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Biomedical subjects

D A Stevens

Publications and source records attributed to D A Stevens.

At least 325 records · Page 18Linked to original sources
PubMed1984

Inhibition and killing of Candida albicans in vitro by five imidazoles in clinical use.

Five imidazoles (clotrimazole, econazole, ketoconazole, miconazole, and tioconazole) in clinical use were compared for their ability to inhibit and kill Candida albicans. Eleven isolates were obtained from patients before therapy. By spectrophotometric determination of 50% growth inhibition, all isolates were inhibited at low concentrations, with clotrimazole slightly less active than the other four drugs. By the conventional MIC determination, tioconazole was more active than all of the others (P less than 0.01) except clotrimazole. In killing (minimum fungicidal concentration [MFC] assay), tioconazole was the most active by several analyses. Studies of the kinetics of killing indicated that the drugs studied could kill under conditions used for the MFC determination and that tioconazole and ketoconazole could kill particularly rapidly. If the drug was washed from the cells before subculturing, concentrations above the MFC were required to kill, but tioconazole could produce a lethal lesion in all cells virtually instantaneously. These findings are pertinent to MFC and killing kinetics methodology and to the observation of drug persistence after topical application. The results differ from some previous in vitro comparisons made with different methods. They are relevant to conclusions about drug mechanisms based on their abilities to inhibit and to kill, and they underscore the need to study various assay methods and fungal species.

Antifungal Agents↗
PubMed1984

Activation of murine polymorphonuclear neutrophils for fungicidal activity with supernatants from antigen-stimulated immune spleen cell cultures.

An in vitro model of in vivo immunological activation of murine polymorphonuclear neutrophils (PMN) was developed. Culture supernatants of spleen cells from Blastomyces dermatitidis-immunized mice stimulated with B. dermatitidis antigens in vitro were studied. Incubation of the supernatants with thioglycolate-elicited PMN enabled the cells to significantly reduce (31 +/- 6%) B. dermatitidis inoculum CFU. Optimum production of active supernatants occurred after 4 to 6 days of stimulation in vitro and required 200 micrograms of nonviable B. dermatitidis cells per ml. Generation of activity by immune spleen cells was shown to be antigen specific in that stimulation with a heterologous antigen or stimulation of nonimmune spleen cells with B. dermatitidis antigen did not produce active supernatants. The activity in supernatants was dose dependent, nondialyzable (molecular weight greater than or equal to 14,000), and relatively heat labile (80 degrees C, 30 min). Activation of PMN by supernatants for fungicidal activity against B. dermatitidis required only a short incubation period (1 h) followed by a 2-h coculture (challenge) period. Stimulation of normal spleen cells with concanavalin A also resulted in the production of supernatants capable of activating PMN for significant fungicidal activity (31.1 +/- 8.5%). These findings demonstrate for the first time a link between soluble factors produced by antigen stimulation of sensitized lymphoid cells and activation of PMN for enhanced microbicidal activity. Such a process defines an additional immune defense mechanism whereby the immune host may clear specific microorganisms.

Animals↗
PubMed1984

Estrogens inhibit mycelium-to-yeast transformation in the fungus Paracoccidioides brasiliensis: implications for resistance of females to paracoccidioidomycosis.

Evidence that disease due to the thermally dimorphic fungus Paracoccidioides brasiliensis occurs post-puberty predominantly in males led us to hypothesize that hormonal factors critically affect its pathogenesis. We show here that estrogens inhibit mycelial- to yeast-form transformation of P. brasiliensis in vitro. Transformation of three isolates was inhibited to 71, 33, and 19% of the control values in the presence of 10(-10), 10(-8), and 10(-6) M 17 beta-estradiol, respectively. The synthetic estrogen diethylstilbestrol was active but less potent than estradiol, whereas testosterone, 17 alpha-estradiol, tamoxifen, and corticosterone were inactive. This function was specifically inhibited, since yeast-to-mycelium transformation, yeast growth, and yeast reproduction by budding were unaffected by 17 beta-estradiol. Of note is the fact that mycelium-to-yeast transformation occurs as the first step in vivo in the establishment of infection. The cytosol of the three isolates studied possesses a steroid-binding protein which has high affinity for 17 beta-estradiol. We believe that this binding protein represents a P. brasiliensis hormone receptor which can also recognize mammalian estrogens. We hypothesize that the ability of estrogen to decrease or delay mycelium-to-yeast transformation at the initial site of infection contributes to or is responsible for the marked resistance of females, and that the binder described is the molecular site of action.

