PubMed Health⌕ Search

Biomedical subjects

S J Ackerman

Publications and source records attributed to S J Ackerman.

At least 109 records · Page 6Linked to original sources
PubMed1986

Effects of glucocorticoids on eosinophil colony growth.

The in vivo and in vitro effects of glucocorticoids on eosinophilopoiesis were examined with soft agar cultures of bone marrow and peripheral blood cells. Prednisone, 10 mg four times daily for three days, administered to normal volunteers, caused a significant drop in circulating eosinophil numbers (p less than 0.005) but did not decrease eosinophil colony numbers in cultures of bone marrow or peripheral blood. A patient with peripheral blood eosinophilia demonstrated a larger percentage decrease in mean eosinophil colony numbers (50%) after prednisone than did any normal volunteer. Incubation of bone marrow cells for 1 hour with either high concentrations of hydrocortisone, up to 3.3 X 10(-4) mol/L, or with postinfusion plasma from volunteers administered intravenous hydrocortisone, plasma levels to 1.6 X 10(-5) mol/L, did not decrease the numbers of eosinophil colonies. At a concentration of 3.3 X 10(-6) mol/L of hydrocortisone, there was a slight but statistically significant stimulation of eosinophil colony numbers. In contrast, incubation with high concentrations of dexamethasone, 3.3 X 10(-4) mol/L or 3.3 X 10(-6) mol/L, significantly reduced eosinophil colony numbers. Prednisone caused a significant reduction in plasma levels of Charcot-Leyden crystal protein but not of eosinophil granule major basic protein. The results indicate that soft agar assay of eosinophil colony growth by blood or bone marrow cells cannot be used to model the in vivo eosinopenic effect of glucocorticoids, levels of dexamethasone in excess of levels commonly administered in clinical practice are required to inhibit eosinophilopoiesis in vitro, and patients with peripheral blood eosinophilia may be more susceptible to the eosinopenic effects of glucocorticoids than normal subjects.

Blood Proteins↗
PubMed1986

Biochemical and functional similarities between human eosinophil-derived neurotoxin and eosinophil cationic protein: homology with ribonuclease.

Eosinophil-derived neurotoxin (EDN) and eosinophil cationic protein (ECP) were isolated from lysates of human eosinophil granules by gel filtration and ion exchange chromatography on heparin-Sepharose. Radioimmunoassay, using monoclonal antibodies, of fractions from the heparin-Sepharose chromatography showed one peak of EDN activity and two peaks of ECP activity (termed ECP-1 and ECP-2). EDN, ECP-1, and ECP-2 each exhibited heterogeneity in charge and molecular weight when analyzed by two-dimensional nonequilibrium pH gradient electrophoresis and NaDodSO4/PAGE. Digestion of EDN with endoglycosidase F (endo F) decreased its molecular weight and charge heterogeneity. Thus, END likely contains a single complex oligosaccharide. Endo F digestion of ECP-1 and ECP-2 decreased the molecular weight of both polypeptides, indicating that both likely contain at least one complex oligosaccharide. Amino acid sequence analyses showed that ECP-1 and ECP-2 are identical from residue 1 through residue 59 and that the sequences of EDN and ECP are highly homologous (37 of 55 residues identical). Both EDN and ECP NH2-terminal sequences showed significant homology to RNase, especially in regions of the RNase molecule involved in ligand binding. EDN, ECP-1, and ECP-2 had neurotoxic activity, causing the Gordon phenomenon at doses down to 0.15 micrograms when injected into the cisterna magna; the proteins were comparable in their activities. These results indicate that EDN and ECP are related proteins and suggest that they derived from genes associated with the RNase family.

Amino Acid Sequence↗
PubMed1985

Dermal deposition of eosinophil-granule major basic protein in atopic dermatitis. Comparison with onchocerciasis.

