RESUMO
BACKGROUND: Phospholipase A2 (PLA2) is associated with a variety of inflammatory processes related to polymorphonuclear neutrophil (PMN)-endothelial cell interactions. However, the cellular and molecular mechanisms underlying the interactions and the causative isoform(s) of PLA2 remain elusive. In addition, we recently showed that calcium-independent PLA2γ (iPLA2γ), but not cytosolic PLA2 (cPLA2), is responsible for the cytotoxic functions of human PMN including respiratory bursts, degranulation, and chemotaxis. We therefore hypothesized that iPLA2γ is a prerequisite for the PMN recruitment cascade into the site of inflammation. The aim of this study was to elucidate the roles of the three major phospholipases A2, iPLA2, cPLA2 and secretory PLA2, in leukocyte rolling and adherence and in the surface expression of ß2-integrins in vivo and in vitro in response to well-defined stimuli. METHODS: Male Wistar rats were pretreated with PLA2 inhibitors selective for iPLA2ß, iPLA2γ, cPLA2, or secretory PLA2. Leukocyte rolling/adherence in the mesenteric venules superfused with platelet-activating factor (PAF) were quantified by intravital microscopy. Furthermore, isolated human PMNs or whole blood were incubated with each PLA2 inhibitor and then activated with formyl-methionyl-leucyl-phenylalanine (fMLP) or PAF. PMN adherence was assessed by counting cells bound to purified fibrinogen, and the surface expression of lymphocyte function-associated antigen 1 and macrophage antigen 1 (Mac-1) was measured by flow cytometry. RESULTS: The iPLA2γ-specific inhibitor almost completely inhibited the fMLP/PAF-induced leukocyte adherence in vivo and in vitro and also decreased the fMLP/PAF-stimulated surface expression of Mac-1 by 60% and 95%, respectively. In contrast, the other inhibitors did not affect these cellular functions. CONCLUSION: iPLA2γ seems to be involved in leukocyte/PMN adherence in vivo and in vitro as well as in the up-regulation of Mac-1 in vitro in response to PAF/fMLP. This enzyme is therefore likely to be a major regulator in the PMN recruitment cascade.
Assuntos
Comunicação Celular , Movimento Celular/fisiologia , Quimiotaxia de Leucócito/fisiologia , Mesentério/ultraestrutura , Fosfolipases A2 Independentes de Cálcio/metabolismo , Animais , Células Cultivadas , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Humanos , Masculino , Neutrófilos/metabolismo , Proteínas de Transferência de Fosfolipídeos/metabolismo , Distribuição Aleatória , Ratos , Ratos Wistar , Papel (figurativo) , Sensibilidade e EspecificidadeRESUMO
The role of lymphatic vessels is to transport fluid, soluble molecules, and immune cells to the draining lymph nodes. Here, we analyze how the aging process affects the functionality of the lymphatic collectors and the dynamics of lymph flow. Ultrastructural, biochemical, and proteomic analysis indicates a loss of matrix proteins, and smooth muscle cells in aged collectors resulting in a decrease in contraction frequency, systolic lymph flow velocity, and pumping activity, as measured in vivo in lymphatic collectors. Functionally, this impairment also translated into a reduced ability for in vivo bacterial transport as determined by time-lapse microscopy. Ultrastructural and proteomic analysis also indicates a decrease in the thickness of the endothelial cell glycocalyx and loss of gap junction proteins in aged lymph collectors. Redox proteomic analysis mapped an aging-related increase in the glycation and carboxylation of lymphatic's endothelial cell and matrix proteins. Functionally, these modifications translate into apparent hyperpermeability of the lymphatics with pathogen escaping from the collectors into the surrounding tissue and a decreased ability to control tissue fluid homeostasis. Altogether, our data provide a mechanistic analysis of how the anatomical and biochemical changes, occurring in aged lymphatic vessels, compromise lymph flow, tissue fluid homeostasis, and pathogen transport.
