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Biomedical subjects

L P Watson

Publications and source records attributed to L P Watson.

At least 19 recordsLinked to original sources
PubMed1999

Superior mesenteric artery dilatation alone does not account for glucose-induced hypotension in human sympathetic denervation.

Haemodynamic and hormonal effects of two oral isovolaemic, isoosmotic solutions of 0.5 g/kg and 1.0 g/kg glucose were studied in 10 humans with sympathetic denervation due to primary autonomic failure (AF). Measurements were made supine for 60 min, and also after 5 min 45 head-up tilt, before and 60 min after glucose. There was a similar fall in blood pressure (BP) after each dose, after 0.5 g/kg from 160+/-12 / 87+/-6 to 143+/-13 / 76+/-6 mm Hg, P < 0.05 and after 1.0 g/kg from 160+/-13 / 90+/-6 to 136+/-9 / 76+/-5 mm Hg, P < 0.05. Heart rate, cardiac index and forearm muscle blood flow did not change after either dose. After 0.5 g/kg, superior mesenteric artery blood flow was unchanged but rose significantly after 1.0 g/kg, from 243 (169-395) to 722 (227-982) ml/min, P < 0.05, 15 min after ingestion. BP fell further on tilt 60 min after each dose, but there was no difference between doses. Plasma glucose was higher after 1.0 g/kg but plasma insulin was similar after each dose. Thus, in AF with sympathetic denervation there was no dose-related effect of glucose on supine or postural hypotension. Supine hypotension after glucose was not attributable solely to increased splanchnic blood flow; other factors, including dilatation in other vascular beds may have contributed.

Adult↗
PubMed1998

Differing haemodynamic and catecholamine responses to exercise in three groups with peripheralautonomic dysfunction: insulin-dependent diabetes mellitus, familial amyloid polyneuropathy and pure autonomic failure.

The haemodynamic and catecholamine responses to supine exercise, and the effect on standing blood pressure (BP), were studied in three groups with peripheral autonomic dysfunction; insulin-dependent diabetes mellitus (IDDM), familial amyloid polyneuropathy (FAP) and pure autonomic failure (PAF). Healthy normal subjects were studied as controls. With exercise, BP increased in controls, was unchanged in IDDM and FAP, and fell in PAF. Heart rate (HR) increased more in controls than IDDM, FAP or PAF. Cardiac index (CI) increased less in IDDM than controls, FAP or PAF. Systemic vascular resistance (SVR) fell similarly in controls and IDDM, with a greater fall in FAP and PAF. Plasma noradrenaline increased in controls and IDDM only; plasma adrenaline did not change and plasma dopamine was undetectable in all groups. On standing, BP was unchanged in controls; BP fell pre- and post-exercise in IDDM, FAP and PAF, with a significantly greater fall post-exercise in FAP and PAF. In conclusion, the haemodynamic responses to supine exercise and to standing after exercise differed in the three groups with peripheral autonomic dysfunction. These differences, and also the similarities, between different forms of peripheral autonomic dysfunction, may be of relevance to the clinical assessment and therapy of these patients.

Adult↗
PubMed1997

Haemodynamic and hormonal effects of two different oral glucose loads in normal human subjects.

Systemic and regional haemodynamics and hormonal responses to two isovolaemic, iso-osmotic solutions of 0.5 and 1.0 g/kg of glucose were compared in ten normal young subjects (mean age 24 +/- 3 years). Measurements were made while subjects were supine before glucose and every 15 min for 60 min after ingestion of each solution. Superior mesenteric artery (SMA) blood flow rose similarly after each dose. There were corresponding reductions in SMA vascular resistance after each dose but no difference between doses. Pulsatility index of the SMA was lower after 1.0 g/kg. There was no change in blood pressure, heart rate, cardiac index or forearm muscle blood flow after either dose. Plasma glucose levels rose after each dose and were higher after 1.0 g/kg. There was no difference in the plasma insulin rise between doses. Plasma levels of noradrenaline and adrenaline did not change after either dose. These results suggest that with glucose loads within the ranges we used, changes in SMA blood flow are not dose-related and larger increases in SMA blood flow or in plasma insulin than we observed are needed to reflexly activate cardiac output and raise plasma noradrenaline levels in young normal subjects.

