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Biomedical subjects

L P Bignold

Publications and source records attributed to L P Bignold.

At least 37 records · Page 2Linked to original sources
PubMed1992

Interleukin-8 and neutrophil leucocytes: adhesion, spreading, polarisation, random motility, chemotaxis and deactivation in assays using 'sparse-pore' polycarbonate (nuclepore) membranes in the Boyden chamber.

Adhesion, spreading, chemotaxis and deactivation of chemotactic responses of separated human peripheral blood neutrophil leucocytes under the influence of interleukin-8 (IL-8) were tested using a previously described Boyden chamber-type method involving 'sparse-pore' polycarbonate (Nuclepore) filtration membrane. The random motilities of neutrophils in similar concentrations of IL-8 were tested using corresponding chambers with polycarbonate membranes of standard pore densities. In addition, polarisation of neutrophils in suspension in various concentrations of IL-8, and the possibility of deactivation of this polarisation response by IL-8 itself or by N-formyl-methionyl-leucyl-phenylalanine (FMLP) were examined. Neutrophils exhibited maximum chemotaxis to IL-8 in a concentration of 100 ng/ml, and to a degree which was similar to the maximum response to FMLP (10(-7) M). This chemotactic response to IL-8 was markedly reduced by pretreatment of the cells with either 100 ng/ml IL-8 (deactivation) or 10(-7) M FMLP (cross-deactivation). On the other hand, the chemotactic response of the neutrophils to FMLP was reduced by pretreatment with FMLP but was not deactivated by pretreatment with IL-8 (i.e. deactivation but not cross-deactivation). Neutrophils in suspension were maximally polarised by 100 ng/ml IL-8, and to lesser degrees by 1, 10 and 1,000 ng/ml IL-8. The detailed morphology of the polar cells was not distinguishable from that induced by 10(-8) M FMLP. Pretreatment of the cells with either IL-8 or FMLP resulted in no reduction of polarisation in response to subsequent exposure to either agent (i.e. neither deactivation nor cross-deactivation).(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Adhesion↗
PubMed1991

Inhibition of chemotaxis of neutrophil leukocytes to interleukin-8 by endotoxins of various bacteria.

The effects of endotoxins from various bacteria (Escherichia coli, Klebsiella pneumoniae, Vibrio cholerae, Shigella flexneri, Salmonella typhosa, and Pseudomonas aeruginosa) on chemotaxis of neutrophil leukocytes to formyl peptide and interleukin-8 were tested in an improved chemotaxis assay involving a "sparse-pore" polycarbonate (Nuclepore) membrane in a Boyden-type chamber. The possible chemotactic activity of the endotoxins themselves were tested by the same technique. In addition, the effects of these substances on random motility of neutrophils were tested with a corresponding assay involving similar chambers fitted with membranes of standard pore density. Possible activation of the complement system of serum by each endotoxin was tested with sheep erythrocyte assays and the maximum endotoxin concentration (100 micrograms/ml) used in the chemotaxis and motility assays. All endotoxins inhibited chemotaxis of neutrophils to interleukin-8. No endotoxin affected chemotaxis to formyl peptide or was itself chemotactic for neutrophils. Endotoxin of S. flexneri inhibited random motility of neutrophils, while the others had no such effect. Endotoxins of K. pneumoniae and of P. aeruginosa produced moderate and marked inhibition, respectively, of total complement, as measured by hemolysis of sheep erythrocytes, without affecting the levels of C3c and C4 in these assays. Endotoxins of the other bacteria had no demonstrable effect in any of these assays of complement activation. These results suggest that chemotaxis to interleukin-8 may be mediated by cellular mechanisms different from those involved in chemotaxis to formyl peptide. Furthermore, the presence of these endotoxins could be significant for the suppression of neutrophil accumulation in inflammatory lesions mediated by interleukin-8.

Bacterial Toxins↗
PubMed1990

Importance of extracellular divalent cations to polarisation of polymorphonuclear leukocytes induced by plasma.

