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P D Walzer

Publications and source records attributed to P D Walzer.

At least 37 records · Page 2Linked to original sources
PubMed1995

Genetic variation among Pneumocystis carinii hominis isolates in recurrent pneumocystosis.

Pneumocystis carinii hominis is a ubiquitous organism that causes pneumonia in immunocompromised persons. Paired P. carinii hominis isolates from human immunodeficiency virus-infected persons who had two episodes of pneumocystosis were examined for genetic heterogeneity. Genetic variation was detected by sequence comparison of a portion of the mitochondrial ribosomal RNA gene. In 5 of 10 patients experiencing two episodes of pneumocystosis, genetically distinct isolates were associated with each episode. These included 4 of 6 patients whose second episode of pneumocytosis occurred > 6 months after their initial bout. The genetic data support the hypothesis that some recurrent episodes of P. carinii hominis pneumonia are caused by reinfection rather than by reactivation of latent infection.

AIDS-Related Opportunistic Infections↗
PubMed1995

Adoptive transfer of lymphocytes sensitized to the major surface glycoprotein of Pneumocystis carinii confers protection in the rat.

Pneumocystis carinii is a major opportunistic pathogen and a leading cause of morbidity in patients with AIDS. CD4+ cells have been shown to be important in host defenses against P. carinii, but the antigen(s) involved with this response have not been identified. We undertook the present study to determine whether the major surface glycoprotein (MSG) of P. carinii contains epitopes that can elicit a protective cellular immune response. Spleen cells and purified CD4+ cells isolated from Lewis rats, pulsed 1-4 d with MSG, and injected into corticosteroid-treated Lewis rats with pneumocystosis resulted in significant reduction in the P. carinii burden, as judged by organism quantitation and lung histology. The protective response demonstrated by the donor cells was dependent on previous exposure to P. carinii, cell concentration, and time of incubation with MSG. In addition, reconstitution with MSG-specific CD4+ cells resulted in an early hyperinflammatory response within the lungs of these animals with a high percentage of mortality. Thus, in this model, MSG can elicit an immune response mediated by CD4+ cells, which has a harmful as well as helpful effect on the host, and these responses occur despite the presence of corticosteroids.

Animals↗
PubMed1994

Fractionation of Pneumocystis carinii developmental stages by counterflow centrifugal elutriation and sequential filtrations.

Immunomagnetic sorting, sequential filtrations, and counterflow centrifugal elutriation were compared for their ability to obtain enriched populations of Pneumocystis carinii developmental stages from infected rat-lung homogenates. Elutriation combined with sequential filtrations resulted in highly (> 95%) enriched populations of P. carinii cysts and trophozoites with excellent viability. This approach offers advantages over previously described methods of obtaining enriched P. carinii cell populations and should have important applications to research on this organism.

Animals↗
PubMed1994

Characterization of multiple unique cDNAs encoding the major surface glycoprotein of rat-derived Pneumocystis carinii.

The major surface glycoprotein (MSG) of rat-derived Pneumocystis carinii represents a group of related molecules that are encoded by multiple genes. We isolated seven unique MSG cDNAs from a library prepared from a single infected rat lung. The cDNAs displayed both conserved and variant regions to previously described cDNAs. These clones contained inserts that ranged in size from 0.4 to 1.8 kb and all contained a poly(A) tail. The largest clone, Pc1410, hybridized to all 15 chromosomes resolved by pulsed-field gel electrophoresis (PFGE). Protein produced by in vitro translation from Pc1410 was immunoprecipitated with affinity-purified MSG antibodies. The clones were characterized by DNA sequencing of their 3' and 5' ends. Analysis of the untranslated and coding regions demonstrated that the clones contained unique and conserved regions of sequence, but none of the clones were identical. Isolation of seven additional unique clones picked from a single screening of a cDNA library suggests that numerous MSG transcripts exist within a population of P. carinii.

Amino Acid Sequence↗
PubMed1994

Analysis of Pneumocystis carinii organism burden, viability and antigens in bronchoalveolar lavage fluid in AIDS patients with pneumocystosis: correlation with disease severity.

