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G J Silverman

Publications and source records attributed to G J Silverman.

At least 73 records · Page 4Linked to original sources
PubMed1988

Incidence of three cross-reactive idiotypes on human rheumatoid factor paraproteins.

The basis for rheumatoid factor (RF) production in autoimmune or lymphoproliferative diseases cannot be understood without defining the molecular factors that dictate RF structure and specificity. Recently three different mAb (6B6.6, 17.109, and G6) have been developed that define cross-reactive idiotypes (CRI) on intact L or H chains of human monoclonal RF cryoglobulins. However, the true incidence of these CRI among RF and their relationship to each other have not been delineated. In the present experiments, a panel of 163 randomly selected IgM paraproteins was evaluated for the expression of the two kappa L chain CRI, 6B6.6 and 17.109, and the H chain CRI, G6. Among the paraproteins with kappa L chains, 14% expressed the 17.109 CRI, and 9% expressed the 6B6.6 CRI. Both ELISA and Western immunoblotting experiments showed that the two L chain CRI were mutually exclusive. Anti-IgG activity was documented in 22 of the IgM-kappa paraproteins, among which mAb 6B6.6 reacted with 7 (32%) and mAb 17.109 with 6 (27%). Both CRI were expressed exclusively by L chains within the kappaIII variable gene subgroup. Although 17.109 CRI+ paraproteins had kappaIIIb L chains, none of the 6B6.6 CRI+ paraproteins possessed L chains with this kappa sub-subgroup specific Ag. The G6 CRI was found predominantly among RF paraproteins and was frequently yet not exclusively associated with the 17.109 CRI+ L chains. Additional experiments were performed on a panel of normal adult human sera and documented the presence of 6B6.6 and 17.109 CRI on a small percentage (0.1 to 2.0%) of IgM from most individuals. These data indicate that 1) the mAb 6B6.6 and 17.109 identify two major and distinct CRI among IgM-RF paraproteins, 2) both CRI are associated exclusively with kappaIII L chains, 3) kappaIIIb and kappaIII non-b L chains are equally prevalent among IgM-RF, 4) the G6 H chain CRI is frequently associated with 17.109 CRI+ L chains, but not with 6B6.6 CRI+ L chains, and 5) although the ability to make 6B6.6 and 17.109 CRI+ kappa L chains is common in humans, these CRI are present in low concentrations in normal IgM.

Adolescent↗
PubMed1988

Idiotypic and subgroup analysis of human monoclonal rheumatoid factors. Implications for structural and genetic basis of autoantibodies in humans.

Rheumatoid factors (RFs) in humans have been studied intensively because of their association with autoimmune and lymphoproliferative diseases. Many human IgM-RFs express cross-reactive idiotypes (CRIs) and have homologous light chains, some of which are encoded by a single V kappa gene, termed V kappa 325. However, although antibody activity generally requires the interaction between heavy and light chain variable regions, much less is known about structural relationships among RF heavy chains. To delineate further the structural and genetic basis of RF autoantibody synthesis, we generated "sequence-dependent" reagents specific for the human heavy and kappa light chain subgroups, and used them to analyze a panel of 27 monoclonal RFs. In addition, these proteins were tested for the expression of a heavy chain-associated CRI (G6), and a light chain-associated CRI (17.109). The results showed that most 17.109-reactive RFs contain heavy chains of the VHI subgroup, which bear the G6 idiotypic marker. However, among the 14 17.109-reactive RFs, two have heavy chains of the VHII subgroup, and another two contain heavy chains of the VHIII subgroup. Previously, we have shown that 17.109 is a phenotypic marker of the human V kappa 325 gene. Accordingly, these results demonstrate that the same human V kappa gene can combine with several VH genes from different VH gene subgroups to generate RF activity.

Amino Acid Sequence↗
PubMed1988

Idiotypic characteristics of rheumatoid factors.

We have used monoclonal anti-idiotypic antibodies and "sequence-dependent" heterologous anti-peptide antibodies of predefined specificities against either the complementarity determining or framework regions of the human immunoglobulin molecules to delineate the structural and genetic basis of human IgM rheumatoid factor (RF) paraprotein formation. Human IgM RF paraproteins were found to be most commonly reactive with anti-kappa III subgroup anti-sera (88%). These RFs could be subdivided into two major light chain cross reactive idiotype (CRI) groups with the monoclonal antibodies 17.109 and 6B6.6. The two CRI positive light chains are likely encoded by two distinct V kappa germline genes, designated respectively Humkv325 (VkRF) and Humkv328. These CRIs are also associated with specificities other than RFs. Heavy chain analysis of RFs revealed that the product of the Humkv325 gene frequently assemble with heavy chains of the VHI subgroup, but will assemble with the heavy chains of other subgroups less frequently. The VHI heavy chains RFs were found to carry frequently the G6 CRI marker.

