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1

HLA-B27 and Human ?2-Microglobulin Affect the Gut Microbiota of Transgenic Rats  

Science.gov (United States)

The HLA-B27 gene is a major risk factor for clinical diseases including ankylosing spondylitis, acute anterior uveitis, reactive arthritis, and psoriatic arthritis, but its mechanism of risk enhancement is not completely understood. The gut microbiome has recently been shown to influence several HLA-linked diseases. However, the role of HLA-B27 in shaping the gut microbiome has not been previously investigated. In this study, we characterize the differences in the gut microbiota mediated by the presence of the HLA-B27 gene. We identified differences in the cecal microbiota of Lewis rats transgenic for HLA-B27 and human ?2-microglobulin (h?2m), compared with wild-type Lewis rats, using biome representational in situ karyotyping (BRISK) and 16S rRNA gene sequencing. 16S sequencing revealed significant differences between transgenic animals and wild type animals by principal coordinates analysis. Further analysis of the data set revealed an increase in Prevotella spp. and a decrease in Rikenellaceae relative abundance in the transgenic animals compared to the wild type animals. By BRISK analysis, species-specific differences included an increase in Bacteroides vulgatus abundance in HLA-B27/h?2m and h?2m compared to wild type rats. The finding that HLA-B27 is associated with altered cecal microbiota has not been shown before and can potentially provide a better understanding of the clinical diseases associated with this gene. PMID:25140823

Lin, Phoebe; Bach, Mary; Asquith, Mark; Lee, Aaron Y.; Akileswaran, Lakshmi; Stauffer, Patrick; Davin, Sean; Pan, Yuzhen; Cambronne, Eric D.; Dorris, Martha; Debelius, Justine W.; Lauber, Christian L.; Ackermann, Gail; Baeza, Yoshiki V.; Gill, Tejpal; Knight, Rob; Colbert, Robert A.; Taurog, Joel D.; Van Gelder, Russell N.; Rosenbaum, James T.

2014-01-01

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Characterization of psoriasiform and alopecic skin lesions in HLA-B27 transgenic rats.  

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We have previously reported a multisystem inflammatory disease in transgenic rat lines with high expression of HLA-B*2705 and human beta 2 microglobulin. Skin disease in these rats includes two predominant lesions: 1) marked psoriasiform dermatitis of the tail and digits; and 2) progressive alopecia of face, neck, trunk, and extremities. Here we present the results of a systematic survey of these lesions. Tail and digit skin showed psoriasiform hyperplasia of the epidermis associated with par...

Yanagisawa, H.; Richardson, J. A.; Taurog, J. D.; Hammer, R. E.

1995-01-01

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Chemically Defined Diet Alters the Protective Properties of Fructo-Oligosaccharides and Isomalto-Oligosaccharides in HLA-B27 Transgenic Rats  

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Non-digestible oligosaccharides (NDO) were shown to reduce inflammation in experimental colitis, but it remains unclear whether microbiota changes mediate their colitis-modulating effects. This study assessed intestinal microbiota and intestinal inflammation after feeding chemically defined AIN-76A or rat chow diets, with or without supplementation with 8 g/kg body weight of fructo-oligosaccharides (FOS) or isomalto-oligosaccharides (IMO). The study used HLA-B27 transgenic rats, a validated model of inflammatory bowel disease (IBD), in a factorial design with 6 treatment groups. Intestinal inflammation and intestinal microbiota were analysed after 12 weeks of treatment. FOS and IMO reduced colitis in animals fed rat chow, but exhibited no anti-inflammatory effect when added to AIN-76A diets. Both NDO induced specific but divergent microbiota changes. Bifidobacteria and Enterobacteriaceae were stimulated by FOS, whereas copy numbers of Clostridium cluster IV were decreased. In addition, higher concentrations of total short-chain fatty acids (SCFA) were observed in cecal contents of rats on rat chow compared to the chemically defined diet. AIN-76A increased the relative proportions of propionate, iso-butyrate, valerate and iso-valerate irrespective of the oligosaccharide treatment. The SCFA composition, particularly the relative concentration of iso-butyrate, valerate and iso-valerate, was associated (P?0.004 and r?0.4) with increased colitis and IL-1 ? concentration of the cecal mucosa. This study demonstrated that the protective effects of fibres on colitis development depend on the diet. Although diets modified specific cecal microbiota, our study indicates that these changes were not associated with colitis reduction. Intestinal inflammation was positively correlated to protein fermentation and negatively correlated with carbohydrate fermentation in the large intestine. PMID:25369019

Valcheva, Rosica; Ganzle, Michael G.; Dieleman, Levinus A.

2014-01-01

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Altered Bone Material Properties in HLA-B27 Rats Include Reduced Mineral to Matrix Ratio and Altered Collagen Cross-Links.  

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Spondyloarthropathy and inflammatory bowel disease (IBD), which includes ulcerative colitis and Crohn's disease, are often associated with severe osteopenia/osteoporosis in both children and adults. HLA-B27 transgenic rats present a phenotype that includes severe colitis and severely accelerated alveolar bone loss. The purpose of this study was to evaluate long bone density status, systemic bone metabolic markers, and intrinsic bone material properties in HLA-B27 transgenic (TG) rats, and compare them with those of age- and sex-matched wild-type (WT) animals. The results indicate that in the HLA-B27 rat, an animal susceptible to both alveolar bone loss (ABL) and long bone osteopenia, there is a statistically significant negative correlation between ABL and long bone bone mineral density (BMD), as well as mineral/matrix ratio at active bone-forming trabecular surfaces. The TG animals had a lower mineral/matrix ratio and higher relative proteoglycan and advanced glycation end product (?-N-Carboxymethyl-L-lysine) content and pyridinoline/divalent collagen cross-link ratio compared with WT. These results may provide better understanding of the interrelationship between osteoporosis and oral bone loss, the underlying causes of the inferior bone strength in the HLA-B27 transgenic animals, and could prove to be a useful model in the elucidation of the pathophysiology of spondyloarthropathy and IBD-associated osteopenia/osteoporosis and in the evaluation of pharmacological intervention(s) against such conditions. © 2014 American Society for Bone and Mineral Research. PMID:24771481

Gamsjaeger, Sonja; Srivastava, Apurva K; Wergedal, Jon E; Zwerina, Jochen; Klaushofer, Klaus; Paschalis, Eleftherios P; Tatakis, Dimitris N

2014-11-01

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HLA-B27 heavy chains contribute to spontaneous inflammatory disease in B27/human beta2-microglobulin (beta2m) double transgenic mice with disrupted mouse beta2m.  

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MHC class I allele, HLA-B27, is strongly associated with a group of human diseases called spondyloarthropathies. Some of these diseases have an onset after an enteric or genitourinary infection. In the present study, we describe spontaneous disease in HLA-B27 transgenic mice where endogenous beta2-microglobulin (beta2m) gene was replaced with transgenic human beta2m gene. These mice showed cell surface expression of HLA-B27 similar to that of human peripheral blood mononuclear cells. In addit...

Khare, S. D.; Hansen, J.; Luthra, H. S.; David, C. S.

1996-01-01

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HLA-B27 Test  

Science.gov (United States)

... name: Human Leukocyte Antigen B27 Related tests: ESR ; C-Reactive Protein ; Rheumatoid Factor ; HLA Testing At a Glance Test ... an ESR (erythrocyte sedimentation rate) or a CRP (C-reactive protein) . HLA-B27 is sometimes ordered to help evaluate ...

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Pathogenicity of Misfolded and Dimeric HLA-B27 Molecules.  

Science.gov (United States)

The association between HLA-B27 and the group of autoimmune inflammatory arthritic diseases, the spondyloarthropathies (SpAs) which include ankylosing spondylitis (AS) and Reactive Arthritis (ReA), has been well established and remains the strongest association between any HLA molecule and autoimmune disease. The mechanism behind this striking association remains elusive; however animal model and biochemical data suggest that HLA-B27 misfolding may be key to understanding its association with the SpAs. Recent investigations have focused on the unusual biochemical structures of HLA-B27 and their potential role in SpA pathogenesis. Here we discuss how these unusual biochemical structures may participate in cellular events leading to chronic inflammation and thus disease progression. PMID:21547037

Antoniou, Antony N; Lenart, Izabela; Guiliano, David B

2011-01-01

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HLA B27 allele types in homogeneous groups of juvenile idiopathic arthritis patients in Latvia  

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Abstract Juvenile idiopathic arthritis (JIA) is a heterogeneous condition and therapeutic strategies vary in different JIA types. The routinely accepted practice to start with Sulphasalazine (SS) as the first line treatment in patients with HLA B27 positive JIA proves to be ineffective in a large proportion of children. Objective to investigate HLA B27 positive JIA patients clinical characteristics, determined HLA B27 allele types and their connection with antirheumati...

Guseinova Dinara; Lazareva Arina; Sochnevs Arturs; Zavadska Dace; Eglite Jelena; Stanevicha Valda; Shantere Ruta; Gardovska Dace

2010-01-01

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HLA-B27 frequency in a group of patients with psoriatic arthritis / Freqüência de HLA-B27 em uma amostra de pacientes com artrite psoriática  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in portuguese FUNDAMENTOS: O HLA-B27 está associado às espondiloartrites, grupo de doenças que engloba, entre outras, a artrite psoriásica. OBJETIVOS: Descrever a freqüência de HLA-B27 em uma amostra de pacientes brasileiros com artrite psoriásica e correlacionar sua presença ou ausência com as manifestações clín [...] icas dos mesmos. MÉTODOS: Estudo transversal avaliando 44 pacientes com artrite psoriásica de um ambulatório de Reumatologia. A avaliação consistia em registro de informações demográficas e sociais, exame clínico da pele e das articulações e pesquisa de HLA-B27. Os dados gerados foram tratados por meio de estatística descritiva e comparativa em Software apropriado. Foram utilizados testes paramétricos e não-paramétricos com significância estatística de 5%. RESULTADOS: O HLA-B27 resultou negativo em 32 dos 44 pacientes estudados (72,7%). A maioria dos pacientes era do sexo masculino, da raça branca, procedente do Rio de Janeiro, portador de psoríase em placas e com idade média de 52,9 anos. Houve associação estatisticamente significativa entre o HLA-B27 positivo e o sexo masculino (p=0,004). O HLA-B27 negativo teve tendência à correlação com artrite de mãos e punhos (p=0,07). Houve correlação inversa significativa entre os valores do HLA e do teste de Schöber (p=0,02). CONCLUSÃO: Apesar do HLA-B27 ser negativo na maioria dos pacientes estudados, esteve significativamente associado ao sexo masculino e inversamente correlacionado ao teste de Schöber. Abstract in english BACKGROUND: HLA-B27 is associated with spondyloarthritis, a group of diseases that includes psoriatic arthritis. OBJECTIVES: To describe the HLA-B27 frequency in a group of Brazilian patients with psoriatic arthritis and correlate its presence or absence with their clinical manifestations. METHODS: [...] Cross-sectional study with 44 psoriatic arthritis patients of a Rheumatology clinic. Demographic and social data were recorded, as were skin and joints clinical examination. HLA-B27 was tested. All data were processed descriptively and comparatively by appropriate software. Parametric and non parametric tests were used with 5% statistical significance. RESULTS: HLA-B27 was negative in 32 of the 44 patients (72,7%). Most of them were male, Caucasian, living in Rio de Janeiro, with plaque type psoriasis and average age of 52,9 years. There was statistical significant correlation between positive HLA-B27 and male gender (p=0,004). Negative HLA-B27 had a tendency to correlate with hands and wrists arthritis (p=0,07). There was an inverse significant correlation between HLA values and Schöber's test (p=0,02). CONCLUSION: Although HLA-B27 is negative in most of patients, it is significantly associated to male gender and inversely correlated with Schöber's test.

Danilo Garcia, Ruiz; Mário Newton Leitão de, Azevedo; Omar, Lupi.

2012-12-01

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HLA-B27 frequency in a group of patients with psoriatic arthritis / Freqüência de HLA-B27 em uma amostra de pacientes com artrite psoriática  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in portuguese FUNDAMENTOS: O HLA-B27 está associado às espondiloartrites, grupo de doenças que engloba, entre outras, a artrite psoriásica. OBJETIVOS: Descrever a freqüência de HLA-B27 em uma amostra de pacientes brasileiros com artrite psoriásica e correlacionar sua presença ou ausência com as manifestações clín [...] icas dos mesmos. MÉTODOS: Estudo transversal avaliando 44 pacientes com artrite psoriásica de um ambulatório de Reumatologia. A avaliação consistia em registro de informações demográficas e sociais, exame clínico da pele e das articulações e pesquisa de HLA-B27. Os dados gerados foram tratados por meio de estatística descritiva e comparativa em Software apropriado. Foram utilizados testes paramétricos e não-paramétricos com significância estatística de 5%. RESULTADOS: O HLA-B27 resultou negativo em 32 dos 44 pacientes estudados (72,7%). A maioria dos pacientes era do sexo masculino, da raça branca, procedente do Rio de Janeiro, portador de psoríase em placas e com idade média de 52,9 anos. Houve associação estatisticamente significativa entre o HLA-B27 positivo e o sexo masculino (p=0,004). O HLA-B27 negativo teve tendência à correlação com artrite de mãos e punhos (p=0,07). Houve correlação inversa significativa entre os valores do HLA e do teste de Schöber (p=0,02). CONCLUSÃO: Apesar do HLA-B27 ser negativo na maioria dos pacientes estudados, esteve significativamente associado ao sexo masculino e inversamente correlacionado ao teste de Schöber. Abstract in english BACKGROUND: HLA-B27 is associated with spondyloarthritis, a group of diseases that includes psoriatic arthritis. OBJECTIVES: To describe the HLA-B27 frequency in a group of Brazilian patients with psoriatic arthritis and correlate its presence or absence with their clinical manifestations. METHODS: [...] Cross-sectional study with 44 psoriatic arthritis patients of a Rheumatology clinic. Demographic and social data were recorded, as were skin and joints clinical examination. HLA-B27 was tested. All data were processed descriptively and comparatively by appropriate software. Parametric and non parametric tests were used with 5% statistical significance. RESULTS: HLA-B27 was negative in 32 of the 44 patients (72,7%). Most of them were male, Caucasian, living in Rio de Janeiro, with plaque type psoriasis and average age of 52,9 years. There was statistical significant correlation between positive HLA-B27 and male gender (p=0,004). Negative HLA-B27 had a tendency to correlate with hands and wrists arthritis (p=0,07). There was an inverse significant correlation between HLA values and Schöber's test (p=0,02). CONCLUSION: Although HLA-B27 is negative in most of patients, it is significantly associated to male gender and inversely correlated with Schöber's test.

Danilo Garcia, Ruiz; Mário Newton Leitão de, Azevedo; Omar, Lupi.

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The radiographic features of rheumatoid arthritis in HLA-B27-positive patients  

International Nuclear Information System (INIS)

Radiographs were reviewed in a group of nine patients with classical seropositive rheumatoid arthritis who on tissue typing were found to express the class I HLA-B27 allele. Radiographs were analyzed with regard to whether or not they demonstrated radiographic features of (1) classical rheumatoid arthritis, (2) seronegative arthritis, or (3) mixed features of rheumatoid and seronegative arthritis. Five patients (55%) displayed radiographic features consistent with a diagnosis of rheumatoid arthritis, two patients (22%) showed radiographic features of seronegative disorder (periostitis and sacroiliitis), and two patients (22%) showed a mixed picture with evidence of both rheumatoid arthritis and a seronegative disorder. Thus, the HLA-B27 allele contributed to the radiographic features in 44% of patients with rheumatoid arthritis and associated HLA-B27. Thus, the wide range of findings in our population indicates that the radiographic attributes are not specific enough to constitute a unique subpopulation of patients with rheumatoid arthritis. (orig.)

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HLA-B27 (antigen in retroperitoneal fibrosis in a family  

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Full Text Available Idiopathic retroperitoneal fibrosis is a rare disease of undetermined aetiology. It is important to distinguish this entity from retroperitoneal fibrosis secondary to malignancy or specific inflammatory disease. There have been no prior means of excluding this condition without surgical exploration and histopathologic study of the excised tissue. A genetic predisposition is suggested for the development of idiopathic primary retroperitoneal fibrosis in patients who are HLA-B27 antigen positive. In this study we present three cases of idiopathic retroperitoneal fibrosis in a family (2 brothers and their grandfather. The presence of HLA-B27 antigen positivity was identified in two of them.

Mohammad Yazdani

2007-03-01

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Functional Interaction of the Ankylosing Spondylitis-associated Endoplasmic Reticulum Aminopeptidase 1 Polymorphism and HLA-B27 in Vivo*  

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The association of ERAP1 with ankylosing spondylitis (AS)1 among HLA-B27-positive individuals suggests that ERAP1 polymorphism may affect pathogenesis by altering peptide-dependent features of the HLA-B27 molecule. Comparisons of HLA-B*27:04-bound peptidomes from cells expressing different natural variants of ERAP1 revealed significant differences in the size, length, and amount of many ligands, as well as in HLA-B27 stability. Peptide analyses suggested that the mechanism of ERAP1/HLA-B27 interaction is a variant-dependent alteration in the balance between epitope generation and destruction determined by the susceptibility of N-terminal flanking and P1 residues to trimming. ERAP1 polymorphism associated with AS susceptibility ensured efficient peptide trimming and high HLA-B27 stability. Protective polymorphism resulted in diminished ERAP1 activity, less efficient trimming, suboptimal HLA-B27 peptidomes, and decreased molecular stability. This study demonstrates that natural ERAP1 polymorphism affects HLA-B27 antigen presentation and stability in vivo and proposes a mechanism for the interaction between these molecules in AS. PMID:22918227

Garcia-Medel, Noel; Sanz-Bravo, Alejandro; Van Nguyen, Dung; Galocha, Begona; Gomez-Molina, Patricia; Martin-Esteban, Adrian; Alvarez-Navarro, Carlos; de Castro, Jose A. Lopez

2012-01-01

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Evaluation of 278 hla-b27 positive patients suspected of seronegative spondyloarthropathies  

International Nuclear Information System (INIS)

To determine HLA-B27 prevalence in patients suspected of Seronegative spondyloarthropathy referred to the Transplantation Department of Blood Transfusion Organization, and to evaluate clinical findings among HLA-B27 positive patients. One thousand six hundred ten patients having clinical manifestation of seronegative SpAs were screened for HLA typing by serological methods from January 1997 to June 2002 at Transplantation Department of Blood Transfusion Organization, Ahwaz, Iran. Serologic-based HLA typing using Antigen-specific sera to determine a person's HLA type was performed. Among these patients, individuals found HLA-B27 positive were investigated regarding clinical findings, age, and sex distribution. In this study the frequency of HLA-B27 antigen was 17.26% (278 cases). The minimum age in males was 10 years and the maximum age in female was 70 years. Median age with seronegative SpAs findings (34.2% including 28.42% females, 71.57% males) was 20-30 years. Based on our results, the most frequent clinical manifestation, was peripheral joints arthritis (58.7%; 34.35% females, 65.65 % males). There were no association between any of the major clinical manifestations and age or sex distribution. These findings confirm the strong association of the HLA B27 allele with various types of spondyloarthritis and suggests that HLA typing would help in the diagnosis of seronagative SpAs, specially ankylosing spondylitis with indeterminate clinical presentation and also in rminate clinical presentation and also in identifying at risk family members. (author)

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HLA-B27 positive juvenile arthritis with cardiac involvement preceding sacroiliac joint changes  

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While cardiovascular disease develops in up to 50% of adult patients with ankylosing spondylitis, it is very uncommon in its juvenile counterpart. Regarding the early stage of the disease, before onset of sacroiliac joint changes, only two cases with aortic incompetence have been published while reports of mitral valve involvement are not available. A 13 year old boy is described with an HLA-B27 positive asymmetric oligoarthritis and enthesitis, without back pain or radiographic evidence of ...

Lee, S.; Im H, Y.; Schueller, W.

2001-01-01

16

The role of HLA-B27 molecules in the pathogenesis of ankylosing spondylitis  

Directory of Open Access Journals (Sweden)

Full Text Available Ankylosing Spondylitis (AS is characterised by the strongest association with an HLA antigen ever described for any disease. It represents therefore the ideal model for the understanding of the link between immune-mediated diseases and the HLA system. The role of HLA-B27 in the pathogenesis of AS will be discussed focusing on the recently described higher expression of these molecules in patients with AS compared with healthy controls.

R. Pala

2011-09-01

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Bilateral macular thickening in mild unilateral anterior uveitis: is HLA-B27 involved?  

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Abstract Background Macular thickening (MT) without clinically recognized macular edema has been described in anterior uveitis (AU). Although fellow-eyes of patients have been used as controls in several studies, little is known about macular thickness in these eyes. We studied the rate and extent of MT in both AU-affected and quiescent fellow-eyes of phakic AU patients with good visual acuity (VA). We also assessed macular thickness related to HLA-B27 presence and to recurre...

Wexler Alexandra; Sand Trond; Elsås Tor B

2012-01-01

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Bilateral macular thickening in mild unilateral anterior uveitis: is HLA-B27 involved?  

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Full Text Available Abstract Background Macular thickening (MT without clinically recognized macular edema has been described in anterior uveitis (AU. Although fellow-eyes of patients have been used as controls in several studies, little is known about macular thickness in these eyes. We studied the rate and extent of MT in both AU-affected and quiescent fellow-eyes of phakic AU patients with good visual acuity (VA. We also assessed macular thickness related to HLA-B27 presence and to recurrence, since these issues have been almost unexplored by previous optical coherence tomography (OCT studies. Methods Patients with AU were prospectively included and macular thickness was measured with OCT initially and on follow up. Macular thickness in patients’ affected eyes (n?=?30 as well as in their quiet fellow-eyes (n?=?28 was compared with eyes of age- and gender matched controls. Inter-ocular differences in macular thickness between AU affected eyes and their fellow-eyes were assessed in patients (n?=?28, also in a subgroup with visual acuity???0.8 (n?=?23 by one-sample Student’s?t-tests. Inter-ocular differences were also assessed related to HLA-B27 presence and related to the status of current AU episode (initial or relapse. Results Subclinical MT is present in both quiet fellow-eyes and AU-affected eyes of patients. MT was found in most cases of AU, even in phakic eyes with good VA. There was a larger increase in macular thickness in HLA-B27-positive than in HLA-B27-negative patients. No differences in macular thickness were found between patients with their first AU episode and patients with recurrent episodes. Conclusions MT probably reflects systemic immune-mediated response to the inflammatory disorder in AU, and it is possible that HLA-B27-related factors are involved in the pathogenesis of AU. These observations are in line with and extend the current understanding of the mechanisms behind MT in AU.

Wexler Alexandra

2012-07-01

19

Phosphorylation of STAT-1 serine 727 is prolonged in HLA-B27-expressing human monocytic cells.  

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A tissue antigen, HLA-B27, is strongly associated with a group of rheumatic diseases called spondyloarthritides. Despite the intensive research, the exact role of HLA-B27 in the pathogenesis of these diseases is still unclear. Here we studied whether HLA-B27 modulates the phosphorylation of signal transducer and activator of transcription 1 (STAT-1) serine 727 residue and the localization of STAT-1 in Salmonella-infected human monocytic cells. In addition, we studied the role of signaling molecule double-stranded RNA activated protein kinase (PKR) in these modulatory effects. U937 human monocytic cell transfectants stably expressing wild type HLA-B27 or mutated HLA-B27 heavy chains with amino acid substitutions in the B pocket were prepared. The PMA-differentiated cells were infected with S. enteritidis. Western blotting was used to detect the phosphorylation of STAT-1, and to visualize the localization of STAT-1 in the cells confocal microscopy was used. Specific inhibitors were employed to study the role of PKR in STAT-1 phosphorylation. We discovered that the phosphorylation of STAT-1 serine 727 is prolonged in cells expressing misfolding forms of HLA-B27 after S. enteritidis infection, whereas in mock cells and in cells expressing mutated, non-misfolding HLA-B27 the phosphorylation of serine 727 is transient. Interestingly, STAT-1 serine 727 phosphorylation is partly dependent on PKR. In addition, more STAT-1 is localized in the nucleus of HLA-B27-expressing cells, even before an external trigger, when compared to mock cells. In conclusion, our results show that the phosphorylation of STAT-1 serine 727 residue is prolonged in HLA-B27-expressing monocyte-macrophage U937 cells after bacterial infection. This is of interest since the phosphorylation of serine 727 on STAT-1 is suggested to contribute to macrophage activation and promote inflammatory responses. Therefore, our results provide a mechanism which explains how the expression of an HLA-B27 molecule can impact the course of Salmonella infection and reactive arthritis. PMID:23349666

Ruuska, Marja; Sahlberg, Anna S; Granfors, Kaisa; Penttinen, Markus A

2013-01-01

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A comparison of self-reported joint symptoms following infection with different enteric pathogens: effect of HLA-B27  

DEFF Research Database (Denmark)

OBJECTIVE: We conducted a case-case comparison study to estimate the attack-rate of reactive joint pain (JPrea) following intestinal infections, and evaluated whether the susceptibility and severity of joint symptoms was associated with the tissue-type HLA-B27. METHODS: Consecutive patients with positive fecal culture for Salmonella, Campylobacter, Yersinia, Shigella, and E. coli were addressed by questionnaires inquiring about gastrointestinal (GI) symptoms and the occurrence of joint pain in a previously healthy joint within 4 weeks after onset of infection. A blood sample was requested for HLA-B27 typing. RESULTS: Of 3146 patients invited, 2105 (67%) responded to the survey questionnaire. The triggering infections were Campylobacter, 1003; Salmonella, 619; E. coli, 290; Shigella, 102; and Yersinia, 91. JPrea was reported by 294 subjects: Campylobacter, 131 (13.1%); Salmonella, 104 (16.8%); Yersinia, 21 (23.1%); Shigella, 10 (9.8%); and E. coli, 28 (9.7%). There was a significant association between severity of gastroenteritis and development of arthralgia (p = 0.001). The odds ratio (OR) for JPrea in an HLA-B27-positive individual was 2.62 (95% CI 1.67-3.93) for the entire group. A significant association between JPrea and HLA-B27 was found for Salmonella, Shigella, and Yersinia; not, however, for Campylobacter and E. coli. HLA-B27-positive patients had a significantly increased risk for severe joint symptoms. CONCLUSION: Our study shows that JPrea after GI infection is positively correlated to severity of GI symptoms. HLA-B27 is not associated with joint pain after Campylobacter. Intestinal E. coli seems to be an arthritogenic pathogen. A significant association between HLA-B27 and severity of joint pain was observed Udgivelsesdato: 2008/3

Schiellerup, P.; Krogfelt, K.A.

2008-01-01

 
 
 
 
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The epidemiology of back pain, axial spondyloarthritis and HLA-B27 in the United States.  

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The concept of inflammatory back pain (IBP) evolved in the 1970s, coincident with the discovery of the HLA-B27 association with ankylosing spondylitis (AS), leading to the development of criteria to determine the presence of IBP. The concept of IBP and it relationship with AS and axial spondyloarthritis (AxSpA) has further evolved, and an instrument developed (the Spondylitis Association of America Back Pain Tool), which was further modified and field tested for use in the 2009 to 2010 National Health and Nutrition Examination Survey (NHANES). This has shown the frequency of chronic back pain to have risen to 19.4%, with nearly one third having IBP. The prevalence of AxSpA has been defined at 1.0% to 1.4% and AS at 0.52% to 0.55%. The national prevalence of HLA-B27 in the United States is 6.1%, and intriguing data from NHANES 2009 suggest a decreasing frequency with increasing age. From this arise new questions and a work agenda ahead. PMID:23841117

Reveille, John D; Weisman, Michael H

2013-06-01

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HLA-A*01:03, HLA-A*24:02, HLA-B*08:01, HLA-B*27:05, HLA-B*35:01, HLA-B*44:02, and HLA-C*07:01 Monochain Transgenic/H-2 Class I Null Mice : Novel Versatile Preclinical Models of Human T Cell Responses  

DEFF Research Database (Denmark)

We have generated a panel of transgenic mice expressing HLA-A*01:03, -A*24:02, -B*08:01, -B*27:05, -B*35:01, -B*44:02, or -C*07:01 as chimeric monochain molecules (i.e., appropriate HLA ?1?2 H chain domains fused with a mouse ?3 domain and covalently linked to human ?2-microglobulin). Whereas surface expression of several transgenes was markedly reduced in recipient mice that coexpressed endogenous H-2 class I molecules, substantial surface expression of all human transgenes was observed in mice lacking H-2 class I molecules. In these HLA monochain transgenic/H-2 class I null mice, we observed a quantitative and qualitative restoration of the peripheral CD8(+) T cell repertoire, which exhibited a TCR diversity comparable with C57BL/6 WT mice. Potent epitope-specific, HLA-restricted, IFN-?-producing CD8(+) T cell responses were generated against known reference T cell epitopes after either peptide or DNA immunization. HLA-wise, these new transgenic strains encompass a large proportion of individuals from all major human races and ethnicities. In combination with the previously created HLA-A*02:01 and -B*07:02 transgenic mice, the novel HLA transgenic mice described in this report should be a versatile preclinical animal model that will speed up the identification and optimization of HLA-restricted CD8(+) T cell epitopes of potential interest in various autoimmune human diseases and in preclinical evaluation of T cell-based vaccines.

Boucherma, Rachid; Kridane-Miledi, Hédia

2013-01-01

23

Dominant influence of an HLA-B27 restricted CD8+ T cell response in mediating HCV clearance and evolution.  

Science.gov (United States)

Virus-specific CD8+ T cell responses play an important role in the natural course of infection; however, the impact of certain CD8+ T cell responses in determining clinical outcome has not been fully defined. A well-defined cohort of women inoculated with HCV from a single source showed that HLA-B27 has a strong association with spontaneous clearance. The immunological basis for this association is unknown. However, the finding is especially significant because HLA-B27 has also been shown to have a protective role in HIV infection. We report the identification of an HLA-B27 restricted hepatitis C virus (HCV)-specific CD8+ T cell epitope that is recognized in the majority of recovered HLA-B27 positive women. In chronically HCV-infected individuals, analysis of the corresponding viral sequence showed a strong association between sequence variations within this epitope and expression of HLA-B27, indicating allele-specific selection pressure at the population level. Functional analysis in 3 chronically HCV-infected patients showed that the emerging variant viral epitopes represent escape mutations. In conclusion, our results suggest a dominant role of HLA-B27 in mediating spontaneous viral clearance as well as viral evolution in HCV infection and mechanistically link both associations to a dominant novel CD8+ T cell epitope. These results support the central role of virus-specific CD8+ T cells and the genetically determined restriction of the virus-specific T cell repertoire in HCV infection. PMID:16496339

Neumann-Haefelin, Christoph; McKiernan, Susan; Ward, Scott; Viazov, Sergei; Spangenberg, Hans Christian; Killinger, Thomas; Baumert, Thomas F; Nazarova, Natalja; Sheridan, Isabelle; Pybus, Oliver; von Weizsäcker, Fritz; Roggendorf, Michael; Kelleher, Dermot; Klenerman, Paul; Blum, Hubert E; Thimme, Robert

2006-03-01

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Ankylosing spondylitis and multiple sclerosis in an HLA-B27 negative patient.  

Science.gov (United States)

A 41-year-old man presented with vertigo and gait disturbance. He gave a 10-year history of definite ankylosing spondylitis with low back pain, limitation of spinal mobility, decreased chest expansion and radiological evidence of bilateral sacroiliitis. The vertigo attacks started 3 years before and he had insidious evolution of bilateral leg weakness, increased muscle tension and walking disability during the past 2 years. The HLA haplotypes of the patient were A2, A33, B14, B49, Bw4, Bw6, Cw7 and he was HLA-B27 negative. The axial and sagittal cranial magnetic resonance imaging (MRI) showed multiple foci of increased signal intensity in the periventricular white matter and cerebellar hemispheres, suggesting a demyelinating disease process. The MRI of the spine showed centromedullar high intensity lesions at C7, Th7-8, Th9-10 levels. The diagnosis was definite MS (primary progressive MS) as the patient had insidious neurological progression, CSF evidence of inthrathecal production of oligoclonal bands, conduction defects at VEP, multiple brain and additional spinal cord lesions on MRI and continued progression for more than 1 year. PMID:15742608

Tan, Funda Uysal; Tellio?lu, Serdar; Aydin, Gülümser; Erdemo?lu, Ali Kemal; Kele?, I?ik

2004-12-01

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A new HLA-B*27 allele (B*2719) identified in a Lebanese patient affected with ankylosing spondylitis.  

Science.gov (United States)

Eighteen different HLA-B*27 alleles (B*2701-B2718) have so far been recognized by the WHO Nomenclature Committee for Factors of the HLA System. Frequency and disease association of these alleles with spondyloarthropathies differ among ethnic groups. We describe here a novel HLA-B*27 subtype identified in a Lebanese patient suffering from ankylosing spondylitis (AS). This new variant differs from the common HLA-B*2705 DNA sequence at five different nucleotide positions. These nucleotide changes lead to three amino acid differences in the alpha2 domain; Thr to Ile at position 94, Leu to Ile at position 95 and Asn to Arg at position 97. Since this novel allele is encountered in an AS patient, the associated sequence changes are not expected to affect significantly neither the presentation of a putative arthritogenic peptide nor the conformation-dependent recognition by effector cells. PMID:11580853

Tamouza, R; Mansour, I; Bouguacha, N; Klayme, S; Djouadi, K; Laoussadi, S; Azoury, M; Dulphy, N; Ramasawmy, R; Krishnamoorthy, R; Toubert, A; Naman, R; Charron, D

2001-07-01

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Structural analysis of an HLA-B27 functional variant, B27d detected in American blacks  

International Nuclear Information System (INIS)

The structure of a new functional variant B27d has been established by comparative peptide mapping and radiochemical sequencing. This analysis complete the structural characterization of the six know histocompatibility leukocyte antigen (HLA)-B27 subtypes. The only detected amino acid change between the main HLA-B27.1 subtype and B27d is that of Try59 to His59. Position 59 has not been previously found to vary among class I HLA or H-2 antigens. Such substitution accounts for the reported isoelectric focusing pattern of this variant. HLA-B27d is the only B27 variant found to differ from other subtypes by a single amino acid replacement. The nature of the change is compatible with its origin by a point mutation from HLB-B27.1. Because B27d was found only American blacks and in no other ethnic groups, it is suggested that this variant originated as a result of a mutation of the B27.1 gene that occurred within the black population. Structural analysis of B27d was done by comparative mapping. Radiochemical sequencing was carried out with 14C-labeled and 3H-labeled amino acids

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Microarray Analysis of Response of Salmonella during Infection of HLA-B27- Transfected Human Macrophage-Like U937 Cells  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Human leukocyte antigen (HLA-B27 is strongly associated with the development of reactive arthritis (ReA in humans after salmonellosis. Human monocytic U937 cells transfected with HLA-B27 are less able to eliminate intracellular Salmonella enterica serovar Enteritidis than those transfected with control HLA antigens (e.g. HLA-A2. To investigate further the mechanisms by which HLA-B27-transfected cells allow increased replication of these bacteria, a DNA-based microarray was used for comparative genomic analysis of S. Enteritidis grown in HLA-B27- or HLA-A2-transfected cells. The microarray consisted of 5080 oligonucleotides from different serovars of Salmonella including S. Enteritidis PT4-specific genes. Bacterial RNA was isolated from the infected HLA-B27- or HLA-A2-transfected cells, reverse-transcribed to cDNA, and hybridized with the oligonucleotides on the microarrays. Some microarray results were confirmed by RT-PCR. Results When gene expression was compared between Salmonella grown in HLA-B27 cells and in HLA-A2 cells, 118 of the 4610 S. Enteritidis-related genes differed in expression at 8 h after infection, but no significant difference was detectable at 2 h after infection. These differentially expressed genes are mainly involved in Salmonella virulence, DNA replication, energy conversion and metabolism, and uptake and metabolism of nutrient substances, etc. The difference suggests HLA-B27-dependent modulation of Salmonella gene expression, resulting in increased Salmonella replication in HLA-B27-positive cells. Among the up-regulated genes were those located in Salmonella pathogenicity island (SPI-2, which play a central role in intracellular survival and replication of Salmonella. Conclusions This is the first report to show the regulation of Salmonella gene expression by HLA-B27 during infection of host cells. This regulation probably leads to increased Salmonella survival and replication in HLA-B27-positive cells. SPI-2 genes seem to contribute significantly to the increased replication.

Hinton Jay CD

2010-07-01

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HLA-B27 and ankylosing spondylitis geographic distribution as the result of a genetic selection induced by malaria endemic? A review supporting the hypothesis.  

Science.gov (United States)

The geographic distribution of HLA-B27 shows a latitude-related gradient inverse to that of malaria endemic. An apparent exception occurs in New Guinea, a region where malaria is present, but where HLA-B27 frequency shows, however, an orographic gradient antithetic to that of malaria incidence. We therefore suggest that Plasmodium falciparum may have exerted a negative selection on this gene. This might be due to a higher susceptibility to severe forms of malaria, associated with HLA-B27 or other close gene(s). In addition, we suggest here that the same selective pressure that has contributed to reduce the HLA-B27 frequency in some regions has favoured the fixing of newly generated B27 subtypes included in more advantageous HLA haplotypes. In some cases, as for B*2709 in Sardinia and B*2706 in Southeast Asia, these haplotypes may harbour factors that protect from Ankylosing Spondylitis, an autoimmune disease strongly associated with HLA-B27, thus offering a novel, powerful tool to dissect disease pathogenesis, and to identify additional genetic factors of susceptibility. PMID:18486928

Mathieu, Alessandro; Cauli, Alberto; Fiorillo, Maria Teresa; Sorrentino, Rosa

2008-05-01

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PECULIARITIES OF BLOOD CYTOKINE SPECTRUM IN THE PATIENTS WITH REACTIVE ARTHRITIS DUE TO ETIOLOGY, INFLAMMATION ACTIVITY AND PRESENCE OF HLA-B27 ANTIGEN  

Directory of Open Access Journals (Sweden)

Full Text Available The aim of our study was to investigate the cytokine profile due to the etiology, inflammation activity, and presence of HLA-B27 antigen in the patients with reactive arthritis. The study showed a direct dependence of IL-4, IL-6 and the TNF-? on the degree of activity of ReA with chronic pyelonephritis. The presence of HLA-B27 antigen in the patients with reactive arthritis and chronic pyelonephritis accompanied by an increasing of proinflammatory cytokines such as IL-1?, IL-6, PNP-?, IL-1R in comparison with the HLA-B27-negative patients. The results of study demonstrated that maximum serum levels of IL-6 and IFN-? in the patients with reactive arthritis that occurred on the background of acute urogenital infection were significantly higher than in the other etiological cases of reactive arthritis.

Khukhlina O. S.

2013-07-01

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Analyse von T-Zell Subpopulationen hinsichtlich Frequenz und spezifischer Zytokinsekretion in HLA B27-positiven AS-Patienten und Kontrollen  

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The association of Ankylosing Spondylitis(AS) and HLA B27 is the strongest association of an autoimmune disease with a certain MHC known so far. Therefore different hypotheses have been developped to explain this conncetion, but still the pathogenensis of this disease remains unknown. In a previous study it has been shown, that HLA B27 positive healthy donors and patients suffering from AS have a lower expression of proinflammatroy cytokines in CD4+ and CD8+ T-cells. Aim of the current st...

Kohler, Siegfried

2010-01-01

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HLA-B27-associated spondyloarthritis and enthesopathy in childhood: clinical, pathologic, and radiographic observations in 58 patients.  

Science.gov (United States)

HLA-B27 typing of all arthritic children helped to identify and focus attention on a subset whose disease was pathogenetically related to and demonstrated clinical features of ankylosing spondylitis and Reiter syndrome, but only rarely fulfilled current diagnostic criteria for those disorders (spondyloarthritis). In contrast to other forms of childhood arthritis, enthesopathy (inflammation at the sites of attachment of ligaments and tendons to bone) was a prominent feature in 75%; a family history of similar arthritis was obtained from 60%; boys were more frequently affected (2:1); urethritis, acute iritis, conjunctivitis, or keratoderma blennorrhagicum occurred at some time in 42%; and the initial attack followed an unexplained febrile illness, known dysentery or urethritis, or severe musculoskeletal trauma in 41%. The arthritis was generally pauciarticular, asymmetric, and primarily in the feet and large joints of the lower extremities. Distinctive radiographic features included periostitis, severe osteopenia, calcaneal erosions, and heel spurs; three of 58 had rapid destruction of a single joint. Only ten patients (all boys) were found to have radiographic sacroiliitis after an average of five years of disease, and only three had the Reiter triad. The lifetime risk of sacroiliitis and spinal ankylosis can only be determined by long-term follow-up of such prospectively identified groups of spondyloarthritic children. PMID:6977633

Jacobs, J C; Berdon, W E; Johnston, A D

1982-04-01

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Susceptibility to ankylosing spondylitis is independent of the Bw4 and Bw6 epitopes of HLA-B27 alleles.  

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We have characterized HLA-B27 alleles in a sample of the population from the Azores (n=46) with the aim of investigating the contribution of different subtypes to ankylosing spondylitis (AS). The study was carried out using PCR-SSOP and in some samples genomic sequencing was conducted. Some significant new finding have arisen from this study. First, B*2705,B*2702,B*2703,B*2707 and B*2708 alleles were found to be represented in this population. The polymorphism of B27 alleles found in a sample of the population from the Azores is higher than the Caucasian groups described. B*2703 and B*2707 have not previously been described to be represented in Caucasians and this could indicate admixtures with different populations of the world. In addition, the B*2708 allele was found to be associated with AS in a large family from the Azores. This association has not been previously reported in either ethnic group and needs to be confirmed in other population studies. This is of considerable interest since has only been described as a rare subtype underrepresented in the British population and has not been previously found to be associated with AS. B*2708 carries the sequence specifying the Bw6 epitope in contrast to most B27 alleles which carry a Bw4 sequence. Differences in this region (residues 77-83) can alter the F-pocket and affect T-cell recognition. The importance that these molecular changes can play in the pathogenesis of AS is discussed. PMID:10203016

Armas, J B; Gonzalez, S; Martinez-Borra, J; Laranjeira, F; Ribeiro, E; Correia, J; Ferreira, M L; Toste, M; López-Vazquez, A; López-Larrea, C

1999-03-01

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Superior oblique tendon (Brown's) syndrome as the presenting finding in childhood onset HLA-B27-related enthesitis and juvenile idiopathic oligoarticular arthritis.  

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We report two patients who presented with Brown's syndrome. The first is a 7-year-old boy who at the time of his diagnosis was also found to have enthesitis and HLA-B27 positivity. The second patient was diagnosed with bilateral Brown's syndrome at 13 months of age. At age 7 she developed a persistent oligoarticular arthritis and unilateral anterior iritis consistent with the oligoarticular Juvenile Idiopatic Arthritis (JIA) phenotype. These cases highlight ophthalmologic findings and diagnostic considerations with respect to Brown's syndrome and associated childhood onset rheumatologic disease. PMID:25376959

Pham, C; Utz, V; Marcotty, A; Zeft, A; Rychwalski, P

2014-01-01

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HLA-B27 Homodimers and Free H Chains Are Stronger Ligands for Leukocyte Ig-like Receptor B2 than Classical HLA Class 1  

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Possession of HLA-B27 (B27), strongly predisposes to the development of spondyloarthritis. B27 forms classical heterotrimeric complexes with beta-2-microglobulin (?2m) and peptide, and (?2m–free) free H chain (FHC) forms including B27 dimers (termed B272) at the cell surface. In this study we characterise the interaction of HLA-B27 with LILR, leukocyte Ig-like receptor (LILR)B1 and LILRB2 biophysically, biochemically and by FACS staining. LILRB1 bound to B27 heterotrimers with a KD of 5.3 ±1.5 ?M but did not bind B27 FHC. LILRB2 bound to B272 and B27 FHC and B27 heterotrimers with KDs of 2.5, 2.6 and 22 ±6?M respectively. Domain exchange experiments showed that B272 bound to the two membrane distal Ig-like domains of LILRB2. In FACS staining experiments, B27 dimer protein and tetramers stained LILRB2 transfectants five times more strongly than B27 heterotrimers. Moreover, LILRB2Fc bound to dimeric and other B27 FHC forms on B27-expressing cell lines more strongly than other HLA-class I FHCs. B27 transfected cells expressing B27 dimers and FHC inhibited IL-2 production by LILRB2-expressing reporter cells to a greater extent than control HLA-class I transfectants. B27 heterotrimers complexed with the L6M variant of the GAG KK10 epitope bound with a similar affinity to complexes with the wild-type KK10 epitope (with KDs of 15.0±0.8 ?M and 16.0±2.0 ?M respectively). Disulfide-dependent B27 H chain dimers and multimers are stronger ligands for LILRB2 than HLA-class I heterotrimers and H chains. The stronger interaction of B27 dimers and FHC forms with LILRB2 compared with other HLA class I could play a role in spondyloarthritis pathogenesis. PMID:22593621

Giles, Joanna; Shaw, Jackie; Piper, Christopher; Wong-Baeza, Isabel; McHugh, Kirsty; Ridley, Anna; Li, Demin; Lenart, Izabela; Antoniou, Antony N.; DiGleria, Katilin; Kuroki, Kimiko; Maenaka, Katsumi; Bowness, Paul; Kollnberger, Simon

2013-01-01

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Peptide-binding motifs associated with MHC molecules common in Chinese rhesus macaques are analogous to those of human HLA supertypes and include HLA-B27-like alleles  

DEFF Research Database (Denmark)

Chinese rhesus macaques are of particular interest in simian immunodeficiency virus/human immunodeficiency virus (SIV/HIV) research as these animals have prolonged kinetics of disease progression to acquired immunodeficiency syndrome (AIDS), compared to their Indian counterparts, suggesting that they may be a better model for HIV. Nevertheless, the specific mechanism(s) accounting for these kinetics remains unclear. The study of major histocompatibility complex (MHC) molecules, including their MHC/peptide-binding motifs, provides valuable information for measuring cellular immune responses and deciphering outcomes of infection and vaccine efficacy. In this study, we have provided detailed characterization of six prevalent Chinese rhesus macaque MHC class I alleles, yielding a combined phenotypic frequency of 29 %. The peptide-binding specificity of two of these alleles, Mamu-A2*01:02 and Mamu-B*010:01, as well as the previously characterized allele Mamu-B*003:01 (and Indian rhesus Mamu-B*003:01), was found tobe analogous to that of alleles in the HLA-B27 supertype family. Specific alleles in the HLA-B27 supertype family, including HLA-B*27:05, have been associated with long-term nonprogression to AIDS in humans. All six alleles characterized in the present study were found to have specificities analogous to HLA supertype alleles. These data contribute to the concept that Chinese rhesus macaque MHC immunogenetics is more similar to HLA than their Indian rhesus macaque counterparts and thereby warrants further studies to decipher the role of these alleles in the context of SIV infection.

Mothé, Bianca R.; Southwood, Scott

2013-01-01

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HLA class I-mediated control of HIV-1 in the Japanese population, in which the protective HLA-B*57 and HLA-B*27 alleles are absent.  

Science.gov (United States)

We investigated the effect of HLA class I alleles on clinical parameters for HIV-1 disease progression in the Japanese population, where two strongly protective alleles, HLA-B*57 and HLA-B*27, are virtually nonexistent. HLA-B alleles showed a dominant role, primarily through HLA-B*67:01 and the HLA-B*52:01-C*12:02 haplotype. Neither a rare-allele nor a heterozygote advantage was found, suggesting that the effect of HLA alleles in the Japanese population is either different from those observed in Africans and Caucasians or undetectable due to limited power. PMID:22811530

Naruto, Takuya; Gatanaga, Hiroyuki; Nelson, George; Sakai, Keiko; Carrington, Mary; Oka, Shinichi; Takiguchi, Masafumi

2012-10-01

37

Transgenic rat models of vasopressin overexpression.  

Science.gov (United States)

Vasopressin has an important role in water metabolism and its impairment induces some clinical disorders such as diabetes insipidus or syndrome of inappropriate antidiuresis (SIAD). SIAD is caused by the overproduction of vasopressin which induces diluting hyponatremia. The accurate diagnosis and appropriate therapy have not settled up to date because its pathophysiology is very complicated. It is meaningful to develop a rat model of SIAD in which human vasopressin gene is overexpressed in order to analyze pathophysiological changes. Several models transgenic for vasopressin including us had been generated. The transgenic rats provide a useful model to investigate various pathophysiological changes resulting from the oversecretion of vasopressin. Some interesting results based on these animal models are reviewed. PMID:14727685

Oiso, Yutaka; Nagasaki, Hiroshi; Yokoi, Hisashi

2003-11-01

38

Generation of transgenic rats through induced pluripotent stem cells.  

Science.gov (United States)

The rat is an important animal model for human disease research. Using inhibitors of glycogen synthase kinase 3 and MAPK signaling pathways, rat embryonic stem cells and rat induced pluripotent stem cells (riPSCs) have been derived. However, unlike rat embryonic stem cells, germ line competent riPSCs have only been derived from Wistar rats at low efficiency. Here, we found that an optimized induction medium containing knock-out serum replacement and vitamin C improved the rate and efficiency of riPSCs generation from Dark Agouti rat fibroblasts and Sertoli cells. riPSCs maintained an undifferentiated status for >30 passages and could differentiate into various cells types including germ cells when injected into rat blastocysts. Moreover, transgenic riPSCs could be generated through the PiggyBac transposon, which could be used to generate transgenic rats through germ line transmission. riPSCs can be used as a novel tool in genetic and genomic studies of the rat. PMID:23926100

Jiang, Ming-Gui; Li, Tianda; Feng, Chunjing; Fu, Rui; Yuan, Yan; Zhou, Quan; Li, Xin; Wan, Haifeng; Wang, Liu; Li, Wei; Xiao, Yamei; Zhao, Xiao-Yang; Zhou, Qi

2013-09-20

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Conditional gene expression systems in the transgenic rat brain  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Turning gene expression on and off at will is one of the most powerful tools for the study of gene function in vivo. While several conditional systems were successful in invertebrates, in mice the Cre/loxP recombination system and the tet-controlled transcription activation system are predominant. Both expression systems allow for spatial and temporal control of gene activities, and, in the case of tet regulation, even for the reversible activation/inactivation of gene expression. Although the rat is the principal experimental model in biomedical research, in particular in studies of neuroscience, conditional rat transgenic systems are exceptionally rare in this species. Results We addressed this lack of technology, and established and thoroughly characterized CreERT2 and tTA transgenic rats with forebrain-specific transgene expression, controlled by the CaMKII alpha promoter. In addition, we developed new universal rat reporter lines for both transcription control systems and established inducible and efficient reporter gene expression in forebrain neurons. Conclusions We demonstrate that conditional genetic manipulations in the rat brain are both feasible and practicable and outline advantages and limitations of the Tet and Cre/loxP system in the rat brain.

Schönig Kai

2012-09-01

40

Lanolin as a treatment option for ringtail in transgenic rats.  

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Ringtail is a condition characterized by dry skin and annular constrictions that sometimes result in loss of portions of the tail. This condition most commonly affects preweanling rats, and low relative humidity is thought to be a principal cause. The use of transgenic rats in our facility has been increasing since 2002, and we recently diagnosed several litters from transgenic Fischer 344 rats (Rattus norvegicus) with ringtail. Treatment was necessary to maintain the health and integrity of the tails to allow genotyping. Lanolin ointment was chosen because it is a nontoxic, inexpensive, effective moisturizer used for treating human skin conditions. We examined 5 litters comprising 37 pups total, ranging in age from 7 to 17 days at the time of presentation. Animals in 3 litters were randomly assigned to a treatment or nontreatment group, and all animals in the remaining 2 litters were treated. Lanolin was applied to the tails of treatment groups once daily for 6 d. Treatment was tolerated well by pups and no animals were rejected by the dams. After treatment, tail condition was scored from 0 to 3, with 0 representing a tail normal in appearance, and 3 representing severe disease. Chi square testing showed marginal statistical significance, with a trend for a higher percentage of treated rats having healthier tails on day 7 compared to untreated pups. The Pearson correlation between treatment and tail condition scores was significant. Results indicate that lanolin was an efficacious treatment option for ringtail. PMID:16539341

Taylor, Douglas K; Rogers, Melissa M; Hankenson, F Claire

2006-01-01

 
 
 
 
41

HIV-1 transgenic rats develop T cell abnormalities  

International Nuclear Information System (INIS)

HIV-1 infection leads to impaired antigen-specific T cell proliferation, increased susceptibility of T cells to apoptosis, progressive impairment of T-helper 1 (Th1) responses, and altered maturation of HIV-1-specific memory cells. We have identified similar impairments in HIV-1 transgenic (Tg) rats. Tg rats developed an absolute reduction in CD4+ and CD8+ T cells able to produce IFN-? following activation and an increased susceptibility of T cells to activation-induced apoptosis. CD4+ and CD8+ effector/memory (CD45RC-CD62L-) pools were significantly smaller in Tg rats compared to non-Tg controls, although the converse was true for the naieve (CD45RC+CD62L+) T cell pool. Our interpretation is that the HIV transgene causes defects in the development of T cell effector function and generation of specific effector/memory T cell subsets, and that activation-induced apoptosis may be an essential factor in this process

42

Neurohypophyseal and fluid homeostasis in transgenic rats expressing a tagged rat vasopressin prepropeptide in hypothalamic neurons.  

Science.gov (United States)

We have developed a transgenic system that, for the first time, facilitates monitoring of the regulatory dynamics of a central peptidergic system from transcription of a neuropeptide gene to the storage and release of the mature secretory product. A rat vasopressin (VP) transgene (5-VCAT-3), the expression of which is restricted to hypothalamic vasopressinergic magnocellular neurons in rats, contains a sequence that, if translated, would place a unique hexadecapeptide (DRSAGYYGLFKDRKEK, abbreviated to DR-12-EK) at the C-terminus of the VP precursor. We have raised an antibody against this "tag" and, using immunohistochemistry, electron microscopy, RIA, and HPLC, have shown for the first time that a VP transgene RNA is translated into a protein product found, in a processed form, in secretory granules in the posterior pituitaries of transgenic rats. Disruption of the C-terminus of the VP precursor by the peptide tag is well tolerated and does not disrupt VP production or disturb salt and water balance. An osmotic stimulus increased hypothalamic DR-12-EK levels, but changes in posterior pituitary DR-12-EK levels were more complex. After 5 days of salt-loading, DR-12-EK levels fell, as would be expected if its release was coordinate with that of VP. However, after 10 days of salt-loading, posterior pituitary DR-12-EK levels increased, despite the lower level of VP. This probably reflects the greater response of the transgene to osmotic challenge at the RNA level, increasing the proportion of DR-12-EK-containing translation products transported to the posterior pituitary relative to those derived from the endogenous gene. The exaggerated response of the tagged transgene to osmotic challenge at both RNA and protein levels affords a new opportunity to study the regulatory dynamics of the VP system at the molecular level, but within the physiologically advantageous context of the intact animal. PMID:8895381

Waller, S; Fairhall, K M; Xu, J; Robinson, I C; Murphy, D

1996-11-01

43

Transgenesis and neuroendocrine physiology: a transgenic rat model expressing growth hormone in vasopressin neurones.  

Science.gov (United States)

Human growth hormone (hGH) and bovine neurophysin (bNP) DNA reporter fragments were inserted into the rat vasopressin (VP) and oxytocin (OT) genes in a 44 kb cosmid construct used to generate two lines of transgenic rats, termed JP17 and JP59. Both lines showed specific hGH expression in magnocellular VP cells in the hypothalamic paraventricular (PVN) and supraoptic nuclei (SON). hGH was also expressed in parvocellular neurones in suprachiasmatic nuclei (SCN), medial amygdala and habenular nuclei in JP17 rats; the rat OT-bNP (rOT-bNP) transgene was not expressed in either line. Immunohistochemistry and radioimmunoassay showed hGH protein in the hypothalamus from where it was transported in varicose fibres via the median eminence to the posterior pituitary gland. Immunogold electron microscopy showed hGH co-stored with VP-NP in the same granules. The VP-hGH transgene did not affect water balance, VP storage or release in vivo. Drinking 2 % saline for 72 h increased hypothalamic transgene hGH mRNA expression, and depleted posterior pituitary hGH and VP stores in parallel. In anaesthetised, water-loaded JP17 rats, hGH was released with VP in response to an acute hypovolumic stimulus (sodium nitrosopentacyano, 400 microg I.V.). JP17 rats had a reduced growth rate, lower anterior pituitary rGH contents, and a reduced amplitude of endogenous pulsatile rGH secretion assessed by automated blood microsampling in conscious rats, consistent with a short-loop feedback of the VP-hGH on the endogenous GH axis. This transgenic rat model enables us to study physiological regulation of hypothalamic transgene protein production, transport and secretion, as well as its effects on other neuroendocrine systems in vivo. PMID:12813157

Wells, Sara E; Flavell, David M; Bisset, Gordon W; Houston, Pamela A; Christian, Helen; Fairhall, Keith M; Robinson, Iain C A F

2003-08-15

44

Transgenic rats with green, red, and blue fluorescence: powerful tools for bioimaging, cell trafficking, and differentiation  

Science.gov (United States)

The rat represents a perfect animal for broadening medical experiments, because its physiology has been well understood in the history of experimental animals. In addition, its larger body size takes enough advantage for surgical manipulation, compared to the mouse. Many rat models mimicking human diseases, therefore, have been used in a variety of biomedical studies including physiology, pharmacology, transplantation, and immunology. In an effort to create the specifically designed rats for biomedical research and regenerative medicine, we have developed the engineered rat system on the basis of transgenic technology and succeeded in establishing various transgenic rat strains. The transgenic rats with green fluorescent protein (GFP) were generated in the two different strains (Wistar and Lewis), in which GFP is driven under the chicken beta-actin promoter and cytomegalovirus enhancer (CAG promoter). Their GFP expression levels were different in each organ, but the Lewis line expressed GFP strongly and ubiquitously in most of the organs compared with that of Wistar. For red fluorescence, DsRed2 was transduced to the Wistar rats: one line specifically expresses DsRed2 in the liver under the mouse albumin promoter, another is designed for the Cre/LoxP system as the double reporter rat (the initial DsRed2 expression turns on GFP in the presence of Cre recombinase). LacZ-transgenic rats represent blue color, and LacZ is driven the CAG (DA) or ROSA26 promoter (Lewis). Our unique transgenic rats" system highlights the powerful performance for the elucidation of many cellular processes in regenerative medicine, leading to innovative medical treatments.

