Haematology of swordtail, Xiphophorus helleri. III. Ultrastructural characterization and peroxidase activity localization of blood cells in kidney and spleen of Xiphophorus helleri

The morphology of blood cells in the kidney and spleen of Xiphophorus helleri was characterized by electron microscopy. Erythrocytes, thrombocytes, lymphocytes, granulocytes and monocytes/macrophages were identified. Thrombocytes were elongate and contained bundles of microtubules and a canalicular system. Lymphocytes were heterogeneous in morphology. Granulocytes were divided into two types, based on the light microscopic results, which were extended by the present electron microscopic study. Neutrophils and eosinophils differed in abundance, cell shape, morphology of the nucleus and granula. Neutrophils displayed a spherical to three-lobulated nucleus and different types of granula. Several intermediate forms of granula were observed. The complement of granules displayed a different composition in the cells. These findings suggest a maturation process of a single type of granulum rather than several types. Eosinophils also contain different types of granula, described as G1–G4. The cell shape was variable and nuclei were small and eccentric. Monocytes and macrophages frequently showed autophagocytotic figures. A peroxidase reaction was observed only in the granula of neutrophils.     

American paddlefish leukocytes demonstrate mammalian‐like cytochemical staining characteristics in lymphoid tissues

American paddlefish Polyodon spathula leukocytes demonstrated cytoplasmic staining patterns very similar to mammalian leukocytes when stained with acid phosphatase, α‐naphthyl butyrate esterase and β‐glucuronidase. American paddlefish monocytes, lymphocytes and granulocytes stained positive for acid phosphatase. Monocytes stained positive for α‐naphthyl butyrate esterase. Lymphocytes that stained positive for α‐naphthyl butyrate esterase were designated type A. Lymphocytes that stained positive with antibodies to the L chain of white sturgeon Acipenser transmontanus immunoglobulin (Ig) were designated type B. Type A and type B lymphocytes stained positive for β‐glucuronidase. All leukocytes observed were negative for Sudan Black B. Monocytes, lymphocytes and granulocytes were present in the renal haematopoietic tissue, spleen, thymus, pericardial myeloid tissue, lamina propria of the spiral valve, and in meningeal myeloid tissue located dorsal to the brain, at the base of the brain and around the notochord. Peyer's patches were present in the gut. Morphological characteristics of leukocytes stained with Wright's and haematoxylin and eosin and appeared very similar to those of other fish species.     

Cytochemical identification of the leukocytes of torpedoes (Torpedo marmorata Risso and Torpedo ocellata Rafinisque)

The authors subjected peripheral blood smears of Torpedoes to cytochemical analysis of lipids, protein, neutral and acid polysaccahrides and of some enzymatic activities, i.e. adenosine triphosphatase (ATP-ase), acid and alkaline phosphatase, aliesterase and peroxidase. It was found that neutrophilic granulocytes are intensely PAS and aliesterase positive and weakly ATP-ase positive. Eosinophilic granulocytes show the presence of neutral polysaccharides in the matrix (which is PAS positive) and strong ATP-ase and acid phosphatase activities in the granules. Lymphocytes sometimes contain weakly PAS and aliesterase positive granules. Monocytes show some small PAS positive granules and weak acid phosphatase and aliesterase activities. Thrombocytes contain some peripheral granules which are PAS positive and slightly ATP-ase positive. There are no transitional forms between the various cellular types. The results confirm the classification of leukocytes of Torpedoes into neutrophilic granulocytes, eosinophilic granulocytes, lymphocytes, monocytes and thrombocytes and contribute some informations about the histoenzymatic content of Elasmobranch leukocytes.     

Phagocytosis of bacterial magnetite by leucocytes

Summary Magnetotactic bacteria were introduced into granulocytes and monocytes by phagocytosis. The number of phagocytes containing bacterial magnetites (magneto-sensitive cells) became constant after 1.5 h incubation, and viable phagocytes contained about 20–40 cells of magnetotactic bacteria. Granulocytes and monocytes containing bacterial magnetites were separated by magnet a Samarium-cobalt from lymphocytes. After separation, 89% of lymphocytes were recovered and 95% of the cells were viable. The contamination of phagocytes in the recovered lymphocytes was below 0.8%. Magneto-sensitive granulocytes and monocytes were removed by applying a magnetic field. The nitro-blue tetrazolium-reducing, chemotactic and phagocytic abilities of phagocytes ingesting magnetotactic bacteria were 84%, 88% and 87% respectively after 1 h incubation.     

