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N R Grande

Publications and source records attributed to N R Grande.

12 recordsLinked to original sources
PubMed1994

Neoformation of blood vessels in association with rat lung fibrosis induced by bleomycin.

We have used intratracheal instillation of bleomycin in rats to study the microanatomical changes of blood vessels associated with lung fibrosis. Bleomycin is a toxic cytostatic drug employed in classical models of lung fibrosis. Wistar rats were submitted to intratracheal injection of 1.5 units of bleomycin and sacrificed 2.5 months later, a timing when marked fibrosis of the lung is observed. We casted the vascular tree of the rat lungs by perfusion with a methacrylate resin. These casts were studied by scanning electron microscopy. Lung tissue was also studied by light microscopy and thin section electron microscopy. The major vascular modifications observed in the bleomycin-treated rats were: (1) neoformation of an elaborate network of vessels located in the peribronchial domains of the lung, and (2) distortion of the architecture of alveolar capillaries. By light microscopy, it was clear that the newly formed vascular network was located in regions of fibrosis (which in the resin casts were digested away). These neoformed vessels appeared to originate from bronchial arteries. Thin section electron microscopy revealed that endothelial cells of the neoformed vessels were plump, presented large nuclei, and showed numerous pinocytotic vesicles that were also observed in subendothelial pericytes. The alveoli of the bleomycin-treated rats were heterogeneous in size and shape in contrast with the homogeneity of alveoli of control animals. The alveolar capillaries of fibrotic lungs appeared to occupy a larger volume of the alveolar wall than alveolar capillaries of control rats. Our findings indicate that lung fibrosis encompasses marked changes of the vascular system, namely, the neoformation of vessels and the rearrangement of alveolar capillaries. These structural changes suggest that fibrotic transformation of the lung is associated with the local generation of angiogenic stimuli.

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PubMed1992

Blood vasculature of the lymph node in the dog: anatomical evidence for participation of extrahilar arterial vessels in the blood supply of the cortex.

The organization of the arterial vessels of dog lymph nodes (LN) was studied using methods of visualization of the vasculature by systemic injection of different tracers (colloidal carbon, Micropaque resin and methylmethacrylate) followed by observation of the samples by light microscopy (after clearing of the thick sections of LN) or scanning electron microscopy (corrosion casts). LN from all of the three groups of nodes studied (tracheobronchial, paratracheal and popliteal) showed an extensive network of arterial vessels encircling the capsule of the organ. We found that branches of these capsular arteries penetrated deeply into the cortical domain of LN. The capsule-originating vessels appeared to have a significant participation in the blood supply of the LN parenchyma at the cortical domains of the organs. Our findings are in contrast with current views on the angiology of the LN that consider that virtually all of the arterial capillaries of the LN parenchyma come from hilar arteries. We propose, therefore, that important segments of the LN cortex receive their blood supply from capsular arteries rather than from hilar vessels.

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PubMed1992

Particle clearance from the canine pleural space into thoracic lymph nodes: an experimental study.

We instilled tungsten powder (CaWO4) into the pleural space of the dog and studied the kinetics and distribution of particle translocation from the pleural space to the thoracic lymph nodes over 1-7 days. We found that the transport of tungsten particles to regional lymph nodes was present at day 1, and reached its peak at day 3. In situ detection of tungsten by elemental particle analysis of lymph node sections by scanning electron microscopy complemented by light microscopy and X-ray analysis allowed precise mapping of the marker in the thoracic nodes. The first lymph nodes to become tungsten-laden was the parasternal group (day 1-3). From day 3 to 7 tungsten inclusions decreased in these parasternal nodes while moderately increasing in the remaining intrathoracic lymph nodes. Retrocardiac pleural folds containing numerous "milky spots" also accumulated prominent amounts of tungsten early after intrapleural injection of CaWO4. These data indicate that 1) particle translocation from the pleural space to regional lymph nodes is a rapid process and is first directed to the parasternal lymph nodal subgroup; 2) particle dissemination to virtually all other lymph nodes within the thorax follows thereafter; 3) retrocardiac pleural folds contribute to the clearance of particles from the pleural space.

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PubMed1992

Microanatomy of the blood vasculature of lymph node follicles in the dog.

We studied the anatomical features of the arterial vasculature of the lymphoid follicles (LF) of tracheobronchial lymph nodes in the dog. The microvascular organization of the lymph node was visualized after systemic perfusion of the canine arterial system with gelatinated colloidal carbon. We found that LF contained a substantial number of blood capillaries forming a vascular network with a density comparable to that of neighboring domains of the node. The density of the vascular supply was greater in the secondary than in the primary LF. The geometric arrangement of LF capillaries was also different in the two types of LF. These findings are in contrast with the current view that the LF is a vascular-poor domain of the lymph node. We also document the plasticity of the LF blood microvasculature that appears to proliferate in conjunction with lymphoblast turnover and immune reactivity characteristic of secondary LF.

