Fleet programming.
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Delivery of Health Care↗
Biomedical subjects
T Daniels
Publications and source records attributed to T Daniels.
Conformation-dependent GAD65 autoantibodies in diabetes.
AIMS/HYPOTHESIS: Conformation-dependent autoantibodies directed against GAD65 are markers of Type 1 diabetes. In this study we aimed to determine whether the substitution of GAD65 with GAD67 amino acids would affect the binding of conformation-dependent GAD65 autoantibodies. METHODS: We used PCR-based site-directed mutagenesis to generate a series of mutated GAD65 cDNA constructs in which specific GAD65 coding sequences for regions of the protein critical for autoantibody binding were replaced with GAD67 coding sequences. RESULTS: The introduction of a point mutation at position 517, substituting glutamic acid with proline, markedly reduced the binding of disease-associated GAD65 antibodies. The binding of GAD65 antibodies to the E517P mutant was reduced in the sera of all newly diagnosed Type 1 diabetes patients ( n=85) by a mean of 72% ( p<0.0001) compared with binding to wild-type GAD65. Patients with latent autoimmune diabetes in adults ( n=24) showed a similar reduction in binding (79% reduction, p<0.0001). First-degree relatives who subsequently progressed to Type 1 diabetes ( n=12) showed a reduction in binding of 80% compared with a reduction of only 65% among relatives who had not progressed to disease ( n=38; p=0.025). In healthy GAD65Ab-positive individuals who did not progress to diabetes during a 9-year follow-up period ( n=51), binding to GAD65-E517P was reduced by only 28% compared with binding to wild-type GAD65. CONCLUSIONS/INTERPRETATION: Differences in autoantibody binding to wild-type GAD65 versus GAD65-E517P may provide predictive information about Type 1 diabetes risk beyond that provided by the presence or absence of GAD65 autoantibodies. Lack of binding to mutant GAD65-E517P defines GAD65-positive individuals who are at higher risk of developing diabetes.
Amino Acid Sequence↗
Caspase cleavage of the nuclear autoantigen LEDGF/p75 abrogates its pro-survival function: implications for autoimmunity in atopic disorders.
Lens epithelium-derived growth factor p75 (LEDGF/p75) is a nuclear autoantigen in atopic disorders implicated in cellular protection against stress-induced apoptosis. We observed that LEDGF/p75 was cleaved during apoptosis into fragments of 65 and 58 kD generated by caspases-3 and -7 cleaving at three sites: DEVPD30/G, DAQD486/G and WEID85/N. Sequence analysis revealed that the DEVPD30/G and WEID85/N sites lie within the highly conserved HATH (homologous to amino terminus of hepatoma-derived growth factor) region, also known as PWWP domain. Alignment of proteins containing this domain failed to reveal conservation of the DEVPD30/G and WEID85/N sites, suggesting that the HATH/PWWP domain of LEDGF/p75 may be specifically targeted by caspases. Overexpression of LEDGF/p75 protected HepG2 cells from serum starvation-induced cell death, whereas expression of the 65 kD fragment failed to protect. The apoptotic cleavage of LEDGF/p75 may contribute to the pathogenesis of atopic disorders by abrogating its pro-survival function and enhancing its immunogenicity.
Apoptosis↗
Insulin-like effects of a physiologic concentration of carnitine on cardiac metabolism.
Pharmacologic (millimolar) levels of carnitine have been reported to increase myocardial glucose oxidation, but whether physiologically relevant concentrations of carnitine affect cardiac metabolism is not known. We employed the isolated, perfused rat heart to compare the effects of physiologic levels of carnitine (50 microM) and insulin (75 mU/l [0.5 nM]) on the following metabolic processes: (1) glycolysis (release of 3H2O from 5-3H-glucose); (2) oxidation of glucose and pyruvate (production of 14CO2 from U-14C-glucose, 1-14C-glucose, 3,4-14C-glucose, 1-14C-pyruvate, and 2-14C-pyruvate); and (3) oxidation of palmitate (release of 3H2O from 9,10-3H-palmitate). We found that addition of carnitine (50 microM) to a perfusate containing both glucose (10 mM) and palmitate (0.5 mM) stimulated glycolytic flux by 20%, nearly doubled the rate of glucose oxidation, and inhibited palmitate oxidation by 20%. These actions of carnitine were uniformly similar to those of insulin. When carnitine and insulin were administered together, their effects on the oxidation of glucose and palmitate, but not on glycolysis, were additive. When pyruvate (1 mM) was substituted for glucose, neither carnitine nor insulin influenced the rate of oxidation of pyruvate or palmitate. In combination, however, carnitine and insulin sharply suppressed pyruvate oxidation (75%) and doubled the rate of palmitate oxidation. None of the responses to carnitine or insulin was affected by varying the isotopic labeling of glucose or pyruvate. The results show that carnitine, at normal blood levels, exerts insulin-like effects on myocardial fuel utilization. They also suggest that plasma carnitine in vivo may interact with insulin both additively and permissively on the metabolism of carbohydrates and fatty acids.
