RESUMO
The reproducibility of research using laboratory animals requires reliable management of their quality, in particular of their genetics, health and environment, all of which contribute to their phenotypes. The point at which these biological materials are transferred between researchers is particularly sensitive, as it may result in a loss of integrity of the animals and/or their documentation. Here, we describe the various aspects of laboratory animal quality that should be confirmed when sharing rodent research models. We also discuss how repositories of biological materials support the scientific community to ensure the continuity of the quality of laboratory animals. Both the concept of quality and the role of repositories themselves extend to all exchanges of biological materials and all networks that support the sharing of these reagents.
Assuntos
Pesquisadores , Animais , Humanos , Reprodutibilidade dos TestesRESUMO
Indirect immunofluorescence labeling of normal rat kidney (NRK) cells with antibodies recognizing a lysosomal glycoprotein (LGP 120; Lewis, V., S.A. Green, M. Marsh, P. Vihko, A. Helenius, and I. Mellman, 1985, J. Cell Biol., 100:1839-1847) reveals that lysosomes accumulate in the region around the microtubule-organizing center (MTOC). This clustering of lysosomes depends on microtubules. When the interphase microtubules are depolymerized by treatment of the cells with nocodazole or during mitosis, the lysosomes disperse throughout the cytoplasm. Lysosomes recluster rapidly (within 30-60 min) in the region of the centrosomes either upon removal of the drug, or, in telophase, when repolymerization of interphase microtubules has occurred. During this translocation process the lysosomes can be found aligned along centrosomal microtubules. Endosomes and lysosomes can be visualized by incubating living cells with acridine orange. We have analyzed the movement of these labeled endocytic organelles in vivo by video-enhanced fluorescence microscopy. Translocation of endosomes and lysosomes occurs along linear tracks (up to 10 microns long) by discontinuous saltations (with velocities of up to 2.5 microns/s). Organelles move bidirectionally with respect to the MTOC. This movement ceases when microtubules are depolymerized by treatment of the cells with nocodazole. After nocodazole washout and microtubule repolymerization, the translocation and reclustering of fluorescent organelles predominantly occurs in a unidirectional manner towards the area of the MTOC. Organelle movement remains unaffected when cells are treated with cytochalasin D, or when the collapse of intermediate filaments is induced by microinjected monoclonal antivimentin antibodies. It can be concluded that translocation of endosomes and lysosomes occurs along microtubules and is independent of the intermediate filament and microfilament networks.
Assuntos
Lisossomos/ultraestrutura , Microtúbulos/ultraestrutura , Organoides/ultraestrutura , Animais , Anticorpos , Linhagem Celular , Imunofluorescência , Glicoproteínas/análise , Rim , Modelos Biológicos , RatosRESUMO
AtT20 (clone D16V) cells develop long neurite-like processes in the growth cones of which secretory granules containing ACTH accumulate. These secretory granules have an acidic pH. Using acridine orange as a vital stain for acidic organelles, in combination with video-enhanced fluorescence microscopy, and subsequent immunolabeling with rabbit antibodies against ACTH, we have shown that these secretory granules move by saltations along the processes. During saltations velocities of 3 to 5 microns/s are achieved. The majority of the secretory granules move in the anterograde direction but some move retrogradely. The growth cones and processes are the site of extensive endocytosis. Using Lucifer Yellow as a vital stain we have shown that most endosomes move by saltations retrogradely. Movement of both secretory granules and endosomes is dependent upon microtubules. Individual secretory granules or endosomes never reverse the direction of their movement as they traverse the processes. Neutralization of the lumen of these acidic organelles with NH4Cl does not inhibit their movement or change its direction.