Antibodies, Monoclonal↗
PubMed1987

Molecular basis for the cross-reactive idiotypes on human anti-IgG autoantibodies (rheumatoid factors).

High titres of anti-IgG autoantibodies (rheumatoid factors, RF) are characteristic of patients with rheumatoid arthritis, Sjögren's syndrome, and mixed cryoglobulinaemia, and may contribute to immune complex formation and tissue damage. The monoclonal RFs from cryoglobulinaemia patients frequently display cross-reactive idiotypes. The genetic basis for the cross-reactive idiotypes on RF autoantibodies has not been determined. To clarify structural and genetic relationships among RFs from unrelated subjects, a series of anti-peptide antibodies have been generated that define primary sequence-dependent idiotypes on RF heavy and light chains. Multiple monoclonal and polyclonal RFs from unrelated individuals have been probed by Western blotting with the anti-idiotypic reagents. The results show that sequences in the kappa light chain variable region represent a common structural element among RF autoantibodies. This hypothesis is confirmed by the cloning and sequencing of the conserved germline variable region gene which encodes human RF kappa chains.

Antibody Specificity↗
PubMed1987

Expression of a germline human kappa chain-associated cross-reactive idiotype after in vitro and in vivo infection with Epstein-Barr virus.

The mouse monoclonal antibody 17.109 recognizes a cross-reactive idiotype (CRI) associated with kappa IIIb light chains of human IgM-rheumatoid factor (RF) paraproteins. The 17.109 idiotypic determinant is encoded by one or a group of closely related V kappa genes. The association of the idiotype with IgM- and IgA-rheumatoid factors in certain autoimmune diseases necessitates an understanding of how human B lymphocytes can be induced to express the idiotype. To investigate the cellular expression of the 17.109 CRI, peripheral blood lymphocytes from normal donors were stimulated in vitro with Epstein-Barr virus (EBV) and pokeweed mitogen (PWM). EBV induced greater expression of IgM-associated 17.109 CRI than did PWM. The 17.109 CRI was preferentially associated with IgM rather than with IgG. In vivo EBV infection was studied in college students with infectious mononucleosis and displayed similar elevation of IgM-associated 17.109 CRI in sera obtained at presentation of clinical illness. Later, IgM levels declined while IgG-associated 17.109 CRI rose. The 17.109 idiotype was unrelated to antibodies against the Epstein-Barr virus nuclear antigen and the viral capsid antigen and was probably due to generalized activation of early B cells. These observations support the hypothesis that the 17.109 CRI is expressed by in vitro and in vivo EBV-infected cells. The 17.109 idiotype identifies a highly conserved V kappa gene product, which is expressed preferentially after EBV infection, but not exclusively with RF autoantibodies.

Animals↗
PubMed1986

Human kappa light chain subgroup analysis with synthetic peptide-induced antisera.

All human kappa light chains belong to one of four subgroups, classified according to their amino acid sequences or by reactivity with adsorbed heteroantisera. The structural basis for the subgroup distinction by antisera is unknown. Therefore, to create anti-kappa subgroup antibodies with predefined specificity, we immunized rabbits with synthetic peptides which correspond to sequences within the first framework region of prototype kappa I, II, III, and IV light chains. The peptide-induced antisera recognized primary sequence-dependent kappa subgroup determinants. They correctly predicted the amino acid sequence in the first framework region of two kappa light chains. By Western immunoblotting and enzyme-linked immunoassay the antisera also identified previously typed, monoclonal light chains of different subgroups with complete specificity. These reagents, define a site of kappa subgroup distinction and represent a potent tool for the characterization of light chain heterogeneity.

Amino Acid Sequence↗
PubMed1980

Herpes zoster in patients with treated Wegener's granulomatosis. A possible role for cyclophosphamide.

In review of the ongoing protocol for the treatment of Wegener's granulomatosis with cyclophosphamide at te National Institutes of Health, an increased incidence of herpes zoster infection was noted. There were a total of nine episodes in seven of a total of 65 patients with a 255 patient year follow-up. The infections occurred while the patients were in complete clinical remission during immunosuppressive therapy. Cutaneous dissemination was noted in two episodes, but no visceral or central nervous system involvement was noted despite continuation of immunosuppressive therapy. The major causal factor of the increased incidence of herpes zoster appeared to be the cyclophosphamide therapy.