Adult↗
PubMed1996

Neurohumoral, peptidergic and biochemical responses to supine exercise in two groups with primary autonomic failure: Shy-Drager syndrome/multiple system atrophy and pure autonomic failure.

The neurohumoral, peptidergic and biochemical responses to supine leg exercise were studied in two groups with primary autonomic failure: Shy-Drager syndrome (SDS, n = 15) and pure autonomic failure (PAF, n = 15), to determine if these accounted for exercise-induced hypotension and the greater blood pressure (BP) fall in PAF. Responses were compared to normal subjects (controls, n = 15), in whom BP rose with exercise. Resting plasma noradrenaline (NA) was higher in controls than SDS, and was lowest in PAF. With exercise, NA increased in controls, with a small rise in SDS, but no change in PAF. Resting plasma adrenaline (A) was higher in controls and SDS than PAF, with no change during exercise. Plasma dopamine was unrecordable at all stages in all groups. Resting plasma renin activity (PRA) was higher in controls than SDS and PAF, and was unchanged with exercise in all groups. Plasma insulin, C-peptide and serum growth hormone (GH) were similar at rest and with exercise in the three groups. Plasma glucose was higher at rest in SDS and PAF, and increased with exercise in all three groups. In conclusion, neither exercise-induced hypotension, nor the differences between SDS and PAF could be related to abnormalities in the release of A, PRA, insulin, glucose or GH. The abnormal NA response to exercise was consistent with the BP fall being due to inadequate compensatory sympathetic activity. In SDS, the small NA increase, in the presence of supersensitivity, may have reduced their BP fall as compared to PAF. These results suggest that impaired sympathetic neural activity is a key factor in exercise-induced hypotension.

Autonomic Nervous System Diseases↗
PubMed1996

Cardiovascular and catecholamine changes induced by supine exercise and upright posture in vasovagal syncope. Comparisons with normal subjects and subjects with sympathetic denervation.

The haemodynamic and catecholamine responses to supine leg exercise were studied in vasovagal syncope (n = 10), pure autonomic failure (n = 10) and in control (n = 10) subjects. With exercise, blood pressure increased in controls; with a smaller rise in vasovagal syncope, and a substantial fall in pure autonomic failure. Heart rate increased similarly in controls and vasovagal syncope, but less in pure autonomic failure. The increase in cardiac index was less in controls and pure autonomic failure than vasovagal syncope; the fall in systemic vascular resistance was greatest in pure autonomic failure, but also fell more in vasovagal syncope than controls. Plasma noradrenaline levels increased in controls; with a smaller rise in vasovagal syncope and no increase in pure autonomic failure. Plasma adrenaline levels increased in vasovagal syncope only. The blood pressure responses to standing before and after exercise were similar in controls and vasovagal syncope, with no postural blood pressure fall; in pure autonomic failure there was a greater postural blood pressure fall post exercise. In conclusion, with supine exercise, blood pressure rose in controls and vasovagal syncope, and fell in pure autonomic failure. Systemic vascular resistance fell more in vasovagal syncope and pure autonomic failure, than controls. Noradrenaline responses differed and adrenaline rose in vasovagal syncope only. Standing post exercise did not induce syncope in vasovagal syncope, but increased postural hypotension in pure autonomic failure. There are clear differences in response to exercise in vasovagal syncope and pure autonomic failure. The differences between vasovagal syncope and control subjects suggest an underlying abnormality which may predispose to vasodepression in subjects with vasovagal syncope.

Adult↗
PubMed1995

Abnormal cardiovascular and catecholamine responses to supine exercise in human subjects with sympathetic dysfunction.

