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R S McCuskey

Publications and source records attributed to R S McCuskey.

At least 55 records · Page 3Linked to original sources
PubMed1988

STM imaging of molecular collagen and phospholipid membranes.

The application of STM to biological materials has been limited by poor conductivity, sample geometry and stability of biological materials. In this paper we describe an STM study of the monomeric helical forms of collagen, a stable, conductive and widely prevalent structural protein. We have also used STM to image artificial Langmuir DPE (dipalmitoyl phosphatidyl ethanolamine) phospholipid membranes. Both molecular collagen and the phospholipid membranes were dried in air on highly oriented pyrolytic graphite (HOPG). Our STM images of collagen dried on HOPG reveal strands 15 A in diameter with a periodicity of about 30 A which correlates with that known to occur in collagen. Spikes which periodically protrude from strands in our STM images of collagen appear to represent pyrrolidine ring structures in the amino acids proline and hydroxyproline. Thus, we report the first STM imaging of native biomolecules revealing intramolecular details and what appear to be specific amino acids. STM imaging of phospholipid membranes show a lattice pattern with densities spaced approximately 4.5 A apart. These are thought to represent individual phospholipid molecules in an artificial membrane formed on the HOPG. We believe STM and its related technologies will have great future utility in biomolecular studies.

Animals↗
PubMed1987

Normal versus abnormal structure: considerations in morphologic responses of teleosts to pollutants.

Consideration of newer more quantitative morphologic approaches to the study of aquatic pollutants can provide opportunity for collaborative/integrated studies with other subdisciplines in toxicology. Current commonly employed morphologic approaches result largely in subjective findings difficult to analyze statistically and often are directed at levels of structural organization inconsistent with biochemical and physiological approaches. We review some of the methods and approaches available for correlated structure/function studies and present examples from normal and altered skin, gill, and liver of teleosts.

Animals↗
PubMed1985

Functional units in rainbow trout (Salmo gairdneri) liver: I. Arrangement and histochemical properties of hepatocytes.

The architectural arrangement and selected histochemical properties of hepatocytes in the rainbow trout (Salmo gairdneri Richardson) were examined. Light and transmission electron microscopic (TEM) examination following fixation by portal venous perfusion revealed a tubular arrangement of hepatocytes. Lobules, as defined in the adult mammal, were absent. Biliary epithelial cells associated with bile preductules and ductules were a prominent feature of trout liver. Patterns and location of reaction products for glucose-6-phosphatase (G-6-Pase), glucose-6-phosphate dehydrogenase (G-6-PDH), and magnesium-dependent adenosine triphosphatase (ATPase), enzymes preferentially distributed in mammalian liver, were demonstrated in trout liver. A slightly heavier staining pattern for G-6-Pase was seen around presumptive portal venules but all other enzyme reaction patterns were uniform throughout the liver parenchyma. Following ATPase localization, four sizes of biliary passageways (canaliculi, bile preductules, ductules, and ducts) were visualized. Maximum glycogen retention was achieved with freeze-drying and glycolmethacrylate embedding and with this method intense, uniform glycogen staining was observed in all areas of the liver. Companion TEM examinations revealed large depots of glycogen within hepatocytes. The results are important for interpretation and description of the effects of toxic/carcinogenic alteration on trout liver.

Adenosine Triphosphatases↗
PubMed1984

Deficient Kupffer cell phagocytosis and lysosomal enzymes in the endotoxin-low-responsive C3H/HeJ mouse.

