularly suited for storage and sharing in databases, systems of data acquisition and repositories are still lacking. To the best of our knowledge, both tests are usually conducted manually, with the data being recorded on a paper form, which may be documented with video recordings, before the data is transferred to a spreadsheet for analysis. The data thus obtained is used to compute global scores, which is the information that usually appears in publications, with a wealth of information being omitted. This information may be relevant to understand locomotion deficits or recovery, or even important aspects of the treatment effects. Therefore, this paper presents a mobile application to record and share Basso Mouse Scale tests, meeting the following criteria: i) user-friendly; ii) few hardware requirements (only a smartphone or tablet with a camera running under Android Operating System); and iii) based on open source software such as SQLite, XML, Java, Android Studio and Android SDK. The BAMOS app can be downloaded and installed from the Google Market repository and the app code is available at the GitHub repository. The BAMOS app demonstrates that mobile technology constitutes an opportunity to develop tools for aiding spinal cord injury scientists in recording and sharing experimental data. Copyright © 2018 Elsevier Ltd. All rights reserved.

MR images of mouse brain using clinical 3T MR scanner and 4CH-Mouse coil

NASA Astrophysics Data System (ADS)

Lim, Soo Mee; Park, Eun Mi; Lyoo, In Kyoon; Lee, Junghyun; Han, Bo Mi; Lee, Jeong Kyong; Lee, Su Bin

2015-07-01

Objectives: Although small-bore high-field magnets are useful for research in small rodent models,this technology, however, has not been easily accessible to most researchers. This current study, thus,tried to evaluate the usability of 4CH-Mouse coil (Philips Healthcare, Best, the Netherlands) forpreclinical investigations in clinical 3T MR scan environment. We evaluated the effects of ischemicpreconditioning (IP) in the mouse stroke model with clinical 3T MR scanner and 4CH-Mouse coil. Materials and Methods: Experiments were performed on male C57BL/6 mice that either received the IP or sham operation (control). Three different MR sequences including diffusion weighted images (DWI), T2-weighted images (T2WI), and fluid attenuated inversion recovery (FLAIR) were performed on the mouse brains following 24, 72 hours of middle cerebral artery occlusion (MCAO) and analyzed for infarct lesions. Results: The images showed that the IP-treated mouse brains had significantly smaller infarct volumes compared to the control group. Of the MR sequences employed, the T2WI showed the highest level of correlations with postmortem infarct volume measurements. Conclusions: The clinical 3T MR scanner turned out to have a solid potential as a practical tool for imaging small animal brains. MR sequences including DWI, T2WI, FLAIR were obtained with acceptable resolution and in a reasonable time constraint in evaluating a mouse stroke model brain.

Mutagenicity testing with transgenic mice. Part I: Comparison with the mouse bone marrow micronucleus test

PubMed Central

Wahnschaffe, U; Bitsch, A; Kielhorn, J; Mangelsdorf, I

2005-01-01

As part of a larger literature study on transgenic animals in mutagenicity testing, test results from the transgenic mutagenicity assays (lacI model; commercially available as the Big Blue® mouse, and the lacZ model; commercially available as the Muta™Mouse), were compared with the results on the same substances in the more traditional mouse bone marrow micronucleus test. 39 substances were found which had been tested in the micronucleus assay and in the above transgenic mouse systems. Although, the transgenic animal mutation assay is not directly comparable with the micronucleus test, because different genetic endpoints are examined: chromosome aberration versus gene mutation, the results for the majority of substances were in agreement. Both test systems, the transgenic mouse assay and the mouse bone marrow micronucleus test, have advantages and they complement each other. However, the transgenic animal assay has some distinct advantages over the micronucleus test: it is not restricted to one target organ and detects systemic as well as local mutagenic effects. PMID:15655069

In vivo characterization of a bigenic fluorescent mouse model of Alzheimer's disease with neurodegeneration.

PubMed

Crowe, Sarah E; Ellis-Davies, Graham C R

2013-07-01

The loss of cognitive function in Alzheimer's disease (AD) patients is strongly correlated with the loss of neurons in various regions of the brain. We have created a new fluorescent bigenic mouse model of AD by crossing "H-line" yellow fluorescent protein (YFP) mice with the 5xFAD mouse model, which we call the 5XY mouse model. The 5xFAD mouse has been shown to have significant loss of L5 pyramidal neurons by 12 months of age. These neurons are transgenically labeled with YFP in the 5XY mouse, which enable longitudinal imaging of structural changes. In the 5XY mice, we observed an appearance of axonal dystrophies, with two distinct morphologies in the early stages of the disease progression. Simple swelling dystrophies are transient in nature and are not directly associated with amyloid plaques. Rosette dystrophies are more complex structures that remained stable throughout all imaging sessions, and always surrounded an amyloid plaque. Plaque growth was followed over 4 weeks, and significant growth was seen between weekly imaging sessions. In addition to axonal dystrophy appearance and plaque growth, we were able to follow spine stability in 4-month old 5XY mice, which revealed no significant loss of spines. 5XY mice also showed a striking shrinkage of the neocortex at older ages (12-14 months). The 5XY mouse model may be a valuable tool for studying specific events in the degeneration of the neocortex, and may suggest new avenues for therapeutic intervention. Copyright © 2013 Wiley Periodicals, Inc.

Transfer of Anti-Rotavirus Antibodies during Pregnancy and in Milk Following Maternal Vaccination with a Herpes Simplex Virus Type-1 Amplicon Vector.

PubMed

Meier, Anita F; Suter, Mark; Schraner, Elisabeth M; Humbel, Bruno M; Tobler, Kurt; Ackermann, Mathias; Laimbacher, Andrea S

2017-02-16

Rotaviruses (RVs) are important enteric pathogens of newborn humans and animals, causing diarrhea and in rare cases death, especially in very young individuals. Rotavirus vaccines presently used are modified live vaccines that lack complete biological safety. Previous work from our laboratory suggested that vaccines based on in situ produced, non-infectious rotavirus-like particles (RVLPs) are efficient while being entirely safe. However, using either vaccine, active mucosal immunization cannot induce protective immunity in newborns due to their immature immune system. We therefore hypothesized that offspring from vaccinated dams are passively immunized either by transfer of maternal antibodies during pregnancy or by taking up antibodies from milk. Using a codon optimized polycistronic gene expression cassette packaged into herpesvirus particles, the simultaneous expression of the RV capsid genes led to the intracellular formation of RVLPs in various cell lines. Vaccinated dams developed a strong RV specific IgG antibody response determined in sera and milk of both mother and pups. Moreover, sera of naïve pups nursed by vaccinated dams also had RV specific antibodies suggesting a lactogenic transfer of antibodies. Although full protection of pups was not achieved in this mouse model, our observations are important for the development of improved vaccines against RV in humans as well as in various animal species.

