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Biomedical subjects

D A Stevens

Publications and source records attributed to D A Stevens.

At least 271 records · Page 15Linked to original sources
PubMed1988

In vivo and in vitro activation of pulmonary macrophages by IFN-gamma for enhanced killing of Paracoccidioides brasiliensis or Blastomyces dermatitidis.

The fungicidal capacity of murine pulmonary macrophages (PuM) activated in vitro with IFN or lymphokines or in vivo with IFN was studied. PuM treated overnight with IFN (1000 U/ml), Con A-stimulated spleen cell culture supernatants, or lymph node cells plus Con A significantly killed yeast cells of the Gar w isolate of Paracoccidioides brasiliensis 45.5 +/- 2.1%, 72.0 +/- 4.2%, and 51.5 +/- 0.7% respectively. Two other isolates of P. brasiliensis (Ru and LA) were also killed (45 and 34%) by PuM activated by lymph node cells plus Con A. Control PuM had lesser but significant capacity for killing of P. brasiliensis isolates, ranging from 15 to 22%. Killing of P. brasiliensis by PuM activated by Con A-stimulated spleen cell culture supernatants could not be significantly inhibited by superoxide dismutase, catalase, or azide. When mice were treated in vivo with 4 X 10(5) IFN U i.p. and PuM isolated 24 h later, the PuM had significantly enhanced ability to kill P. brasiliensis (47.0 +/- 6.3%) compared with PuM from control mice (25.0 +/- 4.2%). PuM thus activated also showed enhanced killing (43%) of a second isolate compared with control PuM (22%). PuM from IFN-treated mice were able to significantly kill Blastomyces dermatitidis (37.5 +/- 0.7%) compared with control PuM (4.5 +/- 6.3%). These results show that PuM can be activated in vitro and in vivo by IFN for enhanced fungicidal activity against two pulmonary fungal pathogens and suggests that immunologic production of IFN could be an important factor in host defenses against these diseases.

Animals↗
PubMed1988

Ketoconazole therapy of progressive coccidioidomycosis. Comparison of 400- and 800-mg doses and observations at higher doses.

One hundred and twelve patients with progressive pulmonary, skeletal, or soft tissue infections caused by Coccidioides immitis were randomly assigned to treatment with 400 or 800 mg per day dosages of ketoconazole. During therapy, if response was unsatisfactory, the protocol provided for treatment with higher doses. With 400 mg, ketoconazole resulted in 23.2 percent successes, which was similar to 32.1 percent successes with 800-mg treatments (p = 0.29). An additional six of 23 patients in whom initial therapy failed and who later received 1,200 or 1,600 mg per day of ketoconazole also showed improvement. However, among patients completing successful courses of therapy, relapses were more frequent in those requiring higher than 400-mg dosages for their success. From these studies, it is concluded that ketoconazole in doses above those currently recommended offer little or no benefit for most patients with non-meningeal forms of coccidioidomycosis.

Adult↗
PubMed1988

Regulation of immune responses by T suppressor cells and by serum in chronic paracoccidioidomycosis.

Regulation of cellular responses was studied during the course of chronic murine disseminated paracoccidioidomycosis. Regulation of peripheral blood lymphocyte (PBL) proliferative responses to concanavalin A (Con A) was studied in vitro by mixing PBL from infected and noninfected mice. PBL from mice infected for 18 weeks had depressed responses to Con A and they depressed the Con A responses of PBL from noninfected mice by 95% when they were mixed in a 1:1 ratio. After treatment of PBL from infected mice with anti-Lyt-2.2 antibody plus complement, the responses to Con A were increased to normal values. The percentage of T-cell subpopulations in PBL from infected mice did not differ significantly from those of normal mice. Immunoregulation of delayed-type hypersensitivity (DTH) responses to antigen by serum from infected animals was studied in mice 1 week after intranasal (i.n.) infection, a time when DTH responses were maximal. DTH responses to antigen 7 days after i.n. infection (10(7) CFU Paracoccidioides brasiliensis) were significantly reduced when 0.5 ml of immune mouse serum (ELISA antibody titer to P. brasiliensis antigens 1:10,240) was given i.v. 1 day before infection (P less than 0.01) or 1 day before skin testing (P less than 0.001). Normal mouse serum did not have this effect. The results indicate that progression of chronic disseminated paracoccidioidomycosis was associated with the development of T-cell suppressor activity for Con A responses of PBL, and that DTH responses to antigen were depressed by the administration of serum with specific high titer antibodies.