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PubMed1992

Blood vessel architecture in lymph nodes of the dog viewed by scanning electron microscopy.

Scanning electron microscopy was used to investigate the submicroscopic organization of the blood vessels of dog lymph nodes (LN). The LN vessels were casted by systemic perfusion of the animals vasculature with two resins of distinct viscosity. This resulted in the retrieval of two types of LN vascular replicas that depicted either the arterial blood system alone (methacrylate casting) or both the arterial and venous blood systems (Mercox casting). We found that the dog LN showed a significantly higher density of arterial vessels in the cortex than in the medulla. In the cortical domain, a subcapsular layer stood out because of its rich content in arterial capillaries. The use of the high-resolution resin (mercox) resulted in excellent structural detail of the luminal surface of the LN vessels that allowed a clear distinction between arterioles and venules based on the geometrical pattern of the imprints left in the replicas by the nuclei of endothelial cells. At the cortex-medulla frontier, most arterioles showed narrowings of their lumen that suggested the existence of sphincters at this level. Our findings document that the microanatomical arrangement of blood vessels in the LN of the dog is different from that of LN from other mammals studied so far, in particular from rodents where vascular-poor microdomains have been reported in the LN cortex. The arteriolar sphincters that we detected at the innermost zone of the cortex may represent the structural counterpart of the physiological modulation of the blood supply of the cortex exerted by arterial branches coming from the hilus.

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PubMed1992

Structural artifacts and advantages of cytocentrifugation of cells as viewed by scanning electron microscopy.

Cytocentrifugation of cell suspensions onto glass slides is a widely used procedure in contemporary cytology. We employed here scanning electron microscopy (SEM) to investigate putative morphological changes induced in cells submitted to cytocentrifugation. The fine structure of murine pleural exudate cells (macrophages mainly) processed by spinning was compared with that of similar cells treated without centrifugation (poly-L-lysine attachment of the cells to glass slides at 1 g). Cells of cytocentrifuged preparations showed a significant increase in diameter and smoothening of the cell surface as compared with the morphology of non-centrifuged cells. Cytocentrifugation also induced the formation of thin elongations coming out of the cellular outlines. The centrifugation-induced flattening of the pleural macrophages improved the detection of large intracellular inclusions (containing tungsten particles): these bodies were readily identified by secondary-electron imaging mode of SEM in cytospinned cells whereas their detection in non-centrifuged spherical cells required the use of the backscattered-electron imaging mode of SEM. We conclude that the cytocentrifugation methodology, on one hand, requires caution on the interpretation of the microanatomy of the cells and, on the other hand, the procedure may be an adequate method to improve the identification of large intracellular inclusions by routine (secondary-electron imaging mode) SEM.

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PubMed1991

Inflammatory macrophages in the dog contain high amounts of intravesicular ferritin and are associated with pouches of connective tissue fibers.

We have studied the subcellular distribution of ferritin in inflammatory macrophages present in regional lymph nodes from dogs subjected to a pulmonary inflammatory reaction. The inflammatory reaction was induced by intrabronchial instillation of calcium tungstate (CaWO4), a water-insoluble powder. Ferritin was identified by electron microscopy, and its electron density was enhanced by the use of a modified Perls method. From day 14 on after the CaWO4 deposition, tungsten-positive lymph node macrophages showed a massive accumulation of ferritin. Most of the ferritin was stored in membrane-bounded vesicles that showed heterogeneous concentrations of the protein. A significant complement of ferritin was also detected in the cytoplasmic ground substance of phagocytes. The cell surface of the ferritin-rich, tungsten-positive macrophages showed deep infoldings that encompassed small pockets of connective tissue fibers. These features were not observed in control samples or in lymph nodes from dogs subjected to CaWO4-induced inflammation for periods shorter than 1 week. Our data indicate that inflammatory macrophages greatly increase their content of ferritin macrophages greatly increased their content of ferritin and that ferritin is stored predominantly by a membrane-bounded vesicular compartment. This is in contrast with suggestions that the inflammation-induced increase in macrophage iron is restricted to the labile pool of iron and it does not involve the iron bound to ferritin molecules. Our observation of nodules of connective-tissue fibers in intimate topographical association with ferritin-rich macrophages may indicate that the increase in intracellular ferritin in the macrophage is in some way related to the secretion of factors by the phagocyte that will stimulate fibrillogenesis by neighboring fibroblats.