The roles of extracellular calcium and magnesium ions in the polarisation of human peripheral blood polymorphonuclear leukocytes (PMN) induced by autologous fresh heparinised plasma were investigated by studying the effects of 5 mM chelators of divalent cations [ethylenediamine tetra-acetic acid (EDTA), ethylenebis-(oxyethylene-nitrilo)-tetra-acetic acid (EGTA) or disodium hydrogen citrate]. In addition, the effects of a blocker of membrane calcium channels (verapamil) were studied. Polarisation of PMN suspended in plasma (84.1 +/- 11.9%) was reduced by each chelating agent over 30 min (to 20.0 +/- 15.6% by EDTA, to 42.5 +/- 19.3% by EGTA and to 29.4 +/- 22.9% by citrate). Polarisation of PMN suspended in plasma treated with EDTA or EGTA was restored by inclusion of equimolar additional Ca2+ ions, and in plasma treated with EDTA, EGTA or citrate, by equimolar additional Mg2+ ions. Additional Mg2+ had no effect on the spherical shape of PMN in Hanks' solution and additional cations had no effects on the polarisation of PMN induced by fMLP. Cells rendered spherical by each chelating agent in plasma for 30 min retained their ability to polarise on addition of fMLP to the plasma-chelator medium. Verapamil (10(-4) M) markedly reduced polarisation in plasma (to 52 +/- 11.3%) but the same drug (10(-5) M) had no such effect. In contrast to the polarisation of cells in plasma, the polarisation response of PMN to N-formyl-methionyl-leucyl-phenylalanine (fMLP, 10(-8) M) in buffered Hanks' solution was not affected by any of the chelating agents or by verapamil, even in high concentration. These results indicate that extracellular divalent cations are necessary for the polarisation of PMN suspended in autologous plasma and that the mechanism of polarisation of PMN in plasma may be different to that of polarisation induced by fMLP.

Blood Physiological Phenomena↗
PubMed1990

Studies of chemotactic, chemotactic movement-inhibiting and random movement-inhibiting effects of interleukin-1 alpha and beta, tumour necrosis factors alpha and beta and interferon gamma on human neutrophils in assays using 'sparse-pore' polycarbonate (Nuclepore) membranes in the Boyden chamber.

Interleukin-1 alpha and beta (IL-1 alpha and beta), tumour necrosis factors alpha and beta (TNF alpha and beta) and interferon gamma (IFN gamma) were tested for their chemotactic effects, their effects on chemotactic movement towards N-formyl-methionyl-leucyl-phenylalanine (FMLP) and their effects on random locomotion of human peripheral blood neutrophils through polycarbonate membranes in Boyden-type chambers. Both IL-1 alpha and beta, but no other cytokine tested were chemotactic for neutrophils using 'sparse-pore' polycarbonate membrane. Both TNFs, but no other cytokine, inhibited neutrophil chemotactic movement towards FMLP using the same membrane. No cytokine influenced random migration of neutrophils through polycarbonate membrane of standard pore density. These results suggest that IL-1 may have a role as a chemotactic mediator of inflammation, but that TNFs may inhibit chemotactic migration of neutrophils into inflammatory lesions.

Biological Factors↗
PubMed1990

Effects of plasma proteins on the adhesion, spreading, polarization in suspension, random motility and chemotaxis of neutrophil leukocytes on polycarbonate (Nuclepore) filtration membrane.