Various substances, including lysosomal enzymes, are produced by Kupffer cells and other macrophages; their release has been implicated in the toxic response to endotoxins. C3H/HeJ mice exhibit little or no response to doses of endotoxin that are lethal in syngeneic C3HeB/FeJ mice. To explore the nature of this deficient response, the Kupffer cells of these mice were studied using in vivo microscopic as well as histochemical and electron microscopical methods. In vivo, the rate of phagocytosis of single 0.8 micron latex particles was measured in individual Kupffer cells as was the number of phagocytic cells per microscopic field. Frozen sections of livers were stained for a variety of lysosomal enzymes and liver specimens also were processed for electron microscopy. In comparison to the endotoxin-sensitive C3HeB/FeJ mice, the livers of the C3H/HeJ mice contained 60% fewer Kupffer cells that phagocytosed latex. However, the rate of phagocytosis by these cells was not statistically different and ranged from 19-26 sec. The volume density of acid-phosphatase-positive Kupffer cells was 40% less in the C3H/HeJ mice. Similar differences were observed with other lysosomal enzymes including cathepsins B and H and dipeptidyl peptidases I and II. However, light and electron microscopy revealed a relatively normal number of Kupffer cells in livers stained for peroxidase, a nonlysosomal enzyme. The results suggest that the insensitivity of C3H/HeJ mice to endotoxin may be related in part to a lysosomal enzyme deficiency and a paucity of phagocytic Kupffer cells in these animals.

Animals↗
PubMed1984

Species differences in Kupffer cells and endotoxin sensitivity.

The relative species sensitivity to Escherichia coli O111:B4 endotoxin was found to be guinea pig greater than hamster greater than mouse greater than rat. The 50% lethal dose of this endotoxin correlated with both the rate at which single latex particles were phagocytosed by individual Kupffer cells and the number of Kupffer cells in hepatic lobules that phagocytosed latex. The results suggests that the intrahepatic density and the level of activation of Kupffer cells participate in determining endotoxin sensitivity.

Animals↗
PubMed1984

Microcinematographic analysis of tethered Leptospira illini.

A model of Leptospira motility was recently proposed. One element of the model states that in translating cells the anterior spiral-shaped end gyrates counterclockwise and the posterior hook-shaped end gyrates clockwise. We tested these predictions by analyzing cells tethered to a glass surface. Leptospira illini was incubated with antibody-coated latex beads (Ab-beads). These beads adhered to the cells, and subsequently some cells became attached to either the slide or the cover glass via the Ab-beads. As previously reported, these cells rapidly moved back and forth across the surface of the beads. In addition, a general trend was observed: cells tethered to the cover glass rotated clockwise around the Ab-bead; cells tethered to the slide rotated counterclockwise around the Ab-bead. A computer-aided microcinematographic analysis of tethered cells indicated that the direction of rotation of cells around the Ab-bead was a function of both the surface of attachment and the shape of the cell ends. The results can best be explained by assuming that the gyrating ends interact with the glass surface to cause rotation around the Ab-beads. The analysis obtained indicates that the hook- and spiral-shaped ends rotate in the directions predicted by the model. In addition, the tethered cell assay permitted detection of rapid, coordinated reversals of the cell ends, e.g., cells rapidly switched from a hook-spiral configuration to a spiral-hook configuration. These results suggest the existance of a mechanism which coordinates the shape of the cell ends of L. illini.

Cell Movement↗
PubMed1984

Hepatic microvascular regulatory mechanisms. V. Effects of lodoxamide tromethamine or phentolamine-HCl on vascular responses elicited by serotonin.

Initial changes in hepatic microvasculature and carotid (arterial) blood pressure were measured in anesthetized Sprague-Dawley rats receiving an endoportal injection or topical application of serotonin alone, or in combination with selected doses of lodoxamide tromethamine or phentolamine-HCl. Although infusion of 1.0 microgram per 100 g b.w. serotonin and/or 10 micrograms per 100 g b.w. lodoxamide produced no change in arterial blood pressure, 10 or 20 micrograms per 100 g b.w. serotonin evoked hypotension within 40 sec. During this period, in vivo microscopy of transilluminated livers revealed that 1.0 or 10 micrograms per 100 b.w. serotonin elicited constriction of portal venules and sinusoids, and a significant increase in the percentage of these microvessels containing decreased cellular flow. None of these responses was antagonized by 10 micrograms per 100 g b.w. lodoxamide. Topical application of 1.0, 10, or 20 micrograms serotonin provoked constriction of sinusoids and decreased sinusoidal and central venous perfusion, but no systemic hypotension. There was no significant difference in the magnitude of responses to these doses of serotonin alone or with 0.1 microgram lodoxamide; however, 100 micrograms phentolamine potentiated responses to 1.0 microgram serotonin. Given these results, and those of previous studies indicating that equivalent doses of these antagonists block mast cell degranulation or catecholamine-induced activation of alpha receptors, it is suggested that the vascular responses are mediated directly by serotonin and not indirectly by serotonin-evoked mast cell degranulation or alpha-adrenergic stimulation. The vascular receptor(s) triggered remain(s) to be elucidated.