Animals↗
PubMed1988

Gamma-interferon activation of macrophages for killing of Paracoccidioides brasiliensis and evidence for nonoxidative mechanisms.

Fungicidal activity of murine peritoneal macrophages for the yeast form of the dimorphic fungal pathogen P. brasiliensis was studied. Killing was assessed by reduction of colony forming units (CFU) using a new medium which has a good plating efficiency. Resident peritoneal macrophages phagocytosed but did not kill P. brasiliensis. Macrophages treated overnight with recombinant gamma-interferon (IFN), lymph node cells plus concanavalin A (Con A) or Con A-stimulated spleen cell culture supernatants (Con A Sup) reproducibly killed three different isolates of P. brasiliensis (35 - 55%, P less than 0.05 - P less than 0.001). This is the first demonstration of killing of this organism by macrophages. Activated macrophages did not show enhanced phagocytosis of P. brasiliensis. Activation of macrophages for killing by IFN was dose-dependent and, varying with the isolate, 100 - 10,000 U/ml was required for inducing significant fungicidal effects against P. brasiliensis. Activation of macrophages by IFN or Con A Sup was abrogated by anti-IFN antibody. These results suggest that immune modulation may be an approach to therapy of paracoccidioidomycosis. Killing was not significantly inhibited in the presence of superoxide dismutase (450 U/ml), catalase (20,000 U/ml), dimethylsulfoxide (300 mM) or azide (1 mM). This indicated that killing mechanism(s) did not depend upon products of the oxidative burst. These results show that P. brasiliensis can be significantly killed by activated macrophages without products of the oxidative burst.

Animals↗
PubMed1988

A Candida albicans dispersed, repeated gene family and its epidemiologic applications.

Candida albicans causes a wide variety of infections but can readily be isolated from the skin and mucosa of healthy individuals. To enable high-resolution epidemiologic studies on this common pathogen, a species-specific DNA probe has been isolated from its genome. There are approximately equal to 10 copies of the sequence dispersed among the chromosome-sized DNA molecules resolved by pulsed-field electrophoresis. New DNA polymorphisms in this gene family arise at high rates. As a consequence, this probe will readily distinguish strains from different patients in the same hospital and from various sites in individual patients. The DNA polymorphisms detected by using this probe are largely due to internal changes in members of the family rather than movement to new genomic locations. This suggests recombination or gene conversion rather than transposition as the mechanism producing the observed variation.

Candida albicans↗
PubMed1988

Clinical and microbiological features of infection with Malassezia pachydermatis in high-risk infants.

Malassezia pachydermatis, a yeast that has not previously been implicated as a cause of human disease, was isolated from cultures of blood from three infants. All infants were 25-27 w of gestational age and had multiple underlying medical problems. The infants had been hospitalized for at least six weeks, had received broad-spectrum antibiotics, and had received parenteral lipid nutrition via a central venous catheter. In one patient, fungemia was accompanied by clinical and laboratory evidence of Broviac catheter infection. During a three-year period, M. pachydermatis was also recovered from fungal cultures of an additional 30 patients, 85% of whom were infants. A pathogenic role for M. pachydermatis recovered from sources other than blood or catheters was not established. Risk factors for and symptoms in infants with M. pachydermatis fungemia appeared to be similar to those described for Malassezia furfur sepsis.

Catheterization, Central Venous↗
PubMed1988

Enhanced oxidative mechanisms in immunologically activated versus elicited polymorphonuclear neutrophils: correlations with fungicidal activity.

Peritoneal polymorphonuclear neutrophils (PMN) from mice were tested for their ability to kill the yeast form of Blastomyces dermatitidis (Bd) in vitro and for their fungicidal mechanisms. PMN elicited from immune mice by the intraperitoneal injection of non-viable Bd (referred to as immunologically activated PMN or ActPMN) showed significantly enhanced fungicidal activity in comparison with PMN elicited with thioglycollate medium (ThioPMN) [means = 44.7% (SD 12.8%) and 16.4% (SD 9.2%) killed; n = 14; p less than 0.001]. Production of superoxide anion (O2-) by ActPMN after stimulation with phorbol myristate acetate was enhanced in comparison with production by ThioPMN. Superoxide dismutase, which removes O2-, inhibited ActPMN killing by 75% (p less than 0.001) when added to cultures immediately before challenge with Bd (optimal concentration: 6000 U/ml). Sodium azide, which inhibits myeloperoxidase and scavenges singlet oxygen (1O2), and catalase, which breaks down hydrogen peroxide (H2O2), inhibited ActPMN killing by 64% (p less than 0.001) and 52% (p less than 0.001), with optimal concentrations of 1 mM and 10,000 U/ml, respectively. Two agents that both scavenge 1O2 and antagonise hypochlorous acid (HOCl-), histidine and tryptophan, were also powerful inhibitors of ActPMN killing. Quenchers of hydroxyl radical (.OH), dimethylsulfoxide and sodium benzoate, had less effect, and required higher concentrations. These data suggest that the enhanced killing of Bd by ActPMN involves one or more oxidative mechanisms, and that there is a prominent role for O2-, either directly or as a precursor of other active oxygen species, a probable role for H2O2, and possible roles for 1O2, HOCl-, and .OH.

