RESUMO
Activated mast cells trigger edema in allergic and inflammatory disease. We report a paracrine mechanism by which mast cell-released heparin increases vascular permeability in vivo. Heparin activated the protease factor XII, which initiates bradykinin formation in plasma. Targeting factor XII or kinin B2 receptors abolished heparin-triggered leukocyte-endothelium adhesion and interfered with a mast cell-driven drop in blood pressure in rodents. Intravital laser scanning microscopy and tracer measurements showed heparin-driven fluid extravasation in mouse skin microvessels. Ablation of factor XII or kinin B2 receptors abolished heparin-induced skin edema and protected mice from allergen-activated mast cell-driven leakage. In contrast, heparin and activated mast cells induced excessive edema in mice deficient in the major inhibitor of factor XII, C1 esterase inhibitor. Allergen exposure triggered edema attacks in hereditary angioedema patients, lacking C1 esterase inhibitor. The data indicate that heparin-initiated bradykinin formation plays a fundamental role in mast cell-mediated diseases.
Assuntos
Bradicinina/biossíntese , Síndrome de Vazamento Capilar/fisiopatologia , Permeabilidade Capilar/fisiologia , Heparina/fisiologia , Mastócitos/metabolismo , Anafilaxia Cutânea Passiva/fisiologia , Animais , Bradicinina/genética , Síndrome de Vazamento Capilar/etiologia , Adesão Celular , Proteína Inibidora do Complemento C1/fisiologia , Edema/etiologia , Edema/fisiopatologia , Células Endoteliais/patologia , Ativação Enzimática , Fator XII/fisiologia , Heparina/metabolismo , Hipotensão/etiologia , Hipotensão/fisiopatologia , Imunoglobulina E/imunologia , Sistema Calicreína-Cinina/fisiologia , Leucócitos/fisiologia , Masculino , Camundongos , Comunicação Parácrina/fisiologia , Plasma , Ratos , Transdução de Sinais/fisiologia , Pele/irrigação sanguíneaRESUMO
Contact to polyanions induces autoactivation of the serine protease factor XII that triggers the kallikrei-kinin system. Recent studies indicate that polysaccharide-induced autoactivation of factor XII has a role in allergy-related vascular leakage, and angioedema. Here, we characterize in vivo effects of the synthetic polysaccharide dextran sulfate in human plasma and in rodent models. Minute amounts of high-molecular-weight dextran sulfate-initiated factor XII-autoactivation and triggered formation of the inflammatory mediator bradykinin via plasma kallikrein-mediated cleavage of high-molecular-weight kininogen. High-molecular-weight kininogen fragments, containing the HKH20 sequence in domain D5H, blocked dextran sulfate-initiated bradykinin-generation by depleting plasma Zn2+ ions. Topical application of high molecular weight dextran sulfate increased leakage in murine skin microvessels, in a bradykinin-dependent manner. Intravital laser scanning microscopy showed a greater than two-fold elevated and accelerated fluid extravasation in C1 esterase inhibitor deficient mice that lack the major inhibitor of factor XII, compared to wild-type controls. Intra-arterial infusion of dextran sulfate induced a rapid transient drop in arterial blood pressure in rats and preinjection of kinin B2 receptor antagonists or HKH20 peptide blunted dextran sulfate-triggered hypotensive reactions. The data characterize dextran sulfate as a potent in vivo activator of factor XII with implications for bradykinin-mediated vascular permeability and blood pressure control.
Assuntos
Bradicinina/metabolismo , Sulfato de Dextrana/farmacologia , Hipotensão/sangue , Hipotensão/patologia , Calicreínas/metabolismo , Zinco/sangue , Animais , Ânions/sangue , Bradicinina/sangue , Permeabilidade Capilar , Sulfato de Dextrana/sangue , Sulfato de Dextrana/química , Fator XII/metabolismo , Fator XIIa/metabolismo , Humanos , Calicreínas/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Ratos Wistar , Zinco/deficiênciaRESUMO
In 2000, the authors found endemic infections of mouse hepatitis virus, minute virus of mice, Syphacia obvelata, and Myobia musculi among mice in a large barrier facility at the University of Mainz. To eliminate the infections, they subdivided the facility into two distinct hygiene units. However, architectural constraints made it impossible to completely separate the HVAC systems of both hygiene units and to establish adequate personnel locks. To compensate for these suboptimal barrier conditions of the two newly established units, the authors replaced the open-top caging and open-servicing system with filter-top cages that were manipulated in cage-changing stations. The authors then depopulated the two units in series, independently eliminating the contaminated mice and restocking the units with SPF animals. In spite of the high infection pressure and the suboptimal barrier conditions, the authors had only a single case of recontamination.
Assuntos
Criação de Animais Domésticos/instrumentação , Criação de Animais Domésticos/métodos , Descontaminação/métodos , Arquitetura de Instituições de Saúde , Abrigo para Animais , Animais , Surtos de Doenças/prevenção & controle , Surtos de Doenças/veterinária , Higiene , Camundongos , Organismos Livres de Patógenos EspecíficosRESUMO
The objective of this study was to evaluate the piglet and the mouse as model systems for preterm cortical development. According to the clinical context, we used non invasive EEG recordings. As a prerequisite, we developed miniaturized Ag/AgCl electrodes for full band EEG recordings in mice and verified that Urethane had no effect on EEG band power. Since mice are born with a "preterm" brain, we evaluated three age groups: P0/P1, P3/P4 and P13/P14. Our aim was to identify EEG patterns in the somatosensory cortex which are distinguishable between developmental stages and represent a physiologic brain development. In mice, we were able to find clear differences between age groups with a simple power analysis of EEG bands and also for phase locking and power spectral density. Interhemispheric coherence between corresponding regions can only be seen in two week old mice. The canolty maps for piglets as well as for mice show a clear PAC (phase amplitude coupling) pattern during development. From our data it can be concluded that analytic tools relying on network activity, as for example PAC (phase amplitude coupling) are best suited to extract basic EEG patterns of cortical development across species.
