RESUMEN
One of the major drawbacks in the use of biomedical materials is the occurrence of biomaterials centered infections. After implantation, the host interacts with a biomaterial by forming a conditioning film on its surface and an immune reaction towards the foreign material. When microorganisms can reach the biomaterials surface they can adhere to it. Adhesion of microorganisms to an implant is mediated by their physico-chemical surface properties and the properties of the biomaterials surface itself. Subsequent surface growth of the microorganisms will lead to a mature biofilm and infection, which is difficult to eradicate by antibiotics. The purpose of this review is to give an overview of the mechanisms involved in biomaterials centered infection and the possible methods to prevent these infections.
RESUMEN
A 62-year-old female patient is described who developed toxic epidermal necrolysis (TEN) after medication with phenytoin and oxazepam. Initially phenytoin was discontinued and dexamethasone pulse therapy (1.5 mg/kg on 3 consecutive days) was initiated on the tenth day of skin disease. This resulted in clinical improvement. Histologically re-epithelialization could be demonstrated below the necrotic epidermis. However, on the eighteenth day of skin disease (10 days after discontinuation of phenytoin and 8 days after the start of dexamethasone pulse therapy), a histologically verified rebound-TEN developed with a detachment of 95%. Oxazepam was stopped and a second series of dexamethasone pulse therapy was given. Re-epithelialization began within 24 h of the start of the second series of dexamethasone pulse therapy, and continued to almost complete recovery within 1 week.
Asunto(s)
Dexametasona/administración & dosificación , Glucocorticoides/administración & dosificación , Hipnóticos y Sedantes/efectos adversos , Oxazepam/efectos adversos , Síndrome de Stevens-Johnson/tratamiento farmacológico , Síndrome de Stevens-Johnson/etiología , Anticonvulsivantes/uso terapéutico , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/secundario , Quimioterapia Combinada , Epidermis/efectos de los fármacos , Epidermis/patología , Femenino , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Persona de Mediana Edad , Fenitoína/uso terapéutico , Quimioterapia por Pulso , Síndrome de Stevens-Johnson/patologíaRESUMEN
Late biomaterial-centered infection is a major complication associated with the use of biomaterial implants. In this study biomaterials that had been implanted subcutaneously in rats were hematogenously challenged with bacteria 4 weeks after implantation. Bacteria were spread either by intravenous injection or by stimulation of bacterial translocation. It was found that none of the biomaterials was infected by hematogenous spread, whereas 5% of the implants were infected by perioperative contamination. We conclude that late hematogenous infection of subcutaneous biomaterials does not occur in the rat. For humans as well, there are growing doubts whether implants actually become infected through hematogenous routes; it is thought that late infections may be caused by delayed appearance of perioperatively introduced bacteria.
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Implantes Experimentales/microbiología , Infecciones por Pseudomonas/microbiología , Infecciones Estafilocócicas/microbiología , Animales , Traslocación Bacteriana/fisiología , Materiales Biocompatibles/efectos adversos , Modelos Animales de Enfermedad , Implantes Experimentales/efectos adversos , Inyecciones Intravenosas , Inyecciones Subcutáneas , Masculino , Ratas , Ratas EndogámicasRESUMEN
The development of transplant arteriosclerosis (TA) is today's most important problem in clinical organ transplantation. Histologically, TA is characterized by perivascular inflammation and progressive intimal thickening. Current thought on this process of vascular remodeling assumes that neointimal vascular smooth muscle (VSM) cells and endothelium in TA are graft-derived, holding that medial VSM cells proliferate and migrate into the subendothelial space in response to signals from inflammatory cells and damaged graft endothelium. Using MHC class I haplotype-specific immunohistochemical staining and single-cell PCR analyses, we show that the neointimal alpha-actin-positive VSM cells in rat aortic or cardiac allografts are of recipient and not of donor origin. In aortic but not in cardiac allografts, recipient-derived endothelial cells (ECs) replaced donor endothelium. Cyclosporine treatment prevents neointima formation and preserves the vascular media in aortic allografts. Recipient-derived ECs do not replace graft endothelium after cyclosporine treatment. We propose that, although it progresses beyond the needs of functional repair, TA reflects the activity of a normal healing process that restores vascular wall function following allograft-induced immunological injury.