Transfer of Anti-Rotavirus Antibodies during Pregnancy and in Milk Following Maternal Vaccination with a Herpes Simplex Virus Type-1 Amplicon Vector

PubMed Central

Meier, Anita F.; Suter, Mark; Schraner, Elisabeth M.; Humbel, Bruno M.; Tobler, Kurt; Ackermann, Mathias; Laimbacher, Andrea S.

2017-01-01

Rotaviruses (RVs) are important enteric pathogens of newborn humans and animals, causing diarrhea and in rare cases death, especially in very young individuals. Rotavirus vaccines presently used are modified live vaccines that lack complete biological safety. Previous work from our laboratory suggested that vaccines based on in situ produced, non-infectious rotavirus-like particles (RVLPs) are efficient while being entirely safe. However, using either vaccine, active mucosal immunization cannot induce protective immunity in newborns due to their immature immune system. We therefore hypothesized that offspring from vaccinated dams are passively immunized either by transfer of maternal antibodies during pregnancy or by taking up antibodies from milk. Using a codon optimized polycistronic gene expression cassette packaged into herpesvirus particles, the simultaneous expression of the RV capsid genes led to the intracellular formation of RVLPs in various cell lines. Vaccinated dams developed a strong RV specific IgG antibody response determined in sera and milk of both mother and pups. Moreover, sera of naïve pups nursed by vaccinated dams also had RV specific antibodies suggesting a lactogenic transfer of antibodies. Although full protection of pups was not achieved in this mouse model, our observations are important for the development of improved vaccines against RV in humans as well as in various animal species. PMID:28212334

Butyrylcholinesterase gene transfer in obese mice prevents postdieting body weight rebound by suppressing ghrelin signaling

PubMed Central

Chen, Vicky Ping; Gao, Yang; Geng, Liyi; Brimijoin, Stephen

2017-01-01

The worldwide prevalence of obesity is increasing at an alarming rate but treatment options remain limited. Despite initial success, weight loss by calorie restriction (CR) often fails because of rebound weight gain. Postdieting hyperphagia along with altered hypothalamic neuro-architecture appears to be one direct cause of this undesirable outcome. In response to calorie deficiency the circulating levels of the appetite-promoting hormone, acyl-ghrelin, rise sharply. We hypothesize that proper modulation of acyl-ghrelin and its receptor’s sensitivity will favorably impact energy intake and reprogram the body weight set point. Here we applied viral gene transfer of the acyl-ghrelin hydrolyzing enzyme, butyrylcholinesterase (BChE), in a mouse model of diet-induced obesity. Our results confirmed that BChE overexpression decreased circulating acyl-ghrelin levels, suppressed CR-provoked ghrelin signaling, and restored central ghrelin sensitivity. In addition to maintaining healthy body weights, BChE treated mice had modest postdieting food intake and showed normal glucose homeostasis. Spontaneous activity and energy expenditure did not differ significantly between treated and untreated mice after body weight rebound, suggesting that BChE gene transfer did not alter energy expenditure in the long term. These findings indicate that combining BChE treatment with CR could be an effective approach in treating human obesity and aiding lifelong weight management. PMID:28973869

Orexin Gene Transfer into Zona Incerta Neurons Suppresses Muscle Paralysis in Narcoleptic Mice

PubMed Central

Liu, Meng; Konadhode, RodaRani; Begum, Suraiya; Pelluru, Dheeraj; Gerashchenko, Dmitry; Sakurai, Takeshi; Yanagisawa, Masashi; van den Pol, Anthony N.

2011-01-01

Cataplexy, a sudden unexpected muscle paralysis, is a debilitating symptom of the neurodegenerative sleep disorder, narcolepsy. During these attacks, the person is paralyzed, but fully conscious and aware of their surroundings. To identify potential neurons that might serve as surrogate orexin neurons to suppress such attacks, the gene for orexin (hypocretin), a peptide lost in most human narcoleptics, was delivered into the brains of the orexin-ataxin-3 transgenic mouse model of human narcolepsy. Three weeks after the recombinant adenoassociated virus (rAAV)-mediated orexin gene transfer, sleep–wake behavior was assessed. rAAV-orexin gene delivery into neurons of the zona incerta (ZI), or the lateral hypothalamus (LH) blocked cataplexy. Orexin gene transfer into the striatum or in the melanin-concentrating hormone neurons in the ZI or LH had no such effect, indicating site specificity. In transgenic mice lacking orexin neurons but given rAAV-orexin, detectable levels of orexin-A were evident in the CSF, indicating release of the peptide from the surrogate neurons. Retrograde tracer studies showed that the amygdala innervates the ZI consistent with evidence that strong emotions trigger cataplexy. In turn, the ZI projects to the locus ceruleus, indicating that the ZI is part of a circuit that stabilizes motor tone. Our results indicate that these neurons might also be recruited to block the muscle paralysis in narcolepsy. PMID:21508228

Technique Selectively Represses Immune System

MedlinePlus

... from attacking myelin in a mouse model of multiple sclerosis. Dr David Furness, Wellcome Images. All rights reserved ... devised a way to successfully treat symptoms resembling multiple sclerosis in a mouse model. With further development, the ...

Lipopolysaccharide-induced endotoxemia in corn oil-preloaded mice causes an extended course of lung injury and repair and pulmonary fibrosis: A translational mouse model of acute respiratory distress syndrome.

PubMed

Wu, Chaomin; Evans, Colin E; Dai, Zhiyu; Huang, Xiaojia; Zhang, Xianming; Jin, Hua; Hu, Guochang; Song, Yuanlin; Zhao, You-Yang

2017-01-01

Acute respiratory distress syndrome (ARDS) is characterized by acute hypoxemia respiratory failure, bilateral pulmonary infiltrates, and pulmonary edema of non-cardiac origin. Effective treatments for ARDS patients may arise from experimental studies with translational mouse models of this disease that aim to delineate the mechanisms underlying the disease pathogenesis. Mouse models of ARDS, however, can be limited by their rapid progression from injured to recovery state, which is in contrast to the course of ARDS in humans. Furthermore, current mouse models of ARDS do not recapitulate certain prominent aspects of the pathogenesis of ARDS in humans. In this study, we developed an improved endotoxemic mouse model of ARDS resembling many features of clinical ARDS including extended courses of injury and recovery as well as development of fibrosis following i.p. injection of lipopolysaccharide (LPS) to corn oil-preloaded mice. Compared with mice receiving LPS alone, those receiving corn oil and LPS exhibited extended course of lung injury and repair that occurred over a period of >2 weeks instead of 3-5days. Importantly, LPS challenge of corn oil-preloaded mice resulted in pulmonary fibrosis during the repair phase as often seen in ARDS patients. In summary, this simple novel mouse model of ARDS could represent a valuable experimental tool to elucidate mechanisms that regulate lung injury and repair in ARDS patients.

