RESUMO
The process of liver regeneration involves the concerted action of certain growth factors, which stimulate hepatocyte proliferation, and other antiproliferative factors, which prevent uncontrolled growth of this organ. Some of the biological actions of insulin-like growth factor-II (IGF-II), a mitogenic polypeptide closely related to insulin, may be mediated by the IGF-II receptor. This receptor consists of a single chain extracellular domain and a very small cytoplasmic domain, and can bind lysosomal enzymes that contain mannose-6-phosphate (M-6-P) residues. Since these enzymes may be involved in remodelling processes in certain tissues, we measured the expression of the IGF-II/M-6-P receptor in the liver after subtotal hepatectomy. Binding of [125I]IGF-II to crude plasma membranes from regenerating liver was maximal 2 days after hepatectomy (4.9% specific binding/60 micrograms protein) and subsequently decreased. Both control livers (livers removed at the time of operation) and sham-operated control livers demonstrated specific [125I]IGF-II binding of 1.1% throughout the experimental period. This increase in binding in regenerating liver was shown to be associated with an increase in the concentration of IGF-II receptor protein by means of Western blot analysis using a polyclonal anti-IGF-II/M-6-P receptor antiserum (3637). Similarly, steady state levels of IGF-II/M-6-P receptor mRNA, measured by solution hybridization/RNase protection assays, were significantly increased in the regenerating liver (2.0-fold over the control value 2 days after hepatectomy). Five and 10 days postsurgery, the levels of IGF-II receptor mRNA were markedly reduced, and they were even lower than the levels in control livers.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Regeneração Hepática/fisiologia , Receptores de Superfície Celular/metabolismo , Animais , Western Blotting , Membrana Celular/metabolismo , Hepatectomia , Fator de Crescimento Insulin-Like II/metabolismo , Fígado/metabolismo , Masculino , Manosefosfatos/metabolismo , Hibridização de Ácido Nucleico , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos , Receptor IGF Tipo 2 , Receptores de Superfície Celular/genética , Receptores de Somatomedina , RibonucleasesRESUMO
Infusing the DR-/dim fraction of bone marrow cells (BMC) from an allogeneic kidney donor into rabbit antithymocyte globulin-treated transplant recipients delivers a tolerogenic signal, leading to functional allograft tolerance in rhesus monkeys without additional drug therapy. Our updated results in an expanded series show a median 131-day graft survival of recipients given DR-/dim donor BMC with a 23% 1-year survival (P < 0.00001 vs. rabbit antithymocyte globulin controls). Removing DRbright cells from donor BMC appeared to have a significant effect (P < 0.05). We have further investigated the tolerogenic mechanism within the experimental framework of the veto hypothesis in this preclinical model. In limiting dilution assays, we demonstrated the donor specificity of clonal inactivation of CTL precursors (CTLp) after in vitro or in vivo exposure to DR-/dim donor BMC, confirming specific tolerance. Additionally, in vitro studies confirmed the allogeneic specificity of CTLp inactivation in 3-cell MLR assays; minimal bystander effects were seen on normal CTLp responses to third party stimulator cells, while CTLp responses to the BMC donor's cells were abrogated in the same cultures. BMC mediating the veto effect were found to be resistant to L-leucyl-L-leucine methyl ester (Leu-leu-OMe), which excluded BMC-mediated cytotoxicity by NK or lymphokine-activated killer cells, CTL, or activated macrophages. In contrast, veto activity was abolished if the BMC were pretreated with either high dose UV-B light irradiation, mitomycin, or gamma-irradiation, indicating that BMC contained a UV-B-sensitive precursor of the veto effector, and that a proliferative step separated the two. Irradiation of DR-/dim donor BMC or administration of cyclophosphamide after infusion of nonirradiated BMC prevented the tolerogenic effect. Only recipients given nonirradiated DR-/dim donor BMC demonstrated PBL chimerism, which associated with functional deletion of antidonor CTLp and duration of graft survival. The Leu-leu-OMe resistance and the other properties of the allogeneic monkey CD3- CD2+ CD8+ BMC subpopulation that exhibits tolerance-promoting activity in vitro and in vivo lead us to postulate that a donor BMC-derived precursor population, possibly a dendritic cell population, may induce allogeneic unresponsiveness in this model.