Asunto(s)
Actinas/metabolismo , Arteriosclerosis/etiología , Músculo Liso Vascular/citología , Trasplante/efectos adversos , Túnica Íntima/metabolismo , Animales , Aorta/trasplante , Arteriosclerosis/fisiopatología , Ciclosporina/farmacología , Femenino , Genes MHC Clase I/genética , Trasplante de Corazón , Humanos , Inmunohistoquímica , Inmunosupresores/farmacología , Masculino , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/metabolismo , Reacción en Cadena de la Polimerasa , Ratas , Ratas Endogámicas , Túnica Íntima/citología , Túnica Íntima/efectos de los fármacosRESUMEN
BACKGROUND: We showed previously that our intrathymic immune modulation protocol induces virtually permanent graft survival of simultaneously transplanted cardiac allografts in MHC-incompatible rat strain combinations. It is, however, unknown whether this procedure prevents the development of graft arterial disease (GAD). METHODS: Male AO recipient rats were intrathymically inoculated with 2.5x10(7) PVG splenocytes immediately followed by heterotopic transplantation of a PVG cardiac allograft (day 0). Immunosuppression consisted of 1 ml of antilymphocyte serum i.p. (day 0) and cyclosporine i.m. (15 mg/kg body weight) on days 1, 2, and 3 posttransplantation. Histological analysis, mixed lymphocyte reactions, and intragraft cytokine mRNA expression were performed at several time points after engraftment. RESULTS: Histological analysis revealed that GAD was already present 14 days after transplantation. At 200 days, virtually all vessels were affected and over 80% of the vessels showed severe intimal lesions. Infiltrate analysis displayed massive parenchymatous infiltrates (CD8+ cells and ED1+ macrophages) 2 weeks after transplantation. At later time points, infiltrates became epicardial and/or blood vessel associated and mainly consisted of CD4+, CD8+, and B cells. Mixed lymphocyte reactions showed nonspecifically decreased responses at 60 days but complete restoration of these responses at later time points (120 to 280 days). Intragraft cytokine mRNA expression showed decreased interleukin-2/interferon-gamma and sustained interleukin-10 expression 2 weeks after transplantation. Transforming growth factor-beta mRNA expression was increased >200 days after transplantation. CONCLUSIONS: Intrathymic immune modulation does not abolish alloreactivity, and despite induction of long-lasting graft survival, this procedure does not prevent and may even facilitate the development of GAD.
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Trasplante de Células , Enfermedad de la Arteria Coronaria/etiología , Rechazo de Injerto/prevención & control , Trasplante de Corazón/efectos adversos , Trasplante de Corazón/inmunología , Bazo/citología , Bazo/inmunología , Timo/inmunología , Enfermedad Aguda , Animales , Enfermedad Crónica , Citocinas/metabolismo , Femenino , Supervivencia de Injerto , Inyecciones , Prueba de Cultivo Mixto de Linfocitos , Masculino , Miocardio/metabolismo , Miocardio/patología , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Tiempo , Trasplante HeterotópicoAsunto(s)
Subgrupos de Linfocitos B/inmunología , Reordenamiento Génico de Cadena Pesada de Linfocito B/genética , Genes de Inmunoglobulinas , Cadenas Pesadas de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/genética , Ganglios Linfáticos/citología , Animales , Antígenos Bacterianos/inmunología , Células de la Médula Ósea/inmunología , Diferenciación Celular , Supresión Clonal , Hematopoyesis , Humanos , Sistema Inmunológico/crecimiento & desarrollo , Memoria Inmunológica , Ligandos , Lipopolisacáridos/inmunología , Ratones , Quimera por Radiación , Ratas , Especificidad de la Especie , Bazo/citología , Bazo/inmunologíaRESUMEN