Endometrial apoptosis and neutrophil infiltration during menstruation exhibits spatial and temporal dynamics that are recapitulated in a mouse model.

PubMed

Armstrong, Gregory M; Maybin, Jacqueline A; Murray, Alison A; Nicol, Moira; Walker, Catherine; Saunders, Philippa T K; Rossi, Adriano G; Critchley, Hilary O D

2017-12-12

Menstruation is characterised by synchronous shedding and restoration of tissue integrity. An in vivo model of menstruation is required to investigate mechanisms responsible for regulation of menstrual physiology and to investigate common pathologies such as heavy menstrual bleeding (HMB). We hypothesised that our mouse model of simulated menstruation would recapitulate the spatial and temporal changes in the inflammatory microenvironment of human menses. Three regulatory events were investigated: cell death (apoptosis), neutrophil influx and cytokine/chemokine expression. Well-characterised endometrial tissues from women were compared with uteri from a mouse model (tissue recovered 0, 4, 8, 24 and 48 h after removal of a progesterone-secreting pellet). Immunohistochemistry for cleaved caspase-3 (CC3) revealed significantly increased staining in human endometrium from late secretory and menstrual phases. In mice, CC3 was significantly increased at 8 and 24 h post-progesterone-withdrawal. Elastase + human neutrophils were maximal during menstruation; Ly6G + mouse neutrophils were maximal at 24 h. Human endometrial and mouse uterine cytokine/chemokine mRNA concentrations were significantly increased during menstrual phase and 24 h post-progesterone-withdrawal respectively. Data from dated human samples revealed time-dependent changes in endometrial apoptosis preceding neutrophil influx and cytokine/chemokine induction during active menstruation. These dynamic changes were recapitulated in the mouse model of menstruation, validating its use in menstrual research.

Development of an Autologous Macrophage-Based Adoptive Gene Transfer Strategy to Treat Posttraumatic Osteoarthritis (PTOA) and Osteoarithritis (OA)

DTIC Science & Technology

2014-09-01

mouse. Clear evidence for intra- articular fractures, existence of substantial subchondral bone erosion at the surface of articular plate, and formation...of bone spurs (small growths called osteophytes ) on the edges is seen in the PTOA joint but not on the intact contralateral knee joint. This

The STR/ort mouse model of spontaneous osteoarthritis - an update.

PubMed

Staines, K A; Poulet, B; Wentworth, D N; Pitsillides, A A

2017-06-01

Osteoarthritis is a degenerative joint disease and a world-wide healthcare burden. Characterized by cartilage degradation, subchondral bone thickening and osteophyte formation, osteoarthritis inflicts much pain and suffering, for which there are currently no disease-modifying treatments available. Mouse models of osteoarthritis are proving critical in advancing our understanding of the underpinning molecular mechanisms. The STR/ort mouse is a well-recognized model which develops a natural form of osteoarthritis very similar to the human disease. In this Review we discuss the use of the STR/ort mouse in understanding this multifactorial disease with an emphasis on recent advances in its genetics and its bone, endochondral and immune phenotypes. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Mouse brain magnetic resonance microscopy: Applications in Alzheimer disease.

PubMed

Lin, Lan; Fu, Zhenrong; Xu, Xiaoting; Wu, Shuicai

2015-05-01

Over the past two decades, various Alzheimer's disease (AD) trangenetic mice models harboring genes with mutation known to cause familial AD have been created. Today, high-resolution magnetic resonance microscopy (MRM) technology is being widely used in the study of AD mouse models. It has greatly facilitated and advanced our knowledge of AD. In this review, most of the attention is paid to fundamental of MRM, the construction of standard mouse MRM brain template and atlas, the detection of amyloid plaques, following up on brain atrophy and the future applications of MRM in transgenic AD mice. It is believed that future testing of potential drugs in mouse models with MRM will greatly improve the predictability of drug effect in preclinical trials. © 2015 Wiley Periodicals, Inc.

Successful mouse cloning of an outbred strain by trichostatin A treatment after somatic nuclear transfer.

PubMed

Kishigami, Satoshi; Bui, Hong-Thuy; Wakayama, Sayaka; Tokunaga, Kenzo; Van Thuan, Nguyen; Hikichi, Takafusa; Mizutani, Eiji; Ohta, Hiroshi; Suetsugu, Rinako; Sata, Tetsutaro; Wakayama, Teruhiko

2007-02-01

Although the somatic cloning technique has been used for numerous applications and basic research of reprogramming in various species, extremely low success rates have plagued this technique for a decade. Further in mice, the "clonable" strains have been limited to mainly hybrid F1 strains such as B6D2F1. Recently, we established a new efficient cloning technique using trichostatin A (TSA) which leads to a 2-5 fold increase in success rates for mouse cloning of B6D2F1 cumulus cells. To further test the validity of this TSA cloning technique, we tried to clone the adult ICR mouse, an outbred strain, which has never been directly cloned before. Only when TSA was used did we obtain both male and female cloned mice from cumulus and fibroblast cells of adult ICR mice with 4-5% success rates, which is comparable to 5-7% of B6D2F1. Thus, the TSA treatment is the first cloning technique to allow us to successfully clone outbred mice, demonstrating that this technique not only improves the success rates of cloning from hybrid strains, but also enables mouse cloning from normally "unclonable" strains.

Better Utilization of Mouse Models of Neurodegenerative Diseases in Preclinical Studies: From the Bench to the Clinic.

PubMed

Janus, Christopher; Hernandez, Carolina; deLelys, Victoria; Roder, Hanno; Welzl, Hans

2016-01-01

The major symptom of Alzheimer's disease is dementia progressing with age. Its clinical diagnosis is preceded by a long prodromal period of brain pathology that encompasses both formation of extracellular amyloid and intraneuronal tau deposits in the brain and widespread neuronal death. At present, familial cases of dementia provide the most promising foundation for modeling neurodegenerative tauopathies, a group of heterogeneous disorders characterized by prominent intracellular accumulation of hyperphosphorylated tau protein. In this chapter, we describe major behavioral hallmarks of tauopathies, briefly outline the genetics underlying familial cases, and discuss the arising implications for modeling the disease in transgenic mouse systems. The selection of tests performed to evaluate the phenotype of a model should be guided by the key behavioral hallmarks that characterize human disorder and their homology to mouse cognitive systems. We attempt to provide general guidelines and establish criteria for modeling dementia in a mouse; however, interpretations of obtained results should avoid a reductionist "one gene, one disease" explanation of model characteristics. Rather, the focus should be directed to the question of how the mouse genome can cope with the over-expression of the protein coded by transgene(s). While each model is valuable within its own constraints and the experiments performed are guided by specific hypotheses, we seek to expand upon their methodology by offering guidance spanning from issues of mouse husbandry to choices of behavioral tests and routes of drug administration that might increase the external validity of studies and consequently optimize the translational aspect of preclinical research.

