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1.
Rev Med Interne ; 42(10): 678-685, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34303547

RESUMO

INTRODUCTION: Aspergillusfumigatus can cause a systemic infection called invasive aspergillosis causing pulmonary and extra-pulmonary damage. Aspergillus endocarditis (AE) is a relatively rare disease but can be life-threatening. CASE REPORTS: We report here on five cases of endocarditis due to invasive aspergillosis: a 58-year-old man receiving immunosuppressive medication following a kidney graft, a 58-year-old man undergoing chemotherapy for chronic lymphocytic leukaemia, a 55-year-old man receiving corticosteroids for IgA vasculitis, a 52-year-old HIV-infected woman under no specific treatment and a 17-year-old boy under immunosuppressive therapy for auto-immune chronic neutropenia. DISCUSSION: Aspergillus accounts for 25-30% of fungal endocarditis and 0.25% to 8.5% of all cases of infectious endocarditis. Aspergillus endocarditis results from invasion of the lung arterioles by hyphae and blood dissemination. It is associated with a very high mortality rate (42-68%). Diagnosing Aspergillus endocarditis is mainly problematic because blood cultures are almost always negative, and fever may be absent. Immunosuppression, haematological malignancies, recent cardiothoracic surgery, negative blood cultures with endocarditis and/or systemic or pulmonary emboli are predictors of AE. In the setting of endocarditis, some clinical characteristics may raise early suspicions of aspergillosis rather than a non-fungal agent: no fever, vegetations affecting the mitral valve, non-valve or aortotomy sites, aortic abscess or pseudo-aneurysm. The identification of invasive aspergillosis is based on a chest CT scan, microscopy/culture or other serological and molecular tests. The treatment of Aspergillus endocarditis requires triazole antifungal drugs, and frequently additional surgical debridement. CONCLUSION: Aspergillus endocarditis is rare but is associated with a very high mortality rate. Knowledge of its predictive factors and key clinical features can help to differentiate aspergillosis from non-fungal endocarditis and may enable improved survival rates.


Assuntos
Aspergilose , Endocardite , Transplante de Rim , Adolescente , Antifúngicos/uso terapêutico , Aspergilose/diagnóstico , Endocardite/diagnóstico , Feminino , Humanos , Transplante de Rim/efeitos adversos , Masculino , Pessoa de Meia-Idade , Valva Mitral
2.
Skin Pharmacol Physiol ; 27(3): 164-71, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24457536

RESUMO

BACKGROUND/AIMS: Silicone excipients are commonly used ingredients because of their emollient and skin-conditioning effects, and their ability to form uniform, water-resistant, yet permeable films. Based on comparisons with organic materials and conflicting knowledge from silicones used in scar treatment, the misconception still exists that silicone topical excipients are occlusive substances that may block the passive loss of water through the upper skin layers. Therefore, 3 types of common silicone excipients and 3 water-in-(oil-plus-silicone) or W/(O + Si) creams, containing 10% (w/w) of the respective silicones, were investigated as a function of time and compared to petrolatum. METHODS: Transepidermal water loss (TEWL) and skin hydration measurements were carried out after a single topical application on forearm skin of 26 healthy young female volunteers. RESULTS: Both petrolatum and silicones significantly decreased TEWL 15 min after application, but the measurements for the silicones were not significantly different from the untreated control values. The tested silicones did not moisturize the skin. Petrolatum formed an occlusive layer, creating an increase in skin hydration for more than 4 h. The results measured for the W/(O + Si) creams indicated that they moisturized the skin, without any effect on TEWL. CONCLUSION: A clear difference was shown between the skin occlusive properties of petrolatum and the water vapor permeability of the common silicone excipient materials.