Assuntos
Envelhecimento/metabolismo , Linfonodos/metabolismo , Linfa/metabolismo , Vasos Linfáticos/química , Proteoma/metabolismo , Sequência de Aminoácidos , Animais , Conexinas/genética , Conexinas/metabolismo , Células Endoteliais/metabolismo , Células Endoteliais/ultraestrutura , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Junções Comunicantes/metabolismo , Junções Comunicantes/ultraestrutura , Glicocálix/química , Glicocálix/metabolismo , Glicosilação , Infecções por Bactérias Gram-Positivas/metabolismo , Infecções por Bactérias Gram-Positivas/microbiologia , Homeostase , Linfonodos/microbiologia , Linfonodos/ultraestrutura , Vasos Linfáticos/metabolismo , Vasos Linfáticos/microbiologia , Vasos Linfáticos/ultraestrutura , Masculino , Mesentério/metabolismo , Mesentério/microbiologia , Mesentério/ultraestrutura , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Mycobacterium smegmatis/fisiologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/ultraestrutura , Proteoma/genética , Ratos , Ratos Endogâmicos F344 , Staphylococcus aureus/fisiologia , Imagem com Lapso de TempoRESUMO
Using the quantitative methods, the remodeling of the cytoarchitectonics of the morpho-functional zones in the grouped lymphoid nodules (GLN) or Peyer's patches and in the mesenteric lymph nodes (MLN) were studied in 30 rats after 30-day-long exposure to hypokinesia and during the period of rehabilitation (30 days after hypokinesia discontinuation). It was found that following the hypokinesia the germinal centers in lymphoid nodules in GLN retained the lymphocytopoiesis, while in the internodular zone the proportion of immature cells was increased and plasma cells appeared. In the similar structural zones of MLN, the complete suppression of lymphocytopoiesis and T-cell maturation was noted. During the rehabilitation period, the cytoarchitectonic indexes recovery was more pronounced in GLN than in MLN. However, the quantitative parameters of their cellular composition did not reach the values found in the group of intact of animals.
Assuntos
Hipocinesia/patologia , Sistema Imunitário , Linfonodos/ultraestrutura , Mesentério/ultraestrutura , Animais , Linfonodos/imunologia , Linfonodos/patologia , Linfopoese , Mesentério/imunologia , Mesentério/patologia , Nódulos Linfáticos Agregados/imunologia , Nódulos Linfáticos Agregados/patologia , Nódulos Linfáticos Agregados/ultraestrutura , Ratos , Baço/imunologia , Linfócitos T/imunologia , Linfócitos T/ultraestruturaRESUMO
To supplement and correct the morphological features of lymphatic labyrinths (LLs) in rat mesenteric lymph node, the distribution, morphology and origin of LLs, and cellular elements in LLs, particularly the organization and integrity of the wall of LLs were examined by silver impregnation, transmission electron microscopy (TEM), scanning electron microscopy (SEM), and immunohistochemistry. LLs consisted of labyrinthine tubules and ran through not only the periphery of the deep cortical unit (DCU) but interfollicular cortex. LLs originated at the edge of the center of the DCU and of the follicle. At the site of their origin, the fibers in the wall of LL were continuous with the fibers located in the follicle and the center of DCU. The wall of LLs was a trilaminar membrane: a layer of flattened lymphatic endothelium; a layer of fibroblastic reticular cells; and amorphous substance and collagen fibers sandwiched between the above two layers. Under SEM and TEM, the whole amoeboid lymphocytes were moving through the pores in the wall of LL, which showed that lymphocytes end their journey through the paracortical cord by migrating into LLs. Immunohistochemical lymphatic vessel endothelial hyaluronan receptor-1 expression was present in cells lining the LLs and intraluminal stellate cells, which may belong to the "sinus endothelial/virgultar cells." LLs are specific channels that are different from lymphatic sinuses. LL may be regarded as a special part of lymphatic vascular system in lymph nodes. We confirm that LLs are important transport pathway of lymphocytes in lymph nodes. The structural framework of LLs facilitates the migration of lymphocytes.
Assuntos
Orelha Interna/anatomia & histologia , Linfonodos/anatomia & histologia , Sistema Linfático/anatomia & histologia , Mesentério/anatomia & histologia , Animais , Orelha Interna/metabolismo , Orelha Interna/ultraestrutura , Técnicas Imunoenzimáticas , Linfonodos/metabolismo , Linfonodos/ultraestrutura , Sistema Linfático/metabolismo , Sistema Linfático/ultraestrutura , Masculino , Mesentério/metabolismo , Mesentério/ultraestrutura , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Ratos , Ratos WistarRESUMO
The aim of this study was to investigate the spatiotemporal development of autonomic nerve fibers and primordial germ cells (PGCs) along their migratory route from the dorsal mesentery to the gonadal ridges in human embryos using immunohistochemical markers and electron microscopy. Autonomic nerve fibers in the dorsal mesentery, the pre-aortic and para-aortic plexuses and in the gonadal ridge were stained for beta III tubulin, neuron specific enolase and the glia fibrillary acidic protein. Electron microscopy demonstrated the presence of neurofilaments and neurotubules in these nerve fibers and their intimate contact with PGCs. PGCs expressed GAGE, MAGE-A4, OCT4 and c-Kit. Serial paraffin sections showed that most PGCs were located inside bundles of autonomic nerve fibers with the majority adjacent to the most peripheral fibers (close to Schwann cells). We also show that both nerve fibers and PGCs arrive at the gonadal ridge between 29 and 33 days pc. In conclusion, our data suggest that PGCs in human embryos preferentially migrate along autonomic nerve fibers from the dorsal mesentery to the developing gonad where they are delivered via a fine nerve plexus.