Animals↗
PubMed1988

Treatment of mycoses with itraconazole.

The efficacy of itraconazole, a new oral triazole, was evaluated at doses of 50-400 mg/day in 64 courses: 39 with coccidioidomycosis, and 25 with other mycoses. Among patients with coccidioidomycosis, 21 had pulmonary disease; 10, bone and joint; 8, lymphatic; 6, skin and soft tissue; 3, meningeal; and 1, urogenital. After a mean of 7.1 months of treatment, 17 of 27 evaluable courses (63%) have shown a full response; 8 (30%), a partial response; and 2 (7%), no response. Response to treatment was higher in patients with a shorter duration of illness and in patients with no previous treatment. After a mean of 12 months of treatment, two of five responders off therapy have relapsed. In patients with other mycoses, diagnoses include aspergillosis in six patients; histoplasmosis and tinea in three each; alternariosis, candidiasis, cryptococcosis, blastomycosis, and chromomycosis in two each; and pseudallescheriosis in one. After a mean of 9.4 months of treatment, 14 of 23 evaluable patients (61%) have shown a full response; 2 (8%), a partial response; and 7 (30%), no response. All patients with histoplasmosis and blastomycosis have responded to therapy. There are no relapses among five evaluable responders after a mean of 9.2 months off therapy. In 512 patient months of therapy, toxicity has been minimal. Mean serum levels in patients receiving 200 mg twice daily range from 3.9 to 5.9 micrograms/ml for 8 hr after dosing, with a peak at 7 hr and wide interpatient variability. Itraconazole is well tolerated and has continued to demonstrate efficacy in the treatment of a variety of systemic and superficial mycoses.

Adult↗
PubMed1988

Ketoconazole treatment of coccidioidal meningitis.

Fifteen patients with coccidioidal meningitis were treated with high doses of ketoconazole for up to 4 years. Five patients were treated with ketoconazole alone. One clinically failed, one developed hepatotoxicity, and three achieved remission of meningitis. One patient received intrathecal AMB in addition to ketoconazole for only 2 weeks before continuing on ketoconazole alone. He improved, but discontinued ketoconazole because of nausea and vomiting, and suffered a lethal relapse. Nine patients received ketoconazole in combination with prolonged courses of intrathecal AMB. Two patients were failures from nausea and vomiting, and the remaining seven either improved or experienced remission. The clinical responses appeared to be similar in patients receiving high-dose ketoconazole, either alone or combined with AMB, suggesting that there is no clinically significant antagonism of the drugs. Nausea and vomiting are significant limitations of high-dose ketoconazole. Ketoconazole alone is effective in some patients with coccidioidomycotic meningitis.

Amphotericin B↗
PubMed1988

Pharmacokinetics of fluconazole in cerebrospinal fluid and serum in human coccidioidal meningitis.

The pharmacokinetics of fluconazole, a new oral azole, were evaluated in cerebrospinal fluid and sera of eight patients with coccidioidal meningitis. At a dose of 50 mg/day, peak concentrations of 2.5 to 3.5 and 2.0 to 2.3 micrograms/ml occurred at 2 to 6 and 4 to 8 h in serum and cerebrospinal fluid, respectively. At 100 mg/day, peak concentrations of 4.5 to 8.0 and 3.4 to 6.2 micrograms/ml occurred at 2 to 4 and 4 to 12 h, respectively. The mean ratios of the concentration in cerebrospinal fluid to that in serum were 73.8% at 50 mg/day and 88.7% at 100 mg/day. Results suggested that there was a prolonged half-life in both cerebrospinal fluid and serum and that it was slightly longer in the former. Minimal toxicity was noted in 34 patient months of therapy (12 months on 50 mg daily; 22 months on 100 mg daily). After a mean of 4.5 months of therapy, five patients responded to therapy and three were unevaluable. The penetration of fluconazole into cerebrospinal fluid was substantial, toxicity was minimal, and early clinical experience was encouraging. Fluconazole holds promise as the sole or adjunctive therapy for fungal meningitis.