Assuntos
Emolientes/química , Excipientes/química , Silicones/química , Pele/efeitos dos fármacos , Administração Cutânea , Adulto , Emolientes/administração & dosagem , Excipientes/administração & dosagem , Feminino , Antebraço , Humanos , Óleos/química , Permeabilidade , Vaselina/administração & dosagem , Vaselina/química , Silicones/administração & dosagem , Pele/metabolismo , Fatores de Tempo , Água , Perda Insensível de Água , Adulto Jovem
3.
J Bioenerg Biomembr ; 35(1): 19-30, 2003 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-12848338

RESUMO

Mice carrying a homozygous germ-line mutation in the nm23-M1 gene that eliminates its protein expression and drives expression of beta-galactosidase by nm23-M1 promoter have been generated. nm23-M1 gene inactivation is not teratogenic and the pups can grow to adult age without apparent health problems. However, they undergo a growth retardation and knocked out females cannot feed their pups. Both effects are background dependent. Beta-galactosidase mapping of nm23-M1 promoter activation during embryogenesis shows that the nm23-M1 gene is principally expressed in epithelial layer of tissues which require inductive epithelial-mesenchymal interactions for their formation. In conclusion, invalidated mice could be interesting models to analyze the role of nm23-M1 on signal transduction pathway regulation, or cancer induction and proliferation.


Assuntos
Mama/metabolismo , Retardo do Crescimento Fetal/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação Enzimológica da Expressão Gênica/genética , Modelos Animais , Núcleosídeo-Difosfato Quinase , Proteínas/genética , Proteínas/metabolismo , Animais , Animais Recém-Nascidos , Clonagem Molecular , Feminino , Retardo do Crescimento Fetal/metabolismo , Camundongos , Camundongos Knockout/embriologia , Camundongos Knockout/crescimento & desenvolvimento , Camundongos Knockout/metabolismo , Nucleosídeo NM23 Difosfato Quinases , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transdução de Sinais/genética , Relação Estrutura-Atividade
4.
J Bacteriol ; 183(13): 3848-54, 2001 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-11395447

RESUMO

Polynucleotide phosphorylase (PNPase) synthesis is translationally autocontrolled via an RNase III-dependent mechanism, which results in a tight correlation between protein level and messenger stability. In cells grown at 18 degrees C, the amount of PNPase is twice that found in cells grown at 30 degrees C. To investigate whether this effect was transcriptional or posttranscriptional, the expression of a set of pnp-lacZ transcriptional and translational fusions was analyzed in cells grown at different temperatures. In the absence of PNPase, there was no increase in pnp-lacZ expression, indicating that the increase in pnp expression occurs at a posttranscriptional level. Other experiments clearly show that increased pnp expression at low temperature is only observed under conditions in which the autocontrol mechanism of PNPase is functional. At low temperature, the destabilizing effect of PNPase on its own mRNA is less efficient, leading to a decrease in repression and an increase in the expression level.


Assuntos
Proteínas de Escherichia coli , Escherichia coli/genética , Polirribonucleotídeo Nucleotidiltransferase/genética , Temperatura Baixa , Endorribonucleases/metabolismo , Escherichia coli/enzimologia , Regulação Bacteriana da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Genes Reporter , Óperon Lac , Polirribonucleotídeo Nucleotidiltransferase/biossíntese , Biossíntese de Proteínas , Estabilidade de RNA , Proteínas Recombinantes de Fusão/biossíntese , Ribonuclease III
5.
Proc Natl Acad Sci U S A ; 97(17): 9508-13, 2000 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-10944220

RESUMO

Several lines of evidence suggest that the serotonin (5-hydroxytryptamine, 5-HT) regulates cardiovascular functions during embryogenesis and adulthood. 5-HT binds to numerous cognate receptors to initiate its biological effects. However, none of the 5-HT receptor disruptions in mice have yet resulted in embryonic defects. Here we show that 5-HT(2B) receptor is an important regulator of cardiac development. We found that inactivation of 5-HT(2B) gene leads to embryonic and neonatal death caused by heart defects. 5-HT(2B) mutant embryos exhibit a lack of trabeculae in the heart and a specific reduction in the expression levels of a tyrosine kinase receptor, ErbB-2, leading to midgestation lethality. These in vivo data suggest that the Gq-coupled receptor 5-HT(2B) uses the signaling pathway of tyrosine kinase receptor ErbB-2 for cardiac differentiation. All surviving newborn mice display a severe ventricular hypoplasia caused by impaired proliferative capacity of myocytes. In adult mutant mice, cardiac histopathological changes including myocyte disarray and ventricular dilation were consistently observed. Our results constitute genetic evidence that 5-HT via 5-HT(2B) receptor regulates differentiation and proliferation of developing and adult heart. This mutation provides a genetic model for cardiopathy and should facilitate studies of both the pathogenesis and therapy of cardiac disorders in humans.