1. The cardiovascular and catecholamine responses to supine leg exercise were measured in fifteen normal subjects (controls) and in three groups with sympathetic dysfunction: fifteen with central failure (Shy-Drager syndrome; SDS), fifteen with peripheral failure (pure autonomic failure; PAF) and two with isolated dopamine beta-hydroxylase deficiency (DBH deficiency). 2. With exercise, blood pressure increased in controls, fell markedly in SDS and PAF and was unchanged in DBH deficiency. After exercise, blood pressure rapidly returned to baseline in controls, but remained low in SDS and PAF. With exercise, heart rate increased more in controls than SDS or PAF; the response varied in DBH deficiency. 3. With exercise, cardiac output increased similarly in controls, SDS and PAF, with a larger increase in DBH deficiency. Vascular resistance fell less in controls than SDS, PAF and DBH deficiency. 4. With exercise, plasma noradrenaline increased in controls only; plasma adrenaline remained unchanged in all groups. In DBH deficiency, plasma noradrenaline and adrenaline were undetectable, but plasma dopamine was elevated and rose further with exercise. 5. Supine exercise substantially lowered blood pressure in sympathetic failure due to SDS and PAF. In DBH deficiency blood pressure was unchanged; this lack of fall may have been due to vasoconstriction induced by dopamine and other substances released from otherwise intact sympathetic terminals, or to preserved cardiac vagal function.

Adult↗
PubMed1995

Effect of the somatostatin analogue, octreotide, on exercise-induced hypotension in human subjects with chronic sympathetic failure.

1. In autonomic failure, supine exercise lowers blood pressure and worsens postural hypotension. The somatostatin analogue, octreotide, reduces post-prandial and postural hypotension, but its effects on exercise-induced hypotension and on postural hypotension post-exercise are unknown. 2. Eighteen subjects with chronic sympathetic denervation were studied; 12 had pure autonomic failure and six had additional neurological features of the Shy-Drager syndrome. Haemodynamic, hormonal and biochemical changes were measured before, during and after incremental supine leg exercise on two occasions: on no treatment and after subcutaneous octreotide. Exercise was performed 120 min after octreotide in eight subjects and 60 min after octreotide in ten subjects. 3. Octreotide did not improve exercise-induced hypotension; the blood pressure fall was greater during exercise, but the blood pressure level was no different than without treatment. Heart rate, stroke distance, cardiac index and systemic vascular resistance were similar at rest and changed to the same degree with exercise on and off octreotide. After octreotide, resting levels of serum growth hormone, plasma noradrenaline, adrenaline and renin were unchanged, but glucose was higher and insulin was lower. There was no change in biochemical and hormone levels during exercise either off or on octreotide. 4. After octreotide, although the rate of blood pressure recovery was similar post-exercise, the levels of blood pressure were higher than in the non-treatment phase and postural hypotension was improved before and after exercise. 5. In conclusion, in primary autonomic failure, octreotide did not improve exercise-induced hypotension in the supine position, suggesting that octreotide-sensitive vasodilatory peptides do not contribute to the blood pressure fall.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1991

Distribution of proteoglycans during the hair growth cycle in human skin.

The involvement of proteoglycans in hair growth has been recognized through the observation of increased hair growth in diseases such as the mucopolysaccharidoses and pre-tibial myxedema, which involve an increase in skin proteoglycan content. In an attempt to understand this, we have examined the distribution of chondroitin 6 sulphate (C6S), unsulphated chondroitin (COS), dermatan sulphate (DS), and heparan sulphate proteoglycans (HSPG) in frozen tissue sections of normal scalp by immunostaining. Results show that during anagen, the thick connective tissue sheath around the follicle strains strongly for C6S, COS, and DS. COS is uniquely associated with this region and is not found beneath the epidermis or infundibular epithelium. HSPG is, however, localized in the basement membrane zone adjacent to the outer root sheath. In addition, all of these proteoglycans are localized in the dermal papilla. In mid-catagen, we observed significant loss of C6S and COS staining from both the dermal papilla and the connective tissue sheath, but no decrease in staining for HSPG. In late catagen, very little staining of C6S and COS was observed. In early anagen, we observed that C6S was again present in the connective tissue sheath and dermal papilla; however, COS staining appeared to be weaker and less closely associated with the follicle. HSPG staining was observed in early anagen in a pattern very similar to that found for other basement membrane components. Results for DS were not obtained for catagen or early anagen. These results provide further evidence that hair growth is associated with the presence of chondroitin proteoglycans in the follicle environment and that the cessation of growth is associated with their removal. Further studies are underway to characterize the relationship between hair growth and proteoglycans.