Assuntos
Hormônio Adrenocorticotrópico/metabolismo , Grânulos Citoplasmáticos/metabolismo , Microtúbulos/metabolismo , Adeno-Hipófise/citologia , Laranja de Acridina , Animais , Transporte Biológico Ativo , Células Cultivadas , Grânulos Citoplasmáticos/ultraestrutura , Endocitose , Camundongos , Adeno-Hipófise/metabolismo , Adeno-Hipófise/ultraestrutura , Gravação de VideoteipeRESUMO
A case of tumoral calcinosis of the hand is reported. The lesion was localized at the fifth digit of the right hand and caused pain and disability. The clinical, diagnostic and therapeutic problems of this rare soft tissue disease are discussed.
Assuntos
Calcinose/diagnóstico por imagem , Articulações dos Dedos/diagnóstico por imagem , Adulto , Artrografia , Calcinose/cirurgia , Feminino , Articulações dos Dedos/cirurgia , Humanos , Artropatias/diagnóstico por imagem , Artropatias/cirurgiaRESUMO
In this work, the authors support the therapeutic validity of gastric-duodenal resection in urgency surgery. This study has been made on 479 cases observed during the last 21 years (1957-78). In 58 of them we have made a sinking of ulcer, in 421 patients we have made, immediatly, a gastric-duodenal resection with B II technique. On the base of the good obtained results, the authors conclude that the contro-indications to this operation (age, general conditions of patient, old pierce, etc.) that were accepted by the most surgeons up to few years ago, must considered of secondary importance.
Assuntos
Úlcera Péptica Perfurada/cirurgia , Adolescente , Adulto , Idoso , Feminino , Humanos , Masculino , Métodos , Pessoa de Meia-Idade , Complicações Pós-Operatórias , PrognósticoRESUMO
The authors describe four cases of penetrating heart wounds: two from slash and puncture injuries and two from gunshot. Hearth lesions involved the left ventricle and pericardium in two cases, the right ventricle in one case, and only the diaphragmatic pericardium in one case. After a review of the literature, the authors discuss clinical and diagnostic problems relative to penetrating chest wounds and particularly those of the heart, stressing the need for very prompt diagnosis; as for actual treatment, they conclude that this can be carried out without any special problems in any Department of general emergency surgery.
Assuntos
Traumatismos Cardíacos/cirurgia , Ferimentos Penetrantes/cirurgia , Adolescente , Adulto , Emergências , Feminino , Traumatismos Cardíacos/patologia , Humanos , Masculino , Ferimentos por Arma de Fogo/patologia , Ferimentos por Arma de Fogo/cirurgia , Ferimentos Penetrantes/patologia , Ferimentos Perfurantes/patologia , Ferimentos Perfurantes/cirurgiaRESUMO
The authors describe 9 cases of primitive tumors of the small gut (3 adenocarcinomas, 2 lymphosarcomas, 1 fibroma, 1 angioma, and 2 leiomyosarcomas, one being the cause of active hemorrhage). In all cases the tumors were diagnosed only at operation, which was invariably necessitated by complications. Seven patients underwent emergency surgery for intestinal obstruction; one underwent an exploratory laparotomy for recurrent intestinal bleeding caused by an angioma of the ileum; and the last of this series (exceptional enough to warrant separate reporting elsewhere) was operated upon as an emergency case when the tumor, precisely a leiomyosarcoma, was perforated and actively bleeding.
Assuntos
Neoplasias Intestinais/complicações , Intestino Delgado/patologia , Adolescente , Adulto , Idoso , Emergências , Feminino , Humanos , Neoplasias Intestinais/patologia , Neoplasias Intestinais/cirurgia , Obstrução Intestinal/etiologia , Obstrução Intestinal/cirurgia , Intestino Delgado/cirurgia , Masculino , Pessoa de Meia-IdadeRESUMO
A case of perforated jejunal diverticulitis in a 87-year-old man is described and the literature is reviewed. Jejunal diverticulosis (JD) is estimated to occur in 0.02% to 1.3% of the adult population and is found most often in the elderly. The acute diverticulitis with perforation has been reported as high as 2.3% among patients with JD and is associated with high mortality. Clinical presentation mimic other more common acute intraperitoneal inflammatory conditions. Enteroclysis and abdominal CT are the most specific diagnostic tests. The common treatment is surgical resection of the involved segment. Laparoscopic resection and medical and medical/radiological approaches have also been proposed. Diagnostic and therapeutical aspects of this pathology are discussed.