Assuntos
Coração/embriologia , Miocárdio/metabolismo , Receptores de Serotonina/metabolismo , Animais , Animais Recém-Nascidos , Diferenciação Celular , Divisão Celular , Embrião de Mamíferos/metabolismo , Embrião de Mamíferos/patologia , Embrião de Mamíferos/fisiopatologia , Feminino , Morte Fetal , Deleção de Genes , Genes erbB-2/genética , Coração/fisiopatologia , Cardiopatias Congênitas/metabolismo , Cardiopatias Congênitas/patologia , Cardiopatias Congênitas/fisiopatologia , Proteínas Heterotriméricas de Ligação ao GTP/genética , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Cinética , Masculino , Camundongos , Camundongos Knockout , Miocárdio/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Receptor 5-HT2B de Serotonina , Receptores de Serotonina/genética , Transdução de Sinais
6.
Hum Mol Genet ; 9(5): 849-58, 2000 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-10749994

RESUMO

Deletion of the murine survival of motor neuron gene (SMN) exon 7, the most frequent mutation found in spinal muscular atrophy (SMA) patients, directed to neurons but not to skeletal muscle, enabled generation of a mouse model of SMA providing evidence that motor neurons are the primary target of the gene defect. Moreover, the mutated SMN protein (SMNDeltaC15) is dramatically reduced in the motor neuron nuclei and causes a lack of gems associated with large aggregates of coilin, a coiled-body-specific protein. These results identify the lack of the nuclear targeting of SMN as the biochemical defect in SMA.


Assuntos
Núcleo Celular/metabolismo , Atrofia Muscular Espinal/genética , Proteínas do Tecido Nervoso/genética , Animais , Sequência de Bases , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico , Primers do DNA , Modelos Animais de Doenças , Éxons , Deleção de Genes , Genes Letais , Homozigoto , Camundongos , Neurônios Motores/metabolismo , Atrofia Muscular Espinal/metabolismo , Proteínas do Tecido Nervoso/química , Fenótipo , Proteínas de Ligação a RNA , Proteínas do Complexo SMN
7.
Nucleic Acids Res ; 27(19): e27, 1999 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-10481039

RESUMO

Spatially and temporally regulated somatic mutations can be achieved by using the Cre/LoxP recombination system of bacteriophage P1. In order to develop gene knockouts restricted to striated muscle, we generated a transgenic mouse line expressing Cre recombinase under the control of the human alpha-skeletal actin promoter. Specific excision of a loxP-flanked gene was demonstrated in striated muscle, heart and skeletal muscle, in a pattern very similar to the expression of the endogenous alpha-skeletal actin gene. Therefore, the reported transgenic line can be used to target inactivation or activation of a given gene to the skeletal muscle lineage.


Assuntos
Marcação de Genes , Integrases/genética , Músculo Esquelético/metabolismo , Proteínas Virais , Actinas/genética , Animais , Galinhas , Humanos , Óperon Lac , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miocárdio/metabolismo , Regiões Promotoras Genéticas
8.
Development ; 126(19): 4295-304, 1999 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-10477297