Identification of peripheral blood leucocytes of the dogfish (Scyliorhinus canicula L.) by electron microscopy

Dogfish peripheral blood leucocytes were examined by electron microscopy after the injection of colloidal carbon. The cells were classified as lymphocytes, plasma cells, monocytes, thrombocytes and granulocytes. The granulocytes were further classified into four types according to the structure of their granules. Monocytes, thrombocytes and two types of the granulocytic cells were phagocytic.     

Modeled microgravity affects motility and cytoskeletal structures

The hypothesis to be tested is that reduced cell-cell interactions between T cells and monocytes are one of the reasons for the observed depression of the "in vitro" activation of human lymphocytes in microgravity. Locomotion is essential for cell-cell contacts. Lymphocytes in suspension are highly motile in microgravity, whereas no data are available so far on the motility of adherent monocytes. It can be argued that an impaired locomotion of monocytes and cytoskeletal changes, both linked to cell contacts, could be responsible for their reduced interaction with T lymphocytes. This study is aimed at revealing how locomotion as well as cytoskeletal structures of adherent monocytes are modified under modeled microgravity conditions using the Random Positioning Machine (RPM, Dutch-Space) as earth based model of spaceflight.     

Ultrastructure of the peripheral blood leucocytes of Oreochromis mossambicus

Peripheral blood leucocytes were examined for their fine structure and their in vivo response to colloidal carbon particles and bacteria, Aeromonas salmonicida . Four main types of leucocyte were identified: lymphocytes, thrombocytes, monocytes and granulocytes. Previously, two types of granulocyte had been identified but at the electron microscope level a third type could be distinguished in the peripheral blood. Monocytes and the type 1 and type 3 granulocytes were found to be phagocytic.     

Granulocyte interactions with GM-CSF and G-CSF secretion by endothelial cells and monocytes

We have, in previous studies, characterized the cytokine and cellular regulation of GM-CSF and G-CSF production by monocytes and endothelial cells. In this study, we investigated the regulatory role of granulocytes. The addition of granulocytes to endotoxin-stimulated monocytes dose-dependently decreased both GM-CSF and G-CSF concentrations, presumably by absorbing the cytokines. A similar dose-dependent decrease in GM-CSF concentration was found when granulocytes were added to IL-1-stimulated endothelial cells. In contrast, G-CSF secretion by endothelial cells responded to granulocytes in a biphasic fashion. At low granulocyte concentrations, endothelial cells responded with an increased G-CSF secretion, but at high concentrations of granulocytes G-CSF secretion was down modulated. Our results suggest that there exist two loops between granulocytes and endothelial cells for regulating G-CSF activity. Granulocytes can stimulate G-CSF secretion by activated endothelial cells but can also decrease the biological activity by absorbing the cytokine. These mechanisms might be involved in the regulation of the local and systemic levels of granulocytes.     

Decreased chemotaxis of human peripheral phagocytes exposed to a strong static magnetic field

The chemotaxis of human peripheral phagocytes, neutrophils and monocytes was examined in a strong static magnetic field (0.317+/-0.012 Tesla). The chemotaxis of the suspension of purified neutrophils and monocytes was tested in the Boyden chamber using C5a as a chemotactic signal. The chambers were placed into a temperature regulated (36.6 degrees C) equipment producing a strong static magnetic field (0.317 Tesla) for 60 minutes. The movement of cells proceeded into a nitrocellulose membrane toward the north-pole of the magnet, i.e. in the direction of the Earth's gravitational pull. The C5a induced chemotaxis of human neutrophils decreased significantly in the strong static magnetic field. Monocytes were not significantly effected. The strong static magnetic field decreased the chemotactic movement of neutrophils and this phenomenon may have implications when humans are exposed to magnetic resonance imaging for extended periods of time.     

