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Biomedical subjects

P Stevens

Publications and source records attributed to P Stevens.

At least 91 records · Page 5Linked to original sources
PubMed1987

Biochemical and immunocytological localization of the neuropeptides FMRFamide, SCPA, SCPB, to neurons involved in the regulation of feeding in Aplysia.

The localization of the neuropeptide FMRFamide in the buccal ganglia and buccal muscles of Aplysia was studied by immunocytology and high-pressure liquid chromatography (HPLC) combined with either a sensitive bioassay or 35S-methionine labeling. Immunocytology with an antiserum directed to FMRFamide stained a large number of fibers, varicosities, and neuronal somata. Two groups of stained neurons were of particular interest. One was the S cells, a group comprised of many small neurons, the majority of which were stained. HPLC of pooled labeled S cells confirmed that at least some of these neurons synthesize FMRFamide. The other group of stained neurons were in the ventral cluster, a group comprised of a small number of large neurons, many of which are motor neurons that innervate the buccal muscles involved in producing biting and swallowing movements. Several of the ventral neurons were previously shown to contain 2 other neuropeptides, the small cardioactive peptides SCPA and SCPB. These neurons are sufficiently large to permit HPLC analyses of the neuropeptides synthesized by individual neurons. This procedure confirmed that individual ventral neurons synthesized FMRFamide, or the SCPs, or all 3 peptides. The coexistence of FMRFamide and the SCPs in the same neuron was confirmed by simultaneous staining of sections from the buccal ganglia with a monoclonal antibody to the SCPs and an antiserum to FMRFamide. The coexistence of the 3 peptides in the same neuron was surprising in light of the observations that these peptides often have opposite biological activity. The ventral neurons are large and potentially identifiable as individuals. Thus, these neurons may be particularly useful for studying the physiological and behavioral roles of neuropeptides in generating complex behaviors.

Animals↗
PubMed1985

The effectiveness of a low molecular weight heparinoid in chronic intermittent haemodialysis.

A new low molecular weight heparinoid, Org 10172 was compared to heparin in a randomized single blind cross-over study in 55 patients with end-stage renal failure undergoing chronic intermittent haemodialysis. The heparinoid administered as a single pre-dialysis i.v. injection of 34.4 anti-Xa units/kg body weight was compared to standard heparin (loading dose 2,500 IU + continuous infusion of 1,800 IU/hr). Mean anti-Xa plasma levels reached were 0.55 and 0.94 anti-Xa units/ml midway dialysis respectively. All 110 dialysis procedures were successfully performed without clotting or bleeding complications. Analysis of the number of clotted hollow-fibres within the dialysers showed a slight statistically calculated advantage in favour of heparin. Clinically no difference was detected. In conclusion, the heparinoid seems to be a good alternative means of anticoagulation in haemodialysis. As it is administered as a single i.v. predialysis injection it will simplify the dialysis procedure.

Adult↗
PubMed1984

Enzyme immunoassay for the quantitation of human immunoglobulin G specific for the glycolipid of Enterobacteriaceae.

An enzyme immunoassay (EIA) is reported that quantitates human immunoglobulin (Ig)G specific for the glycolipid (Re-GL) of Salmonella minnesota Re 595. Re-GL was adsorbed to polystyrene microcuvettes, and 100 microliters of appropriately diluted lots of human plasma or affinity-purified rabbit anti-Re-Gl IgG was added to the cuvettes. The cuvettes were incubated, washed, and then reacted with Staphylococcus aureus protein A-peroxidase. After a final wash the substrate 2,2'-azino-di(3-ethylbenzthiazoline)sulfonic acid (ABTS) was added to yield a blue-green color with a maximum absorption at 405 nm. Using the affinity-purified rabbit anti-Re-IgG as standard, it was possible to quantitate specific levels of anti-Re-GL. The EIA was rapid (2 hr), with an absolute sensitivity of 5 ng of specific anti-Re-GL IgG and an interassay coefficient of variation of 24%. On a unit-mass basis, rabbit anti-Re-GL IgG had 10-fold greater activity than comparable affinity-purified human anti-Re-GL IgG from normal donor plasma. When a rabbit standard was used, the mean level of specific anti-Re-GL IgG in lots of pooled human plasma, which consisted of 18,330 liters, was 1.1 +/- 0.28 micrograms/ml. Of 237 individual donors tested, 24 (10%) had levels of anti-Re-GL IgG that were 10-fold or greater than the mean of 1.1 micrograms/ml. The EIA provided a rapid, reproducible method for accurate quantitation of specific anti-Re-GL IgG in the screening of large volumes of human plasma.