RESUMO

Previous studies have shown that the homeobox gene Otx2 is required first in the visceral endoderm for induction of forebrain and midbrain, and subsequently in the neurectoderm for its regional specification. Here, we demonstrate that Otx2 functions both cell autonomously and non-cell autonomously in neurectoderm cells of the forebrain and midbrain to regulate expression of region-specific homeobox and cell adhesion genes. Using chimeras containing both Otx2 mutant and wild-type cells in the brain, we observe a reduction or loss of expression of Rpx/Hesx1, Wnt1, R-cadherin and ephrin-A2 in mutant cells, whereas expression of En2 and Six3 is rescued by surrounding wild-type cells. Forebrain Otx2 mutant cells subsequently undergo apoptosis. Altogether, this study demonstrates that Otx2 is an important regulator of brain patterning and morphogenesis, through its regulation of candidate target genes such as Rpx/Hesx1, Wnt1, R-cadherin and ephrin-A2.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Homeodomínio , Mesencéfalo/embriologia , Mesencéfalo/metabolismo , Proteínas do Tecido Nervoso/fisiologia , Prosencéfalo/embriologia , Rombencéfalo/embriologia , Transativadores/fisiologia , Animais , Apoptose , Caderinas/metabolismo , Adesão Celular/fisiologia , Quimera/genética , Efrina-A2 , Hibridização In Situ , Proteínas de Membrana/metabolismo , Mesencéfalo/anatomia & histologia , Camundongos , Mutagênese , Fatores de Transcrição Otx , Fenótipo , Prosencéfalo/anatomia & histologia , Rombencéfalo/anatomia & histologia , Células-Tronco/metabolismo , Fatores de Tempo , Fatores de Transcrição/metabolismo
9.
Oncogene ; 18(7): 1419-34, 1999 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-10050879

RESUMO

The MDM2 proto-oncogene is overexpressed in human tumours and regulates the activities of the tumour suppressors p53 and pRB. We created mice that overexpress MDM2 under the control of the CMV promoter. These mice did not display an increased tumour incidence, but rather a specific skin phenotype, characterized by desquamation and hyperkeratosis. Transgenic MDM2 was found to be overexpressed in the epidermis, a tissue that normally expresses high levels of MDM2. The phenotype appeared during the first week after birth and then lessened with age, closely following the level of expression of the transgene. MDM2 overexpression was associated with an increase in proliferation in the basal layer, thickening of the epidermis, altered expression of the differentiation markers cytokeratin CK14, CK10 and CK1, and a decrease in the size and the number of granules that contain products of differentiation. Transgenic mice on a p53 null background displayed similar although not identical changes, showing that the effects of MDM2 are to a certain degree p53 independent. The skin is a major site of MDM2 expression in mice, raising the possibility that MDM2 overexpression perturbs the normal pattern of MDM2 expression and inhibits differentiation of the epidermis.


Assuntos
Proteínas Nucleares , Proteínas Proto-Oncogênicas/biossíntese , Neoplasias Cutâneas/etiologia , Proteína Supressora de Tumor p53/fisiologia , 9,10-Dimetil-1,2-benzantraceno/farmacologia , Animais , Apoptose , Biomarcadores , Carcinógenos/farmacologia , DNA/biossíntese , Epiderme/patologia , Expressão Gênica , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Fenótipo , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-mdm2 , Coelhos , Neoplasias Cutâneas/patologia , Acetato de Tetradecanoilforbol/farmacologia , Proteína Supressora de Tumor p53/genética
10.
Development ; 125(5): 845-56, 1998 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-9449667

RESUMO

The homeobox gene Otx2 is a mouse cognate of the Drosophila orthodenticle gene, which is required for development of the brain, rostral to rhombomere three. We have investigated the mechanisms involved in this neural function and specifically the requirement for Otx2 in the visceral endoderm and the neuroectoderm using chimeric analysis in mice and explant recombination assay. Analyses of chimeric embryos composed of more than 90% of Otx2-/- ES cells identified an essential function for Otx2 in the visceral endoderm for induction of the forebrain and midbrain. The chimeric studies also demonstrated that an anterior neural plate can form without expressing Otx2. However, in the absence of Otx2, expression of important regulatory genes, such as Hesx1/Rpx, Six3, Pax2, Wnt1 and En, fail to be initiated or maintained in the neural plate. Using explant-recombination assay, we could further demonstrate that Otx2 is required in the neuroectodem for expression of En. Altogether, these results demonstrate that Otx2 is first required in the visceral endoderm for the induction, and subsequently in the neuroectoderm for the specification of forebrain and midbrain territories.


