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Biomedical subjects

H C Stevenson

Publications and source records attributed to H C Stevenson.

86 records · Page 5Linked to original sources
PubMed1983

Monoclonal antibody to human eosinophils recognizing 95 kD surface membrane antigen.

Mice were immunized with purified eosinophils obtained from patients with the idiopathic hypereosinophilic syndrome. A hybridoma initially producing an IgM antibody which switched to an IgG1 antibody was selected for cloning and further testing. This IgG1 antibody reacted with human eosinophils, granulocytes, monocytes and large granular lymphocytes, but did not react with T lymphocytes, B lymphocytes, platelets, erythrocytes, or a panel of human leukemia cells and cell lines. Bone marrow analysis revealed staining of myeloid precursor cells but not erythroid precursors or plasma cells. This IgG1 antibody had no effect on aggregation of granulocytes, lysozyme release, superoxide production, chemotaxis, or killing activity; however, there was some stimulation of beta-glucuronidase secretion. While the antibody did not augment the killing of Staphylococcus aureus by granulocytes, the antibody itself was bactericidal. By immunoprecipitation of granulocytes, eosinophils and monocytes, a molecule with a molecular weight of 95 kD was identified.

Animals↗
PubMed1983

Spontaneous cytotoxicity by monocyte-enriched subpopulations of human peripheral blood mononuclear cells against human or mouse anchorage-dependent tumour cell lines. Contribution of NK-like cells.

Human peripheral blood mononuclear cells (PBMNC) were found to be cytotoxic for mouse or human anchorage-dependent target cell lines in a 48-72 h [125I]iododeoxyuridine (IUDR) release assay. Unfractionated, adherent or nonadherent cells had significant levels of cytotoxicity, as did cells fractionated according to size into 'lymphocytes' or 'monocytes' by elutriation. Intermediate size cells, not enriched for monocytes, had high levels of cytotoxicity. In all fractions tested, including adherent populations, some cells with the morphology of large granular cells were observed. Treatment of all fractions with interferon (IFLrA, a purified, recombinant alpha-IFN) boosted cytotoxicity against four target cells lines. Treatment with lymphokines containing putative 'macrophage-activating factor' (MAF) also enhanced cytotoxicity in fractions depleted of monocytes. Culture in fetal bovine serum enhanced cytotoxicity mainly in unfractionated and nonadherent PBMNC. These experiments indicated that NK-like cells can be appreciable contaminants in elutriator-purified monocyte-enriched or adherent cell populations and thereby contribute to observed cytotoxicity, particularly after pretreatment with IFN or other stimulatory factors.

Animals↗
PubMed1983

Characterization by monoclonal antibodies of the cytotoxic effector cells in human peripheral blood mononuclear cells reactive against anchorage-dependent tumour cell lines.

The effector cells for spontaneous cytotoxicity against anchorage-dependent human or mouse tumour cell lines in a 72-h iododeoxyuridine-release assay by normal human peripheral blood cells (PBMNC) or monocyte-enriched fractions were analysed by the use of monoclonal antibodies. PBMNC or adherent or elutriated monocyte-enriched populations of PBMNC were depleted of monoclonal antibody-reactive cells by complement-dependent lysis or separated into monoclonal-antibody-positive or -negative subsets by an indirect rosetting technique followed by Ficoll-Hypaque density gradient separation. The experimental data indicated that in both PBMNC and monocyte-enriched populations, an appreciable proportion of the effector cells with cytolytic activity against adherent human or mouse tumour target cells were positive with B73.1.1 (an antibody with a high degree of selectivity for natural killer (NK) cells), B43.4.1 (or OKM1), and with OKT11a (an antibody recognizing the receptors for sheep erythrocytes), and had the morphology of large granular cells, which have previously been shown to mediate NK activity. These effector cells were mostly negative for BRL.1, BRL.2, B52.1.1, B44.1.1, B13.4.1 and DR antigens, unlike classical monocytes. Some cells which are cytotoxic for the adherent mouse, SV-40-transformed kidney tumour line, TU-5, may bear B52.1.1 or other monocyte-like antigens. Taken together, these results indicate that, in monocyte-enriched populations, both NK cells and monocytes have cytotoxic effector activity against various human and mouse adherent target cell lines.

Animals↗
PubMed1983

Characterization of a human blood monocyte subset with low peroxidase activity.