Adult↗
PubMed1989

Cytochemical properties of osteoblast cell membrane domains.

The interactions of osteoblasts with one another and with the extracellular milieu are of vital importance for cell function. These interactions are mediated by cell membrane-associated components. In the present work, we studied the distribution of several mediators known to be associated with the cell surface, using ultrastructural cytochemistry, to characterize the three cell membrane domains (osteoid, lateral, and vascular) of osteoblasts. Osteoblasts in neonatal rat calvariae were studied for cell surface distribution of alkaline phosphatase (APase), calcium-activated adenosine triphosphatase (Ca2+-ATPase), calcium, soybean agglutinin (SBA)-reactive sites, and peanut agglutinin (PNA)-reactive sites. APase was absent in the osteoid domain but was evenly distributed in the other domains. Ca2+-ATPase was found to be concentrated mainly in the lateral domains. In contrast, calcium was present in all cell membrane domains. Using lectins conjugated to horseradish peroxidase, we demonstrated that SBA binding sites were evenly distributed along the osteoblast cell membrane, whereas PNA binding sites were absent or minimally present in the osteoid and lateral domains but were evenly distributed in the vascular domain. These results suggest that the various functions of osteoblasts may be facilitated by specialized cell membrane domains which are cytochemically distinct. Previous reports have failed to demonstrate the cytochemical differences between the three domains of the osteoblast cell membrane.

Alkaline Phosphatase↗
PubMed1988

Cytochemical changes in hepatocytes of rats with endotoxemia or sepsis: localization of fibronectin, calcium, and enzymes.

Bacterial lipopolysaccharide (LPS) is known to be implicated in the pathogenesis of endotoxemia and septic shock. The liver is the first vital organ to exhibit pathological alterations in shock. The present studies include immunoelectron microscopic localization of tissue fibronectin and cytochemical localization of calcium and enzymes in hepatocytes of animals with LPS-induced endotoxemia or cecal ligation-induced septic shock. The results showed increased staining of fibronectin in the basal (perisinusoidal) surfaces and in the cisternae of rough endoplasmic reticulum and the Golgi complex of hepatocytes in rats with endotoxemia or septic shock. Intracellular calcium content was significantly increased in the LPS-treated or septic rats. Calcium pyroantimonate precipitate was deposited predominantly on the outer surfaces of the RER of hepatocytes. In addition, diminution or depletion of glycogen, reduction of catalase-containing peroxisomes, increase of G-6-Pase activity, and depletion of cytochrome c oxidase in many mitochondria were also observed in hepatocytes of experimental animals. The overall results suggest that LPS stimulates: (a) hepatic synthesis and secretion of fibronectin; (b) uptake of calcium by hepatocytes; and (c) G-6-Pase activity. LPS treatment also leads to reduced numbers of peroxisomes and depletion of cytochrome c oxidase.

Adenosine Triphosphatases↗
PubMed1988

Incorporation of bacterial lipopolysaccharide by human Leu-11a+ natural killer cells. Ultrastructural and functional correlations.