Assuntos
Genes Homeobox , Proteínas de Homeodomínio/genética , Peptídeos e Proteínas de Sinalização Intercelular , Mesencéfalo/embriologia , Mesencéfalo/metabolismo , Proteínas do Tecido Nervoso/genética , Prosencéfalo/embriologia , Prosencéfalo/metabolismo , Transativadores/genética , Animais , Proteínas de Transporte , Linhagem Celular , Quimera/genética , Drosophila/embriologia , Drosophila/genética , Ectoderma/metabolismo , Indução Embrionária/genética , Endoderma/metabolismo , Feminino , Folistatina , Regulação da Expressão Gênica no Desenvolvimento , Genes de Insetos , Glicoproteínas/genética , Hibridização In Situ , Masculino , Mesencéfalo/anormalidades , Camundongos , Camundongos Knockout , Camundongos Mutantes , Fatores de Transcrição Otx , Gravidez , Prosencéfalo/anormalidades , Proteínas/genética
11.
EMBO J ; 17(4): 886-97, 1998 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-9463367

RESUMO

***micro***-, delta- and kappa-opioid receptors are widely expressed in the central nervous system where they mediate the strong analgesic and mood-altering actions of opioids, and modulate numerous endogenous functions. To investigate the contribution of the kappa-opioid receptor (KOR) to opioid function in vivo, we have generated KOR-deficient mice by gene targeting. We show that absence of KOR does not modify expression of the other components of the opioid system, and behavioural tests indicate that spontaneous activity is not altered in mutant mice. The analysis of responses to various nociceptive stimuli suggests that the KOR gene product is implicated in the perception of visceral chemical pain. We further demonstrate that KOR is critical to mediate the hypolocomotor, analgesic and aversive actions of the prototypic kappa-agonist U-50, 488H. Finally, our results indicate that this receptor does not contribute to morphine analgesia and reward, but participates in the expression of morphine abstinence. Together, our data demonstrate that the KOR-encoded receptor plays a modulatory role in specific aspects of opioid function.


Assuntos
(trans)-Isômero de 3,4-dicloro-N-metil-N-(2-(1-pirrolidinil)-ciclo-hexil)-benzenoacetamida/farmacologia , Morfina/efeitos adversos , Dor/induzido quimicamente , Receptores Opioides kappa/agonistas , Receptores Opioides kappa/genética , Síndrome de Abstinência a Substâncias/metabolismo , Animais , Comportamento Animal/efeitos dos fármacos , Encefalinas/biossíntese , Encefalinas/genética , Feminino , Regulação da Expressão Gênica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dor/metabolismo , Dor/psicologia , Pró-Opiomelanocortina/biossíntese , Pró-Opiomelanocortina/genética , Precursores de Proteínas/biossíntese , Precursores de Proteínas/genética , Receptores Opioides kappa/deficiência , Síndrome de Abstinência a Substâncias/genética , Síndrome de Abstinência a Substâncias/psicologia , Vísceras
12.
Nature ; 383(6603): 819-23, 1996 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-8893006

RESUMO

Despite tremendous efforts in the search for safe, efficacious and non-addictive opioids for pain treatment, morphine remains the most valuable painkiller in contemporary medicine. Opioids exert their pharmacological actions through three opioid-receptor classes, mu, delta and kappa, whose genes have been cloned. Genetic approaches are now available to delineate the contribution of each receptor in opioid function in vivo. Here we disrupt the mu-opioid-receptor gene in mice by homologous recombination and find that there are no overt behavioural abnormalities or major compensatory changes within the opioid system in these animals. Investigation of the behavioural effects of morphine reveals that a lack of mu receptors abolishes the analgesic effect of morphine, as well as place-preference activity and physical dependence. We observed no behavioural responses related to delta- or kappa-receptor activation with morphine, although these receptors are present and bind opioid ligands. We conclude that the mu-opioid-receptor gene product is the molecular target of morphine in vivo and that it is a mandatory component of the opioid system for morphine action.