Amino Acids↗
PubMed1984

In vivo and electron microscopic study of the development of cerebral diabetic microangiography.

The cerebral microvasculature of rats rendered diabetic with streptozotocin (75 mg/kg) and vehicle-treated controls paired for age and sex were studied using in vivo and electron microscopic methods at intervals from two weeks to 12 months after induction of diabetes mellitus. By one month, the pial vessels of diabetics were dilated and tortuous; and, the vasoconstrictive responses of arterioles and venules to local increases of PO2 in the artificial CSF bathing these vessels was reduced as was the linear velocity of blood flow. Increased arterio-venous shunting also was observed. By five months the responsiveness to locally increased levels of PO2 was further reduced from control values of 49.9% +/- 5 (SEM) to 6.9% +/- 1.8 (SEM) in arterioles and from 11.5% +/- 2.1 (SEM) to 2.6% +/- 0.7 (SEM) in venules. No further significant change in responsiveness was measured from five to twelve months. At 5 months, the functional changes were no longer reversible; and focal changes were noted in the thickness and density of the vascular basement membrane. Astrocytic end feet were greatly swollen and contained mitochondria having longitudinal rearrangement of their cristae. Basement membranes contained nodules of electron-lucent material which impinged on degenerating smooth muscle cells, pericytes and astrocytes. In some sites these cells appeared to be replaced by an amorphous material laced with collagen fibrils. By 11 months, these focal lesions were more frequent and more pronounced. Additional cellular replacement had taken place which resulted in greatly widened basement membranes which varied in density and content. The ultrastructure of endothelial cells was not noticeably altered except for presence of numerous cytoplasmic vesicles. The tight interendothelial junctions appeared to be intact even though dramatic changes had taken place perivascularly. These architectural and functional changes in the microvasculature are suggested to result not only from metabolic defects in the vascular wall, but also as a response to a relative hypoxia of the brain during the diabetic state.

Animals↗
PubMed1984

Hematopoiesis on cellulose-ester membranes (CEM). VI. Histochemical evaluation of stromal-CEM interactions after stromal enrichment.

Histochemical procedures were used to elucidate differences in the distribution of glycosaminoglycans (GAGs) and glycoproteins (GPs) in tubes made of cellulose-ester membrane (CEM) in six weeks and 12 months following intraperitoneal (i.p.) implantation. After six weeks, CEM tubes coated on their inner surfaces with bone marrow (BM), bone homogenate (BH), or regenerating medullary mesenchyme (M) contained a highly vascularized loose connective tissue and scattered hematopoietic elements. Numerous mononuclear cells were seen at the stromal-CEM interface and infiltrating the CEM. At the site of infiltration, there were accumulations of GPs and nonsulfated acidic GAGs. The remainder of the CEM contained limited amounts of sulfated and nonsulfated GAGs. By 12 months, both coated and uncoated CEMs were saturated with sulfated and nonsulfated acidic GAGs. However, only coated CEM tubes contained trilineal hematopoiesis surrounded by a shell of bone that was incorporated into the structure of the CEM. Exterior to the bone, the CEM was infiltrated with GPs, some of which were acidic and nonsulfated, possibly sialoglycoproteins. No bone or glycoproteins infiltrated the uncoated CEMs. The stroma supporting trilineal hematopoiesis contained GPs and nonsulfated acidic GAGs, but not sulfated acidic GAGs. However, both sulfated and nonsulfated GAGs were found in the fibrous connective tissue filling the lumens of uncoated CEMs. While CEM tubes accumulate peritoneal GAGs that may be conducive for the retention, differentiation, and proliferation of osteogenic and other stromal cells provided by coatings of BM, BH, or M, it is these cells that produce the bone and glycoproteins that appear to contribute to a microenvironment conducive to trilineal hematopoiesis. However, the relative roles of these substances in this process remain to be elucidated.