OBJECTIVES: We examined 96 bronchoalveolar lavage fluid (BALF) specimens from AIDS patients with proven Pneumocystis carinii pneumonia (PCP) in order to compare the relationship of organism burden, viability and antigen expression with disease severity at the time of clinical presentation. METHODS: Tinctorial analysis of BALF specimens with proven PCP using Diff-Quik, cresyl echt violet and erythrosin B stains to evaluate organism burden and viability. P. carinii antigen examination was performed by Western blot analysis. RESULTS: P. carinii cluster ratios were more sensitive than cyst counts as an indicator of organism burden, and correlated well with the alveolar-arterial oxygen gradient as a measure of disease severity. Erythrosin B, the vital stain used to measure P. carinii viability, displayed a wide range of values and provided little useful information. Antigens of 35-45 and 95kD, which were specific for P. carinii, were found by immunoblot analysis in BALF cellular fraction of most patients with pneumocystosis. By contrast, antigens of 52 and 66 kD, which were found in both BALF supernatant and cellular fractions of P. carinii patients and controls, most likely represented albumin and immunoglobulin G heavy chain, respectively, of host origin. The 35-45 kD antigen was found in 88% of the BALF specimens and appeared to represent an important marker of P. carinii infection. The 95 kD antigen was detected in 49% of the specimens. CONCLUSIONS: We conclude that analysis of P. carinii characteristics in BALF specimens of patients with pneumocystis may provide additional information. These data will also be helpful in developing more sensitive assays and in targeting specific P. carinii factors for future investigation.

AIDS-Related Opportunistic Infections↗
PubMed1994

Guanylhydrazones in therapy of Pneumocystis carinii pneumonia in immunosuppressed rats.

Guanylhydrazones are cationic heteroaromatic drugs similar to the diamidines which are effective in the treatment of African trypanosomiasis and pneumocystosis. On the basis of their antitrypanosomal activity, different guanylhydrazones were selected for evaluation in a rat model of Pneumocystis carinii pneumonia. The most active compounds were the 2-(4'-formylphenyl)-1-methylimidazo-[1,2-a] pyridinium guanylhydrazones which, at a dose of 2 mg/kg/day, were about as effective as trimethoprim-sulfamethoxazole at a dose of 50 mg of trimethoprim per kg/day plus 250 mg of sulfamethoxazole per kg/day. The anti-P. carinii activity of these guanylhydrazone derivatives was found with parenteral but not with oral administration. The 1,3-arylene diketone bis(guanylhydrazones) were generally ineffective, although a triacetyl derivative showed some anti-P. carinii activity. Nitroimidazole guanylhydrazone derivatives were also ineffective. Attempts to improve the therapeutic efficacy of the different guanylhydrazones were limited by problems of toxicity. We conclude that some guanylhydrazone derivatives are potent anti-P. carinii drugs and that further studies should be pursued to develop safer compounds and investigate structure-activity relationships.

Animals↗
PubMed1994

Cellular responses to a 55-kilodalton recombinant Pneumocystis carinii antigen.

The host-parasite interaction in Pneumocystis carinii pneumonia is poorly understood. In recent years, two major groups of P. carinii antigens have been identified. One class of antigens is characterized by a broad band of immunoreactivity between 45 and 55 kDa in P. carinii derived from rats. This antigen complex is the P. carinii antigen most commonly found in respiratory tract specimens and most frequently recognized by the host immune response. The availability of a recombinant antigen has permitted studies focusing on the cellular and humoral responses to a single antigen within this class, p55. In this study, we have demonstrated that the p55 antigen elicits a cell-mediated immune response in animals previously exposed to P. carinii. Under conditions of natural exposure, the 5' portion of the molecule, p55(1-200), appears immunologically silent, failing to elicit lymphocyte proliferation or cytokine secretion. Following active immunization, the 5' portion is capable of stimulating lymphocyte proliferation. The 3' portion, p55(268-414), has at least one immunodominant region which contains a 7-amino-acid repeat motif. The cells responding to p55 include a CD4+ T cell which secretes a Th1 cytokine pattern. A detailed understanding of the host-parasite interaction will facilitate the development of immunoprophylaxis and immunotherapy for P. carinii infection.

