Interference of rheumatoid factor activity by aspartame,a dipeptide methyl ester

Circulating autoimmune complexes of IgM rheumatoid factors (RF) bound to the Fc portions of normal, polyclonal IgG antibodies are frequently present in humans with rheumatoid arthritis (RA). The sweet tasting methyl ester of L ‐Asp‐L ‐Phe (aspartame or APM) was found to relieve pain and improve joint mobility in subjects with osteo‐ and mixed osteo/rheumatoid arthritis [Edmundson, A. B. and Manion, C. V. ( 1998 ). Clin. Pharmac. Ther. 63 , 580–593]. These clinical observations prompted the testing of the inhibition by APM of the binding interactions of human IgM RFs with IgG Fc regions. The propensity of APM to inhibit IgM RF binding was assessed by competitive enzyme immunoassays with solid‐phase human IgG. Ten RA serum samples and three purified monoclonal cryoglobulins, all of which had RF activity, were tested in this system. We found that the presence of APM significantly reduced the binding of IgM RFs. The inhibitory propensity of APM with monoclonal RF cryoglobulins was increased by the addition of CaCl₂ to the binding buffer. Similar inhibition of the binding of RA derived RFs to IgG was observed for Asp–Phe and its amidated derivative, indicating that the methyl ester is not required for APM's interaction with IgM antibodies. A human (Mez) IgM known to bind octameric peptides derived from the Fc portion of a human IgG₁ antibody was tested for binding of dipeptides by the Pepscan method of combinatorial chemistry. The relative binding constants of Asp–Phe and Phe–Asp were ranked among the highest values for 400 possible combinations of the 20 most common amino acids. Possible blocking interactions of APM were explored by computer‐assisted docking studies with the model of a complex of an RF Fab with the Fc of a human IgG₄ antibody. Modeling of ternary immune complexes revealed a few key residues, which could act as molecular recognition sites for APM. A structural hypothesis is presented to explain the observed interference with RF reactivity by APM. Extrapolations of the current results suggest that APM may inhibit the binding of IgG in a substantial proportion of IgM RFs. Interference of RF reactivity, especially in RA patients, may alleviate the pain and immobility resulting from chronic inflammation of the joints. Copyright © 1999 John Wiley & Sons, Ltd.     

Effectiveness of AV-1011® to reduce damage to drill-planted rice from blackbirds

We determined efficacy of AV-1011® (a 50% anthraquinone product; Arkion® Life Sciences, New Castle, Delaware) on drill-planted rice seed to reduce blackbird damage and determine residue levels of anthraquinone (AQ) in rice seeds and seedlings and in the mature rice crop under field enclosures at the University of Missouri-Delta Center farm near Portageville, Missouri. Red-winged blackbird (Agelaius phoeniceus) damage was higher for untreated than AV-1011® treated rice seedlings at assessment period 3, 15 days postplanting (F_1,141 = 15.81, P < 0.001), and at assessment period 4, 19 days postplanting (F_1,136 = 11.54, P = 0.001). Blackbird damage to AV-1011-treated seedling for assessment periods 3 and 4 was 8% and 7%, respectively, while blackbird damage to untreated seedlings during the same assessment periods was 52% and 44%. More blackbirds used untreated plots than AV-1011-treated plots during assessment periods 2–4 (F_1,17.8 = 20.02, P < 0.001). Overall concentrations of AQ on seeds averaged 5,993 µg/g or 0.59% during the test period. Concentrations of AQ in mature rice seed and plant collected at harvest averaged 1.22 µg/g and 0.10 µg/g, respectively. AV-1011 offers promise for reducing bird depredations to newly planted rice, but additional testing should be conducted to evaluate this repellent in a large-scale field setting. © 2011 The Wildlife Society.     