Assuntos
Sistema Nervoso Autônomo/embriologia , Movimento Celular , Células Germinativas/fisiologia , Gônadas/embriologia , Mesentério/embriologia , Fibras Nervosas/fisiologia , Células de Schwann/fisiologia , Sistema Nervoso Autônomo/química , Sistema Nervoso Autônomo/ultraestrutura , Biomarcadores/análise , Feminino , Células Germinativas/química , Células Germinativas/ultraestrutura , Idade Gestacional , Gônadas/química , Gônadas/ultraestrutura , Humanos , Imuno-Histoquímica , Mesentério/química , Mesentério/ultraestrutura , Microscopia Eletrônica , Fibras Nervosas/química , Fibras Nervosas/ultraestrutura , Ovário/embriologia , Células de Schwann/química , Células de Schwann/ultraestruturaRESUMO
Peritoneal diseases can be seen in the different imaging modalities either as fluid collections or solid tumors along the ligaments, mesenteries, and spaces of the peritoneal cavity. The broad spectrum of different abnormalities includes inflammatory, infectious, traumatic, and neoplastic diseases. In this article, a large variety of peritoneal abnormalities such as ascites, peritonitis, intraperitoneal hemorrhage, and both primary and secondary peritoneal tumors are discussed. The different imaging modalities, characteristic radiological features, and typical pathways of anatomic spread are explained.
Assuntos
Imageamento por Ressonância Magnética/métodos , Mesentério , Doenças Peritoneais/diagnóstico , Tomografia Computadorizada por Raios X/métodos , Ultrassonografia/métodos , Humanos , Mesentério/diagnóstico por imagem , Mesentério/patologia , Mesentério/ultraestruturaRESUMO
We have investigated the structural basis by which the counterclockwise direction of the amniote gut is established. The chirality of midgut looping is determined by left-right asymmetries in the cellular architecture of the dorsal mesentery, the structure that connects the primitive gut tube to the body wall. The mesenchymal cells of the dorsal mesentery are more condensed on the left side than on the right and, additionally, the overlying epithelium on the left side exhibits a columnar morphology, in contrast to a cuboidal morphology on the right. These properties are instructed by a set of transcription factors: Pitx2 and Isl1 specifically expressed on the left side, and Tbx18 expressed on the right, regulated downstream of the secreted protein Nodal which is present exclusively on the left side. The resultant differences in cellular organization cause the mesentery to assume a trapezoidal shape, tilting the primitive gut tube leftward.
Assuntos
Padronização Corporal/fisiologia , Trato Gastrointestinal/embriologia , Mesentério/embriologia , Rotação , Animais , Forma Celular , Embrião de Galinha , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Células Epiteliais/citologia , Células Epiteliais/ultraestrutura , Proteínas de Homeodomínio/metabolismo , Proteínas com Homeodomínio LIM , Mesentério/citologia , Mesentério/ultraestrutura , Camundongos , Modelos Biológicos , Proteína Nodal , Proteínas com Domínio T/metabolismo , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Proteína Homeobox PITX2RESUMO
A phenomenon that has defied explanation for two decades is the time scale for transient reabsorption in the classic experiments of Michel and Phillips on individually perfused frog mesentery microvessels. One finds that transient reabsorption lasts <2 min before a new steady state of low filtration is established when the lumen pressure is abruptly dropped from a high to a low value. Our experiments in frog and rat venular microvessels under a variety of conditions revealed the same time trend for new steady states to be established as in Michel and Phillips' experiments. In contrast, one theoretically predicts herein that the time required for the tissue albumin concentration to increase to values for a new steady state to be achieved through reabsorption is in the order of several hours. In this paper we propose a new hypothesis and theoretical model for this rapid regulation, namely that pericytes covering the interendothelial cleft exits create small trapped microdomains outside the cleft exits which regulate this transient behavior. Our electron microscopy studies on rat mesenteric venular microvessels reveal an average pericyte coverage of approximately 85%. The theoretical model based on this ultrastructural study predicts an equilibration time on the order of 1 min when the lumen pressure is abruptly lowered. The basic concept of a trapped microdomain can also be extended to microdomains in the interstitial space surrounding skeletal muscle capillaries.