Although blood eosinophilia is commonly present in atopic dermatitis, accumulation of tissue eosinophils is not prominent. To determine whether eosinophil degranulation occurs in lesions of atopic dermatitis, we analyzed tissues by immunofluorescence for the presence of the eosinophil-granule major basic protein. Twenty biopsy specimens from 18 patients with atopic dermatitis were studied, and all showed major basic protein staining outside eosinophils. In 18 specimens, the staining was fibrillar, was located in the upper half of the dermis, and was similar to the distribution of elastic fibers. Twelve specimens with fibrillar staining also showed major basic protein staining in the form of extracellular granules. One specimen from unaffected skin showed minimal faint, fine, fluorescing fibrils, but there was marked deposition of the protein in affected skin. The fibrillar pattern of major basic protein staining in atopic dermatitis was very similar to that seen in lichenified lesions of untreated onchocerciasis. These results suggest that eosinophils commonly release granule proteins in the dermis and that assessment of eosinophil involvement in disease cannot be based simply on numbers of eosinophils in tissue.

Adolescent↗
PubMed1985

Donor sensitivity to basophil activation by eosinophil granule major basic protein.

Leukocytes of 22 individuals were incubated with a suboptimal concentration of eosinophil granule major basic protein and the amount of histamine release was determined. Cells from 20 of the 22 donors had a histamine release of greater than or equal to 15%. The use of selected donors on repeated occasions revealed significant variability in individual donor responsiveness. The findings support a role for major basic protein activation of human basophils in allergic reactions.

Basophils↗
PubMed1985

Increased numbers of hypodense eosinophils in the blood of patients with bronchial asthma.

We investigated the density of blood eosinophils from patients with asthma using polyvinylpyrrolidone-coated silica gel (Percoll) discontinuous density gradient centrifugation of peripheral blood leukocytes. Ten patients with allergic asthma, 10 normal subjects, and 2 patients with the hypereosinophilic syndrome (HES) were studied. The density distribution profiles of eosinophils from normal subjects showed: (1) peaks at densities of 1.085 to 1.090 g/ml and (2) inflection points or nadirs near 1.082 g/ml, below which only 10% of eosinophils were found. On the basis of these results, we divided eosinophils into 2 subpopulations: normodense (greater than 1.082 g/ml) and hypodense (less than 1.082 g/ml). Densities of eosinophils from patients with asthma and HES peaked at 1.083 and 1.076 g/ml (mean values), respectively, significantly lighter than eosinophils from normal donors (p less than 0.005 and p less than 0.001, respectively). The proportions of hypodense eosinophils in patients with asthma and HES were 35 and 95%, respectively, and were significantly greater than that in normal donors (p less than 0.002 and p less than 0.001, respectively). The density distribution profiles of a normal subject were stable over time, but those of asthmatic patients varied with time. For the 22 participants, there was a positive correlation between log-transformed blood eosinophil counts and the percentage of hypodense eosinophils (r = +0.86, p less than 0.001). Similarly, for 15 of them, plasma eosinophil granule major basic protein correlated with the numbers of peripheral blood eosinophils (r = +0.92, p less than 0.0005) and hypodense eosinophils (r = +0.92, p less than 0.0005). Thus, a portion of the eosinophils in asthmatic patients and most of the eosinophils in patients with HES are hypodense.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1985

Comparative toxicity of purified human eosinophil granule cationic proteins for schistosomula of Schistosoma mansoni.

The human eosinophil granule contains several distinctive cationic proteins that have been purified to homogeneity, including major basic protein (MBP), eosinophil cationic protein (ECP), and eosinophil-derived neurotoxin (EDN). Two earlier studies have shown that MBP and ECP both damage schistosomula of Schistosoma mansoni in vitro in a dose-dependent fashion. The present study expands upon these observations by comparing the toxicity of MBP, ECP, as well as EDN when tested at equimolar concentrations (0.03-2 X 10(-5) M). On a molar basis, ECP was 8 to 10 times more potent than MBP, and the ECP-mediated killing of schistosomula was qualitatively different than that of MBP. Purified ECP produced complete fragmentation and disruption of schistosomula, whereas MBP produced a distinctive ballooning and detachment of the tegumental membrane. In contrast, EDN was only marginally toxic at high concentrations and caused crinkling of the tegumental membrane. Heating MBP and ECP for four hr at 56 degrees C caused precipitation and loss of toxicity for MBP, but not for ECP. Native MBP (with reactive sulfhydryl groups intact) and stabilized, reduced and alkylated MBP had comparable toxicity. To determine the relative contribution of MBP, ECP and other potentially helminthotoxic eosinophil granule constituents to schistosomulum damage, fractions of acid soluble granule extracts prepared by chromatography on Sephadex G-50 columns were analyzed for toxicity to schistosomula and for MBP and ECP levels by radioimmunoassay. Schistosomula were killed by fractions containing MBP, and to a much lesser and more variable extent by fractions containing EDN and a 21,000 dalton protein, but not by fractions coincident with the elution of ECP, which contained concentrations of ECP below that required to produce significant killing of schistosomula by the purified protein. Therefore, although ECP is a more potent helminthotoxin for schistosomula than MBP on a molar basis, MBP, by virtue of its abundance in the granule, accounts for the bulk of the toxicity in fractions of acid solubilized granules obtained from eosinophils of patients with marked eosinophilia.