Assuntos
Medula Óssea/imunologia , Transplante de Rim/imunologia , Animais , Sequência de Bases , Quimera , Primers do DNA/química , Antígenos de Histocompatibilidade Classe II/imunologia , Tolerância Imunológica , Depleção Linfocítica , Macaca mulatta , Masculino , Dados de Sequência Molecular , Linfócitos T Citotóxicos/imunologiaRESUMO
The immunotoxicity of methyl isocyanate (MIC) was evaluated in female B6C3F1 mice exposed via inhalation to 0, 1, or 3 ppm for 6 hr per day on 4 consecutive days. The antibody response to sheep erythrocytes and natural killer cell activity were found to be unaffected by MIC exposure. Although lymphoproliferative responses to mitogens were moderately suppressed by MIC, the differences were not statistically significant. The response of splenic lymphocytes to allogeneic leukocytes in a mixed leukocyte response (MLR) was suppressed in a dose-related fashion and was significantly different from the control response at the 3 ppm level. This effect was thought to be secondary and a result of general toxicity, rather than a direct effect of MIC on the immune system. Furthermore, resistance to the infectious agents Listeria monocytogenes, mouse malaria parasite, and influenza virus, or to transplantable tumor cells was not compromised by MIC exposure. Thus, the immune system does not appear to be a primary target for MIC toxicity.
Assuntos
Cianatos/toxicidade , Sistema Imunitário/efeitos dos fármacos , Isocianatos , Animais , Formação de Anticorpos/efeitos dos fármacos , Cianatos/administração & dosagem , Citotoxicidade Imunológica/efeitos dos fármacos , Feminino , Células Matadoras Naturais/efeitos dos fármacos , Ativação Linfocitária/efeitos dos fármacos , Teste de Cultura Mista de Linfócitos , CamundongosRESUMO
Acute exposure of adult mice to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) results in a selective suppression of proliferating cells of the immune system, including hematopoietic stem cells and B cells. Suppression of B cell-mediated or humoral immunity, in turn, results in altered host resistance to the parasite Plasmodium yoelii, a malaria model. Data presented in this study demonstrate a direct effect of TCDD on cultured lymphoctes resulting in a selective inhibition of the differentiation of B cells into antibody-secreting cells. A structure-activity study suggested that this inhibition was mediated by the Ah receptor. As previously defined by receptor binding studies in hepatic cytosol, active congeners were inhibitory, whereas inactive congeners were without effect. Using lymphocytes from congenic mice which differ only at the Ah locus, it was determined that the Ahbb-derived cells were inhibited by TCDD in vitro, whereas the Ahdd-derived cells were not. B cell differentiation thus provides a valuable model for understanding TCDD toxicity as well as the role of the Ah receptor in growth and differentiation.
Assuntos
Linfócitos B/citologia , Dioxinas/farmacologia , Dibenzodioxinas Policloradas/farmacologia , Animais , Formação de Anticorpos/efeitos dos fármacos , Benzofuranos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Feminino , Malária/imunologia , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Relação Estrutura-Atividade , TimectomiaRESUMO
A practical and repeatable method for measurement of pulsatile insulin release in the dog was developed by direct, chronic cannulation of the portal vein cephalad to the pancrease and the entry point of the cranial pancreaticoduodenal vein, with subsequent exteriorization of the sampling catheter. Mixed breed male dogs of various body weights and ages underwent midline laparotomy for exposure of the portal vein system. After exposure the portal vein was dissected free and cannulated through a purse-string suture placed just caudad (approximately 1 cm) to the entry point of the cranial pancreaticoduodenal vein with a cuffed, medical grade, silastic catheter. The catheter was advanced cranial to a point just distal to the level of the liver and secured in place via the purse-string suture and a single stay suture. The catheter was then passed through the dorsolateral abdominal wall and routed subcutaneously to a point between the shoulder blades. After catheter plug attachment the catheter was tested for patency, flushed, heparin locked, then was secured to the subcutaneous tissue between the scapulas. The dorsal skin wound and abdominal incision were closed in a routine manner, and the dogs recovered from anesthesia. Catheter patency and function in all dogs were maintained for periods up to 4 weeks. No postoperative complications such as catheter tract infections occurred, maintenance was minimal, and all dogs were afforded complete freedom of movement and activity throughout the study, with no requirement for cumbersome jackets or wraps.
Assuntos
Coleta de Amostras Sanguíneas/métodos , Cateterismo/métodos , Insulina/sangue , Insulina/metabolismo , Periodicidade , Veia Porta , Procedimentos Cirúrgicos Operatórios/métodos , Animais , Cães , Secreção de Insulina , MasculinoRESUMO
Mice given pharmacological levels of the synthetic estrogen diethylstilbestrol demonstrated a marked increase in susceptibility to infection with Listeria monocytogenes. Experiments were performed in an effort to determine the mechanism(s) by which estrogen treatment increases the susceptibility of mice to L. monocytogenes infection. Estrogen exposure depressed the in vivo proliferative response of splenic lymphocytes to L. monocytogenes, which correlated with the decreased in vitro response of these cells to phytohemagglutinin. Interleukin 2 (IL 2) production by splenic lymphocytes from estrogen-treated mice was decreased, although these cells were capable of proliferating normally in response to exogenous IL 2. Interleukin 1 production by peritoneal macrophages was not depressed by estrogen exposure. The number of bacteria observed in the spleens of estrogen-exposed mice challenged with L. monocytogenes was reduced by IL 2 administration. Thus, estrogens may decrease host resistance to L. monocytogenes by inhibiting IL 2 production and the subsequent proliferation of antigen-sensitized T lymphocytes required for recovery.