Adult↗
PubMed1988

Inhibition by estrogens of conidium-to-yeast conversion in the fungus Paracoccidioides brasiliensis.

Conidia produced by Paracoccidioides brasiliensis are inhibited by mammalian estrogens in their in vitro conversion into yeast-form cells. This was demonstrated with four different isolates. In these experiments, conversion was reduced to 10.7 and 34.4% of the control values by 17-beta-estradiol at 10(-6) and 10(-8) M, respectively. At the same concentrations, the synthetic estrogen diethylstilbestrol was slightly less inhibitory. In contrast, other sex hormones and analogs, i.e., testosterone, 17-alpha-estradiol, tamoxifen, and hydroxytamoxifen, had no effect on conidium-to-yeast conversion. Previous studies have shown that estrogens similarly inhibit mycelium-to-yeast-form transition in P. brasiliensis. Conidia, and not mycelial fragments, are believed to be the natural infectious propagules. These findings with conidia support the hypothesis that estrogens, affecting the initial host-parasite interactions by suppressing conversion to the parasitic form of the organism, are, at least in part, responsible for the greater resistance of females to paracoccidioidomycosis.

Estrogens↗
PubMed1988

Impairment of cellular but not humoral immune responses in chronic pulmonary and disseminated paracoccidioidomycosis in mice.

Humoral and cellular immune responses were measured during the progression of chronic pulmonary and disseminated paracoccidioidomycosis in mice. The chronic disease was established by pulmonary infection of mice with different doses of the yeast form of Paracoccidioides brasiliensis isolate GAP. Levels of antibodies to P. brasiliensis, detected in serum by immunodiffusion and enzyme-linked immunosorbent assay, directly correlated with the size of the infectious challenge. Significant delayed-type hypersensitivity (DTH) responses to antigen were largely restricted to week 1 after pulmonary infection with intranasally administered high doses (5.0 x 10(6) or 1.1 x 10(7) CFU per inoculum). In vitro lymphoproliferative responses of peripheral blood lymphocytes (PBL) to P. brasiliensis antigens were significant only at 2 weeks after infection with intranasally administered 1.1 x 10(7) CFU. Responses of PBL to concanavalin A were depressed (50% of control response) as early as 8 weeks and reached a nadir at 10 to 18 weeks after infection. Infected mice made antibodies to sheep erythrocytes (SRBC) (10(9) intravenously [i.v.]) normally at all times tested after infection. In contrast, infected mice sensitized to SRBC (10(6) i.v.) had significantly depressed DTH responses to SRBC at 9 and 20 weeks postinfection compared with noninfected mice. These results indicated that in this model, normal humoral responses developed to homologous and heterologous antigens. In contrast, the T cellular immune responses were depressed with progression and chronicity of the disease. Thus, this model closely mimics the immunological findings in human paracoccidioidomycosis.

Animals↗
PubMed1988

Dermatophyte-hormone relationships: characterization of progesterone-binding specificity and growth inhibition in the genera Trichophyton and Microsporum.

We reported previously that Trichophyton mentagrophytes contains a cytoplasmic macromolecule which specifically binds progesterone. Progesterone is also an effective inhibitor of growth of the fungus. We report here studies which characterize more fully the specific binding properties and the functional responses of T. mentagrophytes and taxonomically related fungi to a series of mammalian steroid hormones. Scatchard analysis of [3H]progesterone binding in both the + and - mating types of Arthroderma benhamiae and in Microsporum canis revealed a single class of binding sites with approximately the same affinity as that in T. mentagrophytes (Kd, 1 X 10(-7) to 2 X 10(-7) M). Trichophyton rubrum had a protein with a higher binding affinity (Kd, 1.6 X 10(-8) M). Characterization of the [3H]progesterone-binding sites in T. mentagrophytes showed the binder to be a protein which was destroyed by trypsin and heating to 56 degrees C. Previous examination of the steroid-binding specificity in T. mentagrophytes had demonstrated that deoxycorticosterone (DOC) and dihydrotestosterone (DHT) were effective competitors for [3H]progesterone binding. Expansion of this study to include other competitors revealed that R5020 (a synthetic progestin), androstenedione, and dehydroepiandosterone possessed relative binding affinities which were 20, 11, and 9% of that of progesterone, respectively. Other ligands tested were less effective. Competition studies for the binder in M. canis resulted in similar findings: DOC and DHT were effective competitors for [3H]progesterone binding. The growth of A. benhamiae + and -, M. canis, and T. rubrum were all inhibited by progesterone in a dose-responsive manner, with 50% inhibition achieved at concentrations of 9.8 x 10(-6), 1.2 x 10(-5), 1.5 x 10(-5), and 2.7 x 10(-6) M. respectively,.