Facts and speculation about necrotaxis (chemotaxis toward a dying cell)

The name "necrotaxis" has been given to a special type of chemotaxis in which granulocytes and monocytes are attracted to cells in the process of dying. Microirradiation devices (conventional UV and laser) have been used to destroy a target cell. Immediately, leukocytes (neutrophils and monocytes) can be seen to advance towards the damaged cell and to engulf it. Hemoglobin and albumin coagulated in the microbeam are equally necrotactic. The presence of plasma does not seem necessary, as the phenomenon also occurs in a saline environment. The neutrophil nucleus is not necessary for necrotaxis, anucleated cytoplasmic fragments are capable of directionality, rosette formation around the dead cell and phagocytosis. Three specific points are briefly discussed: -- the nature of necrotactic substances -- the recognition mechanism of damaged cells and molecules -- the eventual role of necrotaxis in vivo.     

Eosinophilic granulocytes in the epidermis of Oreochromis mossambicus gill filaments studied in situ

Granular cells were observed in the epidermis of isolated gill filaments using Normaski microscopy at×400 magnification. Granulocytes were motile and chemotactic to histamine (introduced via a micropipette into the filament) and to the products of degranulation of neighbouring granulocytes within the epidermis. The abundance of visible granulocytes within the epidermis of gill filaments was increased in the period following net capture, confirming their migratory ability. Cells isolated from gill tissue are poorly glass adherent. They are also eosinophilic and more dense than macrophages and lymphocytes on a discontinuous percoll gradient. They are identified as eosinophils on the basis of these characteristics.     

Determination of carrier status for the Wiskott-Aldrich syndrome by flow cytometric analysis of Wiskott-Aldrich syndrome protein expression in peripheral blood mononuclear cells

The Wiskott-Aldrich syndrome (WAS) is caused by defects in the WAS protein (WASP) gene on the X chromosome. Previous study disclosed that flow cytometric analysis of intracellular WASP expression (FCM-WASP analysis) in lymphocytes was useful for the diagnosis of WAS patients. Lymphocytes from all WAS patients showed WASPdim instead of WASPbright. Here we report that FCM-WASP analysis in monocytes could be a useful tool for the WAS carrier diagnosis. Monocytes from all nine WAS carriers showed varied population of WASPdim together with WASPbright. None of control individuals possessed the WASPdim population. In contrast, lymphocytes from all the carriers except two lacked the WASPdim population. The difference of the WASPdim population in monocytes and lymphocytes observed in WAS carriers suggests that WASP plays a more critical role in the development of lymphocytes than in that of monocytes. The present studies suggest that a skewed X-chromosomal inactivation pattern observed in WAS carrier peripheral blood cells is not fixed at the hemopoietic stem cell level but progresses after the lineage commitment.     

A cytochemical and light microscopical study of the peripheral blood leucocytes of Oreochromis mossambicus, Cichlidae

The morphological characteristics of the peripheral leucocytes of the cichlid O. mossambicus were studied using light microscopy. Cytochemical procedures and phagocytic studies were employed. Four main types of leucocytes could be recognized: lymphocytes, thrombocytes, monocytes and granulocytes. Two distinct types of granulocyte were identified. Monocytes and one type of granulocyte were found to be phagocytic.     

Separation of leucocytes in the dogfish (Scyliorhinus canicula) using density gradient centrifugation and differential adhesion to glass coverslips

Summary The blood of the dogfish, S. canicula, contains several types of leucocytes, namely thrombocytes, monocytes, lymphocytes and four populations of granulocytes. Three of these granulocyte types, G1, G3 and G4, are eosinophilic while G2 is heterophilic/neutrophilic. All of the leucocyte types, with the exception of G2 granulocytes and monocytes, can be separated by means of their differential adherent properties to glass and by density gradient centrifugation. Thrombocytes, G3 and G4 granulocytes can be separated in good purity by single-step methods while G1 granulocytes and lymphocytes require a combination of density gradient centrifugation followed by adherence to glass to remove contaminating thrombocytes. Depending on the cell type, between 11–45% of cells with consistently high viability can be recovered after separation. Separated populations of the thrombocytes and granulocytes will be especially useful for studies on the role of such cell types in inflammation.     