Assuntos
Estrogênios/farmacologia , Interleucina-1/biossíntese , Interleucina-2/biossíntese , Listeriose/imunologia , Ativação Linfocitária/efeitos dos fármacos , Animais , Dietilestilbestrol/farmacologia , Feminino , Imunidade/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Baço/citologia , Baço/efeitos dos fármacos , Baço/imunologiaRESUMO
Arachidonic acid metabolites have been shown to exert a variety of regulatory effects on cellular activation and proliferation. Recently, a role for these products as regulators of hematopoiesis was suggested and evidence provided that products of the lipoxygenase pathway, specifically leukotrienes, are essential for human myeloid colony formation in vitro. In this report the broader role of these metabolites in hematopoiesis was examined using murine bone marrow stem cell assays for both myeloid and lymphoid cell lines. The effects of lipoxygenase and/or cyclooxygenase pathway inhibitors on stem cell colony formation were evaluated and compared to qualitative and quantitative changes in arachidonic acid metabolism that occurred in similarly treated bone marrow cell cultures. Interruption of the lipoxygenase pathway by esculetin or nordihydroguaiaretic acid resulted in decreased colony formation in both lymphoid and myeloid stem cells. This inhibition of colony growth was partly reversed by the addition of leukotrienes and was particularly evident in B-cell progenitor cultures to which was added LTB4. Inhibition of the cyclooxygenase pathway by indomethacin or ibuprofen had a slight stimulatory effect on myeloid colony formation, while slightly inhibiting the formation of lymphoid colonies. These results support a direct role for lipoxygenase products in myeloid colony formation and lymphoid stem cell proliferation. A more complex role for cyclooxygenase metabolites in the hematopoietic process appears probable.
Assuntos
Ácidos Araquidônicos/metabolismo , Hematopoese/efeitos dos fármacos , Células-Tronco Hematopoéticas/metabolismo , Ácido 12-Hidroxi-5,8,10,14-Eicosatetraenoico , Animais , Linfócitos B/metabolismo , Inibidores de Ciclo-Oxigenase , Granulócitos/metabolismo , Ácidos Hidroxieicosatetraenoicos/análise , Ibuprofeno/farmacologia , Técnicas In Vitro , Indometacina/farmacologia , Leucotrienos/fisiologia , Inibidores de Lipoxigenase , Macrófagos/metabolismo , Masoprocol/farmacologia , Camundongos , Prostaglandinas/análise , Prostaglandinas/fisiologia , Tromboxano B2/análise , Umbeliferonas/farmacologiaRESUMO
Induction of anesthesia is accompanied by modest hyperglycemia and a decreased plasma insulin concentration. Most insulin is secreted in discrete pulses occurring at approximately 6- to 8-min intervals. We sought to test the hypothesis that anesthesia inhibits insulin release by disrupting pulsatile insulin secretion in a canine model by use of direct portal vein sampling. We report that induction of anesthesia causes an abrupt decrease in the insulin secretion rate (1.1 +/- 0.2 vs. 0.7 +/- 0.1 pmol. kg(-1). min(-1), P < 0.05) by suppressing insulin pulse mass (630 +/- 121 vs. 270 +/- 31 pmol, P < 0.01). Anesthesia also elicited an approximately 30% higher increase in insulin pulse frequency (P < 0.01) and more orderly insulin concentration profiles (P < 0.01). These effects were evoked by either sodium thiamylal or nitrous oxide and isoflurane. In conclusion, anesthesia represses insulin secretion through the mechanism of a twofold blunting of pulse mass despite an increase in orderly pulse frequency. These data thus unveil independent amplitude and frequency controls of beta-cells' secretory activity in vivo.
Assuntos
Anestesia , Insulina/metabolismo , Anestésicos Inalatórios/farmacologia , Anestésicos Intravenosos/farmacologia , Animais , Glicemia/metabolismo , Cateterismo Periférico , Cães , Insulina/sangue , Secreção de Insulina , Isoflurano/farmacologia , Óxido Nitroso/farmacologia , Veia Porta/fisiologia , Tiamilal/farmacologiaRESUMO
Transmissible pathogenic and opportunistic zoonotic enteric bacteria comprise a recognized occupational health threat to exposed humans from non-human primates (NHPs). In an effort to evaluate the occurrence of selected enteric organisms with zoonotic and biohazard potential in a research colony setting, we performed a prevalence study examining 61 juvenile and young adult rhesus macaques participating in a transplant immunology project. Primary emphasis was directed specifically to detection of pathogenic enteric Yersinia, less well-documented and reported NHP pathogens possessing recognized significant human disease potential. NHPs were surveyed by rectal culture during routine health monitoring on three separate occasions, and samples incubated using appropriate media and specific selective culture methods. Enteric organisms potentially transmissible to humans were subcultured and identified to genus and species. Significant human pathogens of the Salmonella/Shigella, Campylobacter, and enteric Yersinia groups were not isolated throughout the survey, suggesting prevalence of these organisms may generally be quite low.