Rodent models of congenital and hereditary cataract in man.

PubMed

Tripathi, B J; Tripathi, R C; Borisuth, N S; Dhaliwal, R; Dhaliwal, D

1991-01-01

Because the organogenesis and physiology of the lens are essentially similar in various mammals, an understanding of the etiology and pathogenesis of the formation of cataract in an animal model will enhance our knowledge of cataractogenesis in man. In this review, we summarize the background, etiology, and pathogenesis of cataracts that occur in rodents. The main advantages of using rodent mutants include the well-researched genetics of the animals and the comparative ease of breeding of large litters. Numerous rodent models of congenital and hereditary cataracts have been studied extensively. In mice, the models include the Cts strain, Fraser mouse, lens opacity gene (Lop) strain, Lop-2 and Lop-3 strains, Philly mouse, Nakano mouse, Nop strain, Deer mouse, Emory mouse, Swiss Webster strain, Balb/c-nct/nct mouse, and SAM-R/3 strain. The rat models include BUdR, ICR, Sprague-Dawley, and Wistar rats, the spontaneously hypertensive rat (SHR), the John Rapp inbred strain of Dahl salt-sensitive rat, as well as WBN/Kob, Royal College of Surgeons (RCS), and Brown-Norway rats. Other proposed models for the study of hereditary cataract include the degu and the guinea pig. Because of the ease of making clinical observations in vivo and the subsequent availability of the intact lens for laboratory analyses at different stages of cataract formation, these animals provide excellent models for clinicopathologic correlations, for monitoring of the natural history of the aging process and of metabolic defects, as well as for investigations on the effect of cataract-modulating agents and drugs, including the prospect of gene therapy.

Mouse Genome Informatics (MGI) Is the International Resource for Information on the Laboratory Mouse.

PubMed

Law, MeiYee; Shaw, David R

2018-01-01

Mouse Genome Informatics (MGI, http://www.informatics.jax.org/ ) web resources provide free access to meticulously curated information about the laboratory mouse. MGI's primary goal is to help researchers investigate the genetic foundations of human diseases by translating information from mouse phenotypes and disease models studies to human systems. MGI provides comprehensive phenotypes for over 50,000 mutant alleles in mice and provides experimental model descriptions for over 1500 human diseases. Curated data from scientific publications are integrated with those from high-throughput phenotyping and gene expression centers. Data are standardized using defined, hierarchical vocabularies such as the Mammalian Phenotype (MP) Ontology, Mouse Developmental Anatomy and the Gene Ontologies (GO). This chapter introduces you to Gene and Allele Detail pages and provides step-by-step instructions for simple searches and those that take advantage of the breadth of MGI data integration.

Differential biological effects of dehydroepiandrosterone (DHEA) between mouse (B16F10) and human melanoma (BLM) cell lines.

PubMed

Joshi, Kumud; Hassan, Sherif S; Ramaraj, Pandurangan

2017-01-01

Dehydroepiandrosterone (DHEA) is a weak androgen and had been shown to have anti-cancer, anti-adipogenic and anti-inflammatory effects on mouse and other rodent models, but not on humans, suggesting a systemic level difference between mouse and human. Our previous study on DHEA biological functions involving a variety of cell lines, suggested that the functional differences between mouse and human existed even at the cellular level. Hence, using mouse and human melanoma cell models, in-vitro effects of DHEA on cell growth, mechanism of cell death and mechanism of DHEA action were studied. Results indicated a differential biological effects of DHEA between mouse and human melanoma cell lines. These in-vitro studies also suggested that the differential biological effects observed between these two cell lines could be due to the difference in the way DHEA was processed or metabolized inside the cell.

Current State of Animal (Mouse) Modeling in Melanoma Research.

PubMed

Kuzu, Omer F; Nguyen, Felix D; Noory, Mohammad A; Sharma, Arati

2015-01-01

Despite the considerable progress in understanding the biology of human cancer and technological advancement in drug discovery, treatment failure remains an inevitable outcome for most cancer patients with advanced diseases, including melanoma. Despite FDA-approved BRAF-targeted therapies for advanced stage melanoma showed a great deal of promise, development of rapid resistance limits the success. Hence, the overall success rate of melanoma therapy still remains to be one of the worst compared to other malignancies. Advancement of next-generation sequencing technology allowed better identification of alterations that trigger melanoma development. As development of successful therapies strongly depends on clinically relevant preclinical models, together with the new findings, more advanced melanoma models have been generated. In this article, besides traditional mouse models of melanoma, we will discuss recent ones, such as patient-derived tumor xenografts, topically inducible BRAF mouse model and RCAS/TVA-based model, and their advantages as well as limitations. Although mouse models of melanoma are often criticized as poor predictors of whether an experimental drug would be an effective treatment, development of new and more relevant models could circumvent this problem in the near future.

Identification of novel mRNAs and lncRNAs associated with mouse experimental colitis and human inflammatory bowel disease.

PubMed

Rankin, Carl Robert; Theodorou, Evangelos; Law, Ivy Ka Man; Rowe, Lorraine; Kokkotou, Efi; Pekow, Joel; Wang, Jiafang; Martin, Martin G; Pothoulakis, Charalabos; Padua, David Miguel

2018-06-28

Inflammatory bowel disease (IBD) is a complex disorder that is associated with significant morbidity. While many recent advances have been made with new diagnostic and therapeutic tools, a deeper understanding of its basic pathophysiology is needed to continue this trend towards improving treatments. By utilizing an unbiased, high-throughput transcriptomic analysis of two well-established mouse models of colitis, we set out to uncover novel coding and non-coding RNAs that are differentially expressed in the setting of colonic inflammation. RNA-seq analysis was performed using colonic tissue from two mouse models of colitis, a dextran sodium sulfate induced model and a genetic-induced model in mice lacking IL-10. We identified 81 coding RNAs that were commonly altered in both experimental models. Of these coding RNAs, 12 of the human orthologs were differentially expressed in a transcriptomic analysis of IBD patients. Interestingly, 5 of the 12 of human differentially expressed genes have not been previously identified as IBD-associated genes, including ubiquitin D. Our analysis also identified 15 non-coding RNAs that were differentially expressed in either mouse model. Surprisingly, only three non-coding RNAs were commonly dysregulated in both of these models. The discovery of these new coding and non-coding RNAs expands our transcriptional knowledge of mouse models of IBD and offers additional targets to deepen our understanding of the pathophysiology of IBD.

Gene Regions Responding to Skeletal Muscle Atrophy

NASA Technical Reports Server (NTRS)

Booth, Frank W.