Four-parameter white blood cell differential counting based on light scattering measurements

Measurement of the depolarized orthogonal light scattering in flow cytometry enables one to discriminate human eosinophilic granulocytes from neutrophilic granulocytes. We use this method to perform a four-parameter differential white blood cell analysis. A simple flow cytometer was built equipped with a 5-mW helium neon laser that measures simultaneously four light scattering parameters. Lymphocytes, monocytes, and granulocytes were identified by simultaneously measuring the light scattering intensity at angles between 1.0 degrees and 2.6 degrees and angles between 3.0 degrees and 11.0 degrees. Eosinophilic granulocytes were distinguished from neutrophilic granulocytes by simultaneous measurement of the orthogonal and depolarized orthogonal light scattering. Comparison of a white blood cell differentiation of 45 donors obtained by the Technicon H-6000 and our instrument revealed good correlations. The correlation coefficients (r2) found were: 0.99 for lymphocytes, 0.76 for monocytes, 0.99 for neutrophilic granulocytes, and 0.98 for eosinophilic granulocytes. The results demonstrate that reliable white blood cell differentiation of the four most clinically relevant leukocytes can be obtained by measurement of light scattering properties of unstained leukocytes.     

Countercurrent distribution of lymphocytes from human peripheral blood in an aqueous two-phase system. I. Separation into subsets of lymphocytes bearing distinctive markers

Lymphocytes from human peripheral blood have been separated by countercurrent distribution in a charged aqueous two-phase system composed of Dextran T 500 and polyethylene glycol 6000 with a cell yield of 59–88% and viability above 90%. A highly reproducible partition pattern was seen with four distinct peaks. Lymphocytes with surface membrane immunoglobulin (SmIg) were located in the first part of the distribution corresponding mainly to peak I. T lymphocytes as detected by E rosetting and α-naphthyl acetate esterase (ANAE) staining showed a broad distribution with a maximum in peaks II and III. ANAE-negative lymphocytes were seen in both extremes of the distribution, corresponding to B cells in the first part and to a population of E^? and SmIg^? lymphocytes in the last part. Monocytes were present in all fractions with some enrichment in peaks II–IV. Lymphocytes with low-affinity Fc receptors were found in B-cell-containing fractions in the first part of the distribution, but also in the last part. Lymphocytes with high-affinity Fc receptors were detected mainly in peak IV. It is thus demonstrated that peripheral blood lymphocytes can be fractionated into subpopulations enriched in cells with characteristic markers.     

Cholesterol biosynthesis in human lymphocytes,monocytes, and granulocytes

Lymphocytes, monocytes and granulocytes were separated by counter-flow centrifugation from the blood of normal individuals and were incubated in full serum medium or lipid-depleted medium. The monocytes incorporated about five times more [2-¹⁴C]acetate into sterols than did the lymphocytes in full serum medium and approximately twenty times more than the lymphocytes in lipid-depleted medium. The granulocytes were unable to synthesize sterols from either [2-¹⁴C]acetate or [2-¹⁴C]mevalonate, but they were able to use these substrates for the synthesis of squalene and demonstrated approximately a two fold increase in the incorporation of [2-¹⁴C]acetate (but not [2-¹⁴C]mevalonate) into squalene when incubated in the lipid-depleted medium as compared to the full serum medium.     

Species differences in the expression of carbohydrate differentiation antigens on mammalian blood cells revealed by immunofluorescence with monoclonal antibodies

