Coupling of mu-opioid receptors with adenylate cyclase in naive and morphine tolerant rabbits.
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Adenylyl Cyclase Inhibitors↗
Biomedical subjects
M J Boyer
Publications and source records attributed to M J Boyer.
Lipid-modified surface protein antigens expressing size variation within the species Mycoplasma hyorhinis.
Monoclonal antibodies (MAbs) previously shown to recognize distinct epitopes selectively expressed on the surface of some Mycoplasma hyorhinis strains were used to define two discrete sets of lipid-modified membrane surface proteins showing marked size variation within this species. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis of Triton X-114 phase-fractionated proteins from six isolates of M. hyorhinis defined a set of amphiphilic integral membrane proteins of 23, 50, and 55 kilodaltons (kDa) recognized on respective isolates by one MAb and a second set of integral proteins of 88, 120, and 100 to 150 kDa recognized by another MAb. The first group of proteins all contained a common, amphiphilic 18-kDa limit tryptic polypeptide bearing the epitope. The size- and strain-variant surface antigens identified by the MAbs were shown to be lipid-modified proteins. Phase fractionation of [3H]palmitate-labeled organisms revealed numerous 3H-labeled proteins in all isolates, which partitioned exclusively into the hydrophobic phase. These proteins generally showed pronounced size variation among isolates and included the antigen variants recognized by the two MAbs, as demonstrated directly by immunoprecipitation of correspondingly sized 3H-labeled proteins from each isolate. A third MAb recognized an invariant, lipid-associated surface protein of 70 kDa on all M. hyorhinis isolates. Covalent modification of lipid-associated proteins was confirmed by identifying 3H-labeled methyl palmitate after acid methanolysis of Triton X-114 phase proteins derived from [3H]palmitate-labeled organisms. However, removal of covalently bound lipid from chloroform-methanol-extracted proteins by alkaline hydroxylamine was selective; complete removal was observed with only a few proteins, possibly including the 120-kDa form of one antigen variant. This suggested potential differences in the nature of covalent linkage among lipid-modified M. hyorhinis surface antigens. Intraspecies antigen variants described here in M. hyorhinis share some characteristics with size-variant antigens reported in phylogenetically related gram-positive eubacteria and may contribute to phenotypic diversification and differences in pathogenicity of mycoplasmas.
Antibodies, Monoclonal↗
Association of lipids with integral membrane surface proteins of Mycoplasma hyorhinis.
Triton X-114 (TX-114)-phase fractionation was used to identify and characterize integral membrane surface proteins of the wall-less procaryote Mycoplasma hyorhinis GDL. Phase fractionation of mycoplasmas followed by analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed selective partitioning of approximately 30 [35S]methionine-labeled intrinsic membrane proteins into the TX-114 phase. Similar analysis of [3H]palmitate-labeled cells showed that approximately 20 proteins of this organism were associated with lipid, all of which also efficiently partitioned as integral membrane components into the detergent phase. Immunoblotting and immunoprecipitation of TX-114-phase proteins from 125I-surface-labeled cells with four monoclonal antibodies to distinct surface epitopes of M. hyorhinis identified surface proteins p120, p70, p42, and p23 as intrinsic membrane components. Immunoprecipitation of [3H]palmitate-labeled TX-114-phase proteins further established that surface proteins p120, p70, and p23 (a molecule that mediates complement-dependent mycoplasmacidal monoclonal antibody activity) were among the lipid-associated proteins of this organism. Two of these proteins, p120 and p123, were acidic (pI less than or equal to 4.5), as shown by two-dimensional isoelectric focusing. This study established that M. hyorhinis contains an abundance of integral membrane proteins tightly associated with lipids and that many of these proteins are exposed at the external surface of the single limiting plasma membrane. Monoclonal antibodies are reported that will allow detailed analysis of the structure and processing of lipid-associated mycoplasma proteins.
Antibodies, Monoclonal↗
Triton X-114 phase fractionation of an integral membrane surface protein mediating monoclonal antibody killing of Mycoplasma hyorhinis.
A previously defined immunoglobulin M(kappa) monoclonal antibody reacting with a surface epitope of Mycoplasma hyorhinis is shown in this report to mediate specific, complement-dependent mycoplasmacidal activity. Immunoblot analysis of mycoplasma components and their tryptic cleavage products showed that the epitope recognized was present on a protein with an apparent molecular weight of 23,000 (p23) and on a limit tryptic fragment of this protein with an apparent molecular weight of 18,000 (p18). Both p23 and p18 are shown by Triton X-114 phase fractionation to partition efficiently into the hydrophobic detergent phase. Other antigens bearing epitopes not expressed at the cell surface were present among the numerous hydrophilic proteins found in the aqueous phase. The external orientation and membrane association of the p23 antigen were further established by demonstrating that trypsin treatment of intact mycoplasmas generated the antigenic p18 fragment, which remained tightly associated with the organism. These results localize an epitope responsible for antibody-mediated mycoplasma killing onto a specific, surface-exposed region of an integral membrane protein of this organism. Since the monoclonal antibody used in this study does not bind to the surface of all strains of M. hyorhinis, the epitope identified also defines a structural marker of antigenic surface variation within this species, a feature previously observed during serological classification of the organism. Analysis of the antigenic and structural features of the p23 surface antigen may therefore be useful in establishing mechanisms of surface antigen variation among integral membrane proteins of mycoplasmas that could dictate important antigenic characteristics recognized during chronic disease caused by these agents.
