Biomedical subjects

M Strand

Publications and source records attributed to M Strand.

At least 55 records · Page 3Linked to original sources

PubMed1992

Methyl inosine monophosphate: a potential immunotherapeutic for early human immunodeficiency virus (HIV) infection.

MIMP is a new thymomimetic purine under development for immunorestorative therapy. Lymphocytes were obtained from eight patients with acquired immunodeficiency disease (AIDS), eight with symptomatic pre-AIDS (ARC), and 22 normal controls and were stimulated in vitro with phytohemagglutinin (PHA). AIDS patients (mean CD4 counts of 40) showed PHA responses less than 10% of control while ARC patients (mean CD4 counts of 544) showed responses approximately 50% of the control responses. MIMP (0.1, 1, 10 and 100 micrograms/ml) progressively augmented the PHA responses in all these groups. The augmentation of the responses of the leukocytes of AIDS patients while statistically significant was minimal. The augmentation of the responses of ARC patients was significant and their maximal responses approached control levels. The effect of 1 micrograms/ml MIMP was comparable with that observed with indomethacin (10(-6) M) and interleukin-2 (IL2 - 4 units/ml) and was additive with each of these stimulants. In a parallel manner, MIMP restored the suppression of control lymphocytes induced by the immunosuppressive 17 amino acid fragment of the P41 peptide of HIV. In vivo experiments showed that MIMP significantly delayed death in a murine FLV AIDS model at a dose of 1 mg/kg by the oral or parenteral route. MIMP is under preclinical development for early HIV disease to forestall progression to AIDS by attenuating virus-induced immunosuppression.

Adjuvants, Immunologic↗

PubMed1992

Circulating schistosomal antigen in diagnosis and assessment of cure in individuals infected with Schistosoma mansoni.

The effectiveness of praziquantel in treating schistosomiasis is most commonly assessed by quantitating egg production or anti-schistosome antibodies in serum. We have used a monoclonal antibody (MAb)-based antigen-capture enzyme-linked immunosorbent assay (ELISA) for the serologic diagnosis of schistosomiasis, and to monitor the efficacy of praziquantel therapy in 49 individuals with parasitologically proven schistosomiasis. The MAb used, 128C3/3/21, recognizes a repeating carbohydrate epitope expressed at all stages of parasite development, and antibodies recognizing this epitope are found in the serum of infected humans. The overall sensitivity of the ELISA was 78%, with a sensitivity of 100% for patients excreting greater than 100 eggs/g of feces and 72% for those excreting less than 100 eggs/g of feces. The positivity of the ELISA was directly related to the fecal egg counts obtained on days -3, -2, and -1 before treatment with praziquantel, but there was no correlation between antigen levels and the clinical stage of the disease. After praziquantel treatment, we observed a highly significant correlation (P less than 0.0001) between the time elapsed since treatment and the decrease in antigenemia. Furthermore, although no eggs were detected in any of the stool specimens at week 12 after treatment, the antigen was detected in 21% of the treated patients (seven of 33 ELISA-positive patients). Antigen levels decreased over the 12-week period in six of these patients, whereas the antigen level increased with time in one individual. The persistence of antigenemia suggests that these individuals are either still clearing antigen or remain infected.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1991

A synthetic peptide with sequence identity to the transmembrane protein GP41 of HIV-1 inhibits distinct lymphocyte activation pathways dependent on protein kinase C and intracellular calcium influx.

A synthetic peptide containing env amino acid (aa) sequence 581 to 597 of the transmembrane protein gp41 of human immunodeficiency virus type 1 (HIV-1) was tested for its effect on protein kinase C (PKC) and cytoplasmic free Ca2+ [( Ca2+]i) influx-dependent immune functions. We have previously shown that this peptide inhibits PKC-mediated phosphorylation and T-cell receptor-mediated [Ca2+]i influx as well as lymphoproliferation. In this study we demonstrate that the HIV-1 gp41 peptide aa581-597 inhibits lymphoproliferation stimulated via the distinct T-cell-activation molecules CD3, CD2, and CD28, as well as direct stimulation mediated by phorbol ester combined with ionomycin. Further, aa581-597 inhibits both PKC-dependent interleukin 2 (IL 2) production and the [Ca2+]i influx-dependent but PKC-independent induction of IL 2 receptor expression. The HIV-1 gp41 peptide also induces dramatic morphologic changes in lymphocytes, characterized by cytoplasmic ballooning and the acquisition of adherence to plastic, and these changes are dependent on both the length and the temperature of exposure. The results of this study suggest that the HIV-1 gp41 sequence aa581-597 acts at multiple sites to inhibit both PKC activity and [Ca2+]i influx, resulting in the abrogation of several distinct immune functions that are critical for an intact immune response and are defective in HIV-1-infected individuals.