Assuntos
Diverticulite/diagnóstico , Perfuração Intestinal/diagnóstico , Doenças do Jejuno/diagnóstico , Doença Aguda , Idoso , Idoso de 80 Anos ou mais , Diverticulite/cirurgia , Humanos , Perfuração Intestinal/cirurgia , Doenças do Jejuno/cirurgia , MasculinoRESUMO
Some second malignant neoplasms are increasingly being observed following NHL and a considerable amount of data has accumulated in the literature. The authors describe a case of 65-year old male who presented with submandibular adenopathy. Results of a biopsy of the mass surgically removed revealed low grade non-Hodgkin lymphoma. During the staging workup, a meningioma and a renal cell adenocarcinoma (RCC) were unexpectedly discovered and successively resected. The patient is currently alive with no evidence of metastatic diseases 12 months after diagnosis of non-Hodgkin's lymphoma (NHL), 10 months after meningioma resection and 8 months after RCC resection. The possibility of an underlying pathologic mechanism predisposing to multiple tumours should be considered. RCC and central nervous system (CNS) neoplasms are among second malignancies with higher incidences in non-Hodgkin lymphoma patients whereas with specific regard to meningioma, one of the most common benign intracranial tumours that sometimes shows biological aggressiveness and malignancy, we have currently no data in the literature. Increased risks for several malignancies occur late in the NHL follow-up period and are largely confined to patients receiving either radiation therapy or chemotherapy. On the other hand, increased risks for renal cancer have also been reported at less than one year after diagnosis of NHL and are present in all treatment subgroups (radiation therapy, chemotherapy, other-no treatment). Increased risks for CNS malignant neoplasms have also been reported at less than one year. The authors review the pathogenic significance of this case report neoplasms association in the light of the various explicative hypothesis of this concurrence. Possible immune mechanisms associated with these neoplasm are particularly pointed up.
Assuntos
Adenocarcinoma de Células Claras/diagnóstico , Neoplasias Encefálicas/diagnóstico , Neoplasias Renais/diagnóstico , Linfoma de Células B/cirurgia , Meningioma/diagnóstico , Segunda Neoplasia Primária/diagnóstico , Neoplasias da Glândula Submandibular/cirurgia , Idoso , Humanos , MasculinoRESUMO
Non-motor symptoms in Parkinson's disease (PD) have been often described at different stages of the disease but they are poorly understood. We observed specific phenotypes related to these symptoms in mice lacking the PD-associated GPR37/PAEL receptor. GPR37 is an orphan G-protein-coupled receptor highly expressed in the mammalian central nervous system. It is a substrate of parkin and it is involved in the pathogenesis of PD. GPR37 interacts with the dopamine transporter (DAT), modulating nigro-striatal dopaminergic signaling and behavioral responses to amphetamine and cocaine. GPR37 knockout (KO) mice are resistant to MPTP and exhibit several motor behavioral abnormalities related to altered dopaminergic system function. To evaluate non-motor behavioral domains, adult and aged, male and female GPR37 KO mice and their wild-type (WT) littermates were analyzed in a series of cross-sectional studies. Aged GPR37 KO female mice showed mild improvements in olfactory function, while anxiety and depression-like behaviors appeared to be significantly increased. A reduction of the startle response to acoustic stimuli was observed only in adult GPR37 KO mice of both genders. Furthermore, HPLC analysis of major neurotransmitter levels revealed gender differences in the striatum, hippocampus and olfactory bulb of mutant mice. The absence of GPR37 receptor could have a neuroprotective effect in an age and gender-dependent manner, and the study of this receptor could be valuable in the search for novel therapeutic targets.