Murakami, Takashi; Kobayashi, Eiji

2005-04-01

45

Tet system in the brain: transgenic rats and lentiviral vectors approach.  

Science.gov (United States)

Local and regulated expression of exogenous genes in the central nervous system is one of the major challenges of modern neuroscience. We have approached this issue by applying the inducible tetracycline system to regulate the expression of EGFP reporter gene in double transgenic rats. We have obtained a strong induction of EGFP only in male testes, which correlated with a high level of rtTA expression only in this organ. To overcome the problem of lack of rtTA protein in the transgenic rat brain, we have delivered this Tet system activator with lentiviral vectors into the dentate gyrus of hippocampus of transgenic EGFP rats. As a result, after systemic application of doxycycline we have obtained inducible, stable and restricted to the desired brain region expression of EGFP. An advantage of this strategy is that the transgene is located in the same genetic milieu in every cell of the transgenic organism. This is crucial to obtain uniform expression of the regulated gene within the target brain structure. Combination of rat transgenesis and lentiviral vectors is a novel approach enabling precise spatiotemporal regulation of genes of interest strictly in the brain structure of choice or in other tissues. PMID:19241392

Konopka, Witold; Duniec, Kamila; Klejman, Agata; Wawrzyniak, Marcin; Owczarek, Dorota; Gawrys, Ludwika; Maleszewski, Marek; Mallet, Jacques; Kaczmarek, Leszek

2009-04-01

46

Can organic and transgenic soy be used as a substitute for animal protein by rats?  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We evaluated the protein quality of organic and transgenic soy fed to rats throughout life. Thirty female Wistar rats were divided into three groups (N = 10): organic soy group (OSG) receiving organic soy-based diet, genetically modified soy group (GMSG) receiving transgenic soy-based diet, and a control group (CG) receiving casein-based diet. All animals received water and isocaloric diet (10% protein), ad libitum for 291 days. After this, the weight of GMSG animals (290.9 ± 9.1 g) was sign...

Soares, L. L.; Lucas, A. M. M.; Boaventura, G. T.

2005-01-01

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Development of transgenic rats producing human ?-amyloid precursor protein as a model for Alzheimer's disease: Transgene and endogenous APP genes are regulated tissue-specifically  

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Full Text Available Abstract Background Alzheimer's disease (AD is a devastating neurodegenerative disorder that affects a large and growing number of elderly individuals. In addition to idiopathic disease, AD is also associated with autosomal dominant inheritance, which causes a familial form of AD (FAD. Some instances of FAD have been linked to mutations in the ?-amyloid protein precursor (APP. Although there are numerous mouse AD models available, few rat AD models, which have several advantages over mice, have been generated. Results Fischer 344 rats expressing human APP driven by the ubiquitin-C promoter were generated via lentiviral vector infection of Fischer 344 zygotes. We generated two separate APP-transgenic rat lines, APP21 and APP31. Serum levels of human amyloid-beta (A?40 were 298 pg/ml for hemizygous and 486 pg/ml for homozygous APP21 animals. Serum A?42 levels in APP21 homozygous rats were 135 pg/ml. Immunohistochemistry in brain showed that the human APP transgene was expressed in neurons, but not in glial cells. These findings were consistent with independent examination of enhanced green fluorescent protein (eGFP in the brains of eGFP-transgenic rats. APP21 and APP31 rats expressed 7.5- and 3-times more APP mRNA, respectively, than did wild-type rats. Northern blots showed that the human APP transgene, driven by the ubiquitin-C promoter, is expressed significantly more in brain, kidney and lung compared to heart and liver. A similar expression pattern was also seen for the endogenous rat APP. The unexpected similarity in the tissue-specific expression patterns of endogenous rat APP and transgenic human APP mRNAs suggests regulatory elements within the cDNA sequence of APP. Conclusion This manuscript describes the generation of APP-transgenic inbred Fischer 344 rats. These are the first human AD model rat lines generated by lentiviral infection. The APP21 rat line expresses high levels of human APP and could be a useful model for AD. Tissue-specific expression in the two transgenic rat lines and in wild-type rats contradicts our current understanding of APP gene regulation. Determination of the elements that are responsible for tissue-specific expression of APP may enable new treatment options for AD.

Chan Anthony WS

2008-02-01

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Overexpression of vasopressin in the rat transgenic for the metallothionein-vasopressin fusion gene.  

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Arginine vasopressin (AVP) is a major antidiuretic hormone, the overproduction of which causes diluting hyponatremia in humans and is called the syndrome of inappropriate antidiuresis (SIAD). To study physiological changes resulting from AVP overproduction and to develop an animal model of hyponatremia, the human AVP gene was expressed under the control of the metallothionein promoter in transgenic (Tg) rats. Analyses of AVP immunoreactivity (irAVP) in the tissues revealed that the transgene is expressed mainly in the central nervous system. Gel filtration showed that irAVP in the brain and plasma was properly processed AVP. AVP purified from the brains of both Tg and control rats also exerted equal bioactivity to generate cAMP in LLC-PK1 cells. The founder rats did not show any physical or anatomical abnormalities. Under basal conditions, Tg rats had high plasma AVP levels (Tg 13.8 +/- 2.5 pg/ml; control 2.7 +/- 1.2 pg/ml; n=6 in both groups; means +/- S.E.M.), decreased urine volume, and normal plasma [Na(+)]. Hypertonic saline injected i.p. did not affect AVP secretion in Tg rats. In response to a zinc-supplemented liquid diet, plasma AVP decreased in control rats, but increased in Tg rats (Tg 32.7 +/- 2.7 pg/ml; control 1.0+/-0.1 pg/ml; n=6), resulting in hyponatremia (Tg 135.2 +/- 2.5 mEq/l; control 140.8 +/- 0.4 mEq/l; n=6). To our knowledge, this is the first transgenic animal to show diluting hyponatremia. This transgenic rat may therefore provide a useful model in which to investigate various physiological alterations resulting from the oversecretion of AVP which involve SIAD, stress response, behavior, and blood pressure. PMID:11927382

Nagasaki, H; Yokoi, H; Arima, H; Hirabayashi, M; Ishizaki, S; Tachikawa, K; Murase, T; Miura, Y; Oiso, Y

2002-04-01

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Expression of the Mu Opioid Receptor in the Human Immunodeficiency Virus Type 1 Transgenic Rat Model?  

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Opioids, via the mu opioid receptor (MOR), can exacerbate bacterial infections and the immunopathogenesis of human immunodeficiency virus type 1 (HIV-1) infection. Recently, an HIV-1 transgenic (HIV-1Tg) rat model containing circulating HIV-1 gp120 was created. Using real-time reverse transcription-PCR, we found that MOR mRNA levels were significantly higher in the peritoneal macrophages of the HIV-1Tg rat than those in control animals. Lipopolysaccharide, a bacterial endotoxin, induced secre...

Chang, Sulie L.; Beltran, Jose A.; Swarup, Shilpa

2007-01-01

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Chronic alcohol ingestion exacerbates skeletal muscle myopathy in HIV-1 transgenic rats  

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Full Text Available Abstract Background Separately, chronic alcohol ingestion and HIV-1 infection are associated with severe skeletal muscle derangements, including atrophy and wasting, weakness, and fatigue. One prospective cohort study reported that 41% of HIV-infected patients met the criteria for alcoholism, however; few reports exist on the co-morbid effects of these two disease processes on skeletal muscle homeostasis. Thus, we analyzed the atrophic effects of chronic alcohol ingestion in HIV-1 transgenic rats and identified alterations to several catabolic and anabolic factors. Findings Relative plantaris mass, total protein content, and fiber cross-sectional area were reduced in each experimental group compared to healthy, control-fed rats. Alcohol abuse further reduced plantaris fiber area in HIV-1 transgenic rats. Consistent with previous reports, gene levels of myostatin and its receptor activin IIB were not increased in HIV-1 transgenic rat muscle. However, myostatin and activin IIB were induced in healthy and HIV-1 transgenic rats fed alcohol for 12 weeks. Catabolic signaling factors such as TGF?1, TNF?, and phospho-p38/total-p38 were increased in all groups compared to controls. There was no effect on IL-6, leukemia inhibitory factor (LIF, cardiotrophin-1 (CT-1, or ciliary neurotrophic factor (CNTF in control-fed, transgenic rats. However, the co-morbidity of chronic alcohol abuse and HIV-1-related protein expression decreased expression of the two anabolic factors, CT-1 and CNTF. Conclusions Consistent with previous reports, alcohol abuse accentuated skeletal muscle atrophy in an animal model of HIV/AIDS. While some catabolic pathways known to drive alcoholic or HIV-1-associated myopathies were also elevated in this co-morbid model (e.g., TGF?1, consistent expression patterns were not apparent. Thus, specific alterations to signaling mechanisms such as the induction of the myostatin/activin IIB system or reductions in growth factor signaling via CT-1- and CNTF-dependent mechanisms may play larger roles in the regulation of muscle mass in alcoholic, HIV-1 models.

Bratina Margaux A

2011-08-01

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Nonmammalian gonadotropin-releasing hormone molecules in the brain of promoter transgenic rats  

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Mammalian gonadotropin-releasing hormone (GnRH1) and nonmammalian immunoreactive GnRH subtypes were examined in transgenic rats carrying an enhanced GFP (EGFP) reporter gene driven by a rat GnRH1 promoter. Double-label immunocytochemistry was performed on EGFP+/GnRH1 brain sections by using antisera against GnRH1, GnRH2 (chicken II), GnRH3 (salmon), or seabream GnRH. EGFP+/GnRH1 neurons were in the septal–preoptic hypothalamus but not in the midbrain, consistent with GnRH1-immunopositive ne...

Parhar, Ishwar S.; Soga, Tomoko; Ogawa, Satoshi; Ogawa, Sonoko; Pfaff, Donald W.; Sakuma, Yasuo

2005-01-01

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Retinal Degeneration in Two Lines of Transgenic S334ter Rats  

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Aim of this study was to examine synaptic connectivity changes in the retina and the location and rate of apoptosis in transgenic S334ter line-3 and line-5 rats with photoreceptor degeneration. Heterozygous S334ter-line-3 and line-5 at P11-13, P30, P60, P90 and several control non-dystrophic rats (Long Evans and Sprague-Dawley) at P60, were studied anatomically by immunohistochemistry for various cell and synaptic markers, and by PNA and TUNEL label.- S334ter line-3 exhibited the fastest rate...

Martinez-navarrete, G.; Seiler, M. J.; Aramant, R. B.; Fernandez-sanchez, L.; Pinilla, I.; Cuenca, N.

2011-01-01

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Megaesophagus in a line of transgenic rats: a model of achalasia.  

Science.gov (United States)

Megaesophagus is defined as the abnormal enlargement or dilatation of the esophagus, characterized by a lack of normal contraction of the esophageal walls. This is called achalasia when associated with reduced or no relaxation of the lower esophageal sphincter (LES). To date, there are few naturally occurring models for this disease. A colony of transgenic (Pvrl3-Cre) rats presented with megaesophagus at 3 to 4 months of age; further breeding studies revealed a prevalence of 90% of transgene-positive animals having megaesophagus. Affected rats could be maintained on a total liquid diet long term and were shown to display the classic features of dilated esophagus, closed lower esophageal sphincter, and abnormal contractions on contrast radiography and fluoroscopy. Histologically, the findings of muscle degeneration, inflammation, and a reduced number of myenteric ganglia in the esophagus combined with ultrastructural lesions of muscle fiber disarray and mitochondrial changes in the striated muscle of these animals closely mimic that seen in the human condition. Muscle contractile studies looking at the response of the lower esophageal sphincter and fundus to electrical field stimulation, sodium nitroprusside, and L-nitro-L-arginine methyl ester also demonstrate the similarity between megaesophagus in the transgenic rats and patients with achalasia. No primary cause for megaesophagus was found, but the close parallel to the human form of the disease, as well as ease of care and manipulation of these rats, makes this a suitable model to better understand the etiology of achalasia as well as study new management and treatment options for this incurable condition. PMID:24457157

Pang, J; Borjeson, T M; Muthupalani, S; Ducore, R M; Carr, C A; Feng, Y; Sullivan, M P; Cristofaro, V; Luo, J; Lindstrom, J M; Fox, J G

2014-11-01

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Free access to running wheels abolishes hyperphagia in human growth hormone transgenic rats.  

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Obesity is a major health problem, and increased food intake and decreased physical activity are considered as two major factors causing obesity. Previous studies show that voluntary exercise in a running wheel decreases not only body weight but also food intake of rats. We previously produced human growth hormone transgenic (TG) rats, which are characterized by severe hyperphagia and obesity. To gain more insight into the effects on physical activity to food consumption and obesity, we examined whether voluntary running wheel exercise causes inhibition of hyperphagia and alteration of body composition in TG rats. Free access to running wheels completely abolished hyperphagia in TG rats, and this effect persisted for many weeks as far as the running wheel is accessible. Unexpectedly, though the running distances of TG rats were significantly less than those of wild type rats, it was sufficient to normalize their food consumption. This raises the possibility that rearing environment, which enables them to access to a running wheel freely, rather than the amounts of physical exercises is more important for the maintenance of proper food intake. PMID:24717416

Komatsuda, Mugiko; Yamanouchi, Keitaro; Matsuwaki, Takashi; Nishihara, Masugi

2014-07-01

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Can organic and transgenic soy be used as a substitute for animal protein by rats?  

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Full Text Available We evaluated the protein quality of organic and transgenic soy fed to rats throughout life. Thirty female Wistar rats were divided into three groups (N = 10: organic soy group (OSG receiving organic soy-based diet, genetically modified soy group (GMSG receiving transgenic soy-based diet, and a control group (CG receiving casein-based diet. All animals received water and isocaloric diet (10% protein, ad libitum for 291 days. After this, the weight of GMSG animals (290.9 ± 9.1 g was significantly lower (P <= 0.04 than CG (323.2 ± 7.9 g. The weight of OSG (302.2 ± 8.7 g was between that of the GMSG and the CG. Protein intake was similar for OSG (308.4 ± 6.8 g and GMSG (301.5 ± 2.5 g, and significantly lower (P <= 0.0005 than the CG (358.4 ± 8.1 g. Growth rate was similar for all groups: OSG (0.80 ± 0.02 g, GMSG (0.81 ± 0.03 g and CG (0.75 ± 0.02 g. In addition to providing a good protein intake and inducing less weight gain, both types of soy were utilized in a manner similar to that of casein, suggesting that the protein quality of soy is similar to that of the standard protein casein. The groups fed soy-based diet gained less weight, which may be considered to be beneficial for health. We conclude that organic and transgenic soy can be fed throughout life to rats in place of animal protein, because contain high quality protein and do not cause a marked increase in body weight.

L.L. Soares

2005-04-01

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Effect of HIV-1-related protein expression on cardiac and skeletal muscles from transgenic rats  

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Full Text Available Abstract Background Human immunodeficiency virus type 1 (HIV-1 infection and the consequent acquired immunodeficiency syndrome (AIDS has protean manifestations, including muscle wasting and cardiomyopathy, which contribute to its high morbidity. The pathogenesis of these myopathies remains partially understood, and may include nutritional deficiencies, biochemical abnormalities, inflammation, and other mechanisms due to viral infection and replication. Growing evidence has suggested that HIV-1-related proteins expressed by the host in response to viral infection, including Tat and gp120, may also be involved in the pathophysiology of AIDS, particularly in cells or tissues that are not directly infected with HIV-1. To explore the potentially independent effects of HIV-1-related proteins on heart and skeletal muscles, we used a transgenic rat model that expresses several HIV-1-related proteins (e.g., Tat, gp120, and Nef. Outcome measures included basic heart and skeletal muscle morphology, glutathione metabolism and oxidative stress, and gene expressions of atrogin-1, muscle ring finger protein-1 (MuRF-1 and Transforming Growth Factor-?1 (TGF?1, three factors associated with muscle catabolism. Results Consistent with HIV-1 associated myopathies in humans, HIV-1 transgenic rats had increased relative heart masses, decreased relative masses of soleus, plantaris and gastrocnemius muscles, and decreased total and myosin heavy chain type-specific plantaris muscle fiber areas. In both tissues, the levels of cystine (Cyss, the oxidized form of the anti-oxidant cysteine (Cys, and Cyss:Cys ratios were significantly elevated, and cardiac tissue from HIV-1 transgenic rats had altered glutathione metabolism, all reflective of significant oxidative stress. In HIV-1 transgenic rat hearts, MuRF-1 gene expression was increased. Further, HIV-1-related protein expression also increased atrogin-1 (~14- and ~3-fold and TGF?1 (~5-fold and ~3-fold in heart and plantaris muscle tissues, respectively. Conclusion We provide compelling experimental evidence that HIV-1-related proteins can lead to significant cardiac and skeletal muscle complications independently of viral infection or replication. Our data support the concept that HIV-1-related proteins are not merely disease markers, but rather have significant biological activity that may lead to increased oxidative stress, the stimulation of redox-sensitive pathways, and altered muscle morphologies. If correct, this pathophysiological scheme suggests that the use of dietary thiol supplements could reduce skeletal and cardiac muscle dysfunction in HIV-1-infected individuals.

Guidot David M

2008-04-01

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In vivo genotoxicity of methyleugenol in gpt delta transgenic rats following medium-term exposure.  

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Methyleugenol (MEG), which is commonly used as a fragrance and flavoring agent, has been shown to induce hepatocellular tumors in rodents. However, the role of genotoxicity as a possible mechanism of action is not fully understood even though the DNA-reactive metabolite of MEG has been identified. In this study, a gpt delta transgenic rat model was used to clarify whether genotoxic mechanisms are involved in MEG-induced hepatocarcinogenesis following medium-term exposure. F344 gpt delta rats were subjected to repeated oral administration of MEG at dosages of 0, 10, 30, or 100mg/kg (a carcinogenic dose) for 13 weeks. The relative weight of the liver of the male and female rats that were administered 100mg/kg MEG and the absolute weight of the liver of the male rats that were administered 100mg/kg MEG were significantly increased. In addition, the number and area of glutathione S-transferase placental form (GST-P) positive foci and proliferating cell nuclear antigen (PCNA) positive cell ratios in the hepatocytes were significantly increased in the male and female rats that were administered 100mg/kg MEG compared with the control animals. In the in vivo mutation assays, a significant increase in the gpt and Spi(-) mutant frequencies was observed in both sexes at the carcinogenic dose. These results suggest the possible participation of genotoxic mechanisms in MEG-induced hepatocarcinogenesis. PMID:23074021

Jin, Meilan; Kijima, Aki; Hibi, Daisuke; Ishii, Yuji; Takasu, Shinji; Matsushita, Kohei; Kuroda, Ken; Nohmi, Takehiko; Nishikawa, Akiyoshi; Umemura, Takasi

2013-02-01

58

Angiotensin II induced inflammation in the kidney and in the heart of double transgenic rats  

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Full Text Available Abstract Background We are investigating a double transgenic rat (dTGR model, in which rats transgenic for the human angiotensinogen and renin genes are crossed. These rats develop moderately severe hypertension but die of end-organ cardiac and renal damage by week 7. The heart shows necrosis and fibrosis, whereas the kidneys resemble the hemolytic-uremic syndrome vasculopathy. Surface adhesion molecules (ICAM-1 and VCAM-1 are expressed early on the endothelium, while the corresponding ligands are found on circulating leukocytes. Leukocyte infiltration in the vascular wall accompanies PAI-1, MCP-1, iNOS and Tissue Factor expression. Furthermore we show evidence that Ang II causes the upregulation of NF-kB in our model. Methods We started PDTC-treatment on four weeks old dTGR (200 mg/kg sc and age-matched SD rats.. Blood-pressure- and albuminuria- measurements were monitored during the treatement period (four weeks. The seven weeks old animals were killed, hearts and kidneys were isolated and used for immunohistochemical-and electromobility shift assay analsis. Results Chronic treatment with the antioxidant PDTC decreased blood pressure (162 ± 8 vs. 190 ± 7 mm Hg, p = 0.02. Cardiac hypertrophy index was significantly reduced (4.90 ± 0.1 vs. 5.77 ± 0.1 mg/g, p Conclusion Our data show that inhibition of NF-?B by PDTC markedly reduces inflammation, iNOS expression in the dTGR most likely leading to decreased cytotoxicity, and cell proliferation. Thus, NF-?B activation plays an important role in ANG II-induced end-organ damage.

Haller Hermann

2002-01-01

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A progressive dopaminergic phenotype associated with neurotoxic conversion of ?-synuclein in BAC-transgenic rats  

DEFF Research Database (Denmark)

Conversion of soluble ?-synuclein into insoluble and fibrillar inclusions is a hallmark of Parkinson's disease and other synucleinopathies. Accumulating evidence points towards a relationship between its generation at nerve terminals and structural synaptic pathology. Little is known about the pathogenic impact of ?-synuclein conversion and deposition at nigrostriatal dopaminergic synapses in transgenic mice, mainly owing to expression limitations of the ?-synuclein construct. Here, we explore whether both the rat as a model and expression of the bacterial artificial chromosome construct consisting of human full-length wild-type ?-synuclein could exert dopaminergic neuropathological effects. We found that the human promoter induced a pan-neuronal expression, matching the rodent ?-synuclein expression pattern, however, with prominent C-terminally truncated fragments. Ageing promoted conversion of both full-length and C-terminally truncated ?-synuclein species into insolube and proteinase K-resistant fibres, with strongest accumulation in the striatum, resembling biochemical changes seen in human Parkinson's disease. Transgenic rats develop early changes in novelty-seeking, avoidance and smell before the progressive motor deficit. Importantly, the observed pathological changes were associated with severe loss of the dopaminergic integrity, thus resembling more closely the human pathology.

Nuber, Silke; Harmuth, Florian

2013-01-01

60

Endotoxin-induced cytokine and chemokine expression in the HIV-1 transgenic rat  

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Full Text Available Abstract Background Repeated exposure to a low dose of a bacterial endotoxin such as lipopolysaccharide (LPS causes immune cells to become refractory to a subsequent endotoxin challenge, a phenomenon known as endotoxin tolerance (ET. During ET, there is an imbalance in pro- and anti-inflammatory cytokine and chemokine production, leading to a dysregulated immune response. HIV-1 viral proteins are known to have an adverse effect on the immune system. However, the effects of HIV-1 viral proteins during ET have not been investigated. Methods In this study, HIV-1 transgenic (HIV-1Tg rats and control F344 rats (n = 12 ea were randomly treated with 2 non-pyrogenic doses of LPS (LL to induce ET, or saline (SS, followed by a high challenge dose of LPS (LL+L, SS+L or saline (LL+S, SS+S. The gene expression of 84 cytokines, chemokines, and their receptors in the brain and spleen was examined by relative quantitative PCR using a PCR array, and protein levels in the brain, spleen, and serum of 7 of these 84 genes was determined using an electrochemiluminescent assay. Results In the spleen, there was an increase in key pro-inflammatory (IL1?, IL-1?, IFN-? and anti-inflammatory (IL-10 cytokines, and inflammatory chemokines (Ccl2, Ccl7, and Ccl9, in response to LPS in the SS+L and LL+L (ET groups of both the HIV-1Tg and F344 rats, but was greater in the HIV-1Tg rats than in the F344. In the ET HIV-1Tg and F344 (LL+L rats in the spleen, the LPS-induced increase in pro-inflammatory cytokines was diminished and that of the anti-inflammatory cytokine was enhanced compared to the SS+L group rats. In the brain, IL-1?, as well as the Ccl2, Ccl3, and Ccl7 chemokines were increased to a greater extent in the HIV-1Tg rats compared to the F344; whereas Cxcl1, Cxcl10, and Cxcl11 were increased to a greater extent in the F344 rats compared to the HIV-1Tg rats in the LL+L and SS+L groups. Conclusion Our data indicate that the continuous presence of HIV-1 viral proteins can have tissue-dependent effects on endotoxin-induced cytokine and chemokine expression in the ET state.

Homji Natasha F

2012-01-01

 
 
 
 
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Changes in endovascular trophoblast invasion and spiral artery remodelling at term in a transgenic preeclamptic rat model.  

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As a follow-up to our previous study which revealed a surprisingly deeper endovascular trophoblast (ET) invasion on day 18 in a transgenic preeclamptic (PE) rat model (hAngiotensinogen female symbol x hRenin male symbol) compared to non-PE controls, we examined further changes in ET invasion and associated spiral artery (SA) remodelling at term (day 21). PE transgenic rats and non-PE reversely mated (RM) transgenic rats were compared to normal SD rats (C). Sections were stained to visualize trophoblast, fibrinoid, vascular smooth muscle (VSM) and endothelium. SA were evaluated in three depth levels in the mesometrial triangle (MT) using the KS-400 image analysis system. In separate transgenic rats, Doppler ultrasound was performed in uterine arteries, and the resistance indices (RI) were calculated. Although for the whole MT differences in ET invasion were no longer significant between the PE and C, indicating a partial catching up in C rats, there was still significantly more ET in the deepest level in the PE group as compared to the C and RM groups. At the same time the SA walls in PE rats contained significantly more fibrinoid (versus RM and C) and VSM (versus C). In all SA cross-sections, re-endothelialisation was prominent, but significantly different between PE and C group. The Doppler results showed a significantly lower RI in the arcuate uterine artery of the PE group compared to the C group. There was no evidence of elimination of deeply invaded ET at term, previously considered as a possible mechanism for restriction of vascular remodelling in human PE. The differences in vascular remodelling, previously described on day 18 by histology and Doppler data, were maintained on day 21, but there was extensive endothelial repair in the three groups. Atherosis-like lesions were observed in the three groups, most frequently in the RM group, but were never associated with placental infarcts. PMID:20144482

Geusens, N; Hering, L; Verlohren, S; Luyten, C; Drijkoningen, K; Taube, M; Vercruysse, L; Hanssens, M; Dechend, R; Pijnenborg, R

2010-04-01

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Genotype specific age related changes in a transgenic rat model of Huntington's disease.  

Science.gov (United States)

We aimed to characterize the transgenic Huntington rat model with in vivo imaging and identify sensitive and reliable biomarkers associated with early and progressive disease status. In order to do so, we performed a multimodality (DTI and PET) longitudinal imaging study, during which the same TgHD and wildtype (Wt) rats were repetitively scanned. Surprisingly, the relative ventricle volume was smaller but increased faster in TgHD compared to Wt animals. DTI (mean, axial, radial diffusivity) revealed subtle genotype-specific aging effects in the striatum and its surrounding white matter, already in the presymptomatic stage. Using ¹?F-FDG and ¹?F-Fallypride PET imaging, we were not able to demonstrate genotype-specific aging effects within the striatum. The outcome of this longitudinal study was somewhat surprising as it demonstrated a significant differential aging pattern in TgHD versus Wt animals. Although it seems that the TgHD rat model does not have a sufficient expression of disease yet at the age of 12 months, further validation of this model is highly beneficial since there is still an incomplete understanding of the early disease mechanisms of Huntington's disease. PMID:21767653

Blockx, Ines; Van Camp, Nadja; Verhoye, Marleen; Boisgard, Raphael; Dubois, Albertine; Jego, Benoit; Jonckers, Elisabeth; Raber, Kerstin; Siquier, Karine; Kuhnast, Bertrand; Dollé, Frédéric; Nguyen, Huu Phuc; Von Hörsten, Stephan; Tavitian, Bertrand; Van der Linden, Annemie

2011-10-15

63

Neuronal and astrocytic metabolism in a transgenic rat model of Alzheimer's disease.  

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Regional hypometabolism of glucose in the brain is a hallmark of Alzheimer's disease (AD). However, little is known about the specific alterations of neuronal and astrocytic metabolism involved in homeostasis of glutamate and GABA in AD. Here, we investigated the effects of amyloid ? (A?) pathology on neuronal and astrocytic metabolism and glial-neuronal interactions in amino acid neurotransmitter homeostasis in the transgenic McGill-R-Thy1-APP rat model of AD compared with healthy controls at age 15 months. Rats were injected with [1-(13)C]glucose and [1,2-(13)C]acetate, and extracts of the hippocampal formation as well as several cortical regions were analyzed using (1)H- and (13)C nuclear magnetic resonance spectroscopy and high-performance liquid chromatography. Reduced tricarboxylic acid cycle turnover was evident for glutamatergic and GABAergic neurons in hippocampal formation and frontal cortex, and for astrocytes in frontal cortex. Pyruvate carboxylation, which is necessary for de novo synthesis of amino acids, was decreased and affected the level of glutamine in hippocampal formation and those of glutamate, glutamine, GABA, and aspartate in the retrosplenial/cingulate cortex. Metabolic alterations were also detected in the entorhinal cortex. Overall, perturbations in energy- and neurotransmitter homeostasis, mitochondrial astrocytic and neuronal metabolism, and aspects of the glutamate-glutamine cycle were found in McGill-R-Thy1-APP rats. PMID:24594625

Nilsen, Linn Hege; Witter, Menno P; Sonnewald, Ursula

2014-05-01

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A transgenic Alzheimer rat with plaques, tau pathology, behavioral impairment, oligomeric a?, and frank neuronal loss.  

Science.gov (United States)

Alzheimer's disease (AD) is hallmarked by amyloid plaques, neurofibrillary tangles, and widespread cortical neuronal loss (Selkoe, 2001). The "amyloid cascade hypothesis" posits that cerebral amyloid sets neurotoxic events into motion that precipitate Alzheimer dementia (Hardy and Allsop, 1991). Yet, faithful recapitulation of all AD features in widely used transgenic (Tg) mice engineered to overproduce A? peptides has been elusive. We have developed a Tg rat model (line TgF344-AD) expressing mutant human amyloid precursor protein (APPsw) and presenilin 1 (PS1?E9) genes, each independent causes of early-onset familial AD. TgF344-AD rats manifest age-dependent cerebral amyloidosis that precedes tauopathy, gliosis, apoptotic loss of neurons in the cerebral cortex and hippocampus, and cognitive disturbance. These results demonstrate progressive neurodegeneration of the Alzheimer type in these animals. The TgF344-AD rat fills a critical need for a next-generation animal model to enable basic and translational AD research. PMID:23575824

Cohen, Robert M; Rezai-Zadeh, Kavon; Weitz, Tara M; Rentsendorj, Altan; Gate, David; Spivak, Inna; Bholat, Yasmin; Vasilevko, Vitaly; Glabe, Charles G; Breunig, Joshua J; Rakic, Pasko; Davtyan, Hayk; Agadjanyan, Michael G; Kepe, Vladimir; Barrio, Jorge R; Bannykh, Serguei; Szekely, Christine A; Pechnick, Robert N; Town, Terrence

2013-04-10

65

Generation of topically transgenic rats by in utero electroporation and in vivo bioluminescence screening.  

Science.gov (United States)

In utero electroporation (IUE) is a technique which allows genetic modification of cells in the brain for investigating neuronal development. So far, the use of IUE for investigating behavior or neuropathology in the adult brain has been limited by insufficient methods for monitoring of IUE transfection success by non-invasive techniques in postnatal animals. For the present study, E16 rats were used for IUE. After intraventricular injection of the nucleic acids into the embryos, positioning of the tweezer electrodes was critical for targeting either the developing cortex or the hippocampus. Ventricular co-injection and electroporation of a luciferase gene allowed monitoring of the transfected cells postnatally after intraperitoneal luciferin injection in the anesthetized live P7 pup by in vivo bioluminescence, using an IVIS Spectrum device with 3D quantification software. Area definition by bioluminescence could clearly differentiate between cortical and hippocampal electroporations and detect a signal longitudinally over time up to 5 weeks after birth. This imaging technique allowed us to select pups with a sufficient number of transfected cells assumed necessary for triggering biological effects and, subsequently, to perform behavioral investigations at 3 month of age. As an example, this study demonstrates that IUE with the human full length DISC1 gene into the rat cortex led to amphetamine hypersensitivity. Co-transfected GFP could be detected in neurons by post mortem fluorescence microscopy in cryosections indicating gene expression present at ?6 months after birth. We conclude that postnatal bioluminescence imaging allows evaluating the success of transient transfections with IUE in rats. Investigations on the influence of topical gene manipulations during neurodevelopment on the adult brain and its connectivity are greatly facilitated. For many scientific questions, this technique can supplement or even replace the use of transgenic rats and provide a novel technology for behavioral neuroscience. PMID:24084570

Vomund, Sandra; Sapir, Tamar; Reiner, Orly; Silva, Maria A de Souza; Korth, Carsten

2013-01-01

66

Urinary excretion of nitric oxide, cyclic GMP, and catecholamines during rest and activity period in transgenic hypertensive rats.  

Science.gov (United States)

Dysregulation of the system of nitric oxide (NO)-cyclic 3',5'-guanosine monophosphate (cGMP) might be involved in the development of hypertension in transgenic hypertensive TGR(mREN2)27 (TGR) rats. The present study was performed to determine possible differences in the day-night pattern and the urinary excretion rates of NO and cGMP in TGR rats in comparison to normotensive Sprague-Dawley (SPRD) controls. In addition, the urinary excretion of creatinine and catecholamines was measured in both rat strains. The day-night excretion patterns of NO, cGMP, catecholamines, and creatinine were preserved in TGR rats. Urinary excretion of NO was significantly decreased in TGR rats, whereas cGMP, the second messenger of NO, was elevated in the transgenic animals. Catecholamines and creatinine excretion rates did not differ between the strains. In conclusion, data suggest that a reduced NO synthesis could contribute to the increased blood pressure in the severely hypertensive rats. However, these data make it unlikely that the disturbances in the nitric oxide-cGMP system and the sympathetic nervous system are mainly responsible for the inverse circadian blood pressure rhythm in TGR rats. PMID:10373100

Globig, S; Witte, K; Lemmer, B

1999-05-01

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Meloxicam Blocks Neuroinflammation, but Not Depressive-Like Behaviors, in HIV-1 Transgenic Female Rats  

Science.gov (United States)

Adolescents living with human immunodeficiency virus (HIV) comprise approximately 12% of the HIV-positive population worldwide. HIV-positive adolescents experience a higher rate of clinical depression, a greater risk of sexual and drug abuse behaviors, and a decreased adherence to highly active antiretroviral therapies (HAART). Using adolescent HIV-1 transgenic rats (HIV-1 tg) that display related immune response alterations and pathologies, this study tested the hypothesis that developmental expression of HIV-1-related proteins induces a depressive-like phenotype that parallels a decrease in hippocampal cell proliferation and an increase in pro-inflammatory cytokine expression in the hippocampus. Consistent with this hypothesis, adolescent HIV-1 tg rats demonstrated a depressive-like behavioral phenotype, had decreased levels of cell proliferation, and exhibited elevated expression of monocyte chemotactic protein-1 (Mcp-1) in the hippocampus relative to controls. Subsequently, we tested the ability of meloxicam, a selective COX-2 inhibitor, to attenuate behavioral deficits via inflammatory mechanisms. Daily meloxicam treatments did not alter the behavioral profile despite effectively reducing hippocampal inflammatory gene expression. Together, these data support a biological basis for the co-morbid manifestation of depression in HIV-positive patients as early as in adolescence and suggest that modifications in behavior manifest independent of inflammatory activity in the hippocampus. PMID:25271421

Nemeth, Christina L.; Glasper, Erica R.; Harrell, Constance S.; Malviya, Sanjana A.; Otis, Jeffrey S.; Neigh, Gretchen N.

2014-01-01

68

Food-anticipatory activity and liver per1-luc activity in diabetic transgenic rats  

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The mammalian Per1 gene is an important component of the core cellular clock mechanism responsible for circadian rhythms. The rodent liver and other tissues rhythmically express Per1 in vitro but typically damp out within a few cycles. In the liver, the peak of this rhythm occurs in the late subjective night in an ad lib-fed rat, but will show a large phase advance in response to restricted availability of food during the day. The relationship between this shift in the liver clock and food-anticipatory activity (FAA), the circadian behavior entrained by daily feeding, is currently unknown. Insulin is released during feeding in mammals and could serve as an entraining signal to the liver. To test the role of insulin in the shift in liver Per1 expression and the generation of FAA, per-luciferase transgenic rats were made diabetic with a single injection of streptozotocine. Following 1 week of restricted feeding and locomotor activity monitoring, liver was collected for per-luc recording. In two separate experiments, FAA emerged and liver Per1 phase-shifted in response to daytime 8-h food restriction. The results rule out insulin as a necessary component of this system.

Davidson, Alec J.; Stokkan, Karl-Arne; Yamazaki, Shin; Menaker, Michael

2002-01-01

69

Characterization of protein tyrosine phosphatase activity in rat liver microsomes: suppressive effect of endogenous regucalcin in transgenic rats.  

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The role of regucalcin, a regulatory protein in intracellular signaling system, in the regulation of protein phosphatase activity in rat liver microsomes was investigated. Protein phosphatase activity torward phosphotyrosine, phosphoserine, and phosphothreonine was assayed in a reaction mixture containing the microsomal protein. Protein phosphatase activity toward phosphotyrosine was strong as compared with that of the enzyme activity toward phosphoserine and phosphothreonine, indicating the existence of protein tyrosine phosphatase. Protein phosphatase activity toward three phosphoaminoacids was significantly enhanced by the addition of both calcium chloride (10 micro M) and calmodulin (2.5 or 5 micro g/ml) in the reaction mixture. The presence of ethylene glycol bis (2-amino-ethylether) N, N, N', N'-tetracetic acid (EGTA; 0.1, 1 or 2 mM) or trifluoperazine (TFP; 10, 20 or 50 micro M), an antagonist of calmodulin, did not have a significant effect on protein phosphatase activity toward phosphotyrosine without calcium addition. Microsomal protein tyrosine phosphatase activity was not changed by okadaic acid (10(-6)-10(-4) M). The enzyme activity was significantly decreased by vanadate (10, 50 or 100 micro M). The addition of regucalcin (0.25 or 0.5 micro M) in the reaction mixture caused a significant inhibition of protein tyrosine phosphatase activity in liver microsomes. Western blot analysis showed a remarkable increase in regucalcin protein level in the liver microsomes of regucalcin transgenic (TG) rats. Protein tyrosine phosphatase activity was significantly suppressed in the liver microsomes of TG rats. This study demonstrates that protein tyrosine phosphatase activity is found in the liver microsomes, and that the enzyme activity is suppressed by regucalcin. PMID:15289895

Fukaya, Yuko; Yamaguchi, Masayoshi

2004-09-01

70

Transgenic rats overexpressing the human MrgX3 gene show cataracts and an abnormal skin phenotype  

International Nuclear Information System (INIS)

The human MrgX3 gene, belonging to the mrgs/SNSRs (mass related genes/sensory neuron specific receptors) family, was overexpressed in transgenic rats using the actin promoter. Two animal lines showed cataracts with liquification/degeneration and swelling of the lens fiber cells. The transient epidermal desquamation was observed in line with higher gene expression. Histopathology of the transgenic rats showed acanthosis and focal parakeratosis. In the epidermis, there was an increase in cellular keratin 14, keratin 10, and loricrin, as well as PGP 9.5 in innervating nerve fibers. These phenotypes accompanied an increase in the number of proliferating cells. These results suggest that overexpression of the human MrgX3 gene causes a disturbance of the normal cell-differentiation process

71

Regional gene expression of LOX-1, VCAM-1, and ICAM-1 in aorta of HIV-1 transgenic rats  

DEFF Research Database (Denmark)

BACKGROUND: Increased prevalence of atherosclerotic cardiovascular disease in HIV-infected patients has been observed. The cause of this accelerated atherosclerosis is a matter of controversy. As clinical studies are complicated by a multiplicity of risk-factors and a low incidence of hard endpoints, studies in animal models could be attractive alternatives. METHODOLOGY/PRINCIPAL FINDINGS: We evaluated gene expression of lectin-like oxidized-low-density-lipoprotein receptor-1 (LOX-1), vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) in HIV-1 transgenic (HIV-1Tg) rats; these genes are all thought to play important roles in early atherogenesis. Furthermore, the plasma level of sICAM-1 was measured. We found that gene expressions of LOX-1 and VCAM-1 were higher in the aortic arch of HIV-1Tg rats compared to controls. Also, the level of sICAM-1 was elevated in the HIV-1Tg rats compared to controls, but the ICAM-1 gene expression profile did not show any differences between the groups. CONCLUSIONS/SIGNIFICANCE: HIV-1Tg rats have gene expression patterns indicating endothelial dysfunction and accelerated atherosclerosis in aorta, suggesting that HIV-infection per se may cause atherosclerosis. This transgenic rat model may be a very promising model for further studies of the pathophysiology behind HIV-associated cardiovascular disease.

Hag, Anne Mette Fisker; Kristoffersen, Ulrik Sloth

2009-01-01

72

Optogenetic patterning of whisker-barrel cortical system in transgenic rat expressing channelrhodopsin-2.  

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The rodent whisker-barrel system has been an ideal model for studying somatosensory representations in the cortex. However, it remains a challenge to experimentally stimulate whiskers with a given pattern under spatiotemporal precision. Recently the optogenetic manipulation of neuronal activity has made possible the analysis of the neuronal network with precise spatiotemporal resolution. Here we identified the selective expression of channelrhodopsin-2 (ChR2), an algal light-driven cation channel, in the large mechanoreceptive neurons in the trigeminal ganglion (TG) as well as their peripheral nerve endings innervating the whisker follicles of a transgenic rat. The spatiotemporal pattern of whisker irradiation thus produced a barrel-cortical response with a specific spatiotemporal pattern as evidenced by electrophysiological and functional MRI (fMRI) studies. Our methods of generating an optogenetic tactile pattern (OTP) can be expected to facilitate studies on how the spatiotemporal pattern of touch is represented in the somatosensory cortex, as Hubel and Wiesel did in the visual cortex. PMID:24695456

Honjoh, Tatsuya; Ji, Zhi-Gang; Yokoyama, Yukinobu; Sumiyoshi, Akira; Shibuya, Yuma; Matsuzaka, Yoshiya; Kawashima, Ryuta; Mushiake, Hajime; Ishizuka, Toru; Yawo, Hiromu

2014-01-01

73

Neuronal driven pre-plaque inflammation in a transgenic rat model of Alzheimer's disease.  

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Chronic brain inflammation is associated with Alzheimer's disease (AD) and is classically attributed to amyloid plaque deposition. However, whether the amyloid pathology can trigger early inflammatory processes before plaque deposition remains a matter of debate. To address the possibility that a pre-plaque inflammatory process occurs, we investigated the status of neuronal, astrocytic, and microglial markers in pre- and post-amyloid plaque stages in a novel transgenic rat model of an AD-like amyloid pathology (McGill-R-Thy1-APP). In this model, we found a marked upregulation of several classical inflammatory markers such as COX-2, IL-1?, TNF-?, and fractalkine (CX3CL1) in the cerebral cortex and hippocampus. Interestingly, many of these markers were highly expressed in amyloid beta-burdened neurons. Activated astrocytes and microglia were associated with these A?-burdened neurons. These findings confirm the occurrence of a proinflammatory process preceding amyloid plaque deposition and suggest that A?-burdened neurons play a crucial role in initiating inflammation in AD. PMID:24831823

Hanzel, Cecilia E; Pichet-Binette, Alexa; Pimentel, Luisa S B; Iulita, M Florencia; Allard, Simon; Ducatenzeiler, Adriana; Do Carmo, Sonia; Cuello, A Claudio

2014-10-01

74

Longitudinal analysis of the behavioral phenotype in a novel transgenic rat model of early stages of Alzheimer's disease  

Science.gov (United States)

Intraneuronal accumulation of amyloid ? (iA?) has been linked to mild cognitive impairment that may precede Alzheimer's disease (AD) onset. This neuropathological trait was recently mimicked in a novel animal model of AD, the hemizygous transgenic McGill-R-Thy1-APP (Tg+/?) rat. The characterization of the behavioral phenotypes in this animal model could provide a baseline of efficacy for earlier therapeutic interventions. The aim of the present study was to undertake a longitudinal study of A? accumulation and a comprehensive behavioral evaluation of this transgenic rat model. We assessed exploratory activity, anxiety-related behaviors, recognition memory, working memory, spatial learning and reference memory at 3, 6, and 12 months of age. In parallel, we measured A? by ELISA, Western blots and semiquantitative immunohistochemistry in hippocampal samples. SDS-soluble A? peptide accumulated at low levels (~9 pg/mg) without differences among ages. However, Western blots showed SDS-resistant A? oligomers (~30 kDa) at 6 and 12 months, but not at 3 months. When compared to wild-type (WT), male Tg+/? rats exhibited a spatial reference memory deficit in the Morris Water Maze (MWM) as early as 3 months of age, which persisted at 6 and 12 months. In addition, Tg+/? rats displayed a working memory impairment in the Y-maze and higher anxiety levels in the Open Field (OF) at 6 and 12 months of age, but not at 3 months. Exploratory activity in the OF was similar to that of WT at all-time points. Spatial learning in the MWM and the recognition memory, as assessed by the Novel Object Recognition Test, were unimpaired at any time point. The data from the present study demonstrate that the hemizygous transgenic McGill-R-Thy1-APP rat has a wide array of behavioral and cognitive impairments from young adulthood to middle-age. The low A? burden and early emotional and cognitive deficits in this transgenic rat model supports its potential use for drug discovery purposes in early AD. PMID:25278855

Galeano, Pablo; Martino Adami, Pamela V.; Do Carmo, Sonia; Blanco, Eduardo; Rotondaro, Cecilia; Capani, Francisco; Castano, Eduardo M.; Cuello, A. Claudio; Morelli, Laura

2014-01-01

75

Longitudinal analysis of the behavioral phenotype in a novel transgenic rat model of early stages of Alzheimer's disease.  

Science.gov (United States)

Intraneuronal accumulation of amyloid ? (iA?) has been linked to mild cognitive impairment that may precede Alzheimer's disease (AD) onset. This neuropathological trait was recently mimicked in a novel animal model of AD, the hemizygous transgenic McGill-R-Thy1-APP (Tg(+/-)) rat. The characterization of the behavioral phenotypes in this animal model could provide a baseline of efficacy for earlier therapeutic interventions. The aim of the present study was to undertake a longitudinal study of A? accumulation and a comprehensive behavioral evaluation of this transgenic rat model. We assessed exploratory activity, anxiety-related behaviors, recognition memory, working memory, spatial learning and reference memory at 3, 6, and 12 months of age. In parallel, we measured A? by ELISA, Western blots and semiquantitative immunohistochemistry in hippocampal samples. SDS-soluble A? peptide accumulated at low levels (~9 pg/mg) without differences among ages. However, Western blots showed SDS-resistant A? oligomers (~30 kDa) at 6 and 12 months, but not at 3 months. When compared to wild-type (WT), male Tg(+/-) rats exhibited a spatial reference memory deficit in the Morris Water Maze (MWM) as early as 3 months of age, which persisted at 6 and 12 months. In addition, Tg(+/-) rats displayed a working memory impairment in the Y-maze and higher anxiety levels in the Open Field (OF) at 6 and 12 months of age, but not at 3 months. Exploratory activity in the OF was similar to that of WT at all-time points. Spatial learning in the MWM and the recognition memory, as assessed by the Novel Object Recognition Test, were unimpaired at any time point. The data from the present study demonstrate that the hemizygous transgenic McGill-R-Thy1-APP rat has a wide array of behavioral and cognitive impairments from young adulthood to middle-age. The low A? burden and early emotional and cognitive deficits in this transgenic rat model supports its potential use for drug discovery purposes in early AD. PMID:25278855

Galeano, Pablo; Martino Adami, Pamela V; Do Carmo, Sonia; Blanco, Eduardo; Rotondaro, Cecilia; Capani, Francisco; Castaño, Eduardo M; Cuello, A Claudio; Morelli, Laura

2014-01-01

76

Longitudinal analysis of the behavioral phenotype in a novel transgenic rat model of early stages of Alzheimer’s disease  

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Full Text Available Intraneuronal accumulation of amyloid ? (iA? has been linked to mild cognitive impairment that may precede Alzheimer’s disease (AD onset. This neuropathological trait was recently mimicked in a novel animal model of AD, the hemizygous transgenic McGill-R-Thy1-APP (Tg+/- rat. The characterization of the behavioral phenotypes in this animal model could provide a baseline of efficacy for earlier therapeutic interventions. The aim of the present study was to undertake a longitudinal study of A? accumulation and a comprehensive behavioral evaluation of this transgenic rat model. We assessed exploratory activity, anxiety-related behaviors, recognition memory, working memory, spatial learning and reference memory at 3, 6 and 12 months of age. In parallel, we measured A? by ELISA, Western blots and semiquantitative immunohistochemistry in hippocampal samples. SDS-soluble A? peptide accumulated at low levels (~9 pg/mg without differences among ages. However, Western blots showed SDS-resistant A? oligomers (~30 kDa at 6 and 12 months, but not at 3 months. When compared to wild-type (WT, male Tg+/- rats exhibited a spatial reference memory deficit in the Morris Water Maze (MWM as early as 3 months of age, which persisted at 6 and 12 months. In addition, Tg+/- rats displayed a working memory impairment in the Y-maze and higher anxiety levels in the Open Field (OF at 6 and 12 months of age, but not at 3 months. Exploratory activity in the OF was similar to that of WT at all time points. Spatial learning in the MWM and the recognition memory, as assessed by the Novel Object Recognition Test, were unimpaired at any time point. The data from the present study demonstrate that the hemizygous transgenic McGill-R-Thy1-APP rat has a wide array of behavioral and cognitive impairments from young adulthood to middle-age. The low A? burden and early emotional and cognitive deficits in this transgenic rat model supports its potential use for drug discovery purposes in early AD.

LauraMorelli

2014-09-01

77

Distinct Transcriptional Mechanisms Direct Expression of the Rat Dmrt1 Promoter in Sertoli Cells and Germ Cells of Transgenic Mice1  

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DMRT1 is a transcription factor expressed only in Sertoli cells and undifferentiated spermatogonia of the postnatal testis, where it is required for proper cellular differentiation and fertility. To elucidate the transcriptional regulatory regions that provide DMRT1's cell-specific expression, transgenic mice containing a LacZ reporter gene driven by variable amounts of rat Dmrt1 5? flanking sequence, 9 kb and smaller, were evaluated. Examination of transgene expression by RT-PCR indicated ...

Lei, Ning; Karpova, Tatiana; Hornbaker, Kaori I.; Rice, Daren A.; Heckert, Leslie L.

2009-01-01

78

Roles of glutathione in antioxidant defense, inflammation, and neuron differentiation in the thalamus of HIV-1 transgenic rats.  

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Inflammation and oxidative stress in the brain are major causes of HIV-associated neurocognitive disorders. Previously we have reported high content of glutathione (GSH) in the thalamus of rats with F344 genetic background. In this study, we investigated the changes of GSH metabolism and GSH-dependent antioxidant enzymes in the rat thalamus in response to HIV-1 transgenesis, and their associations with oxidative stress, inflammation, and neuronal development. Male HIV-1 transgenic (HIV-1Tg) rats and wild type F344 rats at 10 months were used in this study, with 5 rats in each group. Parameters measured in this study included: total and oxidized GSH, glutathione peroxidase (GPx), glutathione-S-transferase (GST), gamma-glutamylcysteine synthetase (GCS), gamma-glutamyl transferase (GGT), cysteine/cystine transporters, 4-hydroxynonenal (HNE), interleukin 12 (IL12), neuronal nuclei (NeuN), microtubule-associated protein (MAP2), and glia fibrillary acidic protein (GFAP). The levels of total GSH, oxidized GSH (GSSG) and MAP2 protein, and enzymatic activities of GCS, GPx and GST were significantly higher in HIV-1Tg rats compared with F344 rats, but the ratio of GSSG/GSH, activity of GGT and levels of HNE, NeuN protein and GFAP protein did not change. HIV-1Tg rats showed a lower level of IL12 protein. GSH positively correlated with GCS, GST and MAP2, GSSG/GSH ratio positively correlated with HNE and IL12, the activities of GPx, GST and GCS positively correlated with each other, and negatively correlated with HNE. These findings suggest an important role of the GSH-centered system in reducing oxidative stress and neuroinflammation, and enhancing neuron differentiation in the thalamus of HIV-1Tg rats. PMID:24609977

Pang, Xiaosha; Panee, Jun

2014-06-01

79

Radiographic visualisation of seropositive rheumatoid arthritis in Carriers of HLA-B27  

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A group of 11 B27-positive, seropositive patients with rheumatoid arthritis was compared with 11 matched B27-negative seropositive patients. The radiographs of all limb joints, the sacroiliac joints, and the cervical spine were read blindly. Ten patients in each group were radiographed 2-6 times during observation periods of 3-13 years; one patient in each group was only examined once. The prevailing picture of both groups was that of progressive erosive rheumatoid arthritis, although two small differences were found: Erosions of the apophyseal joints of the cervical spine and slight periosteal new bone formation of the shoulder, hip, and knee regions occurred more often in the B27-positive than in the B27-negative group.

Jurik, A.G.; Carvalho, A. de; Graudal, H.

1987-07-01

80

Radiographic visualisation of seropositive rheumatoid arthritis in Carriers of HLA-B27  

International Nuclear Information System (INIS)

A group of 11 B27-positive, seropositive patients with rheumatoid arthritis was compared with 11 matched B27-negative seropositive patients. The radiographs of all limb joints, the sacroiliac joints, and the cervical spine were read blindly. Ten patients in each group were radiographed 2-6 times during observation periods of 3-13 years; one patient in each group was only examined once. The prevailing picture of both groups was that of progressive erosive rheumatoid arthritis, although two small differences were found: Erosions of the apophyseal joints of the cervical spine and slight periosteal new bone formation of the shoulder, hip, and knee regions occurred more often in the B27-positive than in the B27-negative group. (orig.)

 
 
 
 
81

Response to metal stress of Nicotiana langsdorffii plants wild-type and transgenic for the rat glucocorticoid receptor gene.  

Science.gov (United States)

Recently our findings have shown that the integration of the gene coding for the rat gluco-corticoid receptor (GR receptor) in Nicotiana langsdorffii plants induced morphophysiological effects in transgenic plants through the modification of their hormonal pattern. Phytohormones play a key role in plant responses to many different biotic and abiotic stresses since a modified hormonal profile up-regulates the activation of secondary metabolites involved in the response to stress. In this work transgenic GR plants and isogenic wild type genotypes were exposed to metal stress by treating them with 30ppm cadmium(II) or 50ppm chromium(VI). Hormonal patterns along with changes in key response related metabolites were then monitored and compared. Heavy metal up-take was found to be lower in the GR plants. The transgenic plants exhibited higher values of S-abscisic acid (S-ABA) and 3-indole acetic acid (IAA), salicylic acid and total polyphenols, chlorogenic acid and antiradical activity, compared to the untransformed wild type plants. Both Cd and Cr treatments led to an increase in hormone concentrations and secondary metabolites only in wild type plants. Analysis of the results suggests that the stress responses due to changes in the plant's hormonal system may derive from the interaction between the GR receptor and phytosteroids, which are known to play a key role in plant physiology and development. PMID:23395537

Fuoco, Roger; Bogani, Patrizia; Capodaglio, Gabriele; Del Bubba, Massimo; Abollino, Ornella; Giannarelli, Stefania; Spiriti, Maria Michela; Muscatello, Beatrice; Doumett, Saer; Turetta, Clara; Zangrando, Roberta; Zelano, Vincenzo; Buiatti, Marcello

2013-05-01

82

Development and characterization of a new inbred transgenic rat strain expressing DsRed monomeric fluorescent protein.  

Science.gov (United States)

The inbred rat is a suitable model for studying human disease and because of its larger size is more amenable to complex surgical manipulation than the mouse. While the rodent fulfills many of the criteria for transplantation research, an important requirement is the ability to mark and track donors cells and assess organ viability. However, tracking ability is limited by the availability of transgenic (Tg) rats that express suitable luminescent or fluorescent proteins. Red fluorescent protein cloned from Discosoma coral (DsRed) has several advantages over other fluorescent proteins, including in vivo detection in the whole animal and ex vivo visualization in organs as there is no interference with autofluorescence. We generated and characterized a novel inbred Tg Lewis rat strain expressing DsRed monomeric (DsRed mono) fluorescent protein under the control of a ubiquitously expressed ROSA26 promoter. DsRed mono Tg rats ubiquitously expressed the marker gene as detected by RT-PCR but the protein was expressed at varying levels in different organs. Conventional skin grafting experiments showed acceptance of DsRed monomeric Tg rat skin on wild-type rats for more than 30 days. Cardiac transplantation of DsRed monomeric Tg rat hearts into wild-type recipients further showed graft acceptance and long-term organ viability (>6 months). The DsRed monomeric Tg rat provides marked cells and/or organs that can be followed for long periods without immune rejection and therefore is a suitable model to investigate cell tracking and organ transplantation. PMID:25011565

Montanari, Sonia; Wang, Xing-Hua; Yannarelli, Gustavo; Dayan, Victor; Berger, Thorsten; Zocche, Larissa; Kobayashi, Eiji; Viswanathan, Sowmya; Keating, Armand

2014-10-01

83

Reduced brown adipose tissue thermogenesis during environmental interactions in transgenic rats with ataxin-3-mediated ablation of hypothalamic orexin neurons.  