Assuntos
Analgésicos/farmacologia , Morfina/farmacologia , Entorpecentes/farmacologia , Receptores Opioides mu/metabolismo , Analgésicos/efeitos adversos , Analgésicos/metabolismo , Animais , Comportamento Animal , Linhagem Celular , Ala(2)-MePhe(4)-Gly(5)-Encefalina , Encefalinas/metabolismo , Deleção de Genes , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Morfina/efeitos adversos , Morfina/metabolismo , Dependência de Morfina/metabolismo , Entorpecentes/efeitos adversos , Entorpecentes/metabolismo , Receptores Opioides delta/metabolismo , Receptores Opioides kappa/metabolismo , Receptores Opioides mu/genética , Recompensa , Síndrome de Abstinência a Substâncias/metabolismo
13.
Nat Genet ; 13(3): 370-3, 1996 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-8673141

RESUMO

Cell-extracellular matrix interactions have important roles in many biological processes, including embryonic development, growth control and differentiation. Integrins are the principal receptors for extracellular matrix. They are composed of non-covalently associated alpha and beta chains. Integrin alpha 6 can associate with either beta 1 or beta 4 (refs 2,3). Both integrin complexes are receptors for laminins, major components of basement membranes. The distribution of alpha 6 (refs 4-10) as well as studies using function-blocking antibodies have suggested an essential role for this laminin receptor during embryogenesis, in processes such as endoderm migration or kidney tubule formation9. Here we report that, surprisingly, mice lacking the alpha 6 integrin chain develop to birth. However, they die at birth with severe blistering of the skin and other epithelia, a phenotype reminiscent of the human disorder epidermolysis bullosa. Hemidesmosomes are absent in mutant tissue. This absence is likely to result from the lack of alpha 6/beta 4, the only integrin in hemidesmosomes of stratified squamous and transitional epithelia. Mutations in the genes encoding integrin beta 4 and chains of laminin-5 have been implicated in junctional epidermolysis bullosa. Our study provides evidence that some forms of epidermolysis bullosa may originate from defects of the alpha 6 gene.


Assuntos
Antígenos CD/genética , Epidermólise Bolhosa/genética , Morte Fetal/genética , Animais , Antígenos CD/fisiologia , Moléculas de Adesão Celular/química , Desmossomos/patologia , Epiderme/anatomia & histologia , Epiderme/patologia , Feminino , Fertilidade , Regulação da Expressão Gênica no Desenvolvimento , Vetores Genéticos , Heterozigoto , Integrina alfa6 , Queratinócitos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pele/química , Pele/patologia , Calinina
14.
C R Acad Sci III ; 319(1): 45-50, 1996 Jan.
Artigo em Francês | MEDLINE | ID: mdl-8673619

RESUMO

This work was aimed at generating a novel system for gene transfer to tumor cell, combining the advantages of non-viral gene transfer methods with those of transfer by recombinant retroviruses. We replaced the env gene of an infectious Moloney murine leukemia provirus with the gene coding for the thymidine kinase of Herpes Simplex Virus 1 (HSV1-TK). The sole transfection of this construction allows the production of viral particles, and the encapsidation of a viral genome carrying transgene. We show that this gene is expressed at a level sufficient for conferring sensitivity to ganciclovir, a nucleoside analog that is metabolised in a toxic compound by HSV1-TK. We also show that the complementation of this recombinant defective provirus with a gene coding for a retroviral envelope, either expressed constitutively by the transduced cell, or by co-transfection, leads to the formation of infectious viral particles capable of transducing HSV1-TK into tumor cells.