Amino Acid Sequence↗
PubMed1993

Geographic variation in the humoral response to Pneumocystis carinii.

The reported incidence of Pneumocystis carinii pneumonia varies in different areas of the world, but little is known about geographic variation in the antibody response to specific antigens. The frequency of anti-P. carinii antibodies in the serum of normal and human immunodeficiency virus (HIV)-infected individuals from five different regions was compared. Serum specimens from 948 subjects were assayed for IgG antibodies to human-derived P. carinii by Western blot. The overall prevalence of anti-P. carinii antibodies was 72.9%. Among HIV-seronegative individuals, the rate of seropositivity was similar in all regions (70.5%-82.4%). Antibodies to the 30- to 40-kDa antigen were most commonly detected. The frequency of antibodies to high-molecular-mass antigens (95, 120, and > 120 kDa) demonstrated significant geographic variation. This study demonstrates that antibody responses to P. carinii are common in all areas studied but vary in frequency and pattern.

AIDS-Related Opportunistic Infections↗
PubMed1993

A 55 kDa antigen of Pneumocystis carinii: analysis of the cellular immune response and characterization of the gene.

Rat-derived Pneumocystis carinii contains a major antigen complex of 45-55 kDa. The fusion protein of a cDNA encoding the 3' portion of the 55kDa antigen, which had previously been shown to be recognized by serum antibodies of exposed subjects, was investigated for its ability to stimulate a cellular immune response. Rats exposed to P. carinii via the environment exhibited a vigorous proliferative response to the antigen whereas unexposed rats did not. The full-length cDNA for a 55kDa antigen was cloned and found to contain a 1245bp open reading frame capable of encoding a 414-amino-acid peptide. The gene encoding this protein contained a single 39bp intron and transcribed a 1.45kb RNA message. The cloning and characterization of the 55kDa antigen gene will allow production of the specific immunological reagents necessary to characterize this molecule and study its role in the biology and pathogenesis of P. carinii.

Amino Acid Sequence↗
PubMed1993

Synergistic combinations of Ro 11-8958 and other dihydrofolate reductase inhibitors with sulfamethoxazole and dapsone for therapy of experimental pneumocystosis.

We compared Ro 11-8958, an analog of trimethoprim (TMP) with improved antimicrobial and pharmacokinetic properties, other dihydrofolate reductase (DHFR) inhibitors, sulfamethoxazole (SMX), and dapsone (DAP) in the treatment of Pneumocystis carinii pneumonia in an immunosuppressed rat model. In contrast to previous reports, high dosages of the DHFR inhibitors were used in combination with fixed, low dosages of SMX (3 mg/kg of body weight per day) or DAP (25 mg/kg/day). When administered alone at these dosages, SMX and DAP reduced the median P. carinii cyst count about 5- to 15-fold. Ro 11-8958, TMP, and diaveridine used at a dosage of 20 mg/kg/day with SMX were only slightly more effective than SMX used alone. However, administration of these DHFR inhibitors at a dosage of 100 mg/kg/day with SMX lowered the cyst count about 500- to 1,000-fold, indicating a synergistic effect. Little or no synergism was found when other DHFR inhibitors (pyrimethamine, cycloguanil, and tetroxoprim) were combined with SMX. Regimens of Ro 11-8958 at a dosage of 20 mg/kg/day with DAP and of TMP or diaveridine used at a dosage of 100 mg/kg/day with DAP showed comparable anti-P. carinii activity, lowering the cyst count 100- to 200-fold. By contrast, Ro 11-8958 administered at a dosage of 100 mg/kg/day with DAP reduced the cyst count > 1,000-fold. Thus, the experimental approach used here enables the rat model of pneumocystosis to be used to compare synergistic combinations of antifolate drugs. The favorable results achieved with Ro 11-8958 indicate that it should be considered for clinical trials.

AIDS-Related Opportunistic Infections↗
PubMed1993

Proliferative and cytokine responses to a major surface glycoprotein of Pneumocystis carinii.