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Thermogenesis in brown adipose tissue (BAT) contributes to substantial increases in body temperature evoked by threatening or emotional stimuli. BAT thermogenesis also contributes to increases in body temperature that occur during active phases of the basic rest-activity cycle (BRAC), as part of normal daily life. Hypothalamic orexin-synthesizing neurons influence many physiological and behavioral variables, including BAT and body temperature. In conscious unrestrained animals maintained for 3 days in a quiet environment (24-26°C) with ad libitum food and water, we compared temperatures in transgenic rats with ablation of orexin neurons induced by expression of ataxin-3 (Orx_Ab) with wild-type (WT) rats. Both baseline BAT temperature and baseline body temperature, measured at the onset of BRAC episodes, were similar in Orx_Ab and WT rats. The time interval between BRAC episodes was also similar in the two groups. However, the initial slopes and amplitudes of BRAC-related increases in BAT and body temperature were reduced in Orx_Ab rats. Similarly, the initial slopes and amplitudes of the increases in BAT temperatures induced by sudden exposure to an intruder rat (freely moving or confined to a small cage) or by sudden exposure to live cockroaches were reduced in resident Orx_Ab rats. Constriction of the tail artery induced by salient alerting stimuli was also reduced in Orx_Ab rats. Our results suggest that orexin-synthesizing neurons contribute to the intensity with which rats interact with the external environment, both when the interaction is "spontaneous" and when the interaction is provoked by threatening or salient environmental events. PMID:25324552

Mohammed, Mazher; Ootsuka, Youichirou; Yanagisawa, Masashi; Blessing, William

2014-10-15

84

Early differences in dorsal hippocampal metabolite levels in males but not females in a transgenic rat model of Alzheimer's disease.  

Science.gov (United States)

McGill-R-Thy1-APP rats express the human amyloid precursor protein carrying the Swedish and Indiana mutations. We examined the neurochemical content of the dorsal hippocampus in three-months-old male and female transgenic rats and healthy age- and gender-matched controls using in vivo (1)H MRS in order to assess early metabolite alterations and whether these were similar for both genders. Whereas male and female controls had similar levels of all metabolites, differences were evident between male and female McGill-R-Thy1-APP rats. Compared with McGill-R-Thy1-APP females, McGill-R-Thy1-APP males had lower levels of myo-inositol and N-acetylaspartate (NAA). No differences in metabolite levels were evident when female control and McGill-R-Thy1-APP rats were compared, whereas McGill-R-Thy1-APP males had lower levels of glutamate, NAA and total choline compared with male controls. In addition to metabolite concentrations, metabolite ratios are reported as these are widely used. The results from this preliminary study demonstrate early metabolite alterations in the dorsal hippocampus of males in this rat model of Alzheimer's disease, and imply that very early possible neurochemical markers of the disease are different for males and females. PMID:24338370

Nilsen, L H; Melø, T M; Witter, M P; Sonnewald, U

2014-02-01

85

HIV-1 transgene expression in rats induces differential expression of tumor necrosis factor alpha and zinc transporters in the liver and the lung  

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Full Text Available Abstract Background Highly effective antiviral treatment can suppress HIV-1 infection, but the chronic effects of HIV-1-related viral proteins, including gp120 and Tat, on organs such as the lungs can be damaging. HIV-1 transgenic rodent models are useful for studying the systemic effects of these proteins independently of viral infection. We have previously shown that HIV-1 transgene expression (and therefore, HIV-1-related protein expression in rats decreases alveolar macrophage zinc levels and phagocytic capacity by unknown mechanisms. We hypothesized that HIV-1 transgene expression induces chronic inflammation and zinc sequestration within the liver and thereby decreases zinc bioavailability in the lung. We examined the expression of the pro-inflammatory cytokine, tumor necrosis factor alpha (TNF?, the zinc storage protein, metallothionein (MT1, and the zinc exporter, ZNT1 in the livers and the lungs of wild type and HIV-1 transgenic rats ± dietary zinc supplementation. In addition, we measured zinc levels, the zinc importing protein ZIP1, and the phagocytic capacity in the alveolar macrophages. Results HIV-1 transgene expression increased the liver-specific expression of TNF?, suggesting a chronic inflammatory response within the liver in response to HIV-1-related protein expression. In parallel, HIV-1 transgene expression significantly increased MT1 and ZNT1 expression in the liver as compared to the lung, a pattern that is consistent with zinc sequestration in the liver as occurs during systemic inflammation. Further, HIV-1 transgene expression decreased intracellular zinc levels and increased expression of ZIP1 in the alveolar macrophages, a pattern consistent with zinc deficiency, and decreased their bacterial phagocytic capacity. Interestingly, dietary zinc supplementation in HIV-1 transgenic rats decreased gene expression of TNF?, MT1, and ZNT1 in the liver while simultaneously increasing their expression in the lung. In parallel, zinc supplementation increased alveolar macrophage intracellular zinc levels and bacterial phagocytic capacity in HIV-1 transgenic rats. Conclusion Taken together, these findings suggest that chronic HIV-1-related protein expression causes liver inflammation and zinc sequestration, which in turn limits zinc bioavailability in the lung and thereby impairs alveolar macrophage phagocytic function. Importantly, dietary zinc supplementation decreases liver inflammation and zinc sequestration and restores alveolar macrophage phagocytic function in HIV-1 transgenic rats, a result with potential clinical implications for improving lung health in HIV-1-infected individuals.

Guidot David M

2011-10-01

86

The direct renin inhibitor aliskiren improves vascular remodelling in transgenic rats harbouring human renin and angiotensinogen genes.  

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In the present study, we tested the hypothesis that chronic treatment with the direct rennin inhibitor aliskiren improves the remodelling of resistance arteries in dTGR (double-transgenic rats). dTGR (5 weeks) were treated with aliskiren (3 mg/kg of body mass per day) or ramipril (1 mg/kg of body mass per day) for 14 days and compared with age-matched vehicle-treated dTGR. BP (blood pressure) was similarly reduced in both aliskiren-treated and ramipril-treated rats compared with control dTGR (167±1 and 169±2 mmHg compared with 197±4 mmHg respectively; PACE (angiotensin-converting enzyme) inhibition in improving vascular remodelling through different mechanisms. PMID:23438195

Savoia, Carmine; Arrabito, Emanuele; Parente, Rosa; Sada, Lidia; Madaro, Luca; Nicoletti, Carmine; Zezza, Luigi; Alonzo, Alessandro; Rubattu, Speranza; Michelini, Serena; Muller, Dominik N; Volpe, Massimo

2013-08-01

87

Characterization of dsRed2-positive cells in the doublecortin-dsRed2 transgenic adult rat retina.  

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Doublecortin (DCX) is predominantly expressed in neuronal precursor cells and young immature neurons of the developing and adult brain, where it is involved in neuronal differentiation, migration and plasticity. Moreover, its expression pattern reflects neurogenesis, and transgenic DCX promoter-driven reporter models have been previously used to investigate adult neurogenesis. In this study, we characterize dsRed2 reporter protein-expressing cells in the adult retina of the transgenic DCX promoter-dsRed2 rat model, with the aim to identify cells with putative neurogenic activity. Additionally, we confirmed the expression of the dsRed2 protein in DCX-expressing cells in the adult hippocampal dentate gyrus. Adult DCX-dsRed2 rat retinas were analyzed by immunohistochemistry for expression of DCX, NF200, Brn3a, Sox2, NeuN, calbindin, calretinin, PKC-a, Otx2, ChAT, PSA-NCAM and the glial markers GFAP and CRALBP, followed by confocal laser-scanning microscopy. In addition, brain sections of transgenic rats were analyzed for dsRed2 expression and co-localization with DCX, NeuN, GFAP and Sox2 in the cortex and dentate gyrus. Endogenous DCX expression in the adult retina was confined to horizontal cells, and these cells co-expressed the DCX promoter-driven dsRed2 reporter protein. In addition, we encountered dsRed2 expression in various other cell types in the retina: retinal ganglion cells (RGCs), a subpopulation of amacrine cells, a minority of bipolar cells and in perivascular cells. Since also RGCs expressed dsRed2, the DCX-dsRed2 rat model might offer a useful tool to study RGCs in vivo under various conditions. Müller glial cells, which have previously been identified as cells with stem cell features and with neurogenic potential, did express neither endogenous DCX nor the dsRed2 reporter. However, and surprisingly, we identified a perivascular glial cell type expressing the dsRed2 reporter, enmeshed with the glia/stem cell marker GFAP and colocalizing with the neural stem cell marker Sox2. These findings suggest the so far undiscovered existence of perivascular associated cell with neural stem cell-like properties in the adult retina. PMID:25138677

Trost, A; Schroedl, F; Marschallinger, J; Rivera, F J; Bogner, B; Runge, C; Couillard-Despres, S; Aigner, L; Reitsamer, H A

2014-12-01

88

Overexpression of the rat inducible 70-kD heat stress protein in a transgenic mouse increases the resistance of the heart to ischemic injury.  

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Myocardial protection and changes in gene expression follow whole body heat stress. Circumstantial evidence suggests that an inducible 70-kD heat shock protein (hsp70i), increased markedly by whole body heat stress, contributes to the protection. Transgenic mouse lines were constructed with a cytomegalovirus enhancer and beta-actin promoter driving rat hsp70i expression in heterozygote animals. Unstressed, transgene positive mice expressed higher levels of myocardial hsp70i than transgene negative mice after whole body heat stress. This high level of expression occurred without apparent detrimental effect. The hearts harvested from transgene positive mice and transgene negative littermates were Langendorff perfused and subjected to 20 min of warm (37 degrees C) zero-flow ischemia and up to 120 min of reflow while contractile recovery and creatine kinase efflux were measured. Myocardial infarction was demarcated by triphenyltetrazolium. In transgene positive compared with transgene negative hearts, the zone of infarction was reduced by 40%, contractile function at 30 min of reflow was doubled, and efflux of creatine kinase was reduced by approximately 50%. Our findings suggest for the first time that increased myocardial hsp70i expression results in protection of the heart against ischemic injury and that the antiischemic properties of hsp70i have possible therapeutic relevance. PMID:7706448

Marber, M S; Mestril, R; Chi, S H; Sayen, M R; Yellon, D M; Dillmann, W H

1995-04-01

89

Identification of the responsible proteins for increased selenium bioavailability in the brain of transgenic rats overexpressing selenoprotein M.  

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The present study was conducted to investigate whether the high antioxidant activity induced by selenium (Sel) treatment and selenoprotein M (SelM) overexpression affected the protein profile of the brain cortex. To accomplish this, the changes in global protein expression were measured in transgenic (Tg) rats expressing human SelM (CMV/hSelM) and non?Tg rats using two?dimensional electrophoresis (2?DE). The results revealed that: ?) CMV/hSelM Tg rats showed a high level of enzyme activity for antioxidant protein in the brain cortex compared to non?Tg rats; ?) the high activity of these enzymes induced a decrease in total antioxidant concentration and ??secretase activity in CMV/hSelM Tg rats; ?) five proteins were upregulated and three were downregulated by SelM overexpression; ?) among the five upregulated proteins, two associated with creatine kinase B?type (B?CK) and E3 ubiquitin?protein ligase RING1 (RING finger protein 1) were further increased in the two groups following Sel treatment, whereas synaptotagmin-15 (SytXV), eukaryotic translation initiation factor 4H (eIF-4H) and lactate dehydrogenase B (LDH-B) were increased or decreased under the same conditions; ?) the three downregulated proteins did not induce a significant change in expression following Sel treatment; and ?) the protein expression level alterations of the two selected spots (B?CK and SytXV) identified by 2?DE were extremely similar to the results from western blot analysis. Overall, the results of the present study provide primary novel biological evidence that new functional protein groups and individual proteins in the brain cortex of CMV/hSelM Tg rats are associated with Sel biology, including the response to Sel treatment and SelM overexpression. PMID:25269742

Kim, Yona; Goo, Jun Seo; Kim, Il Yong; Kim, Ji Eun; Kwak, Moon Hwa; Go, Jun; Shim, Sunbo; Hong, Jin Tae; Hwang, Dae Youn; Seong, Je Kyung

2014-12-01

90

Inhibition of soluble epoxide hydrolase is renoprotective in 5/6 nephrectomized Ren-2 transgenic hypertensive rats  

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1. The aim of the present study was to test the hypothesis that increasing kidney tissue concentrations of epoxyeicosatrienoic acids (EETs) by preventing their degradation to the biologically inactive dihydroxyeicosatrienoic acids (DHETEs) using blockade of soluble epoxide hydrolase (sEH) would attenuate the progression of chronic kidney disease (CKD). 2. Ren-2 transgenic rats (TGR) after 5/6 renal mass reduction (5/6 NX) served as a model of CKD associated with angiotensin (Ang) II-dependent hypertension. Soluble epoxide hydrolase was inhibited using cis-4-[4-(3-adamantan-1-yl-ureido)cyclohexyloxy]benzoic acid (c-AUCB; 3 mg/L drinking water) for 20 weeks after 5/6 NX. Sham-operated normotensive transgene-negative Hannover Sprague-Dawley (HanSD) rats served as controls. 3. When applied in TGR subjected to 5/6 NX, c-AUCB treatment improved survival rate, prevented the increase in blood pressure, retarded the progression of cardiac hypertrophy, reduced proteinuria and the degree of glomerular and tubulointerstitial injury and reduced glomerular volume. All these organ-protective actions were associated with normalization of the intrarenal EETs : DHETEs ratio, an index of the availability of biologically active EETs, to levels observed in sham-operated HanSD rats. There were no significant concurrent changes of increased intrarenal AngII content. 4. Together, these results show that 5/6 NX TGR exhibit a profound deficiency of intrarenal availability of active epoxygenase metabolites (EETs), which probably contributes to the progression of CKD in this model of AngII-dependent hypertension, and that restoration of intrarenal availability of EETs using long-term c-AUCB treatment exhibits substantial renoprotective actions. PMID:24471737

Kujal, Petr; Certikova Chabova, Vera; Skaroupkova, Petra; Huskova, Zuzana; Vernerova, Zdena; Kramer, Herbert J; Walkowska, Agnieszka; Kompanowska-Jezierska, Elzbieta; Sadowski, Janusz; Kitada, Kento; Nishiyama, Akira; Hwang, Sung H; Hammock, Bruce D; Imig, John D; Cervenka, Ludek

2014-01-01

91

Differences in acid-induced currents between oxytocin-mRFP1 and vasopressin-eGFP neurons isolated from the supraoptic and paraventricular nuclei of transgenic rats.  

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The hypothalamic supraoptic nucleus (SON) and paraventricular nucleus (PVN) consists of two types of magnocellular neurosecretory cells, oxytocin (OXT) and arginine vasopressin (AVP). We generated and characterized rats that express an OXT-monomeric red fluorescent protein 1 (mRFP1) and an AVP-enhanced green fluorescent protein (eGFP) fusion transgene. These transgenic rats enable the visualization of OXT or AVP neurons. Taking advantage of this, we examined the differences between OXT-mRFP1 neurons and AVP-eGFP neurons in response to acid. Acid-sensing ion channels (ASICs) are neuronal voltage-insensitive cationic channels that are activated by extracellular acidification. Although functional ASICs have been identified in AVP neurons, differences in acid-induced currents between OXT and AVP neurons in SON have not been reported. In the present study, we used the whole-cell patch-clamp technique to investigate differences between OXT-mRFP1 neurons and AVP-eGFP neurons reaction to acid in SON and PVN. In voltage clamp mode, lowering extracellular pH evoked inward currents in both OXT-mRFP1 neurons and AVP-eGFP neurons. In our findings, the acid-induced currents in the OXT-mRFP1 neurons were significantly smaller than those in the AVP-eGFP neurons. These acid-induced currents were inhibited by amiloride, a known blocker of ASICs. Further, to compare the response to acid between OXT-mRFP1 and AVP-eGFP neurons in the same transgenic rat, we used a double transgenic rat by mating an OXT-mRFP1 transgenic rat with an AVP-eGFP transgenic rat. The acid-induced currents of OXT-mRFP1 neurons were significantly smaller than those of AVP-eGFP neurons from the double transgenic rats. These currents were almost completely inhibited by amiloride. The difference of acid-sensitivity between OXT and AVP neurons might contribute to maintaining systematic order in hypothalamic function. PMID:25220704

Ohkubo, Jun-Ichi; Ohbuchi, Toyoaki; Yoshimura, Mitsuhiro; Maruyama, Takashi; Hashimoto, Hirofumi; Matsuura, Takanori; Suzuki, Hideaki; Ueta, Yoichi

2014-11-01

92

Increased neuroinflammatory and arachidonic acid cascade markers, and reduced synaptic proteins, in brain of HIV-1 transgenic rats  

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Full Text Available Abstract Background Cognitive impairment has been reported in human immune deficiency virus-1- (HIV-1- infected patients as well as in HIV-1 transgenic (Tg rats. This impairment has been linked to neuroinflammation, disturbed brain arachidonic acid (AA metabolism, and synapto-dendritic injury. We recently reported upregulated brain AA metabolism in 7- to 9-month-old HIV-1 Tg rats. We hypothesized that these HIV-1 Tg rats also would show upregulated brain inflammatory and AA cascade markers and a deficit of synaptic proteins. Methods We measured protein and mRNA levels of markers of neuroinflammation and the AA cascade, as well as pro-apoptotic factors and synaptic proteins, in brains from 7- to 9-month-old HIV-1 Tg and control rats. Results Compared with control brain, HIV-1 Tg rat brain showed immunoreactivity to glycoprotein 120 and tat HIV-1 viral proteins, and significantly higher protein and mRNA levels of (1 the inflammatory cytokines interleukin-1? and tumor necrosis factor ?, (2 the activated microglial/macrophage marker CD11b, (3 AA cascade enzymes: AA-selective Ca2+-dependent cytosolic phospholipase A2 (cPLA2-IVA, secretory sPLA2-IIA, cyclooxygenase (COX-2, membrane prostaglandin E2 synthase, 5-lipoxygenase (LOX and 15-LOX, cytochrome p450 epoxygenase, and (4 transcription factor NF-?Bp50 DNA binding activity. HIV-1 Tg rat brain also exhibited signs of cell injury, including significantly decreased levels of brain-derived neurotrophic factor (BDNF and drebrin, a marker of post-synaptic excitatory dendritic spines. Expression of Ca2+-independent iPLA2-VIA and COX-1 was unchanged. Conclusions HIV-1 Tg rats show elevated brain markers of neuroinflammation and AA metabolism, with a deficit in several synaptic proteins. These changes are associated with viral proteins and may contribute to cognitive impairment. The HIV-1 Tg rat may be a useful model for understanding progression and treatment of cognitive impairment in HIV-1 patients.

Harry Gaylia

2011-08-01

93

Molecular changes in the early phase of renin-dependent cardiac hypertrophy in hypertensive cyp1a1ren-2 transgenic rats.  

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An early response to high arterial pressure is the development of cardiac hypertrophy. Functional and transcriptional regulation of ion channels and Ca(2+) handling proteins are involved in this process but the relative contribution of each is unclear. In this study, we investigated the expression of genes involved in action potential generation and Ca(2+) homeostasis of cardiomyocytes in hypertensive cyp1a1ren-2 transgenic rats. In this model, the transgene prorenin was induced by indole-3-carbinol for 2 weeks allowing the induction of hypertension. Electrophysiological recordings from cardiomyocytes of hypertensive rats revealed a slight increase in membrane capacitance consistent with cellular hypertrophy. L-type calcium current density was reduced by 30%. Left ventricles of hypertensive rats showed a significant increase in transcript and protein levels of the cation channel TRPC6 and FK506-binding protein, whereas levels of SERCA2 and voltage-dependent potassium channels K(v)4.2 and K(v)4.3 were found to be decreased. Further, a marked nuclear localization of the transcription factors GATA4 and NFATC4 was observed in cardiac tissue of hypertensive rats. The cyp1a1ren-2 transgenic rat thus appears to be a valid model to investigate early changes in cardiac hypertrophy. This study points to roles for TRPC6, FK506BP, SERCA2, K(v)4.2, and K(v)4.3 in the development of cardiac hypertrophy. PMID:23060473

Kunert-Keil, Christiane; Landsberger, Martin; Jantzen, Franziska; Niessner, Felix; Kroemer, Heyo K; Felix, Stephan B; Brinkmeier, Heinrich; Peters, Jörg

2013-03-01

94

Changes in skeletal muscle iron metabolism outpace amyotrophic lateral sclerosis onset in transgenic rats bearing the G93A hmSOD1 gene mutation.  

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Abstract Objective. Recently, iron and the adaptor protein "p66Shc" have been shown to play an important role in the development of amyotrophic lateral sclerosis (ALS) in rats. We hypothesized that changes in muscle p66Shc activity and iron metabolism would appear before visible symptoms of the disease occurred. Methods. In the present study, we used transgenic rats bearing the G93A hmSOD1 gene mutation and their non-transgenic littermates to test this hypothesis. We examined muscle p66Shc phosphorylation and iron metabolism in relation to oxidative stress in animals at three disease stages: asymptomatic (ALS I), disease onset (ALS II), and end-stage disease (ALS III). Results. Significant changes in iron metabolism and markers of lipid and protein oxidation were detected in ALS I animals, which manifested as decreased levels of ferritin H and ferroportin 1 (Fpn1) and increased levels of ferritin L levels. Muscles of ALS I rats possessed increased levels of p66Shc phosphorylated at Ser(36) compared with muscles of control rats. During disease progression, level of ferritin H significantly increased and was accompanied by iron accumulation. Conclusions. This study showed that multiple mechanisms may underlie iron accumulation in muscles of ALS transgenic rats, which include changes in blood hepcidin and muscle Fpn1 and increased level of muscle ferritin H. These data suggest that impaired iron metabolism is not a result of changes in motor activity. PMID:25175826

Halon, M; Kaczor, J J; Ziolkowski, W; Flis, D J; Borkowska, A; Popowska, U; Nyka, W; Wozniak, M; Antosiewicz, J

2014-11-01

95

Cyst formation in the PKD2 (1-703 transgenic rat precedes deregulation of proliferation-related pathways  

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Full Text Available Abstract Background Polycystic Kidney Disease is characterized by the formation of large fluid-filled cysts that eventually destroy the renal parenchyma leading to end-stage renal failure. Although remarkable progress has been made in understanding the pathologic mechanism of the disease, the precise orchestration of the early events leading to cyst formation is still unclear. Abnormal cellular proliferation was traditionally considered to be one of the primary irregularities leading to cyst initiation and growth. Consequently, many therapeutic interventions have focused on targeting this abnormal proliferation, and some have even progressed to clinical trials. However, the role of proliferation in cyst development was primarily examined at stages where cysts are already visible in the kidneys and therefore at later stages of disease development. Methods In this study we focused on the cystic phenotype since birth in an attempt to clarify the temporal contribution of cellular proliferation in cyst development. Using a PKD2 transgenic rat model (PKD2 (1-703 of different ages (0-60 days after birth we performed gene expression profiling and phenotype analysis by measuring various kidney parameters. Results Phenotype analysis demonstrated that renal cysts appear immediately after birth in the PKD2 transgenic rat model (PKD2 (1-703. On the other hand, abnormal proliferation occurs at later stages of the disease as identified by gene expression profiling. Interestingly, other pathways appear to be deregulated at early stages of the disease in this PKD model. Specifically, gene expression analysis demonstrated that at day 0 the RAS system is involved. This is altered at day 6, when Wnt signaling and focal adhesion pathways are affected. However, at and after 24 days, proliferation, apoptosis, altered ECM signaling and many other factors become involved. Conclusions Our data suggest that cystogenesis precedes deregulation of proliferation-related pathways, suggesting that proliferation abnormalities may contribute in cyst growth rather than cyst formation.

Koupepidou Panayiota

2010-09-01

96

HIV-1 Transgenic Female Rat: Synaptodendritic Alterations of Medium Spiny Neurons in the Nucleus Accumbens.  

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HIV-1 associated neurocognitive deficits are increasing in prevalence, although the neuronal basis for these deficits is unclear. HIV-1 Tg rats constitutively express 7 of 9 HIV-associated proteins, and may be useful for studying the neuropathological substrates of HIV-1 associated neurocognitive disorders (HAND). In this study, adult female HIV-1 Tg rats and F344 control rats had similar growth rates, estrous cyclicity and startle reflex inhibition to a visual prepulse stimulus. Medium spiny neurons (MSNs) in the nucleus accumbens (NAcc) were ballistically-labeled utilizing the indocarbocyanine dye DiI. The branching complexity of MSNs in the NAcc was significantly decreased in HIV-1 Tg rats, relative to controls; moreover, the shorter length and decreased volume of dendritic spines, but unchanged head diameter, in HIV-1 Tg rats suggested a reduction of longer spines and an increase in shorter, less projected spines, indicating a population shift to a more immature spine phenotype. Collectively, these results from HIV-1 Tg female rats indicated significant synaptodendritic alterations of MSNs in the NAcc occur as a consequence of chronic, low-level, exposure to HIV-1 associated proteins. PMID:25037595

Roscoe, Robert F; Mactutus, Charles F; Booze, Rosemarie M

2014-12-01

97

Accumulation of chondroitin sulfate proteoglycans in the microenvironment of spinal motor neurons in amyotrophic lateral sclerosis transgenic rats.  

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Chondroitin sulfate proteoglycans (CSPGs) are the major components of extracellular matrix in the central nervous system. In the spinal cord under various types of injury, reactive gliosis emerges in the lesion accompanied by CSPG up-regulation. Several types of CSPG core proteins and their side chains have been shown to inhibit axonal regeneration in vitro and in vivo. In the present study, we examined spatiotemporal expression of CSPGs in the spinal cord of transgenic (Tg) rats with His46Arg mutation in the Cu/Zn superoxide dismutase gene, a model of amyotrophic lateral sclerosis (ALS). Immunofluorescence disclosed a significant up-regulation of neurocan, versican, and phosphacan in the ventral spinal cord of Tg rats compared with age-matched controls. Notably, Tg rats showed progressive and prominent accumulation of neurocan even at the presymptomatic stage. Immunoblotting confirmed the distinct increase in the levels of both the full-length neurocan and their fragment isoforms. On the other hand, the up-regulation of versican and phosphacan peaked at the early symptomatic stage, followed by diminishment at the late symptomatic stage. In addition, double immunofluorescence revealed a colocalization between reactive astrocytes and immunoreactivities for neurocan and phosphacan, especially around residual large ventral horn neurons. Thus, reactive astrocytes are suggested to be participants in the CSPG accumulation. Although the possible neuroprotective involvement of CSPG remains to be investigated, the present results suggest that both the reactive astrocytes and the differential accumulation of CSPGs may create a nonpermissive microenvironment for neural regeneration in neurodegenerative diseases such as ALS. PMID:18438943

Mizuno, Hideki; Warita, Hitoshi; Aoki, Masashi; Itoyama, Yasuto

2008-08-15

98

Bone marrow-derived mesenchymal stem cells expressing the Shh transgene promotes functional recovery after spinal cord injury in rats.  

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Spinal cord injury (SCI) is one of the most disabling diseases. Cell-based gene therapy is becoming a major focus for the treatment of SCI. Bone marrow-derived mesenchymal stem cells (BMSCs) are a promising stem cell type useful for repairing SCI. However, the effects of BMSCs transplants are likely limited because of low transplant survival after SCI. Sonic hedgehog (Shh) is a multifunctional growth factor which can facilitate neuronal and BMSCs survival, promote axonal growth, prevent activation of the astrocyte lineage, and enhance the delivery of neurotrophic factors in BMSCs. However, treatment of SCI with Shh alone also has limited effects on recovery, because the protein is cleared quickly. In this study, we investigated the use of BMSCs overexpressing the Shh transgene (Shh-BMSCs) in the treatment of rats with SCI, which could stably secrete Shh and thereby enhance the effects of BMSCs, in an attempt to combine the advantages of Shh and BMSCs and so to promote functional recovery. After Shh-BMSCs treatment of SCI via the subarachnoid, we detected significantly greater damage recovery compared with that seen in rats treated with phosphate-buffered saline (PBS) and BMSCs. Use of Shh-BMSCs increased the expression and secretion of Shh, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF), improved the behavioral function, enhanced the BMSCs survival, promoted the expression level of neurofilament 200 (NF200), and reduced the expression of glial fibrillary acidic protein (GFAP). Thus, our results indicated that Shh-BMSCs enhanced recovery of neurological function after SCI in rats and could be a potential valuable therapeutic intervention for SCI in humans. PMID:24837681

Jia, Yijia; Wu, Dou; Zhang, Ruiping; Shuang, Weibing; Sun, Jiping; Hao, Haihu; An, Qijun; Liu, Qiang

2014-06-24

99

Milk products containing bioactive tripeptides have an antihypertensive effect in double transgenic rats (dTGR) harbouring human renin and human angiotensinogen genes  

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Tripeptides isoleucyl-prolyl-proline (IPP) and valyl-prolyl-proline (VPP) act as ACE inhibitors in vitro. Double transgenic rats (dTGR) harbouring human renin and human angiotensinogen genes develop malignant hypertension due to increased angiotensin II formation. The present study was aimed to evaluate possible antihypertensive effect of IPP and VPP in this severe model. Four-week-old dTGR were randomized in three groups to receive: (1) water (control), (2) fermented milk containing IPP and ...

Eero Mervaala; Riitta Korpela; Heikki Vapaatalo; Amp Ller, Dominik N. M.; Hannu Kautiainen; Zhong Jian Cheng; Tiina Jauhiainen; Taru Pilvi

2010-01-01

100

Genotoxicity of phenacetin in the kidney and liver of Sprague-Dawley gpt delta transgenic rats in 26-week and 52-week repeated-dose studies.  

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Transgenic rat mutation assays can be used to assess genotoxic properties of chemicals in target organs for carcinogenicity. Mutations in transgenes are genetically neutral and accumulate during a treatment period; thus, assays are suitable for assessing the genotoxic risk of chemicals using a repeated-dose treatment paradigm. However, only a limited number of such studies have been conducted. To examine the utility of transgenic rat assays in repeated-dose studies, we fed male and female Sprague-Dawley gpt delta rats with a 0.5% phenacetin-containing diet for 26 and 52 weeks. A long-term feeding of phenacetin is known to induce renal cancer in rats. Phenacetin administration for 52 weeks in males significantly increased gpt (point mutations) mutant frequency (MF) in the kidney, the target organ of carcinogenesis. In the liver, the nontarget organ of carcinogenesis, gpt MFs were significantly elevated in phenacetin treatment groups of both genders during 26- and 52-week treatments. Furthermore, sensitive to P2 interference (Spi(-)deletions) MF increased in the liver of both genders following 52-week treatment. MFs were higher after treatment for 52 weeks than after treatment for 26 weeks. Frequencies of phenacetin-induced mutations were higher in the liver than in the kidney, suggesting that the intensity of genotoxicity does not necessarily correlate with the induction of tumor formation. Results from gpt delta rat assays of repeated-dose treatments are extremely useful to elucidate the relationship between gene mutations and carcinogenesis in the target organ induced by cancer-causing agents. PMID:25047350

Kawamura, Yuji; Hayashi, Hiroyuki; Masumura, Kenichi; Numazawa, Satoshi; Nohmi, Takehiko

2014-10-01

 
 
 
 
101

Cardiac remodeling during and after renin-angiotensin system stimulation in Cyp1a1-Ren2 transgenic rats.  

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This study investigated renin-angiotensin system (RAS)-induced cardiac remodeling and its reversibility in the presence and absence of high blood pressure (BP) in Cyp1a1-Ren2 transgenic inducible hypertensive rats (IHR). In IHR (pro)renin levels and BP can be dose-dependently titrated by oral administration of indole-3-carbinol (I3C). Young (four-weeks old) and adult (30-weeks old) IHR were fed I3C for four weeks (leading to systolic BP >200 mmHg). RAS-stimulation was stopped and animals were followed-up for a consecutive period. Cardiac function and geometry was determined echocardiographically and the hearts were excised for molecular and immunohistochemical analyses. Echocardiographic studies revealed that four weeks of RAS-stimulation incited a cardiac remodeling process characterized by increased left ventricular (LV) wall thickness, decreased LV volumes, and shortening of the left ventricle. Hypertrophic genes were highly upregulated, whereas in substantial activation a fibrotic response was absent. Four weeks after withdrawal of I3C, (pro)renin levels were normalized in all IHR. While in adult IHR BP returned to normal, hypertension was sustained in young IHR. Despite the latter, myocardial hypertrophy was fully regressed in both young and adult IHR. We conclude that (pro)renin-induced severe hypertension in IHR causes an age-independent fully reversible myocardial concentric hypertrophic remodeling, despite a continued elevated BP in young IHR. PMID:23462119

Heijnen, Bart F J; Pelkmans, Leonie P J; Danser, A H Jan; Garrelds, Ingrid M; Mullins, John J; De Mey, Jo G R; Struijker-Boudier, Harry A J; Janssen, Ben J A

2014-03-01

102

Dynamics of testicular germ cell apoptosis in normal mice and transgenic mice overexpressing rat androgen-binding protein  

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Full Text Available Abstract The number and type of testicular germ cells undergoing apoptosis in different age groups of mice (from 7 to 360 days of age was determined and compared in age-matched wild type (WT control and in a transgenic (TG mice homozygous to rat androgen binding protein (ABP using flow cytometry. Flow cytometric quantification revealed that the total number of germ cells undergoing apoptosis did not differ significantly in WT and TG mice up to Day 14. From Day 21 to Day 60, the number of germ cells undergoing apoptosis was consistently higher in TG than in WT mice. Starting from Day 90, the number of germ cells undergoing apoptosis in TG mice was lower than controls until Day 360. In 21–60 days old TG mice, spermatogonia, S-Phase cells, and primary spermatocytes are the cell types undergoing apoptosis at significantly greater numbers than those in WT mice. However, starting from day 60, the total number of spermatids undergoing apoptosis was significantly lower in TG mice than in age-matched WT controls. TdT-mediated dUTP-biotin nick end labeling (TUNEL in testicular sections from TG mice of 21 and 30 days of age confirmed the presence of increased numbers of apoptotic germ cells compared to their age matched controls. These data indicate that the continuous presence of greater than physiological concentrations of ABP in the mouse testis has a biphasic effect on the frequency of apoptosis in germ cells. The initial pre-pubertal increase in testicular germ cell apoptosis may result from direct or indirect actions of ABP and is likely to determine the subsequent life-death balance of germ cell populations in TG mice, whereas the subsequent reduction may result from maturation depletion. A wave of apoptosis during the pre-pubertal period is required for normal spermatogenesis to develop, and our data indicate that this apoptotic wave may be regulated by ABP and/or androgens.

Petrusz Peter

2003-06-01

103

Electrophysiological characteristics of inhibitory neurons of the prepositus hypoglossi nucleus as analyzed in Venus-expressing transgenic rats.  

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The identification and characterization of excitatory and inhibitory neurons are significant steps in understanding neural network functions. In this study, we investigated the intrinsic electrophysiological properties of neurons in the prepositus hypoglossi nucleus (PHN), a brainstem structure that is involved in gaze holding, using whole-cell recordings in brainstem slices from vesicular GABA transporter (VGAT)-Venus transgenic rats, in which inhibitory neurons express the fluorescent protein Venus. To characterize the intrinsic properties of these neurons, we recorded afterhyperpolarization (AHP) profiles and firing patterns from Venus-expressing [Venus?] and Venus-non-expressing [Venus?] PHN neurons. Although both types of neurons showed a wide variety of AHP profiles and firing patterns, oscillatory firing was specific to Venus? neurons, while a firing pattern showing only a few spikes was specific to Venus? neurons. In addition, AHPs without a slow component and delayed spike generation were preferentially displayed by Venus? neurons, whereas a firing pattern with constant interspike intervals was preferentially displayed by Venus? neurons. We evaluated the mRNAs expression of glutamate decarboxylase (GAD65, GAD67) and glycine transporter 2 (GlyT2) to determine whether the recorded Venus? neurons were GABAergic or glycinergic. Of the 67 Venus? neurons tested, GlyT2 expression alone was detected in only one neuron. Approximately 40% (28/67) expressed GAD65 and/or GAD67 (GABAergic neuron), and the remainder (38/67) expressed both GAD(s) and GlyT2 (GABA&GLY neuron). These results suggest that most inhibitory PHN neurons use either GABA or both GABA and glycine as neurotransmitters. Although the overall distribution of firing patterns in GABAergic neurons was similar to that of GABA&GLY neurons, only GABA&GLY neurons exhibited a firing pattern with a long first interspike interval. These differential electrophysiological properties will be useful for the identification of specific types of PHN neurons. PMID:21952130

Shino, M; Kaneko, R; Yanagawa, Y; Kawaguchi, Y; Saito, Y

2011-12-01

104

Transgenic mouse lines expressing rat AH receptor variants - A new animal model for research on AH receptor function and dioxin toxicity mechanisms  

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Han/Wistar (Kuopio; H/W) rats are exceptionally resistant to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity mainly because of their mutated aryl hydrocarbon receptor (AHR) gene. In H/W rats, altered splicing of the AHR mRNA generates two AHR proteins: deletion (DEL) and insertion (INS) variants, with the INS isoform being predominantly expressed. To gain further insight into their functional properties, cDNAs of these and rat wild-type (rWT) isoform were transferred into C57BL/6J-derived mice by microinjection. The endogenous mouse AHR was eliminated by selective crossing with Ahr-null mice. A single mouse line was obtained for each of the three constructs. The AHR mRNA levels in tissues were generally close to those of C57BL/6 mice in INS and DEL mice and somewhat higher in rWT mice; in testis, however, all 3 constructs exhibited marked overexpression. The transgenic mouse lines were phenotypically normal except for increased testis weight. Induction of drug-metabolizing enzymes by TCDD occurred similarly to that in C57BL/6 mice, but there tended to be a correlation with AHR concentrations, especially in testis. In contrast to C57BL/6 mice, the transgenics did not display any major gender difference in susceptibility to the acute lethality and hepatotoxicity of TCDD; rWT mice were highly sensitive, DEL mice moderately resistant and INS mice highly resistant. Co-expression of mouse AHR and rWT resulted in augmented sensitivity to TCDD and abolished the natural resistance of female C57BL/6 mice, whereas mice co-expressing mouse AHR and INS were resistant. Thus, these transgenic mouse lines provide a novel promising tool for molecular studies on dioxin toxicity and AHR function.

105

Progressive changes in microglia and macrophages in spinal cord and peripheral nerve in the transgenic rat model of amyotrophic lateral sclerosis  

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Full Text Available Abstract Background The role of neuroinflammation in motor neuron death of amyotrophic lateral sclerosis (ALS is unclear. The human mutant superoxide dismutase-1 (hmSOD1-expressing murine transgenic model of ALS has provided some insight into changes in microglia activity during disease progression. The purpose of this study was to gain further knowledge by characterizing the immunological changes during disease progression in the spinal cord and peripheral nerve using the more recently developed hmSOD1 rat transgenic model of ALS. Methods Using immunohistochemistry, the extent and intensity of tissue CD11b expression in spinal cord, lumbar nerve roots, and sciatic nerve were evaluated in hmSOD1 rats that were pre-clinical, at clinical onset, and near disease end-stage. Changes in CD11b expression were compared to the detection of MHC class II and CD68 microglial activation markers in the ventral horn of the spinal cord, as well as to the changes in astrocytic GFAP expression. Results Our study reveals an accumulation of microglia/macrophages both in the spinal cord and peripheral nerve prior to clinical onset based on CD11b tissue expression. The microglia formed focal aggregates in the ventral horn and became more widespread as the disease progressed. Hypertrophic astrocytes were not prominent in the ventral horn until after clinical onset, and the enhancement of GFAP did not have a strong correlation to increased CD11b expression. Detection of MHC class II and CD68 expression was found in the ventral horn only after clinical onset. The macrophages in the ventral nerve root and sciatic nerve of hmSOD1 rats were observed encircling axons. Conclusions These findings describe for the first time in the hmSOD1 rat transgenic model of ALS that enhancement of microglia/macrophage activity occurs pre-clinically both in the peripheral nerve and in the spinal cord. CD11b expression is shown to be a superior indicator for early immunological changes compared to other microglia activation markers and astrogliosis. Furthermore, we suggest that the early activity of microglia/macrophages is involved in the early phase of motor neuron degeneration and propose that studies involving immunomodulation in hmSOD1transgenic models need to consider effects on macrophages in peripheral nerves as well as to microglia in the spinal cord.

Hickey William F

2010-01-01

106

Novel transgenic mouse models develop retinal changes associated with early diabetic retinopathy similar to those observed in rats with diabetes mellitus.  

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Retinal capillary pericyte degeneration has been linked to aldose reductase (AR) activity in diabetic retinopathy (DR). Since the development of DR in mice and rats has been reported to differ and that this may be linked to differences in retinal sorbitol levels, we have established new murine models of early onset diabetes mellitus as tools for investigating the role of AR in DR. Transgenic diabetic mouse models were developed by crossbreeding diabetic C57BL/6-Ins2(Akita)/J (AK) with transgenic C57BL mice expressing green fluorescent protein (GFP), human aldose reductase (hAR) or both in vascular tissues containing smooth muscle actin-? (SMAA). Changes in retinal sorbitol levels were determined by HPLC while changes of growth factors and signaling were investigated by Western Blots. Retinal vascular changes were quantitatively analyzed on elastase-digestion flat mounts. Results show that sorbitol levels were higher in neural retinas of diabetic AK-SMAA-GFP-hAR compared to AK-SMAA-GFP mice. AK-SMAA-GFP-hAR mice showed induction of the retinal growth factors VEGF, IGF-1, bFGF and TGF?, as well as signaling changes in P-Akt, P-SAPK/JNK, and P-44/42 MAPK. Increased loss of nuclei per capillary length and a significant increase in the percentage of acellular capillaries presented in 18 week old AK-SMAA-GFP-hAR mice. These changes are similar to those observed in streptozotocin-induced diabetic rats. Retinal changes in both mice and rats were prevented by inhibition of AR. These studies confirm that the increased expression of AR in mice results in the development of retinal changes associated with the early stages of DR that are similar to those observed in rats. PMID:24370601

Guo, Changmei; Zhang, Zifeng; Zhang, Peng; Makita, Jun; Kawada, Hiroyoshi; Blessing, Karen; Kador, Peter F

2014-02-01

107

Adeno-associated viral vector serotypes 1 and 5 targeted to the neonatal rat and pig striatum induce widespread transgene expression in the forebrain  

DEFF Research Database (Denmark)

Viral vector-mediated gene transfer has emerged as a powerful means to target transgene expression in the central nervous system. Here we characterized the efficacy of serotypes 1 and 5 recombinant adeno-associated virus (rAAV) vectors encoding green fluorescent protein (GFP) after stereotaxic delivery to the neonatal rat and minipig striatum. The efficiency of GFP expression and the phenotype of GFP-positive cells were assessed within the forebrain at different time points up to 12 months after surgery. Both rAAV1-GFP and rAAV5-GFP delivery resulted in transduction of the striatum as well as striatal input and output areas, including large parts of the cortex. In both species, rAAV5 resulted in a more widespread transgene expression compared to rAAV1. In neonatal rats, rAAV5 also transduced several other areas such as the olfactory bulbs, hippocampus, and septum. Phenotypic analysis of the GFP-positive cells, performed using immunohistochemistry and confocal microscopy, showed that most of the GFP-positive cells by either serotype were NeuN-positive neuronal profiles. The rAAV5 vector further displayed the ability to transduce non-neuronal cell types in both rats and pigs, albeit at a low frequency. Our results show that striatal delivery of rAAV5 vectors in the neonatal brain represents a useful tool to express genes of interest both in the basal ganglia and the neocortex. Furthermore, we apply, for the first time, viral vector-mediated gene transfer to the pig brain providing the opportunity to study effects of genetic manipulation in this non-primate large animal species. Finally, we generated an atlas of the Göttingen minipig brain for guiding future studies in this large animal species.

Kornum, Birgitte R; Stott, Simon R W

2010-01-01

108

Combined Renin Inhibition/(Pro)Renin Receptor Blockade in Diabetic Retinopathy- A Study in Transgenic (mREN2)27 Rats  

Science.gov (United States)

Dysfunction of renin-angiotensin system (RAS) contributes to the pathogenesis of diabetic retinopathy (DR). Prorenin, the precursor of renin is highly elevated in ocular fluid of diabetic patients with proliferative retinopathy. Prorenin may exert local effects in the eye by binding to the so-called (pro)renin receptor ((P)RR). Here we investigated the combined effects of the renin inhibitor aliskiren and the putative (P)RR blocker handle-region peptide (HRP) on diabetic retinopathy in streptozotocin (STZ)-induced diabetic transgenic (mRen2)27 rats (a model with high plasma prorenin levels) as well as prorenin stimulated cytokine expression in cultured Müller cells. Adult (mRen2)27 rats were randomly divided into the following groups: (1) non-diabetic; (2) diabetic treated with vehicle; (3) diabetic treated with aliskiren (10 mg/kg per day); and (4) diabetic treated with aliskiren+HRP (1 mg/kg per day). Age-matched non-diabetic wildtype Sprague-Dawley rats were used as control. Drugs were administered by osmotic minipumps for three weeks. Transgenic (mRen2)27 rat retinas showed increased apoptotic cell death of both inner retinal neurons and photoreceptors, increased loss of capillaries, as well as increased expression of inflammatory cytokines. These pathological changes were further exacerbated by diabetes. Aliskiren treatment of diabetic (mRen2)27 rats prevented retinal gliosis, and reduced retinal apoptotic cell death, acellular capillaries and the expression of inflammatory cytokines. HRP on top of aliskiren did not provide additional protection. In cultured Müller cells, prorenin significantly increased the expression levels of IL-1? and TNF-?, and this was completely blocked by aliskiren or HRP, their combination, (P)RR siRNA and the AT1R blocker losartan, suggesting that these effects entirely depended on Ang II generation by (P)RR-bound prorenin. In conclusion, the lack of effect of HRP on top of aliskiren, and the Ang II-dependency of the ocular effects of prorenin in vitro, argue against the combined application of (P)RR blockade and renin inhibition in diabetic retinopathy. PMID:24968134

Batenburg, Wendy W.; Verma, Amrisha; Wang, Yunyang; Zhu, Ping; van den Heuvel, Mieke; van Veghel, Richard; Danser, A. H. Jan; Li, Qiuhong

2014-01-01

109

Benzo[a]pyrene-enhanced mutagenesis by man-made mineral fibres in the lung of lamda-lacI transgenic rats.  

Science.gov (United States)

In an attempt to examine the interaction of man-made mineral fibres with benzo[a]pyrene (B[a]P), homozygous X-lacI transgenic F344 rats were intratracheally treated with rock (stone) wool RWI and glass wool MMVF 10 fibres together with B[a]P. To analyze the induction of gene mutations by fibres and B[a]P in lung, single doses of 1 and 2 mg fibres/animal or multiple doses of 2 mg fibres/animal were administered weekly on 4 consecutive weeks (total dose 8 mg/animal). B[a]P (10 mg/animal) was administered either simultaneously with fibres (for single dose treatment with fibres) or together with the last fiber treatment (for multiple dose treatment with fibres). Animals were scarified 4 weeks after the last treatment. Benzo[a]pyrene administered simultaneously with RW1 fibres exhibited a strong synergistic effect on mutagenicity, the observed mutant frequency (MF) being more than three-fold higher than the net sum of the MF induced after separate administration of both agents. Our data suggest that DNA adducts induced by simultaneous B[a]P and fiber treatment lead to a strong increase in mutatant frequencies. PMID:16375931

Topinka, J; Loli, P; Hurbáková, M; Kováciková, Z; Volkovová, K; Wolff, T; Oesterle, D; Kyrtopoulos, S A; Georgiadis, P

2006-03-20

110

Development of Oral Epithelial Cell Line ROE2 with Differentiation Potential from Transgenic Rats Harboring Temperature-Sensitive Simian Virus40 Large T-Antigen Gene  

Science.gov (United States)

We have developed an immortalized oral epithelial cell line, ROE2, from fetal transgenic rats harboring temperature-sensitive simian virus 40 large T-antigen gene. The cells grew continuously at either a permissive temperature of 33°C or an intermediate temperature of 37°C. At the nonpermissive temperature of 39°C, on the other hand, growth decreased significantly, and the Sub-G1 phase of the cell cycle increased, indicating that the cells undergo apoptosis at a nonpermissive temperature. Histological and immunocytochemical analyses revealed that ROE2 cells at 37°C had a stratified epithelial-like morphology and expressed cytokeratins Krt4 and Krt13, marker proteins for oral nonkeratinized epithelial cells. Global-scale comprehensive microarray analysis, coupled with bioinformatics tools, demonstrated a significant gene network that was obtained from the upregulated genes. The gene network contained 16 genes, including Cdkn1a, Fos, Krt13, and Prdm1, and was associated mainly with the biological process of skin development in the category of biological functions, organ development. These four genes were validated by quantitative real-time polymerase chain reaction, and the results were nearly consistent with the microarray data. It is therefore anticipated that this cell line will be useful as an in vitro model for studies such as physiological functions, as well as for gene expression in oral epithelial cells. PMID:24521861

Tabuchi, Yoshiaki; Wada, Shigehito; Ikegame, Mika; Kariya, Ayako; Furusawa, Yukihiro; Hoshi, Nobuhiko; Yunoki, Tatsuya; Suzuki, Nobuo; Takasaki, Ichiro; Kondo, Takashi; Suzuki, Yoshihisa

2014-01-01

111

Retinal ganglion cell survival and axon regeneration in WldS transgenic rats after optic nerve crush and lens injury  

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Abstract Background We have previously shown that the slow Wallerian degeneration mutation, whilst delaying axonal degeneration after optic nerve crush, does not protect retinal ganglion cell (RGC) bodies in adult rats. To test the effects of a combination approach protecting both axons and cell bodies we performed combined optic nerve crush and lens injury, which results in both enhanced RGC survival as well as axon regeneration past the lesion site in wildtype animals. Results As previously...

Lorber, Barbara; Tassoni, Alessia; Bull, Natalie D.; Moschos, Marilita M.; Martin, Keith R.

2012-01-01

112

Transgenic livestock.  

Science.gov (United States)

Single genes can now be added routinely to the genome of mice by molecular manipulation as simple Mendelian dominants; this complements the normal process of reproduction to give 'transgenic' animals. Success in ruminants is limited to a few examples in sheep and although gene expression has yet to be documented, there is every reason to expect that it will be achieved. The application of this technology to livestock improvement depends on the identification of circumstances in which the phenotype is limited by the deficiency of a single protein. While there is little evidence to indicate that single dominant genes are in general likely to have favourable effects, it is argued that there are likely to be exceptions. These include particular combinations of promoter and structural gene sequences to alter feedback control, for example through a change in tissue specificity, and the alteration of definitive proteins such as those of milk. A mouse model has been established to study the molecular manipulation of sheep milk proteins. The sheep beta lactoglobulin gene has been incorporated and the sheep whey protein is secreted by the mammary gland of transgenic mice. For the future, means to delete or reduce the expression of existing genes are likely to be important, as are more effective means of incorporation such as retroviral based methods and the incorporation of multigene constructs. The resources required to test transgenic livestock will, however, be greater than those required to create them. PMID:3305921

Simons, J P; Land, R B

1987-01-01

113

Lesion-dependent regulation of transgene expression in the rat brain using a human glial fibrillary acidic protein-lentiviral vector.  

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The ability to regulate transgene expression will be crucial for development of gene therapy to the brain. The most commonly used systems are based on a transactivator in combination with a drug, e.g. the tetracycline-regulated system. Here we describe a different method of transgene regulation by the use of the human glial fibrillary acidic protein (GFAP) promoter. We constructed a lentiviral vector that directs transgene expression to astrocytes. Using toxin-induced lesions we investigated ...

Jakobsson, Johan; Georgievska, Biljana; Ericson, Cecilia; Lundberg, Cecilia

2004-01-01

114

Nogo-A-deficient transgenic rats show deficits in higher cognitive functions, decreased anxiety and altered circadian activity patterns  

Directory of Open Access Journals (Sweden)

Full Text Available Decreased levels of Nogo-A dependent signaling have been shown to affect behavior and cognitive functions. In Nogo-A knockout and knock-down laboratory rodents, behavioral alterations were observed, possibly corresponding with human neuropsychiatric diseases of neurodevelopmental origin, particularly schizophrenia. This study offers further insight into behavioral manifestations of Nogo-A knockdown in laboratory rats, focusing on spatial and non-spatial cognition, anxiety levels, circadian rhythmicity and activity patterns. Demonstrated is an impairment of cognitive functions and behavioral flexibility in a spatial active avoidance task, while non-spatial memory in a step-through avoidance task was spared. No signs of anhedonia, typical for schizophrenic patients, were observed in the animals. Some measures indicated lower anxiety levels in the Nogo-A deficient group. Circadian rhythmicity in locomotor activity was preserved in the Nogo-A-knockout rats and their circadian period (tau did not differ from controls. However, daily activity patterns were slightly altered in the knockdown animals. We conclude that a reduction of Nogo-A levels induces changes in CNS development, manifested as subtle alterations in cognitive functions, emotionality and activity patterns.

TomasPetrasek

2014-03-01

115

Defining peripheral nervous system dysfunction in the SOD-1G93A transgenic rat model of amyotrophic lateral sclerosis.  

Science.gov (United States)

Growing evidence indicates that alterations within the peripheral nervous system (PNS) are involved at an early stage in the amyotrophic lateral sclerosis (ALS) pathogenetic cascade. In this study, magnetic resonance imaging (MRI), neurophysiologic analyses, and histologic analyses were used to monitor the extent of PNS damage in the hSOD-1 ALS rat model. The imaging signature of the disease was defined using in vivo MRI of the sciatic nerve. Initial abnormalities were detected in the nerves by an increase in T2 relaxation time before the onset of clinical disease; diffusion MRI showed a progressive increase in mean and radial diffusivity and reduction of fractional anisotropy at advanced stages of disease. Histologic analysis demonstrated early impairment of the blood-nerve barrier followed by acute axonal degeneration associated with endoneurial edema and macrophage response in motor nerve compartments. Progressive axonal degeneration and motor nerve fiber loss correlated with MRI and neurophysiologic changes. These functional and morphologic investigations of the PNS might be applied in following disease progression in preclinical therapeutic studies. This study establishes the PNS signature in this rat ALS model (shedding new light into pathogenesis) and provides a rationale for translating into future systematic MRI studies of PNS involvement in patients with ALS. PMID:24918640

Riva, Nilo; Chaabane, Linda; Peviani, Marco; Ungaro, Daniela; Domi, Teuta; Dina, Giorgia; Bianchi, Francesca; Spano, Giorgia; Cerri, Federica; Podini, Paola; Corbo, Massimo; Carro, Ubaldo Del; Comi, Giancarlo; Bendotti, Caterina; Quattrini, Angelo

2014-07-01

116

Gene expression changes consistent with neuroAIDS and impaired working memory in HIV-1 transgenic rats  

Science.gov (United States)

Background A thorough investigation of the neurobiology of HIV-induced neuronal dysfunction and its evolving phenotype in the setting of viral suppression has been limited by the lack of validated small animal models to probe the effects of concomitant low level expression of multiple HIV-1 products in disease-relevant cells in the CNS. Results We report the results of gene expression profiling of the hippocampus of HIV-1 Tg rats, a rodent model of HIV infection in which multiple HIV-1 proteins are expressed under the control of the viral LTR promoter in disease-relevant cells including microglia and astrocytes. The Gene Set Enrichment Analysis (GSEA) algorithm was used for pathway analysis. Gene expression changes observed are consistent with astrogliosis and microgliosis and include evidence of inflammation and cell proliferation. Among the genes with increased expression in HIV-1 Tg rats was the interferon stimulated gene 15 (ISG-15), which was previously shown to be increased in the cerebrospinal fluid (CSF) of HIV patients and to correlate with neuropsychological impairment and neuropathology, and prostaglandin D2 (PGD2) synthase (Ptgds), which has been associated with immune activation and the induction of astrogliosis and microgliosis. GSEA-based pathway analysis highlighted a broad dysregulation of genes involved in neuronal trophism and neurodegenerative disorders. Among the latter are genesets associated with Huntington’s disease, Parkinson’s disease, mitochondrial, peroxisome function, and synaptic trophism and plasticity, such as IGF, ErbB and netrin signaling and the PI3K signal transduction pathway, a mediator of neural plasticity and of a vast array of trophic signals. Additionally, gene expression analyses also show altered lipid metabolism and peroxisomes dysfunction. Supporting the functional significance of these gene expression alterations, HIV-1 Tg rats showed working memory impairments in spontaneous alternation behavior in the T-Maze, a paradigm sensitive to prefrontal cortex and hippocampal function. Conclusions Altogether, differentially regulated genes and pathway analysis identify specific pathways that can be targeted therapeutically to increase trophic support, e.g. IGF, ErbB and netrin signaling, and reduce neuroinflammation, e.g. PGD2 synthesis, which may be beneficial in the treatment of chronic forms of HIV-associated neurocognitive disorders in the setting of viral suppression. PMID:24980976

2014-01-01

117

The vestibulo- and preposito-cerebellar cholinergic neurons of a ChAT-tdTomato transgenic rat exhibit heterogeneous firing properties and the expression of various neurotransmitter receptors.  

Science.gov (United States)

Cerebellar function is regulated by cholinergic mossy fiber inputs that are primarily derived from the medial vestibular nucleus (MVN) and prepositus hypoglossi nucleus (PHN). In contrast to the growing evidence surrounding cholinergic transmission and its functional significance in the cerebellum, the intrinsic and synaptic properties of cholinergic projection neurons (ChPNs) have not been clarified. In this study, we generated choline acetyltransferase (ChAT)-tdTomato transgenic rats, which specifically express the fluorescent protein tdTomato in cholinergic neurons, and used them to investigate the response properties of ChPNs identified via retrograde labeling using whole-cell recordings in brainstem slices. In response to current pulses, ChPNs exhibited two afterhyperpolarisation (AHP) profiles and three firing patterns; the predominant AHP and firing properties differed between the MVN and PHN. Morphologically, the ChPNs were separated into two types based on their soma size and dendritic extensions. Analyses of the firing responses to time-varying sinusoidal current stimuli revealed that ChPNs exhibited different firing modes depending on the input frequencies. The maximum frequencies in which each firing mode was observed were different between the neurons that exhibited distinct firing patterns. Analyses of the current responses to the application of neurotransmitter receptor agonists revealed that the ChPNs expressed (i) AMPA- and NMDA-type glutamate receptors, (ii) GABAA and glycine receptors, and (iii) muscarinic and nicotinic acetylcholine receptors. The current responses mediated by these receptors of MVN ChPNs were not different from those of PHN ChPNs. These findings suggest that ChPNs receive various synaptic inputs and encode those inputs appropriately across different frequencies. PMID:24593297

Zhang, Yue; Kaneko, Ryosuke; Yanagawa, Yuchio; Saito, Yasuhiko

2014-04-01

118

Prostate cancer in a transgenic mouse.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Progress toward understanding the biology of prostate cancer has been slow due to the few animal research models available to study the spectrum of this uniquely human disease. To develop an animal model for prostate cancer, several lines of transgenic mice were generated by using the prostate-specific rat probasin promoter to derive expression of the simian virus 40 large tumor antigen-coding region. Mice expressing high levels of the transgene display progressive forms of prostatic disease ...