The effects on adhesion, spreading, polarization in suspension, random motility and chemotaxis to N-formyl-methionyl-leucyl-phenyl-alanine (FMLP) of neutrophil leukocytes on polycarbonate (Nuclepore) membrane in the Boyden chamber of purified serum albumin, gamma globulin and fibrinogen, as well as of the Hanks'-soluble components of Cohn fractions III (beta and gamma globulins) and IV (albumin, alpha globulins and transferrin) of plasma proteins were tested in comparison to the corresponding effects of dilute fresh heparinized plasma. Serum albumin, fibrinogen and fibronectin inhibited adhesion of neutrophils, so that the spreading, random motility and chemotaxis of these cells could not be assessed in the presence of these proteins alone. These plasma proteins induced little or no polarization of the cells in suspension. Of the remaining preparations, gamma globulin promoted adhesion and excessive spreading and caused corresponding reduction of random motility and chemotaxis to FMLP of these cells. This plasma protein, even in low concentration, induced marked polarization of neutrophils in suspension. Both Cohn fractions III and IV supported adhesion and permitted random motility and chemotaxis to FMLP approximating that which occurs in dilute plasma. These fractions induced moderate polarization of cells in suspension. Addition of serum albumin or fibronection to the preparations of gamma globulin, or Cohn fractions III or IV did not alter the adhesion, random motility or chemotaxis of the neutrophils in these preparations. These results suggest that alpha and beta globulins, as well as gamma globulins are factors in plasma affecting adhesiveness and polarization of neutrophils and which provide for optimal motility and chemotaxis of these cells on solid substrata.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood Proteins↗
PubMed1989

Use of sparse-pore polycarbonate (Nuclepore) membrane for the measurement of chemotaxis of polymorphonuclear leukocytes. Comparison of 'micro-Boyden' chambers and estimation of 'drop-off' of cells.

Various modified Boyden chambers having upper and lower compartments of small volume ('micro-Boyden' chambers) were tested for their suitability for use with the recently described 'sparse-pore' polycarbonate (Nuclepore) membrane for the measurement of chemotaxis of polymorphonuclear leukocytes in vitro towards N-formyl-methionyl-leucyl-phenylalanine. Previously reported 'open-well' and 'blind-well' chambers, as well as a new chamber having a lower compartment made from perspex and an upper compartment made from silicone rubber were tested. The most satisfactory results were provided by the new chamber, and this is attributed to the easy, accurate filling of the lower compartment, the lack of distortion of membrane, and the reliability of the seals around the edges of the membrane which can be achieved with this chamber. Full humidification during assembly was necessary for obtaining maximal response of the cells through the 'sparse-pore' membrane in all types of micro-Boyden chamber. 'Drop-off' of polymorphonuclear leukocytes from the lower surface of the 'sparse-pore' membrane during incubation was also studied in these experiments and found to be slight.

Chemotaxis, Leukocyte↗
PubMed1989

Proteoglycans synthesized by human polymorphonuclear leucocytes in vitro.

Polymorphonuclear leucocytes (PMN) were assessed in vitro for their ability to synthesize and secrete proteoglycans. The PMN were isolated from human peripheral blood and were found to contain less than 5% mononuclear cells. Following 24 h incubation in the presence of (35S)-sulfate, significant quantities of 35S-labelled macromolecules were detected both within the culture medium and cells. Although the PMN preparations contained some platelets (approximately five platelets:one PMN), culture of platelets alone did not result in the detection of any 35S-labelled macromolecules in either the medium or platelets. 35S/3H-labelled macromolecules from the PMN cultures were identified as proteoglycans on the basis of their degradation by papain, alkaline sodium borohydride, chondroitinase ACII, chondroitinase ABC and nitrous acid. The labelled proteoglycans isolated from the medium and cells eluted from Sepharose CL-4B with a Kav of 0.63; this indicated a small size compared with many other proteoglycans. The glycosaminoglycans associated with the proteoglycans were identified as heparan sulfate, chondroitin sulfate and dermatan sulfate, with chondroitin sulfate being the principal component. The average molecular weight of the glycosaminoglycans was determined to be 16,000. Therefore, the data from this study demonstrate the ability of human PMN to synthesize and secrete proteoglycans in vitro which appear to differ from those synthesized by mesenchymal cells with respect to molecular size and glycosaminoglycan composition.

Chondroitin Sulfates↗
PubMed1989

Cytokines and inflammation: modified "Miles-Wilhelm" criteria for assessing the likely roles of these substances in vivo.