Naturally derived T-cell responses by rats to a 120-kDa major surface glycoprotein (MSG) of rat-derived Pneumocystis carinii were analyzed in vitro. Specific cytokines elicited by the T-cell response to the MSG were also identified. MSG was purified from rat-derived P. carinii by three different techniques: lectin affinity chromatography, sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by electroelution, and size-exclusion high-performance liquid chromatography. The cell-mediated immunity of spleen cells isolated from Lewis rats with and without natural exposure to P. carinii to the purified MSG was studied. Exposure to P. carinii was monitored by the presence or absence of serum antibodies to P. carinii antigens by Western blotting (immunoblotting). A T-cell proliferative response to the MSG was identified only with spleen cells isolated from rats exposed to P. carinii and peaked at 4 days. Flow cytometric analysis revealed that the percentage of CD4 cells was significantly increased during the proliferative response to MSG. MSG also elicited secretion of tumor necrosis factor alpha, interleukin-1, and interleukin-2, with peak activity of these cytokines occurring after 12, 24, and 48 h, respectively, of culture. These findings suggest that MSG is important in host T-cell recognition of and immune response to P. carinii by recruitment of inflammatory cells and cytokine production.

Animals↗
PubMed1993

Regulation of surfactant phosphatidylcholine secretion from alveolar type II cells during Pneumocystis carinii pneumonia in the rat.

We used an immunosuppressed rat model to test the hypothesis that normal mechanisms regulating surfactant phosphatidylcholine synthesis and secretion in alveolar type II cells are aberrant in Pneumocystis carinii pneumonia. Animal groups included: group 1, healthy controls; group 2, immunosuppressed, without pneumocystosis; group 3, immunosuppressed with pneumocystosis; group 4, immunosuppressed with well-established pneumocystosis treated with trimethoprim-sulfamethoxazole (TMP-SMX). Type II cells were isolated from rats in each group and compared for [3H]choline incorporation into phospholipid and response of the type II cells to secretagogues. Incorporation of [3H]choline into phospholipid subclasses exhibited significant differences. Incorporation into phosphatidylcholine fell from 89.3 +/- 2.2% of total incorporation in group 1 control rats to 79.6 +/- 3.1% in group 3 rats with P. carinii pneumonia, while incorporation into sphingomyelin rose from 5.6 +/- 1.2% in group 1 animals to 15.2 +/- 2.7% in group 3 rats. Incorporation of [3H]choline into phospholipid subclasses in cells from group 2 and group 4 animals was not different from incorporation for group 1 animals. Type II cells from group 1 and group 2 (immunosuppressed control) rats responded appropriately to the secretagogues ATP, TPA, and terbutaline with a marked increase in surfactant phosphatidylcholine secretion; the effect of ATP was also blocked by the lectin, concanavalin A. In contrast, type II cells from group 3 rats failed to respond to the secretagogues with a significant increase in phospholipid secretion. Although treatment of group 4 rats with TMP-SMX markedly reduced the P. carinii organism burden, type II cells from these animals also responded poorly to the secretagogues. The depressed type II cell function described here provides a mechanism for the observed decrease in surfactant phospholipids from bronchoalveolar lavage fluid of experimental animals and patients with P. carinii pneumonia. The data also suggest this defect may become irreversible with advanced disease.

Animals↗
PubMed1992

Isolation of Pneumocystis carinii cysts by flow cytometry.

Pneumocystis carinii cysts were separated as enriched populations from suspensions of lung homogenate obtained from infected rats containing all developmental stages of this organism. Isolation of cyst populations was achieved by incubating filtered lung homogenate with the fluorescent phospholipid analog 1-palmitoyl-2-C6-NBD-phosphatidylcholine. Whereas cysts did not fluorescence as a result of outer-wall restraint of lipid integration, trophozoites and young intermediate stages readily incorporated the fluorescently labeled lipid analog into their outer membrane. The two distinct labeling patterns displayed by cysts and other developmental phases of P. carinii constitute a novel, easy, and reproducible means of isolating cysts from infected lung homogenate by flow cytometry.