1997-01-01

Our stated specific aims for this project were: 1) Identify the region(s) of the mouse IIb myosin heavy chain (MHC) promoter necessary for in vivo expression in mouse fast-twitch muscle, and 2) Identify the region(s) of the mouse IIb MHC promoter responsive to immobilization in mouse slow-twitch muscle in vivo. We sought to address these specific aims by introducing various MHC IIb promoter/reporter gene constructs directly into the tibialis anterior and gastrocnemius muscles of living mice. Although the method of somatic gene transfer into skeletal muscle by direct injection has been successfully used in our laboratory to study the regulation of the skeletal alpha actin gene in chicken skeletal muscle, we had many difficulties utilizing this procedure in the mouse. Because of the small size of the mouse soleus and the difficulty in obtaining consistent results, we elected not to study this muscle as first proposed. Rather, our MHC IIb promoter deletion experiments were performed in the gastrocnemius. Further, we decided to use hindlimb unloading via tail suspension to induce an upregulation of the MHC IIb gene, rather than immobilization of the hindlimbs via plaster casts. This change was made because tail suspension more closely mimics spaceflight, and this procedure in our lab results in a smaller loss of overall body mass than the mouse hindlimb immobilization procedure. This suggests that the stress level during tail suspension is less than during immobilization. This research has provided an important beginning point towards understanding the molecular regulation of the MHC lIb gene in response to unweighting of skeletal muscle Future work will focus on the regulation of MHC IIb mRNA stability in response to altered loading of skeletal muscle

A Dynamic Simulation of Musculoskeletal Function in the Mouse Hindlimb During Trotting Locomotion

PubMed Central

Charles, James P.; Cappellari, Ornella; Hutchinson, John R.

2018-01-01

Mice are often used as animal models of various human neuromuscular diseases, and analysis of these models often requires detailed gait analysis. However, little is known of the dynamics of the mouse musculoskeletal system during locomotion. In this study, we used computer optimization procedures to create a simulation of trotting in a mouse, using a previously developed mouse hindlimb musculoskeletal model in conjunction with new experimental data, allowing muscle forces, activation patterns, and levels of mechanical work to be estimated. Analyzing musculotendon unit (MTU) mechanical work throughout the stride allowed a deeper understanding of their respective functions, with the rectus femoris MTU dominating the generation of positive and negative mechanical work during the swing and stance phases. This analysis also tested previous functional inferences of the mouse hindlimb made from anatomical data alone, such as the existence of a proximo-distal gradient of muscle function, thought to reflect adaptations for energy-efficient locomotion. The results do not strongly support the presence of this gradient within the mouse musculoskeletal system, particularly given relatively high negative net work output from the ankle plantarflexor MTUs, although more detailed simulations could test this further. This modeling analysis lays a foundation for future studies of the control of vertebrate movement through the development of neuromechanical simulations. PMID:29868576

Additive interactions between 1-methyl-1,2,3,4-tetrahydroisoquinoline and clobazam in the mouse maximal electroshock-induced tonic seizure model--an isobolographic analysis for parallel dose-response relationship curves.

PubMed

Andres-Mach, Marta; Haratym-Maj, Agnieszka; Zagaja, Mirosław; Luszczki, Jarogniew J

2014-01-01

The aim of this study was to characterize the anticonvulsant effect of 1-methyl-1,2,3,4-tetrahydroisoquinoline (1-MeTHIQ) in combination with clobazam (CLB) in the mouse maximal electroshock-induced seizure (MES) model. The anticonvulsant interaction profile between 1-MeTHIQ and CLB in the mouse MES model was determined using an isobolographic analysis for parallel dose-response relationship curves. Electroconvulsions were produced in albino Swiss mice by a current (sine wave, 25 mA, 500 V, 50 Hz, 0.2-second stimulus duration) delivered via auricular electrodes by a Hugo Sachs generator. There was an additive effect of the combination of 1-MeTHIQ with CLB (at the fixed ratios of 1:3, 1:1 and 3:1) in the mouse MES-induced tonic seizure model. The additive interaction of the combination of 1-MeTHIQ with CLB (at fixed-ratios of 1:3, 1:1 and 3:1) in the mouse MES model seems to be pharmacodynamic in nature and worth of considering in further clinical practice. © 2014 S. Karger AG, Basel.

Improvements and Limitations of Humanized Mouse Models for HIV Research: NIH/NIAID “Meet the Experts” 2015 Workshop Summary

PubMed Central

Akkina, Ramesh; Allam, Atef; Balazs, Alejandro B.; Blankson, Joel N.; Burnett, John C.; Casares, Sofia; Garcia, J. Victor; Hasenkrug, Kim J.; Kitchen, Scott G.; Klein, Florian; Kumar, Priti; Luster, Andrew D.; Poluektova, Larisa Y.; Rao, Mangala; Shultz, Leonard D.; Zack, Jerome A.

2016-01-01

Abstract The number of humanized mouse models for the human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) and other infectious diseases has expanded rapidly over the past 8 years. Highly immunodeficient mouse strains, such as NOD/SCID/gamma chainnull (NSG, NOG), support better human hematopoietic cell engraftment. Another improvement is the derivation of highly immunodeficient mice, transgenic with human leukocyte antigens (HLAs) and cytokines that supported development of HLA-restricted human T cells and heightened human myeloid cell engraftment. Humanized mice are also used to study the HIV reservoir using new imaging techniques. Despite these advances, there are still limitations in HIV immune responses and deficits in lymphoid structures in these models in addition to xenogeneic graft-versus-host responses. To understand and disseminate the improvements and limitations of humanized mouse models to the scientific community, the NIH sponsored and convened a meeting on April 15, 2015 to discuss the state of knowledge concerning these questions and best practices for selecting a humanized mouse model for a particular scientific investigation. This report summarizes the findings of the NIH meeting. PMID:26670361

Behavioral assays with mouse models of Alzheimer’s disease: practical considerations and guidelines

PubMed Central

Puzzo, Daniela; Lee, Linda; Palmeri, Agostino; Calabrese, Giorgio; Arancio, Ottavio

2014-01-01

In Alzheimer’s disease (AD) basic research and drug discovery, mouse models are essential resources for uncovering biological mechanisms, validating molecular targets and screening potential compounds. Both transgenic and non-genetically modified mouse models enable access to different types of AD-like pathology in vivo. Although there is a wealth of genetic and biochemical studies on proposed AD pathogenic pathways, as a disease that centrally features cognitive failure, the ultimate readout for any interventions should be measures of learning and memory. This is particularly important given the lack of knowledge on disease etiology – assessment by cognitive assays offers the advantage of targeting relevant memory systems without requiring assumptions about pathogenesis. A multitude of behavioral assays are available for assessing cognitive functioning in mouse models, including ones specific for hippocampal-dependent learning and memory. Here we review the basics of available transgenic and non-transgenic AD mouse models and detail three well-established behavioral tasks commonly used for testing hippocampal-dependent cognition in mice – contextual fear conditioning, radial arm water maze and Morris water maze. In particular, we discuss the practical considerations, requirements and caveats of these behavioral testing paradigms. PMID:24462904

Icotinib inhibits EGFR signaling and alleviates psoriasis-like symptoms in animal models.