Antibodies, Monoclonal↗
[Screening of essential fatty acid deficiency in children. Respective value of the assay of fatty acids in total lipids and lipid fractions of the serum].
This study was carried out in metabolically stable infants aged from one to six months receiving artificial food. The lipid serum fraction presenting changes characteristic of essential fatty acid (EFA) deficiency was determined. A preliminary study (n = 13 samples) showed that analysis of total fatty acids (TFA) and of phospholipids (PL) was more discriminatory than analysis of free fatty acids (FFA), triglycerides (TG) or esterified cholesterol (EC). Comparison of TFA and PL (n = 25 samples) confirmed literature data; in particular, C18: 2 n-6 and C20: 4 n-6 decreased whereas C20: 3 n-9 increased. These changes were clearer and significantly greater (p less than 0.001) for C20: 3 n-9 and C20: 4 n-6 of the PL, but were also very significant for the TFA compared to healthy controls. The C20: n-9/C20: n-6 ratio was identical for all fractions. TFA analysis by gas-liquid chromatography is faster and less costly than analysis of lipid fractions and provides sufficient data for screening of EFA deficiency.
Chemical Fractionation↗
The use of Histolyn CYL in an enzyme immunoassay to detect Histoplasma capsulatum antibodies.
Histolyn CYL, the yeast phase reagent, has been employed as the antigen in an enzyme immunoassay (ELISA) to detect antibody to Histoplasma capsulatum in sera from experimentally sensitized rabbits and from humans with histoplasmosis. Three different lots of Histolyn CYL were initially evaluated with respect to antibody detection in individual and pooled rabbit serum specimens. Optimal reactivity was obtained with 4.5 micrograms (total dry weight) per ml of lots 2 and 3 of the reagent. The optimally diluted pooled reagent detected antibody titers ranging from 1:16 to 1:8192 in 36 rabbit sera. In contrast, a marked decrease in sensitivity was evidenced when the same sera were assayed by complement fixation and immunodiffusion tests with commercially available reference reagents. Complement fixation titers ranged from 0 to 1:128 and immunodiffusion tests on undiluted sera were positive with only 5 specimens. Twenty-five sera from patients with acute, chronic and disseminated histoplasmosis were also assayed by the ELISA with the reagent described above. Titers ranged from less than 1:16 to 1:20,480 or greater. Complement fixation titers ranged from 0 to 1:1024 (mycelial antigen) and from 0 to 1:256 (whole yeast cell antigen). No antibody was detected in 7 specimens with mycelial antigen or in 3 specimens with the yeast cell antigen. Immunodiffusion tests on undiluted sera were positive with 9 of 11 specimens. Minimal cross reactivity was evidenced when Histolyn CYL was used in the ELISA to detect anti-Coccidioides immitis antibody in sera from infected animals and humans. These data encourage the continued development of this method for the serodiagnosis of human histoplasmal infection.
Animals↗
Communication styles in the cancer consultation: preferences for a patient-centred approach.
OBJECTIVE: Although doctor-patient communication has been the focus of numerous studies, there is a lack of empirical evidence on which to base a curriculum for teaching effective communication skills for use in an oncology setting. Research within the general practice area identifies patient-centred and doctor-centred behaviours as the most important dimensions of doctor-patient communication. This study examined patients and their relatives/friends' preferences for and satisfaction with patient-centred and doctor-centred consulting styles. It was argued that by determining patient preferences for consulting styles, specific recommendations for improving communication in the oncology setting could be formulated. PARTICIPANTS AND METHODS: One hundred and thirteen women who had been treated for breast cancer and 48 of their relatives or friends watched videotaped scenarios of an oncology consultation, using professional actors. Viewers were randomly allocated to either a good prognosis or poor prognosis video, in which the oncologist discussed the patient's diagnosis, treatment and prognosis. These segments were presented in both styles to allow viewers to directly compare and contrast the patient-centred and doctor-centred approach. Outcomes included style preference and satisfaction. Demographic details, information and involvement preferences, anxiety and depression levels were also obtained. RESULTS: Both patients and their relatives or friends significantly preferred a patient-centred consulting style across all aspects of the consultation (p<0.0001), except within the treatment segment of the good prognosis video where there was no significant difference. One third of the viewers preferred a doctor-centred style for the treatment and prognosis segments. Predictors of a patient-centred style preference in the treatment and prognosis segments included watching a poor prognosis video (OR=2.45, 95% CI 1.04-5.81, p=0.04; OR=3.22, 95% CI 1.22-8.50, p=0.02, respectively), and being employed in a professional occupation (OR=2.38, 95% CI 1.02-5.53, p=0.04 for the treatment segment only). Satisfaction ratings varied within and across videos. CONCLUSION: Despite some methodological limitations, this study provides empirical data indicating that patients and their relatives or friends prefer a patient-centred approach to the consultation, particularly when the patient has a poor prognosis. The fact that a substantial minority of patients preferred a doctor-centred style emphasizes the need to enhance physicians' abilities to recognize different patient needs throughout the consultation.