Animals↗
Search for a W' boson via the decay mode W'-->munumu in 1.8 TeV pp collisions.
We report the results of a search for a W' boson produced in pp collisions at a center-of-mass energy of 1.8 TeV using a 107 pb-1 data sample recorded by the Collider Detector at Fermilab. We consider the decay channel W'-->&munumu and search for anomalous production of high transverse mass munumu lepton pairs. We observe no excess of events above background and set limits on the rate of W' boson production and decay relative to standard model W boson production and decay using a fit of the transverse mass distribution observed. If we assume standard model strength couplings of the W' boson to quark and lepton pairs, we exclude a W' boson with invariant mass less than 660 GeV/c2 at 95% confidence level.
Journal Article↗
Plasmapheresis and immunosuppression in stiff-man syndrome with type 1 diabetes: a 2-year study.
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Adult↗
Epitope-specific glutamic acid decarboxylase-65 autoantibodies in intravenous immunoglobulin preparations.
Intravenous immunoglobulin (IVIG) has been used to treat many autoimmune disorders including Stiff-Man Syndrome (SMS). SMS is a neurological disorder associated with an immune-mediated deficiency of gamma-aminobutyric acid (GABA) due to autoantibodies against the GABA synthesizing enzyme glutamic acid decarboxylase-65 (GAD65). GAD65 autoantibodies are present among 1-2% of healthy individuals. It can therefore not be excluded that GAD65 autoantibodies may be present in IVIG, which is prepared from multiple blood donors. We report here that GAD65 but not IA-2 autoantibodies were present in commercial IVIG preparations. The presence of autoantibodies may affect the outcome of IVIG treatment and screening commercial preparations of IVIG for GAD65 autoantibodies is therefore recommended before treating patients.
Antibody Affinity↗
Radioimmunoassay for glutamic acid decarboxylase-65.
BACKGROUND: Glutamic acid decarboxylase-65 (GAD65), the enzyme that catalyzes the formation of gamma-aminobutyric acid (GABA), is the major autoantigen in both type 1 (insulin-dependent) diabetes and stiff-man syndrome (SMS). The observation that GAD65 autoantibodies may be present for years before the clinical onset of diabetes raises the question of when GAD65 is available to initiate an immune response to allow the formation of autoantibodies. In order to address this question it will be necessary to measure GAD65 in tissue, cells, and plasma. METHODS: A radioimmunoassay (RIA) was developed for GAD65 based on the use of a polyclonal rabbit antiserum directed to the N-terminus of GAD65. RESULTS: Using the GAD65 RIA, we have determined the GAD65 content in a human GAD65 gene transfected cell line and in beta-cell preparations from different species. The assay detects an increase of immunoreactive GAD65 after glucose-stimulation and GAD65 that is discharged from rat beta cells after their exposure to the diabetogenic agent streptozotocin. It also measures good recovery of GAD65 added to human plasma samples. CONCLUSIONS: The GAD65 RIA makes it possible to determine both cellular and extracellular levels of GAD65; this might be useful in investigating the mechanisms leading to the formation of GAD65 autoantibodies in type 1 diabetes and SMS patients.
Animals↗
Adverse lipid and coronary heart disease risk profiles in young women with polycystic ovary syndrome: results of a case-control study.