Cyclophosphamide↗
PubMed1979

Effects of glucose, pH, and dissolved-oxygen tension on Bacillus cereus growth and permeability factor production in batch culture.

The production of a Bacillus cereus enterotoxin, measured as rabbit skin permeability factor (PF), in response to differences in glucose availability, pH, and dissolved oxygen tension was studied in a 1-liter batch fermentor system. Glucose had to be present for toxigenesis to occur. In uncontrolled fermentation an increasing inhibition of PF production and growth occurred as pH dropped occurred below 6.5. Optimum pH for toxigenesis was 7.0 to 7.5, and fermentations maintained at this level yielded 10- to 20-fold more PF than comparable uncontrolled fermentations. PF production was appreciably diminished at or below pH 6.0 and at or above pH 8.5. Peak PF titer was associated with a drop in acid output, and the titrant utilization profile could be used as an indication of this point. Productivity was greatest in the early exponential phase of growth and decreased to zero at the transition phase. Differences in dissolved oxygen tension affected both the maximum productivity early in the fermentation and the rate of its decrease as growth progressed. The optimum dissolved oxygen tension for toxigenesis was 0.002 atm, and the most rapid growth occurred at 0.10 atm. Productivity and growth were reduced under anerobic conditions, whereas a hyperoxic environment severely reduced productivity, but not growth. Postexponential-phase loss of toxic activity coincided with a rapid increase in cellular oxygen demand. Neither was inhibited by the presence of glucose. However, PF loss was completely prevented by stringent oxygen limitation. Extracellular proteolytic activity did not appear to be responsible for the loss of toxic activity.

Bacillus cereus↗
PubMed1977

Modified radioimmunoassay determination for staphylococcal enterotoxin B in foods.

The sensitivity of solid-phase radioimmunoassay for the measurement of staphylococcal enterotoxin B (SEB) in foods was decreased by food constituents that react with rabbit anti-SEB with an equivalency of over 2 ng/ml. This activity was minimized by a conditioning step for anti-SEB and by removal of interfering compounds in the sample by extraction. The assay was a sequential solid-phase radioimmunoassay technique in which polystyrene test tubes were initially incubated with antisera and then with bovine serum albumin. The tubes were then conditioned with either a centrifuged aqueous cheese extract or, equally effective, reconstituted nonfat dry milk for 16 h at 4 degrees C. Samples of milk or heat-treated and CHCl(3)-extracted cheese or chicken salad slurries were incubated in the assay tubes for 6 h at 37 degrees C. The samples were replaced by I-labeled SEB and incubated for a further 2 to 4 h before the contents were removed and the tubes were washed and counted. A buffer solution containing known concentrations of toxin served as standards for assaying SEB in the food extracts. The entire assay can be accomplished within 24 h with a sensitivity of 1 ng/ml in milk and in the cheese extract or 1.3 ng/ml in the chicken salad extract.

Journal Article↗
PubMed1976

Synthesis of staphylococcal enterotoxin A and nuclease under controlled fermentor conditions.

The production of enterotoxin A and nuclease by Staphylococcus aureus strain 100 was studied in a 1.0-liter fermentor. The effects of the gas flow rate, pH, and dissolved oxygen were evaluated. Toxin and nuclease secretion occurred under all conditions which permitted growth of the organism. Final yields of toxin and nuclease in cultures grown at constant air flow rates, ranging from 50 to 500 cm3 per min, were higher at successively higher flow rates. An optimum flow rate for either toxin or nuclease production was not observed. When the aeration rate alone or aeration rate and pH were held constant, the dissolved oxygen levels in the culture decreased from the initial 100% level to 0 to 5% 3 to 4 h after inoculation. The O2 demand of the culture then maintained this level for an additional 4 to 5 h. This low dissolved oxygen interval was characterized by rapid growth and extracellular protein production. Controlling the dissolved oxygen at a constant level throughout growth did not increase the final levels of toxin and nuclease above those achieved at the respective constant pH values. Growth under the influence of a constant aeration rate of 500 cm3 per min and a constant pH of 6.5 and 7.0 yielded the highest titers of nuclease (1,550 units/ml) and toxin (10.5 mug/ml) obtained in any of the fermentations conducted in this study. Sparging fermentor cultures with pure oxygen at a rate of 100 cm3 per min yielded growth and extracellular protein levels similar to those achieved at the sparge rate of 500 cm3 of air per min. Controlling the dissolved oxygen at 100% of pure oxygen saturation appeared to inhibit the culture, as the final cultural turbidity as well as the levels of toxin and nuclease were reduced. These data indicate that enterotoxin and nuclease secretions are closely associated with the growth of strain 100. Analyses of the production rates of these components indicated that early log phase was the most efficient production interval in the growth cycle and that this efficiency was increased by pH control at 6.7 to 6.8 and dissolved oxygen control at 10% of air saturation.