Cytosol↗
PubMed1984

Pseudallescheria boydii infections treated with ketoconazole. Clinical evaluations of seven patients and in vitro susceptibility results.

Seven patients infected with Pseudallescheria boydii were treated with oral ketoconazole, 200 to 600 mg/day for one to 13 months. Five patients had pulmonary infections; two had skeletal infections. Improvement of pretreatment abnormalities occurred in five patients, one of whom had concurrent arthrodesis of his infected knee. The other two patients were subsequently healed by surgical resection of their pulmonary lesions. Ketoconazole appeared less active than miconazole against 22 clinical isolates of P boydii when tested by two in vitro methods. We conclude that ketoconazole is effective treatment for some patients infected with P boydii, although this may not be predicted by current in vitro susceptibility tests. Further experience is needed to establish the optimal use of ketoconazole with respect to its dosage, duration of administration and concurrent surgical resection.

Aged↗
PubMed1984

High-dose ketoconazole therapy and adrenal and testicular function in humans.

Ketoconazole, an oral antifungal, when given in conventional doses, transiently blocks testosterone synthesis and adrenal response to corticotropin. Higher therapeutic doses (ie, 800 to 1,200 mg/day), even once daily, caused more prolonged blockade. In some men, the serum testosterone concentrations were always subnormal. Bound and free testosterone values were equally diminished. Oligospermia and azospermia after prolonged therapy were noted. Impotence and decreased libido were found. Gynecomastia appeared more common than with lower doses. Depressed response to corticotropin was pronounced. Urine cortisol excretion was depressed. The blockade appeared related to the serum ketoconazole concentration. Instances of normal hormone levels or responsiveness were associated with low ketoconazole concentrations. The hormonal effects were generally unrelated to duration of therapy, although there may have been partial reversal with continued therapy. These effects appeared reversible with discontinuation of therapy. Patients receiving ketoconazole should be considered potentially unable to mount an adrenal stress response and may require testosterone supplementation.

Adrenal Glands↗
PubMed1984

Chronic murine paracoccidioidomycosis: effect of ketoconazole on clearance of Paracoccidioides brasiliensis and immune response.

In a murine model of chronic pulmonary and disseminated paracoccidioidomycosis, ketoconazole (100 mg kg-1 in 0.3% agar) given by gavage twice daily for 1 or 2 months enabled all mice to clear disseminated Paracoccidioides brasiliensis from the spleen. Clearance of P. brasiliensis from the lungs was more difficult, and was achieved in 60% of the mice treated for 2 months. Sera from agar-treated control mice at days 77 and 103 post-infection demonstrated precipitating antibodies to P. brasiliensis antigens, but sera from ketoconazole-treated mice were precipitin-negative, indicating a favorable prognosis. Delayed hypersensitivity reactions to P. brasiliensis antigens in ketoconazole-treated mice were not significantly greater than in controls; consequently this test correlated less well with response than levels of serum antibody. This is the first use of this animal model of paracoccidioidomycosis to study the effect of antifungal drug protocols on the resolution of the disease. It also demonstrates the utility of this model in addressing clinically relevant questions about this disease and its treatment.

Animals↗
PubMed1984

Paracoccidioidomycosis in the immunosuppressed host: report of a case and review of the literature.