Binding, Competitive↗
PubMed1988

A culture medium for Paracoccidioides brasiliensis with high plating efficiency, and the effect of siderophores.

The plating efficiency of Paracoccidioides brasiliensis on standard mycological media is poor, impairing its isolation and recovery from various sources, particularly infected tissues. We describe a medium that markedly improves P. brasiliensis plating efficiency. It consists of a synthetic medium (modified McVeigh-Morton) supplemented with 4% (v:v) horse serum and 5% (v:v) culture filtrate from stationary phase P. brasiliensis cultures. A commercially available medium (brain-heart infusion), ordinarily inferior to unsupplemented McVeigh-Morton medium, is at least as efficacious as supplemented McVeigh-Morton medium when supplemented in this manner. We show that plating efficiency varies among P. brasiliensis isolates and can even vary with the isolate's history of passage in culture. In contrast, all isolates studied could produce the growth enhancing factors present in culture filtrate. Some siderophores produced by other fungi can be substituted for the culture filtrate, whereas others can be substituted for both the filtrate and serum. The enhancing effect of filtrate and/or serum could be removed by chelating iron. P. brasiliensis-produced siderophores are likely to be the growth enhancing moiety in culture filtrates.

Culture Media↗
PubMed1987

Activation of murine polymorphonuclear neutrophils for fungicidal activity by recombinant gamma interferon.

Recombinant murine gamma-interferon (IFN) was tested for its ability to enhance murine polymorphonuclear neutrophil (PMN) fungicidal activity in vitro. PMNs, elicited by intraperitoneal injection of thioglycollate 4 hr prior to collection, were treated with 0.00003-300,000 units of IFN per milliliter for 1 hr prior to challenge with yeast form Blastomyces dermatitidis. These PMNs were not fungicidal for Blastomyces in the absence of IFN; significant enhancement of PMN fungicidal activity by IFN treatment occurred in a dose-dependent manner with maximal enhancement observed at 30,000 U/ml (21% killing). Pretreatment of IFN with rabbit anti-IFN antiserum before addition to PMNs eliminated the enhancement of fungicidal activity by effective doses of IFN. PMN fungicidal activity against phagocytizable Candida albicans was significantly (P less than .001) higher (71.3 +/- 17.4%) than against B. dermatitidis. Candidacidal activity was not significantly enhanced by IFN treatment of PMNs. Exogenously added lipopolysaccharide, at levels corresponding to those found in this preparation of IFN, did not activate PMNs for enhanced fungicidal capacity. These data indicate a stimulatory role for IFN in the killing of B. dermatitidis by PMNs, suggesting that IFN is an active component of the communication between T lymphocytes and PMNs with respect to antimicrobial resistance. They suggest a natural role for IFN in host defense against blastomycosis and other fungal infections, and a possible therapeutic use for exogenous IFN in fungal disease.

Animals↗
PubMed1987

Effect of a local immune reaction on peripheral blood polymorphonuclear neutrophil microbicidal function: studies with fungal targets.

Peripheral blood polymorphonuclear neutrophils (PMN) from mice immunized with Blastomyces dermatitidis and then stimulated locally (intraperitoneally, ip) with B. dermatitidis antigen had enhanced killing of B. dermatitidis in vitro (54.4 +/- 19.49 of inoculum) compared to nonimmune mice (32.7 +/- 8.7%; P less than 0.02), nonimmune mice given antigen ip (30.6 +/- 14.0%; P less than 0.05), or immune mice not given antigen ip (15.4 +/- 9.9%; P less than 0.01). Peripheral blood PMN from all four groups had marked killing ability against Candida albicans (91.8-99.3% of inoculum). That the killing of B. dermatitidis was due to PMNs was demonstrated by lack of killing by isolated peripheral blood mononuclear cells from all four groups. A local immune reaction can result in enhancement of PMN fungicidal activity, and this is reflected even in peripheral blood PMN. We hypothesize this is an important component of normal host defenses against fungal infection, and likely other microbial infections. Enhancement of PMN microbicidal function by the soluble mediators presumed to be responsible for the effects observed may be an approach to immunomodulating therapy or prophylaxis of infection.

Animals↗