BACKGROUND: Coronary artery disease is today's most important post-heart transplantation problem after the first perioperative year. Histologically, coronary artery disease is characterized by transplant arteriosclerosis. The current view on this vasculopathy is that vascular smooth muscle (VSM) cells from the media of affected arteries proliferate and migrate into the sub-endothelial space (intima) in response to signals from inflammatory cells and damaged graft endothelium. According to this model, the intimal VSM cells in transplant arteriosclerotic lesions should originate from donor tissue. Using recipient-specific polymerase chain reaction (PCR) analysis of microdissected, single, neointimal VSM nuclei, we recently showed that after allogeneic aorta transplantation the neointimal VSM cells are of recipient and not of donor origin. In this study, we analyzed whether VSM-cell replacement with recipient-derived cells also takes place after allogeneic heart transplantation. METHODS: Cardiac allografts, when transplanted from female donors to male immune-modulated recipient rats, eventually developed transplant arteriosclerosis. We microdissected alpha-actin positive neointimal VSM cells from tissue sections and determined the origin (donor or recipient) using recipient-specific (male), single-cell, PCR analysis. RESULTS: In total, we analyzed 35 VSM-cell nuclei from 3 allografts, and PCR analysis revealed that 30/35 (86%) of the samples displayed the male-specific 128 base pair DNA fragment. These results indicate that after allogeneic cardiac transplantation, at least 86% of VSM cells in transplant arteriosclerotic lesions are of recipient origin. CONCLUSIONS: In contrast to current thought, the neointimal VSM cells in cardiac allografts that show transplant arteriosclerosis are of recipient and not of donor origin.
Asunto(s)
Enfermedad de la Arteria Coronaria/patología , Trasplante de Corazón/patología , Músculo Liso Vascular/patología , Túnica Íntima/patología , Actinas/análisis , Animales , Aorta/trasplante , Emparejamiento Base , División Celular , Linaje de la Célula , Movimiento Celular , Núcleo Celular/ultraestructura , Endotelio Vascular/patología , Femenino , Masculino , Microcirugia , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , Estadísticas no Paramétricas , Donantes de Tejidos , Cromosoma Y/genéticaRESUMEN
The present study was performed to analyze whether marginal zone B (MZ-B) cells in nondeliberately immunized adult rats are selected on basis of the specificity of their B cell receptor, and to determine to what extent memory B cells contribute to the MZ-B cell subset. To this end, the Ig PC7183 V(H) gene repertoire was studied among V(H)DJ(H)-mu transcripts expressed in four sequential stages of B cell development, of two individual untreated adult rats. B cell subsets, i.e., pro/pre-B cells and newly formed B (NF-B) cells from bone marrow, and recirculating follicular B cells and MZ-B cells from spleen were sorted by flow cytometry. In addition, from one these rats, cells were microdissected from follicular and MZ areas of the spleen and productive PC7183 V(H) gene rearrangements were analyzed for the presence of somatic mutations. Sequence analysis reveals that most MZ-B cells in the adult rat, either defined by flow cytometry or by their anatomical location in the spleen, express germline encoded V(H) genes (naive MZ-B cells) and a minor fraction (about 20%) of the MZ-B cells carry somatic mutations (memory MZ-B cells). In addition, we show that naive MZ-B cells are a selected population of cells, both based on PC7183 V(H) gene repertoire and on the length of the Ig heavy (H) chain complementarity-determining region 3 (H-CDR3) region, i.e., PC7183 V(H)DJ(H)-mu transcripts of MZ-B cells carry significantly shorter H-CDR3 regions than other B cell subsets.