Greenberg, N. M.; Demayo, F.; Finegold, M. J.; Medina, D.; Tilley, W. D.; Aspinall, J. O.; Cunha, G. R.; Donjacour, A. A.; Matusik, R. J.; Rosen, J. M.

1995-01-01

119

Transgenic bioreactors.  

Science.gov (United States)

1. Although many human therapeutic proteins are currently produced in microbial fermentors using recombinant DNA techniques, it is obvious that microbial processing is not suitable for a large number of bioactive proteins owing to the inability of bacteria to carry out postsynthetic modification reactions required for full biological activity. 2. This disadvantage does not apply to animal cell bioreactors that can generate biologically fully active entities, yet the use of large-scale animal cell cultures for production purposes is prohibitively expensive. 3. With the advent of transgenic technology, the production of valuable human pharmaceuticals in large farm animals (pig, sheep, goat and dairy cattle) has become more and more attractive as a high-quantity, low-cost alternative. By employing targeted gene transfer, e.g. using mammary gland-specific regulatory sequences fused with the desired production genes, it is possible to govern the expression to occur exclusively in the mammary gland and hence the gene product is being ultimately secreted in the milk. 4. While reviewing the remarkable progress in this field that has even led to commercial exploitations, we will outline in somewhat greater detail our strategy for the use of dairy cattle as a bioreactor for valuable proteins of pharmaceutical interest. PMID:8063010

Jänne, J; Hyttinen, J M; Peura, T; Tolvanen, M; Alhonen, L; Sinervirta, R; Halmekytö, M

1994-07-01

120

Activation and Repression Domains Within the Promoter of the Rat Cathepsin L Gene Orchestrate Sertoli Cell-Specific and Stage-Specific Gene Transcription in Transgenic Mice1  

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In murine testes, only Sertoli cells express the cathepsin L (Ctsl) gene, and this expression is restricted to stages V–VIII of the cycle. Our previous transgenic analysis of Tg (?2065/+977) demonstrated that this expression is regulated by a ?2-kb promoter. To begin to elucidate this regulation, we analyzed the in vivo expression of two new transgenes, Tg (?935/+977) and Tg (?451/+977). Tg (?935/+977) was expressed by Sertoli cells but, in contrast to Tg (?2065/+977), was expre...

Visone, Thomas; Charron, Martin; Wright, William W.

2009-01-01

 
 
 
 
121

Detection of a novel HLA-B27 allele, B*2740, in Taiwanese volunteer bone marrow donors by sequence-based typing: curiosity rewarded.  

Science.gov (United States)

We report here a novel HLA-B allele, B*2740, discovered in Taiwanese volunteer marrow donors. The new sequence has nucleotide variation at position 527 (T-->A) as compared to B*2708. The nucleotide change caused an amino acid substitution from valine (V) to glutamic acid (E) at codon 152. Since B*2740 carries sequence confers to HLA-Bw6 public epitope we believe that this novel B*27 allele might have been generated from a gene conversion involving a Bw4-specific allele (probably B*2704) and a Bw6-specific allele. PMID:19476482

Chen, M J; Yang, T C; Chu, C C; Shyr, M H; Lin, C L; Lin, P Y; Yang, K L

2009-08-01

122

HLA-B27 i HLA-DR w prognozowaniu przebiegu m?odzie?czego idiopatycznego zapalenia stawów o pocz?tku uogólnionym  

Directory of Open Access Journals (Sweden)

Full Text Available Przebieg m?odzie?czego idiopatycznego zapalenia stawówo pocz?tku uogólnionym (UMIZS jest zró?nicowany. W artykuleoceniono wp?yw antygenu B27 i serii HLA klasy II – DR na post?pchoroby, ci??ko?? objawów pozastawowych oraz rozwój amyloidozyu 47 chorych z UMIZS z wieloletnim czasem trwania choroby(?rednio 18 ±7,4 roku. G?ównymi parametrami klinicznymi istotniewp?ywaj?cymi na losy chorych, które poddano analizie, by?yzmiany radiologiczne, wydolno?? czynno?ciowa, ci??ko?? zmianstawowych oraz rozwój amyloidozy. U ka?dego pacjenta okre?lonorównie? ci??ko?? objawów uogólnienia, które obserwowanow czasie d?ugoletniej choroby.Wykazano istotn? zale?no?? pomi?dzy obecno?ci? HLA-DR4 a rozwojemzmian radiologicznych w uk?adzie ruchu. HLA-DR4 stwierdzonoznamiennie cz??ciej u chorych ze znacznymi zmianamiradiologicznymi w porównaniu z grup? kontroln? (73,7 vs 23,6%,p < 0,0001, a tak?e w stosunku do chorych bez tych zmian (73,7vs 25%, p < 0,05 (ryc. 4. Antygen DR4 wykrywano równie? znamienniecz??ciej w grupie chorych z najbardziej zaawansowanymizmianami stawowymi w porównaniu z grup? kontroln? (63 vs24%, p < 0,001 (ryc. 2. Istotne powi?zanie z wyst?powaniemobjawów uogólnienia dotyczy?o natomiast HLA-DR3. HLA-DR3istotnie cz??ciej w porównaniu z grup? kontroln? stwierdzono u chorych z objawami uogólnienia wyst?puj?cymi nie tylko napocz?tku choroby, ale równie? w jej dalszym przebiegu (76,2 vs23,6%, p < 0,001, jak i pomi?dzy grup? pacjentów z objawamiuogólnienia ograniczonymi do 2 lat choroby a podgrup? chorychz objawami nawracaj?cymi w dalszym przebiegu choroby (22,7vs 76,2%, p < 0,001 (ryc. 1.Cz?sto?? typowanych antygenów w wyodr?bnionych podgrupach,zarówno w zale?no?ci od wydolno?ci uk?adu ruchu, jak i rozwojuamyloidozy nie ró?ni?a si? istotnie statystycznie.Typowanie HLA mo?e by? pomocne w prognozowaniu dalszegoprzebiegu UMIZS, szczególnie pomaga w identyfikacji chorych,u których dochodzi do zmian destrukcyjnych oraz nawracania objawówuogólnienia w dalszym przebiegu choroby.

Agnieszka Gazda

2011-02-01

123

Angiotensin-converting enzyme inhibition, but not AT(1) receptor blockade, in the solitary tract nucleus improves baroreflex sensitivity in anesthetized transgenic hypertensive (mRen2)27 rats.  

Science.gov (United States)

Transgenic hypertensive (mRen2)27 rats overexpress the murine Ren2 gene and have impaired baroreflex sensitivity (BRS) for control of the heart rate. Removal of endogenous angiotensin (Ang)-(1-7) tone using a receptor blocker does not further lower BRS. Therefore, we assessed whether blockade of Ang II with a receptor antagonist or combined reduction in Ang II and restoration of endogenous Ang-(1-7) levels with Ang-converting enzyme (ACE) inhibition will improve BRS in these animals. Bilateral solitary tract nucleus (nTS) microinjections of the AT(1) receptor blocker, candesartan (CAN, 24?pmol in 120?nl, n=9), or a peptidic ACE inhibitor, bradykinin (BK) potentiating nonapeptide (Pyr-Trp-Pro-Arg-Pro-Gln-Ile-Pro-Pro; BPP9?, 9?nmol in 60?nl, n=12), in anesthetized male (mRen2)27 rats (15-25 weeks of age) show that AT(1) receptor blockade had no significant effect on BRS, whereas microinjection of BPP9? improved BRS over 60-120?min. To determine whether Ang-(1-7) or BK contribute to the increase in BRS, separate experiments using the Ang-(1-7) receptor antagonist D-Ala(7)-Ang-(1-7) or the BK antagonist HOE-140 showed that only the Ang-(1-7) receptor blocker completely reversed the BRS improvement. Thus, acute AT(1) blockade is unable to reverse the effects of long-term Ang II overexpression on BRS, whereas ACE inhibition restores BRS over this same time frame. As the BPP9? potentiation of BK actions is a rapid phenomenon, the likely mechanism for the observed delayed increase in BRS is through ACE inhibition and elevation of endogenous Ang-(1-7). PMID:21937997

Isa, Katsunori; Arnold, Amy C; Westwood, Brian M; Chappell, Mark C; Diz, Debra I

2011-12-01

124

Oxidative Stress-Mediated Mitochondrial Dysfunction Contributes to Angiotensin II-Induced Nonalcoholic Fatty Liver Disease in Transgenic Ren2 Rats  

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Emerging evidence indicates that impaired mitochondrial fatty acid ?-oxidation plays a key role in liver steatosis. We have recently demonstrated that increased angiotensin (ANG) II causes progressive hepatic steatosis associated with oxidative stress; however, the underlying mechanisms remain unclear. We hypothesized that ANG II causes hepatic mitochondrial oxidative damage and impairs mitochondrial ?-oxidation, thereby leading to hepatic steatosis. We used the Ren2 rat with elevated endog...

Wei, Yongzhong; Clark, Suzanne E.; Thyfault, John P.; Uptergrove, Grace M. E.; Li, Wenhan; Whaley-connell, Adam T.; Ferrario, Carlos M.; Sowers, James R.; Ibdah, Jamal A.

2009-01-01

125

Transgenic animal models for the functional analysis of vasoactive peptides  

Directory of Open Access Journals (Sweden)

Full Text Available The interplay of vasoactive peptide systems is an essential determinant of blood pressure regulation in mammals. While the endothelin and the renin-angiotensin systems raise blood pressure by inducing vasoconstriction and sodium retention, the kallikrein-kinin and the natriuretic-peptide systems reduce arterial pressure by eliciting vasodilatation and natriuresis. Transgenic technology has proven to be very useful for the functional analysis of vasoactive peptide systems. As an outstanding example, transgenic rats overexpressing the mouse Ren-2 renin gene in several tissues become extremely hypertensive. Several other transgenic rat and mouse strains with genetic modifications of components of the renin-angiotensin system have been developed in the past decade. Moreover, in recent years gene-targeting technology was employed to produce mouse strains lacking these proteins. The established animal models as well as the main insights gained by their analysis are summarized in this review.

Bader M.

1998-01-01

126

Transgenic animal models for the functional analysis of vasoactive peptides  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english The interplay of vasoactive peptide systems is an essential determinant of blood pressure regulation in mammals. While the endothelin and the renin-angiotensin systems raise blood pressure by inducing vasoconstriction and sodium retention, the kallikrein-kinin and the natriuretic-peptide systems red [...] uce arterial pressure by eliciting vasodilatation and natriuresis. Transgenic technology has proven to be very useful for the functional analysis of vasoactive peptide systems. As an outstanding example, transgenic rats overexpressing the mouse Ren-2 renin gene in several tissues become extremely hypertensive. Several other transgenic rat and mouse strains with genetic modifications of components of the renin-angiotensin system have been developed in the past decade. Moreover, in recent years gene-targeting technology was employed to produce mouse strains lacking these proteins. The established animal models as well as the main insights gained by their analysis are summarized in this review.

M., Bader.

127

Transgenic animal bioreactors.  

Science.gov (United States)

The production of recombinant proteins is one of the major successes of biotechnology. Animal cells are required to synthesize proteins with the appropriate post-translational modifications. Transgenic animals are being used for this purpose. Milk, egg white, blood, urine, seminal plasma and silk worm cocoon from transgenic animals are candidates to be the source of recombinant proteins at an industrial scale. Although the first recombinant protein produced by transgenic animals is expected to be in the market in 2000, a certain number of technical problems remain to be solved before the various systems are optimized. Although the generation of transgenic farm animals has become recently easier mainly with the technique of animal cloning using transfected somatic cells as nuclear donor, this point remains a limitation as far as cost is concerned. Numerous experiments carried out for the last 15 years have shown that the expression of the transgene is predictable only to a limited extent. This is clearly due to the fact that the expression vectors are not constructed in an appropriate manner. This undoubtedly comes from the fact that all the signals contained in genes have not yet been identified. Gene constructions thus result sometime in poorly functional expression vectors. One possibility consists in using long genomic DNA fragments contained in YAC or BAC vectors. The other relies on the identification of the major important elements required to obtain a satisfactory transgene expression. These elements include essentially gene insulators, chromatin openers, matrix attached regions, enhancers and introns. A certain number of proteins having complex structures (formed by several subunits, being glycosylated, cleaved, carboxylated...) have been obtained at levels sufficient for an industrial exploitation. In other cases, the mammary cellular machinery seems insufficient to promote all the post-translational modifications. The addition of genes coding for enzymes involved in protein maturation has been envisaged and successfully performed in one case. Furin gene expressed specifically in the mammary gland proved to able to cleave native human protein C with good efficiency. In a certain number of cases, the recombinant proteins produced in milk have deleterious effects on the mammary gland function or in the animals themselves. This comes independently from ectopic expression of the transgenes and from the transfer of the recombinant proteins from milk to blood. One possibility to eliminate or reduce these side-effects may be to use systems inducible by an exogenous molecule such as tetracycline allowing the transgene to be expressed only during lactation and strictly in the mammary gland. The purification of recombinant proteins from milk is generally not particularly difficult. This may not be the case, however, when the endogenous proteins such as serum albumin or antibodies are abundantly present in milk. This problem may be still more crucial if proteins are produced in blood. Among the biological contaminants potentially present in the recombinant proteins prepared from transgenic animals, prions are certainly those raising the major concern. The selection of animals chosen to generate transgenics on one hand and the elimination of the potentially contaminated animals, thanks to recently defined quite sensitive tests may reduce the risk to an extremely low level. The available techniques to produce pharmaceutical proteins in milk can be used as well to optimize milk composition of farm animals, to add nutriceuticals in milk and potentially to reduce or even eliminate some mammary infectious diseases. PMID:11131009

Houdebine, L M

2000-01-01

128

Calcium electrotransfer for termination of transgene expression in muscle  

DEFF Research Database (Denmark)

Gene electrotransfer is expanding in clinical use, thus we have searched for an emergency procedure to stop transgene expression in case of serious adverse events. Calcium is cytotoxic at high intracellular levels, so we tested effects of calcium electrotransfer on transgene expression in muscle. A clinical grade calcium solution (20 ?l, 168 mM) was injected into transfected mouse or rat tibialis cranialis muscle. Ca(2+) uptake was quantified using calcium 45 ((45)Ca), and voltage and time between injection and pulsation were varied. Extinction of transgene expression was investigated by using both in vivo imaging of infrared fluorescent "Katushka" and erythropoietin evaluated by ELISA and hemoglobin. Histology was performed. Electrotransfer of Katushka and erythropoietin yielded significant expression. Maximal calcium uptake occurred after injection of Ca(2+) before electropulsing using eight high voltage pulses of 1000 V/cm. Using these parameters, in vivo imaging showed that transgene expression significantly decreased 4 hr after Ca(2+) electrotransfer and was eliminated within 24 hr. Similarly, serum erythropoietin was reduced by 46% at 4 hr and to control levels at 2 days. Histological analyses showed muscle damage and subsequent regeneration. Electrotransfer of isotonic CaCl(2) terminates transgenic protein expression in muscles and may be used for contingency elimination of transgene expression.

Hojman, Pernille; Spanggaard, Iben

2011-01-01

129

Expression of human growth hormone in the milk of transgenic rabbits with transgene mapped to the telomere region of chromosome 7q.  

Science.gov (United States)

The advent of transgenic technology has provided methods for the production of pharmaceuticals by the isolation of these proteins from transgenic animals. The mammary gland has been focused on as a bioreactor, since milk is easily collected from lactating animals and protein production can be expressed at very high levels, including hormones and enzymes. We demonstrate here the expression pattern of recombinant human growth hormone (rhGH) in transgenic rabbits carrying hGH genomic sequences driven by the rat whey acidic protein (WAP) promoter. The transgene was mapped to the q26-27 telomere region of chromosome 7q by fluorescence in situ hybridization (FISH). Nearly 30 % of the F1 generation demonstrated the presence of transgene. The recombinant growth hormone was detected in the milk of the transgenic rabbit females, but not in serum, up to the level of 10 ?g/ml. Ectopic expression of the transgene in the brain, heart, kidney, liver, and salivary gland was not observed, indicating that a short sequence of rat WAP promoter (969 bp) contained essential sequences directing expression exclusively to the mammary gland. The biological activity of recombinant growth hormone was measured by immunoreactivity and the capability to stimulate growth of the hormone-dependent Nb211 cell line. PMID:22898896

Lipinski, Daniel; Zeyland, Joanna; Szalata, Marlena; Plawski, Andrzej; Jarmuz, Malgorzata; Jura, Jacek; Korcz, Aleksandra; Smorag, Zdzislaw; Pienkowski, Marek; Slomski, Ryszard

2012-11-01

130

Hypothalamic vasopressin response to stress and various physiological stimuli: visualization in transgenic animal models.  

Science.gov (United States)

Arginine vasopressin (AVP) is involved in the homeostatic responses numerous life-threatening conditions, for example, the promotion of water conservation during periods of dehydration, and the activation of the hypothalamo-pituitary adrenal axis by emotional stress. Recently, we generated new transgenic animals that faithfully express an AVP-enhanced green fluorescent protein (eGFP) fusion gene in the paraventricular nucleus (PVN), the supraoptic nucleus (SON) and the suprachiasmatic nucleus (SCN) of the hypothalamus. In these transgenic rats, marked increases in eGFP fluorescence and fusion gene expression were observed in the magnocellular division of the PVN and the SON, but not the SCN, after osmotic challenges, such as dehydration and salt loading, and both acute and chronic nociceptive stimuli. In the parvocellular division of the PVN, eGFP expression was increased after acute and chronic pain, bilateral adrenalectomy, endotoxin shock and restraint stress. In the extra-hypothalamic areas of the brain, eGFP expression was induced in the locus coeruleus after the intracerebroventricular administration of colchicine. Next, we generated another transgenic rat that expresses a fusion gene comprised of c-fos promoter-enhancer sequences driving the expression of monomeric red fluorescent protein 1 (mRFP1). In these transgenic rats, abundant nuclear fluorescence of mRFP1 was observed in the PVN, the SON and other osmosensitive areas after acute osmotic stimulation. Finally, we generated a double transgenic rat that expresses both the AVP-eGFP and c-fos-mRFP1 fusion genes. In this double transgenic rat, we have observed nuclear mRFP1 fluorescence in eGFP-positive neurons after acute osmotic stimulation. These unique transgenic rats provide an exciting new tool to examine neuroendocrine responses to physiological and stressful stimuli in both in vivo and in vitro preparations. PMID:21185297

Ueta, Yoichi; Dayanithi, Govindin; Fujihara, Hiroaki

2011-02-01

131

[TSA improve transgenic porcine cloned embryo development and transgene expression].  

Science.gov (United States)

Uncompleted epigenetic reprogramming is attributed to the low efficiency of producing transgenic cloned animals. Histone modification associated with epigenetics can directly influence the embryo development and transgene expression. Trichostatin A (TSA), as an inhibitor of histone deacetylase, can change the status of histone acetylation, improve somatic cell reprogramming, and enhance cloning efficiency. TSA prevents the chromatin structure from being condensed, so that transcription factor could binds to DNA sequence easily and enhance transgene expression. Our study established the optimal TSA treatment on porcine donor cells and cloned embryos, 250 nmol/L, 24 h and 40 nmol/L, 24 h, respectively. Furthermore, we found that both the cloned embryo and the donor cell treated by TSA resulted in the highest development efficiency. Meanwhile, TSA can improve transgene expression in donor cell and cloned embryo. In summary, TSA can significantly improve porcine reconstructed embryo development and transgene expression. PMID:22049689

Kong, Qing-Ran; Zhu, Jiang; Huang, Bo; Huan, Yan-Jun; Wang, Feng; Shi, Yong-Qian; Liu, Zhong-Feng; Wu, Mei-Ling; Liu, Zhong-Hua

2011-07-01

132

COMPARISON TRANSGENIC AND NON-TRANSGENIC MILK QUALITY  

Directory of Open Access Journals (Sweden)

Full Text Available Transgenic founder rabbits carrying a gene construct consisting of a 2.5 kb murine whey acidic protein promoter (mWAP, 7.2 kb of the human clotting factor VIII (hFVIII cDNA and 4.6 kb of 3’ flanking sequences of mWAP gene were crossed for five generations. Transgenic females showed high level of recombinant hFVIII (rhFVIII mRNA expression in biopsed mammary gland tissues. The presence of the mWAP-hFVIII transgene in rabbit genome and secretion of rhFVIII into milk of transgenic females (F1, F2, F3, F4 and F5 generation did not have any adverse phenotypic effect on milk quality.

Peter Chrenek

2012-02-01

133

NMR profiling of transgenic peas.  

Science.gov (United States)

A high throughput proton nuclear magnetic resonance spectroscopy method for the metabolite fingerprinting of plants was applied to genetically modified peas (Pisum sativum) to determine whether biochemical changes, so called 'unintended effects', beyond those intended by incorporation of a transgene, were detectable. Multivariate analysis of 1H NMR (nuclear magnetic resonance) spectra obtained from uniformly grown glasshouse plants revealed differences between the transgenic and control group that exceeded the natural variation of the plants. When a larger data set of six related transgenic lines was analysed, including a null segregant in addition to the wild-type control, multivariate analysis showed that the distribution of metabolites in the transgenics was different from that of the null segregant. However, the profile obtained from the wild-type material was diverse in comparison with both the transgenics and the null segregant, suggesting that the primary cause of the observed differences was that the transformation process selects for a subset of individuals able to undergo the transformation and selection procedures, and that their descendants have a restricted variation in metabolite profile, rather than that the presence of the transgene itself generates these differences. PMID:17166140

Charlton, Adrian; Allnutt, Theo; Holmes, Stephen; Chisholm, James; Bean, Samantha; Ellis, Noel; Mullineaux, Phil; Oehlschlager, Sarah

2004-01-01

134

ALIMENTOS TRANSGÉNICOS TRANSGENIC FOODS  

Directory of Open Access Journals (Sweden)

Full Text Available Gracias al gran avance de la tecnología, la ingeniería genética y la biología molecular, se han desarrollado los productos transgénicos. En sus inicios, los productos modificados genéticamente tenían como objeto obtener ventajas en las áreas de la agricultura y ganadería. Posteriormente esta técnica se comenzó a aplicar en el ámbito de la producción de alimentos para el consumo humano. Se ha generado mucha controversia en relación a su utilización. Esta revisión tiene por objeto revisar la información científica disponible en relación a las aplicaciones, ventajas y potenciales riesgos para la salud humana y el medio ambiente asociados al consumo de los alimentos transgénicosDue to the advancements in technology, genetic engineering and molecular biology, have develop transgenic foods. Initially, genetically modified plants were produced to confer advantages in agriculture and animal husbandry. Later this technique was applied to the production of food for human consumption, generating a great deal of controversy. This review discusses the available scientific evidence in relation to the advantages and potential risks of genetically modified foods

María Soledad Reyes S.

2003-04-01

135

ALIMENTOS TRANSGÉNICOS / TRANSGENIC FOODS  

Scientific Electronic Library Online (English)

Full Text Available SciELO Chile | Language: Spanish Abstract in spanish Gracias al gran avance de la tecnología, la ingeniería genética y la biología molecular, se han desarrollado los productos transgénicos. En sus inicios, los productos modificados genéticamente tenían como objeto obtener ventajas en las áreas de la agricultura y ganadería. Posteriormente esta técnica [...] se comenzó a aplicar en el ámbito de la producción de alimentos para el consumo humano. Se ha generado mucha controversia en relación a su utilización. Esta revisión tiene por objeto revisar la información científica disponible en relación a las aplicaciones, ventajas y potenciales riesgos para la salud humana y el medio ambiente asociados al consumo de los alimentos transgénicos Abstract in english Due to the advancements in technology, genetic engineering and molecular biology, have develop transgenic foods. Initially, genetically modified plants were produced to confer advantages in agriculture and animal husbandry. Later this technique was applied to the production of food for human consump [...] tion, generating a great deal of controversy. This review discusses the available scientific evidence in relation to the advantages and potential risks of genetically modified foods

María Soledad, Reyes S.; Jaime, Rozowski N.

136

Transgenic RNA Interference in Mice  

Science.gov (United States)

The discovery that small interfering RNA duplexes (siRNA) can silence gene expression in mammalian cells has revolutionized biomedical research. The most successful application of the discovery has been to study gene function in cultured human or mouse cells. However, the knockdown effect of siRNA is only transient. To achieve a more sustained gene-silencing effect, shRNA (small hairpin RNA) expressed from a vector is preferred. An additional benefit of shRNA is that RNA interference (RNAi) can now be applied in vivo through delivering shRNA-expressing vectors by transgenic technology. Transgenic RNAi not only allows the study of biological processes not present in cultured cells but also offers chronic therapeutic potentials. In this review, we will summarize the developments in the generation of transgenic RNAi mice.

2007-06-01

137

Transgenic agriculture and environmental indicators  

Directory of Open Access Journals (Sweden)

Full Text Available Despite the rapid diffusion of transgenic crops, there are still few environmental impact studies capable of supplying a conclusive scientific response in regard to its technical and economic advantages and disadvantages. Prospective scenarios were elaborated to assist environmental impact assessment, using techniques derived from SWOT (Strength, Weakness, Opportunity, Threat analysis and the DPSIR (Driving Force – human activity, Pressure, State, Impact, Response model, to evaluate the environmental indicators and the relationship between them. Control and management actions were identified, searching the integration of aspects related to the biotechnology applied to transgenic processes, biodiversity, biosafety and intellectual property. It was demonstrated that the DPSIR model is, in fact, an instrument for integrated environmental assessment and the application of the proposed methodology resulted in favorable indicators to the adoption of transgenic agriculture. The elaborated scenarios are useful to develop an Environmental Management System (EMS to agriculture.

Denize Dias de Carvalho

2006-12-01

138

Wading pools, fading memories—place navigation in transgenic mouse models of Alzheimer's disease  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The Morris swim navigation task (“water maze”) has been a primary research tool to assess hippocampal-dependent spatial learning and memory in rodents for three decades. Originally developed for rats, its application to mouse studies has been a tedious process, but nowadays there are more studies performed with the Morris swim task in mice than in rats. The task has proved to be particularly useful in demonstrating age-related memory impairment in transgenic mouse models of Alzheimer's di...

Tanila, Heikki

2012-01-01

139

Complete rescue of obesity, diabetes, and infertility in db/db mice by neuron-specific LEPR-B transgenes  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We have generated mice that carry a neuron-specific leptin receptor (LEPR) transgene whose expression is driven by the rat synapsin I promoter synapsin–LEPR B (SYN-LEPR-B). We have also generated mice that are compound hemizygotes for the transgenes SYN-LEPR-B and neuron-specific enolase–LEPR B (NSE-LEPR-B). We observed a degree of correction in db/db mice that are hemizygous (Syn db/db) and homozygous (Syn/Syn db/db) for the SYN-LEPR-B transgene similar to that previously reported for th...

Luca, Carl; Kowalski, Timothy J.; Zhang, Yiying; Elmquist, Joel K.; Lee, Charlotte; Kilimann, Manfred W.; Ludwig, Thomas; Liu, Shun-mei; Chua, Streamson C.

2005-01-01

140

Temporal Expression of Mutant LRRK2 in Adult Rats Impairs Dopamine Reuptake  

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Full Text Available Parkinson's disease (PD results from progressive degeneration of dopaminergic neurons. Most PD cases are sporadic, but some have pathogenic mutation in the individual genes. Mutation of the leucine-rich repeat kinase-2 (LRRK2 gene is associated with familial and sporadic PD, as exemplified by G2019S substitution. While constitutive expression of mutant LRRK2 in transgenic mice fails to induce neuron death, transient expression of the disease gene by viral delivery causes a substantial loss of dopaminergic neurons in mice. To further assess LRRK2 pathogenesis, we created inducible transgenic rats expressing human LRRK2 with G2019S substitution. Temporal overexpression of LRRK2G2019S in adult rats impaired dopamine reuptake by dopamine transporter (DAT and thus enhanced locomotor activity, the phenotypes that were not observed in transgenic rats constitutively expressing the gene throughout life time. Reduced DAT binding activity is an early sign of dopaminergic dysfunction in asymptomatic subjects carrying pathogenic mutation in LRRK2. Our transgenic rats recapitulated the initiation process of dopaminergic dysfunction caused by pathogenic mutation in LRRK2. Inducible transgenic approach uncovered phenotypes that may be obscured by developmental compensation in constitutive transgenic rats. Finding in inducible LRRK2 transgenic rats would guide developing effective strategy in transgenic studies: Inducible expression of transgene may induce greater phenotypes than constitutive gene expression, particularly in rodents with short life time.

Hongxia Zhou, Cao Huang, Jianbin Tong, Weimin C Hong, Yong-Jian Liu, Xu-Gang Xia

2011-01-01

 
 
 
 
141

Neurobehavioral deficits of epidermal growth factor-overexpressing transgenic mice: impact on dopamine metabolism.  

Science.gov (United States)

Epidermal growth factor (EGF) and its family member neuregulin-1 are implicated in the etiology of schizophrenia. Our recent pharmacological studies indicate that EGF injections to neonatal and adult rats both induce neurobehavioral deficits relevant to schizophrenia. We, however, did not evaluate the genetic impact of EGF transgene on neurobehavioral traits. Here we analyzed transgenic mice carrying the transgene of mouse EGF cDNA. As compared to control littermates, heterozygous EGF transgenic mice had an increase in EGF mRNA levels and showed significant decreases in prepulse inhibition and context-dependent fear learning, but there were no changes in locomotor behaviors and sound startle responses. In addition, these transgenic mice exhibited higher behavioral sensitivity to the repeated cocaine injections. There were neurochemical alterations in metabolic enzymes of dopamine (i.e., tyrosine hydroxylase, dopa decarboxylase, catechol-O-methyl transferase) and monoamine contents in various brain regions of the EGF transgenic mice, but there were no apparent neuropathological signs in the brain. The present findings rule out the indirect influence of anti-EGF antibody production on the reported behavioral deficits of EGF-injected mice. These results support the argument that aberrant hyper-signals of EGF have significant impact on mouse behavioral traits and dopamine metabolism. PMID:23669645

Eda, Takeyoshi; Mizuno, Makoto; Araki, Kazuaki; Iwakura, Yuriko; Namba, Hisaaki; Sotoyama, Hidekazu; Kakita, Akiyoshi; Takahashi, Hitoshi; Satoh, Hiroshi; Chan, Siu-Yuen; Nawa, Hiroyuki

2013-06-28

142

Transposon-mediated transgenesis, transgenic rescue, and tissue-specific gene expression in rodents and rabbits.  

Science.gov (United States)

Germline transgenesis is an important procedure for functional investigation of biological pathways, as well as for animal biotechnology. We have established a simple, nonviral protocol in three important biomedical model organisms frequently used in physiological studies. The protocol is based on the hyperactive Sleeping Beauty transposon system, SB100X, which reproducibly promoted generation of transgenic founders at frequencies of 50-64, 14-72, and 15% in mice, rats, and rabbits, respectively. The SB100X-mediated transgene integrations are less prone to genetic mosaicism and gene silencing as compared to either the classical pronuclear injection or to lentivirus-mediated transgenesis. The method was successfully applied to a variety of transgenes and animal models, and can be used to generate founders with single-copy integrations. The transposon vector also allows the generation of transgenic lines with tissue-specific expression patterns specified by promoter elements of choice, exemplified by a rat reporter strain useful for tracking serotonergic neurons. As a proof of principle, we rescued an inborn genetic defect in the fawn-hooded hypertensive rat by SB100X transgenesis. A side-by-side comparison of the SB100X- and piggyBac-based protocols revealed that the two systems are complementary, offering new opportunities in genome manipulation. PMID:23195032

Katter, Katharina; Geurts, Aron M; Hoffmann, Orsolya; Mátés, Lajos; Landa, Vladimir; Hiripi, László; Moreno, Carol; Lazar, Jozef; Bashir, Sanum; Zidek, Vaclav; Popova, Elena; Jerchow, Boris; Becker, Katja; Devaraj, Anantharam; Walter, Ingrid; Grzybowksi, Michael; Corbett, Molly; Filho, Artur Rangel; Hodges, Matthew R; Bader, Michael; Ivics, Zoltán; Jacob, Howard J; Pravenec, Michal; Bosze, Zsuzsanna; Rülicke, Thomas; Izsvák, Zsuzsanna

2013-03-01

143

Transgenic Arabidopsis Gene Expression System  

Science.gov (United States)

The Transgenic Arabidopsis Gene Expression System (TAGES) investigation is one in a pair of investigations that use the Advanced Biological Research System (ABRS) facility. TAGES uses Arabidopsis thaliana, thale cress, with sensor promoter-reporter gene constructs that render the plants as biomonitors (an organism used to determine the quality of the surrounding environment) of their environment using real-time nondestructive Green Fluorescent Protein (GFP) imagery and traditional postflight analyses.

Ferl, Robert; Paul, Anna-Lisa

2009-01-01

144

Transgenic trees and forestry biosafety  

Scientific Electronic Library Online (English)

Full Text Available SciELO Chile | Language: English Abstract in english The benefits from the development of transgenic trees are expected from the improvement of traits as growth and form, wood quality, industrial processes, disease and insect resistance, herbicide tolerance, ecological restoration, rooting ability, etc. One of the first reported field trials with gene [...] tically modified forest trees was established in Belgium in 1988 and the characteristic evaluated was herbicide tolerance in poplars. Since then, there have been more than 200 reported trials, involving at least 15 forest species. The majority of the field trials have been carried out in the USA (64%). More than 50% of the field trials are done with Populus species and the main target traits are herbicide tolerance (31%), followed by marker genes (23%) and insect resistance (14%). Until today, there is only one report on commercial-scale production of transgenic forest trees which is Populus nigra with the Bt gene release in China in 2002 and established on commercial plantations in 2003. Operational application of GMO's in forestry depends on technical, economical, political and public aspects, but the development of adequate regulatory frameworks and public acceptance of transgenic trees will define the future of this technology in forestry.

Sofía, Valenzuela; Claudio, Balocchi; Jaime, Rodríguez.

145

Transgenic trees and forestry biosafety  

Scientific Electronic Library Online (English)

Full Text Available SciELO Chile | Language: English Abstract in english The benefits from the development of transgenic trees are expected from the improvement of traits as growth and form, wood quality, industrial processes, disease and insect resistance, herbicide tolerance, ecological restoration, rooting ability, etc. One of the first reported field trials with gene [...] tically modified forest trees was established in Belgium in 1988 and the characteristic evaluated was herbicide tolerance in poplars. Since then, there have been more than 200 reported trials, involving at least 15 forest species. The majority of the field trials have been carried out in the USA (64%). More than 50% of the field trials are done with Populus species and the main target traits are herbicide tolerance (31%), followed by marker genes (23%) and insect resistance (14%). Until today, there is only one report on commercial-scale production of transgenic forest trees which is Populus nigra with the Bt gene release in China in 2002 and established on commercial plantations in 2003. Operational application of GMO's in forestry depends on technical, economical, political and public aspects, but the development of adequate regulatory frameworks and public acceptance of transgenic trees will define the future of this technology in forestry.

Sofía, Valenzuela; Claudio, Balocchi; Jaime, Rodríguez.

2006-06-01

146

Determining gene flow in transgenic cotton.  

Science.gov (United States)

Gene flow is one of the major concerns associated with the release of transgenic plants into the environment. Unrestricted gene flow can results in super weeds, reduction in species fitness and genetic diversity, and contamination of traditional plants and foods. Thus, it is important and also necessary to evaluate the extent of gene flow in the field for transgenic plants already released or being considered for a release. Transgenic cotton is among the first transgenic crops for commercialization, which are widely cultivated around the world. In this chapter, we use transgenic insect resistant cotton and herbicide-tolerant cotton as two examples to present a field practice method for determining transgene flow in cotton. The procedure includes three major sections: (1) field design, (2) seed collection, and (3) field and lab bioassay. PMID:23143499

Pan, Xiaoping

2013-01-01

147

A Built-In Strategy for Containment of Transgenic Plants: Creation of Selectively Terminable Transgenic Rice  

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Plant transgenic technology has been widely utilized for engineering crops for trait improvements and for production of high value proteins such as pharmaceuticals. However, the unintended spreading of commercial transgenic crops by pollination and seed dispersal is a major concern for environmental and food safety. Simple and reliable containment strategies for transgenes are highly desirable. Here we report a novel method for creating selectively terminable transgenic rice. In this method, ...

Lin, Chaoyang; Fang, Jun; Xu, Xiaoli; Zhao, Te; Tu, Juming; Ye, Gongyin; Shen, Zhicheng

2008-01-01

148

Expression of genomic and cDNA transgenes after co-integration in transgenic mice.  

Science.gov (United States)

In general, genomic transgenes are expressed efficiently in mice, while their cDNA-based transgenes are frequently silent. Clark et al. (1992) have shown that silent cDNA transgenes under the control of the sheep beta-lactoglobulin promoter can be activated after co-injecting them with a genomic sheep beta-lactoglobulin transgene. We have tested the general utility of this concept using mouse whey acidic protein (WAP) transgenes. Here we show that WAP cDNA transgenes are virtually silent in transgenic mice. In contrast, WAP transgenes containing all introns are expressed in approximately 50% of the lines at levels ranging from 1% to more than 100% of the endogenous RNA (McKnight et al., 1992). When a WAP-genomic transgene was co-injected with a WAP-cDNA, basal activation of the cDNA was possible. However, the activity of the WAP-cDNA transgene did not exceed 1% of the WAP-genomic transgene. This suggests that a permissive integration site capable of supporting basal level transcription can be established, but further events are required for full activation of the transgene. PMID:7881461

McKnight, R A; Wall, R J; Hennighausen, L

1995-01-01

149

A built-in strategy for containment of transgenic plants: creation of selectively terminable transgenic rice.  

Science.gov (United States)

Plant transgenic technology has been widely utilized for engineering crops for trait improvements and for production of high value proteins such as pharmaceuticals. However, the unintended spreading of commercial transgenic crops by pollination and seed dispersal is a major concern for environmental and food safety. Simple and reliable containment strategies for transgenes are highly desirable. Here we report a novel method for creating selectively terminable transgenic rice. In this method, the gene(s) of interest is tagged with a RNA interference cassette, which specifically suppresses the expression of the bentazon detoxification enzyme CYP81A6 and thus renders transgenic rice to be sensitive to bentazon, a herbicide used for rice weed control. We generated transgenic rice plants by this method using a new glyphosate resistant 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene from Pesudomonas putida as the gene of interest, and demonstrated that these transgenic rice plants were highly sensitive to bentazon but tolerant to glyphosate, which is exactly the opposite of conventional rice. Field trial of these transgenic rice plants further confirmed that they can be selectively killed at 100% by one spray of bentazon at a regular dose used for conventional rice weed control. Furthermore, we found that the terminable transgenic rice created in this study shows no difference in growth, development and yield compared to its non-transgenic control. Therefore, this method of creating transgenic rice constitutes a novel strategy of transgene containment, which appears simple, reliable and inexpensive for implementation. PMID:18350155

Lin, Chaoyang; Fang, Jun; Xu, Xiaoli; Zhao, Te; Cheng, Jiaan; Tu, Juming; Ye, Gongyin; Shen, Zhicheng

2008-01-01

150

[The use of transgenic animals in biomedical research in Germany. Part 1: Status Report 2001-2003].  

Science.gov (United States)

While the German Federal Government has set itself the goal to make an active contribution to reducing animal experiments, the use of transgenic animals in biomedical research continuously increases every year. It is against this background that the study at hand aimed at providing an overview over the goals and the contents of research projects performed in Germany, in the course of which transgenic animals were produced or used in experimental procedures. Specifically, it was envisaged to spell out those specific areas of research, for which transgenic animals mainly were being used. Subsequently it was evaluated whether the research goals revealed might also be pursued with non animal test methods. In a literature survey, a total of 577 scientific publications relevant for the purposes of the study were collected. This material enables conclusions on those scientific areas, in which transgenic animals are used, applying to fundamental research, but not on their use in routine procedures in applied research or for the maintenance of transgenic breeds, since such purposes do not tend to be the subject of publications in scientific journals. According to the topics covered by the publications, main areas of biomedical research with transgenic animals can be found in the fields of neurobiology, immunology, cardiology, embryology and oncology. However their use can be discerned in all other areas of fundamental biomedical research as well. In accordance with the official German laboratory animal statistics, the vast majority of transgenic animals used were mice, followed by rats and pigs. Additionally, singular research projects with fish, rabbits and chicken were recorded. (In the official German laboratory animals statistics, very small numbers of transgenic hamsters, sheep and amphibians were also recorded in the past years.) A high percentage of the rats were used in cardiovascular research, whereas transgenic pigs as a rule were produced and bred as organ donors in xenotransplantation research. The majority of research projects either dealt with the experimental use of already established transgenic animal lines, or they described that transgenic animals specifically were produced for the purpose of the respective research project. Mostly, transgenesis was initiated by inserting the foreign gene into the germ cell genome. In some research projects, it was reported that the transgenic material was inserted into normally bred animals some time after parturition. PMID:16344907

Sauer, Ursula G; Kolar, Roman; Rusche, Brigitte

2005-01-01

151

Production of transgenic livestock: promise fulfilled.  

Science.gov (United States)

The introduction of specific genes into the genome of farm animals and its stable incorporation into the germ line has been a major technological advance in agriculture. Transgenic technology provides a method to rapidly introduce "new" genes into cattle, swine, sheep, and goats without crossbreeding. It is a more extreme methodology, but in essence, not really different from crossbreeding or genetic selection in its result. Methods to produce transgenic animals have been available for more than 20 yr, yet recently lines of transgenic livestock have been developed that have the potential to improve animal agriculture and benefit producers and/or consumers. There are a number of methods that can be used to produce transgenic animals. However, the primary method to date has been the microinjection of genes into the pronuclei of zygotes. This method is one of an array of rapidly developing transgenic methodologies. Another method that has enjoyed recent success is that of nuclear transfer or "cloning." The use of this technique to produce transgenic livestock will profoundly affect the use of transgenic technology in livestock production. Cell-based, nuclear transfer or cloning strategies have several distinct advantages for use in the production of transgenic livestock that cannot be attained using pronuclear injection of DNA. Practical applications of transgenesis in livestock production include enhanced prolificacy and reproductive performance, increased feed utilization and growth rate, improved carcass composition, improved milk production and/or composition, and increased disease resistance. One practical application of transgenics in swine production is to improve milk production and/or composition. To address the problem of low milk production, transgenic swine over-expressing the milk protein bovine alpha-lactalbumin were developed and characterized. The outcomes assessed were milk composition, milk yield, and piglet growth. Our results indicate that transgenic overexpression of milk proteins may provide a means to improve swine lactation performance. PMID:15000404

Wheeler, M B

2003-01-01

152

Plant Transformation: Needs and Futurity of the Transgenes  

Directory of Open Access Journals (Sweden)

Full Text Available To produce transgenic plants which have various applications in agricultural and non-agricultural fields, a marker gene is necessary to recover a viable transgenic plant. To express or transcribe of transgenes, utilization of promoters is also unavoidable. Analysis of transgenes includes copy number, insertion site, integration stability, expression and it`s pattern and variability is immensely important in order to develop a successful transgenic event. This review presents the necessities for better recovery of transgenic plants, transcription or expression of transgenes, as well as methods to analyze transgenes.

Behrooz Darbani

2008-01-01

153

Combining M-FISH and Quantum Dot technology for fast chromosomal assignment of transgenic insertions  

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Full Text Available Abstract Background Physical mapping of transgenic insertions by Fluorescence in situ Hybridization (FISH is a reliable and cost-effective technique. Chromosomal assignment is commonly achieved either by concurrent G-banding or by a multi-color FISH approach consisting of iteratively co-hybridizing the transgenic sequence of interest with one or more chromosome-specific probes at a time, until the location of the transgenic insertion is identified. Results Here we report a technical development for fast chromosomal assignment of transgenic insertions at the single cell level in mouse and rat models. This comprises a simplified 'single denaturation mixed hybridization' procedure that combines multi-color karyotyping by Multiplex FISH (M-FISH, for simultaneous and unambiguous identification of all chromosomes at once, and the use of a Quantum Dot (QD conjugate for the transgene detection. Conclusions Although the exploitation of the unique optical properties of QD nanocrystals, such as photo-stability and brightness, to improve FISH performance generally has been previously investigated, to our knowledge this is the first report of a purpose-designed molecular cytogenetic protocol in which the combined use of QDs and standard organic fluorophores is specifically tailored to assist gene transfer technology.

Yusuf Mohammed

2011-12-01

154

Transposon-mediated chromosomal integration of transgenes in the parasitic nematode Strongyloides ratti and establishment of stable transgenic lines.  

Science.gov (United States)

Genetic transformation is a potential tool for analyzing gene function and thereby identifying new drug and vaccine targets in parasitic nematodes, which adversely affect more than one billion people. We have previously developed a robust system for transgenesis in Strongyloides spp. using gonadal microinjection for gene transfer. In this system, transgenes are expressed in promoter-regulated fashion in the F1 but are silenced in subsequent generations, presumably because of their location in repetitive episomal arrays. To counteract this silencing, we explored transposon-mediated chromosomal integration of transgenes in S. ratti. To this end, we constructed a donor vector encoding green fluorescent protein (GFP) under the control of the Ss-act-2 promoter with flanking inverted tandem repeats specific for the piggyBac transposon. In three experiments, free-living Strongyloides ratti females were transformed with this donor vector and a helper plasmid encoding the piggyBac transposase. A mean of 7.9% of F1 larvae were GFP-positive. We inoculated rats with GFP-positive F1 infective larvae, and 0.5% of 6014 F2 individuals resulting from this host passage were GFP-positive. We cultured GFP-positive F2 individuals to produce GFP-positive F3 L3i for additional rounds of host and culture passage. Mean GFP expression frequencies in subsequent generations were 15.6% in the F3, 99.0% in the F4, 82.4% in the F5 and 98.7% in the F6. The resulting transgenic lines now have virtually uniform GFP expression among all progeny after at least 10 generations of passage. Chromosomal integration of the reporter transgenes was confirmed by Southern blotting and splinkerette PCR, which revealed the transgene flanked by S. ratti genomic sequences corresponding to five discrete integration sites. BLAST searches of flanking sequences against the S. ratti genome revealed integrations in five contigs. This result provides the basis for two powerful functional genomic tools in S. ratti: heritable transgenesis and insertional mutagenesis. PMID:22912584

Shao, Hongguang; Li, Xinshe; Nolan, Thomas J; Massey, Holman C; Pearce, Edward J; Lok, James B

2012-01-01

155

Generation of NSE-MerCreMer transgenic mice with tamoxifen inducible Cre activity in neurons  

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To establish a genetic tool for conditional deletion or expression of gene in neurons in a temporally controlled manner, we generated a transgenic mouse (NSE-MerCreMer), which expressed a tamoxifen inducible type of Cre recombinase specifically in neurons. The tamoxifen inducible Cre recombinase (MerCreMer) is a fusion protein containing Cre recombinase with two modified estrogen receptor ligand binding domains at both ends, and is driven by the neural-specific rat neural specific enolase (NS...

Kam, Mandy Ka Man; Lee, King Yiu; Tam, Paul Kwong Hang; Lui, Vincent Chi Hang

2012-01-01

156

Rapid characterization of transgenic and non-transgenic soybean oils by chemometric methods using NIR spectroscopy  

Science.gov (United States)

Near infrared (NIR) spectroscopy and multivariate classification were applied to discriminate soybean oil samples into non-transgenic and transgenic. Principal Component Analysis (PCA) was applied to extract relevant features from the spectral data and to remove the anomalous samples. The best results were obtained when with Support Vectors Machine-Discriminant Analysis (SVM-DA) and Partial Least Squares-Discriminant Analysis (PLS-DA) after mean centering plus multiplicative scatter correction. For SVM-DA the percentage of successful classification was 100% for the training group and 100% and 90% in validation group for non transgenic and transgenic soybean oil samples respectively. For PLS-DA the percentage of successful classification was 95% and 100% in training group for non transgenic and transgenic soybean oil samples respectively and 100% and 80% in validation group for non transgenic and transgenic respectively. The results demonstrate that NIR spectroscopy can provide a rapid, nondestructive and reliable method to distinguish non-transgenic and transgenic soybean oils.

Luna, Aderval S.; da Silva, Arnaldo P.; Pinho, Jéssica S. A.; Ferré, Joan; Boqué, Ricard

157

Generation of transgenic hydra by embryo microinjection.  

Science.gov (United States)

As a member of the phylum Cnidaria, the sister group to all bilaterians, Hydra can shed light on fundamental biological processes shared among multicellular animals. Hydra is used as a model for the study of regeneration, pattern formation, and stem cells. However, research efforts have been hampered by lack of a reliable method for gene perturbations to study molecular function. The development of transgenic methods has revitalized the study of Hydra biology(1). Transgenic Hydra allow for the tracking of live cells, sorting to yield pure cell populations for biochemical analysis, manipulation of gene function by knockdown and over-expression, and analysis of promoter function. Plasmid DNA injected into early stage embryos randomly integrates into the genome early in development. This results in hatchlings that express transgenes in patches of tissue in one or more of the three lineages (ectodermal epithelial, endodermal epithelial, or interstitial). The success rate of obtaining a hatchling with transgenic tissue is between 10% and 20%. Asexual propagation of the transgenic hatchling is used to establish a uniformly transgenic line in a particular lineage. Generating transgenic Hydra is surprisingly simple and robust, and here we describe a protocol that can be easily implemented at low cost. PMID:25285460

Juliano, Celina E; Lin, Haifan; Steele, Robert E

2014-01-01

158

Rescuing transgene expression by co-integration.  

Science.gov (United States)

To test whether foreign gene expression can be improved in transgenic mice by manipulating the site of integration, we co-integrated the efficiently expressed sheep beta-lactoglobulin gene with two poorly expressed beta-lactoglobulin-derived hybrid genes encoding human proteins. In each case, we observed a significant improvement in the frequency and level of expression of the hybrid gene. "Rescuing" transgene expression by co-integration may provide a general solution for improving the efficiency of heterologous gene expression in transgenic animals. PMID:1281647

Clark, A J; Cowper, A; Wallace, R; Wright, G; Simons, J P

1992-11-01

159

Effects of a GTP-insensitive mutation of glutamate dehydrogenase on insulin secretion in transgenic mice.  

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Glutamate dehydrogenase (GDH) plays an important role in insulin secretion as evidenced in children by gain of function mutations of this enzyme that cause a hyperinsulinism-hyperammonemia syndrome (GDH-HI) and sensitize beta-cells to leucine stimulation. GDH transgenic mice were generated to express the human GDH-HI H454Y mutation and human wild-type GDH in islets driven by the rat insulin promoter. H454Y transgene expression was confirmed by increased GDH enzyme activity in islets and decreased sensitivity to GTP inhibition. The H454Y GDH transgenic mice had hypoglycemia with normal growth rates. H454Y GDH transgenic islets were more sensitive to leucine- and glutamine-stimulated insulin secretion but had decreased response to glucose stimulation. The fluxes via GDH and glutaminase were measured by tracing 15N flux from [2-15N]glutamine. The H454Y transgene in islets had higher insulin secretion in response to glutamine alone and had 2-fold greater GDH flux. High glucose inhibited both glutaminase and GDH flux, and leucine could not override this inhibition. 15NH4Cl tracing studies showed 15N was not incorporated into glutamate in either H454Y transgenic or normal islets. In conclusion, we generated a GDH-HI disease mouse model that has a hypoglycemia phenotype and confirmed that the mutation of H454Y is disease causing. Stimulation of insulin release by the H454Y GDH mutation or by leucine activation is associated with increased oxidative deamination of glutamate via GDH. This study suggests that GDH functions predominantly in the direction of glutamate oxidation rather than glutamate synthesis in mouse islets and that this flux is tightly controlled by glucose. PMID:16574664

Li, Changhong; Matter, Andrea; Kelly, Andrea; Petty, Thomas J; Najafi, Habiba; MacMullen, Courtney; Daikhin, Yevgeny; Nissim, Ilana; Lazarow, Adam; Kwagh, Jae; Collins, Heather W; Hsu, Betty Y L; Nissim, Itzhak; Yudkoff, Marc; Matschinsky, Franz M; Stanley, Charles A

2006-06-01

160

LKB1 deletion with the RIP2.Cre transgene modifies pancreatic ?-cell morphology and enhances insulin secretion in vivo  

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The tumor suppressor liver kinase B1 (LKB1), also called STK11, is a protein kinase mutated in Peutz-Jeghers syndrome. LKB1 phosphorylates AMP-activated protein kinase (AMPK) and several related protein kinases. Whereas deletion of both catalytic isoforms of AMPK from the pancreatic ?-cell and hypothalamic neurons using the rat insulin promoter (RIP2).Cre transgene (?AMPKdKO) diminishes insulin secretion in vivo, deletion of LKB1 in the ?-cell with an inducible Pdx-1.CreER transgene enhanc...

Sun, Gao; Tarasov, Andrei I.; Mcginty, James A.; French, Paul M.; Mcdonald, Angela; Leclerc, Isabelle; Rutter, Guy A.

2010-01-01

 
 
 
 
161

Transgenic Wheat, Barley and Oats: Future Prospects  

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Following the success of transgenic maize and rice, methods have now been developed for the efficient introduction of genes into wheat, barley and oats. This review summarizes the present position in relation to these three species, and also uses information from field trial databases and the patent literature to assess the future trends in the exploitation of transgenic material. This analysis includes agronomic traits and also discusses opportunities in expanding areas such as biofuels and biopharming.

Dunwell, Jim M.

162

Reversing insect adaptation to transgenic insecticidal plants.  

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The refuge-high-dose strategy for delaying insect adaptation to transgenic plants produces non-transgenic plants that enable survival of susceptible individuals. Previous theoretical work has suggested three requirements for success of the refuge-high-dose strategy: a low initial frequency of the resistance allele, extensive mating between resistant and susceptible adults and recessive inheritance of resistance. In order to understand an observed decrease in resistance frequency and improve t...

Carrie?re, Y.; Tabashnik, B. E.

2001-01-01

163

Transgenic animals and their application in medicine  

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Full Text Available Transgenic animals are animals that are genetically altered to have traits that mimic symptoms of specific human pathologies. They provide genetic models of various human diseases which are important in understanding disease and developing new targets. In early 1980 Gordon and co-workers described the first gene addition experiment using the microinjection technology and since then the impact of transgenic technology on basic research has been significant. Within 20 years of its inception, ATryn the first drug approved by USFDA from transgenic animals was developed and it has opened door to drugs from transgenic animals. In addition, they are looked upon as potential future donors for xenotransplantation. With increasing knowledge about the genetics and improvements in the transgenetic technology numerous useful applications like biologically safe new-generation drugs based on human regulatory proteins are being developed.Various aspects of concern in the coming years are the regulatory guidelines, ethical issues and patents related to the use of transgenic animals. This modern medicine is on the threshold of a pharmacological revolution. Use of transgenic animals will provide solutions for drug research, xenotransplantation, clinical trials and will prove to be a new insight in drug development.

Bagle TR, Kunkulol RR, Baig MS, More SY

2013-01-01

164

TRANSGENIC FISH MODEL IN ENVIRONMENTAL TOXICOLOGY  

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Full Text Available A number of experiments and the use of drugs have been performed in fish. The fish may be used as model organism in various biological experiments, including environmental toxicology. Aquatic animals are being engineered to increase aquaculture production, for medical and industrial research, and for ornamental reasons. Fish have been found to play an important role in assessing potential risks associated with exposure to toxic substances in aquatic environment. Hence, it has been thought that the development of transgenic fish can enhance the use of fish in environmental toxicology. India has developed experimental transgenics of rohu fish, zebra fish, cat fish and singhi fish. Genes, promoters and vectors of indigenous origin are now available for only two species namely rohu and singhi for engineering growth. Development of fish model carrying identical transgenes to those found in rodents is beneficial and has shown that several aspects of in vivo mutagenesis are similar between the two classes of vertebrates. Fish shows the frequencies of spontaneous mutations similar to rodents and respond to mutagen exposure consistent with known mutagenic mechanisms. The feasibility of in vivo mutation analysis using transgenic fish has been demonstrated and the potential value of transgenic fish as a comparative animal model has been illustrated. Therefore, the transgenic fish can give the significant contribution to study the environmental toxicity in animals as a whole.

Madhuri Sharma

2012-05-01

165

Transgenic technologies to induce sterility  

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Full Text Available Abstract The last few years have witnessed a considerable expansion in the number of tools available to perform molecular and genetic studies on the genome of Anopheles mosquitoes, the vectors of human malaria. As a consequence, knowledge of aspects of the biology of mosquitoes, such as immunity, reproduction and behaviour, that are relevant to their ability to transmit disease is rapidly increasing, and could be translated into concrete benefits for malaria control strategies. Amongst the most important scientific advances, the development of transgenic technologies for Anopheles mosquitoes provides a crucial opportunity to improve current vector control measures or design novel ones. In particular, the use of genetic modification of the mosquito genome could provide for a more effective deployment of the sterile insect technique (SIT against vector populations in the field. Currently, SIT relies on the release of radiation sterilized males, which compete with wild males for mating with wild females. The induction of sterility in males through the genetic manipulation of the mosquito genome, already achieved in a number of other insect species, could eliminate the need for radiation and increase the efficiency of SIT-based strategies. This paper provides an overview of the mechanisms already in use for inducing sterility by transgenesis in Drosophila and other insects, and speculates on possible ways to apply similar approaches to Anopheles mosquitoes.