Animals↗
PubMed1985

Facial edema and eosinophilia. Evidence for eosinophil degranulation.

Two patients had recurrent facial edema and peripheral blood eosinophilia. One patient showed a marked increase in the serum level of the eosinophil granule major basic protein. In both patients, skin biopsy samples showed nonspecific mononuclear cell inflammation with few eosinophils. However, immunofluorescence staining showed extracellular localization of the major basic protein within the dermis, similar to that previously shown in chronic urticaria and the recently described syndrome of episodic angioedema with eosinophilia. These observations provide further evidence that degranulation of eosinophils occurs in the skin and suggest that eosinophil mediators may play a role in the development of cutaneous edema.

Blood Proteins↗
PubMed1985

Heterogeneity of human peripheral blood eosinophil-type colonies: evidence for a common basophil-eosinophil progenitor.

We have recently shown that a proportion of previously designated human eosinophil "(Eo)-type" colonies in methylcellulose contain basophils and histamine (Denburg et al Blood 61:775, 1983). In the present studies, individual Eo-type colonies have been analyzed by cell morphology as well as by biochemical assays for histamine, Charcot-Leyden crystal protein (CLC), and eosinophil granule major basic protein (MBP). Clonal origin of single Eo-type colonies was confirmed by G6PD isoenzyme analysis. Morphological observations of such colonies revealed the existence of two distinct colony types: (1) Eo type containing 100% basophils and (2) Eo type containing mixtures of basophils and eosinophils, including cells with mixed basophil-eosinophil granulation. Histamine was not detected in pure, mature peripheral blood eosinophils. Immunofluorescent studies demonstrated bright staining for CLC and MBP in 95% +/- 3% of cells in Eo-type colonies but only in 5% +/- 4% of cells in GM-type colonies. Radioimmunoassay for MBP was positive in 5/9 Eo-type and 0/10 neutrophil-macrophage ("GM-type") colonies, with a mean level (nanogram/colony) of 11.6 +/- 4.2 per Eo-type colony; four of the latter colonies were doubly positive for both histamine and MBP. These and previous findings point out the morphological and biochemical heterogeneity of peripheral blood Eo-type colonies and provide direct evidence for the existence of a common, circulating basophil-eosinophil progenitor.

Basophils↗
PubMed1985

Calcium ionophore A23187 calcium-dependent cytolytic degranulation in human eosinophils.

The divalent cation ionophore A23187 is frequently used for studies of eosinophil degranulation. Nonetheless, the mechanism whereby A23187 induces degranulation in human eosinophils is still unclear. In the present experiments, A23187 caused human eosinophils to release a granule protein, eosinophil-derived neurotoxin (EDN) and a membrane-associated protein, Charcot-Leyden crystal (CLC) protein in a calcium and a concentration-dependent manner. However, A23187 at a concentration (1 microgram/ml) that caused 15% EDN release and 30% CLC protein release also produced release of the cytoplasmic enzyme lactic dehydrogenase (LDH) and loss of cell viability, both of which were calcium dependent. CLC protein release preceded EDN release and was detectable even at 15 min after the addition of 1 microgram/ml A23187, whereas EDN release occurred after a lag period of 30 min, and coincided with LDH release. At 1 microgram/ml A23187, neither the release of LDH nor the loss of viability occurred with purified neutrophils obtained in the same blood sample as a by-product of eosinophil purification. Electron microscopic examination demonstrated that exposure to A23187 for 15 min resulted in an increase and elongation of microridges on the cell surface, and exposure for 45 min caused cell disruption followed by extrusion of membrane-bound granules through breaks in the plasma membrane. Only once was granule exocytosis observed. These results indicate that A23187 treatment of eosinophils causes an initial release of membrane-associated CLC protein by a noncytolytic mechanism, and causes degranulation as a result of eosinophil lysis.