Asunto(s)
Subgrupos de Linfocitos B/inmunología , Subgrupos de Linfocitos B/metabolismo , Células de la Médula Ósea , Cadenas Pesadas de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/genética , Bazo/citología , Secuencia de Aminoácidos , Animales , Diversidad de Anticuerpos/genética , Secuencia de Bases , Células de la Médula Ósea/inmunología , Células de la Médula Ósea/metabolismo , Separación Celular , Regiones Determinantes de Complementariedad/biosíntesis , Regiones Determinantes de Complementariedad/genética , Análisis Mutacional de ADN , Disección , Citometría de Flujo , Regulación de la Expresión Génica/inmunología , Reordenamiento Génico de Cadena Pesada de Linfocito B , Mutación de Línea Germinal , Cadenas Pesadas de Inmunoglobulina/biosíntesis , Cadenas J de Inmunoglobulina/biosíntesis , Cadenas J de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/biosíntesis , Cadenas mu de Inmunoglobulina/biosíntesis , Cadenas mu de Inmunoglobulina/genética , Ligandos , Masculino , Micromanipulación , Datos de Secuencia Molecular , Familia de Multigenes/inmunología , Ratas , Ratas Endogámicas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Bazo/inmunología , Bazo/metabolismo , Transcripción Genética/inmunologíaRESUMEN
The salivary gland is the preferred organ for cytomegalovirus (CMV) replication and viral persistence. In order to identify the nature of infected cells and to study viral replication in more detail, several experiments were conducted. Using the rat CMV (RCMV) model, acutely infected young adult rats (6 weeks of age) and new-born rats (3 days of age) were infected, and submandibular, parotid and sublingual glands were harvested at different time points after infection. For identification of the nature of infected cells, immunohistochemistry, in situ hybridisation and electron microscopic techniques were used. In young adult animals, the submandibular gland was the preferred organ for RCMV replication. The parotid and sublingual glands contained fewer viruses than the submandibular gland. In contrast, in new-born rats, the main site of RCMV replication was the sublingual gland, while the submandibular and parotid glands contained low amounts of virus. No virus could be detected in the parotid glands. In all glands of RCMV-infected animals, the infection was exclusively confined to striated duct cells. Infection resulted in a cellular inflammatory response which was mostly located in the interlobular duct region, whereas only few inflammatory cells were found in the neighbourhood of infected striated duct cells. This phenomenon may contribute to the long persistence of the virus in this organ.
Asunto(s)
Citomegalovirus/aislamiento & purificación , Glándulas Salivales/virología , Replicación Viral , Animales , Citomegalovirus/fisiología , Inmunohistoquímica , Masculino , Microscopía Electrónica , Ratas , Ratas Endogámicas Lew , Glándulas Salivales/ultraestructuraRESUMEN
Using light and transmission electron microscopy, we observed novel structures in the rabbit vitreous body. They were found in 18 out of 27 eyes from rabbits 0.5-36 months of age. These structures are scattered throughout the entire vitreous matrix. By light and transmission electron microscopy, they appear to be made up of the same structural components. Based upon their morphological appearance, they can be subdivided into two groups which we provisionally named 'intravitreal structure type 1 and 2' or 'IVS-1' and 'IVS-2'. IVS-1 has a highly variable morphology (e.g. star-shaped, round, oval), whereas IVS-2 is tubular. The dimensions of IVS-1 vary in relation to the mesh diameters of the collagen matrix, while those of IVS-2 do not. In adult rabbit eyes, we observed transitions between IVS-1 and intravitreal ghost vessels (acellular remnants of blood vessels), and between IVS-1 and IVS-2. In very young rabbits (14 days) we observed intravitreal ghost vessels consisting of tightly-packed IVS-1. Therefore, we concluded that IVS-1 and 2 are related structures presumably of vascular origin. It appears that they represent fragmented and non-fragmented acellular remnants of hyaloid blood vessels. The presence of vascular remnants throughout the entire vitreous matrix of adult rabbit eyes is in conflict with existing theories on the embryonic development of the vitreous body, which describe a strict spatial separation between the primary (vascular) and secondary (avascular) vitreous. However, it strongly supports an alternative theory that explains the formation of the secondary vitreous by a process of continuous remodelling of the primary vitreous.