Two human monocyte subsets from the peripheral blood of healthy donors have been isolated in greater than 90% purity by countercurrent centrifugal elutration and human serum albumin gradients and their functional capabilities have been assessed. We have demonstrated that one subset ("regular" monocytes, RM) showed intense cytoplasmic peroxidase staining and contained substantial peroxidase activity. In contrast, another subset ("intermediate" monocytes, IM) stained poorly for peroxidase and had low peroxidase activity. By electron microscopic analysis combined with peroxidase localization, it was found that IM had fewer peroxidase-positive granules per cell than did RM. IM coelutriated with some lymphocytes and by cell sizing analysis were shown to be slightly smaller than RM. Functional and cytochemical analysis of these subsets indicated that IM had less activity than RM in assays such as accessory cell function for mitogen-induced T lymphocyte proliferation and antibody-dependent cellular cytotoxicity, and that fewer IM expressed OKM1 antigen and pokeweed mitogen (PWM) receptors on their membranes than did RM. The subset of IM not bearing either the PWM receptor or the OKM1 antigen had very low peroxidase activity. IM also were found to have a greater sensitivity to polyriboinosinic and polyribocytidilic acid (100 micrograms/ml)-induced secretion of interferon. There was no significant difference in the phagocytic capability, the percentage of Fc receptor-positive cells, 5'-nucleotidase activity, DR antigen expression, or the responsiveness to migration inhibitory factor of IM as compared with RM. Furthermore, it was found that the ratio of IM to RM increased after prolonged cytapheresis, which suggests that IM are more mobilizable than RM from the extravascular reservoirs of human monocytes.

Adult↗
PubMed1983

Prostaglandin E synthesis and release by murine macrophages and human monocytes after in vitro treatment with biological response modifiers.

Prostaglandins of the E series (PGE), produced by mononuclear phagocytes, have many biological activities and are involved in the regulation of myelopoiesis and of the cytotoxic activities of macrophage (M phi) and natural killer (NK) cells. Since one of the possible effects of biological response modifiers (BRMs) on immune regulation might be modulation of PGE production, resident peritoneal murine M phi (mM phi) and elutriated human blood monocytes (hM) were incubated with BRMs and the supernatants were then assayed for PGE. Control mM phi produced about 4.9 ng PGE/10(5) mM phi over a 24 hr period. Lipopolysaccharide (LPS, 1-5 micrograms/ml), polyinosinic-polycytidylic acid complexed with poly-L-lysine (Poly ICLC, 0.1-10 micrograms/ml) and murine alpha, beta-interferon, (IFN, 10-1000 U/ml) all caused a highly significant increase in PGE-secretion. BM41.332, a 2-cyanoaziridine (25-50 micrograms/ml), was found to be less stimulatory, whereas the pyran copolymer MVE-2 (25-50 micrograms/ml), and Azimexone (25-50 micrograms/ml) had marginal effects on PGE-production. Kinetic studies showed that the plateau of PGE-production by mM phi occurred during the first 24 hr of incubation, and mM phi which were washed after a 24 hr incubation period could not be restimulated to produce more PGE. PGE release by hM was increased after stimulation with LPS, Poly IGLC and BM41.332, whereas human IFNs (alpha and beta) induced a slight decrease in PGE production. As in the murine studies, Azimexone and MVE-2 had no detectable effect. The ability of some BRMs to augment PGE-secretion by mM phi and hM may contribute to their negative effects on host responses, such as suppression of NK cell activity.

Adjuvants, Immunologic↗
PubMed1983

Activation of human monocyte cytotoxicity by natural and recombinant immune interferon.

Human T cell hybridomas were established by fusion of SH9 cells, the 6-thioguanine-resistant mutant line of human T lymphoma Hut 102-B2, with concanavalin A-stimulated human peripheral blood lymphocytes. Hybridoma line L38 produced a macrophage activating factor (MAF) with the ability to activate human peripheral blood monocytes to show enhanced cytotoxicity against human colon adenocarcinoma HT-29 cells in a 72-hr 125iododeoxyuridine-release assay. The L38 line was then cloned by the limiting dilution technique and two sublines, L38B and L38D, were found to produce high levels of MAF constitutively. Interferon activity was also detected in L38B and L38D supernatants. When interferon activity was neutralized with specific antiserum to purified human immune interferon (IFN-gamma), MAF activity was abrogated. To confirm that the MAF activity is indeed due to IFN-gamma, IFN-gamma was purified from the culture supernatant of another human T cell hybridoma, L265K2, a cell line known to produce high levels of IFN-gamma. Two highly purified IFN-gamma fractions with m.w. of 20,000 and 25,000, respectively, were obtained by NaDodSO4/polyacrylamide gel electrophoresis (SDS-PAGE). Similar fractions were obtained from IFN-gamma derived from human peripheral blood lymphocyte (PBL) cultures induced with 12-0-tetradecanoylphorbol-13-acetate (TPA) and phytohemagglutinin (PHA). In comparison, Escherichia coli-derived recombinant human IFN-gamma separated by SDS-PAGE yielded two major active fractions with m.w. of 17,000 and 34,000. With all three types of preparations, a close correlation was found between the presence of IFN-gamma activity demonstrable in an antiviral assay and MAF activity in individual fractions. Substantial quantitative differences were observed in the ability of various human IFN to activate monocytes. Although no MAF activity was detected with IFN-alpha and IFN-beta at concentrations up to 200 U/ml, both natural and recombinant IFN-gamma showed marked MAF activity at concentrations as low as 0.3 to 1 U/ml.

