Development of monoclonal mouse antibodies specific for allergenic components in ryegrass (Lolium perenne) pollen

The development of monoclonal mouse antibodies against ryegrass (Lolium perenne) pollen allergens is described. Hybridoma colonies secreting antibodies specific for allergenic components were detected using an enzyme-linked immunosorbent assay. Positive colonies were cloned and expanded. The pollen components with which the monoclonal antibodies interact were identified and characterised following sodium dodecyl sulphate polyacrylamide gel electrophoresis and electrophoretic transfer to nitrocellulose. In this paper six monoclonal mouse antibodies are described. Three antibodies interact with a single molecule of between 30,000 and 35,000 daltons. One antibody interacts with a component of 16,000 daltons whereas the remaining two antibodies react with more than one component, one reacting with two components at 28,000 and 30,000, and the other with five components having molecular weights between 18,000 and 71,000 daltons.     

The in vitro transformation of thymocytes and lymphocytes from humans, rabbits and guinea-pigs and from thymomas

The response of human, rabbit and guinea-pig thymocytes in culture to blastogenic stimulants such as phytohaemagglutinin (PHA) and staphylococcal filtrate (SF) was usually considerably less than that seen with equivalent numbers of peripheral blood and lymph node lymphocytes.     

Expression of silencer of death domains and death-receptor-3 in normal human kidney and in rejecting renal transplants

We have previously reported the pattern of cellular expression of tumor necrosis factor receptors (TNFR) in human kidney and their altered expression in transplant rejection. We have extended our studies to examine the expression of Silencer of Death Domains (SODD), a protein that binds to the cytoplasmic portion of TNFR1 to inhibit signaling in the absence of ligand. In normal human kidney SODD is expressed in glomerular endothelial cells where it colocalizes with TNFR1. During acute rejection both SODD and TNFR1 are lost from glomeruli, but we found strong expression of SODD on the luminal surface of tubular epithelial cells. This occurs in the absence of detectable TNFR1 expression, suggesting that SODD could interact with other proteins at these sites. Several other members of the TNF superfamily, including Fas and death receptors (DR)-3, -4, and -5, also contain intracellular death domains, but SODD only interacts with the death domain of DR3. We therefore studied the expression of DR3 in human kidney, and report that this death receptor is up-regulated in renal tubular epithelial cells and endothelial cells of some interlobular arteries, in parallel with SODD, during acute transplant rejection. In less severe rejection episodes, DR3 and SODD were more focally induced, generally at sites of mononuclear cell infiltrates. In ischemic allografts, eg, with acute tubular necrosis but no cellular rejection, DR3 was induced on tubular epithelial cells and on glomerular endothelial cells. These data confirm that TNF receptor family members are expressed in a regulated manner during renal transplant rejection, and identify DR3 as a potential inducible mediator of tubular inflammation and injury.     

Functional MRI of visuospatial working memory in schizotypal personality disorder: a region-of-interest analysis

BACKGROUND: Functional MRI studies have begun to identify neural networks implicated in visuo-spatial working memory in healthy volunteers and patients with schizophrenia. The study of schizotypal personality disorder (SPD) provides regional analysis in unmedicated patients in the schizophrenia spectrum. METHOD: Unmedicated patients with SPD by DSM-IV criteria and normal controls were assessed with fMRI while performing a visuo-spatial working-memory task. It required the subjects to retain the location of three dots located on the circumference of an imaginary circle and then respond to a query display in which one dot was presented and the subject required to press a button to indicate whether the probe dot location was previously displayed. Subject groups did not differ significantly in spatial memory scores. The exact Talairach and Tournoux coordinates of brain areas previously reported to show activation with spatial memory tasks were assessed. RESULTS: The majority of these locations showed BOLD response activation significantly less in patients during the memory retention period, including the left ventral prefrontal cortex, superior frontal gyrus, intraparietal cortex and posterior inferior gyrus. Regions in the right middle prefrontal and prestriate cortex showed greater activation at a trend level for patients with SPD than for normal controls. In addition, we replicated the findings of increased activation with the task in healthy volunteers in the premotor areas, ventral prefrontal cortex and parietal cortex. CONCLUSIONS: SPD patients show decreased activation compared to healthy volunteers in key frontal regions and we also provided a partial replication of findings reported in healthy subjects.     

A nephelometric study of the reaction of monoclonal rheumatoid factor with heat aggregated gamma globulin and sera from patients with immune complex diseases.

Monoclonal rheumatoid factors (MCRF) have previously been used in a variety of assays for the detection of IgG-containing circulating immune complexes. We have isolated a MCRF from a patient with a lymphoproliferative disorder and have used a nephelometric technique to characterize its reaction with heat-aggreagated gammaglobulin (HAGG) used as a source of artificial immune complexes. The method is simple, economical and rapid and will detect as little as 6 microgram/ml of HAGG over a wide range of physicochemical conditions. A clinical study demonstrated that the sera from thirty-five out of fifty-eight patients (59%) with rheumatoid arthritis and twenty-one out of seventy-four patients (28%) with systemic lupus erythematosus (SLE) gave increased precipitation with MCRF compared with 232 blood donors. However, in marked contrast to previous studies, sucrose gradient ultracentrifugal analysis of nine strongly precipitating sera revealed that in eight the MCRF precipitated with material sedimenting in the monomeric IgG position. In only one specimen did the MCRF react with material sedimenting in heavier regions. It is suggested that different MCRFs vary in the specificity for binding IgG complexes and these reagents should be carefully characterized before becoming established in nephelometric assays for circulating immune complexes.     