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PubMed1990

Time course and distribution of tungsten-laden macrophages in the hilar lymph nodes of the dog lung after experimental instillation of calcium tungstate into the left apical bronchus.

We sprayed a tungsten powder (CaWO4) into the airway of a single lobe (left apical) of the dog lung in order to study: (a) the kinetics of particle translocation from the bronchoalveolar lining to hilar lymph nodes, and (b) the sorting in lung lymph nodes of inhaled microcrystals. We found that the transport of the tungsten particles to the regional lymph node takes at least 24 hours and reaches its peak at day 7. In situ detection of tungsten by elemental particle analysis of lymph node sections by scanning electron microscopy allowed precise mapping of the marker in the node; the method was complemented by light microscopy and thin-section electron microscopy of the same nodes. Virtually all of the lymph node tungsten was located inside macrophages. The first tungsten-positive macrophages seen in the regional lymph nodes (day 1 to day 3) were restricted to the subcapsular space. This was followed by massive filling of the same sinus and of the narrow interfollicular areas by the particle-laden macrophages (day 3 to day 7). The even distribution of the tungsten-bearing phagocytes found in these anatomical regions of the node indicated that the subcapsular area in the dog was a continuous domain rather than the segmented region observed in nodes of common laboratory animals such as the rat. By day 7 after tungsten instillation, a moderate number of tungsten-positive macrophages was also detected in the paracortical region of the node. Finally, the presence of tungsten-bearing macrophages was extended to the outer lymph node medulla (day 7 to day 14); here, the macrophages were located in association with cords of plasmacytes and showed interdigitations with these lymphocytes. Only minimal amounts of tungsten were detected inside lymphoid follicles in association with dendritic cells. Some of the tungsten initially deposited in the airway of the apical left lung lobe was detected in contralateral hilar lymph nodes. We conclude that: (i) particle translocation from the alveolus to regional lymph nodes is a slow process that is mediated by pulmonary macrophages, in agreement with the findings of Harmsen et al Science 230:1277, 1985); (ii) in the lymph node, particle-bearing macrophages are sorted through narrow interfollicular sinuses into the outer medulla where they interact extensively with plasma cells; (iii) the migrating macrophages cannot penetrate the follicular domains of the node; minute quantities of exogenous particles may, nevertheless, be transferred from macrophages to follicular dendritic cells; and (iv) contralateral drainage may be a feature of the lymphatic system in the lung.

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PubMedDate not supplied

Cellular kinetics of inflammation in the pleural space of mice in response to the injection of exogenous particles.

CD-1 mice were used to study the cellular kinetics of the inflammatory response of the pleural space to the injection of 250 micrograms of silica or of tungsten microparticles. The pleural exudates were collected by lavage of the serous cavity of mice that were sacrificed at 30 min and up to 7 days after the intrapleural instillation of the particles. The samples were studied by light and electron microscopy (transmission and scanning modes); the quantitative cellular kinetics of the inflammation was determined by leukocyte counting in exudates using cytocentrifuge preparations. The normal resident population of cells of CD-1 mice was made up of (2.47 +/- 0.37) x 10(6) cells. It consisted mostly of macrophage-like cells ((2.03 +/- 0.26) x 10(6) cells, 82% of total cells), some lymphocytes ((0.37 +/- 0.07) x 10(6) cells, 15% of total cells), a few mast cells and eosinophilic granulocytes (1-2% of total cells). The initial inflammatory reaction (30-60 min after injection) was characterized by a decrease in the number of cells harvested from the pleural space. This was followed by an intense recruitment of granulocytes and monocytes that resulted in a peak of intrapleural cells at 24 h ((16.8 +/- 4.0) x 10(6) cells induced by silica particles and (18.3 +/- 4.2) x 10(6) cells induced by tungsten particles). In tungsten-injected mice (but not in silica-treated animals) the enhancement in the number of intrapleural macrophages continued up to 72 h after particle injection. The highest percentage of macrophages with ingested tungsten (50% of total macrophages) was found early (6 h) and decreased thereafter; at day 7 it encompassed just 17% of the macrophages. Injection of any of the two particulates led to the disappearance of mast cells from the pleural space of mice. Silica particles attracted a high number of eosinophils to the pleural cavity of mice. Light and electron microscopy documented that pleural macrophages underwent striking morphological changes during the inflammatory response: the phagocytes showed marked increase in size and in number of surface processes, and their cytoplasm often contained large amounts of the injected particles and also of cellular debris. This study establishes the mouse as a reliable animal model to study the dynamics of the pleural space and it offers a precise definition of the cellular kinetics of inflammation in this serous cavity. The data indicate that the kinetics of experimental pleural inflammation induced by particulates may depend on the nature of the injected particles.

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