Antigens, CD↗

PubMed1991

Characterization of cDNA clones encoding a novel calcium-activated neutral proteinase from Schistosoma mansoni.

To identify and characterize Schistosoma mansoni proteins that are recognized by infected hosts, we have used a pool of sera from infected humans to screen cDNA libraries constructed from poly(A)+ mRNA of adult S. mansoni. The deduced amino acid sequences of the three isolated clones showed a high degree of similarity to the large subunit of calcium-activated neutral proteinase (CANP) from humans and chicken. These overlapping clones, which include a nearly full-length clone with an open reading frame of 758 amino acid residues, together encode the entire large subunit of CANP. The deduced sequence of this S. mansoni protein can be divided into four domains (I-IV) that include the two domains characteristic of other large subunits of CANP: a thiol-protease domain (II) and a calcium-binding domain (IV) containing EF hand motifs. However, the schistosome protein is unique in having only three EF hand motifs in the calcium-binding domain and in having an additional EF hand motif that is shared between domains II and III. We have shown that these EF hand motifs are capable of binding 45Ca2+. Furthermore, the large subunit is S. mansoni contains an NH2-terminal sequence of 28 residues that is absent from the mammalian CANPs and has a high degree of similarity to the presumed receptor binding sequence of colicin Ia and Ib.

Amino Acid Sequence↗

PubMed1991

Characterization of a conserved T cell epitope in HIV-1 gp41 recognized by vaccine-induced human cytolytic T cells.

A human CTL epitope located in a region of the HIV-1 envelope protein gp41 that is highly conserved among various HIV-1 strains was identified. This epitope was recognized by CD4+ CTL clones that were induced in seronegative humans by immunization with recombinant gp160. Fusion proteins carrying portions of the HIV-1 env gene and synthetic peptides were used to localize this epitope to amino acids 584-595 of the HIV-1 BRU env sequence. Only two positions within this epitope showed variation among North American HIV-1 isolates, and the substitutions were conservative in nature. The Lys to Arg substitution at position 593 abolished recognition, probably by interfering with the peptide-MHC interactions. This epitope was recognized in association with at least one subtype of the widely distributed human class II MHC specificity DPw4, namely DPw4.2. The relatively high frequency of this allele (27.2% among Caucasians) makes it likely that a larger fraction of the population would generate a response directed at this epitope than would be the case for epitopes recognized in the context of gene products of most other class II and class I loci. Interestingly, the closely related DP beta-chain allele types 4.1 and 2.1, which differ from 4.2 by 3 and 1 amino acids, respectively, were unable to present this gp41 peptide to DPw4.2-restricted clones. Comparison of the structure of this epitope with that of other peptides recognized in the context of DPw4.2 led to the identification of a consensus sequence for DPw4.2 binding peptides. Because the gp41 CTL epitope 584-595 identified here is highly conserved and is recognized in the context of a common DP allele, it may represent an important target region for vaccine development. Our results indicate that vaccines containing this epitope may induce in a significant fraction of those immunized CTL active against at least half of all HIV-1 strains.

Amino Acid Sequence↗

PubMed1991

Human respiratory mucosa in a nonadhesive stationary organ culture system.

Fragments of human adenoid tissue were transferred to a nonadhesive, stationary organ culture system. The culture period was 40 days. In culture, beating cilia could be observed at the surface of the fragments. Light microscopy, scanning electron microscopy, and transmission electron microscopy showed that the tissue fragments were covered by a multilayered, pseudostratified, ciliated epithelium. Beneath the epithelium was a basement membrane. At the start of the culture period, the central parts of the fragments were dominated by lymphocytes. These lymphocytes gradually disappeared and were replaced by a collagen-containing stroma with scattered fibroblasts. The tissue fragments can be used as an organ culture model for normal respiratory mucosa.