Hydrophobic IgG-containing immune complexes in the plasma of autoimmune MRL/lpr mice, lactate dehydrogenase-elevating virus-infected mice, and pigs: association with transforming growth factor-beta and pH-dependent amplification

Persistent infection of mice with lactate dehydrogenase-elevating virus (LDV) is associated with polyclonal B cell activation, autoimmunity, and circulating hydrophobic IgG-containing immune complexes (ICs), which bind to the surfaces of uncoated ELISA plates in the presence of 0.05% Tween 20. We demonstrate here that hydrophobic IgG-containing ICs also appear naturally in the plasma of autoimmune MRL/lpr mice. These and the similar hydrophobic ICs of LDV-infected mice as well as pigs coincide on ELISA plate surfaces with TGF-beta, apparently in the form of an IgG-TGF-beta complex. Circulating hydrophobic IgG-containing ICs are also susceptible to considerable amplification in vitro by exposure to alkaline conditions. By this latter method, the fraction of in vivo hydrophobic IgG, relative to the maximum in vitro chemically inducible IgG, was found to be about 20% in the plasma of LDV-infected mice, 5% in normal mouse plasma, and less than about 2% in pig plasma. These results indicate the potential for both chemically induced and protein-binding contributions to the generation of hydrophobic IgG-containing molecules, and have implications for immunopathological mechanisms in autoimmunity and persistent virus infections.     

Parkin localizes to the Lewy bodies of Parkinson disease and dementia with Lewy bodies

Mutations in alpha-synuclein (alpha S) and parkin cause heritable forms of Parkinson disease (PD). We hypothesized that neuronal parkin, a known E3 ubiquitin ligase, facilitates the formation of Lewy bodies (LBs), a pathological hallmark of PD. Here, we report that affinity-purified parkin antibodies labeled classical LBs in substantia nigra sections from four related human disorders: sporadic PD, inherited alphaS-linked PD, dementia with LBs (DLB), and LB-positive, parkin-linked PD. Anti-parkin antibodies also detected LBs in entorhinal and cingulate cortices from DLB brain and alphaS inclusions in sympathetic gangliocytes from sporadic PD. Double labeling with confocal microscopy of DLB midbrain sections revealed that approximately 90% of anti-alpha S-reactive LBs were also detected by a parkin antibody to amino acids 342 to 353. Accordingly, parkin proteins, including the 53-kd mature isoform, were present in affinity-isolated LBs from DLB cortex. Fluorescence resonance energy transfer and immunoelectron microscopy showed that alphaS and parkin co-localized within brainstem and cortical LBs. Biochemically, parkin appeared most enriched in cytosolic and postsynaptic fractions of adult rat brain, but also in purified, alpha S-rich presynaptic elements that additionally contained parkin's E2-binding partner, UbcH7. We conclude that parkin and UbcH7 are present with alphaS in subcellular compartments of normal brain and that parkin frequently co-localizes with alpha S aggregates in the characteristic LB inclusions of PD and DLB. These results suggest that functional parkin proteins may be required during LB formation.     

Blood ferritin concentrations in newborn infants and the sudden infant death syndrome.

Liver iron concentrations have been shown to be higher in victims of SIDS than in postmortem controls suggesting that high levels of tissue iron may be implicated in SIDS. To determine whether infants who subsequently die from SIDS are born with greater iron stores than those who do not, the iron stores in newborn infants were assessed retrospectively by measuring blood ferritin concentration in spots from Guthrie cards (collected from almost all infants born in the UK in the first week of life). A method for extracting and measuring ferritin from stored blood spots is described. Eighteen cases of SIDS were identified in South Glamorgan along with four controls for each case. Ferritin concentrations did not differ in SIDS victims and controls suggesting that victims of SIDS are not born with abnormal concentrations of stored iron. If iron stores are found to be higher in SIDS victims than in healthy live infants of the same age then it is more likely that the iron will have been acquired after birth.     

Pharyngeal size in snorers, nonsnorers, and patients with obstructive sleep apnea

We measured pharyngeal cross-sectional area and its change with alterations in lung volume in 10 subjects who snored and had obstructive sleep apnea, 6 subjects who snored and did not have obstructive sleep apnea, and 9 subjects who did not snore. Pharyngeal area was measured with use of an acoustic-reflection technique. We found that snorers with and without sleep apnea had a significantly smaller mean (+/- SE) pharyngeal cross-sectional area (4.1 +/- 0.2 and 3.7 +/- 0.9 cm2, respectively) at functional residual capacity than nonsnorers (5.4 +/- 0.5 cm2, P less than 0.025). When lung volume decreased from functional residual capacity to residual volume, both nonsnorers and snorers with sleep apnea had a decrease in pharyngeal area (from 5.4 +/- 0.5 to 4.5 +/- 0.4 cm2 and 4.1 +/- 0.2 to 3.4 +/- 0.2 cm2, respectively), whereas snorers without sleep apnea had no such decrease, suggesting that their pharynxes were less collapsible at low lung volumes. We conclude that snorers with and without sleep apnea have smaller pharyngeal cross-sectional areas than nonsnorers and that snorers with sleep apnea have a further decrease as lung volume falls.