Blood Proteins↗
PubMed1984

Charcot-Leyden crystal protein and eosinophil granule major basic protein in sputum of patients with respiratory diseases.

Sputum eosinophilia is characteristic of bronchial asthma, and sputum specimens from patients with asthma contain eosinophil-derived substances including Charcot-Leyden crystals (CLC) and the eosinophil granule major basic protein (MBP). Prior observations have indicated that an elevated sputum level of MBP is associated with asthma. To determine whether CLC protein was also elevated in asthma, we measured by specific radioimmunoassays the quantities of MBP and CLC protein in sputum specimens from 106 consecutive patients with various respiratory diseases and in sputum specimens from 10 patients hospitalized for asthma. The CLC protein was detected in all sputum samples, and the levels in the 116 samples averaged 3.5 micrograms/ml (range, 0.01 to 25 micrograms/ml). The CLC protein was significantly elevated in sputum specimens from patients with acute asthma and from patients with certain other respiratory diseases when associated with bronchopulmonary infection. In contrast, MBP levels in the 116 samples averaged 0.32 micrograms/ml (range, 0.01 to 8.8 micrograms/ml) and were significantly elevated only in patients with asymptomatic or acute asthma. Sputum MBP was not elevated in patients with bronchopulmonary infections unless acute asthma was also present. Thus, an elevated sputum MBP level was specifically associated with asthma, whereas an elevated CLC protein level was associated with both asthma and with bronchopulmonary infection in certain patient groups.

Asthma↗
PubMed1984

Stimulation of basophil and rat mast cell histamine release by eosinophil granule-derived cationic proteins.

Major basic protein (MBP), an arginine-rich basic polypeptide that constitutes the crystalloid core of the large specific eosinophil granule, has previously been shown to stimulate noncytolytic histamine release from human basophils and rat mast cells by an IgE-independent mechanism. Two additional basic polypeptides present in eosinophil granules, eosinophil cationic protein (ECP) and eosinophil-derived neurotoxin (EDN), were examined for similar activity in the present study. Acid-solubilized eosinophil granules were fractionated by chromatography on a Sephadex G-50 column. Incubation of basophil-containing human mononuclear cells with the individual column fractions demonstrated that histamine release occurred only with the fractions that contained MBP. The selectivity of the basophil response for MBP was confirmed by using equimolar concentrations of purified MBP, ECP, and EDN. In contrast, both MBP and ECP, but not EDN, stimulated histamine release from purified rat peritoneal mast cells. Reduction and alkylation of the MBP molecule diminished the response of human basophils to MBP but enhanced the potency of the molecule with rat mast cells. The distinct potency of MBP as a stimulus for histamine secretion from human basophils suggests that eosinophil release of MBP may be a specific event in the augmentation of immediate hypersensitivity reactions and other disorders characterized by eosinophilia.

Animals↗
PubMed1984

Deposition of eosinophil granule major basic protein onto microfilariae of Onchocerca volvulus in the skin of patients treated with diethylcarbamazine.