Animals↗
PubMed1983

In vivo microscopic observations of the responses of Kupffer cells and the hepatic microcirculation to Mycobacterium bovis BCG alone and in combination with endotoxin.

Kupffer cell function and hepatic microvascular hemodynamics were studied by in vivo microscopy in Mycobacterium bovis BCG-infected NMRI mice before and after treatment with minute (0.01 mu mg) tolerance-producing doses and doses causing 70% lethality (0.5 micrograms) of Escherichia coli 0111:B5 endotoxin alone and in combination. BCG-induced granulomas distorted the hepatic microvasculature and impeded blood flow in many sinusoids; flow also was altered further by leukocytes adhering to the sinusoidal walls and by enlarged Kupffer cells that bulged into the lumen. Nevertheless, in BCG-infected mice, the ratio of Kupffer cells which phagocytosed latex to sinusoids containing blood flow and capable of delivering these particulates to Kupffer cells was significantly greater than that in uninfected mice. The phagocytosis of single latex particles by individual Kupffer cells also was more rapid. This indicated an expansion of the numbers and activation of Kupffer cells. In this hyperreactive state, the tolerance-inducing dose of endotoxin produced no change in the rate of phagocytosis after 2 h. In contrast, the 70% lethal dose reduced the rate by 123%, unless tolerance was induced, in which case there was no reduction in the rate of phagocytosis. Twenty-four hours after injection of the tolerance-inducing dose, however, the rate of phagocytosis was accelerated slightly (17%). This suggested that the Kupffer cells had been activated and perhaps were more effective in clearing subsequent endotoxin from the blood but without sufficient release of toxic substances to be lethal. That some mediators were released, however, was suggested by the microvascular alterations that accompanied the above phagocytic responses. These results further support the concept of a central role for Kupffer cells in endotoxin-mediated, nonspecific host defense mechanisms.

Animals↗
PubMed1983

A compartmental analysis of the splenic circulation in rat.

Isolated, acellular washout experiments of 125I-labeled bovine serum albumin (BSA) from control, anemic, and polycythemic rat spleens were used to develop a model of the splenic plasma circulation. The results indicated that the plasma circulation can be described adequately by two compartments. As in red blood cell (RBC) washouts [Am. J. Physiol. 239 (Heart Circ. Physiol. 8): H272-H277, 1980] the fast compartment represents intrasplenic vessels that bypass the red pulp, whereas the intermediate/slow compartment represents plasma flow through the red pulp (filter). The combined plasma and RBC parameters suggest the rat spleen is not an RBC reservoir and that splenic RBC filtration capacity decreases during polycythemia and anemia. The ratio of fast compartment to systemic hematocrit indicates hemodilution occurs, supporting the concept of plasma skimming. A small plasma holdup occurs in the red pulp of anemic and polycythemic spleens probably due to RBC congestion. This congestion, in turn, might be due to reticulocyte sequestration and/or erythropoiesis in anemic spleens and RBC sequestration and/or destruction in polycythemic spleens. There is plasma redistribution in polycythemic spleens possibly to meet the increased metabolic demand.

Anemia↗
PubMed1983

Scanning electron microscope observations on the structure of portal veins, sinusoids and central veins in rat liver.

Portal veins can be recognized in SEM by the presence of an accompanying element such as the bile ductule. Endothelial cells within these vessels possess small marginal microvilli. Portal veins show few sinusoidal inlets (direct connections to sinusoids) and no attached "wandering" cells. Sinusoids in periportal areas are narrower and more tortuous than the wider and straighter central ones. Periportal endothelial fenestrae have a larger diameter (111 nm) than central ones (105 nm), but their number per square micron, and therefore the porosity (% open area), is higher in centrolobular sinusoids (7.94 vs 5.96%). Central veins have smoother endothelial cells and show numerous sinusoidal outlets (direct sinusoidal connections). Cells are seen attached to the wall and are probably migrating out of the liver. In the orifices of some sinusoidal outlets, traversing bars can be observed, which obstruct the flow of blood from the sinusoids into the central vein. From our observations and measurements the following conclusions can be drawn: a. In the narrower periportal sinusoids, moving blood cells will be forced against the wall, causing a massage of the space of Disse. This will influence the exchange of fluids and particles through the endothelial fenestrae, and it will cause stirring of the fluid in the space of Disse. b. Centrolobular sinusoids have a larger perimeter and an endothelial lining with a higher degree of porosity. These structural factors will favour transport and uptake processes in the centrolobular parenchyma. c. Sinusoidal blood flow might be expected to be slower in central areas, since the number of sinusoids appear to equal that of the portal region, but their lumen is wider.