Cytokines (including lymphokines, interleukins, tumour necrosis factors, interferons and hemopoietic colony-stimulating factors) are a loose group of endogenous proteins which are presently being investigated for an increasingly wide range of bioactivities. Recently, cytokines have been suggested to mediate various aspects of inflammation, but the data on which these suggestions are based are often fragmentary and derive from limited experimental in vitro and in-vivo models of inflammation. Therefore, the roles of cytokines in the mediation of naturally occurring inflammatory lesions in man and animals remain unclear. This article traces the development of notions of endogenous mediators of inflammation over the last hundred years and reviews previously published ways of assessing the relevance of experimental data concerning mediators of inflammation to naturally occurring inflammatory lesions. Emphasis is given to the "criteria" advanced for this purpose by Miles and Wilhelm in the 1950s and 1960s, and additional criteria appropriate to the assessment of data concerning cytokines as mediators of inflammation are discussed.

Animals↗
PubMed1988

Kinetics of chemo-attraction of polymorphonuclear leukocytes towards N-formyl peptide studied with a novel polycarbonate (Nucleopore) membrane in the Boyden chamber.

The motile responses of human polymorphonuclear leukocytes (PMN) to N-formyl-methionyl-leucyl-phenylalanine (FMLP) in the Boyden chamber using a new sparse-pore polycarbonate membrane (pores 3 micron in diameter and occupying 0.1% of surface area) were compared with those demonstrated by using a standard polycarbonate (Nuclepore) filtration membrane (pores 3 micron in diameter and occupying 5% of surface area). Motility of PMN in gradients of FMLP using the new membrane was not influenced by chemokinetic effects of the factor, and the 'background' migration of the cells was minimal. However, motility of PMN in gradients of FMLP using the standard membrane was found to be influenced by chemokinetic effects of the chemotactic factor, and the 'background' or 'control' migration (in the absence of chemotactic factor) of the cells was substantial. Greater directional migration of PMN according to steepness of the gradient of chemotactic factor was demonstrated with the use of the new membrane. The new membrane may be of considerable value in the further study of the chemotactic responses of PMN.

Chemotaxis, Leukocyte↗
PubMed1988

Generalised angiosarcoma of the heart.

A primary angiosarcoma, which formed multiple ventricular pericardial and myocardial nodules without atrial involvement, occurred in the heart of an 80-year old Caucasian female and caused death within three weeks of the onset of symptoms. There were neither cutaneous lesions to suggest Kaposi's sarcoma nor serological evidence of infection by Human T-cell Leukemia Virus and the rapidly fatal course of the patient's illness was similar to that of the typical atrial cardiac angiosarcoma. The distinction between primary cardiac angiosarcoma and Kaposi's sarcoma affecting the heart is discussed.

Aged↗
PubMed1988

Effects of cytochalasin B, N-formyl peptide and plasma on polarisation, zeiosis (blebbing) and degranulation of polymorphonuclear leukocytes in suspension.

The effects were studied of cytochalasin B and N-formyl peptide (FMLP) in various concentrations on the morphology and degranulation (release of the granule contents lysozyme and beta-glucuronidase) of polymorphonuclear leukocytes (PMN) suspended in either Hanks' solution or 100% fresh heparinised plasma. PMN in low concentrations of FMLP in Hanks' solution or in plasma alone showed "long" polarisation and did not degranulate. Cytochalasin B caused the PMN in low concentrations of FMLP or in plasma to become spherical, but no degranulation of the cells occurred. High concentrations of FMLP in Hanks' solution induced "short" polarisation of PMN with slight degranulation of the cells. Cytochalasin B together with high concentrations of FMLP in Hanks' solution induced zeiosis ("blebbing") and marked degranulation of the cells. However, cytochalasin B and high concentrations of FMLP in plasma caused PMN to exhibit "short" polarised morphology and markedly degranulate. These results suggest that degranulation of PMN can be associated with either the "short" polarised shape of the cells or zeiosis, but not the "long" polarised form. Furthermore, the results indicate that plasma, although capable of causing "long" polarisation of the cells, inhibits zeiosis without affecting the degranulation of the cells induced by cytochalasin B.

Blood↗