Assuntos
Técnicas de Transferência de Genes , Vetores Genéticos , Provírus/genética , Transgenes , Antivirais/farmacologia , Ganciclovir/farmacologia , Herpesvirus Humano 1/genética , Técnicas In Vitro , Vírus da Leucemia Murina de Moloney/genética , Recombinação Genética , Timidina Quinase/efeitos dos fármacos , Timidina Quinase/genética
15.
Nature ; 377(6548): 424-8, 1995 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-7566118

RESUMO

Dopaminergic neuronal pathways arise from mesencephalic nuclei and project axons to the striatum, cortex, limbic system and hypothalamus. Through these pathways dopamine affects many physiological functions, such as the control of coordinated movement and hormone secretion. Here we have studied the physiological involvement of the dopamine D2 receptors in dopaminergic transmission, using homologous recombination to generate D2-receptor-deficient mice. Absence of D2 receptors leads to animals that are akinetic and bradykinetic in behavioural tests, and which show significantly reduced spontaneous movements. This phenotype presents analogies with symptoms characteristic of Parkinson's disease. Our study shows that D2 receptors have a key role in the dopaminergic control of nervous function. These mice have therapeutic potential as a model for investigating and correcting dysfunctions of the dopaminergic system.


Assuntos
Doença de Parkinson/metabolismo , Receptores de Dopamina D2/fisiologia , Animais , Encéfalo/metabolismo , Linhagem Celular , Clonagem Molecular , Modelos Animais de Doenças , Dopamina/metabolismo , Feminino , Hibridização In Situ , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Atividade Motora , Neuropeptídeos/biossíntese , Neuropeptídeos/genética , Ovário/metabolismo , Doença de Parkinson/fisiopatologia , RNA/metabolismo , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/deficiência , Testículo/metabolismo
16.
Proc Natl Acad Sci U S A ; 92(1): 310-4, 1995 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-7816839

RESUMO

The Hoxd-11 gene was disrupted by homologous recombination in embryonic stem cells. We found that Hoxd-11-/- mutant mice are viable and display homeotic transformations of their sacral vertebrae, while their forelimbs present abnormalities of some metacarpals and of the first row of carpal bones. These results are discussed in the light of current models of tetrapod axial skeleton and limb patterning.


Assuntos
Osso e Ossos/anormalidades , Membro Anterior/anormalidades , Genes Homeobox , Medula Espinal/anormalidades , Animais , Cosmídeos , Éxons , Camundongos , Camundongos Mutantes , Fenótipo , Recombinação Genética , Mapeamento por Restrição
17.
Nucleic Acids Res ; 22(3): 397-403, 1994 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-7510392

RESUMO

Polynucleotide phosphorylase, a 3' to 5' processive exoribonuclease is post-transcriptionally autocontrolled and it was previously shown that this control is dependent on a 5' processing by RNase III. In this paper, the mechanism of regulation is analyzed by studying the properties of a pnp-lacZ translational gene fusion. It is shown that this message is stable, even when processed by RNase III, and that the degradation rate is directly linked to the intracellular concentration of polynucleotide phosphorylase or to the pnp-lacZ messenger translation rate. Mutations able to decrease the level of repression are all located in the ribosome loading site. Taken together, these results suggest that polynucleotide phosphorylase is able to recognize specifically the processed messenger and to prevent its translation, thus allowing degradation of the message.


Assuntos
Proteínas de Escherichia coli , Escherichia coli/enzimologia , Regulação Bacteriana da Expressão Gênica , Polirribonucleotídeo Nucleotidiltransferase/genética , RNA Mensageiro/metabolismo , Sequência de Bases , Endorribonucleases/metabolismo , Escherichia coli/genética , Ligação de Hidrogênio , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Conformação de Ácido Nucleico , Iniciação Traducional da Cadeia Peptídica , Biossíntese de Proteínas , RNA Bacteriano/genética , RNA Bacteriano/metabolismo , Ribonuclease III , Relação Estrutura-Atividade
18.
EMBO J ; 11(7): 2633-41, 1992 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-1628624