PubMed

Tan, Fenlai; Yang, Guiqun; Wang, Yanping; Chen, Haibo; Yu, Bo; Li, He; Guo, Jing; Huang, Xiaoling; Deng, Yifang; Yu, Pengxia; Ding, Lieming

2018-02-01

To investigate the effects of icotinib hydrochloride and a derivative cream on epidermal growth factor receptor (EGFR) signaling and within animal psoriasis models, respectively. The effect of icotinib on EGFR signaling was examined in HaCaT cells, while its effect on angiogenesis was tested in chick embryo chorioallantoic membranes (CAM). The effectiveness of icotinib in treating psoriasis was tested in three psoriasis models, including diethylstilbestrol-treated mouse vaginal epithelial cells, mouse tail granular cell layer formation, and propranolol-induced psoriasis-like features in guinea pig ear skin. Icotinib treatment blocked EGFR signaling and reduced HaCaT cell viability as well as suppressed CAM angiogenesis. Topical application of icotinib ameliorated psoriasis-like histological characteristics in mouse and guinea pig psoriasis models. Icotinib also significantly inhibited mouse vaginal epithelium mitosis, promoted mouse tail squamous epidermal granular layer formation, and reduced the thickness of the horny layer in propranolol treated auricular dorsal surface of guinea pig. We conclude that icotinib can effectively inhibit psoriasis in animal models. Future clinical studies should be conducted to explore the therapeutic effects of icotinb in humans. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Pathogenicity of swine influenza viruses possessing an avian or swine-origin PB2 polymerase gene evaluated in mouse and pig models.

PubMed

Ma, Wenjun; Lager, Kelly M; Li, Xi; Janke, Bruce H; Mosier, Derek A; Painter, Laura E; Ulery, Eva S; Ma, Jingqun; Lekcharoensuk, Porntippa; Webby, Richard J; Richt, Jürgen A

2011-02-05

PB2 627K is a determinant of influenza host range and contributes to the pathogenicity of human-, avian-, and mouse-adapted influenza viruses in the mouse model. Here we used mouse and pig models to analyze the contribution of a swine-origin and avian-origin PB2 carrying either 627K or 627E in the background of the classical swine H1N1 (A/Swine/Iowa/15/30; 1930) virus. The results showed PB2 627K is crucial for virulence in the mouse model, independent of whether PB2 is derived from an avian or swine influenza virus (SIV). In the pig model, PB2 627E decreases pathogenicity of the classical 1930 SIV when it contains the swine-origin PB2, but not when it possesses the avian-origin PB2. Our study suggests the pathogenicity of SIVs with different PB2 genes and mutation of codon 627 in mice does not correlate with the pathogenicity of the same SIVs in the natural host, the pig. Copyright © 2010 Elsevier Inc. All rights reserved.

An Immunocompetent Mouse Model of Zika Virus Infection.

PubMed

Gorman, Matthew J; Caine, Elizabeth A; Zaitsev, Konstantin; Begley, Matthew C; Weger-Lucarelli, James; Uccellini, Melissa B; Tripathi, Shashank; Morrison, Juliet; Yount, Boyd L; Dinnon, Kenneth H; Rückert, Claudia; Young, Michael C; Zhu, Zhe; Robertson, Shelly J; McNally, Kristin L; Ye, Jing; Cao, Bin; Mysorekar, Indira U; Ebel, Gregory D; Baric, Ralph S; Best, Sonja M; Artyomov, Maxim N; Garcia-Sastre, Adolfo; Diamond, Michael S

2018-05-09

Progress toward understanding Zika virus (ZIKV) pathogenesis is hindered by lack of immunocompetent small animal models, in part because ZIKV fails to effectively antagonize Stat2-dependent interferon (IFN) responses in mice. To address this limitation, we first passaged an African ZIKV strain (ZIKV-Dak-41525) through Rag1 -/- mice to obtain a mouse-adapted virus (ZIKV-Dak-MA) that was more virulent than ZIKV-Dak-41525 in mice treated with an anti-Ifnar1 antibody. A G18R substitution in NS4B was the genetic basis for the increased replication, and resulted in decreased IFN-β production, diminished IFN-stimulated gene expression, and the greater brain infection observed with ZIKV-Dak-MA. To generate a fully immunocompetent mouse model of ZIKV infection, human STAT2 was introduced into the mouse Stat2 locus (hSTAT2 KI). Subcutaneous inoculation of pregnant hSTAT2 KI mice with ZIKV-Dak-MA resulted in spread to the placenta and fetal brain. An immunocompetent mouse model of ZIKV infection may prove valuable for evaluating countermeasures to limit disease. Copyright © 2018 Elsevier Inc. All rights reserved.

Histologic scoring of gastritis and gastric cancer in mouse models.

PubMed

Rogers, Arlin B

2012-01-01

Histopathology is a defining endpoint in mouse models of experimental gastritis and gastric adenocarcinoma. Presented here is an overview of the histology of gastritis and gastric cancer in mice experimentally infected with Helicobacter pylori or H. felis. A modular histopathologic scoring scheme is provided that incorporates relevant disease-associated changes. Whereas the guide uses Helicobacter infection as the prototype challenge, features may be applied to chemical and genetically engineered mouse models of stomach cancer as well. Specific criteria included in the combined gastric histologic activity index (HAI) include inflammation, epithelial defects, oxyntic atrophy, hyperplasia, pseudopyloric metaplasia, and dysplasia or neoplasia. Representative photomicrographs accompany descriptions for each lesion grade. Differentiation of genuine tumor invasion from pseudoinvasion is highlighted. A brief comparison of normal rodent versus human stomach anatomy and physiology is accompanied by an introduction to mouse-specific lesions including mucous metaplasia and eosinophilic droplets (hyalinosis). In conjunction with qualified pathology support, this guide is intended to assist research scientists, postdoctoral fellows, graduate students, and medical professionals from affiliated disciplines in the interpretation and histologic grading of chronic gastritis and gastric carcinoma in mouse models.

Understanding the biology of ex vivo-expanded CD8 T cells for adoptive cell therapy: role of CD62L.