Lipopolysaccharides (LPS) of gram-negative bacteria are known to augment the ability of macrophages and natural killer (NK) cells to lyse susceptible target cells. In the present studies, we sought correlations between the ultrastructural changes and function of Leu-11a+ cells from the peripheral blood mononuclear cells which occurred as a result of their incorporation of LPS. We also studied the effect of LPS on the NK activity of purified Leu-11a+ cells. LPS samples from E. coli and Pseudomonas aeruginosa were utilized. A double-immunolabeling technique employing immunogold and immunoperoxidase markers was used to identify the Leu-11a+ cells with incorporated LPS. Pinocytosis, phagocytosis, and direct insertion into membrane bilayers appeared to be the routes for incorporation of LPS by Leu-11a+ cells. Subsequent ultrastructural effects included dilation of intracellular membrane compartments, formation of tubuloreticular inclusions and increased acid phosphatase activity. Opsonized Staphylococcus aureus were ingested both by control and LPS-treated Leu-11a+ cells; however, the number of Leu-11a+ cells with bacteria increased significantly as a result of LPS treatment. Autoradiography combined with immunolabeling showed no [3H] thymidine incorporation by either control or LPS-stimulated Leu-11a+ cells. NK cell-mediated cytotoxicity was significantly increased as compared with the control (p less than or equal to 0.02) when isolated Leu-11a+ cells were treated with LPS for 24 hours. This result suggests that LPS may have direct effect on the NK cell activity. Tests of peripheral blood mononuclear cell samples after LPS treatment showed up to a 2-fold increase in NK cytotoxicity and a dose-related increase of in vitro interferon production. This latter finding together with the ultrastructural observations of tubuloreticular inclusions in Leu-11a+ cells suggests that, in addition to any direct effect of LPS, cytotoxic activity could be indirectly augmented as a result of autocrine or paracrine interferon, or lymphokine production.

Acid Phosphatase↗
PubMed1988

Effects of lipopolysaccharide, lipid A, lipid X, and phorbol ester on cultured bovine endothelial cells.

In pursuing the mechanism of endotoxin action, we examined the effect of lipopolysaccharide (LPS) and its chemically defined components, lipid A and lipid X on cultured bovine endothelial cells. We report that LPS and lipid A caused detachment and altered morphology of endothelial cells while lipid X did not. Phorbol myristate acetate, a compound known to activate protein kinase C, also caused endothelial cell detachment. Morphologic changes were readily apparent in the endothelial cells after 6 hours of exposure to lipopolysaccharide (1 microgram/ml); at that time many of the cells had contracted and formed bleblike structures on the surface. Large vacuoles, dense bodies, and pyknotic nuclei were found in the detaching cells, indicating necrosis or cell death. Preceding the morphologic changes and actual detachment, endothelial cell DNA and RNA synthesis was impaired by LPS. The changes in DNA and RNA synthesis occurred within 4 hours of exposure to 1 microgram/ml of LPS when the cells were still able to maintain normal levels of ATP. In addition to the inhibition of nucleic acid synthesis, protein synthesis was inhibited after 6 and 8 hours of LPS exposure. DNA, RNA, and protein synthesis returned to control levels after 24 hours of exposure. Investigation on the cultured bovine endothelial cells as a model for LPS action was useful in that these cells are sensitive to relatively low levels of LPS and the endothelium may be an important target in sepsis.

Animals↗
PubMed1988

Ultrastructural and functional effects of lipopolysaccharide and interleukin-2 on human NK cells.

Bacterial endotoxin (lipopolysaccharide, LPS) and interleukin-2 (IL-2) are known to stimulate NK cell mediated cytotoxicity against tumor cells. In the present report we sought to correlate the stimulatory effect of LPS and IL-2 on NK cell activity with ultrastructural changes which occurred as a result of such stimulation. Peripheral blood mononuclear cells (PBMC) were purified from healthy donors by a Ficoll-Hypaque density gradient technique. Leu-11a+ NK cells were isolated by flow microfluorometry using a monoclonal FITC conjugated anti-Leu-11a antibody and a FACS II cell sorter. The PBMC were incubated, respectively, with E. coli LPS or recombinant IL-2 (IL-2) for various time periods. Sorted Leu-11a+ NK cells were incubated with LPS for 24 hours. The NK cytotoxicity in the PBMC and sorted Leu-11a+ cells was assessed by a 51Cr release technique using K562 tumor cells as targets. Leu-11a+ NK cells were identified by immunoelectron microscopy using anti-Leu-11a antibody and labeling with horseradish peroxidase or colloidal gold. Results showed that both LPS and IL-2 significantly enhanced the cytotoxic activity of PBMC. The cytotoxicity of sorted Leu-11a+ cells was augmented by LPS. Recombinant IL-2 induced a significant increase in the number of dense granules, hypertrophy of Golgi apparatus and rough endoplasmic reticulum, and mitosis of Leu-7+ cells and Leu-11a+ cells 4 or 7 days after stimulation. These data indicate that: (1) the effect of LPS on the enhancement of NK cytotoxicity in PBMC may be a direct and/or indirect process involving production of lymphokines; (2) LPS has a direct effect on sorted Leu-11a+ cells; (3) IL-2 stimulates mitosis of Leu-7+ cells and Leu-11a+ cells; and (4) the LPS or IL-2 induced ultrastructural changes in Leu-11a+ cells are consistent with the enhanced NK cytotoxicity.