RESUMO

It has been previously shown that the pnp messenger RNAs are cleaved by RNase III at the 5' end and that these cleavages induce a rapid decay of these messengers. A translational fusion between pnp and lacZ was introduced into the chromosome of a delta lac strain to study the expression of pnp. In the presence of increased cellular concentrations of polynucleotide phosphorylase, the level of the hybrid beta-galactosidase is repressed, whereas the synthesis rate of the corresponding message is not significantly affected. In the absence of pnp, the level of the hybrid protein increases strongly. Thus, polynucleotide phosphorylase is post-transcriptionally autocontrolled. However, autocontrol is totally abolished in strains where the RNase III site on the pnp message has been deleted or in strains devoid of RNase III. These results suggest that polynucleotide phosphorylase requires RNase III cleavages to autoregulate the translation of its message. Other mutations in the ribosome binding site region support the hypothesis that this 3' to 5' processive enzyme could recognize a specific repressor binding site at the 5' end of pnp mRNA. Implications of these results on the mechanism of regulation and on messenger degradation are discussed.


Assuntos
Endorribonucleases/metabolismo , Proteínas de Escherichia coli , Escherichia coli/enzimologia , Regulação Bacteriana da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Polirribonucleotídeo Nucleotidiltransferase/genética , Sequência de Bases , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Óperon , Plasmídeos , Polirribonucleotídeo Nucleotidiltransferase/biossíntese , Regiões Promotoras Genéticas , RNA Mensageiro/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Mapeamento por Restrição , Ribonuclease III , beta-Galactosidase/genética
19.
J In Vitro Fert Embryo Transf ; 8(4): 222-4, 1991 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-1753169

RESUMO

Microinjection of capacitated sperm into the perivitelline space of oocyte was offered to one couple with persistent infertility of mixed origin. The husband's semen was subnormal, whereas his wife had definitive tubal occlusion and polycystic ovaries. Four previous in vitro fertilization (IVF) attempts were performed but no fertilization was obtained. After superovulation, 13 oocytes were collected. Ten were submitted to microinjection and two were damaged during the procedure. One of the remaining eight had two pronuclei 18 hr after microinjection and progressed to a four-cell embryo after 48 hr. After reimplantation, a normal pregnancy was initiated, caryotype (46XX) was checked at 17 weeks. A normal and healthy girl has been delivered at term.


Assuntos
Fertilização in vitro/métodos , Microinjeções/métodos , Oócitos , Adulto , Transferência Embrionária , Feminino , Humanos , Masculino , Capacitação Espermática , Interações Espermatozoide-Óvulo
20.
Biologicals ; 18(3): 191-8, 1990 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-2257132

RESUMO

A number of therapeutic plasma proteins are synthesized by human hepatocytes. Since many of these proteins undergo liver-specific post-translational modifications which are required for full biological activity, it may therefore be necessary to develop hepatocyte-based expression systems for their production. Using transgenic mice we have developed a transimmortalisation technique for the isolation of differentiated hepatic cell lines, already engineered to secrete human alpha 1 antitrypsin (alpha 1 AT), a plasma protein which is produced mainly in liver cells. This was achieved by co-expression of the mouse c-myc proto-oncogene and a genomic copy of the human alpha 1 AT gene, both under the control of the human alpha 1 AT promoter. Transgenic mice carrying this construct developed hepatomas producing human alpha 1 AT. Under defined culture conditions, cell lines secreting active alpha 1 AT were derived from these tumours. These cells maintain a differentiated hepatic phenotype and continue to secrete human alpha 1 AT for at least 40 generations.


Assuntos
Transformação Celular Neoplásica/genética , Fígado/citologia , Células Tumorais Cultivadas/metabolismo , alfa 1-Antitripsina/biossíntese , Animais , Sequência de Bases , Meios de Cultura , Genes myc/genética , Fígado/metabolismo , Neoplasias Hepáticas Experimentais/metabolismo , Camundongos , Camundongos Transgênicos , Dados de Sequência Molecular , Fenótipo , Regiões Promotoras Genéticas , Processamento de Proteína Pós-Traducional , Proto-Oncogene Mas , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , alfa 1-Antitripsina/genética
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