A case of reactivated paracoccidioidomycosis in an immunosuppressed host is described, the first association of this disease with renal transplantation. His presentation and clinical course was similar to the general experience with this mycosis. his response to ketoconazole was delayed when compared with that in non-immunosuppressed patients. Ketoconazole produced a dramatic resolution of all symptoms within 7 months. Serial determinations of paracoccidioides complement fixing antibodies showed falling titers, accurately reflecting his clinical status. Comparisons are made with other endemic mycoses in immunosuppressed patients. More cases of paracoccidioidomycosis in immunosuppressed patients can be expected to occur as immunosuppressive therapy becomes more common in areas endemic for Paracoccidioides brasiliensis, and with current trends in migration and travel.

Complement Fixation Tests↗
PubMed1984

Murine model of paracoccidioidomycosis. Production of fatal acute pulmonary or chronic pulmonary and disseminated disease: immunological and pathological observations.

A model of fatal acute pulmonary or chronic pulmonary and disseminated paracoccidioidomycosis was developed by intranasal challenge of young (3-4-week-old) mice with high doses (2.5-10 X 10(7) units) or low doses (0.1-1 X 10(7)) of yeast-phase Paracoccidioides brasiliensis. Fatal acute paracoccidioidomycosis was dose dependent; 10, 5, and 2.5 X 10(7) viable units of P. brasiliensis produced 100%, 66%, and 17% mortality, respectively, within 11 days. The pathologic picture was that of consolidation with a neutrophil infiltrate. Infection of adult (7-8-week-old) mice with even 10 X 10(7) units did not produce acute fatal paracoccidioidomycosis. The model of fatal acute paracoccidioidomycosis lends itself well to studies of therapeutic intervention. Increasing degrees of chronic pulmonary-disseminated paracoccidioidomycosis were produced by pulmonary infection of young mice with 0.1-1 X 10(7) viable units of P. brasiliensis. Although mice in all groups appeared healthy, pathology in lungs, liver, and spleen was evident at 7 and 10 weeks postinfection. Histopathologic observation revealed acinonodular granulomatous inflammation. At 12 weeks postinfection, there was evidence of less pathology, and of clearing of abscesses. In contrast to the pathology produced by 0.1 X 10(7) P. brasiliensis in young mice, no pathology evident on gross examination was produced by a similar dose in adult mice. Immunological evaluations of mice with chronic pulmonary-disseminated disease showed that spleen cells, but not lymph node cells, had significantly depressed blastogenic responses to concanavalin A (ConA) early after infection. However, at 12 weeks postinfection, when pathological examination indicated beginning resolution of infection, spleen cell responses to ConA were normal. The model of chronic disease is of interest for further immunological studies as well.

Acute Disease↗
PubMed1983

Ketoconazole binds to the intracellular corticosteroid-binding protein in Candida albicans.

Ketoconazole is a broad-spectrum, orally-active antifungal agent that has been shown to inhibit sterol synthesis in susceptible fungi. We have previously demonstrated the presence of an intracellular protein in several Candida species that binds mammalian corticosteroids with high affinity. In this paper we report that ketoconazole competitively displaces [3H]corticosterone from the Candida corticosteroid-binding protein at concentrations readily achieved in therapeutic settings. Ketoconazole was at least 50-100 times more potent than structurally related imidazole compounds. Additional data suggest, however, that the binding of ketoconazole and related drugs to this Candida protein is not critical for the in vitro antifungal activity of these drugs.

Binding, Competitive↗
PubMed1983

Experience with ketoconazole in three major manifestations of progressive coccidioidomycosis.