Adenoids↗

PubMed1991

Selective release of a glycosylphosphatidylinositol-anchored antigen from the surface of Schistosoma mansoni.

We have recently demonstrated that a 200-kDa antigen that serves as a target of antibodies acting in synergy with praziquantel is linked to the surface membrane of Schistosoma mansoni by a glycosylphosphatidylinositol (GPI) anchor. In the present study we have examined the potential role of this GPI anchor in the therapeutic action of praziquantel by monitoring the release of surface antigens from living adult schistosomes cultured in the presence or absence of praziquantel and exogenous phospholipases. Phosphatidylinositol-specific phospholipase C (PIPLC) selectively released the 200-kDa antigen from the surface of adult schistosomes, as determined by immunoprecipitation experiments; none of the other GPI-anchored proteins, including alkaline phosphatase and a 22-kDa protein, were released by this enzyme. Anti-cross-reacting determinant antiserum (anti-CRD), which recognizes an epitope on GPI-anchored proteins only after the anchor has been removed by PIPLC, specifically precipitated the 200-kDa antigen, confirming the cleavage of its anchor. When the worms were exposed to both praziquantel and PIPLC, the amount of 200-kDa cleaved from the worms was increased five-fold. The selective release of this antigen was also detected by indirect immunofluorescent labeling of praziquantel-exposed adult worms cultured in the presence of phospholipases. Taken together these observations suggest that modulation of the phospholipase-mediated release of GPI-anchored antigens by praziquantel may contribute to the therapeutic action of the drug.

Animals↗

PubMed1991

Characterization of proteins and immunogens released by adult Schistosoma mansoni.

The proteins released in vitro by metabolically radiolabeled adult Schistosoma mansoni were identified by 2-dimensional gel electrophoresis. To determine the origin of these proteins, adult worms were fractionated into surface membrane, tegument, and remaining body components, and the electrophoretic patterns of the proteins in the 3 fractions were compared to those of the released proteins. The immunogens present in these fractions then were identified by immunoprecipitation with sera from humans infected with S. mansoni. This analysis indicated that essentially all of the proteins released from the worm were immunogenic, whereas most of the major membrane and tegumental proteins were not reactive with the immune sera. Thus, it appears that the adult worm is defended against immune attack by detection of the host's antibody response against released proteins rather than against proteins-exposed on the worm's surface.

Animals↗

PubMed1990

Preparation of proteosome-based vaccines. Correlation of immunogenicity with physical characteristics.

In order to facilitate the use of proteosome-based vaccines, we have identified and analyzed the parameters that affect their immunogenicity. As a model system we used synthetic peptides (LCF6) containing sequences from the immunodominant (NANP)n tandem repeat region of the P. falciparum circumsporozoite protein, hydrophobically complexed to multimeric protein preparations (proteosomes) of meningococcal outer membrane proteins (OMP), since we have previously shown that high levels of anti-(NANP)n IgG can be elicited in mice by use of this novel adjuvant system (Lowell et al., 1988a). We have now examined these preparations by velocity sedimentation and measured their ability to elicit an IgG response in mice. Velocity sedimentation of freshly mixed OMP and LCF6, without dialysis, produced a limited number of small complexes, whereas dialysis of the mixture for 4 d yielded heterogeneously sized complexes that became more homogeneous when the dialysis was carried out for 7 or 10 days. The most homogeneous of these peptide-proteosome complexes (those dialyzed for 10 days) induced substantial levels of anti-(NANP)n IgG in mice, and shorter periods of dialysis resulted in vaccines that induced proportionately lower titers. Analysis of a series of preparations with varying LCF6: OMP ratios (w/w) showed that the degree of peptide substitution of the proteosomes was inversely proportional to the rate of sedimentation of the complexes and that there exists an optimal degree of lipopeptide complexing to the proteosomes. Our results suggest that the parameters affecting the immunogenicity of the peptide-proteosome complexes are: (i) hapten density, and (ii) size of the complex. Furthermore, sedimentation analysis of peptide-proteosome immunogens may serve as a rapidly performed assay of immunogenic potency.