We investigated the association between eosinophil degranulation, as evidenced by the deposition of granule major basic protein (MBP), and the killing of microfilariae of Onchocerca volvulus in vivo following treatment with diethylcarbamazine (DEC). Utilizing an immunofluorescence procedure for the cellular and extracellular localization of eosinophil MBP in formalin-fixed, paraffin-embedded tissues, we studied skin biopsies from onchocerciasis patients before and during treatment with topically or orally administered DEC. Before DEC, there was little or no inflammatory response in either dermis or epidermis and microfilariae were essentially intact. Immunofluorescent staining for MBP revealed some filamentous fluorescence associated with dermal collagen fibers, very few eosinophils, and no fluorescence in association with intact microfilariae. In contrast, during treatment with DEC, immunofluorescent staining for MBP revealed extensive eosinophil infiltrates in both dermis and epidermis with numerous intraepidermal eosinophil abscesses containing degenerating microfilariae. An intense extracellular immunofluorescence for MBP surrounded degenerating microfilariae in the dermis and epidermis in both the presence and absence of eosinophil infiltrates as early as 4.5 hours after starting therapy. Many intact nondegenerating microfilariae were also present, but they did not show immunofluorescent staining for MBP nor a surrounding inflammatory infiltrate. The results show that immediately following administration of DEC, eosinophils localize and degranulate around microfilariae in the skin and release granule MBP onto or in close proximity to the parasite's surface. Because of the striking association between eosinophil localization, degranulation, and deposition of MBP onto microfilarial surfaces, and the degeneration of microfilariae in the skin, these observations support the hypothesis that the eosinophil, through helminthotoxic granule proteins such as MBP, damages the microfilariae of O. volvulus.

Adult↗
PubMed1984

Evidence for secretion of human eosinophil granule major basic protein and Charcot-Leyden crystal protein during eosinophil maturation.

Prior electron microscopic studies have suggested that immature eosinophils degranulate during normal maturation in human bone marrow. This hypothesis was tested by measuring levels of eosinophil granule major basic protein (MBP) and Charcot-Leyden crystal (CLC) protein in bone marrow sinusoidal blood. MBP and CLC protein levels were elevated initially in bone marrow sinusoidal blood from normal volunteers when compared with peripheral blood, and levels of both proteins decreased during serial sampling; CLC protein levels remained significantly elevated, while MBP levels declined to equal those of peripheral blood. To investigate whether MBP and CLC protein were secreted during eosinophil maturation, bone marrow cells were cultured in soft agar; MBP and CLC protein levels were measured in culture supernatants by RIA. There was a significant positive correlation between eosinophil colony growth and levels of each protein. Electron micrographs of cells in soft agar colonies provided ultrastructural evidence for secretion of granule products by immature eosinophils. These results support prior observations that eosinophil promyelocytes degranulate during maturation.

Adult↗
PubMed1983

Elevated serum levels in human pregnancy of a molecule immunochemically similar to eosinophil granule major basic protein.

We have shown that serum levels of a molecule immunochemically similar to eosinophil granule major basic protein (MBP) are elevated in pregnant women throughout gestation. MBP levels increase during gestation and plateau at approximately 7,500 ng/ml by the 20th wk (greater than 10-fold above normal). Levels return to normal after delivery, with a T1/2 of 13.7 d. The MBP in pregnancy serum is remarkably similar to the eosinophil granule MBP in that: (a) pregnancy MBP fully inhibits the binding of radiolabeled MBP standard in a double antibody radioimmunoassay; (b) this inhibition reaction is specific for human MBP because pregnancy serum produces no inhibition of the binding of radiolabeled guinea pig MBP in the guinea pig MBP radioimmunoassay; (c) in a two-site immunoradiometric assay for MBP, slopes of dose-response curves for pregnancy serum, purified MBP, and serum from a patient with hypereosinophilic syndrome are identical, and maximal binding is comparable; (d) reduction and alkylation of pregnancy sera increases measured MBP 100-fold, as previously shown for eosinophil granule MBP in serum; and (e) the MBP in pregnancy serum demonstrates the same pattern of heat lability as has been previously reported for MBP. Four observations have raised the possibility that the eosinophil is not the source of the MBP in pregnancy serum: (a) no correlation between serum MBP level and peripheral blood eosinophil count exists in pregnant women, in contrast to previous studies of patients with eosinophilia; (b) levels of three other eosinophil-associated proteins are normal or low in pregnancy sera, whereas the serum levels of these proteins are elevated in patients with eosinophilia; (c) the slopes of dose-response curves for pregnancy sera and MBP standards differ in the double antibody radioimmunoassay; and (d) the molecule in pregnancy serum elutes from Sephadex G-50 columns at the void volume, while eosinophil granule MBP and the MBP in serum of patients with eosinophilia elute at a volume consistent with the previously established molecular weight of 9,300. These findings suggest that the MBP in pregnancy serum is derived from a source other than the eosinophil.