Forty-seven courses of oral ketoconazole therapy administered to 46 patients with progressive coccidioidomycosis were evaluated. Responses, of varying degrees evaluated by a point system, occurred in 81, 94, and 91 percent of courses evaluated for skeletal, chronic pulmonary, and cutaneous disease, respectively. Presumed side effects occurred in 26 percent of courses and were generally minor and reversible. Relapse occurred in 33, 33, and 11 percent of evaluable responding patients with skeletal, chronic pulmonary and cutaneous disease, respectively. From this experience, ketoconazole appears to be an important advance in the therapy of chronic coccidioidomycosis. Whether cure is possible will require further study and follow-up.

Adult↗
PubMed1983

Immunostimulation with muramyl dipeptide and its desmethyl analogue: Studies of non-specific resistance to pulmonary blastomycosis in inbred mouse strains.

N-acetyl-muramyl-L-alanyl-D-isoglutamine (MDP) protected against pulmonary blastomycosis when given prophylactically to BALB/c mice. Its desmethyl analogue (DM-MDP) had a similar effect. In C3H/HeJ, the effect was less marked. Early treatment after infection, with MDP and DM-MDP, had a modest effect in C3H/HeJ and BALB/c respectively, whereas late treatment had no effect in any mouse strain. No effect could be demonstrated with challenge sizes producing too lethal a model or minimal lethality, or in DBA/2J or young BALB/c mice. The effects in various strains do not correlate with differing effects on nonspecific immunostimulation in these strains. Immunostimulation with glycopeptides deserves further study in prophylaxis or therapy of fungal infection.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed1983

A cephalosporin active in vivo against Nocardia: efficacy of cefotaxime in murine model of acute pulmonary nocardiosis.

Cefotaxime, a cephalosporin drug, has been shown to be active in vitro against nocardiae, a finding confirmed in this study. Pharmacokinetic studies were performed in mice to define regimens which provided peak serum levels comparable to that achieved in man with currently used doses. These regimens were shown to be effective with only short courses of therapy of rapidly progressive and highly lethal N. asteroides infection, produced by pulmonary challenge of mice. This suggests the possible utility of this drug in human nocardiosis.

Acute Disease↗
PubMed1983

Estradiol binds to a receptor-like cytosol binding protein and initiates a biological response in Paracoccidioides brasiliensis.

Paracoccidioidomycosis, a disease caused by Paracoccidioides brasiliensis, which is endemic to Latin America, is much more common in men than women, suggesting a role for hormonal factors. We recently showed that two other yeasts possess steroid binding proteins and postulated that these receptor-like molecules represented a mechanism by which the hormonal milieu of the host might influence an infecting pathogen. Therefore, we examined P. brasiliensis for a sex steroid binding protein. Because tritiated steroids rapidly dissociated from the other fungal binding proteins, we developed a fast binding method with Sephadex G-50 microcolumns speeded by centrifugation. This method detected specific binding of [3H]estradiol in P. brasiliensis cytosol. Other tritiated steroid hormones, including testosterone and corticosterone, failed to exhibit specific binding. Scatchard analysis of [3H]estradiol binding showed an apparent dissociation constant (Kd) of 1.7 X 10(-8) M and a maximal binding capacity (Nmax) of 235 fmol/mg of protein. Susceptibility to trypsin indicated the binding site was protein in nature. The protein had a Stokes radius of approximately equal to 32 A by HPLC exclusion column and a sedimentation coefficient of 4.4 S by sucrose gradient, consistent with an apparent Mr of approximately equal to 60,000. Competition experiments revealed that estrone, estriol, and progesterone had 25% of the affinity of estradiol, whereas diethylstilbestrol, androgens, and corticosteroids had low affinity. Investigation of steroid hormone actions in P. brasiliensis indicated that estradiol inhibited the fungal transformation from mycelial form to yeast form, the initial step of infection. This suppressive effect was dose-dependent and not found with testosterone. We hypothesize that endogenous estrogens in the host, acting through the cytosol binding protein in the fungus, inhibit mycelial-to-yeast transformation, thus explaining the resistance of women to paracoccidioidomycosis.

Binding, Competitive↗
PubMed1983

Correlation of in vitro susceptibility test results with in vivo response: flucytosine therapy in a systemic candidiasis model.