Specific binding of the C-peptide of proinsulin to cultured B-cells from a transplantable rat islet cell tumor

Specific binding of the C-peptide of proinsulin was evaluated using a transplantable NEDH rat islet cell tumour predominantly composed of insulin-secreting B-cells. Cultured tumour B-cells exhibited greater than 90% viability assessed by trypan blue exclusion, and retained the ability to form tumours with accompanying hypoglycaemia and hyperinsulinaemia after reimplantation. During binding experiments with synthetic rat C-peptide I and iodinated tyrosylated rat C-peptide I, turnout B-cells exhibited 54±6% specific binding. Displacement of tracer increased with increasing concentrations of unlabelled rat C-peptide I (0.25–1,000 ng/ml), and the specificity of binding was substantiated by reduced displacement with human C-peptide. Scatchard analysis of specific C-peptide binding revealed a curvilinear plot with upward concavity. The demonstration of specific C-peptide binding to insulin-secreting B-cells provides evidence for a physiological role of proinsulin C-peptide.     

Extraction of cellular DNA from human cells and tissues fixed in ethanol

DNA can be extracted from ethanol-fixed lymphoid cells and tissues. The fixation procedure is simple and rapid, and the DNA extraction itself is the same as that normally used for fresh tissue or cells. DNA extracted from ethanol-fixed material is indistinguishable from DNA extracted from fresh samples based on its purity, its ability to be digested with restriction endonucleases, and its ability to specifically hybridize to DNA probes. The capability to extract DNA from ethanol-fixed cells and tissues eliminates the need for stringent handling and storage requirements of fresh or frozen specimens.     

Human erythrocyte Band-3 has an altered N terminus in malaria-resistant Melanesion ovalocytosis

There is a high prevalence of the erythrocyte polymorphism ovalocytosis associated with reduced susceptibility to malaria in Papua New Guinea. The major erythrocyte integral membrane protein, Band-3, showed markedly increased phosphorylation in whole cells or isolated ghosts from ovalocytic individuals. The cytoplasmic domain of the ovalocyte Band-3 was found to be approx. 3 kDa largen than the normocytic protein. The N-terminal sequence of the ovalocytic Band-3 was different from the reported sequence for human Band-3, suggesting that the increased size results from an N-terminal extension. Since this is the region of Band-3 which is phosphorylated and interacts with the red cell cytoskeleton, it is likely that this alteration in ovalocytic Band-3 is the underlying cause of the diverse alterations in ovalocytic cells including increased phosphorylation, increased membraned rigidity, decreased agglutinability by blood group antibodies and refractoriness to invasion by malarial parasites.     

Verification of a proteomic biomarker panel to diagnose minor stroke and transient ischaemic attack: phase 1 of SpecTRA,a large scale translational study

Objective: To derive a plasma biomarker protein panel from a list of 141 candidate proteins which can differentiate transient ischaemic attack (TIA)/minor stroke from non-cerebrovascular (mimic) conditions in emergency department (ED) settings.

Design: Prospective clinical study (#NCT03050099) with up to three timed blood draws no more than 36?h following symptom onset. Plasma samples analysed by multiple reaction monitoring-mass spectrometry (MRM-MS).

Participants: Totally 545 participants suspected of TIA enrolled in the EDs of two urban medical centres.

Outcomes: 90-day, neurologist-adjudicated diagnosis of TIA informed by clinical and radiological investigations.

Results: The final protein panel consists of 16 proteins whose patterns show differential abundance between TIA and mimic patients. Nine of the proteins were significant univariate predictors of TIA [odds ratio (95% confidence interval)]: L-selectin [0.726 (0.596–0.883)]; Insulin-like growth factor-binding protein 3 [0.727 (0.594–0.889)]; Coagulation factor X [0.740 (0.603–0.908)]; Serum paraoxonase/lactonase 3 [0.763 (0.630–0.924)]; Thrombospondin-1 [1.313 (1.081–1.595)]; Hyaluronan-binding protein 2 [0.776 (0.637–0.945)]; Heparin cofactor 2 [0.775 (0.634–0.947)]; Apolipoprotein B-100 [1.249 (1.037–1.503)]; and von Willebrand factor [1.256 (1.034–1.527)]. The scientific plausibility of the panel proteins is discussed.

Conclusions: Our panel has the potential to assist ED physicians in distinguishing TIA from mimic patients.