RESUMO
Germ cell tumors (GCTs) are among the most common malignancies in young men. We have previously documented that patients with GCT frequently produce serum antibodies directed against proteins encoded by human endogenous retrovirus (HERV) type K sequences. Transcripts originating from the env gene of HERV-K, including the rec-relative of human immunodeficiency virus rev, are highly expressed in GCTs. We report here that mice that inducibly express HERV-K rec show a disturbed germ cell development and may exhibit, by 19 months of age, changes reminiscent of carcinoma in situ, the predecessor lesion of classic seminoma in humans. This provides the first direct evidence that the expression of a human endogenous retroviral gene previously established as a marker in human germ cell tumors may contribute to organ-specific tumorigenesis in a transgenic mouse model.
Assuntos
Carcinoma in Situ/etiologia , Retrovirus Endógenos/genética , Células Germinativas/citologia , Neoplasias Embrionárias de Células Germinativas/metabolismo , Proteínas do Envelope Viral/fisiologia , Processamento Alternativo , Animais , Apoptose , Western Blotting , Carcinoma in Situ/virologia , Linhagem Celular Tumoral , Germinoma/metabolismo , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Microscopia de Fluorescência , Modelos Genéticos , Estrutura Terciária de Proteína , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Túbulos Seminíferos/metabolismo , Fatores de Tempo , Proteínas do Envelope Viral/metabolismoRESUMO
O(6)-Substituted guanine derivatives are powerful agents used for tumor cell sensitization by inhibition of the DNA repair enzyme O(6)-methylguanine-DNA methyltransferase (MGMT). To provide targeted accumulation of MGMT inhibitors in tumor tissue as well as tools for in vivo imaging, we synthesized iodinated C(8)-alkyl-linked glucose conjugates of 2-amino-6-(5-iodothenyl)-9H-purine (O(6)-(5-iodothenyl) guanine, ITG) and 2-amino-6-(3-iodobenzyloxy)-9H-purine (O(6)-(5-iodobenzyl) guanine, IBG). These compounds have MGMT inhibitor constants (IC(50) values) of 0.8 and 0.45 microM for ITGG and IBGG, respectively, as determined in HeLa S3 cells after 2-h incubation with inhibitor. To substantiate that the (131)I-(hetero)arylmethylene group at the O(6)-position of guanine is transferred to MGMT, both the glucose conjugated inhibitors ITGG and IBGG and the corresponding nonglucose conjugated compounds ITG and IBG were labeled with iodine-131. The radioiodinations of all compounds with [(131)I]I(-) were performed with radiochemical yields of >70% for the destannylation of the corresponding tri-n-butylstannylated precursors. The binding ability of [(131)I]ITGG, [(131)]IBGG, [(131)I]ITG, and [(131)I]IBG to purified MGMT was tested. All radioactive compounds were substrates for MGMT, as demonstrated using a competitive repair assay. The newly synthesized radioactive inhibitors were utilized to study ex vivo biodistribution in mice, and the tumor-to-blood ratio of tissue uptake of [(131)I]IBG and [(131)I]IBGG was determined to be 0.24 and 0.76 after 0.5 h, respectively.
Assuntos
Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Glucose/química , Guanina/análogos & derivados , Guanina/síntese química , Guanina/farmacologia , O(6)-Metilguanina-DNA Metiltransferase/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Inibidores Enzimáticos/química , Guanina/química , Células HeLa , Humanos , Técnicas In Vitro , Isótopos de Iodo , Camundongos , Camundongos Nus , Estrutura Molecular , Relação Estrutura-Atividade , Fatores de Tempo , Transplante Heterólogo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND/AIMS: Interferon gamma (IFNgamma) controls hepatitis B virus replication. As systemic application may cause severe adverse effects, approaches of liver-directed IFNgamma gene therapy may represent an attractive alternative for treatment of chronic viral hepatitis B and thus needs testing in vivo in suitable animal models. METHODS: We therefore crossbred Alb-1HBV transgenic mice overexpressing the large HBV surface protein (LHBs) in their livers and developing LHBs storage disease and ground glass hepatocyte appearance with SAP-IFNgamma transgenic animals previously shown to exhibit constitutive hepatic IFNgamma expression, and analyzed the resulting double-transgenic offspring. RESULTS: We found that IFNgamma coexpression significantly reduced hepatic LHBs expression and thereby inhibited hepatocellular LHBs storage disease and ground glass hepatocyte appearance. The beneficial antiviral IFNgamma effects as observed in Alb1-HBV SAP-IFNgamma double-transgenic livers were associated with significantly elevated serum ALT concentrations, massive mononuclear cell infiltrates, appearance of Councilman bodies, and increased alpha-PARP (poly(ADP-ribose) polymerase cleavage). CONCLUSIONS: Exacerbation of hepatic necroinflammation and increased hepatocellular apoptosis rate in IFNgamma-expressing Alb1-HBV transgenic livers suggest that special precautions be taken for testing approaches of liver-specific IFNgamma expression in patients with chronic hepatitis B.