Assuntos
Endotélio Vascular/metabolismo , Mesentério/metabolismo , Pericitos/metabolismo , Albumina Sérica/química , Albumina Sérica/metabolismo , Vênulas/metabolismo , Animais , Aorta , Endotélio Vascular/fisiologia , Endotélio Vascular/ultraestrutura , Masculino , Mesentério/irrigação sanguínea , Mesentério/ultraestrutura , Microcirculação/metabolismo , Microcirculação/ultraestrutura , Pericitos/ultraestrutura , Estrutura Terciária de Proteína/fisiologia , Transporte Proteico/fisiologia , Rana pipiens , Rana temporaria , Ratos , Ratos Sprague-Dawley , Albumina Sérica/fisiologia , Vênulas/fisiologia , Vênulas/ultraestruturaRESUMO
Interstitial Cajal-like Cells (ICLC) were recently recognized in a plethora of non-digestive organs. Here, we describe a cell type of rat mesentery sharing ultrastructural and immunohistochemical features with ICLC. Mesenteric ICLC were demonstrated by transmission electron microscopy (TEM) and further tested by light microscope immunohistochemistry. The cell described here fulfils the TEM diagnostic criteria accepted for ICLC: location in the connective interstitium; close vicinity to nerves, capillaries and other interstitial cells; characteristic long, moniliform cell processes; specialized cell-to-cell junctions; caveolae; mitochondria at 5-10% of cytoplasmic volume; rough endoplasmic reticulum at about 1-2%; intermediate and thin filaments, microtubules; undetectable thick filaments. The processes of this mesenteric ICLC were particularly long, with a mean length of 24.91 microm (10.27-50.83 micorm), and a convolution index of 2.32 (1.37-3.63) was calculated in order to measure their potential length. Mean distances versus main target cells of ICLC-nerve bundles, vessels, adipocytes and macrophages-were 110.69, 115.80, 205.07 and 34.65 nm, respectively. We also tested the expression of CD117/c-kit, CD34, vimentin, alpha-smooth muscle actin, nestin, NK-1, tryptase and chymase and the antigenic profile of the mesenteric ICLC was comparable if not identical with that recently observed in ICLC from other extra-digestive tissues. Due to the peculiar aspect of the mesenteric ICLC processes it can be hypothesized that these cells form a three-dimensional network within the mesentery that is at the same time resistant and deformable following stretches consequent to intestine movements, mainly avoiding blood vessels closure or controlling blood vessels rheology. It remains, however, to be established if and how such cells are connected with the archetypal enteric ICC.
Assuntos
Células do Tecido Conjuntivo/ultraestrutura , Junções Intercelulares/ultraestrutura , Mesentério/ultraestrutura , Animais , Antígenos CD34/metabolismo , Biomarcadores/metabolismo , Células Cultivadas , Células do Tecido Conjuntivo/metabolismo , Retículo Endoplasmático/metabolismo , Técnicas Imunoenzimáticas , Masculino , Mesentério/metabolismo , Microscopia Eletrônica de Transmissão , Microtúbulos/metabolismo , Proteínas Proto-Oncogênicas c-kit/metabolismo , Ratos , Ratos Wistar , Vimentina/metabolismoRESUMO
BACKGROUND: Rat mesentery has been widely used to study microvascular functions. The goal of this work is to extend this animal model to monitor blood and lymph microvessel function during lymphedema. METHODS AND RESULTS: Lymphedema is created by microsurgical removal of regional lymph nodes (lymphadenectomy) or ligation of the collecting vein. Water volume in mesenteric tissue, microvessel diameters, phasic contraction, valve function, lymph flow velocity, and cell migration were analyzed during lymphedema development. Dynamic observation of water amount after lymphadenectomy revealed increasing edema from 30 min to 1 week; greatest degree of edema at one week, and gradual decrease in edema from 1 to 11 weeks. These effects were accompanied by acute constriction of lymph vessels and slowing of lymph flow velocity, switching to dilation and appearance of new blood capillaries at week 1, progressing to dilation and degenerative changes of the microlymphatic wall at week 4, and, finally, leading to lymphatic fibrosis and lymphangiogenesis at week 11. Acute venous insufficiency (30 min after vein ligation) led to significant edema, decreasing blood flow velocity to stasis, and output of erythrocytes from venules to interstitium, with further movement to microlymphatics and regional lymph nodes. CONCLUSIONS: Rat mesentery as an animal model in combination with an advanced optical imaging system is valuable in studying microlymphatic disturbances in mesentery during the development of experimental lymphedema from latent period to chronic stages, including monitoring of individual cell dislocation with high resolution optical imaging.