Wimmer Ernst A

2009-11-01

166

Transgenic technologies to induce sterility.  

Science.gov (United States)

The last few years have witnessed a considerable expansion in the number of tools available to perform molecular and genetic studies on the genome of Anopheles mosquitoes, the vectors of human malaria. As a consequence, knowledge of aspects of the biology of mosquitoes, such as immunity, reproduction and behaviour, that are relevant to their ability to transmit disease is rapidly increasing, and could be translated into concrete benefits for malaria control strategies. Amongst the most important scientific advances, the development of transgenic technologies for Anopheles mosquitoes provides a crucial opportunity to improve current vector control measures or design novel ones. In particular, the use of genetic modification of the mosquito genome could provide for a more effective deployment of the sterile insect technique (SIT) against vector populations in the field. Currently, SIT relies on the release of radiation sterilized males, which compete with wild males for mating with wild females. The induction of sterility in males through the genetic manipulation of the mosquito genome, already achieved in a number of other insect species, could eliminate the need for radiation and increase the efficiency of SIT-based strategies. This paper provides an overview of the mechanisms already in use for inducing sterility by transgenesis in Drosophila and other insects, and speculates on possible ways to apply similar approaches to Anopheles mosquitoes. PMID:19917077

Catteruccia, Flaminia; Crisanti, Andrea; Wimmer, Ernst A

2009-01-01

167

Selenoprotein-Transgenic Chlamydomonas reinhardtii  

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Full Text Available Selenium (Se deficiency is associated with the occurrence of many diseases. However, excessive Se supplementation, especially with inorganic Se, can result in toxicity. Selenoproteins are the major forms of Se in vivo to exert its biological function. Expression of those selenoproteins, especially with the application of a newly developed system, is thus very important for studying the mechanism of Se in nutrition. The use of Chlamydomonas reinhardtii (C. reinhardtii as a biological vector to express an heterogeneous protein is still at the initial stages of development. In order to investigate the possibility of using this system to express selenoproteins, human 15-KDa selenoprotein (Sep15, a small but widely distributed selenoprotein in mammals, was chosen for the expression platform test. Apart from the wild-type human Sep15 gene fragment, two Sep15 recombinants were constructed containing Sep15 open reading frame (ORF and the selenocysteine insertion sequence (SECIS element from either human Sep15 or C. reinhardtii selenoprotein W1, a highly expressed selenoprotein in this alga. Those Sep15-containing plasmids were transformed into C. reinhardtii CC-849 cells. Results showed that Sep15 fragments were successfully inserted into the nuclear genome and expressed Sep15 protein in the cells. The transgenic and wild-type algae demonstrated similar growth curves in low Se culture medium. To our knowledge, this is the first report on expressing human selenoprotein in green alga.

Jiazuan Ni

2013-02-01

168

Transgenic cotton: from biotransformation methods to agricultural application.  

Science.gov (United States)

Transgenic cotton is among the first transgenic plants commercially adopted around the world. Since it was first introduced into the field in the middle of 1990s, transgenic cotton has been quickly adopted by cotton farmers in many developed and developing countries. Transgenic cotton has offered many important environmental, social, and economic benefits, including reduced usage of pesticides, indirect increase of yield, minimizing environmental pollution, and reducing labor and cost. Agrobacterium-mediated genetic transformation method is the major method for obtaining transgenic cotton. However, pollen tube pathway-mediated method is also used, particularly by scientists in China, to breed commercial transgenic cotton. Although transgenic cotton plants with disease-resistance, abiotic stress tolerance, and improved fiber quality have been developed in the past decades, insect-resistant and herbicide-tolerant cotton are the two dominant transgenic cottons in the transgenic cotton market. PMID:23143479

Zhang, Baohong

2013-01-01

169

BAC manipulations for making BAC transgene arrays.  

Science.gov (United States)

Chromosome tagging using lac or tet operator repeats for in vivo visualization of chromosome dynamics has now become a standard methodology used in a range of organisms. One variation of this approach has been to build transgene arrays creating artificial chromosome blocks to study various aspects of chromatin structure, transcription, replication, or DNA repair. Previously, plasmid transgenes with or without subsequent gene amplification have been used to build these arrays. However, plasmid arrays typically show heterochromatic properties, while gene amplification typically results in chromosome instability of the amplified regions. To avoid these problems, we are now building transgene arrays from large genomic DNA inserts cloned in bacterial artificial chromosomes (BAC). These BAC transgenes show transcriptional levels within several fold of endogenous genes while also exhibiting targeting to specific nuclear compartments similar to the targeting of the endogenous genes. Here we describe Tn5 transposition and BAC recombineering methods used to retrofit BACs for their use in building BAC transgene arrays. This includes insertion of operator repeats and selectable markers into these BACs as well as targeted insertion or deletion of BAC sequences. PMID:23980009

Khanna, Nimish; Bian, Qian; Plutz, Matt; Belmont, Andrew S

2013-01-01

170

Transgenic technology and applications in swine.  

Science.gov (United States)

The introduction of foreign DNA into the genome of livestock and its stable integration into the germ line has been a major technical advance in agriculture. Production of transgenic livestock provides a method to rapidly introduce "new" genes into cattle, swine, sheep and goats without crossbreeding. It is a more extreme methodology, but in essence, not really different from crossbreeding or genetic selection in its result. Several recent developments will profoundly impact the use of transgenic technology in livestock production. These developments are: 1) the ability to isolate and maintain in vitro embryonic stem (ES) cells from preimplantation embryos, embryonic germ (EG) and somatic cells from fetuses; and somatic cells from adults, and 2) the ability to use these embryonic and somatic cells as nuclei donors in nuclear transfer or "cloning" strategies. Cell based (ES, EG, and somatic cells) strategies have several distinct advantages for use in the production of transgenic livestock that cannot be attained using pronuclear injection of DNA. There are many potential applications of transgenic methodology to develop new and improved strains of livestock. Practical applications of transgenesis in livestock production include enhanced prolificacy and reproductive performance, increased feed utilization and growth rate, improved carcass composition, improved milk production and/or composition and increased disease resistance. Development of transgenic farm animals will allow more flexibility in direct genetic manipulation of livestock. PMID:11758888

Wheeler, M B; Walters, E M

2001-11-01

171

Characterization of cardiac phenotype in SERCA2a transgenic rats  

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Cellular Ca2+ regulation disorders play an essential pathogenic role in diastolic and systolic dysfunction typically occurring in hypertrophy and insufficiency of heart muscle. Decreased SERCA2a mediated sarcoplasmic reticulum (SR) Ca2+ transport has been identified as an important factor in myocardial Ca2+ dysregulation. Therefore, suitable interventions have been intensively searched to enhance activity of SR Ca2+ transport. Permanent or inducible SERCA2 overexpression has been developed in...

Weiß, Wolfgang

2010-01-01

172

[Induced phytophthora resistance in transgenic potato tubers].  

Science.gov (United States)

Resistance of transgenic cultivars based on the expression of one or more resistance genes is sooner or later broken by pathogens whose race-producing rates are high. Thus, combining transgenesis with elicitor-induced resistance is a promising approach. The elicitor-induced resistance is based on the expression of multiple resistance genes, which can prevent the adaptation of pathogens to transgenic races, maintain the stability of cultivars, and increase their lifespan. In this work, we used transgenic potato cultivars Temp and Superior transformed with Bacillus thuringiensis delta-endotoxin gene and Lukyanovskii transformed with leukocyte alpha-interferon gene. Arachidonic acid (10(-8) M) and soluble chitosan (5 kDa, 100 micrograms/ml) were used as elicitors for tuber treatment. Our data showed that pretreatment with elicitors causes a 15-25% increase in both the systemic prolonged resistance of potato tubers to Phytophthora infestans and their ability to repair mechanical damage. PMID:12391758

Ozeretskovskaia, O L; Vasiukova, N I; Chalenko, G I; Gerasimova, N G; Grishanina, A N; Khromova, L Ia; Iakovleva, G A; Varlamov, V P; Skriabin, K G

2002-01-01

173

Toxins for transgenic resistance to hemipteran pests.  

Science.gov (United States)

The sap sucking insects (Hemiptera), which include aphids, whiteflies, plant bugs and stink bugs, have emerged as major agricultural pests. The Hemiptera cause direct damage by feeding on crops, and in some cases indirect damage by transmission of plant viruses. Current management relies almost exclusively on application of classical chemical insecticides. While the development of transgenic crops expressing toxins derived from the bacterium Bacillus thuringiensis (Bt) has provided effective plant protection against some insect pests, Bt toxins exhibit little toxicity against sap sucking insects. Indeed, the pest status of some Hemiptera on Bt-transgenic plants has increased in the absence of pesticide application. The increased pest status of numerous hemipteran species, combined with increased prevalence of resistance to chemical insecticides, provides impetus for the development of biologically based, alternative management strategies. Here, we provide an overview of approaches toward transgenic resistance to hemipteran pests. PMID:22822455

Chougule, Nanasaheb P; Bonning, Bryony C

2012-06-01

174

Comparative metallomics of transgenic and non-transgenic soybeans using HPLC-ICP-MS  

International Nuclear Information System (INIS)

kDa; 800-420 kDa; 420-120 kDa; 100-23 kDa; 23-7 kDa; 7-2 kDa; 2-0.4 kDa and 0.4-0.2 kDa, respectively) identified in the transgenic and non-transgenic soybeans after 95 min of separation using a flow rate of 0.25 mL.min-1. A wide range of elements could be identified in all the fractions, including: Cu, Zn, Mn, Mg, Ni, Cr, Hg, Fe and Pb. Differences in the detectability of elements in the transgenic and non-transgenic soybeans were found, specially for Hg where the counts were two times higher in the transgenic soybean. Elements were found in the two samples that were not common for both of them, such as Sr identified only in fraction 2 of the non-transgenic soybean and Th in fraction 4 of the transgenic soybean. Financial support from Fundacao de Amparo a Pesquisa do Estado de Sao Paulo - FAPESP and Conselho Nacional de Desenvolvimento Cientifico e Tecnologico - CNPq are highly acknowledged.

175

Generation of transgenic non-human primates with germline transmission.  

Science.gov (United States)

The common marmoset (Callithrix jacchus) is increasingly attractive for use as a non-human primate animal model in biomedical research. It has a relatively high reproduction rate for a primate, making it potentially suitable for transgenic modification. Although several attempts have been made to produce non-human transgenic primates, transgene expression in the somatic tissues of live infants has not been demonstrated by objective analyses such as polymerase chain reaction with reverse transcription or western blots. Here we show that the injection of a self-inactivating lentiviral vector in sucrose solution into marmoset embryos results in transgenic common marmosets that expressed the transgene in several organs. Notably, we achieved germline transmission of the transgene, and the transgenic offspring developed normally. The successful creation of transgenic marmosets provides a new animal model for human disease that has the great advantage of a close genetic relationship with humans. This model will be valuable to many fields of biomedical research. PMID:19478777

Sasaki, Erika; Suemizu, Hiroshi; Shimada, Akiko; Hanazawa, Kisaburo; Oiwa, Ryo; Kamioka, Michiko; Tomioka, Ikuo; Sotomaru, Yusuke; Hirakawa, Reiko; Eto, Tomoo; Shiozawa, Seiji; Maeda, Takuji; Ito, Mamoru; Ito, Ryoji; Kito, Chika; Yagihashi, Chie; Kawai, Kenji; Miyoshi, Hiroyuki; Tanioka, Yoshikuni; Tamaoki, Norikazu; Habu, Sonoko; Okano, Hideyuki; Nomura, Tatsuji

2009-05-28

176

Adenovirus hexon protein enhances nuclear delivery and increases transgene expression of polyethylenimine/plasmid DNA vectors.  

Science.gov (United States)

Inefficient nuclear delivery restricts transgene expression using polyelectrolyte DNA vectors. To increase transfer from the cytoplasm to the nucleus, we have covalently linked adenovirus hexon protein to polyethylenimine (PEI, 800 kDa). Activity of the conjugate was compared with PEI and PEI linked to albumin. Hexon-containing complexes gave 10-fold greater transgene expression in HepG2 cells than PEI/DNA or complexes containing albumin, without increasing cell uptake. Following cytoplasmic injection into Xenopus laevis oocytes, hexon-containing complexes showed reporter gene expression to be elevated by 10-fold compared with PEI/DNA. The ability of hexon to promote nuclear delivery of PEI/DNA nanoparticles was compared with that of classical nuclear localization sequences (NLS) by measuring transgene expression following intracytoplasmic microinjection of hexon-PEI/DNA complexes and NLS-albumin-PEI/DNA complexes in rat-1 fibroblasts. The resulting nuclear transfer efficiency was in the following order: hexon-PEI/DNA>NLS-albumin-PEI/DNA>PEI/DNA>DNA alone>albumin-PEI/DNA. The activities of both NLS-albumin-PEI and hexon-PEI were abolished by co-injection of wheat germ agglutinin, suggesting that both act by means of the nuclear pore complex (NPC); in contrast, excess free NLS-albumin abolished transgene expression with NLS-albumin-PEI/DNA, but only partially inhibited hexon-PEI/DNA. Nuclear transfer efficiency following cytoplasmic injection was dependent on DNA concentration for all materials, although hexon conjugates showed much better activity than NLS-albumin at low DNA doses (500-1000 plasmids/cell). Our data are consistent with hexon mediating nuclear delivery of plasmid complexes by means of the NPC, using mechanisms that are only partially dependent on the classical NLS import pathway. The hexon-mediated mechanism of nuclear import enables substantially better transgene expression, particularly when DNA concentrations in the cytoplasm are limiting. PMID:11708884

Carlisle, R C; Bettinger, T; Ogris, M; Hale, S; Mautner, V; Seymour, L W

2001-11-01

177

Transgene Mausmodelle zur Untersuchung von Nervenzellwanderungen  

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The transcription factor Pax6 has been found to be associated with neuronal migrations. To further elucidate the role of Pax6 in these migrations, transgenic mouse models were developed. Additionally, an organotypic culture system was established that allows to visualize embryonic migrations in vitro.In the YAC transgenic mouse line PhPax6-taulacZ, expression of the reporter gene taulacZ is controlled by the Pax6 promotor. In this line, cells expressing Pax6 in their nucleus will be marked by...

Benzing, Karsten

2002-01-01

178

Transgenic cultures: from the economic viewpoint  

Directory of Open Access Journals (Sweden)

Full Text Available The introduction of transgenic seeds for agricultural purposes poses modification to their production, due to the potential for reaching desired characteristics such as greater yield, this being fundamental in an economic environment characterised by open market conditions. However, acceptance of products resulting from genetic engineering is far from becoming a simple process; discussion relating to the predominance of private sector interests, the monopoly of knowledge and the safety of such seeds/food is currently in the spotlight. This article presents the main points of debate regarding adoption of transgenic cultures, contributing to discussion about this topic for Colombia.

Mauricio Mosquera

2011-12-01

179

Detection of potential transgenic plant DNA recipients among soil bacteria  

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The likelihood of gene transfer from transgenic plants to bacteria is dependent on gene number and the presence of homologous sequences. The large number of transgene copies in transplastomic (transgenes contained in the chloroplast genome) plant cells as well as the prokaryotic origin of the transgene, may thus significantly increase the likelihood of gene transfer to bacteria that colonize plant tissues. In order to assess the probability of such transfer, the length of homologous DNA seque...

Monier, Jean-michel; Bernillon, Dominique; Kay, Elizabeth; Faugier, Aure?lie; Rybalka, Oleksandra; Dessaux, Yves; Simonet, Pascal; Vogel, Timothy

2007-01-01

180

The rat as an animal model of Alzheimer's disease  

DEFF Research Database (Denmark)

As a disease model, the laboratory rat has contributed enormously to neuroscience research over the years. It has also been a popular animal model for Alzheimer's disease but its popularity has diminished during the last decade, as techniques for genetic manipulation in rats have lagged behind that of mice. In recent years, the rat has been making a comeback as an Alzheimer's disease model and the appearance of increasing numbers of transgenic rats will be a welcome and valuable complement to the existing mouse models. This review summarizes the contributions and current status of the rat as an animal model of Alzheimer's disease.

Benedikz, Eirikur; Kloskowska, Ewa

2009-01-01

 
 
 
 
181

Production of recombinant proteins in milk of transgenic and non-transgenic goats  

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Full Text Available Among all the transgenic mammalians produced so far, goats have represented an excellent model of transgenesis when considering the factors such as the market demand for protein, volume of milk produced per lactation and reproductive rate. Various recombinant proteins have been obtained from the transgenic and non-transgenic goats, and among these, human antithrombin, produced by the transgenic goats, was the first recombinant protein of animal origin to be released as a drug for the clinical use in humans. This review reports the aspects inherent to the production of recombinant proteins in the goats, from the production of the animal bioreactors up to the expression of these proteins in their milk.

Raylene Ramos Moura

2011-10-01

182

Production of pharmaceutical proteins from transgenic animals.  

Science.gov (United States)

Different systems are being studied and used to prepare recombinant proteins for pharmaceutical use. The blood, and still more the milk, from transgenic animals appear a very attractive source of pharmaceuticals. The cells from these animals are expected to produce well-matured proteins in potentially huge amounts. Several problems remain before this process becomes used in a large scale. Gene transfer remains a difficult and costly task for farm animals. The vectors carrying the genes coding for the proteins of interest are of unpredictable efficiency. Improvement of these vectors includes the choice of efficient promoters, introns and transcription terminators, the addition of matrix attached regions (MAR) and specialized chromatin sequences (SCS) to enhance the expression of the transgenes and to insulate them from the chromatin environment. Mice are routinely used to evaluate the gene constructs to be transferred into larger animals. Mice can also be utilized to prepare amounts as high as a few hundred mg of recombinant proteins from their milk. Rabbit appears adequate for amounts not higher than 1 kg per year. For larger quantities, goat, sheep, pig and cow are required. No recombinant proteins extracted from the blood or milk of transgenic animals are yet on the market. The relatively slow but real progress to improving the efficiency of this process inclines to be reasonably optimistic. Predictive reports suggest that 10% of the recombinant proteins, corresponding to a 100 million dollars annual market, will be prepared from the milk of transgenic animals by the end of the century. PMID:7764960

Houdebine, L M

1994-05-31

183

Transgenic Animals: Their Benefits To Human Welfare  

Science.gov (United States)

The issue-focused, reviewed, student article is about how transgenic animals, i.e., engineered to carry genes from other species, have the potential to improve human welfare in: agriculture, such as larger sheep that grow more wool, medicine, such as cows that produce insulin in their milk, andindustry, such as goats that produce spider silk for materials production.

Endang Tri Margawati (Bogor Agricultural University, Indonesia;)

2003-01-01

184

Monitoring transgenic plants using in vivo markers  

Energy Technology Data Exchange (ETDEWEB)

The gene coding for green fluorecent protein (GFP), isolated and cloned from the jellyfish Aequorea victoria, is an ideal transgene for the monitoring of any plant species. It has the ability to fluoresce without added substrate, enzyme, or cofactor; it does not introduce morphological or sexual aberrations when expressed. 7 refs., 1 fig.

Stewart, C.N. Jr. [Univ. of North Carolina, Greensboro, NC (United States)

1996-06-01

185

Transgenic plants with increased calcium stores  

Science.gov (United States)

The present invention provides transgenic plants over-expressing a transgene encoding a calcium-binding protein or peptide (CaBP). Preferably, the CaBP is a calcium storage protein and over-expression thereof does not have undue adverse effects on calcium homeostasis or biochemical pathways that are regulated by calcium. In preferred embodiments, the CaBP is calreticulin (CRT) or calsequestrin. In more preferred embodiments, the CaBP is the C-domain of CRT, a fragment of the C-domain, or multimers of the foregoing. In other preferred embodiments, the CaBP is localized to the endoplasmic reticulum by operatively associating the transgene encoding the CaBP with an endoplasmic reticulum localization peptide. Alternatively, the CaBP is targeted to any other sub-cellular compartment that permits the calcium to be stored in a form that is biologically available to the plant. Also provided are methods of producing plants with desirable phenotypic traits by transformation of the plant with a transgene encoding a CaBP. Such phenotypic traits include increased calcium storage, enhanced resistance to calcium-limiting conditions, enhanced growth and viability, increased disease and stress resistance, enhanced flower and fruit production, reduced senescence, and a decreased need for fertilizer production. Further provided are plants with enhanced nutritional value as human food or animal feed.

Wyatt, Sarah (Inventor); Tsou, Pei-Lan (Inventor); Robertson, Dominique (Inventor); Boss, Wendy (Inventor)

2004-01-01

186

Transgenic trees: how far the pollen flies?  

Directory of Open Access Journals (Sweden)

Full Text Available Implications of extended gene flow is discussed under the light of new laws issued, regulating the admixtures among organic, traditional and biotec agriculture. Recent scientific literature is reviewed on the role of gene flow in gene escape in transgenic forest trees.

Leonardi S

2006-01-01

187

Metal resistance sequences and transgenic plants  

Science.gov (United States)

The present invention provides nucleic acid sequences encoding a metal ion resistance protein, which are expressible in plant cells. The metal resistance protein provides for the enzymatic reduction of metal ions including but not limited to divalent Cu, divalent mercury, trivalent gold, divalent cadmium, lead ions and monovalent silver ions. Transgenic plants which express these coding sequences exhibit increased resistance to metal ions in the environment as compared with plants which have not been so genetically modified. Transgenic plants with improved resistance to organometals including alkylmercury compounds, among others, are provided by the further inclusion of plant-expressible organometal lyase coding sequences, as specifically exemplified by the plant-expressible merB coding sequence. Furthermore, these transgenic plants which have been genetically modified to express the metal resistance coding sequences of the present invention can participate in the bioremediation of metal contamination via the enzymatic reduction of metal ions. Transgenic plants resistant to organometals can further mediate remediation of organic metal compounds, for example, alkylmetal compounds including but not limited to methyl mercury, methyl lead compounds, methyl cadmium and methyl arsenic compounds, in the environment by causing the freeing of mercuric or other metal ions and the reduction of the ionic mercury or other metal ions to the less toxic elemental mercury or other metals.

Meagher, Richard Brian (Athens, GA); Summers, Anne O. (Athens, GA); Rugh, Clayton L. (Athens, GA)

1999-10-12

188

Progress in Xenotransplantation Research Employing Transgenic Pigs  

Directory of Open Access Journals (Sweden)

Full Text Available Microinjection of foreign DNA into pronuclei of a fertilized oocyte has predominantly been used for the generation of transgenic livestock. This technology works reliably, but is inefficient and results in random integration and variable expression patterns in the transgenic offspring. Nevertheless, remarkable achievements have been made with this technology with regard to xenotransplantation. Transgenic pigs that express human complement regulating proteins have been tested in their ability to serve as donors in human organ transplantation (i.e. xenotransplantation. In vitro and in vivo data convincingly show that the hyperacute rejection response can be overcome in a clinically acceptable manner by successfully employing this strategy. The recent developments in nuclear transfer and its merger with the growing genomic data allow targeted and regulatable transgenesis. Systems for efficient homologous recombination in somatic cells are being developed and the first knock-out pigs, carrying a deletion in the a-galactosyltransferase gene, were recently generated. It is anticipated that poly-transgenic pigs will be available as donors for functional xenografts within a few years. Similarly, pigs may serve as donors for a variety of xenogenic cells and tissues. The availability of these technologies is essential to maintain "genetic security" and to ensure absence of unwanted side effects.

H. Niemann

2003-06-01

189

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

International Nuclear Information System (INIS)

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

190

Evaluation of haematological, biochemical and histopathological parameters of transgenic rabbits.  

Science.gov (United States)

The aim of our study was to compare the hFVIII mRNA expression in different organs, pathological changes and selected haematological and biochemical blood parameters between transgenic and non-transgenic rabbits from F3 generation. Selected physiological parameters of 3- to 4-month-old transgenic rabbits from F3 generation carrying human factor VIII gene (hFVIII) were analysed and compared with those of non-transgenic ones. Before slaughtering, the blood for haematological and biochemical analysis was taken from the central ear artery. Pathological and histological examination of vital organs and RT-PCR analysis of several tissue organs of transgenic and non-transgenic animals were performed after slaughtering. Except for the mammary gland tissue, slight hfVIII mRNA expression in the spleen, lung and brain and none expression in the liver, kidney, skeletal muscle and heart of rabbits were recorded. pathological examination of vital organs showed some pathological changes in both transgenic and non-transgenic rabbits which were confirmed by histological qualitative evaluations. Statistically significant lower values of blood haemoglobin in blood of transgenic (11.86+/-0.86) animals compared with non-transgenic (12.41+/-1.02, Pbiochemical serum parameters of transgenic rabbits were higher in comparison with non-transgenic ones. Significant differences were found in the concentration of the urea, AST and GMT between transgenic and non-transgenic animals (P<0.001) and in the total protein content, the difference was significant at P<0.05. In conclusion, our results showed that no considerable impact on the general health was found in transgenic rabbits. PMID:17931230

Jurcik, R; Suvegova, K; Hanusova, E; Massanyi, P; Ryban, L; Chrenek, P

2007-11-01

191

Split-Cre Complementation Restores Combination Activity on Transgene Excision in Hair Roots of Transgenic Tobacco  

Science.gov (United States)

The Cre/loxP system is increasingly exploited for genetic manipulation of DNA in vitro and in vivo. It was previously reported that inactive ‘‘split-Cre’’ fragments could restore Cre activity in transgenic mice when overlapping co-expression was controlled by two different promoters. In this study, we analyzed recombination activities of split-Cre proteins, and found that no recombinase activity was detected in the in vitro recombination reaction in which only the N-terminal domain (NCre) of split-Cre protein was expressed, whereas recombination activity was obtained when the C-terminal (CCre) or both NCre and CCre fragments were supplied. We have also determined the recombination efficiency of split-Cre proteins which were co-expressed in hair roots of transgenic tobacco. No Cre recombination event was observed in hair roots of transgenic tobacco when the NCre or CCre genes were expressed alone. In contrast, an efficient recombination event was found in transgenic hairy roots co-expressing both inactive split-Cre genes. Moreover, the restored recombination efficiency of split-Cre proteins fused with the nuclear localization sequence (NLS) was higher than that of intact Cre in transgenic lines. Thus, DNA recombination mediated by split-Cre proteins provides an alternative method for spatial and temporal regulation of gene expression in transgenic plants. PMID:25329460

Wen, Mengling; Gao, Yuan; Wang, Lijun; Ran, Lingyu; Li, Jiahui; Luo, Keming

2014-01-01

192

Primary transgenic bovine cells and their rejuvenated cloned equivalents show transgene-specific epigenetic differences.  

Science.gov (United States)

Cell-mediated transgenesis, based on somatic cell nuclear transfer (SCNT), provides the opportunity to shape the genetic make-up of cattle. Bovine primary fetal fibroblasts, commonly used cells for SCNT, have a limited lifespan, and complex genetic modifications that require sequential transfections can be challenging time and cost-wise. To overcome these limitations, SCNT is frequently used to rejuvenate the cell lines and restore exhausted growth potential. We have designed a construct to be used in a 2-step cassette exchange experiment. Our transgene contains a puromycin resistance marker gene and an enhanced green fluorescence protein (EGFP) expression cassette, both driven by a strong mammalian promoter, and flanked by loxP sites and sequences from the bovine ?-casein locus. Several transgenic cell lines were generated by random insertion into primary bovine cell lines. Two of these original cell lines were rederived by SCNT and new primary cells, with the same genetic makeup as the original donors, were established. While the original cell lines were puromycin-resistant and had a characteristic EGFP expression profile, all rejuvenated cell lines were sensitive to puromycin, and displayed varied EGFP expression, indicative of various degrees of silencing. When the methylation states of individual CpG sites within the transgene were analyzed, a striking increase in transgene-specific methylation was observed in all rederived cell lines. The results indicate that original transgenic donor cells and their rejuvenated derivatives may not be equivalent and differ in the functionality of their transgene sequences. PMID:22532863

Alonso-González, Lucia; Couldrey, Christine; Meinhardt, Marcus W; Cole, Sally A; Wells, David N; Laible, Götz

2012-01-01

193

[Animal welfare problems concerning the use of transgenic animals  

Science.gov (United States)

Using transgenic animals as clinical models pose certain problems since they can suffer. Yet in single cases transgenic animals can reduce the suffering of (other) animals. The permission to generate transgenic animals is not yet clearly regulated in Switzerland. The term "dignity of creature", as formulated in the Swiss Constitution, has to be defined for the Swiss animal protection law. We present the recommendations of the commission for ethical questions concerning transgenic animals appointed by the Federal Council. Partly, these recommendations shall also be applied to the traditional breeding methods. We support the nomination of a national ethics committee for transgenic animals. PMID:11208267

Mani, Peter

1998-01-01

194

Glucose metabolic gene expression in growth hormone transgenic coho salmon.  

Science.gov (United States)

Salmonids are generally known to be glucose intolerant. However, previous studies have shown that growth hormone (GH) transgenic coho salmon display modified nutritional regulation of glycolysis and lipogenesis compared to non-transgenic fish, suggesting the potential for better use of glucose in GH transgenic fish. To examine this in detail, GH transgenic and non-transgenic coho salmon were subjected to glucose tolerance test and subsequent metabolic assessments. After intra-peritoneal injection of 250mg/kg glucose, we analysed post-injection kinetics of glycaemia and expression of several key target genes highly involved in glucose homeostasis in muscle and liver tissues. Our data show no significant differences in plasma glucose levels during peak hyperglycaemia (3-6h after injection), demonstrating a similar glucose tolerance between transgenic and non transgenic. However, and unrelated to the hyperglycaemic episode, GH transgenic fish return to a slightly lower basal glycaemia values 24h after injection. Correspondingly, GH transgenic fish exhibited higher mRNA levels of glucokinase (GK) and glucose-6-phosphate dehydrogenase (G6PDH) in liver, and glucose transporter (GLUT4) in muscle. These data suggest that these metabolic actors may be involved in different glucose use in GH transgenic fish, which would be expected to influence the glucose challenge response. Overall, our data demonstrate that GH transgenic coho salmon may be a pertinent animal model for further study of glucose metabolism in carnivorous fish. PMID:24486143

Panserat, Stéphane; Kamalam, Biju Sam; Fournier, Jeanne; Plagnes-Juan, Elisabeth; Woodward, Krista; Devlin, Robert H

2014-04-01

195

Assessment of peanut quality and compositional characteristics among transgenic sclerotinia blight-resistant and non-transgenic susceptible cultivars.  

Science.gov (United States)

This study presents the results of a comparison that includes an analysis of variance and a canonical discriminant analysis to determine compositional equivalence and similarity between transgenic, sclerotinia blight-resistant and non-transgenic, susceptible cultivars of peanut in 3 years of field trials. Three Virginia-type cultivars (NC 7, Wilson, and Perry) and their corresponding transgenic lines (N70, W73, and P39) with a barley oxalate oxidase gene were analyzed for differences in key mineral nutrients, fatty acid components, hay constituents, and grade characteristics. Results from both analyses demonstrated that transgenic lines were compositionally similar to their non-transgenic parent cultivar in all factors as well as market-grade characteristics and nutritional value. Transgenic lines expressing oxalate oxidase for resistance to sclerotinia blight were substantially equivalent to their non-transgenic parent cultivar in quality and compositional characteristics. PMID:24972023

Hu, Jiahuai; Telenko, Darcy E P; Phipps, Patrick M; Grabau, Elizabeth A

2014-08-01

196

Green tea polyphenols control dysregulated glutamate dehydrogenase in transgenic mice by hijacking the ADP activation site.  

Science.gov (United States)

Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of L-glutamate and, in animals, is extensively regulated by a number of metabolites. Gain of function mutations in GDH that abrogate GTP inhibition cause the hyperinsulinism/hyperammonemia syndrome (HHS), resulting in increased pancreatic ?-cell responsiveness to leucine and susceptibility to hypoglycemia following high protein meals. We have previously shown that two of the polyphenols from green tea (epigallocatechin gallate (EGCG) and epicatechin gallate (ECG)) inhibit GDH in vitro and that EGCG blocks GDH-mediated insulin secretion in wild type rat islets. Using structural and site-directed mutagenesis studies, we demonstrate that ECG binds to the same site as the allosteric regulator, ADP. Perifusion assays using pancreatic islets from transgenic mice expressing a human HHS form of GDH demonstrate that the hyperresponse to glutamine caused by dysregulated GDH is blocked by the addition of EGCG. As observed in HHS patients, these transgenic mice are hypersensitive to amino acid feeding, and this is abrogated by oral administration of EGCG prior to challenge. Finally, the low basal blood glucose level in the HHS mouse model is improved upon chronic administration of EGCG. These results suggest that this common natural product or some derivative thereof may prove useful in controlling this genetic disorder. Of broader clinical implication is that other groups have shown that restriction of glutamine catabolism via these GDH inhibitors can be useful in treating various tumors. This HHS transgenic mouse model offers a highly useful means to test these agents in vivo. PMID:21813650

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

2011-09-30

197

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

Science.gov (United States)

Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of l-glutamate and, in animals, is extensively regulated by a number of metabolites. Gain of function mutations in GDH that abrogate GTP inhibition cause the hyperinsulinism/hyperammonemia syndrome (HHS), resulting in increased pancreatic ?-cell responsiveness to leucine and susceptibility to hypoglycemia following high protein meals. We have previously shown that two of the polyphenols from green tea (epigallocatechin gallate (EGCG) and epicatechin gallate (ECG)) inhibit GDH in vitro and that EGCG blocks GDH-mediated insulin secretion in wild type rat islets. Using structural and site-directed mutagenesis studies, we demonstrate that ECG binds to the same site as the allosteric regulator, ADP. Perifusion assays using pancreatic islets from transgenic mice expressing a human HHS form of GDH demonstrate that the hyperresponse to glutamine caused by dysregulated GDH is blocked by the addition of EGCG. As observed in HHS patients, these transgenic mice are hypersensitive to amino acid feeding, and this is abrogated by oral administration of EGCG prior to challenge. Finally, the low basal blood glucose level in the HHS mouse model is improved upon chronic administration of EGCG. These results suggest that this common natural product or some derivative thereof may prove useful in controlling this genetic disorder. Of broader clinical implication is that other groups have shown that restriction of glutamine catabolism via these GDH inhibitors can be useful in treating various tumors. This HHS transgenic mouse model offers a highly useful means to test these agents in vivo. PMID:21813650

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

2011-01-01

198

Transgenic arthropods and the sterile insect technique  

International Nuclear Information System (INIS)

The Sterile Insect Technique can benefit from transgenesis in three ways by creating; (1) genetically marked strains, (2) genetic sexing strains and (3) strains that induce molecular sterility in the field. Experience with the development of genetic sexing strains based on indicates that caution is required during the experimental evaluation of any potential transgenic strain. Two major scientific concerns involve the overall fitness of transgenic strains and their stability over time. The latter being very important especially when the extremely large numbers of insects that are mass reared is taken into account. Currently transformation events are random and it will probably be necessary to select suitable strains from many that are induced. The success of transformation itself in many insect species will enable many new strategies to be developed and tested. (author)

199

Transgenics and vertebrate cloning as tools for species conservation.  

Science.gov (United States)

It has been suggested that transgenics and vertebrate cloning have a role to play in conservation. Now is the time to evaluate their risks and benefits, before these technologies are widely implemented in our field. Direct risks of transgenics include escape and introgression of transgenes into wild populations; weedy invasion by transgenic organisms; toxicity or pathogenicity of engineered organisms and their products; and human error in the field testing and tracking of transgenic organisms. Indirect risks include environmental effects of increased herbicide use; the danger that engineered organisms may aid the development of bioweapons; the likelihood that gene patenting will lead to the privatization of natural resources; and the diversion of support from less glamorous forms of conservation. Formal risk assessments are commonly used to evaluate transgenic procedures, but our incomplete understanding of both ecosystem processes and the action of transgenes renders most of these assessments scientifically and socially unjustified. Nevertheless, a few, low-risk applications of transgenics may be possible: for example, "super-sterile" ornamental cultivars. Vertebrate cloning poses little risk to the environment, but it can consume scarce conservation resources, and its chances of success in preserving species seem poor To date, the conservation benefits of transgenics and vertebrate cloning remain entirely theoretical, but many of the risks are known and documented. Conservation biologists should devote their research and energies to the established methods of conservation, none of which require transgenics or vertebrate cloning. PMID:16909565

Ehrenfeld, David

2006-06-01

200

Transgene mobilization and regulatory uncertainty for non-GE fruit products of transgenic rootstocks.  

Science.gov (United States)

Genetically engineered (GE) rootstocks may offer some advantages for biotechnology applications especially in woody perennial crops such as grape or walnut. Transgrafting combines horticultural grafting practices with modern GE methods for crop improvement. Here, a non-GE conventional scion (upper stem portion) is grafted onto a transgenic GE rootstock. Thus, the scion does not contain the genetic modification present in the rootstock genome. We examined transgene presence in walnut and tomato GE rootstocks and non-GE fruit-bearing scions. Mobilization of transgene DNA, protein, and mRNA across the graft was not detected. Though transgenic siRNA mobilization was not observed in grafted tomatoes or walnut scions, transgenic siRNA signal was detected in walnut kernels. Prospective benefits from transgrafted plants include minimized risk of GE pollen flow (Lev-Yadun and Sederoff, 2001), possible use of more than one scion per approved GE rootstock which could help curb the estimated US$136 million (CropLife International, 2011) cost to bring a GE crop to international markets, as well as potential for improved consumer and market acceptance since the consumable product is not itself GE. Thus, transgrafting provides an alternative option for agricultural industries wishing to expand their biotechnology portfolio. PMID:22749907

Haroldsen, Victor M; Chi-Ham, Cecilia L; Bennett, Alan B

2012-10-31

 
 
 
 
201

Regulation of transgene expression in genetic immunization  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english The use of mammalian gene expression vectors has become increasingly important for genetic immunization and gene therapy as well as basic research. Essential for the success of these vectors in genetic immunization is the proper choice of a promoter linked to the antigen of interest. Many genetic im [...] munization vectors use promoter elements from pathogenic viruses including SV40 and CMV. Lymphokines produced by the immune response to proteins expressed by these vectors could inhibit further transcription initiation by viral promoters. Our objective was to determine the effect of IFN-g on transgene expression driven by viral SV40 or CMV promoter/enhancer and the mammalian promoter/enhancer for the major histocompatibility complex class I (MHC I) gene. We transfected the luciferase gene driven by these three promoters into 14 cell lines of many tissues and several species. Luciferase assays of transfected cells untreated or treated with IFN-g indicated that although the viral promoters could drive luciferase production in all cell lines tested to higher or lower levels than the MHC I promoter, treatment with IFN-g inhibited transgene expression in most of the cell lines and amplification of the MHC I promoter-driven transgene expression in all cell lines. These data indicate that the SV40 and CMV promoter/enhancers may not be a suitable choice for gene delivery especially for genetic immunization or cancer cytokine gene therapy. The MHC I promoter/enhancer, on the other hand, may be an ideal transgene promoter for applications involving the immune system.

J.S., Harms; S.C., Oliveira; G.A., Splitter.

202

Transgenic nonhuman primates for neurodegenerative diseases  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Animal models that represent human diseases constitute an important tool in understanding the pathogenesis of the diseases, and in developing effective therapies. Neurodegenerative diseases are complex disorders involving neuropathologic and psychiatric alterations. Although transgenic and knock-in mouse models of Alzheimer's disease, (AD, Parkinson's disease (PD and Huntington's disease (HD have been created, limited representation in clinical aspects has been recognized and the rodent models lack true neurodegeneration. Chemical induction of HD and PD in nonhuman primates (NHP has been reported, however, the role of intrinsic genetic factors in the development of the diseases is indeterminable. Nonhuman primates closely parallel humans with regard to genetic, neuroanatomic, and cognitive/behavioral characteristics. Accordingly, the development of NHP models for neurodegenerative diseases holds greater promise for success in the discovery of diagnoses, treatments, and cures than approaches using other animal species. Therefore, a transgenic NHP carrying a mutant gene similar to that of patients will help to clarify our understanding of disease onset and progression. Additionally, monitoring disease onset and development in the transgenic NHP by high resolution brain imaging technology such as MRI, and behavioral and cognitive testing can all be carried out simultaneously in the NHP but not in other animal models. Moreover, because of the similarity in motor repertoire between NHPs and humans, it will also be possible to compare the neurologic syndrome observed in the NHP model to that in patients. Understanding the correlation between genetic defects and physiologic changes (e.g. oxidative damage will lead to a better understanding of disease progression and the development of patient treatments, medications and preventive approaches for high risk individuals. The impact of the transgenic NHP model in understanding the role which genetic disorders play in the development of efficacious interventions and medications is foreseeable.

Chan Anthony WS

2004-06-01

203

Transgenic animals as bioproducers of therapeutic proteins.  

Science.gov (United States)

Many human therapeutic proteins are currently produced with the aid of recombinant DNA technology in microbial bioreactors and a few also in large-scale animal cell cultures. Although extremely cost-efficient, the microbial production system has many inherent limitations. Micro-organisms, such as bacteria, can read the universal genetic code and hence produce human proteins with correct amino acid sequence, but cannot carry out post-translational modifications, such as glycosylation, or fold the newly synthesized protein properly to ultimately generate a biologically active entity. Moreover, even though the production of the proteins as such is inexpensive, the downstream processing of the final product may be extremely difficult and costly. Many of these disadvantages, especially the lack of post-translational modifications, can be overcome by employing large-scale animal cell cultures for the production of proteins of pharmaceutical interest. However, due to the long generation time and the requirement for rich culture media, the use of animal cell bioreactors is unacceptably expensive. With the advent of transgenic technology, the production of human pharmaceuticals in large transgenic animals has become more and more attractive. The use of targeted gene transfer, the expression of the transgene of interest can be directed to occur in the mammary gland of large farm animals, such as pigs, sheep, goats or dairy cattle, and hence the transgene product is ultimately being secreted into the milk. Although not yet in commercial use, the last few years have witnessed a remarkable progress in this area and proved the feasibility of the use of 'molecular farming' in high-quantity, low-cost production of valuable therapeutic or industrial proteins. While reviewing the progress of the field over the past few years, we discuss in somewhat greater detail aspects connected with the use of dairy cattle as bioproducers of human therapeutic proteins. PMID:1389089

Jänne, J; Hyttinen, J M; Peura, T; Tolvanen, M; Alhonen, L; Halmekytö, M

1992-08-01

204

Regulation of transgene expression in genetic immunization  

Directory of Open Access Journals (Sweden)

Full Text Available The use of mammalian gene expression vectors has become increasingly important for genetic immunization and gene therapy as well as basic research. Essential for the success of these vectors in genetic immunization is the proper choice of a promoter linked to the antigen of interest. Many genetic immunization vectors use promoter elements from pathogenic viruses including SV40 and CMV. Lymphokines produced by the immune response to proteins expressed by these vectors could inhibit further transcription initiation by viral promoters. Our objective was to determine the effect of IFN-g on transgene expression driven by viral SV40 or CMV promoter/enhancer and the mammalian promoter/enhancer for the major histocompatibility complex class I (MHC I gene. We transfected the luciferase gene driven by these three promoters into 14 cell lines of many tissues and several species. Luciferase assays of transfected cells untreated or treated with IFN-g indicated that although the viral promoters could drive luciferase production in all cell lines tested to higher or lower levels than the MHC I promoter, treatment with IFN-g inhibited transgene expression in most of the cell lines and amplification of the MHC I promoter-driven transgene expression in all cell lines. These data indicate that the SV40 and CMV promoter/enhancers may not be a suitable choice for gene delivery especially for genetic immunization or cancer cytokine gene therapy. The MHC I promoter/enhancer, on the other hand, may be an ideal transgene promoter for applications involving the immune system.

J.S. Harms

1999-02-01

205

Regulation of transgene expression in genetic immunization  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english The use of mammalian gene expression vectors has become increasingly important for genetic immunization and gene therapy as well as basic research. Essential for the success of these vectors in genetic immunization is the proper choice of a promoter linked to the antigen of interest. Many genetic im [...] munization vectors use promoter elements from pathogenic viruses including SV40 and CMV. Lymphokines produced by the immune response to proteins expressed by these vectors could inhibit further transcription initiation by viral promoters. Our objective was to determine the effect of IFN-g on transgene expression driven by viral SV40 or CMV promoter/enhancer and the mammalian promoter/enhancer for the major histocompatibility complex class I (MHC I) gene. We transfected the luciferase gene driven by these three promoters into 14 cell lines of many tissues and several species. Luciferase assays of transfected cells untreated or treated with IFN-g indicated that although the viral promoters could drive luciferase production in all cell lines tested to higher or lower levels than the MHC I promoter, treatment with IFN-g inhibited transgene expression in most of the cell lines and amplification of the MHC I promoter-driven transgene expression in all cell lines. These data indicate that the SV40 and CMV promoter/enhancers may not be a suitable choice for gene delivery especially for genetic immunization or cancer cytokine gene therapy. The MHC I promoter/enhancer, on the other hand, may be an ideal transgene promoter for applications involving the immune system.

J.S., Harms; S.C., Oliveira; G.A., Splitter.

1999-02-01

206

Transgenic approaches to western corn rootworm control.  

Science.gov (United States)

The western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae) is a significant corn pest throughout the United States corn belt. Rootworm larvae feed on corn roots causing yield losses and control expenditures that are estimated to exceed US$1 billion annually. Traditional management practices to control rootworms such as chemical insecticides or crop rotation have suffered reduced effectiveness due to the development of physiological and behavioral resistance. Transgenic maize expressing insecticidal proteins are very successful in protecting against rootworm damage and preserving corn yield potential. However, the high rate of grower adoption and early reliance on hybrids expressing a single mode of action and low-dose traits threatens the durability of commercialized transgenic rootworm technology for rootworm control. A summary of current transgenic approaches for rootworm control and the corresponding insect resistance management practices is included. An overview of potential new modes of action based on insecticidal proteins, and especially RNAi targeting mRNA coding for essential insect proteins is provided. PMID:23604211

Narva, Kenneth E; Siegfried, Blair D; Storer, Nicholas P

2013-01-01

207

Transgenic oil palm: production and projection.  

Science.gov (United States)

Oil palm is an important economic crop for Malaysia. Genetic engineering could be applied to produce transgenic oil palms with high value-added fatty acids and novel products to ensure the sustainability of the palm oil industry. Establishment of a reliable transformation and regeneration system is essential for genetic engineering. Biolistic was initially chosen as the method for oil palm transformation as it has been the most successful method for monocotyledons to date. Optimization of physical and biological parameters, including testing of promoters and selective agents, was carried out as a prerequisite for stable transformation. This has resulted in the successful transfer of reporter genes into oil palm and the regeneration of transgenic oil palm, thus making it possible to improve the oil palm through genetic engineering. Besides application of the Biolistics method, studies on transformation mediated by Agrobacterium and utilization of the green fluorescent protein gene as a selectable marker gene have been initiated. Upon the development of a reliable transformation system, a number of useful targets are being projected for oil palm improvement. Among these targets are high-oleate and high-stearate oils, and the production of industrial feedstock such as biodegradable plastics. The efforts in oil palm genetic engineering are thus not targeted as commodity palm oil. Due to the long life cycle of the palm and the time taken to regenerate plants in tissue culture, it is envisaged that commercial planting of transgenic palms will not occur any earlier than the year 2020. PMID:11171275

Parveez, G K; Masri, M M; Zainal, A; Majid, N A; Yunus, A M; Fadilah, H H; Rasid, O; Cheah, S C

2000-12-01

208

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

International Nuclear Information System (INIS)

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

209

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

Energy Technology Data Exchange (ETDEWEB)

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

Ham, Young-Mi, E-mail: youngmi_ham@hms.harvard.edu [College of Life Science and Biotechnology, Korea University, Seoul (Korea, Republic of); Department of Cell Biology, Harvard Medical School, Boston, MA 02115 (United States); Mahoney, Sarah Jane [Department of Cell Biology, Harvard Medical School, Boston, MA 02115 (United States)

2013-06-10

210

In situ methods to localize transgenes and transcripts in interphase nuclei: a tool for transgenic plant research  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Genetic engineering of commercially important crops has become routine in many laboratories. However, the inability to predict where a transgene will integrate and to efficiently select plants with stable levels of transgenic expression remains a limitation of this technology. Fluorescence in situ hybridization (FISH is a powerful technique that can be used to visualize transgene integration sites and provide a better understanding of transgene behavior. Studies using FISH to characterize transgene integration have focused primarily on metaphase chromosomes, because the number and position of integration sites on the chromosomes are more easily determined at this stage. However gene (and transgene expression occurs mainly during interphase. In order to accurately predict the activity of a transgene, it is critical to understand its location and dynamics in the three-dimensional interphase nucleus. We and others have developed in situ methods to visualize transgenes (including single copy genes and their transcripts during interphase from different tissues and plant species. These techniques reduce the time necessary for characterization of transgene integration by eliminating the need for time-consuming segregation analysis, and extend characterization to the interphase nucleus, thus increasing the likelihood of accurate prediction of transgene activity. Furthermore, this approach is useful for studying nuclear organization and the dynamics of genes and chromatin.

Shaw Peter

2006-11-01

211

Comparing the Agronomic and Grain Quality Characteristics of Transgenic Rice Lines Expressing cry1Ab vs. Non-Transgenic Controls  

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Full Text Available This study aimed to investigate and compare the agronomic and grain quality attributes of three advanced backcross-derived transgenic rice lines expressing synthetic cry1Ab gene vs. non-transgenic control in a Randomized Complete Block Design (RCBD under field conditions. The data exhibited that transgenic rice lines, Neda and Nemat were higher in height, earlier in maturity and highly resistant to striped stem borer (Chilo suppressalis in comparing with non-transgenic varieties. In contrast, no significant difference was observed for transgenic Khazar as compared to its control, except for 1000-grain weight. Laboratory tests for grain physicochemical properties showed no significant variations between transgenic lines and non-transgenic controls. However, some variations for traits like Amylose Content (AC and Gel Consistency (GC were seen for transgenic Neda and Khazar, respectively. As regards the rice striped stem borer natural infestation, field-release experiment indicated that all three transgenic rice lines conferred a very high degree of resistance to rice striped stem borer as compared to non-transgenic check varieties.

G. Kiani

2009-01-01

212

Production of recombinant proteins in milk of transgenic and non-transgenic goats  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Among all the transgenic mammalians produced so far, goats have represented an excellent model of transgenesis when considering the factors such as the market demand for protein, volume of milk produced per lactation and reproductive rate. Various recombinant proteins have been obtained from the tra [...] nsgenic and non-transgenic goats, and among these, human antithrombin, produced by the transgenic goats, was the first recombinant protein of animal origin to be released as a drug for the clinical use in humans. This review reports the aspects inherent to the production of recombinant proteins in the goats, from the production of the animal bioreactors up to the expression of these proteins in their milk.

Raylene Ramos, Moura; Luciana Magalhães, Melo; Vicente José de Figueirêdo, Freitas.

213

Dual reporter transgene driven by 2.3Col1a1 promoter is active in differentiated osteoblasts  

Science.gov (United States)

AIM: As quantitative and spatial analyses of promoter reporter constructs are not easily performed in intact bone, we designed a reporter gene specific to bone, which could be analyzed both visually and quantitatively by using chloramphenicol acetyltransferase (CAT) and a cyan version of green fluorescent protein (GFPcyan), driven by a 2.3-kb fragment of the rat collagen promoter (Col2.3). METHODS: The construct Col2.3CATiresGFPcyan was used for generating transgenic mice. Quantitative measurement of promoter activity was performed by CAT analysis of different tissues derived from transgenic animals; localization was performed by visualized GFP in frozen bone sections. To assess transgene expression during in vitro differentiation, marrow stromal cell and neonatal calvarial osteoblast cultures were analyzed for CAT and GFP activity. RESULTS: In mice, CAT activity was detected in the calvaria, long bone, teeth, and tendon, whereas histology showed that GFP expression was limited to osteoblasts and osteocytes. In cell culture, increased activity of CAT correlated with increased differentiation, and GFP activity was restricted to mineralized nodules. CONCLUSION: The concept of a dual reporter allows a simultaneous visual and quantitative analysis of transgene activity in bone.

Marijanovic, Inga; Jiang, Xi; Kronenberg, Mark S.; Stover, Mary Louise; Erceg, Ivana; Lichtler, Alexander C.; Rowe, David W.

2003-01-01

214

An industry perspective on the utility of short-term carcinogenicity testing in transgenic mice in pharmaceutical development.  

Science.gov (United States)

International guidelines allow for use of a short-term cancer bioassay (twenty-six weeks) in transgenic mice as a substitute for one of the two required long-term rodent bioassays in the preclinical safety evaluation of pharmaceuticals. The two models that have gained the widest acceptance by sponsors and regulatory authorities are the CB6F1-RasH2 mouse hemizygous for a human H-ras transgene and the B6.129N5-Trp53 mouse heterozygous for a p53 null allele. The p53(+/-) model is of particular value for compounds with residual concern that genotoxic activity may contribute to tumorigenesis. The rasH2 model is an appropriate alternative without regard to evidence of genotoxic potential. Since results from a short-term bioassay can be obtained relatively early in drug development, there is the potential for more timely assessment of cancer risk for individuals in long-term clinical trials. Use of these models in preclinical safety evaluation also significantly reduces animal use, time, and manpower. Preliminary findings indicate that prediction of two-year rat bioassay outcomes based on data from chronic rat toxicity studies, together with early assessment of carcinogenic potential in short-term transgenic models, may have the potential to increase the timeliness and efficiency of strategies for the identification of human carcinogenic hazards. PMID:19893055

Storer, Richard D; Sistare, Frank D; Reddy, M Vijayaraj; DeGeorge, Joseph J

2010-01-01

215

Perspectives on the state of insect transgenics.  

Science.gov (United States)

Genetic transformation is a critical component to the fundamental genetic analysis of insect species and holds great promise for establishing strains that improve population control and behavior for practical application. This is especially so for insects that are disease vectors, many of which are currently subject to genomic sequence analysis, and intensive population control measures that must be improved for better efficacy and cost-effectiveness. Transposon-mediated germ-line transformation has been the ultimate goal for most fundamental and practical studies, and impressive strides have been made in recent development of transgene vector and marker systems for several mosquito species. This has resulted in rapid advances in functional genomic sequence analysis and new strategies for biological control based on conditional lethality. Importantly, advances have also been made in our ability to use these systems more effectively in terms of enhanced stability and targeting to specific genomic loci. Nevertheless, not all insects are currently amenable to germ-line transformation techniques, and thus advances in transient somatic expression and paratransgenesis have also been critical, if not preferable for some applications. Of particular importance is how this technology will be used for practical application. Early ideas for population replacement of indigenous pests with innocuous transgenic siblings by transposon-vector spread, may require reevaluation in terms of our current knowledge of the behavior of transposons currently available for transformation. The effective implementation of any control program using released transgenics, will also benefit from broadening the perspective of these control measures as being more mainstream than exotic. PMID:18510010

O'Brochta, David A; Handler, Alfred M

2008-01-01

216

Transgene detection by digital droplet PCR.  

Science.gov (United States)

Somatic gene therapy is a promising tool for the treatment of severe diseases. Because of its abuse potential for performance enhancement in sports, the World Anti-Doping Agency (WADA) included the term 'gene doping' in the official list of banned substances and methods in 2004. Several nested PCR or qPCR-based strategies have been proposed that aim at detecting long-term presence of transgene in blood, but these strategies are hampered by technical limitations. We developed a digital droplet PCR (ddPCR) protocol for Insulin-Like Growth Factor 1 (IGF1) detection and demonstrated its applicability monitoring 6 mice injected into skeletal muscle with AAV9-IGF1 elements and 2 controls over a 33-day period. A duplex ddPCR protocol for simultaneous detection of Insulin-Like Growth Factor 1 (IGF1) and Erythropoietin (EPO) transgenic elements was created. A new DNA extraction procedure with target-orientated usage of restriction enzymes including on-column DNA-digestion was established. In vivo data revealed that IGF1 transgenic elements could be reliably detected for a 33-day period in DNA extracted from whole blood. In vitro data indicated feasibility of IGF1 and EPO detection by duplex ddPCR with high reliability and sensitivity. On-column DNA-digestion allowed for significantly improved target detection in downstream PCR-based approaches. As ddPCR provides absolute quantification, it ensures excellent day-to-day reproducibility. Therefore, we expect this technique to be used in diagnosing and monitoring of viral and bacterial infection, in detecting mutated DNA sequences as well as profiling for the presence of foreign genetic material in elite athletes in the future. PMID:25375130

Moser, Dirk A; Braga, Luca; Raso, Andrea; Zacchigna, Serena; Giacca, Mauro; Simon, Perikles

2014-01-01

217

Transgene Detection by Digital Droplet PCR  

Science.gov (United States)

Somatic gene therapy is a promising tool for the treatment of severe diseases. Because of its abuse potential for performance enhancement in sports, the World Anti-Doping Agency (WADA) included the term ‘gene doping’ in the official list of banned substances and methods in 2004. Several nested PCR or qPCR-based strategies have been proposed that aim at detecting long-term presence of transgene in blood, but these strategies are hampered by technical limitations. We developed a digital droplet PCR (ddPCR) protocol for Insulin-Like Growth Factor 1 (IGF1) detection and demonstrated its applicability monitoring 6 mice injected into skeletal muscle with AAV9-IGF1 elements and 2 controls over a 33-day period. A duplex ddPCR protocol for simultaneous detection of Insulin-Like Growth Factor 1 (IGF1) and Erythropoietin (EPO) transgenic elements was created. A new DNA extraction procedure with target-orientated usage of restriction enzymes including on-column DNA-digestion was established. In vivo data revealed that IGF1 transgenic elements could be reliably detected for a 33-day period in DNA extracted from whole blood. In vitro data indicated feasibility of IGF1 and EPO detection by duplex ddPCR with high reliability and sensitivity. On-column DNA-digestion allowed for significantly improved target detection in downstream PCR-based approaches. As ddPCR provides absolute quantification, it ensures excellent day-to-day reproducibility. Therefore, we expect this technique to be used in diagnosing and monitoring of viral and bacterial infection, in detecting mutated DNA sequences as well as profiling for the presence of foreign genetic material in elite athletes in the future. PMID:25375130

Moser, Dirk A.; Braga, Luca; Raso, Andrea; Zacchigna, Serena; Giacca, Mauro; Simon, Perikles

2014-01-01

218

Characterization of a Maize Wip1 Promoter in Transgenic Plants  

Directory of Open Access Journals (Sweden)

Full Text Available The Maize Wip1 gene encodes a wound-induced Bowman-Birk inhibitor (BBI protein which is a type of serine protease inhibitor, and its expression is induced by wounding or infection, conferring resistance against pathogens and pests. In this study, the maize Wip1 promoter was isolated and its function was analyzed. Different truncated Wip1 promoters were fused upstream of the GUS reporter gene and transformed into Arabidopsis, tobacco and rice plants. We found that (1 several truncated maize Wip1 promoters led to strong GUS activities in both transgenic Arabidopsis and tobacco leaves, whereas low GUS activity was detected in transgenic rice leaves; (2 the Wip1 promoter was not wound-induced in transgenic tobacco leaves, but was induced by wounding in transgenic rice leaves; (3 the truncated Wip1 promoter had different activity in different organs of transgenic tobacco plants; (4 the transgenic plant leaves containing different truncated Wip1 promoters had low GUS transcripts, even though high GUS protein level and GUS activities were observed; (5 there was one transcription start site of Wip1 gene in maize and two transcription start sites of GUS in Wip1::GUS transgenic lines; (6 the adjacent 35S promoter which is present in the transformation vectors enhanced the activity of the truncated Wip1 promoters in transgenic tobacco leaves, but did not influence the disability of truncated Wip1231 promoter to respond to wounding signals. We speculate that an ACAAAA hexamer, several CAA trimers and several elements similar to ACAATTAC octamer in the 5'-untranslated region might contribute to the strong GUS activity in Wip1231 transgenic lines, meanwhile, compared to the 5'-untranslated region from Wip1231 transgenic lines, the additional upstream open reading frames (uORFs in the 5'-untranslated region from Wip1737 transgenic lines might contribute to the lower level of GUS transcript and GUS activity.