Enterotoxins↗
PubMed1976

Heterogeneity of Staphylococcus aureus enterotoxin B as a function of growth stage: implications for surveillance of foods.

Staphylococcus aureus was grown in a fermentor under controlled conditions of pH, oxygenation, and temperature, while the higher-molecular-weight products of its growth were continuously removed across ultrafiltration membranes. These products were examined by single and double gel diffusion and immunoelectrophoresis against a variety of available anti-enterotoxin B antisera. All antisera examined were polyvalent for S. aureus antigens. However, two electrophoretically distinct proteins were the major reactants with the antisera. One of these was present in early- to mid-log-phase cultures. After mid-log growth was achieved, both were present but in continuously changing proportions. This observation was repeated with a variety of growth conditions and media. A significant part of the physicochemical heterogeneity of enterotoxin B observed over the past 20 years is thus correlated with the growth phase of the organism. Taken together, these facts are used to argue for a two-step rationale for the detection of food-borne staphylococcal disease: (i) screening for a presumptive hazard by analysis for any antigen, toxin, or enzyme of S. aureus in a foodstuff and (ii) confirmation of the hazard by identifying the presence of an enterotoxin using a combination of physicochemical and serological techniques.

Antigens, Bacterial↗
PubMed1976

Volatiles produced by microorganisms isolated from refrigerated chicken at spoilage.

Volatile components present at spoilage of refrigerated chicken breasts were identified using high-vacuum-low-temperature distillation techniques followed by analysis with combined temperature-programmed gas chromatography and mass spectrometry. A comparison was made of the compounds detected from both irradiated and non-irradiated muscle stored at 2 and 10 degrees C under both aerobic and anaerobic conditions. Isolates were randomly selected from the spoiled poultry, identified, and evaluated for their ability to produce volatile spoilage noted when grown on radiation-sterilized chicken. Several isolates that produced off-odors on sterile chicken breasts were examined. Twenty-two compounds were associated with spoilage. Some of the compounds found on both irradiated and unirradiated samples were considered to play only a minor role in the spoilage aroma or were present in low concentrations, since the aroma of spoiled irradiated chicken lacked the harsh odor notes typical of spoiled unirradiated chicken. Fifteen of the 22 compounds were considered to be unique to unirradiated, aerobically spoiled samples. Nine of these compounds, hydrogen sulfide, methyl mercaptan, dimethyl sulfide, dimethyl disulfide, methyl acetate, ethyl acetate, heptadiene, methanol, and ethanol, were found on chicken spoiled at both 2 and 10 degrees C. xylene, benzaldehyde, and 2,3-dithiahexane were detected only in samples stored at 2 degrees C and methyl thiolacetate, 2-butanone, and ethyl propionate were associated with 10 degrees C spoilage. Fifty-eight isolates randomly selected from fresh, radiation-pasteurized, and unirradiated spoiled poultry were classified taxonomically, and 10 of them, which produced spoilage odors on sterilized chicken breasts, were selected for subsequent analysis of their volatiles. Isolates identified as Pseudomonas putrefaciens and Pseudomonas species that were members of groups I and II of Shewan's classification, as well as Flavobacterium and oxidative Moraxella, produced a number of the compounds found in the aroma of spoiled chicken. A total of 17 compounds were identified. Whereas no isolate produced all of the aroma compounds found in the aroma of spoiled chicken, together they did produce the nine found in unirradiated samples spoiled at either 2 or 10 degrees C, as well as methyl thiolacetate and xylene. Six compounds were present in the volatiles produced by the isolates but were absent in the volatiles identified from spoiled chicken. These were hydrogen cyanide, methyl isopropyl sulfide, 2-propane thiol, methyl propionate, ethyl benzene, and an unidentified compound.

Aerobiosis↗