Assuntos
Capilares/ultraestrutura , Modelos Animais de Doenças , Imageamento Tridimensional , Vasos Linfáticos/ultraestrutura , Linfedema/patologia , Animais , Masculino , Mesentério/irrigação sanguínea , Mesentério/cirurgia , Mesentério/ultraestrutura , Ratos , Ratos Sprague-DawleyRESUMO
Transmission electron microscopy was employed to study structural changes in the lantern muscles occurring during the transition from young to adult in the sea urchin Strongylocentrotus nudus. A comparative examination of four major lantern muscles (compass depressors, compass elevators, protractors and retractors) suggests that myogenesis involves four consecutive stages. At the initial stage, the muscles show the organization of a mesentery delimited by pseudostratified coelomic epithelia, which are composed of peritoneal cells spanning the whole height of each epithelium, and myoepithelial cells, which are clustered together to fill the interstices between the basal processes of the peritoneal cells. During the next stage, the clusters of myoepithelial cells partly "sink" into the underlying connective tissue. At the third stage of muscularization, the myoepithelial cells increase in size and further invade the underlying connective tissue so that the myoepithelium splits into an apical peritoneal layer and a deeper mass of myoepithelial cells immersed in the connective tissue. However, these two layers are connected by a continuous basal lamina. This is thus the first description of an intermediate developmental stage between pseudostratified myoepithelim and genuine echinoderm muscles. For such a myoepithelium, we propose the term "immersed myoepithelium". At the most advanced stage of myogenesis, the myocytes detach completely from the epithelium to form subepithelial muscle bundles. Myogenesis in the sea urchin takes a long time during which continuous myogenic differentiation occurs in the coelomic epithelium and the newly formed myocytes and associated neurons penetrate into the underlying connective tissue.
Assuntos
Desenvolvimento Muscular/fisiologia , Músculo Esquelético/citologia , Músculo Esquelético/crescimento & desenvolvimento , Strongylocentrotus/citologia , Fatores Etários , Animais , Membrana Basal/ultraestrutura , Diferenciação Celular/fisiologia , Movimento Celular/fisiologia , Tecido Conjuntivo/ultraestrutura , Células do Tecido Conjuntivo/citologia , Células do Tecido Conjuntivo/ultraestrutura , Desmossomos/ultraestrutura , Células Epiteliais/citologia , Células Epiteliais/ultraestrutura , Epitélio/ultraestrutura , Mesentério/citologia , Mesentério/ultraestrutura , Microscopia Eletrônica de Transmissão , Modelos Biológicos , Células Musculares/citologia , Células Musculares/ultraestrutura , Músculo Esquelético/ultraestrutura , Fibras Nervosas/ultraestrutura , Neurônios/ultraestrutura , Peritônio/citologia , Peritônio/ultraestrutura , Strongylocentrotus/crescimento & desenvolvimento , Strongylocentrotus/ultraestruturaRESUMO
Holothurians (sea cucumbers) have been known from ancient times to have the capacity to regenerate their internal organs. In the species Holothuria glaberrima, intestinal regeneration involves the formation of thickenings along the free mesentery edge; these thickenings will later give rise to the regenerated organ. We have previously documented that a remodeling of the extracellular matrix and changes in the muscle layer occur during the formation of the intestinal primordium. In order to analyze these changes in depth, we have now used immunocytochemical techniques and transmission electron microscopy. Our results show a striking disorganization of the muscle layer together with myocyte dedifferentiation. This dedifferentiation involves nucleic activation, disruptions of intercellular junctions, and the disappearance of cell projections, but more prominently, the loss of the contractile apparatus by the formation and elimination of spindle-like structures. Muscle dedifferentiation can be seen as early as 2 days following evisceration and continues during the next 2 weeks of the regeneration process. Dedifferentiation of myocytes might result in cells that proliferate and give rise to new myocytes. Alternatively, dedifferentiating myocytes could give rise to cells with high nuclear-to-cytoplasmic ratios, with some being eliminated by apoptosis. Our results, together with those in other regeneration models, show that myocyte dedifferentiation is a common event in regeneration processes and that the dedifferentiated cells might play an important role in the formation of the new tissues or organs.