Polycystic ovary syndrome (PCOS), a disorder of hyperandrogenism and chronic anovulation affects 5%-10% of all women. Women with PCOS often have elevated cardiovascular risk factors. A total of 244 PCOS cases were identified through the Division of Reproductive Endocrinology at Magee-Womens Hospital and were age-matched to 244 neighborhood controls. The average age of cases and controls was 35.3 +/- 7.4 and 36.7 +/- 7.7. Women with PCOS compared to controls had substantially higher LDL-C and total cholesterol levels at each age group under 45 years after adjustment for body mass index, hormone use, and insulin levels. In the over 40-year age group, little difference was noted between cases and controls. Among cases and controls (<40), PCOS predicted LDL-C, total cholesterol and triglycerides, but did not have a significant effect on lipid levels in older cases and controls after controlling for the other variables. The primarily pre- to perimenopausal PCOS cases > or =40 years of age have similar LDL-C and total cholesterol levels as their age-matched controls, probably reflecting the LDL-C increase with age among controls.
Adult↗
Clinical and angiographic findings in black patients with suspected coronary artery disease.
To determine the clinical variables and coronary angiographic findings in black patients with suspected coronary artery disease, we analyzed the data on consecutive black patients undergoing their first coronary angiogram over a three year period at the Cook County Hospital, Chicago, Illinois. We compared these findings to those of black and white patients from previous studies. There were 654 patients with a mean age+/-standard deviation of 56+/-10 years; 309 (47%) were men. Two hundred nineteen patients (33%) presented with unstable angina, 75 patients (12%) with acute myocardial infarction and 338 patients (52%) with chronic stable angina. Three hundred forty-six patients (53%) had 50% or greater stenosis in at least one of the major vessels. Among the patients with coronary artery disease, 128 patients (37%) had one vessel disease, 102 patients (29%) had two-vessel disease, and 116 patients (34%) had three-vessel disease. Black patients who undergo coronary angiography for suspected coronary artery disease have a high frequency of normal coronary angiogram or non-obstructive coronary artery disease. The frequency of 1-, 2- and 3-vessel disease in blacks with coronary artery disease is comparable to those observed in whites in previous reports.
Adult↗
Diagnostic sensitivity of immunodominant epitopes of glutamic acid decarboxylase (GAD65) autoantibodies in childhood IDDM.
The prevalence and titre of epitope-specific autoantibodies to glutamic acid decarboxylase (GAD65) in 155 insulin-dependent diabetic (IDDM) and 9 GAD65 antibody (Ab)-positive healthy children were determined using four GAD65/67 chimaeric molecules which discriminate among the N-terminal (N), middle (M) and C-terminal (C) epitopes of GAD65. Radioligand binding assays for IgGAb used immunoprecipitation of in vitro translated 35S-GAD. We found autoantibodies to GAD65 in 116 of 155 (75%), to GAD67 in 19 of 155 (12%) (p < 0.0001) and to the GAD65-N-67 chimaera in 25 of 155 (16%) (p < 0.0001) IDDM sera. GAD67Ab were found almost exclusively (17 of 19, 89%) in GAD65Ab-positive sera and the levels of GAD67Ab correlated with those of GAD65Ab (r2 = 0.5913; p = 0.009). GAD65Ab directed to GAD65-M were found in 104 of 155 (67%), to GAD65-C in 104 of 155 (67%) and to GAD65-M + C in 116 of 155 (75%) of IDDM sera, and indicated reactivity to at least two distinct epitopes. Among the nine GAD65Ab-positive healthy children, two (22%) were also positive with GAD67, nine (100%) with GAD65-M + C, seven (78%) with GAD65-M, eight (89%) with GAD65-C and two (22%) with GAD65-N-67. Titres of GAD65Ab (p = 0.007), GAD65-C-Ab (p = 0.002) and GAD65-C + M-Ab (p = 0.003), but not of GAD65-M-Ab (p = 0.101) were significantly higher in IDDM than in healthy children. We conclude that GAD65Ab in IDDM and healthy children are directed to middle and C-terminal epitopes, and propose that levels of antibodies specifically directed to the carboxy-terminal end of GAD65 may distinguish IDDM from healthy children.
Adolescent↗
Soft tissue infections and the diabetic foot.