Antigens, Surface↗
PubMed1981

Human blood monocytes: characterization of negatively selected human monocytes and their suspension cell culture derivatives.

Normal human monocytes were negatively selected from leucapheresis cell suspensions by countercurrent centrifugation-elutriation in high yield with a mean purity of 93.5%. The combination of the novel methods of negative cell selection and suspension cell culture has provided the opportunity to study serially over several days the morphologic and functional changes of monocytes from a single donor as they matured in culture to typical macrophages. Human monocytes nearly double in size during the first week of culture, experiencing near daily increases in cell volume. This was associated with changes in the ultrastructure of these cells, including the development of numerous small knob-like projections on the cell membrane and the proliferation of microtubules and filamentous structures within the cell cytoplasm during the first 6 days of culture. Peroxidase activity declined during the first 4 days of culture, whereas 5'-nucleotidase activity was acquired during the first 48 h of culture. Lysozyme activity in the cultures increased form day 2 to day 6 of culture. The phagocytic capacity of monocytes for igG-coated erythrocytes increased dramatically during the first week of culture, but the cytotoxic capability of monocytes against similar targets in an antibody-dependent cytotoxicity assay declined to nearly half of base-line levels by day 2 of culture and remained at this diminished level during subsequent days of culture.

Antibody-Dependent Cell Cytotoxicity↗
PubMed1980

Activation of human B lymphocytes. XII. Differential effects of in vitro cyclophosphamide on human lymphocyte subpopulations involved in B-cell activation.

The differential effects of in vitro cyclophosphamide (CY) on subpopulations of normal human peripheral blood lymphocytes involved in the pokeweed mitogen-induced plaque-forming cell (PFC) response against sheep red blood cells were examined. It was found that the plaque-forming B cells in this system are sensitive to CY over a wide concentration range including concentrations which have a minimal effect on overall cell viability. Kinetic experiments revealed that CY exerts its inhibitory effect on the PFC response only if added very early in culture. Thus, it appears that in vitro CY must exert its inhibitory influence on an early phase of polyclonal B-cell activation. When T-cell enriched (TCE) populations were incubated overnight with high concentration CY and then added back in co-culture to fresh autologous B cells, significant enhancement of PFC responses was observed suggesting a selective inhibition or elimination of a regulatory suppressor cell population found in TCE lymphocyte preparations. Helper T cells are relatively resistant to the inhibitory actions of CY. Thus, human B cells appear to be most sensitive to CY, followed in sensitivity by the suppressor cell populations in the T-cell fraction with relative resistance of the helper T cells. These observations have direct relevance in understanding the mechanisms of selective action of CY on normal human lymphocyte subpopulations with possible application to disease states in man.

Antibody Formation↗
PubMedDate not supplied

Low density lipoprotein metabolism in human macrophages stimulated with microbial or microbial-related products.

To determine whether stimulation of macrophages with products related to, or released as a consequence of, infectious processes could play a role in inducing the formation of foam cells, we studied the metabolism of native and acetylated low density lipoprotein (LDL) by human macrophages stimulated with lipopolysaccharide (LPS), muramyl dipeptide (MDP), polyinosinic:polycytidilic acid (Poly I:Poly C) and gamma-interferon. Cholesteryl ester (CE) synthesis by macrophages stimulated with LPS, MDP and Poly I:Poly C was markedly increased when the cells were incubated with native LDL (p less than 0.05). When incubated with acetylated LDL, LPS-stimulated macrophages showed a depression in CE synthesis (p less than 0.05). When incubated with acetyl-LDL, macrophages stimulated with Poly I:Poly C and gamma interferon showed a significant increase (p less than 0.05) in CE synthesis. The increase in CE synthesis by LPS-stimulated macrophages exposed to native LDL and by gamma-interferon-stimulated macrophages exposed to acetylated LDL was paralleled by an increase in cholesterol ester mass. The increase in CE synthesis and accumulation observed in LPS-stimulated macrophages incubated with native LDL seems to be due to an increase in the receptor mediated uptake of LDL. LPS inhibited and gamma-interferon activated the expression of the scavenger pathway in human macrophages. This may explain the changes observed in CE synthesis and accumulation when macrophages activated by the above stimuli were incubated with acetylated LDL. In conclusion, activation of human macrophages by some products released during, or as a consequence of, infectious processes led to an increase in CE synthesis and accumulation that may be relevant to the formation of "foam cells".

Acetylmuramyl-Alanyl-Isoglutamine↗