Assuntos
Intestinos/fisiologia , Mesentério/fisiologia , Músculo Liso/fisiologia , Regeneração/fisiologia , Pepinos-do-Mar/fisiologia , Animais , Imuno-Histoquímica , Mucosa Intestinal/citologia , Mucosa Intestinal/fisiologia , Mucosa Intestinal/ultraestrutura , Intestinos/citologia , Intestinos/ultraestrutura , Mesentério/citologia , Mesentério/ultraestrutura , Músculo Liso/citologia , Músculo Liso/ultraestrutura , Pepinos-do-Mar/anatomia & histologiaRESUMO
Flow cytometry is a well-established, powerful technique for studying cells in artificial flow in vitro. This review covers a new potential application of this technique for studying normal and abnormal cells in their native condition in blood or lymph flow in vivo. Specifically, the capabilities of the label-free photothermal (PT) technique for detecting and imaging cells in the microvessel network of rat mesentery are analyzed from the point of view of overcoming the problems of flow cytometry in vivo. These problems include, among others, the influences of light scattering and absorption in vessel walls and surrounding tissues, instability of cell velocity, and cells numbers and positions in a vessel's cross-section. The potential applications of this new approach in cell biochemistry and medicine are discussed, including molecular imaging; studying the metabolism and pathogenesis of many diseases at a cellular level; and monitoring and quantifying metastatic and apoptotic cells, and/or their responses to therapeutic interventions (e.g., drug or radiation), in natural biological environments.
Assuntos
Separação Celular/métodos , Citometria de Fluxo/métodos , Mesentério/ultraestrutura , Fotometria/métodos , Termografia/métodos , Animais , Ouro/química , Humanos , Interpretação de Imagem Assistida por Computador/métodos , Linfa/fisiologia , Mesentério/irrigação sanguínea , Camundongos , Ratos , Análise Espectral/métodosAssuntos
Meningioma/diagnóstico , Mesentério/patologia , Neoplasias de Bainha Neural/diagnóstico , Neoplasias Peritoneais/diagnóstico , Proliferação de Células , Colágeno Tipo IV/análise , Diagnóstico Diferencial , Feminino , Humanos , Imuno-Histoquímica , Meningioma/química , Meningioma/ultraestrutura , Mesentério/química , Mesentério/ultraestrutura , Microscopia Eletrônica de Transmissão , Pessoa de Meia-Idade , Mucina-1/análise , Neoplasias de Bainha Neural/química , Neoplasias de Bainha Neural/ultraestrutura , Proteínas do Tecido Nervoso/análise , Neoplasias Peritoneais/química , Neoplasias Peritoneais/ultraestrutura , Receptores de Fatores de Crescimento/análise , Receptores de Fator de Crescimento Neural , Vimentina/análiseRESUMO
Complex comparative analysis of the organization of smooth muscle (SM) forming the wall of lymphatic vessels in bovine small intestinal mesenterium was performed using the methods of morphometry, quantitative histochemistry (including the analysis of nuclear DNA content, and cytoplasmic protein content) and electron microscopy. SM cells (SMC) isolated by dissociation were studied and were found to possess various levels of differentiation, associated with specific morphometric and metabolic characteristics. The structure of SMC population was shown to vary in both different parts of lymphatic bed and within the wall of an individual lymphangion. The results obtained indicate the cellular heteromorphism of lymphatic bed SM. The peculiarities of SM organization in lymphatic vessels are functionally dependent and are determined not only by the level of SM representation in their wall but also by the proportions of different SMC types.
Assuntos
Vasos Linfáticos/citologia , Miócitos de Músculo Liso/citologia , Animais , Bovinos , DNA/análise , Intestino Delgado/ultraestrutura , Vasos Linfáticos/ultraestrutura , Masculino , Mesentério/ultraestrutura , Miócitos de Músculo Liso/ultraestruturaRESUMO
The infrequent occurrence of septic shock in patients with inherited deficiencies of the terminal complement components experiencing meningococcal disease led us to suspect that the terminal complement complex is involved in vascular leakage. To this end, the permeabilizing effect of the cytolytically inactive soluble terminal complement complex (SC5b-9) was tested in a Transwell system measuring the amount of fluorescein-labeled BSA (FITC-BSA) leaked through a monolayer of endothelial cells. The complex caused increased permeability to FITC-BSA after 15 min as opposed to the prompt response to bradykinin (BK). The effect of SC5b-9 was partially reduced by HOE-140 or CV-3988, two selective antagonists of BK B2 and platelet-activating factor receptors, respectively, and was completely neutralized by the mixture of the two antagonists. Also, DX-88, a specific inhibitor of kallikrein, partially inhibited the activity of SC5b-9. The permeabilizing factor(s) released after 30 min of incubation of endothelial cells with SC5b-9 caused a prompt leakage of albumin like BK. Intravital microscopy confirmed both the extravasation of circulating FITC-BSA across mesenteric microvessels 15 min after topical application of SC5b-9 and the complete neutralization by the mixture of HOE-140 and CV-3988. SC5b-9 induced opening of interendothelial junctions in mesenteric endothelium documented by transmission electron microscopy.