Assuntos
Animais de Laboratório , Infecções por Enterobacteriaceae/transmissão , Macaca mulatta , Saúde Ocupacional , Yersiniose/transmissão , Yersinia/isolamento & purificação , Zoonoses , Técnicos em Manejo de Animais , Animais , Coleta de Dados , Sistema Digestório/microbiologia , Infecções por Enterobacteriaceae/veterinária , Humanos , Masculino , Prevalência , Yersiniose/veterináriaRESUMO
Delivery of intradermally focused nanosecond laser pulses with small energy as an alternate technique applicable to clinical procedures in dermatological and plastic surgery is an area of relatively new interest with multiple potential applications. We assessed this approach on common tattoo pigments in dermis in an in vivo study using a wavelength of 1064 nm. Paired micropigs were tattooed with standard blue, black, green and red pigments. The tattoos were allowed to mature and then treated by 12 ns pulses in a focused beam of 11.4 degrees cone angle. Visual observation and histological analysis of biopsies were performed to evaluate results. Significant reduction in pulse energy and collateral damage was achieved with pulse energy ranging between 38 to 63 mJ. Blue and black tattoos were found to respond well from a clinical standpoint. The depth dependence of tissue response and pigment redistributions at 1 hour, 1 week and 1 month after laser treatment was quantitatively analysed through biopsies and a strong relationship was demonstrated between tattoo response and laser-induced dermal vacuolation. The optical absorption coefficients of the four tattoo pigments were measured to be approximately the same and the laser-induced plasma is suggested to be responsible for the pigment redistribution. As we hypothesised, intradermal focusing of nanosecond pulses significantly reduced required pulse energy for tattoo ablation to about 60 mJ or less. These results stimulate a number of additional questions relevant not only to clinical applications but also to the understanding of the fundamental process of laser-pigment interaction in the dermis as it relates to tattoo removal.
Assuntos
Terapia a Laser , Tatuagem , Animais , Corantes , Procedimentos Cirúrgicos Dermatológicos , Feminino , Pele/patologia , Suínos , Porco MiniaturaRESUMO
The relationship between immune function and chemically induced hepatocarcinogenesis was studied employing an in vivo murine model. Neonatal B6C3F1 mice were given a single carcinogenic dose of diethylnitrosamine (DEN) and the time-response kinetics for the early (foci of alteration) and late (adenomas/carcinomas) phases of hepatocellular carcinogenesis were compared to changes in hematopoiesis and immune functions associated with immune surveillance and natural resistance. Increases in hematopoiesis occurred just prior to or concurrent with the appearance of hepatocellular carcinomas, while increased macrophage and natural killer cell cytotoxicity and suppression of cell-mediated immunity occurred following tumor appearance and progressed with increasing tumor burden. Neither immunological nor hematopoietic changes were associated with early phases of hepatocarcinogenesis, as monitored by the appearance of altered hepatocellular foci. Although changes in hematopoiesis may represent an early indicator for hepatocarcinogenesis in the mouse tumor model, the data suggest that altered immune surveillance and natural resistance are not factors in the development of chemically induced hepatocellular tumors, and the changes in immune function are probably secondary to tumor development.
Assuntos
Neoplasias Hepáticas Experimentais/imunologia , Animais , Dietilnitrosamina , Feminino , Hematopoese , Células Matadoras Naturais/imunologia , Neoplasias Hepáticas Experimentais/induzido quimicamente , Masculino , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Linfócitos T/imunologiaRESUMO
The effects of methyl isocyanate (MIC) on systemic immunity were evaluated in female B6C3F1 mice exposed via inhalation to 0, 1, or 3 ppm for 6 hr per day on four consecutive days. Humoral immunity, measured as the antibody response to sheep erythrocytes, and natural killer cell activity were not affected by MIC. Furthermore, resistance to the infectious agents Listeria monocytogenes, mouse malaria parasite, and influenza virus, or to B16F10 transplantable tumor cells, was not compromised by MIC exposure. Although lymphoproliferative responses to mitogens were not significantly suppressed, the response of splenic lymphocytes to allogeneic leukocytes in a mixed leukocyte response (MLR) was suppressed in a dose-related fashion and differed significantly from the control response at the 3-ppm level. These studies indicate that MIC exposure in mice does not severely alter systemic immunity. The moderate changes detected in immune function may be a secondary consequence of respiratory toxicity which occurred in these animals.