Adjuvants, Immunologic↗

PubMed1990

Improved in vivo stability and tumor targeting of bismuth-labeled antibody.

We have used a series of bifunctional chelating agents to prepare 206Bi-labeled monoclonal antibody and have assessed the in vivo stability and tumor targeting of these conjugates in the Rauscher murine erythroleukemia model. Several derivatives of diethylenetriaminepentaacetic acid [the dicyclic dianhydride of diethylenetriaminepentaacetic acid (ca-DTPA), 2-(p-isothiocyanatobenzyl)diethylenetriaminepentaacetic acid (SCNBzDTPA), and 2-(p-isothiocyanatobenzyl)-5(6)-methyl-diethylenetriaminepentaacet ic acid (MxDTPA)], as well as a macrocyclic polyazacycloalkane-N-acetic acid [2-(p-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-N ,N',N",N"'- tetraacetic acid (DOTA)], were conjugated to monoclonal antibody 103A, which is specific for gp70 expressed on Rauscher virus-infected cells. The stability in vivo of 206Bi chelate-103A conjugates was first evaluated in normal mice by determining the levels of 206Bi in blood and kidney, since these were the organs in which free 206Bi, 206Bi-caDTPA-103A, and 35S-103A accumulated. The biodistribution of 206Bi administered as a chelate of caDTPA-103A was virtually indistinguishable from that of free 206Bi, indicating a low degree of in vivo stability of this bismuth chelate when compared to biosynthetically labeled 35S-103A. There was a progressive increase in the 206Bi levels observed in blood when the series of 103A conjugates prepared using SCNBzDTPA, MxDTPA, and DOTA was compared to 206Bi administered free or as a caDTPA-103A chelate. At 1 h after injection into normal mice, the blood level of 206Bi-DOTA-103A was 25-fold greater than that observed for 206Bi-caDTPA-103A and the level in kidney was 6-fold less, values that did not differ significantly from those observed for 35S-103A. Targeting to leukemic spleen was increased by 10-fold when the DOTA conjugate was used; the tumor level was 90% injected dose/g for DOTA, as compared to only 9% injected dose/g for caDTPA-103A at 1 h after injection. Use of the DOTA chelator also reduced by 7-fold the level of uptake by the kidney in the leukemic animals. We, therefore, conclude that the chelator DOTA is a promising reagent for the delivery of 212Bi-antibody conjugates to vascularized tumors under conditions that require targeting via the circulatory system.

Animals↗

PubMed1990

Inhibition of protein kinase C and anti-CD3-induced Ca2+ influx in Jurkat T cells by a synthetic peptide with sequence identity to HIV-1 gp41.

We have previously shown that a synthetic peptide containing env residues 581-597 from HIV-1 inhibits lymphoproliferation of human PBMC. We have investigated the molecular mechanism(s) by which this HIV-1-derived peptide inhibits CD3-mediated signal transduction. We show that the peptide containing residues 581-597 from the HIV-1 transmembrane protein gp41 specifically inhibited the intracellular Ca2+ influx in Jurkat cells stimulated by the mAb OKT3 whereas it had no effect on the production of inositol triphosphate. In addition, the peptide inhibited protein kinase C (pkC)-mediated phosphorylation of the CD3 gamma-chain in intact cells and directly inhibited partially purified pkC. The inhibition was noncompetitive with respect to the substrates histone and ATP and independent of the regulatory domain of the enzyme. Furthermore, the peptide required internalization for inhibitory activity because no inhibition of lymphoproliferation was observed when cells were treated with peptide at 4 degrees C. Based on these results obtained with the peptide aa581-597, we postulate that the transmembrane protein gp41 of HIV-1 may inhibit pkC activity and thus block pkC-dependent immune function contributing to the immunosuppression of HIV-1-infected individuals.

Amino Acid Sequence↗

PubMed1990

Radioimmunotherapy in experimental animal models: principles derived from models.