Cell Separation↗
PubMed1987

Immunoultrastructural studies of human NK cells: II. Effector-target cell binding and phagocytosis.

The binding of NK cells to a target cell appears to be a necessary step for NK cell-mediated cytolysis. In this report, we demonstrated effector-target binding by immunoelectron microscopy by using monoclonal antibodies against NK cells (Leu-7, Leu-11a) and T-cell subsets (Leu-2a/T8, Leu-3a/T4). The surfaces of NK and K562 cells were characterized by antitransferrin receptor antibody and various lectins. In addition, the controversial phagocytic activity of NK cells was studied by incubation of peripheral blood mononuclear cells with opsonized Staphylococcus aureus and labeling with anti-Leu-7 or anti-Leu-11a antibody. Results showed that only Leu-11a+ cells displayed a broad cell-to-cell contact with the target by a shallow intercellular interdigitation of cytoplasmic projections, while Leu-7+, Leu-2a+, or Leu3a+ cells showed only a partial contact with target without interdigitation. The Leu-11a+ cells were frequently observed in small clusters and in close association with monocytes. Cluster formation and association with monocytes were not observed in other NK and T-cell immunophenotypes. In Leu-11a+ cells conjugated with target cells, membrane-bound granules, small vesicles, parallel tubular arrays, Golgi apparatus, endoplasmic reticulum, and small vacuoles were evident and concentrated toward the target. The surface of NK cells was intensely stained for glycoprotein by chromic acid-phosphotungstic acid, whereas target cells were not stained. Transferrin receptors were stained only on the surface of target cells. Only the lectins RCA and UEA labeled the surfaces of both NK and target cells. Phagocytic vacuoles containing cell debris or fragments and ingested bacteria were found in the cytoplasm of Leu-11a+ cells but not in Leu-7+ cells. NK cells were also found within the cytoplasm of K562 target cells. All these findings suggest that Leu-11a+ cells are the true functional NK cells involved in NK cell-mediated cytolysis, phagocytosis, and emperipolesis. Therefore, the NK cell is probably "a phagocyte in lymphocyte's clothing." The presence of peroxidase in the small vesicles of NK cells and endocytotic vesicles of target cells at the effector-target contact area indicates that cytolytic enzymes or factors derived from NK cells may be transported into the target by endocytosis.

Antibodies, Monoclonal↗
PubMed1985

Immunoelectron microscopic identification of human NK cells by FITC-conjugated anti-Leu-11a and biotinylated anti-Leu-7 antibodies.