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Vasos Sanguíneos/ultraestructura , Cuerpo Vítreo/irrigación sanguínea , Animales , Vasos Sanguíneos/crecimiento & desarrollo , Colágeno/ultraestructura , Microscopía Electrónica , Adhesión en Plástico , Conejos , Coloración y Etiquetado , Cuerpo Vítreo/crecimiento & desarrolloRESUMEN
The purpose of this study was to evaluate the composition of ghost vessels and the newly identified intravitreal structures type 1 and 2 (IVS-1 and 2) observed in the rabbit vitreous body. Rabbit eyes (n = 10, 0.5- approximately 36 months of age) were fixed and embedded in plastic. Post-embedding immuno transmission electron microscopy and enzyme digestion methods specifically directed at vascular extracellular matrix components (collagen IV, elastin and hyaluronan) were used in order to confirm the postulated vascular origin of IVS-1 and 2. In addition, markers of vitreous extracellular matrix components (collagen II, hyaluronan) were used. The postulated vascular nature of ghost vessels and IVS-1 was confirmed by a positive labelling with anti-collagen IV, whereas the demonstration of elastin (by anti-elastin antibodies and elastase digestion) in IVS-1 and 2 confirms their arterial origin. These vascular remnants were also labelled with a hyaluronan marker and with anti-collagen II. The presence of remnants of the hyaloid artery system throughout the vitreous matrix is in conflict with a strict spatial separation between the primary and secondary vitreous during embryonic development as proposed in the literature. It strongly supports an alternative theory which suggests an interactive remodelling of this matrix. The presence of hyaluronan in remnants of the hyaloid system is inconclusive, since hyaluronan is a component both of the adult vitreous matrix and of the vascular extracellular matrix. The presence of collagen II in vascular structures is highly interesting, since it supports another challenging theory, which suggests that lamellae develop alongside tracts formerly occupied by the larger hyaloid vessels.
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Vasos Sanguíneos/metabolismo , Cuerpo Vítreo/irrigación sanguínea , Animales , Biomarcadores , Vasos Sanguíneos/crecimiento & desarrollo , Pruebas Enzimáticas Clínicas , Matriz Extracelular/metabolismo , Ácido Hialurónico/metabolismo , Inmunohistoquímica , Microscopía Electrónica , Adhesión en Plástico , Conejos , Coloración y Etiquetado , Cuerpo Vítreo/crecimiento & desarrolloRESUMEN
In chronic allografts, graft vessels eventually develop so-called "transplant vascular sclerosis" or "intimal hyperplasia". A major question is whether the cells in the neointima are donor or recipient derived. The process of transplant vascular sclerosis closely resembles the remodeling of the vascular wall as seen when synthetic biodegradable small caliber vascular grafts are implanted. In this model, the cells in the newly developing neointima as well as neomedia are, by definition, recipient derived. By using cardiac allografts as well as aortic allografts exchanged between a female donor and a male recipient (rats), the origin of the neointimal vascular smooth muscle cells could be traced by looking for the Y-chromosome in isolated (alpha-actin positive) intimal cells using PCR. In both models these intimal cells were found to be of recipient-origin. It is proposed, that, basically, this remodeling process is part of a normal healing process. Whereas in biodegradable grafts this "healing process" appears to be self limiting, in allografts the process goes on beyond the needs of functional repair, eventually, in some cases, leading to total vascular occlusion. Future therapeutic protocols might try and aim at controlling this essentially normal repair process.
Asunto(s)
Implantes Absorbibles/efectos adversos , Implantación de Prótesis Vascular/efectos adversos , Rechazo de Injerto/inmunología , Animales , Aorta/cirugía , Vasos Coronarios/cirugía , Femenino , Humanos , Masculino , Trasplante HomólogoRESUMEN
Previous studies involving the function and development of peripheral T cells have proposed that, in the rat, CD4(+)CD45RC(+)RT6(-) and CD4(+)CD45RC(-)RT6(+) T-cell subsets may represent Th1 and Th2 cells, respectively. Here we tested this hypothesis directly by analyzing frequencies of IFN-gamma- and IL-4-producing cells in these two subpopulations using ELISPOT assays. We found that the CD4(+)CD45RC(-)RT6(+) subset showed higher numbers of IL-4-producing cells than the CD4(+)CD45RC(+)RT6(-) subset and, though less pronounced, that the latter demonstrated higher numbers of IFN-gamma producers. Therefore, we conclude that our results provide evidence for the existence of phenotypically defined Th1 and Th2 cells in the rat. This is supported by the finding that the ratios of IFN-gamma/IL-4 and CD45RC/RT6 correlated positively among various rat strains. Finally, rat strains susceptible to induction of a Th1-mediated autoimmune disease showed the highest CD45RC/RT6 ratio, whereas the reverse was true for strains susceptible to a Th2-mediated autoimmune disease.