Animals↗
PubMed1992

Activities of antifolate, antiviral, and other drugs in an immunosuppressed rat model of Pneumocystis carinii pneumonia.

The efficacy of antifolate, antiviral, and other drugs was compared in an experimental model of pneumocystosis. Sulfamethoxazole (SMX) administered alone in doses of greater than or equal to 60 mg/kg/day was highly effective in treatment and prophylaxis. Low (less than or equal to 15 mg/kg/day) doses of SMX showed limited, dose-related anti-Pneumocystis carinii activity in therapy but were more effective in prophylaxis. The dihydrofolate reductase (DHFR) inhibitors trimethoprim (TMP), pyrimethamine, and trimetrexate exhibited little anti-P. carinii activity when administered alone and did not enhance the efficacy of SMX; the effects of the DHFR inhibitors could not be related to the dose or the concentration in serum. These data suggested that the rat model is an excellent system for studying the anti-P. carinii activity of sulfonamides but is of limited value in studying DHFR inhibitors. The antiviral drugs azidothymidine, dideoxyinosine, inosine pranobex (Isoprinosine), amantadine, and acyclovir displayed little or no activity against P. carinii; however, azidothymidine did not impair the efficacy of SMX or TMP-SMX. These results supported the clinical practice of giving antiviral agents together with antifolate drugs to patients infected with human immunodeficiency virus and suggested that the beneficial effects of antiviral agents on the occurrence of pneumocystosis are due mainly to their effects on the virus or the host immune response. In contrast to the antiviral drugs, 9-deazainosine, a nucleoside analog with antiprotozoal properties, demonstrated marked activity against P. carinii which was related to dose and route of administration. These data raised the possibility that anti-P. carinii activity is a general property of purine nucleosides and suggested that further exploration of this class of compounds might lead to clinically useful agents.

Animals↗
PubMed1992

Treatment of experimental pneumocystosis: review of 7 years of experience and development of a new system for classifying antimicrobial drugs.

Over a 7-year period, we analyzed 261 dose regimens of antimicrobial drugs in the treatment and prevention of Pneumocystis carinii pneumonia in an immunosuppressed rat model. These compounds ranged from drugs in clinical use to newly synthesized agents. Drug efficacy was expressed as the magnitude of the reduction in median P. carinii cyst or nucleus counts on a scale ranging from inactive (less than 5-fold) to very markedly active (greater than or equal to 1,000-fold). The classification system was reproducible and allowed drugs studied at different times to be compared with each other. The system demonstrated a hierarchy of anti-P. carinii activity not only among classes of compounds but also among individual members of a drug class. Sulfonamides, sulfones, and diamidines were the most active agents; some purine nucleosides and nitrofurans also showed promising activity; and most antiparasitic, antifungal, antibacterial, and antiviral drugs were inactive. We conclude that this classification system represents a simple, quantitative method of comparing the activities of antimicrobial drugs against P. carinii. Information gained from this system should be helpful in developing new anti-P. carinii compounds and establishing standard procedures for their evaluation.

Animals↗
PubMed1992

Isolation and characterization of a recombinant antigen of Pneumocystis carinii.

Pneumocystis carinii contains a major group of antigens which migrates as a broad band of 45 to 55 kDa and 35 to 45 kDa in organisms derived from rats and humans, respectively. This complex is among the most common P. carinii antigens found in the respiratory tract and is recognized by serum antibodies of infected individuals. We have isolated a cDNA clone encoding the 3' portion of a 45- to 55-kDa antigen of rat-derived P. carinii. The predicted protein encoded by this cDNA contains a distinctive domain composed of 10 copies of a 7-amino-acid sequence motif rich in glutamic acid residues. Affinity-purified antibodies to this peptide reacted with the 45- to 55-kDa band of rat-derived P. carinii and with the 35- to 45-kDa band of human-derived P. carinii, indicating shared epitopes. The fusion protein was recognized by serum antibodies from rats and humans with natural exposure to P. carinii and by human immunodeficiency virus patients with P. carinii pneumonia. The production of this recombinant protein should allow more detailed studies of the host-parasite relationship of this important opportunistic infection.

Amino Acid Sequence↗