Experimental animal models have made it possible to study some of the biological, biochemical, and pharmacological parameters involved in the use of radiolabeled monoclonal antibody for therapy and detection. Although such models are less appropriate for studies of dosimetry and the host's immune response to the monoclonal antibody, some general principles have been derived from the various model systems that have largely held true in studies in patients. Some of the points learned from experimental animal models will be illustrated in this paper.

Animals↗

PubMed1990

Identification and characterization of glycosylphosphatidylinositol-linked Schistosoma mansoni adult worm immunogens.

Metabolic radiolabeling of adult worms of Schistosoma mansoni with [3H]myristic acid has revealed that the fatty acid is incorporated into more than 15 proteins. We have shown that two of these proteins, a 200-kDa glycoprotein known to be exposed on the surface of the adult worm following praziquantel treatment and a 22-kDa glycoprotein that shows an enhanced immune reactivity with sera of vaccinated mice, are anchored to the adult worm membrane via a glycosylphosphatidylinositol (GPI) linkage. Both antigens partitioned preferentially into the detergent phase of Triton X-114 and were susceptible, following immunoaffinity purification, to hydrolysis by phosphatidylinositol-specific phospholipase C (PIPLC) from Bacillus thuringiensis and phospholipase C from Bacillus cereus. Diacylglycerol (DAG) was released following hydrolysis by bacterial PIPLC; however, Trypanosoma brucei GPIPLC failed to release the diacylglycerol from either protein. Treatment with nitrous acid generated phosphatidylinositol (PI) from both proteins, and phospholipase D from rat serum cleaved phosphatidic acid from the 200-kDa protein. Although the functional significance of these GPI-anchored proteins is unknown, their release from the surface of the schistosome may contribute to immune evasion.

Animals↗

PubMed1990

Identification of a decapeptide region of human interferon-alpha with antiproliferative activity and homology to an immunosuppressive sequence of the retroviral transmembrane protein P15E.

We have identified a 10-amino acid sequence (amino acid, 9-18) from interferon-alpha (IFN-alpha) that when presented as a synthetic peptide inhibits both the proliferation of the Daudi lymphoblastoid cell line and the antigen receptor-stimulated proliferation of fresh human T lymphocytes. This sequence, which was identified by virtue of its sequence similarity (70% identity) to the immunosuppressive sequence of the retroviral transmembrane protein p15E, represents the smallest fragment of IFN-alpha that has been shown to date to be biologically active.

Amino Acid Sequence↗

PubMed1990

Inhibition of lymphoproliferation and protein kinase C by synthetic peptides with sequence identity to the transmembrane and Q proteins of visna virus.

A peptide sequence in the transmembrane protein of visna virus has been identified that bears a high degree of similarity to a sequence within the transmembrane protein gp41 of human immunodeficiency virus that we have previously shown to be immunosuppressive. Also within the Q (vif/sor) open reading frame of the visna virus genome is a sequence that is highly similar to the immunosuppressive sequence from the retroviral transmembrane protein p15E. We synthesized peptides containing these visna virus sequences and tested them for immunosuppressive activity, comparing them with their human immunodeficiency virus and leukemia retrovirus counterparts. Both the Q- and transmembrane-derived visna virus peptides inhibited lymphoproliferation stimulated by either interleukin-2 or the T-cell antigen receptor in a dose-dependent and sequence-specific manner. The two visna virus peptides also inhibited the enzymatic activity of protein kinase C, thus providing a possible molecular mechanism by which they inhibit immune function.

Amino Acid Sequence↗

PubMed1990

A double-blind, controlled trial in primary care patients with generalized anxiety: a comparison between buspirone and oxazepam.