Animals↗
PubMed1983

Hepatic microvascular regulatory mechanisms. III. Aminergic mechanisms as related to mast cells.

The responses of the hepatic microvasculature to aminergic stimulation were evaluated in Sprague-Dawley rats anesthetized with urethane or pentobarbital. Various concentrations (10(-10)-10(-4) g/ml) of serotonin, histamine or compound 48/80 (a mast cell degranulator) alone or in combination with appropriate blockers were administered topically to the livers of these rats while changes in the microvasculature were measured for a period of 15 min using in vivo microscopic methods. The influence of compound 48/80 on hepatic mast cells was verified by histochemical methods. Histamine induced dilatation of portal venules which was not antagonized by diphenhydramine (H1-blocker) or metiamide (H2-blocker). Constriction was produced by serotonin in sinusoids and by 48/80 in sinusoids and central venules, but not in other parts of the microvasculature. Vasoconstriction was accompanied by enlargement and bulging of the nuclear region of cells lining sinusoids. Concomitantly, platelets and leukocytes adhered to the endothelium of sinusoids, and central and sublobular venules. None of these responses was antagonized by methysergide. Given these results and histochemical demonstration of the release of mast cell constituents following 48/80-induced degranulation, it is hypothesized that the responses in sinusoids and hepatic venules are mediated by serotonin and other mast cell constituents (e.g., prostaglandins, PAF, heparin), but at sites other than histamine and serotonin-specific receptors. These results support the involvement of mast cell constituents in hepatic microvascular regulatory mechanisms.

Animals↗
PubMed1982

In vivo microscopic studies of the responses of the liver to endotoxin.

In vivo microscopic methods concomitant with electron microscopic and histochemical procedures are being used to explore the sequelae of responses of Kupffer cells and the hepatic microvasculature to endotoxins. To gain further insight into the role of the liver in host defense and nonspecific resistance, the effects of endotoxin also are being studied in animals sensitized to endotoxin (BCG infection) or tolerant to endotoxin (pretreated with detoxified endotoxin, low doses of endotoxin, or in C3H/HeJ mice). The results to date, have demonstrated that endotoxin induces significant alterations in the hepatic microcirculation due to swelling of Kupffer and endothelial cells and the adhesion of leukocytes and platelets to the sinusoid wall. Lymphocytes frequently are associated with the Kupffer cells. Phagocytosis also is affected; following a brief period of stimulation, the rate of phagocytosis by Kupffer cells is depressed. In BCG infected animals all of these responses are exaggerated but can be minimized by pretreatment with detoxified endotoxin or minute concentrations of endotoxin 24 h prior to the challenge dose of endotoxin. The responses are not seen in the endotoxin low-responder, C3H/HeJ mouse which was found to have a deficiency in lysosomal enzymes and a paucity of functional Kupffer cells. The results provide some insight into the sequelae of cellular and microvascular events that occur in the liver during endotoxemia, endotoxin-related host defense mechanisms and non-specific resistance. In addition, support is provided for the central role of Kupffer cells in these events and that lysosomal enzymes participate in the toxic response elicited by endotoxin.

Animals↗
PubMed1981

Selective extravascular escape of albumin into the cerebral cortex of the diabetic rat.

The extravasation of plasma proteins (albumin, IgG, and complement C3) into the cerebral cortex was studied in streptozotocin-induced diabetic rats using immunohistochemical techniques. The results indicate that albumin, but not IgG or complement C3, selectively enters the cerebral cortex within 2 wk after induction of diabetes. It is suggested that albumin may be an oncotic substance that contributes to diabetic cerebral microangiopathy, astrocytic swelling, and the cerebral edema that occasionally is seen during fluid and insulin therapy of juvenile ketoacidotic diabetes.

Animals↗