The in vitro susceptibility of Candida albicans isolates to flucytosine was compared to therapeutic effect in experimental murine candidiasis (candidosis). Four groups of 10 isolates were chosen, based upon their broth dilution minimal inhibitory concentrations (MICs), from a group of 402 isolates from patients without prior flucytosine therapy. Group I MICs were less than 12.5 micrograms/ml after seven days, whereas group II, III, and IV MICs exceeded 12.5 micrograms/ml on days 7, 2, and 1, respectively. Pilot experiments selected challenge inocula of similar virulence. Mice were infected intravenously and given various flucytosine doses. Significant prolongation of survival correlated with MICs and with agar disk-diffusion zone diameters (P less than 0.05). In vivo response to therapy was more favorable for group I isolates compared with group IV isolates (P less than 0.01). The present study demonstrates in this animal model that in vitro susceptibility does correlate with in vivo response to therapy, although exceptions occur with individual isolates.

Animals↗
PubMed1983

An evaluation of recently developed antibiotics.

A number of newer antibiotics, broad-spectrum penicillins and cephalosporins, have been evaluated against Gram-negative rods. The organisms were selected for multi-resistance and transferable resistance factors. None of the broad-spectrum penicillins was much use against most of the organisms. Ceftriaxone, cefotaxime, latamoxef (moxalactam) and N-formimidoyl thienamycin were all highly effective against most multi-resistant Gram-negative bacilli; cefoperazone being inferior to them. Enterobacter and Serratia strains were relatively resistant to all the agents mentioned and against Acinetobacter only N-formimidoyl thienamycin showed much activity. Thus, use of these drugs may increase the proportion of infections due to organisms such as Serratia or Acinetobacter.

Acinetobacter↗
PubMed1983

Distribution of a corticosteroid-binding protein in Candida and other fungal genera.

Using [3H]corticosterone as a probe, corticosteroid-binding protein (CBP) was detected in eight out of eight isolates of Candida albicans, of both A and B serotypes. The apparent dissociation constant (Kd) in the various isolates ranged between 8 and 19 nM; the binding capacity varied from 122 to over 2400 fmol (mg cytosol protein)-1. There was no correlation between the amount or affinity of CBP and isolate virulence for murine hosts. Further analysis revealed demonstrable CBP in six out of six Candida species other than C. albicans. One isolate of C. tropicalis has been identified which fails to bind [3H]corticosterone. Saccharomyces cerevisiae, Neurospora crassa and Paracoccidioides brasiliensis also failed to bind [3H]corticosterone. Preliminary attempts were made to determine functions mediated by CBP in Candida, but in vitro growth, phase conversion and glucose oxidation by Candida were unaffected by the addition of a variety of steroid hormones. These data indicate that the presence of CBP in Candida does not correlate with either virulence or serotype. The physiological significance of CBP remains to be determined.

Candida↗
PubMed1983

Ketoconazole inhibition of testicular secretion of testosterone and displacement of steroid hormones from serum transport proteins.

In vivo perfusion of canine testes with ketoconazole inhibited the stimulation of testosterone production by human chorionic gonadotropin in a dose-dependent manner. Ketoconazole also selectively displaced steroids from serum-binding globulins. Dihydrotestosterone and estradiol binding to sex hormone-binding globulin were inhibited by ketoconazole. Cortisol binding to corticosteroid-binding globulin was unaffected. The concentrations of ketoconazole that inhibited human chorionic gonadotropin stimulation of testicular androgen production and displaced sex steroids from sex hormone-binding globulin were in the range of blood levels found in patients on higher therapeutic dosage regimens. Suppression of testicular testosterone synthesis and displacement of estrogens from sex hormone-binding globulin may decrease the androgen/estrogen ratio of the blood and contribute to the development of gynecomastia that has been reported in some ketoconazole-treated patients.

Animals↗