Two hundred thirty patients with generalized anxiety and Hamilton Rating Scale for Anxiety (HAM-A) scores greater than or equal to 18 were subdivided at random, according to a double-blind design, into one group treated with 5-10 mg of oral buspirone t.i.d. or one group treated with 10-20 mg of oral oxazepam t.i.d. for 6 weeks. No anxiolytic treatment was allowed 3 months prior to trial entry. Analysis of demographic variables revealed no significant imbalance between the two treatment groups. Twenty patients were excluded from efficacy analysis because of treatment withdrawal before the first efficacy evaluation on Day 7. Another 4 patients were excluded because they were taking concomitant psychotropic medication. The remaining 206 patients displayed a decrease in HAM-A scores (mean +/- SD) from 23.9 +/- 4.1 to 10.6 +/- 7.7 in the buspirone group and from 23.9 +/- 4.2 to 11.5 +/- 8.0 in the oxazepam group. The two treatment groups were also found to be virtually identical in an "intent to treat" analysis of all 230 patients as well as in other ratings (Hamilton Rating Scale for Depression, Raskin Depression Scale, Covi Anxiety Scale, Physicians Questionnaire, global ratings, and Hopkins Symptom Checklist [HSCL]-56). However, oxazepam was never superior to buspirone in any of the efficacy analyses. Of the 230 patients, 127 spontaneously reported adverse events, including drowsiness, dizziness, headache, nausea, and nervousness. Adverse events were relatively similar in the two groups. In conclusion, buspirone and oxazepam appear to be equally effective in the treatment of generalized anxiety encountered by general practitioners. This outcome, in addition to a previously documented absence of any dependency liability, makes buspirone a clinically important anxiolytic drug.

Adolescent↗

PubMed1989

Crystallization and preliminary X-ray diffraction studies of an anti-4-hydroxy-3-nitrophenylacetic acid monoclonal antibody Fab fragment complexed with immunizing and heteroclitic haptens.

The Fab fragment of the anti-4-hydroxy-3-nitrophenylacetic acid monoclonal antibody, 88C6/12 has been crystallized in the presence of the eliciting hapten, 4-hydroxy-3-nitrophenacetyl-epsilon-aminocaproic acid (NP-aminocap) and the heteroclitic iodinated analog, 4-hydroxy-3-iodo-5-nitrophenylacetyl-epsilon-aminocaproic acid (NIP-aminocap). Crystals obtained by precipitation with 32% (w/v) polyethylene glycol 3400 in the presence of 40 to 400 microM of either NP-aminocap or NIP-aminocap, belong to the orthorhombic space group P2(1)2(1)2(1) with cell dimensions a = 81.2 A, b = 86.9 A, c = 131.1 A. The cell volume suggests the presence of two molecules of the complex per asymmetric unit. Analysis of the Patterson function indicates that these two molecules are related by a local 2-fold axis parallel to the crystallographic b axis located at x = 0.218 and z = 0.25.

Antibodies, Monoclonal↗

PubMed1989

Role of host antibody in the chemotherapeutic action of praziquantel against Schistosoma mansoni: identification of target antigens.

We have demonstrated previously in a mouse model that effective chemotherapy against Schistosoma mansoni with praziquantel (PZQ) is dependent upon an intact host antibody response. In the same study, it was found that worms recovered from PZQ-treated animals display surface-bound antibodies. In order to identify the target antigens of the antibodies involved in the synergy between PZQ and the immune response, monoclonal antibodies (mAbs) and polyclonal antisera recognizing different tegumental components were tested by indirect immunofluorescence (IF) assay for their ability to bind in vitro to the surface of 6-week-old schistosomes perfused from nude (athymic) mice 1 h after PZQ treatment. Nude mice were used as hosts because worms from these animals were found to lack bound anti-schistosome antibodies. Only 5 of the 21 antibodies tested reacted with drug-treated worms. This indicated that the damage caused by PZQ to the schistosome tegument is restricted to specific tegumental components. Of the positive reactions, one group of antibodies gave IF patterns different from, whereas the other group gave IF reactions similar to those seen with worms perfused from immunologically intact mice. Antibodies against a schistosome esterase and alkaline phosphatase produced reaction patterns in the former category. In contrast, two out of three monoclonal antibodies recognizing different epitopes on a 200-kDa glycoprotein abundant in worm tubercles gave IF patterns very similar to those observed on schistosomes from drug-treated, intact mice. The biological significance of these reactions was confirmed by demonstrating that transfer of one of the positive monoclonal antibodies to 6-week-infected, B cell-depleted (mu-suppressed) mice reconstitutes the efficacy of PZQ treatment to normal levels. The above results suggest that the antibodies involved in the mechanism of action of PZQ react with a limited set of antigens. Furthermore, they implicate the 200-kDa tubercle protein as a major target of this response in naturally infected hosts.

Animals↗