Animals↗
PubMed1983

Localization of eosinophil granule major basic protein in human basophils.

In experiments using an immunofluorescent method to localize human eosinophil granule major basic protein (MBP) in cells and tissues, a small number of cells stained for MBP that subsequently could not be identified as eosinophils. Because the Charcot-Leyden crystal protein, another eosinophil protein, was recently identified in basophils, we tested whether MBP might also be a constituent of blood basophils. Highly purified, eosinophil-free basophil suspensions were prepared using the fluorescence-activated cell sorter (FACS) to sort basophil-containing mononuclear cell preparations stained with fluorescein-conjugated sheep IgG anti-human IgE antibody. Using these FACS-purified basophils, we found that: (a) enrichment for surface IgE-positive cells (greater than 95% basophils) by FACS also enriched for cells staining for MBP by immunofluorescence; (b) MBP appeared to be localized in the histamine-, heparin-containing granules; (c) significant quantities of MBP were measurable by radioimmunoassay (RIA) in freeze-thaw detergent extracts of purified basophils; and (d) RIA dose-response curves for extracts of purified eosinophils and basophils had identical slopes. The MBP content of basophils from normal individuals averaged 140 ng/10(6) cells, whereas purified eosinophils from normal donors and patients with the hyper-eosinophilic syndrome averaged 4,979 and 824 ng/10(6) cells, respectively. MBP was also detected by immunofluorescence and RIA in cells obtained from a patient with basophil leukemia, although the MBP content for basophil leukemia cells was lower than that for normal basophils. We conclude that basophils contain a protein that is immunochemically indistinguishable from eosinophil granule MBP.

Basophils↗
PubMed1983

Activation of basophil and mast cell histamine release by eosinophil granule major basic protein.

Major basic protein (MBP) is a primary constituent of eosinophil granules. In this report, we demonstrate that MBP from human eosinophil granules initiates a nonlytic histamine release from human leukocytes. A direct effect of MBP on basophils was confirmed using purified human basophils. The kinetics of release were similar to those reported for poly-L-arginine, although MBP was less potent than poly-L-arginine of similar molecular weight. Reduction and alkylation of MBP diminished both the potency and efficacy of the molecule. Native MBP also stimulated histamine secretion from purified rat peritoneal mast cells in a manner characteristic of other polycations. These results emphasize the bidirectional nature of the basophil/mast cell-eosinophil regulatory axis.

Animals↗
PubMed1983

Distinctive cationic proteins of the human eosinophil granule: major basic protein, eosinophil cationic protein, and eosinophil-derived neurotoxin.

The human eosinophil granule contains a number of cationic proteins that have been identified and purified to homogeneity, including the major basic protein (MBP), the eosinophil cationic protein (ECP), and the eosinophil-derived neurotoxin (EDN). Because of confusion in the literature regarding the distinctiveness of MBP and ECP, we investigated the immunochemical and physicochemical properties of these purified proteins by electrophoresis on sodium dodecyl sulfate-polyacrylamide gels (SDS-PAGE), by specific double antibody radioimmunoassays (RIA) for MBP and ECP, and by fractionation of acid-solubilized eosinophil granules on Sephadex G-50 columns. Analysis of a mixture of the three purified proteins by SDS-PAGE showed that they migrated as three distinct bands with differing m.w. Comparison by specific RIA for MBP and ECP did not demonstrate any appreciable immunochemical cross-reactivities among the three proteins. Sephadex G-50 column fractions of acid-solubilized eosinophil granules were analyzed by RIA and by SDS-PAGE analysis of individual column fractions. MBP, ECP, and EDN eluted at different volumes from Sephadex G-50 columns as determined by RIA and SDS-PAGE. Soluble extracts of eosinophil granules from patients with the hypereosinophilic syndrome contained between six and 64 times more MBP than ECP on a weight basis. These observations demonstrate that MBP, ECP, and EDN are distinctive cationic proteins of the human eosinophil granule and that eosinophil granules from patients with eosinophilia contain considerably greater quantities of MBP than ECP.

Binding, Competitive↗