Adaptation, Psychological↗
Prolonged survival after the diagnosis of metastatic hepatic epitheliod haemangioendothelioma.
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Adult↗
Plasma phospholipid fatty acids and urinary excretion of prostaglandins PGE1 and PGE2 in infants during total parenteral nutrition, with continuous or sequential administration of fat emulsion.
During total parenteral nutrition, using an identical supply of fat emulsion (350 mg/kg/24 hr) to correct essential fatty acid deficiency in children, the efficacy of two methods of administration was studied: continuous over 24 hr, or discontinuous 3 hr/day. At the beginning of the study, all the infants (1-4 months old) had proven essential fatty acid deficiency. After at least 1 month of one of the two nutritional protocols (continuous or discontinuous), plasma phospholipid fatty acid composition and PGE1 and PGE2 urinary excretion were measured. The results obtained indicate better utilization of the fat emulsion when it is administered almost every day, in continuous infusion over 24 hr (1 g/kg/24 hr of Intralipid 20%).
Alprostadil↗
[Multicenter evaluation on different analyzers of three methods for direct HDL-cholesterol assay].
Most frequently, in routine laboratories, C-HDL is measured in the supernatant after precipitation of apolipoprotein B-containing lipoproteins by the sodium phosphotungstate/magnesium chloride reagent (PTA). This method involves precipitation, centrifugation and decantation steps which prevent full automation of the measurement and decrease the accuracy of the results. Recently, three direct assays for C-HDL including alpha-cyclodextrin sulphate (alpha-CD), polyanions/detergents (PA-D) or antibodies anti-beta-lipoproteins (AC) have been commercialized, in which all steps are fully managed by automated analyzers. These new methods have been compared to the conventional procedure (PTA), in multicenter studies among six laboratories using different analyzers. The C-HDL values measured by the alpha-CD and PA-D assays correlated well with those of the PTA method (r > 0.98), on most of the analyzers. With the AC assay, only the results obtained with the Hitachi 717 analyzer were correlated with C-HDL values of the PTA method. The linearity and specificity studies were evaluated in the laboratory A on a Kone Specific analyzer. The alpha-CD and PA-D assays were linear for C-HDL values from 0 to 5.56 mmol/l, as observed by increasing amounts of HDL2 + HDL3 or serum without lipoprotein isolated by ultracentrifugation. The specificity of these two methods was evaluated simultaneously, by adding various amounts of lipoproteins isolated by sequential ultracentrifugation. No interference was observed when adding chylomicrons up to 13.4 mmol/l of triglycerides for both methods. Inversely, increased C-HDL values were observed with added VLDL from 6 mmol/l of triglycerides for the PA-D assay and from 8 mmol/l for the alpha-CD assay. No interference was observed with added LDL up to 11.5 mmol/l of C-LDL for the alpha-CD assay and up to 6.7 mmol/l for the PA-D assay. In conclusion, the present multicenter evaluation demonstrates that the new procedures for the direct automation of C-HDL are easy and accurate and most of them correlated well with the classical precipitation method. In addition the study provides arguments for a choice between the different direct C-HDL methods.
Antibodies↗
[Glutathione peroxidases: value of their determination in clinical biology].
The role of glutathione peroxidase in the oxidative metabolism and recent advances in the demonstration of the consequences of the desequilibrium in the proxidant/antioxidant balance on biological molecules oxidation, intracellular signals transduction, apoptosis and necrosis, have led to new approach in the knowledge of many pathological processes. Methods for determining antioxidant capacity have been developed. The measurement of glutathione peroxidase activity is a key step in the study of oxidative stress. Its determination in clinical biology needs optimal conditions for standardised assays which will be used for epidemiological studies aimed to evaluate the role of nutritional factors involved in the pathogeny of diseases caused or accompanied by oxidative stress.
Adolescent↗