Human natural killer (NK) cells have been reported to express various surface antigens. The majority and the most functionally potent NK cells are Leu-11a (NKP-15) positive cells. Only a small number of functional NK cells express Leu-7 (HNK-1) antigen. In the present study, we have established techniques for immunoelectron microscopic identification of NK cells by mouse monoclonal FITC-conjugated anti-Leu-11a and biotinylated anti-Leu-7 antibodies. Ficoll-Hypaque-isolated peripheral blood lymphocytes (PBL) were reacted with the specific antibodies before or after fixation in a 1% glutaraldehyde/1% paraformaldehyde fixative. Prefixation labeling of viable cells with the antibodies was carried out at 4 degrees C or 37 degrees C. Cells prelabeled with anti-Leu-11a antibody were reacted with secondary antibodies either before or after fixation. Anti-Leu-7 antibody was stained directly via an avidin-biotin-peroxidase (ABC) system, anti-Leu-11a antibody was stained indirectly by the ABC immunoperoxidase procedure via a biotinylated anti-mouse IgG secondary antibody or by a 10 nm or 40 nm colloidal gold-labeled anti-mouse IgG antibody. Results indicate that Leu-7 antigen could be localized by incubation with the specific antibody either before or after 20 min fixation; however, Leu-11a antigen was totally abrogated following the same fixation procedure. The Leu-11a antigen was well stained by the methods of prefixation labeling of cells with anti-Leu-11a antibody and incubation with a biotinylated secondary antibody and the ABC system after fixation. With respect to colloidal gold labeling, better results were obtained when cells were reacted with the gold-labeled antibodies immediately after incubation with anti-Leu-11a antibody but before fixation. Ultrastructurally both Leu-7 positive (+) and Leu-11a positive (+) cells shared common ultrastructural features associated with large granular lymphocytes. Using the above described techniques, we found approximately 2-5% Leu-7+ and 9-15% Leu-11a+ cells in the PBL of healthy donors. The overall results suggest that Leu-11a antigen is more sensitive to glutaraldehyde/paraformaldehyde fixation than Leu-7, since it can be localized only by prefixation labeling procedures; the ABC immunoperoxidase procedure is an ideal technique for labeling NK cells for light and electron microscopic enumeration; the immunogold method provides an adequate technique for labeling NK cells which are designated for ultracytochemical studies.

Adult↗
PubMed1982

Immunoglobulin determinants on the surface of mouse splenic lymphocytes from Trypanosoma musculi-infected mice.

The splenic B-lymphocyte population of C57BL/6 mice was analyzed by flow microfluorometry to determine the relative density of surface immunoglobulin (sIg) during the course of infection with Trypanosoma musculi and after the injection of T. musculi derivatives. Cells were stained with fluorescent-conjugated antisera directed against 7S mouse immunoglobulins or the major sIg components, IgM and IgD. Evaluation of relative density of sIg was made through observation of histograms plotted by the Fluorescence Activated Cell Sorter (FACS). The FACS also provided the percentage of fluorescence-positive cells detected with each stain. The relative density of sIg was altered in all experimental groups as evidenced by abnormal histograms. These alterations in relative density of sIg persisted only on B cells of the trypanosome-infected and not the T. musculi-derivative-treated animals. The changes in sIg presumably resulted from sIgD modulation, because the histograms resulting from cells stained with anti-IgM remained unchanged. In the trypanosome-infected animals, there also was a reduction in the percentage of sIgD-positive cells near the end of the parasitemia. Observations of spleen sizes showed the same pattern of change with splenomegaly as with relative density of sIg. These data may be the result of the persistence of the parasite in host tissues after clearance of the blood infection.

Animals↗
PubMed1980

Preparation of microbiological specimens for scanning electron microscopy.

This tutorial paper describes techniques for processing microorganisms for scanning electron microscopy (SEM). For ease of discussion, the subject is divided into two sections: A. General Processing Requirements, and B. Specific Processing Techniques. The objective of the first section is to outline processing requirements, explain their purpose, and point out where variations are possible. The following basic steps are discussed: (1) specimen selection, (2) prefixation treatment, (3) chemical fixation, (4) dehydration, (5) critical-point drying and freeze-drying, (6) coating, and (7) viewing. The second section describes methods of manipulating microorganisms through the processing steps. Instructions will cover microorganisms processed for SEM (1) in suspension, (2) in tissues. (3) in tissue culture, and (4) on agar. We are relying heavily on our own experiences in the laboratory and the results are illustrated by use of bacteria, mycoplasmas, rickettsiae, fungi, free-living protozoa, and parasitic protozoa. This tutorial is intended to be a general reference for processing microorganisms for study with the scanning electron microscope. Although we have described basic requirements and several specific techniques, it must be emphasized that there is a wide range of preparation flexibility permissible, depending upon the objectives of the study.

Animals↗