Following recent observations using monoclonal antibodies that carbohydrate structures behave as differentiation antigens of man and mouse, we have made a preliminary survey of the expression of 8 monoclonal antibody-defined carbohydrate antigens on blood cell smears of man, baboon, mouse, rat, rabbit, pig, and dog. There are considerable species differ-ences in the patterns of antigen expression. However, certain generalizations can be made as follows: the i and I antigens, associated with linear and branched carbohydrate chains consisting of repeating N-acetyl-lactosamine sequences (GalB1-4GIcNAc, termed Type-2 backbone sequences) are widely distributed among granulocytes and lymphocytes of all the species studied, and on erythrocytes, monocytes, and piateiets of some of them. Substantial amounts of Type-1 backbone sequences (GalB1-3GlcNAc) may occur on rabbit lymphocytes. The N-acetylneuraminic acid-containing antigens, Pr 2 and Gd, are also expressed to varying degrees on blood cells. On the other hand, antigens based on mono- and diiucosylated N-acetyllactosamine, termed SSEA-1 (or X-hapten) and C14 (or Y-hapten) are predominantly granulocyte/monocyte-associated antigens. The former antigen is expressed in overt form only on untreated human granulocytes but occurs in cryptic state , masked by sialic acid, on human monocytes, and on the granulocytes and monocytes of baboon, rabbit, and dog but not on those of mouse, rat, and pig. The latter antigen is expressed on human granulocytes and on neuraminidase-treated monocytes and granulocytes oi dog. Lymphocytes of dog are unusual in their expression of C14 antigen, in cryptic state, masked by sialic acid residues. Although the physiological roles of these various carbohydrate structures , in vivo, are not yet known, they seem excellent candidates as determinants of species and cell-type differences in susceptibilities to infective agents.     

Immunologic studies with LFA-1- and Mo1-deficient lymphocytes from a patient with recurrent bacterial infections

A patient and his parents, deficient for lymphocyte function associated antigen-1 (LFA-1) and Mo1 (OKM1), were studied with respect to leukocyte surface marker expression and functional properties. The patient had a history of severe recurrent bacterial infections. Two siblings had already died of bacterial infections. The patient's granulocytes, monocytes, and lymphocytes expressed low but detectable amounts (less than or equal to 10%) of LFA-1 and Mo1. Intracellularly, LFA-1 and Mo1 (OKM1) were detectable and LFA-1 expression was enhanced on patient T cells stimulated with phytohemagglutinin. Granulocytes and monocytes of both the patient's parents expressed markedly decreased amounts of LFA-1 and Mo1. Lymphocytes of the mother expressed 40 to 60% of the amount of LFA-1 expressed on control lymphocytes, but his father's lymphocytes showed a normal LFA-1 expression. Granulocytes of the patient and of his deceased sister showed normal phagocytosis, but they had a dysfunction in the activation of the oxidative metabolism. Functional activities mediated by patient T cells were all normal. Moreover, all lymphocyte functions, including killer (K), natural killer (NK), cytotoxic T cell activity, helper activity for in vitro immunoglobulin (Ig) production by normal B cells, and PHA-induced proliferation were inhibitable by anti-LFA-1 monoclonal antibodies. K and NK activity mediated by patient leukocytes was 100-fold more sensitive to the inhibiting effect of anti-LFA-1 antibody than K and NK activity of normal donor leukocytes. Thus, although the amount of LFA-1 expressed was strongly reduced, it was still sufficient and required for the functional activity exhibited by patient T cells. The major functional defect observed with leukocytes of the patient and his father was an apparent B cell defect. B cells of the father and of the patient failed to produce Ig in the pokeweed mitogen (PWM)-driven system. The B cells of patient and of his father only produced Ig when cultured with T cells of the father, and not with normal donor T cells or T cells of the mother, in the presence of exogenous interleukin 2 (IL 2). In addition, the father's B cells produced Ig when cocultivated with patient T cells in the IL 2-driven system. This restriction of helper T cell activity is noteworthy because PWM- and IL 2-driven Ig synthesis by normal lymphocytes show no histocompatibility requirements between cooperating T and non-T cell populations.(ABSTRACT TRUNCATED AT 400 WORDS)     

Beta-endorphin in granulocytes

The literature indicates that beta-endorphin can be found in the mononuclear cells of peripheral blood. In the present experiments the endorphin content of granulocytes was studied, compared to lymphocytes as reference cells. Granulocytes as well, as lymphocytes contain endorphin. The granulocytes' endorphin content is much higher. Both lymphocytes and granulocytes are also able to take up endorphin from the milieu.