Shengxue Zhang

2013-12-01

219

Comparative characterization of mesenchymal stem cells from eGFP transgenic and non-transgenic mice  

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Full Text Available Abstract Background Adipose derived- and bone marrow-derived murine mesenchymal stem cells (mMSCs may be used to study stem cell properties in an in vivo setting for the purposes of evaluating therapeutic strategies that may have clinical applications in the future. If these cells are to be used for transplantation, the question arises of how to track the administered cells. One solution to this problem is to transplant cells with an easily identifiable genetic marker such as enhanced green fluorescent protein (eGFP. This protein is fluorescent and therefore does not require a chemical substrate for identification and can be visualized in living cells. This study seeks to characterize and compare adipose derived- and bone marrow-derived stem cells from C57Bl/6 mice and eGFP transgenic C57Bl/6 mice. Results The expression of eGFP does not appear to affect the ability to differentiate along adipogenic or osteogenic lineages; however it appears that the tissue of origin can influence differentiation capabilities. The presence of eGFP had no effect on cell surface marker expression, and mMSCs derived from both bone marrow and adipose tissue had similar surface marker profiles. There were no significant differences between transgenic and non-transgenic mMSCs. Conclusion Murine adipose derived and bone marrow derived mesenchymal stem cells from non-transgenic and eGFP transgenic C57Bl/6 mice have very similar characterization profiles. The availability of mesenchymal stem cells stably expressing a genetic reporter has important applications for the advancement of stem cell research.

Bunnell Bruce A

2009-01-01

220

Expression systems and species used for transgenic animal bioreactors.  

Science.gov (United States)

Transgenic animal bioreactors can produce therapeutic proteins with high value for pharmaceutical use. In this paper, we compared different systems capable of producing therapeutic proteins (bacteria, mammalian cells, transgenic plants, and transgenic animals) and found that transgenic animals were potentially ideal bioreactors for the synthesis of pharmaceutical protein complexes. Compared with other transgenic animal expression systems (egg white, blood, urine, seminal plasma, and silkworm cocoon), the mammary glands of transgenic animals have enormous potential. Compared with other mammalian species (pig, goat, sheep, and cow) that are currently being studied as bioreactors, rabbits offer many advantages: high fertility, easy generation of transgenic founders and offspring, insensitivity to prion diseases, relatively high milk production, and no transmission of severe diseases to humans. Noticeably, for a small- or medium-sized facility, the rabbit system is ideal to produce up to 50 kg of protein per year, considering both economical and hygienic aspects; rabbits are attractive candidates for the mammary-gland-specific expression of recombinant proteins. We also reviewed recombinant proteins that have been produced by targeted expression in the mammary glands of rabbits and discussed the limitations of transgenic animal bioreactors. PMID:23586046

Wang, Yanli; Zhao, Sihai; Bai, Liang; Fan, Jianglin; Liu, Enqi

2013-01-01

 
 
 
 
221

Transgenes sustain epigeal insect biodiversity in diversified vegetable farm systems.  

Science.gov (United States)

Many ecological studies have focused on the effects of transgenes in field crops, but few have considered multiple transgenes in diversified vegetable systems. We compared the epigeal, or soil surface-dwelling, communities of Coleoptera and Formicidae between transgenic and isoline vegetable systems consisting of sweet corn, potato, and acorn squash, with transgenic cultivars expressing Cry1(A)b, Cry3, or viral coat proteins. Vegetables were grown in replicated split plots over 2 yr with integrated pest management (IPM) standards defining insecticide use patterns. More than 77.6% of 11,925 insects from 1,512 pitfall traps were identified to species, and activity density was used to compare dominance distribution, species richness, and community composition. Measures of epigeal biodiversity were always equal in transgenic vegetables, which required fewer insecticide applications than their near isolines. There were no differences in species richness between transgenic and isoline treatments at the farm system and individual crop level. Dominance distributions were also similar between transgenic and isoline farming systems. Crop type, and not genotype, had a significant influence on Carabidae and Staphylinidae community composition in the first year, but there were no treatment effects in the second year, possibly because of homogenizing effects of crop rotations. Communities were more influenced by crop type, and possibly crop rotation, than by genotype. The heterogeneity of crops and rotations in diversified vegetable farms seems to aid in preserving epigeal biodiversity, which may be supplemented by reductions in insecticide use associated with transgenic cultivars. PMID:17349138

Leslie, T W; Hoheisel, G A; Biddinger, D J; Rohr, J R; Fleischer, S J

2007-02-01

222

Rheumatic manifestations of inflammatory bowel disease  

Directory of Open Access Journals (Sweden)

Full Text Available This article reviews the literature concerning rheumatic manifestations of inflammatory bowel disease (IBD, including common immune-mediated pathways, frequency, clinical course and therapy. Musculoskeletal complications are frequent and well-recognized manifestations in IBD, and affect up to 33% of patients with IBD. The strong link between the bowel and the osteo-articular system is suggested by many clinical and experimental observations, notably in HLA-B27 transgenic rats. The autoimmune pathogenic mechanisms shared by IBD and spondyloarthropathies include genetic susceptibility to abnormal antigen presentation, aberrant recognition of self, the presence of autoantibodies against specific antigens shared by the colon and other extra-colonic tissues, and increased intestinal permeability. The response against microorganisms may have an important role through molecular mimicry and other mechanisms. Rheumatic manifestations of IBD have been divided into peripheral arthritis, and axial involvement, including sacroiliitis, with or without spondylitis, similar to idiopathic ankylosing spondylitis. Other periarticular features can occur, including enthesopathy, tendonitis, clubbing, periostitis, and granulomatous lesions of joints and bones. Osteoporosis and osteomalacia secondary to IBD and iatrogenic complications can also occur. The management of the rheumatic manifestations of IBD consists of physical therapy in combination with local injection of corticosteroids and nonsteroidal anti-inflammatory drugs; caution is in order however, because of their possible harmful effects on intestinal integrity, permeability, and even on gut inflammation. Sulfasalazine, methotrexate, azathioprine, cyclosporine and leflunomide should be used for selected indications. In some cases, tumor necrosis factor-? blocking agents should be considered as first-line therapy.

Tatiana Sofía Rodríguez-Reyna, Cynthia Martínez-Reyes, Jesús Kazúo Yamamoto-Furusho

2009-11-01

223

[Biofuels, food security and transgenic crops].  

Science.gov (United States)

Soaring global food prices are threatening to push more poor people back below the poverty line; this will probably become aggravated by the serious challenge that increasing population and climate changes are posing for food security. There is growing evidence that human activities involving fossil fuel consumption and land use are contributing to greenhouse gas emissions and consequently changing the climate worldwide. The finite nature of fossil fuel reserves is causing concern about energy security and there is a growing interest in the use of renewable energy sources such as biofuels. There is growing concern regarding the fact that biofuels are currently produced from food crops, thereby leading to an undesirable competition for their use as food and feed. Nevertheless, biofuels can be produced from other feedstocks such as lingo-cellulose from perennial grasses, forestry and vegetable waste. Biofuel energy content should not be exceeded by that of the fossil fuel invested in its production to ensure that it is energetically sustainable; however, biofuels must also be economically competitive and environmentally acceptable. Climate change and biofuels are challenging FAO efforts aimed at eradicating hunger worldwide by the next decade. Given that current crops used in biofuel production have not been domesticated for this purpose, transgenic technology can offer an enormous contribution towards improving biofuel crops' environmental and economic performance. The present paper critically presents some relevant relationships between biofuels, food security and transgenic plant technology. PMID:19722000

Acosta, Orlando; Chaparro-Giraldo, Alejandro

2009-01-01

224

[Potential ecological risks of transgenic trees].  

Science.gov (United States)

A new approach to genetic improvement of trees has been introduced with the birth of gene engineering technique. Compared to that in crops, gene introduction in trees has bigger potential ecological risk in environmental release and extension, because trees, most of which are wind-dispersed, grow on various habitats, have longer life span, and subject to relatively more extensive management. Extensive plantation of transgenic trees may reduce the genetic diversity of the trees concerned. Co-evolution of pests and pathogens is likely to be caused under the pressure of long-term and continuous selection of the trees derived from gene transferring. Escaping of exogenous gene may have a certain kind of influence on fitness of plants naturally generated, and as a result, have influence on species diversity in the natural world. It is not reasonable for China, a developing country, to reject gene introduction as an approach to promote forestry development. It is also important, on the other hand, to take future ecological safety into consideration because it is unwise to get present profit at the cost of future profit. To strengthen basic study on gene transferring, adopting safe management of varieties generated from gene transferring and increasing funds on research and management of transgenic trees are believed to be measures to decrease, to the greatest extent, ecological risks brought about by gene transferring of trees, and to quicken transformation of products of trees derived from gene-transferring into merchandises. PMID:15506115

Kang, Xiangyang; Liu, Zhimin; Li, Shenggong

2004-07-01

225

Transgenic resistance to tobacco ringspot virus.  

Science.gov (United States)

The coat protein (CP) gene including the 3'-untranslated region (UTR) of RNA2 of a cherry isolate of Tobacco ringspot virus (TRSV) was utilized in a CP-mediated resistance (CP-MR) strategy. To facilitate construction of plant expression vectors the sequence context of the CP gene translation initiation codon was modified at the 5'-end of the coding sequence by including an initiation codon. The gene was ligated to a version of the Cauliflower mosaic virus (CaMV) 35S promoter with a duplicated enhancer. The cloned CP gene was used to transform Nicotiana tabacum cv. Xanthi, as a systemic and local lesion host. The transgenic plants showed different level of resistance ranging from complete resistance to reduction in symptom severity post inoculation with the cherry isolate of TRSV. A CP gene transcript was detected in different tissue of transgenic lines, but translation product was undetectable by Western blot analysis or enzyme-linked immunosorbent assay (ELISA). Interestingly, 100% of seed transmission was blocked in a resistant line, which offers important prospects for engineering TRSV into economically important crops as soybean with 100% seed transmission. PMID:15595207

Zadeh, A Hamdollah; Foster, G D

2004-01-01

226

Advancing environmental risk assessment for transgenic biofeedstock crops  

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Full Text Available Abstract Transgenic modification of plants is a key enabling technology for developing sustainable biofeedstocks for biofuels production. Regulatory decisions and the wider acceptance and development of transgenic biofeedstock crops are considered from the context of science-based risk assessment. The risk assessment paradigm for transgenic biofeedstock crops is fundamentally no different from that of current generation transgenic crops, except that the focus of the assessment must consider the unique attributes of a given biofeedstock crop and its environmental release. For currently envisioned biofeedstock crops, particular emphasis in risk assessment will be given to characterization of altered metabolic profiles and their implications relative to non-target environmental effects and food safety; weediness and invasiveness when plants are modified for abiotic stress tolerance or are domesticated; and aggregate risk when plants are platforms for multi-product production. Robust risk assessments for transgenic biofeedstock crops are case-specific, initiated through problem formulation, and use tiered approaches for risk characterization.

Wolt Jeffrey D

2009-11-01

227

Illegal gene flow from transgenic creeping bentgrass: the saga continues.  

Science.gov (United States)

Ecologists have paid close attention to environmental effects that fitness-enhancing transgenes might have following crop-to-wild gene flow (e.g. Snow et al. 2003). For some crops, gene flow also can lead to legal problems,especially when government agencies have not approved transgenic events for unrestricted environmental release.Creeping bentgrass (Agrostis stolonifera), a common turf grass used in golf courses, is the focus of both areas of concern. In 2002, prior to expected deregulation (still pending), The Scotts Company planted creeping bentgrass with transgenic resistance to the herbicide glyphosate,also known as RoundUp, on 162 ha in a designated control area in central Oregon (Fig. 1).Despite efforts to restrict gene flow, wind-dispersed pollen carried transgenes to florets of local A. stolonifera and A. gigantea as far as 14 km away, and to sentinel plants placed as far as 21 km away (Watrud et al. 2004).Then, in August 2003, a strong wind event moved transgenic seeds from wind rows of cut bentgrass into nearby areas. The company’s efforts to kill all transgenic survivors in the area failed: feral glyphosate-resistant populations of A. stolonifera were found by Reichman et al.(2006), and 62% of 585 bentgrass plants had the telltale CP4 EPSPS transgene in 2006 (Zapiola et al. 2008; Fig. 2).Now, in this issue, the story gets even more interesting as Zapiola & Mallory-Smith (2012) describe a transgenic,intergeneric hybrid produced on a feral, transgenic creeping bentgrass plant that received pollen from Polypogon monspeliensis (rabbitfoot grass). Their finding raises a host of new questions about the prevalence and fitness of intergeneric hybrids, as well as how to evaluate the full extent of gene flow from transgenic crops. PMID:23009646

Snow, Allison A

2012-10-01

228

Inheritance of transgenes in transgenic Bt lines resistance to Helicoerpa armigera in upland cotton.  

Science.gov (United States)

Six transgenic Bt cotton cultivars (lines) including GKsu12, GK19, MR1, GK5, 109B, and SGK1 are highly resistant to bollworm from the seedling to boll-setting stages in bioassays with detached cotton leaves, though there are differences in resistant level and Bt toxin content in these transgenic cottons. Genetics analysis reveals that the resistance to Helicoverpa armigera in these six transgenic Bt cotton cultivars (lines) are controlled by one pair of dominant genes. Allelic tests further demonstrate some populations are in Mendel segregation for two nonallelic genes, i.e., the inserted Bt gene in GKsu12 is nonallelic to that of SGK1, GK5, 109B, and GK19 and Bt genes in GK19 and SGK1 are likely inserted in the same or in close proximity (genetically closely linked), while some F(2) produce abnormal segregation patterns, with a segregation of resistance to Helicoerpa armigera vary between 15:1 and 3:1, though their Bt segregation fit into 15:1 by PCR analysis, suggesting Bt gene silence in these populations. Two genes silence may occur in these populations due to the homologous sequence by crossing since the silenced individuals accounted for 1/16 of the F(2) populations for allelic test. To those silenced populations, one of their parents all showed high resistance to bollworm. PMID:23143492

Zhang, Baolong; Guo, Wangzhen; Zhang, Tianzhen

2013-01-01

229

Evaluating potential risks of transgenic arthropods for pest management programmes  

International Nuclear Information System (INIS)

Genetic modification using recombinant DNA methods can now be used, almost routinely, to transform pest and beneficial arthropods and such genetically engineered insects and mites could be used to improve pest management programs. Genetic manipulation with recombinant DNA techniques may generate concerns about risk, requiring additional time and resources to resolve. Risk assessments must be conducted prior to releasing transgenic arthropods into the environment for either short term experiments or permanent establishment. Potential risk issues to be resolved include whether: the inserted gene(s) (trait) is stable; the traits can be horizontally transferred to other populations or species; released arthropods will perform as expected (especially with regard to their geographic distribution, host or prey specificity; released arthropods will have unintended environmental effects; and, in the case of short term releases, the released arthropods can be recovered from field sites. If the transgenic arthropods strain(s) perform well in preliminary, short term releases and risk assessments are completed satisfactorily, permanent releases into the environment may follow. Many pest management programs, especially those involving replacement of pest populations by the transgenic population, will require permanent establishment in the environment and the use of 'drive mechanisms', have been proposed to achieve this. Because efficacy can be severely compromised by 'transgene sil be severely compromised by 'transgene silencing', plant molecular biologists are now attempting to stabilize gene expression by building in 'insulators'. Transgene silencing occurs in Drosophila and will no doubt be a factor in other transgenic arthropods. (author)

230

Welfare assessment in transgenic pigs expressing green fluorescent protein (GFP)  

DEFF Research Database (Denmark)

Since large animal transgenesis has been successfully attempted for the first time about 25 years ago, the technology has been applied in various lines of transgenic pigs. Nevertheless one of the concerns with the technology—animal welfare—has not been approached through systematic assessment and statements regarding the welfare of transgenic pigs have been based on anecdotal observations during early stages of transgenic programs. The main aim of the present study was therefore to perform an extensive welfare assessment comparing heterozygous transgenic animals expressing GFP with wildtype animals along various stages of post natal development. The protocol used covered reproductory performance and behaviour in GFP and wildtype sows and general health and development, social behaviour, exploratory behaviour and emotionality in GFP and wildtype littermates from birth until an age of roughly 4 months. The absence of significant differences between GFP and wildtype animals in the parameters observed suggests that the transgenic animals in question are unlikely to suffer from deleterious effects of transgene expression on their welfare and thus support existing anecdotal observations of pigs expressing GFP as healthy. Although the results are not surprising in the light of previous experience, they give a more solid fundament to the evaluation of GFP expression as being relatively non-invasive in pigs. The present study may furthermore serve as starting point for researchers aiming at a systematic characterization of welfare relevant effects in the line of transgenic pigs they are working with.

Huber, Reinhard C.; Remuge, Liliana

2012-01-01

231

Ultrastructure of vitrified rabbit transgenic embryos.  

Science.gov (United States)

The aim of the study was to determine the viability of rabbit transgenic (enhanced green fluorescent protein (EGFP)-positive) embryos cultured in vitro and compare with gene-microinjected (Mi) non-transgenic (EGFP-negative) embryos following vitrification. Non-microinjected and non-vitrified embryos were used as the control. Morphological signs of injury to embryo organelles were determined at the ultrastructural level using transmission electron microscopy (TEM). Morphometric evaluation was performed on cellular organelles using microphotographs obtained by TEM. Intact and Mi embryos recovered from in vivo fertilized eggs at 19-20 hours post coitum (hpc) were cultured for up to 72 hpc (morula stage), evaluated for the EGFP gene integration and then vitrified in 0.25 ml insemination straws in modified EFS (40% ethylene glycol + 18% Ficoll 70 + 0.3 M sucrose) vitrification solution. After 1-3 days the embryos were devitrified, a representative selection of embryos was analyzed by TEM and the remaining embryos were subjected to additional in vitro culture. Observations by TEM showed that the vitrified/warmed EGFP-positive and EGFP-negative embryos had a slight accumulation of cellular debris and lipid droplets compared with the control intact embryos. More severe changes were detected in the membrane structures of the treated embryos, mostly in the cytoplasmic envelope, trophoblastic microvilli, junctional contacts and mitochondria. We suggest that the higher proportion of deteriorated cell structures and organelles in the treated embryos may be due to the vitrification process rather than to mechanical violation (the gene-microinjection procedure), as a detailed inspection of ultrastructure revealed that most damage occurred in the cell membrane structures. PMID:24152610

Chrenek, P; Makarevich, A V; Popelková, M; Schlarmannová, J; Toporcerová, S; Ostró, A; Ziv?ák, J; Bosze, Zs

2014-11-01

232

Genetic load and transgenic mitigating genes in transgenic Brassica rapa (field mustard × Brassica napus (oilseed rape hybrid populations  

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Full Text Available Abstract Background One theoretical explanation for the relatively poor performance of Brassica rapa (weed × Brassica napus (crop transgenic hybrids suggests that hybridization imparts a negative genetic load. Consequently, in hybrids genetic load could overshadow any benefits of fitness enhancing transgenes and become the limiting factor in transgenic hybrid persistence. Two types of genetic load were analyzed in this study: random/linkage-derived genetic load, and directly incorporated genetic load using a transgenic mitigation (TM strategy. In order to measure the effects of random genetic load, hybrid productivity (seed yield and biomass was correlated with crop- and weed-specific AFLP genomic markers. This portion of the study was designed to answer whether or not weed × transgenic crop hybrids possessing more crop genes were less competitive than hybrids containing fewer crop genes. The effects of directly incorporated genetic load (TM were analyzed through transgene persistence data. TM strategies are proposed to decrease transgene persistence if gene flow and subsequent transgene introgression to a wild host were to occur. Results In the absence of interspecific competition, transgenic weed × crop hybrids benefited from having more crop-specific alleles. There was a positive correlation between performance and number of B. napus crop-specific AFLP markers [seed yield vs. marker number (r = 0.54, P = 0.0003 and vegetative dry biomass vs. marker number (r = 0.44, P = 0.005]. However under interspecific competition with wheat or more weed-like conditions (i.e. representing a situation where hybrid plants emerge as volunteer weeds in subsequent cropping systems, there was a positive correlation between the number of B. rapa weed-specific AFLP markers and seed yield (r = 0.70, P = 0.0001, although no such correlation was detected for vegetative biomass. When genetic load was directly incorporated into the hybrid genome, by inserting a fitness-mitigating dwarfing gene that that is beneficial for crops but deleterious for weeds (a transgene mitigation measure, there was a dramatic decrease in the number of transgenic hybrid progeny persisting in the population. Conclusion The effects of genetic load of crop and in some situations, weed alleles might be beneficial under certain environmental conditions. However, when genetic load was directly incorporated into transgenic events, e.g., using a TM construct, the number of transgenic hybrids and persistence in weedy genomic backgrounds was significantly decreased.

Warwick Suzanne I

2009-10-01

233

[Effects of phytase transgenic corn planting on soil nematode community].  

Science.gov (United States)

A healthy soil ecosystem is essential for nutrient cycling and energy conversion, and the impact of exogenous genes from genetically modified crops had aroused wide concerns. Phytase transgenic corn (i. e., the inbred line BVLA430101) was issued a bio-safety certificate on 27 September 2009 in China, which could improve the efficiency of feed utilization, reduce environmental pollution caused by animal manure. In this study, the abundance of trophic groups, community structure and ecological indices of soil nematodes were studied over the growing cycle of phytase transgenic corn (ab. transgenic corn) and control conventional parental corn (ab. control corn) in the field. Totally 29 and 26 nematode genera were isolated from transgenic corn and control corn fields, respectively. The abundances of bacterivores and omnivores-predators, the total number of soil nematodes, and the Shannon index (H) were significantly greater under transgenic corn than under control corn, while the opposite trend was found for the relative abundance of herbivores and the maturity index (Sigma MI) of soil nematodes. Repeated-measures analysis of variance (ANOVA) did not detect any significant effects of transgenic corn on the composition and abundance of nematode trophic groups and ecological indices of soil nematodes. Furthermore, the Student-T test showed that the abundances of bacterivores and omnivores-predators and the total number of soil nematodes during the milk-ripe stage were significant higher in the transgenic corn field than in the control corn field. The effects of transgenic corn planting on soil nematodes might be related to the increase in the nitrogen content of field soil under transgenic corn compared to control corn. PMID:25011306

Zhao, Zong-Chao; Su, Ying; Mou, Wen-Ya; Liu, Man-Qiang; Chen, Xiao-Yun; Chen, Fa-Jun

2014-04-01

234

Identification of the Offspring of Vegfr2-luc Transgenic Mouse  

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Full Text Available Background and objective A transgenic mouse, Vegfr2-luc, in which a luciferase reporter (luc is under control of the murine VEGFR2 promoter, can be used to track angiogenesis in vivo. The aim of this study is to identify the offspring of Vegfr2-luc transgenic mouse. Methods Luc was detected with PCR in genomic DNA of the new-born mouse. Luc expression in the offspring of Vegfr2-luc transgenic mouse was monitored with IVIS in vivo imaging system during post-natal development. Wound-healing models of Vegfr2-luc transgenic mouse offspring were established and the expression of luc was monitored during the wound-healing process. Luc activity and VEGFR2 mRNA expression in different organs were detected with luc Assay System and Real-time PCR respectively. Results PCR showed that 50% (56/112 of the offspring of Vegfr2-luc transgenic mouse carry luc. IVIS in vivo imaging results demonstrated that luc expression in Vegfr2-luc transgenic mouse dropped dramatically with age increase (P<0.001 and luc expression in the wound first increased and then decreased during the wound-healing process (P<0.001. Luc activity in female Vegfr2-luc transgenic mouse organs was positively correlated with VEGFR2 mRNA expression (r=0.948, P<0.001. Except testis, luc activity in male Vegfr2-luc transgenic mouse organs was also positively correlated with VEGFR2 mRNA expression (r=0.836, P<0.001. Conclusion The offspring of Vegfr2-luc transgenic mouse is applicable to tracking angiogenesis in vivo.

Qinghua ZHOU

2011-05-01

235

Truncated beta-lactoglobulin transgenes are expressed in the kidney.  

Science.gov (United States)

The major milk whey protein of ruminants is beta-lactoglobulin. Ovine beta-lactoglobulin-encoding gene expression is restricted to the sheep mammary gland. This report describes the expression profile of a truncated beta-lactoglobulin transgene which, although not expressed in the mammary gland, is expressed in the kidney in the majority of lines generated. The high frequency of ectopic kidney expression may relate to the ability of the larger beta-lactoglobulin transgenes to be expressed in a position-independent manner in the mammary gland of transgenic mice. PMID:8921908

Whitelaw, C B

1996-10-31

236

Transgenic Rice Expressing Amyloid ?-peptide for Oral Immunization  

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Full Text Available Various vaccine therapies for Alzheimer's disease (AD have been investigated. Here we report transgenic rice expressing amyloid ?-peptide (A?. The A?42 gene fused with a green fluorescent protein gene was introduced into rice using the Agrobacterium method. When transgenic brown rice expressing A? was orally administered to mice, serum anti-A? antibody titers were elevated. The same results were observed when mice were fed boiled, transgenic brown rice. The results indicate that an edible vaccine against AD using rice may be feasible. A vaccine derived from rice would be far cheaper than existing medical vaccines.

Taiji Yoshida, Eiichi Kimura, Setsuo Koike, Jun Nojima, Eugene Futai, Noboru Sasagawa, Yuichiro Watanabe, Shoichi Ishiura

2011-01-01

237

Localization of a position effect element that affects alcohol dehydrogenase transgene expression in Drosophila melanogaster.  

Science.gov (United States)

The molecular basis for the abnormal expression of an alcohol dehydrogenase (Adh) transgene, introduced into the 25 C-D region of an Adh negative strain of Drosophila melanogaster, was investigated using a transient expression assay. A 14-kb genomic clone from the transformed line, MM-50, containing the transgene was isolated. A position effect element was identified upstream of the Adh gene within a 1.7-kb EcoRI fragment. This element acts as a silencer, greatly reducing the Adh transcriptional activity. Sequence analysis reveals that the silencer element is 1.18 kb long. There are no long open reading frames present in the sequence, suggesting that it is derived from a noncoding region. A 20-bp sequence (17/20 nucleotides matching) containing a high T/C content is repeated within the silencer region. Within each of these homologous regions there is an internal repeat of the sequence CTCTC. The repeated sequences in MM-50 contain a consensus motif, CCTCTC, for silencer elements found in the rat collagen II gene and several other vertebrate genes, including the human beta-interferon (beta-IFN) gene. Comparison of the sequence of the silencer region to the D. melanogaster whole genome sequence shows that the silencer is located within a clone derived from the 25C5-25C10 region of chromosome 2L. PMID:12162808

Castronuevo, Patria; Martin, Presley F

2002-07-01

238

Polycythemia in transgenic mice expressing the human erythropoietin gene  

International Nuclear Information System (INIS)

Erythropoietin is a glycoprotein hormone that regulates mammalian erythropoiesis. To study the expression of the human erythropoietin gene, EPO, 4 kilobases of DNA encompassing the gene with 0.4 kilobase of 5' flanking sequence and 0.7 kilobase of 3' flanking sequence was microinjected into fertilized mouse eggs. Transgenic mice were generated that are polycythemic, with increased erythrocytic indices in peripheral blood, increased numbers of erythroid precursors in hematopoietic tissue, and increased serum erythropoietin levels. Transgenic homozygotes show a greater degree of polycythemia than do heterozygotes as well as striking extramedullary erythropoiesis. Human erythropoietin RNA was found not only in fetal liver, adult liver, and kidney but also in all other transgenic tissues analyzed. Anemia induced increased human erythropoietin RNA levels in liver but not kidney. These transgenic mice represent a unique model of polycythemia due to increased erythropoietin levels

239

Areas of concern for the evaluation of transgenic arthropods  

International Nuclear Information System (INIS)

e environment and other organisms should the transgene be transmitted to another host. These factors must be considered individually, their interaction with one another, and also in the context of transformant strain persistence in the field. (author)

240

Designer proton-channel transgenic algae for photobiological hydrogen production  

Science.gov (United States)

A designer proton-channel transgenic alga for photobiological hydrogen production that is specifically designed for production of molecular hydrogen (H.sub.2) through photosynthetic water splitting. The designer transgenic alga includes proton-conductive channels that are expressed to produce such uncoupler proteins in an amount sufficient to increase the algal H.sub.2 productivity. In one embodiment the designer proton-channel transgene is a nucleic acid construct (300) including a PCR forward primer (302), an externally inducible promoter (304), a transit targeting sequence (306), a designer proton-channel encoding sequence (308), a transcription and translation terminator (310), and a PCR reverse primer (312). In various embodiments, the designer proton-channel transgenic algae are used with a gas-separation system (500) and a gas-products-separation and utilization system (600) for photobiological H.sub.2 production.

Lee, James Weifu (Knoxville, TN)

2011-04-26

 
 
 
 
241

Hybridization between transgenic and wild plants: environmental risk  

Directory of Open Access Journals (Sweden)

Full Text Available Genetically modified products are widely commercialized in agricultural production. These include resistant plants to diseases, insects or herbicides, plants with capacity for longer storing times or better nutritional quality. However, there are some concerns and critics from environmental organizations on the risk associated to transgenic plants or organisms genetically modified (OGM. This review discusses the vertical gene transfer (plant/Plant within the OGM context. Although transgenic hybrids have been reported between transgenic plants and their wild relatives, the extent of the environmental risk has not been evaluated per se. The risk depends on the plant species involved, the transgenes, and the ecosystem where the plants are located. Studies on biosafety assessment must be evaluated case by case. Biotechnology and conventional methods allow to control gen flow and decrease the risk of gene transfer among species.

Alejandro Chaparro Giraldo

2011-12-01

242

Enhanced Malignant Tumorigenesis in Cdk4-Transgenic Mice  

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In a previous study, we reported that overexpression of CDK4 in mouse epidermis results in epidermal hyperplasia, hypertrophy and severe dermal fibrosis. In this study, we have investigated the susceptibility to skin tumor formation by forced expression of CDK4. Skin tumors from transgenic mice showed a dramatic increase in the rate of malignant progression to squamous cell carcinomas (SCC) in an initiation-promotion protocol. Histopathological analysis of papillomas from transgenic mice show...

Miliani Marval, Paula L.; Macias, Everardo; Conti, Claudio J.; Rodriguez-puebla, Marcelo L.

2004-01-01

243

Experimental Meningococcal Sepsis in Congenic Transgenic Mice Expressing Human Transferrin  

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Severe meningococcal sepsis is still of high morbidity and mortality. Its management may be improved by an experimental model allowing better understanding of its pathophysiology. We developed an animal model of meningococcal sepsis in transgenic BALB/c mice expressing human transferrin. We studied experimental meningococcal sepsis in congenic transgenic BALB/c mice expressing human transferrin by transcriptional profiling using microarray analysis of blood and brain samples. Genes encoding a...

Szatanik, Marek; Hong, Eva; Ruckly, Corinne; Ledroit, Morgan; Giorgini, Dario; Jopek, Katarzyna; Nicola, Marie-anne; Deghmane, Ala-eddine; Taha, Muhamed-kheir

2011-01-01

244

Properties of beta-propeller phytase expressed in transgenic tobacco  

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Phytases are enzymes that liberate inorganic phosphates from phytate. In a previous study, a beta-propeller phytase (168phyA) from Bacillus subtilis was introduced into transgenic tobacco, which resulted in certain phenotypic changes. In the study described herein, the recombinant phytase (t168phyA) was purified from transgenic tobacco to near homogeneity by a three-step purification scheme. The biochemical properties and kinetic parameters of t168phyA were compared with those of its counterp...

Lim, Bl; Chan, Wl; Lung, Sc

2006-01-01

245

Generation of tissue-specific transgenic birds with lentiviral vectors  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Birds are of great interest for a variety of research purposes, and effective methods for manipulating the avian genome would greatly accelerate progress in fields that rely on birds as model systems for biological research, such as developmental biology and behavioral neurobiology. Here, we describe a simple and effective method for producing transgenic birds. We used lentiviral vectors to produce transgenic quails that express GFP driven by the human synapsin gene I promoter. Expression of ...

Scott, Benjamin B.; Lois, Carlos

2005-01-01

246

Enhanced Malignant Tumorigenesis in Cdk4-Transgenic Mice  

Science.gov (United States)

In a previous study, we reported that overexpression of CDK4 in mouse epidermis results in epidermal hyperplasia, hypertrophy and severe dermal fibrosis. In this study, we have investigated the susceptibility to skin tumor formation by forced expression of CDK4. Skin tumors from transgenic mice showed a dramatic increase in the rate of malignant progression to squamous cell carcinomas (SCC) in an initiation-promotion protocol. Histopathological analysis of papillomas from transgenic mice showed an elevated number of premalignant lesions characterized by dysplasia and marked atypia. Interestingly, transgenic mice also developed tumors in initiated but not promoted skin, demonstrating that CDK4 replaced the action of tumor promoters. These results suggest that expression of cyclin D1 upon ras activation synergizes with CDK4 overexpression. However, cyclin D1 transgenic mice and double transgenic mice for cyclin D1 and CDK4 did not show increased malignant progression in comparison to CDK4 transgenic mice. Biochemical analysis of tumors showed that CDK4 sequesters the CDK2 inhibitors p27Kip1 and p21Cip1 suggesting that indirect activation of CDK2 plays an important role in tumor development. These results indicate that, contrary to the general assumption, the catalytic subunit, CDK4, has higher oncogenic activity than cyclin D1, revealing a potential use of CDK4 as therapeutic target. PMID:14647432

Miliani de Marval, Paula L.; Macias, Everardo; Conti, Claudio J.; Rodriguez-Puebla, Marcelo L.

2010-01-01

247

Transgene rescue in the mammary gland is associated with transcription but does not require translation of BLG transgenes.  

Science.gov (United States)

Many transgenes, particularly those comprising cDNA sequences fail to be expressed when they are introduced into transgenic mice. We have previously shown that this problem can be overcome in the mammary gland by co-integrating a poorly expressed cDNA transgene, comprising the sheep beta-lactoglobulin promoter, with the efficiently expressed, unmodified beta-lactoglobulin gene. In this report we demonstrate that the transcription of the beta-lactoglobulin gene is associated with this effect because co-integration with a non-transcribed beta-lactoglobulin gene fails to rescue expression. By contrast, co-integration with a translationally inactivated beta-lactoglobulin transgene does rescue the expression of the second gene, but without the co-production of beta-lactoglobulin protein. PMID:9032973

Yull, F; Binas, B; Harold, G; Wallace, R; Clark, A J

1997-01-01

248

Transgenic approaches to a non-transgenic release of sterile male Lepidoptera  

International Nuclear Information System (INIS)

rge (3 pairs), large (3 pairs), medium (15 pairs), small (5 pairs), and dot-like (2 pairs). Females are heterogametic with a W-Z sex chromosome pair, males are homogametic with two Z chromosomes. The W and Z chromosomes represent the two largest elements in female chromosome complements. While the Z is composed of euchromatin and resembles to autosomes, the W consists largely of heterochromatin. For successful development of transgenic sexing strains in the codling moth, it is required to insert a conditional dominant lethal mutation (a transgene) into the W chromosome. Theoretically, the transgene insertion is a matter of probability, which is dependent on the size of the W relative to the rest of the genome. In the codling moth, the W is one of two largest chromosomes, comprising about 4% of the female genome, which should make it a good target for transgenesis with the probability of insertion of 1 in 25 (if only females are included) or with the overall probability of 1 in 50 (since both female and male embryos are exposed). However, since the W chromosome is mainly composed of heterochromatin, silencing of the transgene expression might be a serious problem. Different ways how to overcome this problem are discussed. For further characterisation of the codling moth W chromosome we employed advanced methods of molecular cytogenetics, genomic in situ hybridisation (GISH) and comparative genomic hybridisation (CGH). GISH detected the W chromosome by strong binding of the Cy3-labelled, female-derived DNA probe. With CGH, both the Cy3-labelled female-derived probe and Fluor-X labelled male-derived probe evenly bound to the W. This suggested that the W is composed predominantly of repetitive DNA sequences occurring scattered in other chromosomes but accumulated in the W. Finally, we prepared W-specific probes by laser microdissection of the W chromatin followed by DOP-PCR and PCR labelling. The probes stained the W with a high specificity in a chromosome-painting manner. DNA fragments of the microdissected W chromatin were cloned and sequenced. The W-sequence analysis revealed no homology to any DNA sequenced so far. Several cloned sequences were found to originate exclusively from the W chromosome. These unique sequences can be very useful as molecular markers of the W chromosome in codling moth transgenesis. The demonstrated ways of W chromosome identification will facilitate the development of genetic sexing strains in the codling moth

249

Enhanced neuroinflammation and pain hypersensitivity after peripheral nerve injury in rats expressing mutated superoxide dismutase 1  

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Full Text Available Abstract Background Neuroinflammation and nitroxidative stress are implicated in the pathophysiology of neuropathic pain. In view of both processes, microglial and astroglial activation in the spinal dorsal horn play a predominant role. The present study investigated the severity of neuropathic pain and the degree of glial activation in an inflammatory- and nitroxidative-prone animal model. Methods Transgenic rats expressing mutated superoxide dismutase 1 (hSOD1G93A are classically used as a model for amyotrophic lateral sclerosis (ALS. Because of the associated inflammatory- and nitroxidative-prone properties, this model was used to study thermal and mechanical hypersensitivity following partial sciatic nerve ligation (PSNL. Next to pain hypersensitivity assessment, microglial and astroglial activation states were moreover characterized, as well as inflammatory marker gene expression and the glutamate clearance system. Results PSNL induced thermal and mechanical hypersensitivity in both wild-type (WT and transgenic rats. However, the degree of thermal hypersensitivity was found to be exacerbated in transgenic rats while mechanical hypersensitivity was only slightly and not significantly increased. Microglial Iba1 expression was found to be increased in the ipsilateral dorsal horn of the lumbar spinal cord after PSNL but such Iba1 up-regulation was enhanced in transgenic rats as compared WT rats, both at 3 days and at 21 days after injury. Moreover, mRNA levels of Nox2, a key enzyme in microglial activation, but also of pro-inflammatory markers (IL-1? and TLR4 were not modified in WT ligated rats at 21 days after PSNL as compared to WT sham group while transgenic ligated rats showed up-regulated gene expression of these 3 targets. On the other hand, the PSNL-induced increase in GFAP immunoreactivity spreading that was evidenced in WT rats was unexpectedly found to be attenuated in transgenic ligated rats. Finally, GLT-1 gene expression and uptake activity were shown to be similar between WT sham and WT ligated rats at 21 days after injury, while both parameters were significantly increased in the ipsilateral dorsal region of the lumbar spinal cord of hSOD1G93A rats. Conclusions Taken together, our findings show that exacerbated microglial activation and subsequent inflammatory and nitroxidative processes are associated with the severity of neuropathic pain symptoms.

Lavand'homme Patricia

2011-04-01

250

Composite potato plants with transgenic roots on non-transgenic shoots: a model system for studying gene silencing in roots  

DEFF Research Database (Denmark)

Composite plants, with transgenic roots on a non-transgenic shoot, can be obtained by shoot explant transformation with Agrobacterium rhizogenes. The aim of this study was to generate composite potato plants (Solanum tuberosum) to be used as a model system in future studies on root-pathogen interactions and gene silencing in the roots. The proportion of transgenic roots among the roots induced was high (80-100 %) in the four potato cultivars tested (Albatros, Desirée, Sabina and Saturna). No wild-type adventitious roots were formed at mock inoculation site. All strains of A. rhizogenes tested induced phenotypically normal roots which, however, showed a reduced response to cytokinin as compared with non-transgenic roots. Nevertheless, both types of roots were infected to a similar high rate with the zoospores of Spongospora subterranea, a soilborne potato pathogen. The transgenic roots of composite potato plants expressed significantly higher amounts of ?-glucuronidase (GUS) than the roots of a GUS-transgenic potato line event. Silencing of the uidA transgene (GUS) was tested by inducing roots on the GUS-transgenic cv. Albatros event with strains of A. rhizogenes over-expressing either the uidA sense or antisense transcripts, or inverted-repeat or hairpin uidA RNA. The three last mentioned constructs caused 2.5-4.0 fold reduction in the uidA mRNA expression. In contrast, over-expression of uidA resulted in over 3-fold increase in the uidA mRNA and GUS expression, indicating that sense-mediated silencing (co-suppression) was not functional in roots. The results suggest that composite plants offer a useful experimental system for potato research, which has gained little previous attention.

Horn, Patricia; Santala, Johanna

2014-01-01

251

Superoxide dismutase activity in transgenic canola.  

Science.gov (United States)

Superoxide dismutase (SOD) activity was investigated in leaves of transgenic canola plants which expressed heterologous genes of different origin, namely 1) herbicide resistance genes (bar and simultaneously bar and epsps); 2) DesC desaturase gene (desC) of cyanobacterium Synechococcus vulcanus; 3) human interferon alpha2b gene (huLFN-alpha2b); 4) esxA::fbpB(deltaTMD) fused gene, encoding ESAT-6 and Ag85b Mycobacterium tuberculosis proteins, inducing immune response against tuberculosis; 5) cyp11A1 gene of cytochrome P450(scc) from bovine adrenal cortex mitochondria. Introduction of herbicide resistance genes as well as desaturase gene of cyanobacterium and mycobacterium's genes did not change leaf SOD activity. At the same time it was shown that cyp11A1 and huIFN-alpha2b canola have increased leaf SOD activity up 58 and 33%, respectively, compared with control ones in non-stress conditions. It may be a prerequisite for improved resistance of these plants to the stressors of different origin. PMID:25016823

Sakhno, L O; Slyvets, M S

2014-01-01

252

Culturable endophytic filamentous fungi from leaves of transgenic imidazolinone-tolerant sugarcane and its non-transgenic isolines.  

Science.gov (United States)

The diversity of endophytic filamentous fungi from leaves of transgenic imidazolinone-tolerant sugarcane plants and its isoline was evaluated by cultivation followed by amplified rDNA restriction analysis (ARDRA) of randomly selected strains. Transgenic and non-transgenic cultivars and their crop management (herbicide application or manual weed control) were used to assess the possible non-target effects of genetically modified sugarcane on the fungal endophytic community. A total of 14 ARDRA haplotypes were identified in the endophytic community of sugarcane. Internal transcribed spacer (ITS) sequencing revealed a rich community represented by 12 different families from the Ascomycota phylum. Some isolates had a high sequence similarity with genera that are common endophytes in tropical climates, such as Cladosporium, Epicoccum, Fusarium, Guignardia, Pestalotiopsis and Xylaria. Analysis of molecular variance indicated that fluctuations in fungal population were related to both transgenic plants and herbicide application. While herbicide applications quickly induced transient changes in the fungal community, transgenic plants induced slower changes that were maintained over time. These results represent the first draft on composition of endophytic filamentous fungi associated with sugarcane plants. They are an important step in understanding the possible effects of transgenic plants and their crop management on the fungal endophytic community. PMID:20191263

Stuart, Rodrigo Makowiecky; Romão, Aline Silva; Pizzirani-Kleiner, Aline Aparecida; Azevedo, João Lúcio; Araújo, Welington Luiz

2010-04-01

253

Divergent Phenotypes in Mutant TDP-43 Transgenic Mice Highlight Potential Confounds in TDP-43 Transgenic Modeling  

Science.gov (United States)

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

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

2014-01-01

254

Survival of Skin Graft between Transgenic Cloned Dogs and Non-Transgenic Cloned Dogs  

Science.gov (United States)

Whereas it has been assumed that genetically modified tissues or cells derived from somatic cell nuclear transfer (SCNT) should be accepted by a host of the same species, their immune compatibility has not been extensively explored. To identify acceptance of SCNT-derived cells or tissues, skin grafts were performed between cloned dogs that were identical except for their mitochondrial DNA (mtDNA) haplotypes and foreign gene. We showed here that differences in mtDNA haplotypes and genetic modification did not elicit immune responses in these dogs: 1) skin tissues from genetically-modified cloned dogs were successfully transplanted into genetically-modified cloned dogs with different mtDNA haplotype under three successive grafts over 63 days; and 2) non-transgenic cloned tissues were accepted into transgenic cloned syngeneic recipients with different mtDNA haplotypes and vice versa under two successive grafts over 63 days. In addition, expression of the inserted gene was maintained, being functional without eliciting graft rejection. In conclusion, these results show that transplanting genetically-modified tissues into normal, syngeneic or genetically-modified recipient dogs with different mtDNA haplotypes do not elicit skin graft rejection or affect expression of the inserted gene. Therefore, therapeutically valuable tissue derived from SCNT with genetic modification might be used safely in clinical applications for patients with diseased tissues. PMID:25372489

Kim, Geon A; Oh, Hyun Ju; Kim, Min Jung; Jo, Young Kwang; Choi, Jin; Park, Jung Eun; Park, Eun Jung; Lim, Sang Hyun; Yoon, Byung Il; Kang, Sung Keun; Jang, Goo; Lee, Byeong Chun

2014-01-01

255

Tissue folate binding protein levels in transgenic mice with tumors and in non-transgenic controls.  

Science.gov (United States)

Localized folate deficiency may be a risk factor for cancer. Since, folate binding proteins (FBP) and reduced folate carrier proteins (RFC) mediate cellular transport of folate, we compared FBP concentrations in several organs from tumor-bearing transgenic (TBT) mice and tumor-free non-transgenic controls (NTC) of the same strain, age, and fed identical diets. Liver, spleen, brain, small intestine and kidney were individually homogenized in phosphate-buffered saline (PBS) and separated into membrane, cytoplasmic, mitochondrial/lysomal and nuclear fractions (confirmed with marker enzymes). Homogenates and fractions was analyzed for total protein, and FBP. We used rabbit anti-bovine milk antibody and ELISA to measure FBP. FBP concentrations in kidney, small intestine, and spleen of TBT mice were higher than those of NTC mice; the opposite was true in liver and lung. FBP seemed to be upregulated in kidneys (all fractions), small intestine (all fractions), and spleen (cytoplasmic and nuclear fractions only) of TBT mice compared to NTC mice; the opposite appeared true in liver (all fractions) and lung (all fractions). FBP concentrations in brain, heart, and muscle of TBT mice were not different from those in brain, heart and muscle of NTC mice. A longitudinal study will determine if these changes in FBP concentrations precede tumor onset. PMID:10390055

Au, T V; So-Lui, M C; Zhang, Y; Arjomand, A; Lin, Y; Dueker, S R; Ho, Y K; Clifford, A J

1999-05-01

256

Adenosine (A)(2A)receptor modulation of nicotine-induced locomotor sensitization. A pharmacological and transgenic approach.  

Science.gov (United States)

Preclinical evidence indicates an important role of adenosine (A)(2A) receptors in drug addiction while their therapeutic relevance is still a matter of debate. We examined the influence of the A(2A) receptor agonist CGS 21680 and the antagonist KW 6002 on nicotine sensitization and conditioned locomotor activity in adult (8-week old) male Sprague-Dawley rats (WT). Moreover, behavioral responses to nicotine were studied in rats overexpressing A(2A) receptors under the control of the neuronal specific enolase (NSE) promotor. Changes in the levels of dopamine, glutamate and ?-aminobutyric acid in wild type (WT) and NSEA(2A) rats were determined with using LC-MS. KW 6002 significantly enhanced expression of nicotine sensitization and conditioned locomotion, while CGS 21680 reduced all these effects in WT rats. A reduction of the expression of nicotine-evoked conditioned locomotor activity was also observed in the NSEA(2A) animals. The transgenic rats displayed a reduced basal tissue level of glutamate in the prefrontal cortex and hippocampus while dopamine basal levels in the nucleus accumbens were raised. Chronic nicotine treatment caused a significant reduction in the glutamate tissue level in the dorsal and ventral striatum, prefrontal cortex and cerebellum in wild type rats. In NSEA(2A) animals the same drug treatment instead produced a rise of glutamate levels in the hippocampus and dorsal striatum. Taken together, A(2A) receptor signaling in the rat brain can counteract locomotor sensitization and conditioned locomotion to nicotine which are related to nicotine reward-learning. It is suggested that treatment with A(2A) receptor agonists can help counteract the abuse actions of nicotine. PMID:24632528

Jastrz?bska, Joanna; Nowak, Ewa; Smaga, Irena; Bystrowska, Beata; Frankowska, Ma?gorzata; Bader, Michael; Filip, Ma?gorzata; Fuxe, Kjell

2014-06-01

257

Transgenic Vegetable Breeding for Nutritional Quality and Health Benefits  

Directory of Open Access Journals (Sweden)

Full Text Available Vegetables are essential for well-balanced diets. About 3 billion people in the world are malnourished due to imbalanced diets. Vegetables can contribute to the prevention of malnutrition disorders. Genetic engineering enables vegetable breeders to incorporate desired transgenes into elite cultivars, thereby improving their value considerably. It further offers unique opportunities for improving nutritional quality and bringing other health benefits. Many vegetable crops have been genetically modified to improve traits such as higher nutritional status or better flavour, and to reduce bitterness or anti-nutritional factors. Transgenic vegetables can be also used for vaccine delivery. Consumers could benefit further from eating more nutritious transgenic vegetables, e.g. an increase of crop carotenoids by metabolic sink manipulation through genetic engineering appears feasible in some vegetables. Genetically engineering carrots containing increase Ca levels may boost Ca uptake, thereby reducing the incidence of Ca deficiencies such as osteoporosis. Fortified transgenic lettuce with zinc will overcome the deficiency of this micronutrient that severely impairs organ function. Folates deficiency, which is regarded as a global health problem, can also be overcomed with transgenic tomatoes with folate levels that provide a complete adult daily requirement. Transgenic lettuce with improved tocopherol and resveratrol composition may prevent coronary disease and arteriosclerosis and can contribute to cancer chemopreventative activity. Food safety and health benefits can also be enhanced through transgenic approaches, e.g. rural African resource-poor consumers will benefit eating cyanide-free cultivars of cassava. Biotechnology-derived vegetable crops will succed if clear advantages and safety are demonstrated to both growers and consumers.

João Silva Dias

2012-09-01

258

Overview on the field trials of transgenic plums in Romania  

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Full Text Available Transgenic clones C2, C3, C4, C5, C6, PT3 and PT5 of Prunus domestica L. transformed with the Plum pox virus coat protein gene (PPV-CP were evaluated for Sharka resistance under high infection pressure in field natural conditions in Romania. Transgenic clone C5, subsequently named “HoneySweet”, showed high resistance to PPV. None of the C5 trees became naturally infected by aphids for more than ten years. Known to develop the post-transcriptional gene silencing (PTGS mechanism, we assessed the effect of heterologous viruses on the efficacy and stability of PTGS displayed by the C5 plum against PPV. In this way, C5 trees were graftinoculated with different combinations of Prunus necrotic ringspot virus (PNRSV, Prune dwarf virus (PDV and PPV-D strain. The engineered resistance to PPV in C5 transgenic plums was stable and was not suppressed by the presence of the assayed heterologous viruses over a threeyear experimental period. Because the constitutive transcription of PPV-CP sequences naturally occurs in transgenic C5 plums, the environmental safety issues have been expressed on potential hazards concerning the emergence of PPV variants. In order to analyze this potential environmental effect was compared the serological and molecular variability of PPV detected in the transgenic trees versus those found in conventional plums. This risk assessment revealed a high similarity between PPV isolates from transgenic and conventional plums, and hence the transgenic plums utilized in this study do not affect the diversity of indigenous PPV populations.

Ioan Zagrai

2009-01-01

259

Rice transgene flow: its patterns, model and risk management.  

Science.gov (United States)

Progress has been made in a 12 year's systemic study on the rice transgene flow including (i) with experiments conducted at multiple locations and years using up to 21 pollen recipients, we have elucidated the patterns of transgene flow to different types of rice. The frequency to male sterile lines is 10(1) and 10(3) higher than that to O. rufipogon and rice cultivars. Wind speed and direction are the key meteorological factors affecting rice transgene flow. (ii) A regional applicable rice gene flow model is established and used to predict the maximum threshold distances (MTDs) of gene flow during 30 years in 993 major rice producing counties of southern China. The MTD0.1% for rice cultivars is basically ?5 m in the whole region, despite climate differs significantly at diverse locations and years. This figure is particularly valuable for the commercialization and regulation of transgenic rice. (iii) The long-term fate of transgene integrated into common wild rice was investigated. Results demonstrated that the F1 hybrids of transgenic rice/O. rufipogon gradually disappeared within 3-5 years, and the Bt or bar gene was not detectable in the mixed population, suggesting the O. rufipogon may possess a strong mechanism of exclusiveness for self-protection. (iv) The flowering time isolation and a 2-m-high cloth-screen protection were proved to be effective in reducing transgene flow. We have proposed to use a principle of classification and threshold management for different types of rice. PMID:25431202

Jia, Shirong; Yuan, Qianhua; Pei, Xinwu; Wang, Feng; Hu, Ning; Yao, Kemin; Wang, Zhixing

2014-12-01

260

Hematological and biochemical indexes in blood of HBV transgenic mice  

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Full Text Available Objective To study the effects of gene integration of HBV on the biochemical and hematological indices in blood of transgenic mice.Methods The venous blood was collected from orbital venous plexus of 28 normal mice born in the same brood(HBsAg negative and 50 HBV transgenic mice(HBsAg positive,6-8 weeks in age.The blood routine examination was performed,including white blood cells(WBC,red blood cells(RBC,hemoglobin(Hb,platelets(PLT,lymphocyte percentage(L%,intermediate cell percentage(M%,the percentage of leaf cells(G% and blood biochemical parameters including glucose(GLU,urea nitrogen(BUN,creatinine(CREA,total protein(ALB,albumin(ALB,total bilirubin(TBIL,alanine aminotransferase(ALT,aspartate aminotransferase(AST,total cholesterol(CHOL and triglyceride(TG.The differential indexes were analyzed in HBsAg positive and negative mice,also the mice of different sex.Results Significant differences were found between the transgenic and normal mice in blood GLU,BUN,CREA,RBCs,Hb and PLT(P < 0.05,while no significant differences were found in other indices.No difference was found in the above 6 indices between the different sex of normal mice,while there was significant difference in blood GLU between males and females in transgenic mice,and it was higher in males than in females(P < 0.05.The blood GLU,CREA and BUN were higher in transgenic male mice than in normal male mice,and the contents of RBCs,Hb and PLT were higher in transgenic female mice than in normal female mice(P < 0.05.Conclusion HBV DNA may throw some influences on blood biochemical and hematological indexes of HBV transgenic mice,and it may provide some valuable references to the study of HBV pathogenesis in HBV animal models.

Feng-jiao ZHENG

2011-09-01

 
 
 
 
261

Additive transgene expression and genetic introgression in multiple green-fluorescent protein transgenic crop x weed hybrid generations.  