PubMed

Díaz-Montero, C Marcela; Zidan, Abdel-Aziz; Pallin, Maria F; Anagnostopoulos, Vasileios; Salem, Mohamed L; Wieder, Eric; Komanduri, Krishna; Montero, Alberto J; Lichtenheld, Mathias G

2013-12-01

CD62L governs the circulation of CD8(+) T cells between lymph nodes and peripheral tissues, whereby the expression of CD62L by CD8(+) T cells promotes their recirculation through lymph nodes. As such, CD62L participates in the fate of adoptively transferred CD8(+) T cells and may control their effectiveness for cancer immunotherapy, including settings in which host preconditioning results in the acute lymphopenia-induced proliferation of the transferred cells. Indeed, previous studies correlated CD62L expression by donor CD8(+) cells with the success rate of adoptive cell therapy (ACT). Here, we analyzed the functions and fate of ex vivo-activated, tumor-specific CD62L(-/-) CD8(+) T cells in a mouse melanoma model for ACT. Unexpectedly, we observed that CD62L(-/-) CD8(+) T cells were functionally indistinguishable from CD62L(+/+) CD8(+) T cells, i.e., both greatly expanded in cyclophosphamide preconditioned animals, controlled subcutaneously and hematogenously spreading tumors, and generated anti-tumor-specific CD8(+) T cell memory. Moreover, even in hosts with rudimentary secondary lymphoid organs (LT(-/-) animals), CD8(+) T cells with and without CD62L expanded equivalently to those adoptively transferred into wild-type animals. These results put into question the utility of CD62L as a predictive biomarker for the efficacy of ex vivo-expanded T cells after ACT in lymphopenic conditions and also offer new insights into the homing, engraftment, and memory generation of adoptively transferred ex vivo-activated CD8(+) T cells.

Adoptive transfer of MART-1 T cell receptor transgenic lymphocytes and dendritic cell vaccination in patients with metastatic melanoma

PubMed Central

Chodon, Thinle; Comin-Anduix, Begonya; Chmielowski, Bartosz; Koya, Richard C; Wu, Zhongqi; Auerbach, Martin; Ng, Charles; Avramis, Earl; Seja, Elizabeth; Villanueva, Arturo; McCannel, Tara A.; Ishiyama, Akira; Czernin, Johannes; Radu, Caius G.; Wang, Xiaoyan; Gjertson, David W.; Cochran, Alistair J.; Cornetta, Kenneth; Wong, Deborah J.L.; Kaplan-lefko, Paula; Hamid, Omid; Samlowski, Wolfram; Cohen, Peter A.; Daniels, Gregory A.; Mukherji, Bijay; Yang, Lili; Zack, Jerome A.; Kohn, Donald B.; Heath, James R.; Glaspy, John A.; Witte, Owen N.; Baltimore, David; Economou, James S.; Ribas, Antoni

2014-01-01

Purpose It has been demonstrated that large numbers of tumor-specific T cells for adoptive cell transfer (ACT) can be manufactured by retroviral genetic engineering of autologous peripheral blood lymphocytes and expanding them over several weeks. In mouse models, this therapy is optimized when administered with dendritic cell (DC) vaccination. We developed a short one-week manufacture protocol to determine the feasibility, safety and antitumor efficacy of this double cell therapy. Experimnetal Design A clinical trial (NCT00910650) adoptively transferring MART-1 T cell receptor (TCR) transgenic lymphocytes together with MART-1 peptide pulsed DC vaccination in HLA-A2.1 patients with metastatic melanoma. Autologous TCR transgenic cells were manufactured in 6 to 7 days using retroviral vector gene transfer, and re-infused with (n = 10) or without (n = 3) prior cryopreservation. Results 14 patients with metastatic melanoma were enrolled and nine out of 13 treated patients (69%) showed evidence of tumor regression. Peripheral blood reconstitution with MART-1-specific T cells peaked within two weeks of ACT indicating rapid in vivo expansion. Administration of freshly manufactured TCR transgenic T cells resulted in a higher persistence of MART-1-specific T cells in the blood as compared to cryopreserved. Evidence that DC vaccination could cause further in vivo expansion was only observed with ACT using non-cryopreserved T cells. Conclusion Double cell therapy with ACT of TCR engineered T cells with a very short ex vivo manipulation and DC vaccines is feasible and results in antitumor activity, but improvements are needed to maintain tumor responses. PMID:24634374

A Simplified, Langendorff-Free Method for Concomitant Isolation of Viable Cardiac Myocytes and Nonmyocytes From the Adult Mouse Heart

PubMed Central

Ackers-Johnson, Matthew; Li, Peter Yiqing; Holmes, Andrew P.; O’Brien, Sian-Marie; Pavlovic, Davor; Foo, Roger S.

2018-01-01

Rationale Cardiovascular disease represents a global pandemic. The advent of and recent advances in mouse genomics, epigenomics, and transgenics offer ever-greater potential for powerful avenues of research. However, progress is often constrained by unique complexities associated with the isolation of viable myocytes from the adult mouse heart. Current protocols rely on retrograde aortic perfusion using specialized Langendorff apparatus, which poses considerable logistical and technical barriers to researchers and demands extensive training investment. Objective To identify and optimize a convenient, alternative approach, allowing the robust isolation and culture of adult mouse cardiac myocytes using only common surgical and laboratory equipment. Methods and Results Cardiac myocytes were isolated with yields comparable to those in published Langendorff-based methods, using direct needle perfusion of the LV ex vivo and without requirement for heparin injection. Isolated myocytes can be cultured antibiotic free, with retained organized contractile and mitochondrial morphology, transcriptional signatures, calcium handling, responses to hypoxia, neurohormonal stimulation, and electric pacing, and are amenable to patch clamp and adenoviral gene transfer techniques. Furthermore, the methodology permits concurrent isolation, separation, and coculture of myocyte and nonmyocyte cardiac populations. Conclusions We present a novel, simplified method, demonstrating concomitant isolation of viable cardiac myocytes and nonmyocytes from the same adult mouse heart. We anticipate that this new approach will expand and accelerate innovative research in the field of cardiac biology. PMID:27502479

Astonishing advances in mouse genetic tools for biomedical research.

PubMed

Kaczmarczyk, Lech; Jackson, Walker S

2015-01-01

The humble house mouse has long been a workhorse model system in biomedical research. The technology for introducing site-specific genome modifications led to Nobel Prizes for its pioneers and opened a new era of mouse genetics. However, this technology was very time-consuming and technically demanding. As a result, many investigators continued to employ easier genome manipulation methods, though resulting models can suffer from overlooked or underestimated consequences. Another breakthrough, invaluable for the molecular dissection of disease mechanisms, was the invention of high-throughput methods to measure the expression of a plethora of genes in parallel. However, the use of samples containing material from multiple cell types could obfuscate data, and thus interpretations. In this review we highlight some important issues in experimental approaches using mouse models for biomedical research. We then discuss recent technological advances in mouse genetics that are revolutionising human disease research. Mouse genomes are now easily manipulated at precise locations thanks to guided endonucleases, such as transcription activator-like effector nucleases (TALENs) or the CRISPR/Cas9 system, both also having the potential to turn the dream of human gene therapy into reality. Newly developed methods of cell type-specific isolation of transcriptomes from crude tissue homogenates, followed by detection with next generation sequencing (NGS), are vastly improving gene regulation studies. Taken together, these amazing tools simplify the creation of much more accurate mouse models of human disease, and enable the extraction of hitherto unobtainable data.