Assuntos
Bradicinina/fisiologia , Permeabilidade Capilar/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/fisiologia , Fluoresceína-5-Isotiocianato/análogos & derivados , Fator de Ativação de Plaquetas/fisiologia , Animais , Bradicinina/farmacologia , Permeabilidade Capilar/efeitos dos fármacos , Linhagem Celular , Complexo de Ataque à Membrana do Sistema Complemento/administração & dosagem , Complexo de Ataque à Membrana do Sistema Complemento/farmacologia , Citotoxicidade Imunológica/efeitos dos fármacos , Relação Dose-Resposta Imunológica , Endotélio Vascular/citologia , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/imunologia , Endotélio Vascular/metabolismo , Fluoresceína-5-Isotiocianato/administração & dosagem , Fluoresceína-5-Isotiocianato/farmacologia , Humanos , Íleo/irrigação sanguínea , Íleo/imunologia , Íleo/ultraestrutura , Junções Intercelulares/efeitos dos fármacos , Junções Intercelulares/imunologia , Masculino , Mesentério/irrigação sanguínea , Mesentério/imunologia , Mesentério/ultraestrutura , Perfusão , Ratos , Ratos Endogâmicos WKY , Soroalbumina Bovina/administração & dosagem , Soroalbumina Bovina/farmacologia , SolubilidadeRESUMO
OBJECTIVE AND DESIGN: Unregulated chronic inflammatory process partly due to an estrogen deficiency may render postmenopausal women vulnerable to degenerative conditions such as arthritis, osteoporosis, atherosclerosis, and Alzheimer's disease. Current confusion regarding therapeutic efficacy of estrogen replacement therapy may be due to different estrogen formulations used, short term therapy, as well as advanced stage of the disease. MATERIALS AND METHODS: We compared anti-inflammatory activities of two major estrogen preparations, conjugated equine estrogen (CEE) and 17-beta estradiol, using an animal model (rat mesentery) of in vivo inflammatory reaction to intravenously infused amyloid-beta, examined by video recording and subsequently analyzed by transmission electron microscopy. Cellular markers of inflammation were monitored: leukocyte migration, platelet activation, mast cell activation/degranulation, and endothelial disruption. RESULTS: Low doses of CEE (0.3 mg/kg for 3 weeks) demonstrated significant anti-inflammatory activity, whereas even at high doses (2.0 mg) 17-beta estradiol had only minimal activity. CONCLUSION: CEE, a mixture of several compounds, may have some component(s) with significant anti-inflammatory activity. The anti-inflammatory activity of CEE may have a role in prevention of several degenerative diseases associated with menopause.
Assuntos
Anti-Inflamatórios/farmacologia , Estradiol/farmacologia , Estrogênios Conjugados (USP)/farmacologia , Estrogênios/farmacologia , Peptídeos beta-Amiloides/metabolismo , Animais , Plaquetas/ultraestrutura , Vasos Sanguíneos/ultraestrutura , Adesão Celular , Feminino , Inflamação , Leucócitos/ultraestrutura , Mastócitos/ultraestrutura , Mesentério/ultraestrutura , Microscopia Eletrônica , Microscopia de Vídeo , Ratos , Ratos Sprague-DawleyRESUMO
Recent electrophysiological studies have suggested that there is a subpopulation of cells in lymphatic vessels which act as pacemakers controlling the characteristic spontaneous contractile activity in this tissue. In this study, electron microscopy and immunohistochemical techniques were used on sheep mesenteric lymphatic vessels to investigate the morphology of the cells comprising the lymphatic wall. The smooth muscle cells were not orientated in circular and longitudinal layers as is seen in the gastrointestinal tract, but were arranged in bundles which interlock and cross over in a basket-weave fashion. Antibodies to Kit and vimentin, which are widely used to label specialised pacemaking cells in the gastrointestinal tract (known as interstitial cells of Cajal), demonstrated the existence of an axially orientated subpopulation of cells lying between the endothelium and the bulk of the smooth muscle. Examination of this area using electron microscopy showed cells which were electron dense compared to the underlying smooth muscle and contained caveolae, Golgi complexes, mitochondria, 10-nm filaments, a well-developed endoplasmic reticulum and a basal lamina. The smooth muscle cells typically contained caveolae, dense bodies, mitochondria, abundant filaments, sER and basal laminae. Cells dispersed for patch-clamp studies were also stained for vimentin and myosin. Myosin-staining cells had the typical spindle appearance of smooth muscle cells whereas the vimentin-positive cells could either be branched or more closely resemble the smooth muscle cells. The present study provides the first morphological evidence that specialised cells exist within the vascular system which have the ultrastructural characteristics of pacemaker cells in other tissues and are vimentin and Kit positive.