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ADP Ribosa Transferasas/biosíntesis , Citocinas/biosíntesis , Antígenos Comunes de Leucocito/biosíntesis , Glicoproteínas de Membrana/biosíntesis , Células TH1/metabolismo , Células Th2/metabolismo , Animales , Antígenos de Diferenciación de Linfocitos T/biosíntesis , Antígenos CD4/biosíntesis , Citocinas/sangre , Interferón gamma/biosíntesis , Interferón gamma/sangre , Interleucina-4/biosíntesis , Interleucina-4/sangre , Masculino , Ratas , Ratas Endogámicas BN , Ratas Endogámicas BUF , Ratas Endogámicas F344 , Ratas Endogámicas Lew , Ratas Sprague-Dawley , Especificidad de la Especie , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismoRESUMEN
The present study shows the lymphatic distribution of the negatively charged anti-HIV-1 agents succinylated or aconytilated human serum albumins (HSAs) in rats. Quantitation of blood and lymphatic concentrations of these proteins was performed through fluorescence detection of the fluorescein isothiocyanate (FITC)-labeled proteins. At several time points after i.v. injection, samples were taken from the cannulated thoracic duct and the carotid artery. Distribution of the negatively charged albumins (NCAs) to lymph was much more rapid than that of albumin itself and was dependent on the total net negative charge added to the protein: the half-life times of lymphatic equilibration were 15, 30, and 120 min for FITC-labeled aconytilated HSA, FITC-labeled succinylated HSA, and FITC-labeled HSA, respectively. Lymph to blood concentration ratios of the studied compounds obtained at steady state approached unity. In addition, the fluorescence in both body fluids was shown to represent unchanged labeled proteins. It was therefore inferred that the NCAs efficiently passed the endothelial barrier from blood to the interstitial compartment. Subsequently, we studied whether a specialized process was involved in the endothelial passage of the NCAs to the lymph. The following observations supported such a mechanism: a) preinjection of the scavenger receptor blockers polyinosinic- and formaldehyde-treated HSA reduced the transport from blood to the lymphatic compartment of FITC-labeled aconytilated HSA by more than 90%; b) the rate of lymphatic distribution was largely reduced when the body temperature of the rat was lowered to 28 degrees; and c) pre-administration of chloroquine resulted in a significant reduction in the lymphatic distribution of the NCAs. These data collectively indicate that a scavenger receptor-mediated process is involved in the transendothelial transport of NCAs. In situ localization in lymph nodes of the rat showed that FITC-labeled aconytilated and succinylated HSA are mainly present in the germinal center and parafollicular zones. The efficient distribution of these anionized proteins to the lymphatic system is of particular interest for HIV therapy, taking into account that replication of HIV mainly takes place in the lymphoid system. The observation that macromolecules, through charge modification, can extravasate through a receptor-mediated transcytotic process is potentially of major importance for the delivery of drugs with macromolecular carriers to cells not directly in contact with the blood.