Mutational landscape of a chemically-induced mouse model of liver cancer.

PubMed

Connor, Frances; Rayner, Tim F; Aitken, Sarah J; Feig, Christine; Lukk, Margus; Santoyo-Lopez, Javier; Odom, Duncan T

2018-06-26

Carcinogen-induced mouse models of liver cancer are used extensively to study pathogenesis of the disease and have a critical role in validating candidate therapeutics. These models can recapitulate molecular and histological features of human disease. However, it is not known if the genomic alterations driving these mouse tumour genomes are comparable to those found in human tumours. Here, we provide a detailed genomic characterisation of tumours from a commonly used mouse model of hepatocellular carcinoma (HCC). We analysed whole exome sequences of liver tumours arising in mice exposed to diethylnitrosamine (DEN). DEN-initiated tumours had a high, uniform number of somatic single nucleotide variants (SNVs), with few insertions, deletions or copy number alterations, consistent with the known genotoxic action of DEN. Exposure of hepatocytes to DEN left a reproducible mutational imprint in resulting tumour exomes which we could computationally reconstruct using six known COSMIC mutational signatures. The tumours carried a high diversity of low-incidence, non-synonymous point mutations in many oncogenes and tumour suppressors, reflecting the stochastic introduction of SNVs into the hepatocyte genome by the carcinogen. We identified four recurrently mutated genes that were putative oncogenic drivers of HCC in this model. Every neoplasm carried activating hotspot mutations either in codon 61 of Hras, in codon 584 of Braf or in codon 254 of Egfr. Truncating mutations of Apc occurred in 21% of neoplasms, which were exclusively carcinomas supporting a role for deregulation of Wnt/β-catenin signalling in cancer progression. Our study provides detailed insight into the mutational landscape of tumours arising in a commonly-used carcinogen model of HCC, facilitating the future use of this model to understand the human disease. Mouse models are widely used to study the biology of cancer and to test potential therapies. Here, we have described the mutational landscape of tumours arising in a carcinogen-induced mouse model of liver cancer. Since cancer is a disease caused by genomic alterations, information about the patterns and types of mutations in the tumours in this mouse model should facilitate its use to study human liver cancer. Copyright © 2018 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

SU-F-T-669: Commissioning of An Electronic Brachytherapy System for Targeted Mouse Irradiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Culberson, W; Micka, J; Carchman, E

Purpose: The aim of this study was to commission the Xoft Axxent™ electronic brachytherapy (eBT) source and 10 mm diameter surface applicator with NIST traceability for targeted irradiations of mouse anal carcinomas. Methods: The Xoft Axxent™ electronic brachytherapy (eBT) and 10 mm diameter surface applicator was chosen by the collaborating physician as a radiation delivery mechanism for mouse anal carcinomas. The target dose was 2 Gy at a depth of 3 mm in tissue to be delivered in a single fraction. To implement an accurate and reliable irradiation plan, the system was commissioned by first determining the eBT source outputmore » and corresponding dose rate at a depth of 3 mm in tissue. This was determined through parallel-plate ion chamber measurements and published conversion factors. Well-type ionization chamber measurements were used to determine a transfer coefficient, which correlates the measured dose rate at 3 mm to the NIST-traceable quantity, air-kerma rate at 50 cm in air, for eBT sources. By correlating these two quantities, daily monitoring in the well chamber becomes an accurate and efficient quality assurance technique. Once the dose-rate was determined, a treatment recipe was developed and confirmed with chamber measurements to deliver the requested dose. Radiochromic film was used to verify the dose distribution across the field. Results: Dose rates at 3 mm depth in tissue were determined for two different Xoft Axxent™ sources and correlated with NIST-traceable well-type ionization chamber measurements. Unique transfer coefficients were determined for each source and the treatment recipe was validated by measurements. Film profiles showed a uniform dose distribution across the field. Conclusion: A Xoft Axxent™ eBT system was successfully commissioned for use in the irradiation of mouse rectal tumors. Dose rates in tissue were determined as well as other pertinent parameters to ensure accurate delivery of dose to the target region.« less

The influence of the structural orientation of amide linkers on the serum compatibility and lung transfection properties of cationic amphiphiles.

PubMed

Srujan, Marepally; Chandrashekhar, Voshavar; Reddy, Rakesh C; Prabhakar, Rairala; Sreedhar, Bojja; Chaudhuri, Arabinda

2011-08-01

Understanding the structural parameters of cationic amphiphiles which can influence gene transfer efficiencies of cationic amphiphiles continues to remain important for designing efficient liposomal gene delivery reagents. Previously we demonstrated the influence of structural orientation of the ester linker (widely used in covalently tethering the polar head and the non-polar tails) in modulating in vitro gene transfer efficiencies of cationic amphiphiles. However, our previously described cationic amphiphiles with ester linkers failed to deliver genes under in vivo conditions. Herein we report on the development of a highly serum compatible cationic amphiphile with circulation stable amide linker which shows remarkable selectivity in transfecting mouse lung. We also demonstrate that reversing structural orientation of the amide linker adversely affects both serum compatibility and the lung selective gene transfer property. Dynamic laser light scattering and atomic force microscopic studies revealed smaller average hydrodynamic sizes of the liposomes of transfection efficient lipid than those for the liposomes of transfection incompetent analog (148 ± 1 nm vs 214 ± 4 nm). Average surface potential of the liposomes of transfection competent amphiphiles were found to be significantly higher than that for the liposomes of transfection incompetent analog (10.7 ± 5.4 mV vs 2.8 ± 1.3 mV, respectively). Findings in fluorescence resonance energy transfer and dye entrapment experiments support lower rigidity and higher biomembrane fusogenicity of the liposomes of the transfection efficient amphiphiles. Importantly, cationic lipoplexes of the novel amide-linker based amphiphile exhibited higher mouse lung selective gene transfer properties than DOTAP, one of the widely used commercially available liposomal lung transfection kits. In summary, the present findings demonstrate for the first time that amide linker structural orientation profoundly influences the serum compatibility and lung transfection efficiencies of cationic amphiphiles. Copyright © 2011 Elsevier Ltd. All rights reserved.

Chip Based Magnetic Imager for Molecular Profiling of Ovarian Cancer Cells

DTIC Science & Technology

2016-12-01

2015) Genome-wide CRISPR screen in a mouse model of tumor growth and metastasis. Cell 160:1246-1260. PMC4380877, PMID:25748654. Acknowledgement of...Weissleder R, Lee H, Zhang F, Sharp PA (2015) Genome-wide CRISPR screen in a mouse model of tumor growth and metastasis. Cell 160:1246-1260. 5. Im H, Shao H...Lett 32(10):1229–1231. 6 of 6 | www.pnas.org/cgi/doi/10.1073/pnas.1501815112 Im et al. Resource Genome-wide CRISPR Screen in a Mouse Model of Tumor