Assuntos
Ansiedade/genética , Depressão/genética , Fenótipo , Receptores Acoplados a Proteínas G/genética , Fatores Etários , Animais , Encéfalo/metabolismo , Encéfalo/fisiologia , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurotransmissores/genética , Neurotransmissores/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Reflexo de Sobressalto/genética , Fatores Sexuais , Olfato/genéticaAssuntos
Hipertensão Renal/cirurgia , Obstrução da Artéria Renal/cirurgia , Adulto , Fatores Etários , Arteriosclerose/complicações , Prótese Vascular , Endarterectomia , Seguimentos , Humanos , Hipertensão Renal/mortalidade , Nefrectomia , Renografia por Radioisótopo , Obstrução da Artéria Renal/mortalidade , Trimetafano/farmacologiaAssuntos
Diuréticos/uso terapêutico , Hipertensão/tratamento farmacológico , Fitoterapia , Plantas Medicinais , Rauwolfia/uso terapêutico , Adulto , Idoso , Ensaios Clínicos como Assunto , Diuréticos/efeitos adversos , Combinação de Medicamentos , Tolerância a Medicamentos , Eletrocardiografia , Estudos de Avaliação como Assunto , Feminino , Humanos , Hipertensão Renal/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Náusea/induzido quimicamente , Rauwolfia/efeitos adversos , Vômito/induzido quimicamenteAssuntos
Arteriopatias Oclusivas/tratamento farmacológico , Arteriosclerose Obliterante/tratamento farmacológico , Flavonoides/uso terapêutico , Músculos/irrigação sanguínea , Vasodilatadores/uso terapêutico , Xenônio , Adulto , Idoso , Velocidade do Fluxo Sanguíneo , Preparações de Ação Retardada/uso terapêutico , Humanos , Perna (Membro)/irrigação sanguínea , Masculino , Pessoa de Meia-Idade , Extratos Vegetais/uso terapêutico , RadioisótoposAssuntos
Anormalidades Múltiplas , Axila/anormalidades , Mama/anormalidades , Adolescente , Feminino , HumanosRESUMO
The structure of a late endosomal compartment, which contains the bulk of the cation-independent mannose 6-phosphate receptor (MPR) in NRK cells, is documented using immunocytochemistry and cryo-sections, as well as conventional Epon sections. This compartment, which we refer to as the prelysosomal compartment (PLC), has a complex three-dimensional structure consisting of tubuloreticular domains in continuity with vesicular parts. The latter are characterized by a high density of internal membranes, which may be either tubular or sheet-like, that label extensively for the MPR. This structural organization was also maintained after fractionation in sucrose gradients. The amount of MPR immunolabelling was then quantitated with respect to the membrane surface areas of the four compartments where it is found: namely, the plasma membrane, early endosomes, the trans Golgi network and the PLC. The results showed that in NRK cells 90% of the labelling for the receptor was found in the PLC, with the rest distributed over the other three compartments. Cytochemical studies indicated that the PLC is the first structure along the endocytic pathway that gives a significant reaction for acid phosphatase. However, the PLC is clearly distinct from the MPR-negative lysosomes, which are also acid phosphatase-positive, since the two organelles could be physically separated from each other after fractionation on Percoll gradients.