Animals↗
PubMed1984

Inhibition of phagocytosis and chemiluminescence in human leukocytes by a lipid soluble factor in normal tissues.

Homogenates of normal rat tissues inhibited several functional parameters of normal human peripheral blood leukocytes, including luminol-dependent chemiluminescence induced by both soluble (phorbol myristate acetate) and particulate (Escherichia coli) stimuli; in vitro uptake of radiolabeled E. coli; and in vitro phagocytosis and killing of E. coli. The doses of rat tissue protein that caused a 50% inhibition of leukocyte chemiluminescence were ca. 6.2 micrograms for small intestine, 83 micrograms for kidney; 100 micrograms for heart; 132 micrograms for liver, 190 micrograms for skeletal muscle, and 307 micrograms for brain. The putative phagocytosis inhibitor (PI) in rat liver was more plentiful in particulate fractions than in the cytosol. The PI activity in the original or Miranol-solubilized rat liver homogenate was nondialyzable, and it was reduced substantially by heating at 90 degrees C for 30 min but not at 56 degrees C for 30 min. It was unaffected by aprotinin, a potent inhibitor of proteolytic activity. Treatment of tissues with trypsin did not reduce PI activity, whereas treatment with phospholipase A2 clearly increased it. The bulk (up to 88%) of PI in rat liver or small intestine could be extracted by lipid solvents, e.g., diethyl ether. Purified fatty acids were potent inhibitors of leukocyte chemiluminescence; other lipids had little or no inhibiting activity. The various data suggest that (i) normal tissues contain a potent PI and (ii) that the PI is a lipid moiety.

Blood Bactericidal Activity↗
PubMed1984

The role of protein synthesis in the chemotaxis and chemiluminescence of human polymorphonuclear leukocytes.

We investigated the role of protein synthesis in human polymorphonuclear leukocyte (PMN) chemotaxis and luminol-dependent chemiluminescence (CL). We used cycloheximide and puromycin to inhibit protein synthesis, and determined the extent of synthesis by measurement of 14C-amino acid incorporation. With 2-hour incubations both puromycin at 9.0 X 10(-6) M and cycloheximide at 1.8 X 10(-6) M inhibited PMN protein synthesis. At concentrations of 2-4 X 10(-5) M both cycloheximide and puromycin inhibited PMN-chemotaxis 40 and 55%, respectively. However, inhibition was observed only when using zymosan-activated serum and not formyl-methionyl-phenylalanine as a chemoattractant. Using 2-hour incubations, PMN-CL was also suppressed by puromycin and cycloheximide, at 20 and 40%, respectively. The data demonstrated that protein synthesis had an important role in chemotaxis that was dependent on the chemoattractant and perhaps the cellular receptor involved in that process. CL did not require de novo protein synthesis but appeared to depend on protein(s) with a relatively rapid turnover.

Carbon Radioisotopes↗
PubMed1983

Correlation of E. coli K-1 bacteremia and capsular polysaccharide antigenemia in acute and chronic infection.

The K-1 polysaccharide is an important virulence factor in human E. coli infections. Using E. coli 016K1, we investigated the kinetic association of bacteremia and K-1 antigenemia in acute lapine and canine infections and in a chronic infection model of neutropenic rats. Additionally, we assessed the presence of K-1 antigenemia in E. coli K-1 bacteremic patients. K-1 was measured by a solid phase radioimmunoassay (RIA) using cross-reactive equine anti-Group B meningococcal IgM. In acute infections, none of the dogs or rabbits developed antigenemia even with a bacteremia of 2 X 10(4) CFU/ml or 5 X 10(5) CFU/ml, respectively. Antigenemia appeared in the rabbit only with an infecting dose of greater than or equal to 5 X 10(8) CFU. In the rat model we observed an initial bacteremia of 10(3) CFU/ml, which increased to 10(6) CFU/ml at 24 hrs. However, antigenemia was most often delayed, appearing in only greater than or equal to 30 hrs postinfection. Percent mortality was directly associated with the degree of bacteremia and antigenemia. In acute human E. coli K-1 bacteremia, 11 of 22 (50%) of patients were positive for K-1 antigenemia. The data demonstrated that K-1 polysaccharide was not usually detectable in the early stages of bacteremia, but occurred only after prolonged infection or very high infecting doses. The RIA to measure K-1 antigenemia would not be a useful diagnostic tool.

Acute Disease↗
PubMed1983

Opsonization of various capsular (K) E. coli by the alternative complement pathway.