Science.gov (United States)

The level of transgene expression in crop x weed hybrids and the degree to which crop-specific genes are integrated into hybrid populations are important factors in assessing the potential ecological and agricultural risks of gene flow associated with genetic engineering. The average transgene zygosity and genetic structure of transgenic hybrid populations change with the progression of generations, and the green fluorescent protein (GFP) transgene is an ideal marker to quantify transgene expression in advancing populations. The homozygous T(1) single-locus insert GFP/ Bacillus thuringiensis (Bt) transgenic canola ( Brassica napus, cv Westar) with two copies of the transgene fluoresced twice as much as hemizygous individuals with only one copy of the transgene. These data indicate that the expression of the GFP gene was additive, and fluorescence could be used to determine zygosity status. Several hybrid generations (BC(1)F(1), BC(2)F(1)) were produced by backcrossing various GFP/Bt transgenic canola ( B. napus, cv Westar) and birdseed rape ( Brassica rapa) hybrid generations onto B. rapa. Intercrossed generations (BC(2)F(2) Bulk) were generated by crossing BC(2)F(1) individuals in the presence of a pollinating insect ( Musca domestica L.). The ploidy of plants in the BC(2)F(2) Bulk hybrid generation was identical to the weedy parental species, B. rapa. AFLP analysis was used to quantify the degree of B. napus introgression into multiple backcross hybrid generations with B. rapa. The F(1) hybrid generations contained 95-97% of the B. napus-specific AFLP markers, and each successive backcross generation demonstrated a reduction of markers resulting in the 15-29% presence in the BC(2)F(2) Bulk population. Average fluorescence of each successive hybrid generation was analyzed, and homozygous canola lines and hybrid populations that contained individuals homozygous for GFP (BC(2)F(2) Bulk) demonstrated significantly higher fluorescence than hemizygous hybrid generations (F(1), BC(1)F(1) and BC(2)F(1)). These data demonstrate that the formation of homozygous individuals within hybrid populations increases the average level of transgene expression as generations progress. This phenomenon must be considered in the development of risk-management strategies. PMID:13679991

Halfhill, M D; Millwood, R J; Weissinger, A K; Warwick, S I; Stewart, C N

2003-11-01

262

Transgênicos sem maniqueísmo Transgenics without manichaeism  

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Full Text Available Vivemos em uma época marcada pela hegemonia da ciência e da tecnologia, carregada de questões à espera de respostas, para que o futuro da humanidade seja alcançado de forma segura e sustentável. O desenvolvimento de processos agroindustriais - especificamente, a produção de alimentos - com tecnologia de DNA recombinante tem trazido perspectivas de bons lucros apenas para os grandes conglomerados da biotecnologia e para produtores rurais com alto grau de desenvolvimento tecnológico. Discordamos de uma moratória para a tecnologia do DNA recombinante. Além disso, afirmações de ausência de risco podem levar a conclusões precipitadas, pois pouquíssimos testes foram realizados até hoje. É premente que se estabeleça uma política nacional de biossegurança, que envolva a sociedade civil organizada e todos os órgãos de governo (federal e estaduais responsáveis pela fiscalização, e que se implantem um programa de biovigilância e um código de ética de manipulações genéticas.We live in an era characterized by the hegemony of science and technology, an era fraught with questions awaiting answers which would enable a safe and sustainable future for humankind. The development of agro-industrial processes - food products in particular - through recombinant DNA technology has enhanced the profit prospects of the few big biotechnology companies and of large-scale farmers who have access to the latest technological developments. We thus oppose a moratorium on recombinant DNA technology. Moreover, hasty statements about risk-free transgenics may be misleading in the absence of extensive safety tests. There is a pressing need for the establishment of a biosafety policy in this country involving the organized civil society and every government agency responsible for monitoring such matters. There is also the need to put in place a bio-surveillance and a code of ethics regarding genetic manipulation.

Silvio Valle

2000-10-01

263

Transgênicos sem maniqueísmo / Transgenics without manichaeism  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: Portuguese Abstract in portuguese Vivemos em uma época marcada pela hegemonia da ciência e da tecnologia, carregada de questões à espera de respostas, para que o futuro da humanidade seja alcançado de forma segura e sustentável. O desenvolvimento de processos agroindustriais - especificamente, a produção de alimentos - com tecnologi [...] a de DNA recombinante tem trazido perspectivas de bons lucros apenas para os grandes conglomerados da biotecnologia e para produtores rurais com alto grau de desenvolvimento tecnológico. Discordamos de uma moratória para a tecnologia do DNA recombinante. Além disso, afirmações de ausência de risco podem levar a conclusões precipitadas, pois pouquíssimos testes foram realizados até hoje. É premente que se estabeleça uma política nacional de biossegurança, que envolva a sociedade civil organizada e todos os órgãos de governo (federal e estaduais) responsáveis pela fiscalização, e que se implantem um programa de biovigilância e um código de ética de manipulações genéticas. Abstract in english We live in an era characterized by the hegemony of science and technology, an era fraught with questions awaiting answers which would enable a safe and sustainable future for humankind. The development of agro-industrial processes - food products in particular - through recombinant DNA technology ha [...] s enhanced the profit prospects of the few big biotechnology companies and of large-scale farmers who have access to the latest technological developments. We thus oppose a moratorium on recombinant DNA technology. Moreover, hasty statements about risk-free transgenics may be misleading in the absence of extensive safety tests. There is a pressing need for the establishment of a biosafety policy in this country involving the organized civil society and every government agency responsible for monitoring such matters. There is also the need to put in place a bio-surveillance and a code of ethics regarding genetic manipulation.

Silvio, Valle.

2000-10-01

264

Cloning of transgenic tobacco BY-2 cells; an efficient method to analyse and reduce high natural heterogeneity of transgene expression  

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Full Text Available Abstract Background Phenotypic characterization of transgenic cell lines, frequently used in plant biology studies, is complicated because transgene expression in individual cells is often heterogeneous and unstable. To identify the sources and to reduce this heterogeneity, we transformed tobacco (Nicotiana tabacum L. BY-2 cells with a gene encoding green fluorescent protein (GFP using Agrobacterium tumefaciens, and then introduced a simple cloning procedure to generate cell lines derived from the individual transformed cells. Expression of the transgene was monitored by analysing GFP fluorescence in the cloned lines and also in lines obtained directly after transformation. Results The majority (~90% of suspension culture lines derived from calli that were obtained directly from transformation consisted of cells with various levels of GFP fluorescence. In contrast, nearly 50% of lines generated by cloning cells from the primary heterogeneous suspensions consisted of cells with homogenous GFP fluorescence. The rest of the lines exhibited "permanent heterogeneity" that could not be resolved by cloning. The extent of fluorescence heterogeneity often varied, even among genetically identical clones derived from the primary transformed lines. In contrast, the offspring of subsequent cloning of the cloned lines was uniform, showing GFP fluorescence intensity and heterogeneity that corresponded to the original clone. Conclusion The results demonstrate that, besides genetic heterogeneity detected in some lines, the primary lines often contained a mixture of epigenetically different cells that could be separated by cloning. This indicates that a single integration event frequently results in various heritable expression patterns, which are probably accidental and become stabilized in the offspring of the primary transformed cells early after the integration event. Because heterogeneity in transgene expression has proven to be a serious problem, it is highly advisable to use transgenes tagged with a visual marker for BY-2 transformation. The cloning procedure can be used not only for efficient reduction of expression heterogeneity of such transgenes, but also as a useful tool for studies of transgene expression and other purposes.

Fischer Lukas

2009-04-01

265

Development of transgenic watermelon resistant to Cucumber mosaic virus and Watermelon mosaic virus by using a single chimeric transgene construct.  

Science.gov (United States)

Watermelon, an important fruit crop worldwide, is prone to attack by several viruses that often results in destructive yield loss. To develop a transgenic watermelon resistant to multiple virus infection, a single chimeric transgene comprising a silencer DNA from the partial N gene of Watermelon silver mottle virus (WSMoV) fused to the partial coat protein (CP) gene sequences of Cucumber mosaic virus (CMV), Cucumber green mottle mosaic virus (CGMMV) and Watermelon mosaic virus (WMV) was constructed and transformed into watermelon (cv. Feeling) via Agrobacterium-mediated transformation. Single or multiple transgene copies randomly inserted into various locations in the genome were confirmed by Southern blot analysis. Transgenic watermelon R(0) plants were individually challenged with CMV, CGMMV or WMV, or with a mixture of these three viruses for resistance evaluation. Two lines were identified to exhibit resistance to CMV, CGMMV, WMV individually, and a mixed inoculation of the three viruses. The R(1) progeny of the two resistant R(0) lines showed resistance to CMV and WMV, but not to CGMMV. Low level accumulation of transgene transcripts in resistant plants and small interfering (si) RNAs specific to CMV and WMV were readily detected in the resistant R(1) plants by northern blot analysis, indicating that the resistance was established via RNA-mediated post-transcriptional gene silencing (PTGS). Loss of the CGMMV CP-transgene fragment in R1 progeny might be the reason for the failure to resistant CGMMV infection, as shown by the absence of a hybridization signal and no detectable siRNA specific to CGMMV in Southern and northern blot analyses. In summary, this study demonstrated that fusion of different viral CP gene fragments in transgenic watermelon contributed to multiple virus resistance via PTGS. The construct and resistant watermelon lines developed in this study could be used in a watermelon breeding program for resistance to multiple viruses. PMID:22203520

Lin, Ching-Yi; Ku, Hsin-Mei; Chiang, Yi-Hua; Ho, Hsiu-Yin; Yu, Tsong-Ann; Jan, Fuh-Jyh

2012-10-01

266

Handmade cloned transgenic piglets expressing the nematode fat-1 gene  

DEFF Research Database (Denmark)

Abstract Production of transgenic animals via somatic cell nuclear transfer (SCNT) has been adapted worldwide, but this application is somewhat limited by its relatively low efficiency. In this study, we used handmade cloning (HMC) established previously to produce transgenic pigs that express the functional nematode fat-1 gene. Codon-optimized mfat-1 was inserted into eukaryotic expression vectors, which were transferred into primary swine donor cells. Reverse transcriptase PCR (RT-PCR), gas chromatography, and chromosome analyses were performed to select donor clones capable of converting n-6 into n-3 fatty acids. Blastocysts derived from the clones that lowered the n-6/n-3 ratio to approximately 1:1 were transferred surgically into the uteri of recipients for transgenic piglets. By HMC, 37% (n=558) of reconstructed embryos developed to the blastocyst stage after 7 days of culture in vitro, with an average cell number of 81±36 (n=14). Three recipients became pregnant after 408 day-6 blastocysts were transferred into four naturally cycling females, and a total of 14 live offspring were produced. The nematode mfat-1 effectively lowered the n-6/n-3 ratio in muscle and major organs of the transgenic pig. Our results will help to establish a reliable procedure and an efficient option in the production of transgenic animals.

Zhang, Peng; Zhang, Yidi

2012-01-01

267

Design rules for efficient transgene expression in plants.  

Science.gov (United States)

Sustained expression of transgenes in specified developmental patterns is commonly needed in plant biotechnology, but obstructed by transgene silencing. Here, we present a set of gene design rules, tested on the silencing-susceptible beetle luc and bacterial ims genes, expressed in sugarcane. Designs tested independently or in combination included removal of rare codons, removal of RNA instability sequences, blocking of likely endogenous sRNA binding sites and randomization of non-rare codons. Stable transgene expression analyses, on multiple independent lines per construct, showed greatest improvement from the removal of RNA instability sequences, accompanied by greatly reduced transcript degradation evident in northern blot analysis. We provide a set of motifs that readily can be eliminated concurrently with rare codons and undesired structural features such as repeat sequences, using Gene Designer 2.0 software. These design rules yielded 935- and 5-fold increased expression in transgenic callus, relative to the native luc and ims sequences; and gave sustained expression under the control of sugarcane and heterologous promoters over several years in greenhouse and field trials. The rules can be applied easily with codon usage tables from any plant species, providing a simple and effective means to achieve sustained expression of otherwise silencing-prone transgenes in plants. PMID:24854834

Jackson, Mark A; Sternes, Peter R; Mudge, Stephen R; Graham, Michael W; Birch, Robert G

2014-09-01

268

A selectively terminable transgenic rice line expressing human lactoferrin.  

Science.gov (United States)

Human lactoferrin (hLF) is a multifunctional milk protein which could be utilized for promoting human health. Transgenic rice has been used as a bioreactor for mass production of recombinant hLF. However, one major concern over such transgenic rice is the risk of its unintended spreading into environment and into our food supplies. Here we report the development of selectively terminable transgenic rice expressing human lactoferrin in seeds. These transgenic rice plants could be selectively terminated by bentazon, a common herbicide used for rice weed control. The hLF expression cassette was constructed into a T-DNA containing the RNA interference cassette suppressing the expression of the rice gene CYP81A6 which detoxifies herbicide bentazon, and the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) cassette which confers to glyphosate tolerance. A transgenic line, named as G281, was identified for its high sensitivity to bentazon, high tolerance to glyphosate, and high expression of hLF. Southern analysis suggested G281 is a single copy insertion event. Field tests demonstrated that G281 plants can be completely killed by a single spray of bentazon at 1000 mg/L, which is safe to regular rice and represents only half of the dose recommended by manufacturer for rice field weed control. Therefore, any G281 contaminations in regular rice could be selectively terminated to make sure it will not enter food or feed supplies. PMID:20433928

Lin, Chaoyang; Nie, Peng; Lu, Wei; Zhang, Qing; Li, Jing; Shen, Zhicheng

2010-11-01

269

Reversible methylation and silencing of homologous transgenes in tobacco plants  

International Nuclear Information System (INIS)

Homology dependent gene silencing in transgenic plants occurs frequently when multiple copies of a transgene or a transgene with homology to an endogenous plant gene are present in a plant nucleus. The multiple copies can be arranged in cis on the same DNA molecule, or they can be present on different DNA molecules, either in allelic or non-allelic locations (trans inactivation). Although the phenomenon of silencing is well established, the mechanism by which it occurs are not completely understood. At present, different silencing systems can be distinguished by three major features: (1) the region of homology involved in the interaction (promoter or protein coding region); (2) the level at which silencing occurs (transcriptional or post-transcriptional); and (3) the degree of meiotic heritability of the silenced/methylated states after segregation of two interacting homologous loci in progeny. Interactions that lead to the silencing and methylation of partially homologous transgenes in tobacco have been studied. The transgenes share homology only in promoter regions; both the nopaline synthase promoter and the 35S promoter of the cauliflower mosaic virus have been used to drive the expression of various selectable and biochemical marker genes. The properties of these silencing systems are discussed, with particular reference to the features mentioned above. (author). 13 refs

270

Clock Gene Expression during Chronic Inflammation Induced by Infection with Trypanosoma brucei brucei in Rats  

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African sleeping sickness is characterized by alterations in rhythmic functions. It is not known if the disease affects the expression of clock genes, which are the molecular basis for rhythm generation. We used a chronic rat model of experimental sleeping sickness, caused by the extracellular parasite Trypanosoma brucei brucei (Tb brucei), to study the effects on clock gene expression. In tissue explants of pituitary glands from Period1-luciferase (Per1-luc) transgenic rats infected with Tb ...

Lundkvist, Gabriella B. S.; Sellix, Michael T.; Nyga?rd, Mikael; Davis, Erin; Straume, Marty; Kristensson, Krister; Block, Gene D.

2010-01-01

271

A set of highly informative rat simple sequence length polymorphism (SSLP markers and genetically defined rat strains  

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Full Text Available Abstract Background The National Bio Resource Project for the Rat in Japan (NBRP-Rat is focusing on collecting, preserving and distributing various rat strains, including spontaneous mutant, transgenic, congenic, and recombinant inbred (RI strains. To evaluate their value as models of human diseases, we are characterizing them using 109 phenotypic parameters, such as clinical measurements, internal anatomy, metabolic parameters, and behavioral tests, as part of the Rat Phenome Project. Here, we report on a set of 357 simple sequence length polymorphism (SSLP markers and 122 rat strains, which were genotyped by the marker set. Results The SSLP markers were selected according to their distribution patterns throughout the whole rat genome with an average spacing of 7.59 Mb. The average number of informative markers between all possible pairs of strains was 259 (72.5% of 357 markers, showing their high degree of polymorphism. From the genetic profile of these rat inbred strains, we constructed a rat family tree to clarify their genetic background. Conclusion These highly informative SSLP markers as well as genetically and phenotypically defined rat strains are useful for designing experiments for quantitative trait loci (QTL analysis and to choose strategies for developing new genetic resources. The data and resources are freely available at the NBRP-Rat web site 1.

Yamasaki Ken-ichi

2006-04-01

272

Differences in social interaction- vs cocaine reward in rat vs mouse  

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Considering that human addicts regularly prefer drugs of abuse to drug-free social interaction, the present findings suggest that our experimental paradigm of concurrent CPP for cocaine vs social interaction is of even greater translational power if performed in C57BL/6 mice, the genetic background for most transgenic rodent models, than in rats.

Kai K Kummer

2014-10-01

273

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

Science.gov (United States)

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

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

1995-07-01

274

Expression of human alpha 1 antitrypsin in transgenic sheep.  

Science.gov (United States)

We have recently described the production of large amounts (milk of transgenic sheep (Wright et al., 1991). Here, we describe in more detail the expression of the human protein in the milk of these animals throughout the lactation period. Human alpha 1 antitrypsin is also found at much lower levels in the plasma of transgenic ewes before, during and after lactation. It is also detected in male plasma at very low levels. We have previously shown human alpha 1 antitrypsin purified from transgenic sheep milk to be indistinguishable from commercially available human plasma derived alpha 1 antitrypsin in terms of gross sugar content and in vitro activity. Here we extend this comparison to more detailed analyses of glycosylation state, amino-terminal sequence, pI value, and molecular weight determination by mass spectrometry. PMID:1369184

Carver, A; Wright, G; Cottom, D; Cooper, J; Dalrymple, M; Temperley, S; Udell, M; Reeves, D; Percy, J; Scott, A

1992-01-01

275

Transgene-induced pleiotropic effects in transplastomic plants.  

Science.gov (United States)

Since the first demonstration of stable transgene integration in the plastid genome (plastome) of higher plants, plastid transformation has been used for a wide range of purposes, including basic studies as well as biotechnological applications, showing that transplastomic plants are an effective system to produce recombinant proteins. Compared to nuclear transformation, the main advantages of this technology are the high and stable production level of proteins as well as the natural containment of transgenes. To date, more than 100 transgenes have been successfully expressed in plant chloroplasts. In some cases, however, unintended pleiotropic effects on plant growth and physiology were shown in transplastomic plants. In this paper, we review such effects and discuss some of the technologies developed to overcome them. PMID:24101241

Scotti, N; Cardi, T

2014-02-01

276

Identification of transgenic foods using NIR spectroscopy: A review  

Science.gov (United States)

The utilization of chemometric methods in the quantitative and qualitative analysis of feeds, foods, medicine and so on has been accompanied with the great evolution in the progress and in the near infrared spectroscopy (NIRS). Hence, recently the application of NIR spectroscopy has extended on the context of genetics and transgenic products. The aim of this review was to investigate the application of NIR spectroscopy to identificate transgenic products and to compare it with the traditional methods. The results of copious researches showed that the application of NIRS technology was successful to distinguish transgenic foods and it has advantages such as fast, avoiding time-consuming, non-destructive and low cost in relation to the antecedent methods such as PCR and ELISA.

Alishahi, A.; Farahmand, H.; Prieto, N.; Cozzolino, D.

2010-01-01

277

Molecular control of transgene escape from genetically modified plants.  

Science.gov (United States)

Potential risks of gene escape from transgenic crops through pollen and seed dispersal are being actively discussed and have slowed down full utilization of gene technology in crop improvement. To ban the transgene flow, barren zones and 'terminator' technology were developed as GMO risk management technologies in transgenic crops. Unfortunately, the technologies have not protected reliably the transgene migration to wild relatives. The present study offers a novel molecular technique to eliminate gene flow from transgenic plants to wild relatives by recoverable block of function (RBF). The RBF consists of a blocking sequence linked to the gene of interest and a recovering sequence, all in one transformable construct. The blocking sequence blocks a certain molecular or physiological function of the host plant. Action of the blocking sequence leads to the death of the host plant or to an alteration in its phenotype resulting in inability for sexual reproduction in nature. The recovering construct recovers the blocked function of the host plant. The recovering construct is regulated externally by a specific chemical or physical treatment of the plants and does not act under natural conditions. In nature, hybrids of the transgenic plants with its wild relatives carrying the RBF will die or be unable to reproduce because of the blocking construct action. A working model of RBF is described in this report as one example of the RBF concept. This RBF example is based on barnase (the blocking construct) and barstar (the recovering construct) gene expression in tobacco under sulfhydryl endopeptidase (SH-EP) and a heat shock (HS) promoter, respectively. PMID:11166439

Kuvshinov, V; Koivu, K; Kanerva, A; Pehu, E

2001-02-01

278

Establishment and detection of HBV transgenic mice with YMDD mutation  

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Full Text Available Objective To establish the hepatitis B virus(HBV transgenic mice with YMDD mutation,and provide an animal model for research of HBV prevention and therapeutic approach.Methods 1.3 copies HBV genome containing YMDD mutation associated with lamivudine resistance was injected into the zygote of FVB/N female mice by microinjection.Integration and passage of exogenous gene in transgenic mice was confirmed by PCR.The expression of HBsAg in liver and kidney tissues in transgenic mice was identified by ELISA and immunohistochemistry.Results A total of 3401 zygotes were injected and 269 F0 pups were born.PCR analysis indicated that 33 out of 269 pups were positive,and the integrating rate of exogenous gene was 12.3% in F0.Fluorescent quantitative PCR showed that HBV DNA was weakly positive in serum samples in 9 transgenic mice,less than 103copies/ml.The expression of HBsAg in transgenic mice was observed in liver and kidney tissues by immunohistochemistry,and it was higher in kidney than in liver.The target gene was detected by PCR in 27.6% of 47 F1 offsprings.The expression of HBsAg could be observed in liver and kidney tissues in F1,which was similar to that in F0.Conclusion 1.3 copies HBV transgenic mice model containing YMDD mutation associated with lamivudine resistance is successfully produced by microinjection,and HBsAg expression can be transmitted through germline cells.

Yu-qin YOU

2011-09-01

279

Differential subcellular targeting of recombinant human ??-proteinase inhibitor influences yield, biological activity and in planta stability of the protein in transgenic tomato plants.  

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The response of protein accumulation site on yield, biological activity and in planta stability of therapeutic recombinant human proteinase inhibitor (??-PI) was analyzed via targeting to different subcellular locations, like endoplasmic reticulum (ER), apoplast, vacuole and cytosol in leaves of transgenic tomato plants. In situ localization of the recombinant ??-PI protein in transgenic plant cells was monitored by immunohistochemical staining. Maximum accumulation of recombinant ??-PI in T? and T? transgenic tomato plants was achieved from 1.5 to 3.2% of total soluble protein (TSP) by retention in ER lumen, followed by vacuole and apoplast, whereas cytosolic targeting resulted into degradation of the protein. The plant-derived recombinant ??-PI showed biological activity for elastase inhibition, as monitored by residual porcine pancreatic elastase (PPE) activity assay and band-shift assay. Recombinant ??-PI was purified from transgenic tomato plants with high yield, homogeneity and biological activity. Purified protein appeared as a single band of ?48-50 kDa on SDS-PAGE with pI value ranging between 5.1 and 5.3. Results of mass spectrometry and optical spectroscopy of purified recombinant ??-PI revealed the structural integrity of the recombinant protein comparable to native serum ??-PI. Enzymatic deglycosylation and lectin-binding assays with the purified recombinant ??-PI showed compartment-specific N-glycosylation of the protein targeted to ER, apoplast and vacuole. Conformational studies based on urea-induced denaturation and circular dichroism (CD) spectroscopy revealed relatively lower stability of the recombinant ??-PI protein, compared to its serum counterpart. Pharmacokinetic evaluation of plant derived recombinant and human plasma-purified ??-PI in rat, by intravenous route, revealed significantly faster plasma clearance and lower area under curve (AUC) of recombinant protein. Our data suggested significance of protein sorting sequences and feasibility to use transgenic plants for the production of stable, glycosylated and biologically active recombinant ??-PI for further therapeutic applications. PMID:23017899

Jha, Shweta; Agarwal, Saurabh; Sanyal, Indraneel; Jain, G K; Amla, D V

2012-11-01

280

Transgenic zebrafish recapitulating tbx16 gene early developmental expression.  

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We describe the creation of a transgenic zebrafish expressing GFP driven by a 7.5 kb promoter region of the tbx16 gene. This promoter segment is sufficient to recapitulate early embryonic expression of endogenous tbx16 in the presomitic mesoderm, the polster and, subsequently, in the hatching gland. Expression of GFP in the transgenic lines later in development diverges to some extent from endogenous tbx16 expression with the serendipitous result that one line expresses GFP specifically in commissural primary ascending (CoPA) interneurons of the developing spinal cord. Using this line we demonstrate that the gene mafba (valentino) is expressed in CoPA interneurons. PMID:21720556

Wells, Simon; Nornes, Svanhild; Lardelli, Michael

2011-01-01

 
 
 
 
281

Male-specific insecticide resistance and mosquito transgene dispersal.  

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There is a need to develop methods to spread disease-blocking transgenes through mosquito populations. This article discusses the possibility of linking transgenes to insecticide-resistant alleles engineered to be expressed only in males. The resulting increase in mean longevity of males carrying the construct under insecticide treatment could easily outweigh any fitness costs in females, so that the construct would spread rapidly. It should be possible to produce constructs where any potential risk of loss of male-specific expression would be negligible. PMID:15324731

Sinkins, Steven P; Hastings, Ian M

2004-09-01

282

Enhanced transgene expression from chromatinized plasmid DNA in mouse liver.  

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Plasmid DNA was chromatinized with core histones (H2A, H2B, H3, and H4) in vitro and was delivered into mouse liver by hydrodynamics-based administration. Transgene expression from the chromatinized plasmid DNA was more efficient than that from plasmid DNA delivered in the naked form. The use of acetylation-enriched histones isolated from cells treated with a histone deacetylase inhibitor (trichostatin A) seemed to be more effective. These results indicated that chromatinized plasmid DNA is useful for efficient transgene expression in vivo. PMID:23247018

Kamiya, Hiroyuki; Miyamoto, Shiho; Goto, Hitomi; Kanda, Genki N; Kobayashi, Miwako; Matsuoka, Ichiro; Harashima, Hideyoshi

2013-01-30

283

Neuron Loss in Transgenic Mouse Models of Alzheimer's Disease  

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Full Text Available Since their initial generation in the mid 1990s, transgenic mouse models of Alzheimers's disease (AD have been proven to be valuable model systems which are indispensable for modern AD research. Whereas most of these models are characterized by extensive amyloid plaque pathology, inflammatory changes and often behavioral deficits, modeling of neuron loss was much less successful. The present paper discusses the current achievements of modeling neuron loss in transgenic mouse models based on APP/A? and Tau overexpression and provides an overview of currently available AD mouse models showing these pathological alterations.

Oliver Wirths

2010-01-01

284

Transgenic rice endosperm as a bioreactor for molecular pharming.  

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Plants provide a promising expression platform for producing recombinant proteins with several advantages in terms of high expression level, lower production cost, scalability, and safety and environment-friendly. Molecular pharming has been recognized as an emerging industry with strategic importance that could play an important role in economic development and healthcare in China. Here, this review represents the significant advances using transgenic rice endosperm as bioreactor to produce various therapeutic recombinant proteins in transgenic rice endosperm and large-scale production of OsrHSA, and discusses the challenges to develop molecular pharming as an emerging industry with strategic importance in China. PMID:24413763

Ou, Jiquan; Guo, Zhibin; Shi, Jingni; Wang, Xianghong; Liu, Jingru; Shi, Bo; Guo, Fengli; Zhang, Chufu; Yang, Daichnag

2014-04-01

285

Complex patterns of inheritance of an imprinted murine transgene suggest incomplete germline erasure  

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Here we report a transgenic mouse line that exhibits significant deviations from a classic pattern of parental imprinting. When the transgene is passed through the female germline, it is completely silenced in some offspring while in others expression is reduced. This variable expressivity does not appear to be the result of differences in the presence of unlinked modifiers. Female transmission of the transgene is associated with hypermethylation. The transgene is generally reactivated on pas...

Kearns, Margot; Preis, Jost; Mcdonald, Margaret; Morris, Christine; Whitelaw, Emma

2000-01-01

286

Efficient integration of transgenes into a defined locus in human embryonic stem cells  

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Random integration is one of the more straightforward methods to introduce a transgene into human embryonic stem (ES) cells. However, random integration may result in transgene silencing and altered cell phenotype due to insertional mutagenesis in undefined gene regions. Moreover, reliability of data may be compromised by differences in transgene integration sites when comparing multiple transgenic cell lines. To address these issues, we developed a genetic manipulation strategy based on homo...

Sakurai, Kenji; Shimoji, Miho; Tahimic, Candice G. T.; Aiba, Kazuhiro; Kawase, Eihachiro; Hasegawa, Kouichi; Amagai, Yuji; Suemori, Hirofumi; Nakatsuji, Norio

2010-01-01

287

Nucleus-targeted Dmp1 transgene fails to rescue dental defects in Dmp1 null mice.  

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Dentin matrix protein 1 (DMP1) is essential to odontogenesis. Its mutations in human subjects lead to dental problems such as dental deformities, hypomineralization and periodontal impairment. Primarily, DMP1 is considered as an extracellular matrix protein that promotes hydroxyapatite formation and activates intracellular signaling pathway via interacting with ?v?3 integrin. Recent in vitro studies suggested that DMP1 might also act as a transcription factor. In this study, we examined whether full-length DMP1 could function as a transcription factor in the nucleus and regulate odontogenesis in vivo. We first demonstrated that a patient with the DMP1 M1V mutation, which presumably causes a loss of the secretory DMP1 but does not affect the nuclear translocation of DMP1, shows a typical rachitic tooth defect. Furthermore, we generated transgenic mice expressing (NLS)DMP1, in which the endoplasmic reticulum (ER) entry signal sequence of DMP1 was replaced by a nuclear localization signal (NLS) sequence, under the control of a 3.6?kb rat type I collagen promoter plus a 1.6?kb intron 1. We then crossbred the (NLS)DMP1 transgenic mice with Dmp1 null mice to express the (NLS)DMP1 in Dmp1-deficient genetic background. Although immunohistochemistry demonstrated that (NLS)DMP1 was localized in the nuclei of the preodontoblasts and odontoblasts, the histological, morphological and biochemical analyses showed that it failed to rescue the dental and periodontal defects as well as the delayed tooth eruption in Dmp1 null mice. These data suggest that the full-length DMP1 plays no apparent role in the nucleus during odontogenesis. PMID:25105818

Lin, Shu-Xian; Zhang, Qi; Zhang, Hua; Yan, Kevin; Ward, Leanne; Lu, Yong-Bo; Feng, Jian-Quan

2014-09-01

288

Crossing the divide: gene flow produces intergeneric hybrid in feral transgenic creeping bentgrass population.  

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Gene flow is the most frequently expressed public concern related to the deregulation of transgenic events (Snow 2002; Ellstrand 2003). However, assessing the potential for transgene escape is complex because it depends on the opportunities for unintended gene flow, and establishment and persistence of the transgene in the environment (Warwick et al. 2008). Creeping bentgrass (Agrostis stolonifera L.), a turfgrass species widely used on golf courses, has been genetically engineered to be resistant to glyphosate, a nonselective herbicide. Outcrossing species, such as creeping bentgrass (CB), which have several compatible species, have greater chances for gene escape and spontaneous hybridization (i.e. natural, unassisted sexual reproduction between taxa in the field), which challenges transgene containment. Several authors have emphasized the need for evidence of spontaneous hybridization to infer the potential for gene flow (Armstrong et al. 2005). Here we report that a transgenic intergeneric hybrid has been produced as result of spontaneous hybridization of a feral-regulated transgenic pollen receptor (CB) and a nontransgenic pollen donor (rabbitfoot grass, RF, Polypogon monspeliensis (L.) Desf.). We identified an off-type transgenic seedling and confirmed it to be CB × RF intergeneric hybrid. This first report of a transgenic intergeneric hybrid produced in situ with a regulated transgenic event demonstrates the importance of considering all possible avenues for transgene spread at the landscape level before planting a regulated transgenic crop in the field. Spontaneous hybridization adds a level of complexity to transgene monitoring, containment, mitigation and remediation programmes. PMID:22625177

Zapiola, María L; Mallory-Smith, Carol A

2012-10-01

289

Genetic and Molecular Analysis of Transgenic Rice cv. Rojolele Expressing Lactoferrin  

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Full Text Available In a previous study, human lactoferrin gene have introduced into Javanica rice cv. Rojolele by Agrobacterium-mediated transformation. Lactoferrin (LF is an 80 kDa iron-binding glycoprotein that has been proposed to have many biological roles such as protection against microbial and virus infection. This study aims to analyze the integration and level of lactoferrin gene expression of transgenic rice cv. Rojolele. The study also aims to examine the genetic character of transgenic rice expressing recombinant lactoferrin. Stability expression of recombinant lactoferrin transgenic rice seeds over generations were analyzed by ELISA, while the integration stability of recombinant hLF gene in transgenic plants performed by PCR. The mitotic time, cell cycle and chromosome characterization of transgenic and non-transgenic rice cv. Rojolele were determined. Chromosome characterization of the trangenic and non transgenic rice cv. Rojolele was investigated to determine the genetic variation. All of the above efforts were aimed to evaluate the genetically engineered rice containing recombinant lactoferrin as a nutraceutical food. The results showed that the expression was stable through three consecutive generations. The expression of the hLF gene increased during grain-filling period. The active time of mitotic cells of transgenic rice rojolele was longer than the cells of non-transgenic rice. In addition, the cycle cell of transgenic and non-transgenic rojolele contained prophase, prometaphase, metaphase, anaphase, telophase and interphase. The result showed that all of the transgenic lines had diploid (2n chromosome number = 24.

Diah Rachmawati

2014-02-01

290

Rearrangement of only one human IGHV gene is sufficient to generate a wide repertoire of antigen specific antibody responses in transgenic mice.  

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In recent years, mice carrying human IG transgenes are being generated for the production of human monoclonal antibodies as an alternative approach to the conventional use of mouse or chimeric-humanized antibodies. Theoretically, the size of the repertoire of human antibodies that these mice could produce would be critically dependent on the number of human V genes introduced in the transgene. This could be the case for BABkappa and BABkappa,lambda transgenic mice, which carry several genes from the human IGK (BABkappa), and IGK and IGL (BABkappa,lambda) loci, but only five human IGHV genes and the entire IGHD-IGHJ cluster linked to two human IGHC (IGHM-IGHD) genes. We analyzed the expressed human IG genes in 30 IgM-secreting hybridomas generated from transgenic mice immunized either with soluble proteins (human IgM coupled to KLH) or with cells (human PBMC, tumour cell lines or rat cells transfected with human CD69). The results show that all hybridoma cells analyzed rearranged exclusively the IGHV1-2 gene, in contrast with naive spleen B cells that used three out of the five IGHV genes present in the transgene. The configuration of the rearranged CDR3 region revealed a much higher heterogeneity in the heavy chains. A variety of IGHJ and IGHD genes were used in hybridomas, and somatic mutations were also seen in some hybrids. Regarding the rearranged light chains genes, it was a much higher variety in the use of V and J genes in both, kappa and lambda chains, than in the heavy chain, and also in the level of mutation. The results indicate that only one IGHV gene is sufficient to generate a wide repertoire of antigen specific antibody responses. Thus, efforts aimed at the generation of new transgenic mice should focus more on the integrity of the D/J region and on the DNA regions regulating somatic hypermutation, rather than on the number of V genes present in the transgene. PMID:16343622

Suárez, Eduardo; Magadán, Susana; Sanjuán, Irene; Valladares, Mónica; Molina, Ana; Gambón, Francisco; Díaz-Espada, Fernando; González-Fernández, Africa

2006-04-01

291

Time-Course Expression Profiles of Hair Cycle-Associated Genes in Male Mini Rats after Depilation of Telogen-Phase Hairs  

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Jcl:WistarTGN(ARGHGEN)1Nts rat (Mini rat) is a growth hormone (GH)-deficient transgenic rat. The hair cycle in the dorsal skin of male Mini rats enters a long-lasting telogen phase after eights weeks of age, but depilation can induce a transient hair cycle again. In this study, a time-course profiling of genes expression was done on the dorsal skin of male Mini rats along the progression of depilation-induced hair cycle using DNA microarray analysis. As a result, 1,215 probe sets including 1,...

Aya Umeda-Ikawa; Isao Shimokawa; Kunio Doi

2009-01-01

292

Efficient expression of transgenes in adult zebrafish by electroporation  

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Full Text Available Abstract Background Expression of transgenes in muscle by injection of naked DNA is widely practiced. Application of electrical pulses at the site of injection was demonstrated to improve transgene expression in muscle tissue. Zebrafish is a precious model to investigate developmental biology in vertebrates. In this study we investigated the effect of electroporation on expression of transgenes in 3–6 month old adult zebrafish. Results Electroporation parameters such as number of pulses, voltage and amount of plasmid DNA were optimized and it was found that 6 pulses of 40 V·cm-1 at 15 ?g of plasmid DNA per fish increased the luciferase expression 10-fold compared to controls. Similar enhancement in transgene expression was also observed in Indian carp (Labeo rohita. To establish the utility of adult zebrafish as a system for transient transfections, the strength of the promoters was compared in A2 cells and adult zebrafish after electroporation. The relative strengths of the promoters were found to be similar in cell lines and in adult zebrafish. GFP fluorescence in tissues after electroporation was also studied by fluorescence microscopy. Conclusion Electroporation after DNA injection enhances gene expression 10-fold in adult zebrafish. Electroporation parameters for optimum transfection of adult zebrafish with tweezer type electrode were presented. Enhanced reporter gene expression upon electroporation allowed comparison of strengths of the promoters in vivo in zebrafish.

Rao S Hari

2005-10-01

293

Pre-clinical applications of transgenic mouse mammary cancer models.  

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Breast cancer is a leading cause of cancer morbidity and mortality. Given that the majority of human breast cancers appear to be due to non-genetic factors, identifying agents and mechanisms of prevention is key to lowering the incidence of cancer. Genetically engineered mouse models of mammary cancer have been important in elucidating molecular pathways and signaling events associated with the initiation, promotion, and the progression of cancer. Since several transgenic mammary models of human breast cancer progress through well-defined cancer stages, they are useful pre-clinical systems to test the efficacy of chemopreventive and chemotherapeutic agents. This review outlines several oncogenic pathways through which mammary cancer can be induced in transgenic models and describes several types of preventive and therapeutic agents that have been tested in transgenic models of mammary cancer. The effectiveness of farnesyl inhibitors, aromatase inhibitors, differentiating agents, polyamine inhibitors, anti-angiogenic inhibitors, and immunotherapeutic compounds including vaccines have been evaluated in reducing mammary cancer and tumor progression in transgenic models. PMID:12509137

Kavanaugh, C J; Desai, K V; Calvo, A; Brown, P H; Couldrey, C; Lubet, R; Green, J E

2002-12-01

294

Transgenic maize plants expressing a fungal phytase gene.  

Science.gov (United States)

Maize seeds are the major ingredient of commercial pig and poultry feed. Phosphorus in maize seeds exists predominantly in the form of phytate. Phytate phosphorus is not available to monogastric animals and phosphate supplementation is required for optimal animal growth. Undigested phytate in animal manure is considered a major source of phosphorus pollution to the environment from agricultural production. Microbial phytase produced by fermentation as a feed additive is widely used to manage the nutritional and environmental problems caused by phytate, but the approach is associated with production costs for the enzyme and requirement of special cares in feed processing and diet formulation. An alternative approach would be to produce plant seeds that contain high phytase activities. We have over-expressed Aspergillus niger phyA2 gene in maize seeds using a construct driven by the maize embryo-specific globulin-1 promoter. Low-copy-number transgenic lines with simple integration patterns were identified. Western-blot analysis showed that the maize-expressed phytase protein was smaller than that expressed in yeast, apparently due to different glycosylation. Phytase activity in transgenic maize seeds reached approximately 2,200 units per kg seed, about a 50-fold increase compared to non-transgenic maize seeds. The phytase expression was stable across four generations. The transgenic seeds germinated normally. Our results show that the phytase expression lines can be used for development of new maize hybrids to improve phosphorus availability and reduce the impact of animal production on the environment. PMID:17932782

Chen, Rumei; Xue, Guangxing; Chen, Ping; Yao, Bin; Yang, Wenzhu; Ma, Qianli; Fan, Yunliu; Zhao, Zuoyu; Tarczynski, Mitchell C; Shi, Jinrui

2008-08-01

295

Visualization of C. elegans transgenic arrays by GFP  

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Full Text Available Abstract Background Targeting the green fluorescent protein (GFP via the E. coli lac repressor (LacI to a specific DNA sequence, the lac operator (lacO, allows visualization of chromosomes in yeast and mammalian cells. In principle this method of visualization could be used for genetic mosaic analysis, which requires cell-autonomous markers that can be scored easily and at single cell resolution. The C. elegans lin-3 gene encodes an epidermal growth factor family (EGF growth factor. lin-3 is expressed in the gonadal anchor cell and acts through LET-23 (transmembrane protein tyrosine kinase and ortholog of EGF receptor to signal the vulval precursor cells to generate vulval tissue. lin-3 is expressed in the vulval cells later, and recent evidence raises the possibility that lin-3 acts in the vulval cells as a relay signal during vulval induction. It is thus of interest to test the site of action of lin-3 by mosaic analysis. Results We visualized transgenes in living C. elegans by targeting the green fluorescent protein (GFP via the E. coli lac repressor (LacI to a specific 256 sequence repeat of the lac operator (lacO incorporated into transgenes. We engineered animals to express a nuclear-localized GFP-LacI fusion protein. C. elegans cells having a lacO transgene result in nuclear-localized bright spots (i.e., GFP-LacI bound to lacO. Cells with diffuse nuclear fluorescence correspond to unbound nuclear localized GFP-LacI. We detected chromosomes in living animals by chromosomally integrating the array of the lacO repeat sequence and visualizing the integrated transgene with GFP-LacI. This detection system can be applied to determine polyploidy as well as investigating chromosome segregation. To assess the GFP-LacI•lacO system as a marker for mosaic analysis, we conducted genetic mosaic analysis of the epidermal growth factor lin-3, expressed in the anchor cell. We establish that lin-3 acts in the anchor cell to induce vulva development, demonstrating this method's utility in detecting the presence of a transgene. Conclusion The GFP-LacI•lacO transgene detection system works in C. elegans for visualization of chromosomes and extrachromosomal transgenes. It can be used as a marker for genetic mosaic analysis. The lacO repeat sequence as an extrachromosomal array becomes a valuable technique allowing rapid, accurate determination of spontaneous loss of the array, thereby allowing high-resolution mosaic analysis. The lin-3 gene is required in the anchor cell to induce the epidermal vulval precursors cells to undergo vulval development.

Sternberg Paul W

2006-06-01

296

In utero recombinant adeno-associated virus gene transfer in mice, rats, and primates  

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Full Text Available Abstract Background Gene transfer into the amniotic fluid using recombinant adenovirus vectors was shown previously to result in high efficiency transfer of transgenes into the lungs and intestines. Adenovirus mediated in utero gene therapy, however, resulted in expression of the transgene for less than 30 days. Recombinant adenovirus associated viruses (rAAV have the advantage of maintaining the viral genome in daughter cells thus providing for long-term expression of transgenes. Methods Recombinant AAV2 carrying green fluorescent protein (GFP was introduced into the amniotic sac of fetal rodents and nonhuman primates. Transgene maintenance and expression was monitor. Results Gene transfer resulted in rapid uptake and long-term gene expression in mice, rats, and non-human primates. Expression and secretion of the reporter gene, GFP, was readily demonstrated within 72 hours post-therapy. In long-term studies in rats and nonhuman primates, maintenance of GFP DNA, protein expression, and reporter gene secretion was documented for over one year. Conclusions Because only multipotential stem cells are present at the time of therapy, these data demonstrated that in utero gene transfer with AAV2 into stem cells resulted in long-term systemic expression of active transgene roducts. Thus, in utero gene transfer via the amniotic fluid may be useful in treatment of gene disorders.

Marrero Luis

2003-09-01

297

Transgene integration - an analysis in autotransgenic Labeo rohita Hamilton (Pisces: Cyprinidae)  

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Transgenic Labeo rohita founder population was analyzed for the presence of autotransgene having histone 3 promoter and growth hormone (GH) cDNA (LRH3-GHcDNA) or total GH gene (LRH3-GH2.8) by PCR with transgene specific primers. Transgene specific amplification was seen with LRH3-GHcDNA in five out of seven individuals and all three fishes with LRH3-GH2.8, indicating their transgenic nature. Transgene integration was also studied by Southern hybridization of DNA isolated from blood of the tra...

Rajesh, R.; Majumdar, K. C.

2005-01-01

298

Tilapia chromosomal growth hormone gene expression accelerates growth in transgenic zebrafish (Danio rerio)  

Scientific Electronic Library Online (English)

Full Text Available SciELO Chile | Language: English Abstract in english Gene transfer is economically important and model fish species has produced a great impact in modern biology and biotechnology. Transgenic zebrafish (Danio rerio) were generated through the co-injection of a GFP-expressing plasmid and an "all fish" transgene composed by the carp beta-actin promoter [...] and the chromosomal tilapia (Oreochromis hornorum) growth hormone gene. The GFP expression was a good indicator of stable transformation and allowed for high efficiency selection of transgenic fish. Transgenic F1 zebrafish grew 20% faster than full sibling non-transgenic controls.

Reynold, Morales; María Teresa, Herrera; Amílcar, Arenal; Asterio, Cruz; Oscar, Hernández; Rafael, Pimentel; Isabel, Guillén; Rebeca, Martínez; Mario P, Estrada.

2001-08-15

299

Colored medaka and zebrafish: transgenics with ubiquitous and strong transgene expression driven by the medaka ?-actin promoter.  

Science.gov (United States)

Conditional cell labeling, cell tracing, and genetic manipulation approaches are becoming increasingly important in developmental and regenerative biology. Such approaches in zebrafish research are hampered by the lack of an ubiquitous transgene driver element that is active at all developmental stages. Here, we report the isolation and characterization of the medaka fish (Oryzias latipes) ?-actin (Olactb) promoter, which drives constitutive transgene expression during all developmental stages, and the analysis of adult organs except blood cell types. Taking advantage of the compact medaka promoter, we succeeded in generating a zebrafish transgenic (Tg) line with unprecedentedly strong and widespread transgene expression from embryonic to adult stages. Moreover, the Tg carries a pair of loxP sites, which enables the reporter fluorophore to switch from DsRed2 to enhanced green fluorescent protein (EGFP). We induced Cre/loxP recombination with Tg(hsp70l: mCherry-t2a-Cre(ERt2) ) in the double Tg embryo and generated a Tg line that constitutively expresses EGFP. We further demonstrate the powerful application of Olactb-driven Tgs for cell lineage tracing using transplantation experiments with embryonic cells at the shield stage and adult cells of regenerating fin. Thus, the use of promoter elements from medaka is an alternative approach to generate Tgs with stronger and even novel expression patterns in zebrafish. The Olactb promoter and the Tg lines presented here represent an important advancement for the broader use of Cre/loxP-based Tg applications in zebrafish. PMID:23157381

Yoshinari, Nozomi; Ando, Kazunori; Kudo, Akira; Kinoshita, Masato; Kawakami, Atsushi

2012-12-01

300

Field performance of transgenic sugarcane produced using Agrobacterium and biolistics methods.  

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Future genetic improvement of sugarcane depends, in part, on the ability to produce high-yielding transgenic cultivars with improved traits such as herbicide and insect resistance. Here, transgenic sugarcane plants generated by different transformation methods were assessed for field performance over 3 years. Agrobacterium-mediated (Agro) transgenic events (35) were produced using four different Agrobacterium tumefaciens strains, while biolistic (Biol) transgenic events (48) were produced using either minimal linearized DNA (LDNA) transgene cassettes with 5', 3' or blunt ends or whole circular plasmid (PDNA) vectors containing the same transgenes. A combined analysis showed a reduction in growth and cane yield in Biol, Agro as well as untransformed tissue culture (TC) events, compared with the parent clone (PC) Q117 (no transformation or tissue culture) in the plant, first ratoon and second ratoon crops. However, when individual events were analysed separately, yields of some transgenic events from both Agro and Biol were comparable to PC, suggesting that either transformation method can produce commercially suitable clones. Interestingly, a greater percentage of Biol transformants were similar to PC for growth and yield than Agro clones. Crop ratoonability and sugar yield components (Brix%, Pol%, and commercial cane sugar (CCS)) were unaffected by transformation or tissue culture. Transgene expression remained stable over different crop cycles and increased with plant maturity. Transgene copy number did not influence transgene expression, and both transformation methods produced low transgene copy number events. No consistent pattern of genetic changes was detected in the test population using three DNA fingerprinting techniques. PMID:24330327

Joyce, Priya; Hermann, Scott; O'Connell, Anthony; Dinh, Quang; Shumbe, Leonard; Lakshmanan, Prakash

2014-05-01

 
 
 
 
301

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

Science.gov (United States)

Genetically modified mice have played an important part in elucidating gene function in vivo. However, conclusions from transgenic studies may be compromised by complications arising from the site of transgene integration into the genome and, in inducible systems, the non-innocuous nature of inducer molecules. The aim of the present study was to use the vascular system to validate a technique based on the bacterial lac operon system, in which transgene expression can be repressed and de-repressed by an innocuous lactose analogue, IPTG. We have modified an endothelium specific promoter (TIE2) with synthetic LacO sequences and made transgenic mouse lines with this modified promoter driving expression of mutant forms of connexin40 and an independently translated reporter, EGFP. We show that tissue specificity of this modified promoter is retained in the vasculature of transgenic mice in spite of the presence of LacO sequences, and that transgene expression is uniform throughout the endothelium of a range of adult systemic and cerebral arteries and arterioles. Moreover, transgene expression can be consistently down-regulated by crossing the transgenic mice with mice expressing an inhibitor protein LacI(R), and in one transgenic line, transgene expression could be de-repressed rapidly by the innocuous inducer, IPTG. We conclude that the modified bacterial lac operon system can be used successfully to validate transgenic phenotypes through a simple breeding schedule with mice homozygous for the LacI(R) protein. PMID:24755679

Morton, Susan K; Chaston, Daniel J; Baillie, Brett K; Hill, Caryl E; Matthaei, Klaus I

2014-01-01

302

An efficient and rapid transgenic pollen screening and detection method using flow cytometry.  

Science.gov (United States)

Assaying for transgenic pollen, a major vector of transgene flow, provides valuable information and essential data for the study of gene flow and assessing the effectiveness of transgene containment. Most studies have employed microscopic screening methods or progeny analyses to estimate the frequency of transgenic pollen. However, these methods are time-consuming and laborious when large numbers of pollen grains must be analyzed to look for rare transgenic pollen grains. Thus, there is an urgent need for the development of a simple, rapid, and high throughput analysis method for transgenic pollen analysis. In this study, our objective was to determine the accuracy of using flow cytometry technology for transgenic pollen quantification in practical application where transgenic pollen is not frequent. A suspension of non-transgenic tobacco pollen was spiked with a known amount of verified transgenic tobacco pollen synthesizing low or high amounts of green fluorescent protein (GFP). The flow cytometric method detected approximately 75% and 100% of pollen grains synthesizing low and high amounts of GFP, respectively. The method is rapid, as it is able to count 5000 pollen grains per minute-long run. Our data indicate that this flow cytometric method is useful to study gene flow and assessment of transgene containment. PMID:21154436

Moon, Hong S; Eda, Shigetoshi; Saxton, Arnold M; Ow, David W; Stewart, C Neal

2011-01-01

303

Affinity maturation leads to differential expression of multiple copies of a kappa light-chain transgene.  

Science.gov (United States)

Transgenic animals containing rearranged heavy or light chains are used to study the process of hypermutation, which characterizes the maturation of the antibody response. LK6 mice contain five copies of a transgene coding for a light chain produced in response to the hapten 2-phenyloxazolone. We have selected hybridomas from secondary responses that express the transgene as the only light chain. Some of these hybridomas contain transgene copies carrying mutations known to improve antibody affinity. We have analysed the expression of the five transgene copies in those hybridomas. We report here that the somatic hypermutation process can affect the successful expression of antibody light-chain transgenes. When mutations that improve the antibody affinity appear in one transgene copy, antigenic selection favours cells that downregulate the other copies at multiple levels of gene expression, including examples where nonsense mutations correlate with a drop in messenger RNA level. PMID:8487865

Lozano, F; Rada, C; Jarvis, J M; Milstein, C

1993-05-20

304

Transgenic plants expressing HC-Pro show enhanced virus sensitivity while silencing of the transgene results in resistance.  

Science.gov (United States)

Nicotiana benthamiana plants were engineered to express sequences of the helper component-proteinase (HC-Pro) of Cowpea aphid-borne mosaic potyvirus (CABMV). The sensitivity of the transgenic plants to infection with parental and heterologous viruses was studied. The lines expressing HC-Pro showed enhanced symptoms after infection with the parental CABMV isolate and also after infection with a heterologous potyvirus, Potato virus Y (PVY) and a comovirus, Cowpea mosaic virus (CPMV). On the other hand, transgenic lines expressing nontranslatable HC-Pro or translatable HC-Pro with a deletion of the central domain showed wild type symptoms after infection with the parental CABMV isolate and heterologous viruses. These results showed that CABMV HC-Pro is a pathogenicity determinant that conditions enhanced sensitivity to virus infection in plants, and that the central domain of the protein is essential for this. The severe symptoms in CABMV-infected HC-Pro expressing lines were remarkably followed by brief recovery and subsequent re-establishment of infection, possibly indicating counteracting effects of HC-Pro expression and a host defense response. One of the HC-Pro expressing lines (h48) was found to contain low levels of transgenic HC-Pro RNA and to be resistant to CABMV and to recombinant CPMV expressing HC-Pro. This indicated that h48 was (partially) posttranscriptionally silenced for the HC-Pro transgene inspite of the established role of HC-Pro as a suppressor of posttranscriptional gene silencing. Line h48 was not resistant to PVY, but instead showed enhanced symptoms compared to nontransgenic plants. This may be due to relief of silencing of the HC-Pro transgene by HC-Pro expressed by PVY. PMID:12206307

Mlotshwa, Sizolwenkosi; Verver, Jan; Sithole-Niang, Idah; Prins, Marcel; Van Kammen, A B; Wellink, Joan

2002-01-01

305

Hemolytic C-type lectin CEL-III from sea cucumber expressed in transgenic mosquitoes impairs malaria parasite development.  

Science.gov (United States)

The midgut environment of anopheline mosquitoes plays an important role in the development of the malaria parasite. Using genetic manipulation of anopheline mosquitoes to change the environment in the mosquito midgut may inhibit development of the malaria parasite, thus blocking malaria transmission. Here we generate transgenic Anopheles stephensi mosquitoes that express the C-type lectin CEL-III from the sea cucumber, Cucumaria echinata, in a midgut-specific manner. CEL-III has strong and rapid hemolytic activity toward human and rat erythrocytes in the presence of serum. Importantly, CEL-III binds to ookinetes, leading to strong inhibition of ookinete formation in vitro with an IC(50) of 15 nM. Thus, CEL-III exhibits not only hemolytic activity but also cytotoxicity toward ookinetes. In these transgenic mosquitoes, sporogonic development of Plasmodium berghei is severely impaired. Moderate, but significant inhibition was found against Plasmodium falciparum. To our knowledge, this is the first demonstration of stably engineered anophelines that affect the Plasmodium transmission dynamics of human malaria. Although our laboratory-based research does not have immediate applications to block natural malaria transmission, these findings have significant implications for the generation of refractory mosquitoes to all species of human Plasmodium and elucidation of mosquito-parasite interactions. PMID:18159942

Yoshida, Shigeto; Shimada, Yohei; Kondoh, Daisuke; Kouzuma, Yoshiaki; Ghosh, Anil K; Jacobs-Lorena, Marcelo; Sinden, Robert E

2007-12-01

306

DNA vaccine elicits an efficient antitumor response by targeting the mutant Kras in a transgenic mouse lung cancer model.  

Science.gov (United States)

Mutant Kras (V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog) is observed in more than 20% of non-small-cell lung cancers; however, no effective Kras target therapy is available at present. The Kras DNA vaccine may represent as a novel immunotherapeutic agent in lung cancer. In this study, we investigated the antitumor efficacy of the Kras DNA vaccine in a genetically engineered inducible mouse lung tumor model driven by Kras(G12D). Lung tumors were induced by doxycycline, and the therapeutic effects of Kras DNA vaccine were evaluated with delivery of Kras(G12D) plasmids. Mutant Kras(G12D) DNA vaccine significantly decreased the tumor nodules. A dominant-negative mutant Kras(G12D)N17, devoid of oncogenic activity, achieved similar therapeutic effects. The T-helper 1 immune response was enhanced in mice treated with Kras DNA vaccine. Splenocytes from mice receiving Kras DNA vaccine presented an antigen-specific response by treatment with peptides of Kras but not Hras or OVA. The number of tumor-infiltrating CD8(+) T cells increased after Kras vaccination. In contrast, Kras DNA vaccine was not effective in the lung tumor in transgenic mice, which was induced by mutant L858R epidermal growth factor receptor. Overall, these results indicate that Kras DNA vaccine produces an effective antitumor response in transgenic mice, and may be useful in treating lung cancer-carrying Ras mutation. PMID:25077772

Weng, T-Y; Yen, M-C; Huang, C-T; Hung, J-J; Chen, Y-L; Chen, W-C; Wang, C-Y; Chang, J-Y; Lai, M-D

2014-10-01

307

Glyoxalase-1 overexpression reduces endothelial dysfunction and attenuates early renal impairment in a rat model of diabetes  

DEFF Research Database (Denmark)

In diabetes, advanced glycation end-products (AGEs) and the AGE precursor methylglyoxal (MGO) are associated with endothelial dysfunction and the development of microvascular complications. In this study we used a rat model of diabetes, in which rats transgenically overexpressed the MGO-detoxifying enzyme glyoxalase-I (GLO-I), to determine the impact of intracellular glycation on vascular function and the development of early renal changes in diabetes.

Brouwers, Olaf; Niessen, Petra M G

2014-01-01

308

Immune selection of tumor cells in TCR ?-chain transgenic mice.  