Assuntos
Endocitose/fisiologia , Hexosefosfatos/metabolismo , Manosefosfatos/metabolismo , Organelas/ultraestrutura , Receptores de Superfície Celular/metabolismo , Animais , Fracionamento Celular , Células Cultivadas , Imuno-Histoquímica , Rim/ultraestrutura , Lisossomos/ultraestrutura , Microscopia Eletrônica , Ratos , Receptor IGF Tipo 2RESUMO
Lysosomes as well as a prelysosomal compartment rich in the mannose 6-phosphate receptor are clustered close to the Golgi apparatus in the perinuclear region of the microtubule organizing center in interphase human skin fibroblasts. The spatial organization of these organelles depends on an intact microtubule network. Depolymerization of the microtubules by treatment of cells with nocodazole leads to random scattering of Golgi elements, the prelysosomal compartment, and lysosomes throughout the cytoplasm. To test whether microtubules and the spatial organization of these organelles are important for efficient transport of lysosomal enzymes, the effect of microtubule depolymerization on the maturation of newly synthesized cathepsin D was studied. An up to fivefold inhibition of proteolytic maturation of cathepsin D was observed in drug-treated cells. This effect was due to a decreased rate of transport of cathepsin D from the Golgi apparatus to lysosomes. Depolymerization of microtubules did not inhibit transport of cathepsin D from the endoplasmic reticulum to the trans-Golgi network. Furthermore, synthesis of the phosphomannosyl marker present on cathepsin D was not affected by nocodazole. These results suggest that efficient transport of cathepsin D from the Golgi apparatus to a prelysosomal compartment and lysosomes is facilitated by microtubules and the spatial organization of these organelles.
Assuntos
Catepsina D/metabolismo , Complexo de Golgi/metabolismo , Lisossomos/metabolismo , Microtúbulos/fisiologia , Transporte Biológico/efeitos dos fármacos , Células Cultivadas , Eletroforese em Gel de Poliacrilamida , Fibroblastos , Imunofluorescência , Humanos , Microtúbulos/efeitos dos fármacos , Nocodazol , Receptor IGF Tipo 2 , Receptores de Superfície Celular/metabolismoRESUMO
We report the cloning of the mouse ortholog of the human GPR37 gene, which encodes an orphan G-protein-coupled receptor highly expressed in brain tissues and homologous to neuropeptide-specific receptors (D. Marazziti et al., 1997, Genomics 45: 68-77; Z. Zeng et al., 1997, Biochem. Biophys. Res. Commun. 233: 559-567). The genomic organization of the GPR37 gene is conserved in both mouse and human species with a single intron interrupting the receptor-coding sequence within the presumed third transmembrane domain. Comparative genetic mapping of the GPR37 gene showed that it maps to a conserved chromosomal segment on proximal mouse chromosome 6 and human chromosome 7q31. The mouse Gpr37 gene contains an open reading frame coding for a 600-amino-acid protein 83% identical to the human GPR37 gene product. The predicted mouse GPR37 protein contains seven putative hydrophobic transmembrane domains, as well as a long (249 amino acid residues), arginine- and proline-rich amino-terminal extracellular domain, which is also a distinctive feature of the human GPR37 receptor. Northern blot analysis of mouse tissues with Gpr37-specific probes revealed a main 3.8-kb mRNA and a much less abundant 8-kb mRNA, both expressed in the brain. A 3-kb mRNA is also expressed in the testis. Both the mouse and the human GPR37 genes may belong to a class of highly conserved mammalian genes encoding a novel type of G-protein-coupled receptor predominantly expressed in the brain.
Assuntos
Encéfalo/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Proteínas de Membrana , Proteínas do Tecido Nervoso/genética , Receptores de Superfície Celular/genética , Receptores Acoplados a Proteínas G , Testículo/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Mapeamento Cromossômico , Cromossomos Humanos Par 7/genética , Clonagem Molecular , Primers do DNA/genética , DNA Complementar/genética , Expressão Gênica , Humanos , Masculino , Camundongos , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Superfície Celular/química , Homologia de Sequência de Aminoácidos , Especificidade da EspécieRESUMO
We have developed a replica filter assay that permits the direct identification of bacterial colonies expressing membrane receptors. E.coli transformed with appropriate phage or plasmid vectors containing human beta-adrenergic receptor cDNAs were grown on LB/agar plates. Bacterial colonies transferred onto nitrocellulose filters showed specific [125I]-iodocyanopindolol binding. beta-adrenergic receptors expressed in bacteria retained their pharmacological properties when transferred onto filters. This strategy, which is considerably simplified and more rapid compared to similar methods based upon expression of receptor genes in eukaryotic cells, may be a useful tool for cloning membrane receptors.