A virulence factor of E. coli K-1 is its capacity to avoid opsonization via the alternative complement pathway (ACP). Since it is not known if E. coli with other capsular (K) antigens have similar properties we examined various capsular E. coli for opsonization by the ACP. To assess opsonization we used whole blood luminol-dependent chemiluminescence (CL) and the magnesium salt of ethyleneglycol tetraacetic acid to block the classical pathway. E. coli K-types 6, 7, 27, 30, 42, 53, 57 and 75 were effectively opsonized via the ACP (greater than 65% of CL obtained with unchelated normal serum). K types 2 and 13 were opsonized by the ACP in both the high range greater than 65% and intermediate range 36-65%. Only K-types 1, 3, 5, 12 and 92 were poorly opsonized (less than 35%) by the ACP. The data demonstrate that most E. coli K-types were opsonized via the ACP. The poor opsonization of E. coli K3, 5, 12 and 92 by the ACP may be virulence factor for these bacteria.

Antigens, Bacterial↗
PubMed1982

Gram-negative pathogens in septicaemic infections.

For the last two decades, gram-negative rod bacteremia has been the major infectious disease problem developing in patients hospitalized in the United States and many European countries. Mortality rates are high and have exceeded 30% in patients with serious underlying diseases. The most commonly isolated organisms are Escherichia coli, Klebsiella-Enterobacter-Serratia, and Pseudomonas aeruginosa but the latter organism has consistently been associated with the highest case fatality ratios. Stool sampling surveys in normal individuals reveal low carriage rates of both K. Pneumoniae and Pseudomonas, but prospective studies in neutropenic patients using the surveillance culture technique has indicated a higher prevalence of gastrointestinal colonization. In neutropenic patients asymptomatic colonization may occur weeks prior to onset of gram-negative rod bacteremia. The patients most likely to develop systemic infection are neutropenic subjects who have P. aeruginosa colonizing the gastrointestinal tract. Microbial pathogenetic factors associated with gram-negative bacillary infections are poorly understood. Antigens such as the O and K antigens appear to confer the property of resistance to phagocytosis. Both antibody deficiency and restricted activation of the alternative complement pathway are responsible for impaired phagocytosis of some bacteremic strains in both fresh normal human serum as well as in the serum or plasma of bacteremic patients. A number of studies have shown a direct correlation between type-specific and cross-reacting serum antibody levels with survival. Additionally, Exotoxin A of P. aeruginosa is a likely virulence factors of this species and levels of humoral antitoxic antibodies measured at onset of bacteremia correlate with improved prognosis. Antibodies to capsular antigens like K-1 antigen of E. coli enhance phagocytosis of strains with different O types. Despite a substantial amount of circumstantial evidence, a role for circulating toxins or other candidate virulence factors has not bee convincingly established in human sepsis. Bioassays like those involving the limulus lysate gelation have identified some patients with endotoxemia, but this test may be negative if circulating immune complexes are present. Additionally, other factors in serum may inactivate endotoxins of complex with endotoxins. Several human and experimental studies suggest that prophylaxis or treatment of gram-negative infections may be effective with antiserum against endotoxins antigens. The high mortality from gram-negative bacteremia in some patients strongly justifies further research in this area.

Cross Infection↗
PubMed1981

Sensitive measurement of endotoxin by radio-rocket immunoelectrophoresis using [125I] Staphylococcus aureus protein A.

Antibody directed against the core glycolipid antigen (CGL) of the mutant Salmonella minnesota Re 595 has been shown to cross-react with endotoxin from bacteria within the group Enterobacteriaceae. Using this cross-reactive CGL antibody we have developed a sensitive (250 pg) radio-rocket immunoelectrophoretic technique to measure endotoxin. We used the principles of rocket immunoelectrophoresis and increased the sensitivity by using 125 I-labelled staphylococcal protein A which serves as a sensitive probe to bind to the Fc portion of the IgG complexed with antigen. The rocket-shaped [125I] protein A labelled immune complexes were detected by radioautography. The sensitivity is 100-fold greater than conventional Coomassie brilliant blue staining. Measurement of CGL was inhibited by normal human serum. However, the assay had the capacity to quantitate endotoxin in buffer extracts of clinically isolated Escherichia coli, Serratia marcescens, Klebsiella pneumoniae but not Pseudomonas aeruginosa. Analysis of various preparations of CGL obtained from different investigators demonstrated wide variation in their immunoreactivity. Because of the significant cross-reaction to detect various endotoxins this method has the potential to measure endotoxemia and assess the immunochemical quality of various endotoxin preparations. Additionally, the technique of using [125I] protein A has wide applicability for the sensitive measurement of other antigens.

Antigens, Bacterial↗