Science.gov (United States)

Abstract The concept of immunological surveillance implies that immunogenic variants of tumor cells arising in the organism can be recognized by the immune system. Tumor progression is provided by somatic evolution of tumor cells under the pressure of the immune system. The loss of MHC Class I molecules on the surface of tumor cells is one of the most known outcomes of immune selection. This study developed a model of immune selection based on the immune response of TCR 1d1 single ?-chain transgenic B10.D2(R101) (K(d)I(d)D(b)) mice to allogeneic EL4 (H-2(b)) thymoma cells. In wild-type B10.D2(R101) mice, immunization with EL4 cells induced a vigorous CTL response targeted to the H-2K(b) molecule and results in full rejection of the tumor cells. In contrast, transgenic mice developed a compromised proliferative response in mixed-lymphocyte response assays and were unable to reject transplanted allogeneic EL4 cells. During the immune response to EL4 cells, CD8(+) T-lymphocytes with endogenous ?-chains accumulated predominantly in the spleen of transgenic mice and only a small part of the T-lymphocytes expressing transgenic ?-chains became CD8(+)CD44(+)CD62L(-) effectors. Then, instead of a full elimination of tumor cells as in wild-type mice, a reproducible prolonged equilibrium phase and subsequent escape was observed in transgenic mice that resulted in death of 90% of the mice in 40-60 days after grafting. Prolonged exposure of tumor cells to the pressure of the immune system in transgenic mice in vivo resulted in a stable loss of H-2K(b) molecules on the EL4 cell surface. Genetic manipulation of the T-lymphocyte repertoire was sufficient to reproduce the classic pattern of interactions between tumor cells and the immune system, usually observed in reliable syngeneic models of anti-tumor immunity. This newly-developed model could be used in further studies of immunoregulatory circuits common for transplantational and anti-tumor immune responses. PMID:24308870

Silaeva, Yulia Yu; Grinenko, Tatyana S; Vagida, Murad S; Kalinina, Anastasia A; Khromykh, Ludmila M; Kazansky, Dmitry B

2014-10-01

309

Improvement of spinal non-viral IL-10 gene delivery by D-mannose as a transgene adjuvant to control chronic neuropathic pain  

Science.gov (United States)

Background Peri-spinal subarachnoid (intrathecal; i.t.) injection of non-viral naked plasmid DNA encoding the anti-inflammatory cytokine, IL-10 (pDNA-IL-10) suppresses chronic neuropathic pain in animal models. However, two sequential i.t. pDNA injections are required within a discrete 5 to 72-hour period for prolonged efficacy. Previous reports identified phagocytic immune cells present in the peri-spinal milieu surrounding the i.t injection site that may play a role in transgene uptake resulting in subsequent IL-10 transgene expression. Methods In the present study, we aimed to examine whether factors known to induce pro-phagocytic anti-inflammatory properties of immune cells improve i.t. IL-10 transgene uptake using reduced naked pDNA-IL-10 doses previously determined ineffective. Both the synthetic glucocorticoid, dexamethasone, and the hexose sugar, D-mannose, were factors examined that could optimize i.t. pDNA-IL-10 uptake leading to enduring suppression of neuropathic pain as assessed by light touch sensitivity of the rat hindpaw (allodynia). Results Compared to dexamethasone, i.t. mannose pretreatment significantly and dose-dependently prolonged pDNA-IL-10 pain suppressive effects, reduced spinal IL-1? and enhanced spinal and dorsal root ganglia IL-10 immunoreactivity. Macrophages exposed to D-mannose revealed reduced proinflammatory TNF-?, IL-1?, and nitric oxide, and increased IL-10 protein release, while IL-4 revealed no improvement in transgene uptake. Separately, D-mannose dramatically increased pDNA-derived IL-10 protein release in culture supernatants. Lastly, a single i.t. co-injection of mannose with a 25-fold lower pDNA-IL-10 dose produced prolonged pain suppression in neuropathic rats. Conclusions Peri-spinal treatment with D-mannose may optimize naked pDNA-IL-10 transgene uptake for suppression of allodynia, and is a novel approach to tune spinal immune cells toward pro-phagocytic phenotype for improved non-viral gene therapy. PMID:24884664

2014-01-01

310

Protease inhibitors of Manduca sexta expressed in transgenic cotton.  

Science.gov (United States)

To explore the effectiveness of insect derived protease inhibitors in protecting plants against insect feeding, anti-trypsin, anti-chymotrypsin and anti-elastase protease inhibitor (PI) genes from Manduca sexta L. were expressed in transgenic cotton (Gossypium hirsutum L.). From 198 independent transformants, 35 elite lines were further analyzed. Under the control of the 35S promoter of CaMV, PI accumulated to approximately 0.1% of total protein, depending on the tissue analyzed. Using cell-flow cytometry, DNA content/ nuclei of transgenic and non-transformed cotton were identical. On cotton plants expressing PIs, fecundity of Bemisia tabaci (Genn.), the sweetpotato whitefly, was reduced compared to controls. Expression of these protease inhibitors may reduce the developmental rate of B. tabaci and other insects, and provide a strategy for cotton protection. PMID:24186707

Thomas, J C; Adams, D G; Keppenne, V D; Wasmann, C C; Brown, J K; Kanost, M R; Bohnert, H J

1995-10-01

311

Transgenic approaches for development of disease resistance in banana  

International Nuclear Information System (INIS)

Banana (Musa spp.) is an important food and cash crop worldwide. Diseases and pests pose the most serious constraint to banana cultivation. Among the diseases, Fusarium wilt and Banana Bunchy Top Virus (BBTV) are the most important economically. We have explored different transgenic approaches for development of efficient resistance in banana against these two diseases. For countering Fusarium wilt, we have over expressed Petunia floral defensins using a strong constitutive promoter in transgenic banana plants. We have also tested a host induced gene silencing strategy targeting two vital fungal genes to obtain Fusarium resistant banana plants. For development of BBTV resistant banana plants also, we have used a host-induced gene silencing approach utilizing the full and partial coding sequence of the viral replication initiation protein. Successful bioassays performed in controlled greenhouse conditions have shown the efficacy of using these strategies to develop disease resistant banana plants. (author)

312

Establishment and use of HBV-replication transgenic mice  

Directory of Open Access Journals (Sweden)

Full Text Available Despite the existence of a preventive vaccine,hepatitis B virus(HBV infection is still a major worldwide health problem,especially in China.As HBV naturally Despite of the existence of a preventive vaccine,hepatitis B virus(HBV infection is still a major worldwide healthy problem,especially in China.As HBV naturally infects only human and chimpanzees,many issues pertaining to the biology and the therapeutic of HBV infection remain unresolved due to the limitation of the establishment of a HBV model.However,the establishment of HBV-replication transgenic mice has greatly improved our understanding of life cycle,immunobiology and pathogensis of HBV.The establishment of HBV transgenic mice and its use in assessing the antiviral potential of pharmacological agents and HBV immunopathogenesis are herewith briefly described in the present paper.

Xiang-ping KONG

2011-09-01

313

Effects of Transgenic Rice on Life History Traits of Daphnia magna in Life Table Experiments  

Directory of Open Access Journals (Sweden)

Full Text Available To investigate the impacts of transgenic rice on freshwater organisms, we conducted two life tableexperiments using Daphnia magna for fifteen and twenty days, respectively. We examined life history traits suchas population growth rates (r, reproductive rates (R0, generation times, and survivorship. In the first experiment,we used non-drought-stressed transgenic and non-transgenic rice harvested in 2005. In the second study, weused non-transgenic and transgenic rice harvested in 2006 following drought stress. Each experiment involvedthree treatments in which D. magna neonates were fed with Selenastrum capricornutum (control treatment andS. capricornutum with 5% aqueous extracts of non-transgenic rice (N-T and transgenic rice (T. In the firstexperiment, D. magna showed reduced population growth rates and lowered fecundity in the N-T and Ttreatments. In the second experiment, D. magna receiving both transgenic and non-transgenic rice extractsshowed very high mortality, low population growth rates and reproduction rates. We could not detect anysignificant negative effects of extracts from transgenic rice on D. magna life history traits at 95%.

Nam, Sungjin

2007-11-01

314

FAS-Dependent Cell Death in ?-Synuclein Transgenic Oligodendrocyte Models of Multiple System Atrophy  

DEFF Research Database (Denmark)

Multiple system atrophy is a parkinsonian neurodegenerative disorder. It is cytopathologically characterized by accumulation of the protein p25? in cell bodies of oligodendrocytes followed by accumulation of aggregated ?-synuclein in so-called glial cytoplasmic inclusions. p25? is a stimulator of ?-synuclein aggregation, and coexpression of ?-synuclein and p25? in the oligodendroglial OLN-t40-AS cell line causes ?-synuclein aggregate-dependent toxicity. In this study, we investigated whether the FAS system is involved in ?-synuclein aggregate dependent degeneration in oligodendrocytes and may play a role in multiple system atrophy. Using rat oligodendroglial OLN-t40-AS cells we demonstrate that the cytotoxicity caused by coexpressing ?-synuclein and p25? relies on stimulation of the death domain receptor FAS and caspase-8 activation. Using primary oligodendrocytes derived from PLP-?-synuclein transgenic mice we demonstrate that they exist in a sensitized state expressing pro-apoptotic FAS receptor, which makes them sensitive to FAS ligand-mediated apoptosis. Immunoblot analysis shows an increase in FAS in brain extracts from multiple system atrophy cases. Immunohistochemical analysis demonstrated enhanced FAS expression in multiple system atrophy brains notably in oligodendrocytes harboring the earliest stages of glial cytoplasmic inclusion formation. Oligodendroglial FAS expression is an early hallmark of oligodendroglial pathology in multiple system atrophy that mechanistically may be coupled to ?-synuclein dependent degeneration and thus represent a potential target for protective intervention.

Kragh, Christine L; Fillon, Gwenaëlle

2013-01-01

315

Recent advances in development of marker-free transgenic plants: regulation and biosafety concern.  

Science.gov (United States)

During the efficient genetic transformation of plants with the gene of interest, some selectable marker genes are also used in order to identify the transgenic plant cells or tissues. Usually, antibiotic- or herbicide-selective agents and their corresponding resistance genes are used to introduce economically valuable genes into crop plants. From the biosafety authority and consumer viewpoints, the presence of selectable marker genes in released transgenic crops may be transferred to weeds or pathogenic microorganisms in the gastrointestinal tract or soil, making them resistant to treatment with herbicides or antibiotics, respectively. Sexual crossing also raises the problem of transgene expression because redundancy of transgenes in the genome may trigger homology-dependent gene silencing. The future potential of transgenic technologies for crop improvement depends greatly on our abilities to engineer stable expression of multiple transgenic traits in a predictable fashion and to prevent the transfer of undesirable transgenic material to non-transgenic crops and related species. Therefore, it is now essential to develop an efficient marker-free transgenic system. These considerations underline the development of various approaches designed to facilitate timely elimination of transgenes when their function is no longer needed. Due to the limiting number of available selectable marker genes, in future the stacking of transgenes will be increasingly desirable. The production of marker-free transgenic plants is now a critical requisite for their commercial deployment and also for engineering multiple and complex trait. Here we describe the current technologies to eliminate the selectable marker genes (SMG) in order to develop marker-free transgenic plants and also discuss the regulation and biosafety concern of genetically modified (GM) crops. PMID:22357214

Tuteja, Narendra; Verma, Shiv; Sahoo, Ranjan Kumar; Raveendar, Sebastian; Reddy, I N Bheema Lingeshwara

2012-03-01

316

Transgenic rabbit that expresses a functional human lipoprotein (a)  

Science.gov (United States)

A transgenic rabbit which has in its genomic DNA sequences that encode apolipoprotein (a) and apolipoprotein B polypeptides which are capable of combining to produce lipoprotein (a), a process for creating such a rabbit, and the use of the rabbit to identify compounds which are effective in the treatment of human diseases which are associated with, induced and/or exacerbated by Lp(a) expression.

Rouy, Didier (Thiais, FR); Duverger, Nicolas (Paris, FR); Emmanuel, Florence (Aubervilliers, FR); Denefle, Patrice (Saint Maur, FR); Houdebine, Louis-Marie (Buc, FR); Viglietta, Celine (Versailles, FR); Rubin, Edward M. (Berkeley, CA); Hughes, Steven D. (Oakland, CA)

2003-01-01

317

Restriction of neuroblastoma to the prostate gland in transgenic mice.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Male transgenic mice that carry a construct containing 5'-flanking sequences of the gp91-phox gene linked to the early region of the simian virus 40 (SV40) genome reproducibly develop tumors arising from the prostate gland. As gp91-phox is expressed exclusively in terminally differentiating hematopoietic cells of the myelomonocytic lineage, the induction of tumors arising from the prostate gland was unexpected. These lesions appear to be due to a novel transcription signal that was generated ...

Skalnik, D. G.; Dorfman, D. M.; Williams, D. A.; Orkin, S. H.

1991-01-01

318

Environmental and transgene expression effects on the barley seed proteome  

DEFF Research Database (Denmark)

The barley (Hordeum vulgare) cultivar Golden Promise is no longer widely used for malting, but is amenable to transformation and is therefore a valuable experimental cultivar. Its characteristics include high salt tolerance, however it is also susceptible to several fungal pathogens. Proteome analysis was used to describe the water-soluble protein fraction of Golden Promise seeds in comparison with the modern malting cultivar Barke. Using 2D-gel electrophoresis to visualise several hundred proteins in the pH ranges 4-7 and 6-11, 16 protein spots were found to differ between the two cultivars. Eleven of these were identified by mass spectrometric peptide mass mapping, including an abundant chitinase implicated in defence against fungal pathogens and a small heat-shock protein. To enable a comparison with transgenic seed protein patterns, differences in spot patterns between field and greenhouse-grown seeds were analysed. Four spots were observed to be increased in intensity in the proteome of greenhouse-grown seeds, three of which may be related to nitrogen availability during grain filling and total protein content of the seeds, since they also increased in field grown seeds supplied with extra nitrogen. Finally, the fate of transgene products in barley seeds was followed. Spots containing two green fluorescent protein constructs and the herbicide resistance marker phosphinothricin acetyltransferase were observed in 2D-gel patterns of transgenic seeds and identified by mass spectrometry. Phosphinothricin acetyltransferase was observed in three spots differing in pI suggesting that post-translational modification of the transgene product had occurred.

Finnie, Christine; Svensson, Birte

2004-01-01

319

Probing Pineal-specific Gene Expression with Transgenic Zebrafish†  

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The pineal gland of zebrafish (Danio rerio) contains lightsensitive photoreceptor cells and plays an important role in the neuroendocrine system. The zebrafish exorhodopsin gene encodes a pineal-specific photoreceptive protein, whose promoter region harbors a cis-acting element, pineal expression-promoting element (PIPE), directing pineal-specific gene expression. For in vivo genetic studies on PIPE-binding proteins and their regulatory mechanisms, we generated a transgenic zebrafish line, Tg...

Kojima, Daisuke; Dowling, John E.; Fukada, Yoshitaka

2008-01-01

320

Bacteriophage 5' untranslated regions for control of plastid transgene expression.  

Science.gov (United States)

Expression of foreign proteins from transgenes incorporated into plastid genomes requires regulatory sequences that can be recognized by the plastid transcription and translation machinery. Translation signals harbored by the 5' untranslated region (UTR) of plastid transcripts can profoundly affect the level of accumulation of proteins expressed from chimeric transgenes. Both endogenous 5' UTRs and the bacteriophage T7 gene 10 (T7g10) 5' UTR have been found to be effective in combination with particular coding regions to mediate high-level expression of foreign proteins. We investigated whether two other bacteriophage 5' UTRs could be utilized in plastid transgenes by fusing them to the aadA (aminoglycoside-3'-adenyltransferase) coding region that is commonly used as a selectable marker in plastid transformation. Transplastomic plants containing either the T7g1.3 or T4g23 5' UTRs fused to Myc-epitope-tagged aadA were successfully obtained, demonstrating the ability of these 5' UTRs to regulate gene expression in plastids. Placing the Thermobifida fusca cel6A gene under the control of the T7g1.3 or T4g23 5' UTRs, along with a tetC downstream box, resulted in poor expression of the cellulase in contrast with high-level accumulation while using the T7g10 5' UTR. However, transplastomic plants with the bacteriophage 5' UTRs controlling the aadA coding region exhibited fewer undesired recombinant species than plants containing the same marker gene regulated by the Nicotiana tabacum psbA 5' UTR. Furthermore, expression of the T7g1.3 and T4g23 5' UTR::aadA fusions downstream of the cel6A gene provided sufficient spectinomycin resistance to allow selection of homoplasmic transgenic plants and had no effect on Cel6A accumulation. PMID:23053542

Yang, Huijun; Gray, Benjamin N; Ahner, Beth A; Hanson, Maureen R

2013-02-01

 
 
 
 
321

Proteotypic classification of spontaneous and transgenic mammary neoplasms  

Science.gov (United States)

Introduction Mammary tumors in mice are categorized by using morphologic and architectural criteria. Immunolabeling for terminal differentiation markers was compared among a variety of mouse mammary neoplasms because expression of terminal differentiation markers, and especially of keratins, provides important information on the origin of neoplastic cells and their degree of differentiation. Methods Expression patterns for terminal differentiation markers were used to characterize tumor types and to study tumor progression in transgenic mouse models of mammary neoplasia (mice overexpressing Neu (Erbb2), Hras, Myc, Notch4, SV40-TAg, Tgfa, and Wnt1), in spontaneous mammary carcinomas, and in mammary neoplasms associated with infection by the mouse mammary tumor virus (MMTV). Results On the basis of the expression of terminal differentiation markers, three types of neoplasm were identified: first, simple carcinomas composed exclusively of cells with a luminal phenotype are characteristic of neoplasms arising in mice transgenic for Neu, Hras, Myc, Notch4, and SV40-TAg; second, 'complex carcinomas' displaying luminal and myoepithelial differentiation are characteristic of type P tumors arising in mice transgenic for Wnt1, neoplasms arising in mice infected by the MMTV, and spontaneous adenosquamous carcinomas; and third, 'carcinomas with epithelial to mesenchymal transition (EMT)' are a characteristic feature of tumor progression in Hras-, Myc-, and SV40-TAg-induced mammary neoplasms and PL/J and SJL/J mouse strains, and display de novo expression of myoepithelial and mesenchymal cell markers. In sharp contrast, EMT was not detected in papillary adenocarcinomas arising in BALB/cJ mice, spontaneous adenoacanthomas, neoplasms associated with MMTV-infection, or in neoplasms arising in mice transgenic for Neu and Wnt1. Conclusions Immunohistochemical profiles of complex neoplasms are consistent with a stem cell origin, whereas simple carcinomas might originate from a cell committed to the luminal lineage. In addition, these results suggest that the initiating oncogenic events determine the morphologic features associated with cancer progression because EMT is observed only in certain types of neoplasm. PMID:15535849

Mikaelian, Igor; Blades, Natalie; Churchill, Gary A; Fancher, Karen; Knowles, Barbara B; Eppig, Janan T; Sundberg, John P

2004-01-01

322

Development of phytase transgenic Nile tilapia (Oreochromis niloticus)  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The feeding of plant proteins to fish results in generation of a high amount of phytate because fish cannot effectively digest it. The hydrolysis of phytate by microbes in the environment release P which can stimulate eutrophication. This study was conducted to express phytase gene from fungi, Aspergillus niger in the gastrointestinal tract of tilapia and to conduct feeding trials with the transgenic fish produced. The DNA construct used consist of CMV promoter, secretion signal, phy A gene a...

Kemeh, Settor

2004-01-01

323

Transgenic control of vectors: the effects of interspecific interactions  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The control of insect vectors through conventional sterile insect or transgenic technologies (e.g., RIDL®) is an intense focus of research in the combat against vector-borne disease. While the population dynamic implications of these control strategies are reasonably well-established, the effects of interspecific competition between different vectors and control strategies have not previously been explored. Different control intervention methods can affect the interaction and potential coexi...

Bonsall, Mb; Yakob, L.; Alphey, N.; Alphey, L.

2010-01-01

324

Permeability barrier dysfunction in transgenic mice overexpressing claudin 6.  

Science.gov (United States)

A defective epidermal permeability barrier (EPB) in premature birth remains a leading cause of neonatal death as a result of its associated complications, which include poor temperature stability, infection by micro-organisms through the skin, and the outflow of water. Despite its importance in survival, the mechanisms involved in the formation and maintenance of the EPB are not well understood. To address the possibility that claudins, a new superfamily of tight junctional molecules, are involved, we engineered transgenic mice with claudin 6 (Cldn6) overexpressed via the involucrin (Inv) promoter. Interestingly, the Inv-Cldn6 transgenic animals die within 2 days of birth, apparently due to the lack of an intact EPB as evidenced by increased water loss and the penetration of X-gal through the skin. Barrier dysfunction was manifested biochemically by the aberrant expression of late epidermal differentiation markers, including K1, filaggrin, loricrin, transglutaminase 3, involucrin, repetin, members of the SPRR family and the transcriptional regulator Klf4. The overall claudin profile of the epidermis was also modified. Our data suggest that repetin and SPRR1A and 2A are downregulated in response to the downregulation of Klf4 in the transgenic animals, which would contribute to decreased protein crossbridging leading to fragile, defective cornified envelopes. These results provide new insights into the role of claudin 6 in epithelial differentiation and EPB formation. In addition, the epidermal phenotype of these transgenic mice, which is very reminiscent of that in pre-term infant skin, suggest that they will be an important and novel model for studies on human premature EPB-related morbidity. PMID:11923212

Turksen, Kursad; Troy, Tammy-Claire

2002-04-01

325

Spinal cord pathology in alpha-synuclein transgenic mice  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Accumulation of ?-synuclein is observed in neurodegenerative diseases like Parkinson's disease and Multiple System Atrophy. In previous studies with transgenic C57BL/6 mice overexpressing ?-synuclein carrying the mutations A53T and A30P found in Parkinson's disease or with a parkin-null background, we reported severe mitochondrial impairments in neurons and to a larger extent in glial cells of the mesencephalon. Neuron death was not observed in the brain. Here we show that the mice show sev...

Amp Bbert, Hermann L.; Daniel Segelcke; Xin-Ran Zhu; Teresa Sczepan; Sonja Mendritzki; Saskia Schmidt

2010-01-01

326

Optical modulation of transgene expression in retinal pigment epithelium  

Science.gov (United States)

Over a million people in US alone are visually impaired due to the neovascular form of age-related macular degeneration (AMD). The current treatment is monthly intravitreal injections of a protein which inhibits Vascular Endothelial Growth Factor, thereby slowing progression of the disease. The immense financial and logistical burden of millions of intravitreal injections signifies an urgent need to develop more long-lasting and cost-effective treatments for this and other retinal diseases. Viral transfection of ocular cells allows creation of a "biofactory" that secretes therapeutic proteins. This technique has been proven successful in non-human primates, and is now being evaluated in clinical trials for wet AMD. However, there is a critical need to down-regulate gene expression in the case of total resolution of retinal condition, or if patient has adverse reaction to the trans-gene products. The site for genetic therapy of AMD and many other retinal diseases is the retinal pigment epithelium (RPE). We developed and tested in pigmented rabbits, an optical method to down-regulate transgene expression in RPE following vector delivery, without retinal damage. Microsecond exposures produced by a rapidly scanning laser vaporize melanosomes and destroy a predetermined fraction of the RPE cells selectively. RPE continuity is restored within days by migration and proliferation of adjacent RPE, but since the transgene is not integrated into the nucleus it is not replicated. Thus, the decrease in transgene expression can be precisely determined by the laser pattern density and further reduced by repeated treatment without affecting retinal structure and function.

Palanker, D.; Lavinsky, D.; Chalberg, T.; Mandel, Y.; Huie, P.; Dalal, R.; Marmor, M.

2013-03-01

327

Gene expression in the mammary glands of transgenic animals.  

Science.gov (United States)

The gene encoding the milk protein beta-lactoglobulin in sheep is expressed in the mammary gland in a tissue-specific manner during pregnancy and lactation. The unmodified sheep gene behaves appropriately in transgenic mice, and we have shown that many of the cis-acting elements that mediate this pattern of expression are located in the proximal 400 bp of the promotor. Using a combination of approaches we have identified a number of discrete cis-acting elements and their corresponding trans-acting factors that control the responsiveness of this gene in vivo. The beta-lactoglobulin promoter elements can be used to target the expression of foreign genes to the mammary gland in transgenic mice. We have used this approach in basic studies of mammary gland biology and for the production of therapeutic proteins in the milk of transgenic animals. In these circumstances, however, the promoter rarely functions optimally, and it may even be silenced; consequently, we have had to develop a number of strategies to overcome this problem. Nevertheless, foreign proteins do appear to be appropriately post-translationally modified when they are expressed in the mammary gland. PMID:9513717

Clark, A J

1998-01-01

328

[Implications of transgenics for environmental and agricultural sustainability].  

Science.gov (United States)

The potential risks of GMOs, their impact on human and animal health, and on the environment, as well as their socioeconomic effects, have generated a worldwide discussion which is far from drawing to a close for lack of sufficient information. Part of this information supports risk-hypotheses previously put forward. Thus the presence of transgenic plant genes in other plants and in other organisms has been confirmed in several occasions. Therefore, gene dissemination to plants of the same species as well as to widely different species is already regarded as an actual risk. The principle of substantial equivalence has opened the way for the liberation of transgenic plants for commercial crops, despite short-term tests, which are quantitatively and qualitatively insufficient to certify that the foods deriving from those plants are healthy and safe. Thus, the adoption of the so-called precautionary principle (PP) has turned out to be the most adequate safety measure to date, or else until scientific data should be able to demonstrate the actual impact of transgenic plants on human and animal health, and on the environment. PMID:16680899

Nodari, R O; Guerra, M P

2000-01-01

329

Elucidation of Factors Effecting Enzymatic Saccharification using Transgenic Hardwoods  

Science.gov (United States)

Three groups of transgenic wood samples were used as starting materials to elucidate the recalcitrance of enzymatic saccharification with/without pretreatments. The first group of transgenic wood samples is low lignin P. trichocarpa. The second group is low xylan P. trichocarpa. The third one is 12 hybrid poplars which have different levels of S/V ratio and lignin content. Four pretreatments were carried out in this research including dilute sulfuric acid, green liquor, auto hydrolysis and ozone delignification. The behavior among pretreatments as a function of removal of lignin appears to be different. Lignin is the major factor of recalcitrance of the lignocellulosic material to ethanol conversion process. Xylan also plays key role in this process. In addition, the crude milled wood lignin was isolated from these three groups of transgenic samples. Lignin carbohydrate complexes was characterized by 1H-13C HMQC and 13C NMR. Thus the effect of LCCs on enzymatic saccharification was elucidated. High S/V ratio propels the lignin removal during pretreatments however; high S/V ratio retards the enzymatic saccharification on the lignocellulosic material without pretreatments. The level of LCCs linkages accounts for additional recalcitrance of the lignocellulosic material to ethanol conversion process. The amount of LCCs linkages is affected by xylan content, lignin content and S/V ratio.

Min, Douyong

330

Early flowering and genetic containment studies in transgenic poplar  

Directory of Open Access Journals (Sweden)

Full Text Available Despite of the immense potential of gene technologies for tree breeding, release of genetic modified trees is still very rare. Biosafety concerns have hitherto limited application of gene technologies. The potential risks of transgenic trees, in particular transfer of recombinant DNA into the gene pool of a given species via vertical gene transfer, have been motive of concern. Biosafety research may allow avoiding potential risks of this technology. However, the evaluation of strategies for prevention of vertical gene transfer, probably the most important concern toward transgenic trees, has been hindered by the long time they require to reach the reproductive phase. We tested different strategies for promoting early flowering in poplar, aiming the development of a system for biosafety studies on gene containment. Early flowering poplar containing the 35S::LFY or HSP::FT gene constructs allowed first approaches for the faster evaluation of gene containment. However, some drawbacks, e.g., disturbed vegetative growth and flower development, still limit their potential application on biosafety research. A non-transgenic hybrid aspen showing a short vegetative phase was successfully used for the evaluation of the PrMALE1::STS sterility gene construct.

Fladung M

2012-06-01

331

Mouse transgenes in human cells detect specific base substitutions  

International Nuclear Information System (INIS)

The authors describe a system of transgenic human cell lines that detects and identifies specific point mutations at defined positions within a gene. The target transgenome is a mouse adenine phosphoribosyltransferase (APRT) gene rendered nonfunctional by introduction of a substitution at either of two bases that comprise a splice acceptor site. Reversion at a mutated site results in the expression of wild-type mouse APRT and consequent growth of APRT+ transgenic cell colonies. Site-specific reversion to wild-type sequence is confirmed by regeneration of a previously destroyed diagnostic Pst I site. Two independent cell clones, each with mutant transgenomes bearing an A ? G transition, exhibited an up to 7,500-fold, dose-dependent induction of reversion following treatment with ethyl methanesulfonate. Treatment of these clones with 2-aminopurine resulted in no induction of revertants. In contrast, another transgenic cell clone, bearing a G ? A transition, reverted as a consequence of 2-aminopurine, but not ethyl methanesulfonate, treatment. These data confirm for human cells the proposed mechanisms of action of these mutagens and provide evidence for the utility of their site-specific reversion method for mutagenesis studies

332

Generation and characterization of alpha-chymase-Cre transgenic mice.  

Science.gov (United States)

The Cre-loxP technology allows the introduction of somatic gene alterations in a tissue and/or cell type specific manner. The development of transgenes that target Cre expression to specific cell types is a critical component in this system. Here, we describe the generation and characterization of transgenic mouse lines expressing Cre recombinase under the control of the baboon alpha-chymase promoter, designated Chm:Cre, in order to direct Cre expression specifically to mouse mast cells. Chm:Cre expression was detected in mast cells in lung and colon tissue. Cre-mediated recombination in these mice identified a population of mature tissue resident mast cells using ROSA26R reporter mice. No Cre-expression and Cre-mediated recombination was induced in in vitro generated bone marrow derived mast cells or mast cells isolated from the peritoneal cavity indicating that Cre-expression under the control of the alpha-chymase promoter is solely activated in tissue resident mast cells. These Chm:Cre transgenic mice represent a useful tool to specifically inactivate genes of interest in mast cells of these tissues. PMID:18327770

Müsch, Werner; Wege, Anja K; Männel, Daniela N; Hehlgans, Thomas

2008-03-01

333

Intragenesis and cisgenesis as alternatives to transgenic crop development  

DEFF Research Database (Denmark)

One of the major concerns of the general public about transgenic crops relates to the mixing of genetic materials between species that cannot hybridize by natural means. To meet this concern, the two transformation concepts cisgenesis and intragenesis were developed as alternatives to transgenesis. Both concepts imply that plants must only be transformed with genetic material derived from the species itself or from closely related species capable of sexual hybridization. Furthermore, foreign sequences such as selection genes and vector-backbone sequences should be absent. Intragenesis differs from cisgenesis by allowing use of new gene combinations created by in vitro rearrangements of functional genetic elements. Several surveys show higher public acceptance of intragenic/cisgenic crops compared to transgenic crops. Thus, although the intragenic and cisgenic concepts were introduced internationally only 9 and 7 years ago, several different traits in a variety of crops have currently been modified according to these concepts. Five of these crops are now in field trials and two have pending applications for deregulation. Currently, intragenic/cisgenic plants are regulated as transgenic plants worldwide. However, as the gene pool exploited by intragenesis and cisgenesis are identical to the gene pool available for conventional breeding, less comprehensive regulatory measures are expected. The regulation of intragenic/cisgenic crops is presently under evaluation in the EU and in the US regulators are considering if a subgroup of these crops should be exempted from regulation. It is accordingly possible that the intragenic/cisgenic route will be of major significance for future plant breeding.

Holme, Inger; Wendt, Toni

2013-01-01

334

Transgenic rodent assay for quantifying male germ cell mutant frequency.  

Science.gov (United States)

De novo mutations arise mostly in the male germline and may contribute to adverse health outcomes in subsequent generations. Traditional methods for assessing the induction of germ cell mutations require the use of large numbers of animals, making them impractical. As such, germ cell mutagenicity is rarely assessed during chemical testing and risk assessment. Herein, we describe an in vivo male germ cell mutation assay using a transgenic rodent model that is based on a recently approved Organisation for Economic Co-operation and Development (OECD) test guideline. This method uses an in vitro positive selection assay to measure in vivo mutations induced in a transgenic ?gt10 vector bearing a reporter gene directly in the germ cells of exposed males. We further describe how the detection of mutations in the transgene recovered from germ cells can be used to characterize the stage-specific sensitivity of the various spermatogenic cell types to mutagen exposure by controlling three experimental parameters: the duration of exposure (administration time), the time between exposure and sample collection (sampling time), and the cell population collected for analysis. Because a large number of germ cells can be assayed from a single male, this method has superior sensitivity compared with traditional methods, requires fewer animals and therefore much less time and resources. PMID:25145276

O'Brien, Jason M; Beal, Marc A; Gingerich, John D; Soper, Lynda; Douglas, George R; Yauk, Carole L; Marchetti, Francesco

2014-01-01

335

Transgenic plants expressing HC-Pro show enhanced virus sensitivity while silencing of the transgene results in resistance  

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Nicotiana benthamiana plants were engineered to express sequences of the helper component-proteinase (HC-Pro) of Cowpea aphid-borne mosaic potyvirus (CABMV). The sensitivity of the transgenic plants to infection with parental and heterologous viruses was studied. The lines expressing HC-Pro showed enhanced symptoms after infection with the parental CABMV isolate and also after infection with a heterologous potyvirus, Potato virus Y (PVY) and a comovirus, Cowpea mosaic virus (CPMV). On the oth...

Mlotshwa, S.; Verver, J.; Sithole-niang, I.; Prins, M.; Kammen, A.; Wellink, J.

2002-01-01

336

[Expression of recombinant proteins in the milk of transgenic animals].  

Science.gov (United States)

The bulky production of recombinant proteins can be achieved by procaryotes or eucaryotes cells. Cells from higher eucaryotes may be required when proteins have to be modified post-transcriptionally (glycosylation phosphorylation, cleavage, folding...). Cells from higher vertebrates in culture are used to prepare proteins like human factor VIII and erythropoietin. The use of transgenic organism has been suggested to reach the same goal. Indeed a whole living organism allows a very potent amplification, the number of cells involved in the biosynthesis of the recombinant proteins being very numerous and in the best metabolic conditions. Biological fluids (blood, milk, insect hemolymph, egg white...) and possibly organs from transgenic animals are a priori the best sources of recombinant proteins. Blood is abundant and it is a by-product of slaughter house. Its composition is relatively complex and the circulating recombinant proteins may heavily alter health of animals. Milk is very abundant, its composition is relatively simple, it is poor in proteolytic enzymes and it can be collected easily. Hemolymph from insects is relatively scarce. Egg white will be a possible source of recombinant proteins, when transgenesis has become more accessible in birds. Organs from transgenic animals should be solicited only when a particular cell type is required for the biosynthesis of the recombinant proteins. Milk appears therefore, presently, as the best source of recombinant proteins from transgenic animals. About 15 public and private laboratories try to use these techniques. They consist in preparing vectors containing regulatory regions of one of the milk proteins genes and the coding part (cDNA or gene) of the corresponding proteins to be produced. The transfer of these gene constructs to mouse, rabbit, sheep, goat, pig, shows that these techniques are indeed very promising. A single protein, human alpha 1-antitrypsin produced in milk of transgenic sheep, has presently reached the preparation at an industrial scale. This method has two theoretical limitations: 1) some of the proteins secreted in milk may be not matured as their native counterparts. Experiments carried out so far (about 20 proteins has been produced at an experimental scale) indicate that the mammary cell is able to achieve glycosylation in a correct way; 2) a significant proportion of the recombinant proteins migrate from the alveolar compartment of the mammary gland to blood circulation and they can alter health of lactating animals.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:8476491

Houdebine, L M

1993-01-01

337

Expression of the Huntington's disease transgene in neural stem cell cultures from R6/2 transgenic mice.  

Science.gov (United States)

Huntington's disease (HD) is an inherited neurodegenerative disorder resulting in neuronal cell death in discrete brain regions due to an expanded CAG repeat of the huntingtin gene. The transgenic mouse model R6/2 expresses exon 1 of the human huntingtin gene with >150 CAG repeats, which produces mutant HD protein with an expanded poly-glutamine tract. We have established a neuronal stem cell system deriving from transgenic HD R6/2 neonatal brains as a renewable source for neurons and glia to facilitate studies of HD neuropathology and therapies. These R6/2 stem cell cultures can be cryopreserved and revived. Thawed neural progenitors can be expanded, established as continuous cell lines, and induced to differentiate into glia and neurons. Using standard culture conditions, there was no detectable morphological difference between wild type and HDR6/2 cells. Western analysis reveals that R6/2, but not wild type neurospheres, express the expanded repeat transgenic protein. Immunocytochemistry reveals that at a higher antibody concentration, huntingtin can be localized in the nucleus and the cytoplasm of wild type and R6/2 cells. We conclude that the R6/2 neuronal stem cell culture is a valuable tool for investigating HD pathogenesis and potential genetic or pharmacological interventions. PMID:11719265

Chu-LaGraff, Q; Kang, X; Messer, A

338

Production of brown/yellow patches in the SLC7A11 transgenic sheep via testicular injection of transgene.  

Science.gov (United States)

The gene, SLC7A11, which encodes the solute carrier family 7 member 11 (anionic amino acid transporter light chain, xCT), has been reported to be implicated in multiple processes such as in pheomelanin production, cell proliferation and migration, Kaposi's sarcoma herpesvirus (KSHV) entry into the host cells, learning and memory. Its involvement in cancer cell proliferation and metastasis has been widely studied. Its role in pheomelanogenesis is likely conserved in sheep. The full-length cDNA of sheep SLC7A11 was cloned from sheep skin fibroblasts for evaluating its role in regulating sheep coat color. The complete open reading frame of sheep xCT (sxCT) is 1512 bp in length, encoding a 503 amino acid polypeptide. We explored its function on pheomelanogenesis in vitro and in vivo. In the melan-a non-agouti mouse melanocytes that mainly produce eumelanin, overexpressed sxCT reduced the content of eumelanin. Using a testicular injection transgenic method, sxCT-transgenic sheep were generated and exhibited patches of brown/yellow coat, suggesting that sxCT can be selectively expressed to increase the pheomelanin production in wool. Our studies suggest that testicular injection of transgene can be used to genetically modify sheep coat color. PMID:22749016

He, Xin; Li, Hongtao; Zhou, Zhiyong; Zhao, Zongsheng; Li, Wei

2012-06-20

339

Transgenic plants expressing GLK1 and CCA1 having increased nitrogen assimilation capacity  

Science.gov (United States)

Provided herein are compositions and methods for producing transgenic plants. In specific embodiments, transgenic plants comprise a construct comprising a polynucleotide encoding CCA1, GLK1 or bZIP1, operably linked to a plant-specific promote, wherein the CCA1, GLK1 or bZIP1 is ectopically overexpressed in the transgenic plants, and wherein the promoter is optionally a constitutive or inducible promoter. In other embodiments, transgenic plants in which express a lower level of CCA1, GLK1 or bZIP1 are provided. Also provided herein are commercial products (e.g., pulp, paper, paper products, or lumber) derived from the transgenic plants (e.g., transgenic trees) produced using the methods provided herein.

Coruzzi, Gloria (New York, NY); Gutierrez, Rodrigo A. (Santiago, CL); Nero, Damion C. (Woodside, NY)

2012-04-10

340

Visceral obesity without insulin resistance in late-onset obesity rats.  

Science.gov (United States)

We describe a line of transgenic rats in which the males develop a unique autosomal dominant, late-onset obesity (LOB) phenotype. LOB males gradually accumulate fat specifically in visceral, but not peripheral, fat depots despite a normal intake of a low fat diet. LOB females normally develop only mild obesity with advanced age. However, the phenotype can be induced rapidly in young females by ovariectomy and prevented by estrogen replacement. LOB males are highly sensitive to dietary fat. Young, nonobese LOB males gain more weight on a 30% fat diet and lose more weight when treated with the lipase inhibitor, Orlistat, than their nontransgenic littermates. Remarkably, despite severe visceral obesity, LOB rats have normal fasting blood glucose, insulin, and corticosterone; show normal or increased insulin sensitivity in glucose and insulin tolerance tests; have increased plasma adiponectin levels; and display a heightened response to treatment with rosiglitazone. Their visceral adiposity reflects a specific increase in visceral adipocyte number, not size. Analysis of the transgene in LOB rats revealed a deletion in the gene encoding the S26 subunit of the mitochondrial ribosome that results in the production of a truncated protein, which we show to be imported into mitochondria. However, the transgene integrant is complex, so whether this is the sole molecular disruption underlying this phenotype remains to be established. Nevertheless, LOB rats provide a valuable new model of late-onset, male-preponderant, visceral-specific obesity, clearly dissociated from insulin resistance. PMID:15033913

Bains, Randip K; Wells, Sara E; Flavell, Dave M; Fairhall, Keith M; Strom, Molly; Le Tissier, Paul; Robinson, Iain C A F

2004-06-01

 
 
 
 
341

Repression of retrovirus-mediated transgene expression by interferons: implications for gene therapy.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Retrovirus-mediated gene transfer is commonly used in gene therapy protocols and has the potential to provide long-term expression of the transgene. Although expression of a retrovirus-delivered transgene is satisfactory in cultured cells, it has been difficult to achieve consistent and high-level expression in vivo. In this investigation, we explored the possibility of modulating transgene expression by host-derived cytokines. Normal human keratinocytes and dermal fibroblasts were transduced...

Ghazizadeh, S.; Carroll, J. M.; Taichman, L. B.

1997-01-01

342

Development of transgenic Ambystoma mexicanum (axolotl) to study cell fate during development and regeneration  

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The establishment of transgenesisi in axolotls is crucial for studying development and regeneration, as it would allow for long-term fate tracing as well as gene expression analysis, therefore we were interested in both obtaining animals expresing the transgene with little mosaicism in F0 generation and transgenesis. We demonstrate here that plasmid injection into one cell stage axolotl embryo generates transgenic animals that display germline transmission of a transgene. However, the efficie...

Sobkow, Lidia

2006-01-01

343

Mutations in the pqe-1 gene enhance transgene expression in Caenorhabditis elegans.  

Science.gov (United States)

Although various genetic tools have been developed and used as transgenes, the expression of the transgenes often is hampered by negative regulators. Disrupting such negative regulators of gene expression is potentially a way to overcome the common problem of low expression of transgenes. To find such regulators whose mutations enhance transgene expression in Caenorhabditis elegans, we took advantage of a newly developed reporter transgene, lin-11pA?::venus. This transgene induces expression of a fluorescent protein, Venus, in specific neurons including AIZ, where the expression was stochastic. The frequency of reporter expression in AIZ seemed to be correlated with the strength of transgene expression. By using this system, in which a moderate increase of expression was converted to all-or-none expression states, we describe here a forward genetic screen for mutations that enhance the expression of transgenes. Through the screen, we found that mutations in the pqe-1 gene, which encodes a Q/P-rich nuclear protein with an exonuclease domain, increase the chance of reporter expression in AIZ. The fluorescence intensity in RIC, in which all lin-11pA?::venus animals show reporter expression, was increased in pqe-1 mutants, suggesting that pqe-1 reduces the expression level of the transgene. Expression of transgenes with other promoters, 3'UTR, or reporter genes was also enhanced by the pqe-1 mutation, suggesting that the effect was not specific to a particular type of transgenes, whereas the effect did not seem to extend to endogenous genes. We propose that pqe-1 mutants can be used to increase the expression of various useful transgenes. PMID:22870397

Yamada, Koji; Tsuchiya, Jun-ichi; Iino, Yuichi

2012-07-01

344

The polyomavirus early region gene in transgenic mice causes vascular and bone tumors.  

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Transgenic mice carrying the entire polyomavirus (Py) early region consistently develop both vascular and bone tumors. This tumor spectrum represents a subset of the tumors found in mice infected with Py and an expansion of the vascular tumor spectrum seen in Py middle T antigen (MT) transgenic mice (V. L. Bautch, S. Toda, J. A. Hassell, and D. Hanahan, Cell 51:529-538, 1987). Transgenic mice of three independent lineages develop these pathologies, and mice of individual lineages also develop...

Wang, R.; Bautch, V. L.

1991-01-01

345

Glomerulosclerosis in mice transgenic for Insulin-like Growth Factor Binding Protein-1  

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Glomerulosclerosis in mice transgenic for human insulin-like growth factor-binding protein-1. Background The growth hormone (GH)/insulin-like growth factor (IGF) system is thought to participate in the glomerulosclerosis process. Because IGF-binding proteins (IGFBPs) modulate IGF actions and hence GH secretion, this study assessed whether mice transgenic for human IGFBP-1 have altered susceptibility to glomerulosclerosis. Methods A line of transgenic mice that express human IGFBP-1 mRN...

Doublier, Sophie Michelle

2000-01-01

346

Neuroblastoma in a transgenic mouse carrying a metallothionein/ret fusion gene.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We have recently succeeded in producing transgenic mice carrying a hybrid gene consisting of mouse metallothionein promoter-enhancer and the ret oncogene (MT/ret). (Iwamoto et al., 1991b). A retroperitoneal tumour developed in one of 17 MT/ret transgenic founder mice. Histological analysis revealed that the tumour consisted of undifferentiated neuroblasts and differentiated ganglion cells, the latter of which were strongly positive for neuron specific enolase. Expression of the ret transgene ...

Iwamoto, T.; Taniguchi, M.; Wajjwalku, W.; Nakashima, I.; Takahashi, M.

1993-01-01

347

Vldlr overexpression causes hyperactivity in rats  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Reelin regulates neuronal positioning in cortical brain structures and neuronal migration via binding to the lipoprotein receptors Vldlr and Lrp8. Reeler mutant mice display severe brain morphological defects and behavioral abnormalities. Several reports have implicated reelin signaling in the etiology of neurodevelopmental and psychiatric disorders, including autism, schizophrenia, bipolar disorder, and depression. Moreover, it has been reported that VLDLR mRNA levels are increased in the post-mortem brain of autistic patients. Methods We generated transgenic (Tg rats overexpressing Vldlr, and examined their histological and behavioral features. Results Spontaneous locomotor activity was significantly increased in Tg rats, without detectable changes in brain histology. Additionally, Tg rats tended to show performance deficits in the radial maze task, suggesting that their spatial working memory was slightly impaired. Thus, Vldlr levels may be involved in determining locomotor activity and memory function. Conclusions Unlike reeler mice, patients with neurodevelopmental or psychiatric disorders do not show striking neuroanatomical aberrations. Therefore, it is notable, from a clinical point of view, that we observed behavioral phenotypes in Vldlr-Tg rats in the absence of neuroanatomical abnormalities.

Iwata Keiko

2012-10-01

348

Production of human CD59-transgenic pigs by embryonic germ cell nuclear transfer  

Energy Technology Data Exchange (ETDEWEB)

Research highlights: {yields} Human CD59 (hCD59) gene was introduced into porcine embryonic germ (EG) cells. {yields} hCD59-transgenic EG cells were resistant to hyperacute rejection in cytolytic assay. {yields} hCD59-transgenic pigs were produced by EG cell nuclear transfer. -- Abstract: This study was performed to produce transgenic pigs expressing the human complement regulatory protein CD59 (hCD59) using the nuclear transfer (NT) of embryonic germ (EG) cells, which are undifferentiated stem cells derived from primordial germ cells. Because EG cells can be cultured indefinitely in an undifferentiated state, they may provide an inexhaustible source of nuclear donor cells for NT to produce transgenic pigs. A total of 1980 NT embryos derived from hCD59-transgenic EG cells were transferred to ten recipients, resulting in the birth of fifteen piglets from three pregnancies. Among these offspring, ten were alive without overt health problems. Based on PCR analysis, all fifteen piglets were confirmed as hCD59 transgenic. The expression of the hCD59 transgene in the ten living piglets was verified by RT-PCR. Western analysis showed the expression of the hCD59 protein in four of the ten RT-PCR-positive piglets. These results demonstrate that hCD59-transgenic pigs could effectively be produced by EG cell NT and that such transgenic pigs may be used as organ donors in pig-to-human xenotransplantation.

Ahn, Kwang Sung; Won, Ji Young [Department of Physiology, Dankook University School of Medicine, Cheonan (Korea, Republic of); Park, Jin-Ki [Animal Biotechnology Division, National Institute of Animal Science, Suwon (Korea, Republic of); Sorrell, Alice M. [Department of Physiology, Dankook University School of Medicine, Cheonan (Korea, Republic of); Heo, Soon Young; Kang, Jee Hyun [Department of Nanobiomedical Science, Dankook University, Cheonan (Korea, Republic of); Woo, Jae-Seok [Animal Biotechnology Division, National Institute of Animal Science, Suwon (Korea, Republic of); Choi, Bong-Hwan [Genomics and Bioinformatics Division, National Institute of Animal Science, Suwon (Korea, Republic of); Chang, Won-Kyong [Animal Biotechnology Division, National Institute of Animal Science, Suwon (Korea, Republic of); Shim, Hosup, E-mail: shim@dku.edu [Department of Nanobiomedical Science, Dankook University, Cheonan (Korea, Republic of); Institute of Tissue Regeneration Engineering, Dankook University, Cheonan (Korea, Republic of)

2010-10-01

349

Production of human CD59-transgenic pigs by embryonic germ cell nuclear transfer  

International Nuclear Information System (INIS)

Research highlights: ? Human CD59 (hCD59) gene was introduced into porcine embryonic germ (EG) cells. ? hCD59-transgenic EG cells were resistant to hyperacute rejection in cytolytic assay. ? hCD59-transgenic pigs were produced by EG cell nuclear transfer. -- Abstract: This study was performed to produce transgenic pigs expressing the human complement regulatory protein CD59 (hCD59) using the nuclear transfer (NT) of embryonic germ (EG) cells, which are undifferentiated stem cells derived from primordial germ cells. Because EG cells can be cultured indefinitely in an undifferentiated state, they may provide an inexhaustible source of nuclear donor cells for NT to produce transgenic pigs. A total of 1980 NT embryos derived from hCD59-transgenic EG cells were transferred to ten recipients, resulting in the birth of fifteen piglets from three pregnancies. Among these offspring, ten were alive without overt health problems. Based on PCR analysis, all fifteen piglets were confirmed as hCD59 transgenic. The expression of the hCD59 transgene in the ten living piglets was verified by RT-PCR. Western analysis showed the expression of the hCD59 protein in four of the ten RT-PCR-positive piglets. These results demonstrate that hCD59-transgenic pigs could effectively be produced by EG cell NT and that such transgenic pigs may be used as organ donors in pig-to-human xenotransplantation.

350

MicroRNA-150-regulated vectors allow lymphocyte-sparing transgene expression in hematopoietic gene therapy.  

Science.gov (United States)

Endogenous microRNA (miRNA) expression can be exploited for cell type-specific transgene expression as the addition of miRNA target sequences to transgenic cDNA allows for transgene downregulation specifically in cells expressing the respective miRNAs. Here, we have investigated the potential of miRNA-150 target sequences to specifically suppress gene expression in lymphocytes and thereby prevent transgene-induced lymphotoxicity. Abundance of miRNA-150 expression specifically in differentiated B and T cells was confirmed by quantitative reverse transcriptase PCR. Mono- and bicistronic lentiviral vectors were used to investigate the effect of miRNA-150 target sequences on transgene expression in the lymphohematopoietic system. After in vitro studies demonstrated effective downregulation of transgene expression in murine B220(+) B and CD3(+) T cells, the concept was further verified in a murine transplant model. Again, marked suppression of transgene activity was observed in B220(+) B and CD4(+) or CD8(+) T cells whereas expression in CD11b(+) myeloid cells, lin(-) and lin(-)/Sca1(+) progenitors, or lin(-)/Sca1(+)/c-kit(+) stem cells remained almost unaffected. No toxicity of miRNA-150 targeting in transduced lymphohematopoietic cells was noted. Thus, our results demonstrate the suitability of miRNA-150 targeting to specifically suppress transgene expression in lymphocytes and further support the concept of miRNA targeting for cell type-specific transgene expression in gene therapy approaches. PMID:21975463

Lachmann, N; Jagielska, J; Heckl, D; Brennig, S; Pfaff, N; Maetzig, T; Modlich, U; Cantz, T; Gentner, B; Schambach, A; Moritz, T

2012-09-01

351

Development of potato transgenic plants and molecular and genetic analyses of their integration stability and transgene expression efficiency using vegetative and sexual propagation  

International Nuclear Information System (INIS)

Agrobacterium mediated transformation of potato (Solanum tuberosum L.) and the stability of integration, expression and inheritance of transgenes were investigated. Over 400 kanamycin resistant transgenic plants with the nptII, gus, tt (insect toxin of Bacillus thuringiensis, variety kurstaki) and tg (fusion of tt and gus under the 35S promoter) foreign genes were obtained from eight potato genotypes by co-cultivation of the leaf discs with A. tumefaciens LBA 4404 (pBI 121), A. tumefaciens 3850 (pGV 941tg) and A. rchizogenes A4 (pGV 941tt). Southern blot analysis showed that more than 80% of the regenerating plants had one or several copies of the foreign genes. Selection for kanamycin resistance and Southern blot analysis revealed the stability of inheritance of transgenes by vegetative propagation of the plants via cloning and through tubers. However, 0.3% of the plants lost the nptII gene during the cloning process. About 500 self-pollinated seeds of four independently obtained transgenic clones were harvested. Analysis of the phenotypic segregation of the transgenic progeny on selective medium with kanamycin (50 mg/L) was made and compared with the controls. Inheritance and expression of the nptII transgene in the progeny of all four transgenic clones were shown, although the character of segregation appeared to differ in the various clones

352

The sensitivity of Crepis capillaris transgenic roots to mutagenic factors  

International Nuclear Information System (INIS)

Rapid progress has recently been made in biotechnology, especially in the development of transformation techniques including transgenic roots - hairy roots. Transgenic root cultures are used not only as a source of secondary metabolites, but have also proven to be a very good material for cytogenetic analysis of cytotoxic and mutagenic effect of environmental factors on plant cells. In the present study, the sensitivity of transgenic roots of Crepis capillaris has been analysed. The agents used for treatment were X-rays, maleic hydrazid acid - mutagen effective in the S phase, and oryzaline - herbicide that causes depolimerization of mitotic spindle microtubules. Crepis capillaris is a very good model plant for those studies because it has three pairs of relatively big and morphologically differentiated chromosomes. The transformed Crepis roots grow quickly and are easy to culture in vitro. This makes it very convenient for the analysis of the influence of mutagenic and stress factors on plant cell. The effect of mutagenic treatments was estimated using two mutagenic tests namely micronucleus test and anaphase - telophase test. Additionally, fluorescence in situ hybridization with 45S rDNA was applied in order to analyse the involvement of NOR chromosomes in the formation of micronuclei, fragments and other types of chromosomal aberrations. Sister chromatid exchanges (SCE) were analysed to test the structural chromosome changes. The analysis of tural chromosome changes. The analysis of the changes in cells of Crepis capillaris after oryzaline treatment encompassed also the observation of microtubules of cytoskeleton and of mitotic spindle using the immunocytochemical approach. The results show different sensitivity of cells in 'hairy roots' and primary roots of Crepis capillaris to mutagenic factors. (author)

353

Strategies to facilitate transgene expression in Chlamydomonas reinhardtii.  

Science.gov (United States)

The unicellular green alga Chlamydomonas reinhardtii has been identified as a promising organism for the production of recombinant proteins. While during the last years important improvements have been developed for the production of proteins within the chloroplast, the expression levels of transgenes from the nuclear genome were too low to be of biotechnological importance. In this study, we integrated endogenous intronic sequences into the expression cassette to enhance the expression of transgenes in the nucleus. The insertion of one or more copies of intron sequences from the Chlamydomonas RBCS2 gene resulted in increased expression levels of a Renilla-luciferase gene used as a reporter. Although any of the three RBCS2 introns alone had a positive effect on expression, their integration in their physiological number and order created an over-proportional stimulating effect observed in all transformants. The secretion of the luciferase protein into the medium was achieved by using the export sequence of the Chlamydomonas ARS2 gene in a cell wall deficient strain and Renilla-luciferase could be successfully concentrated with the help of attached C-terminal protein tags. Similarly, a codon adapted gene variant for human erythropoietin (crEpo) was expressed as a protein of commercial relevance. Extracellular erythropoietin produced in Chlamydomonas showed a molecular mass of 33 kDa probably resulting from post-translational modifications. Both, the increased expression levels of transgenes by integration of introns and the isolation of recombinant proteins from the culture medium are important steps towards an extended biotechnological use of this alga. PMID:19127370

Eichler-Stahlberg, Alke; Weisheit, Wolfram; Ruecker, Ovidiu; Heitzer, Markus

2009-03-01

354

Transgenic chickens expressing human urokinase-type plasminogen activator.  

Science.gov (United States)

Urokinase-type plasminogen activator is a serine protease that is clinically used in humans for the treatment of thrombolytic disorders and vascular diseases such as acute ischemic stroke and acute peripheral arterial occlusion. This study explored the feasibility of using chickens as a bioreactor for producing human urokinase-type plasminogen activator (huPA). Recombinant huPA gene, under the control of a ubiquitous Rous sarcoma virus promoter, was injected into the subgerminal cavity of freshly laid chicken eggs at stage X using the replication-defective Moloney murine leukemia virus (MoMLV)-based retrovirus vectors encapsidated with VSV-G (vesicular stomatitis virus G) glycoprotein. A total of 38 chicks, out of 573 virus-injected eggs, hatched and contained the huPA gene in their various body parts. The mRNA transcript of the huPA gene was present in various organs, including blood and egg, and was germ-line transmitted to the next generation. The level of active huPA protein was 16-fold higher in the blood of the transgenic chicken than in the nontransgenic chicken (P rooster showed reduced (P < 0.05) fertility, as revealed by reduced volume of semen ejaculate, sperm concentration, and sperm viability. Taken together, our data suggest that huPA transgenic chickens could be successfully produced by the retroviral vector system. Transgenic chickens, expressing the huPA under the control of a ubiquitous promoter, may not only be used as a bioreactor for pharming of the huPA drug but also be useful for studying huPA-induced bleeding and other disorders. PMID:23960123

Lee, Sung Ho; Gupta, Mukesh Kumar; Ho, Young Tae; Kim, Teoan; Lee, Hoon Taek

2013-09-01

355