Asunto(s)
Albúminas/farmacocinética , Fármacos Anti-VIH/farmacocinética , Sistema Linfático/metabolismo , Albúminas/química , Albúminas/farmacología , Animales , Fármacos Anti-VIH/química , Transporte Biológico , Células Cultivadas , Sistemas de Liberación de Medicamentos , Electroquímica , Fluoresceína/metabolismo , VIH/efectos de los fármacos , Humanos , Sistema Linfático/virología , Masculino , Ratas , Ratas Wistar , Distribución TisularRESUMEN
Even though the rabbit is a frequently used animal model for studies on vitreous function and pathobiology, data on the structural organization of the rabbit vitreous are scarce. The aim of the present study is to give a detailed description of rabbit vitreous structure in order to provide a basis for studies on changes in vitreous organization induced by pathophysiological processes. We studied the vitreous body of adult rabbit eyes by complementary anatomical evaluation methods, by light microscopy and by transmission electron microscopy. Regional and local variations in vitreous matrix organization were observed. Regionally, a cortex, an intermediate area, and a centre were distinguished. In addition, specific structures were locally observed. Lamellae run through the intermediate area in a funnel-like pattern, converging upon the asymmetrically positioned optic disc. A central channel (Cloquet's channel) was found in all eyes. We demonstrated a novel structure, attached to Cloquet's channel and to the medullary rays. Because of its wing-shaped sheet-like morphology, we named it the 'alae canalis Cloqueti'.
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Conejos/anatomía & histología , Cuerpo Vítreo/anatomía & histología , Animales , Colágeno/análisis , Disección , Tinta , Microscopía Electrónica , Cuerpo Vítreo/química , Cuerpo Vítreo/ultraestructuraRESUMEN
The marginal zone is a unique compartment that is only found in the spleen. Rat marginal zone B cells (MZ-B) can be distinguished from other B cells, e.g. recirculating follicular B cells (RF-B), by several phenotypic characteristics. Typically MZ-B cells are surface (s)IgMhi, sIgDlo and CD45R(B220)lo, whereas RF-B cells are sIgMlo, sIgDhi and CD45Rhi. In addition, MZ-B cells stain strongly with HIS57, a newly developed monoclonal antibody. The developmental pathway and origin of MZ-B cells are not exactly known. However, previous studies indicate that recirculating (i. e. thoracic duct) B cells can give rise to MZ-B cells. Here the origin of (naive) MZ-B cells was studied using adriamycin (doxorubicin)-induced B cell depletion. Using three-color flow cytometry and immunohistology we show that 2 days after a single i.v. injection of the anti-tumor drug adriamycin only RF-B cells can be detected, while all other B cell subpopulations are depleted, including all bone marrow precursor B cells. By studying the sequential reappearance of various B cell subsets and their precursors after adriamycin administration we show that MZ-B cells and the splenic marginal zone can be detected at a time point at which newly generated B cells (immature B cells) are not yet present. Given the observation that only RF-B cells were present at this time, we conclude that RF-B cells are the immediate MZ-B precursor cells.
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Linfocitos B/citología , Animales , Subgrupos de Linfocitos B/inmunología , Linfocitos B/clasificación , Linfocitos B/inmunología , Diferenciación Celular , Doxorrubicina , Citometría de Flujo , Linfocitos , Masculino , Ratones , FN-kappa B/metabolismo , Ratas , Bazo/citologíaRESUMEN
OBJECTIVE: To validate proliferating cell nuclear antigen (PCNA) expression and flow cytometry as proliferation markers in regenerating rat liver containing metastases. STUDY DESIGN: Rats containing colorectal liver metastases were killed at various days after 70% partial hepatectomy or a sham operation. [3H]thymidine and 5-bromo-2'deoxyuridine (BrdU) incorporation, PCNA expression and flow cytometry were used to evaluate liver cell proliferation. RESULTS: The assessment of proliferating liver cells by PCNA expression and BrdU incorporation was more reliable than autoradiography. PCNA expression correlated well with BrdU incorporation (r = .68, P = .003) and autoradiography (r = .57, P = .02) in regenerating liver. BrdU incorporation and PCNA expression were higher in hepatectomized rats as compared to sham-operation rats at days 1-4 after hepatectomy. Flow cytometry of propidium-stained nuclei from livers of hepatectomized rats showed a higher proportion of S-phase nuclei as compared to S-phase nuclei in control rats. The correlation coefficients of the number of S-phase nuclei, BrdU-positive nuclei and PCNA-positive nuclei were .39 (P < .02) and .56 (P < .0005), respectively. CONCLUSION: Flow cytometry and PCNA expression are simple and reliable methods of studying proliferation in metastases containing rat liver after partial hepatectomy.