Assuntos
Relógios Biológicos/fisiologia , Sistema Linfático/ultraestrutura , Mesentério/ultraestrutura , Contração Muscular/fisiologia , Miócitos de Músculo Liso/ultraestrutura , Proteínas Proto-Oncogênicas c-kit/metabolismo , Animais , Tamanho Celular/fisiologia , Feminino , Imuno-Histoquímica , Sistema Linfático/metabolismo , Masculino , Mesentério/metabolismo , Microscopia Eletrônica , Miócitos de Músculo Liso/metabolismo , Miosinas/metabolismo , Organelas/metabolismo , Organelas/ultraestrutura , Vimentina/metabolismoRESUMO
In single perfused mesenteric microvessels of pithed frogs, we assessed wall strength from the critical pressure, PB, which has to be applied within the vessel in order to induce openings in the walls through which fluid and cells can extravasate. PB was determined in capillaries and venules of tissues at 12-20 The P(B) (mean +/- S.E.M.) in 22 vessels between 12 and 20 degrees C, P(B) was 92.0 +/- 7.40 cm H2O which was significantly higher than at room temperatures (P<0.001). The compliance of the vessel wall was estimated using both the red cell method and the oil meniscus technique. There was no measurable effect of temperature on wall compliance. The compliance of vessels from which the cells had been removed by previous perfusion with detergent solutions was very similar to that of intact vessels between 12 and 20 degrees C and between 0 and 5 degrees C. The negligible effects of temperature upon compliance suggest that microvessel walls have to be distended to a greater extent in cold tissue before P(B) is reached. This, together with their rapid closure, is consistent with the hypothesis that pressure-induced openings in microvascular walls are dependent on an active response of the endothelium rather than being the result of stress failure of the basement membrane.
Assuntos
Capilares/fisiologia , Mesentério/irrigação sanguínea , Temperatura , Resistência Vascular/fisiologia , Vênulas/fisiologia , Animais , Membrana Basal/fisiologia , Velocidade do Fluxo Sanguíneo/efeitos dos fármacos , Pressão Sanguínea/fisiologia , Permeabilidade Capilar/fisiologia , Complacência (Medida de Distensibilidade) , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/fisiologia , Endotélio Vascular/ultraestrutura , Eritrócitos/citologia , Técnicas In Vitro , Mesentério/citologia , Mesentério/ultraestrutura , Microcirculação/efeitos dos fármacos , Microcirculação/fisiologia , Microcirculação/ultraestrutura , Microscopia Eletrônica , Octoxinol/administração & dosagem , Resistência Vascular/efeitos dos fármacosRESUMO
Tumor microvessels are hyperpermeable to plasma proteins, a consequence of tumor cell-secreted vascular permeability factor/vascular endothelial growth factor (VPF/VEGF). However, the pathways by which macromolecules extravasate from tumor vessels have been little investigated. To characterize tumor vessels more precisely and to elucidate the pathways by which macromolecules extravasated from them, we studied two well-defined, VPF/VEGF-secreting murine carcinomas, MOT and TA3/St. Whether grown in ascites or solid form, MOT tumors induced large, pericyte-poor "mother" vessels whose lining endothelium developed fenestrae that involved 1.8-5.6% of the surface. Fenestrae developed in parallel with markedly reduced endothelial cell vesiculo-vacuolar organelles (VVOs). TA3/St tumors, which secreted more VPF/VEGF than MOT tumors, elicited mother vessels with unchanged VVOs and without fenestrae. In both tumors, a plasma protein tracer, ferritin, extravasated through VVOs and in MOT tumors ferritin also extravasated through fenestrae. Endothelial gaps were not observed in either tumor. Thus, not all VPF/VEGF-secreting tumors induce fenestrated endothelium. Also, VVOs provide an internal store of membrane that can be transferred to the endothelial cell surface to provide the substantial increase in plasma membrane necessary for mother vessel formation in MOT tumors. Such transfer was apparently unnecessary in TA3/St tumors in which extensive early endothelial cell division provided the increased plasma membrane necessary for forming mother vessels.