Soft tissue infections are classified as local or spreading. Spreading soft tissue infections are potentially life-threatening conditions, requiring prompt diagnosis and treatment. The information presented is based on a literature review and the authors' clinical experience. Diagnosis of soft tissue infections is aimed at determining the level of infection (skin, fascia, muscle) and whether necrosis is present. The bacteriology of these infections is varied and is of secondary importance. Treatment of skin infections that have no dead tissue is with antibiotics alone. Infections at the fascial or muscle level and those with necrosis at any level require surgical debridement and adjuvant antibiotics. The feet of diabetic patients are prone to plantar forefoot ulcers associated with tissue destruction and infection. The vast majority are caused by mechanical factors. If local immune defenses are adequate, bacterial colonization occurs without infection. Most diabetic foot ulcers will respond to relief of pressure, which may require total contact casting. Antibiotics and debridement are required in infected or deep ulcers, or when the ulcer does not respond to total contact casting.
Anti-Bacterial Agents↗
Thyroiditis in the BB rat is associated with lymphopenia but occurs independently of diabetes.
The spontaneously diabetic BB rat is an excellent and well studied model for human insulin-dependent diabetes (IDDM), sharing many important features with the human disease. Similarities include an equal frequency of IDDM in males and females, production of antibodies against pancreatic cell antigens, and an MHC disease association. In addition, the BB rat shares with human IDDM patients an increased frequency of autoantibodies against the parietal cells of the stomach and colloid cells of the thyroid gland. Here we investigate the genetic basis of thyroiditis in the BB rat. Based on crosses between BB, Lewis and Fischer rats, we show that two susceptibility factors for diabetes--the lymphopenia trait present in diabetes prone BB rats and the MHC--also appear to be risk factors for thyroiditis. However, the nature of the susceptibility was different for the two autoimmune diseases, with lymphopenia being absolutely required for diabetes although it only conferred increased risk for thyroiditis. Also, in contrast to IDDM, the MHC conferred dominant susceptibility to thyroiditis. Despite these shared risk factors, diabetes per se did not show significant correlation with thyroiditis.
Animals↗
Use of solid phase automated sequencing to define HLA disparity between bone marrow donors and recipients.
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Alleles↗
Identification of autoantibody epitopes of glutamic acid decarboxylase in stiff-man syndrome patients.
Stiff-man syndrome is a neurologic disorder characterized by progressive rigidity of skeletal muscles. Deficiency of the neurotransmitter gamma-aminobutyric acid and autoantibodies to glutamic acid decarboxylase (GAD), the enzyme synthesizing gamma-aminobutyric acid, are closely associated with the disorder, although the relevant antigenic epitopes have not been identified. In the present study, sera from two patients with SMS was used in an immunoblotting assay with recombinant GAD67 (M(r) 67,000) and GAD65 (M(r) 65,000) isoforms to test whether SMS sera can recognize specific epitopes. We found that both SMS sera recognized the GAD65, but not the GAD67, isoform. Using 13 different synthetic GAD peptides to block the autoantibodies, two GAD65 epitopes were identified. One epitope recognized by both patients' sera, was blocked by the peptide representing amino acid residues 354-368. In one patient only, blocking was also observed by a peptide representing residues 390-402, which includes the binding site of the GAD cofactor, pyridoxal 5'-phosphate. A single amino acid substitution in GAD65 at position 401 (leucine to proline) and representing the analogous GAD67 sequence in this region significantly reduced the peptide's inhibitory effect. These findings suggest that SMS GAD autoantibodies share distinct GAD65 linear epitopes and that some SMS patients' autoantibodies may block the active site, explaining SMS GABA deficiency.
Adult↗
A novel radioligand binding assay to determine diagnostic accuracy of isoform-specific glutamic acid decarboxylase antibodies in childhood IDDM.
Insulin-dependent diabetes mellitus (IDDM) is associated with autoreactivity against GAD but the diagnostic sensitivity (positivity in disease) and specificity (negativity in health) of isoform-specific GAD antibodies have yet to be defined in assay systems suitable for screening large number of samples. One set of IDDM patient (n = 10) and control (n = 50) standard sera were used to develop quantitative antibody assays with in vitro synthesized recombinant 35S-methionine-labelled GAD65 and GAD67, respectively, and protein A-Sepharose to separate free from antibody-bound ligand. Binding levels were not normally distributed (p < 0.0001) and therefore, the diagnostic accuracy of GAD antibodies was analysed by the ROC plots in population-based, consecutively-diagnosed, recent onset, 0-14 year-old patients (n = 105), and matched, healthy control subjects (n = 157). The ROC plots showed that the diagnostic sensitivity of GAD65 antibodies was 77% and the specificity 92% compared with 8% and 98%, respectively for GAD67 antibodies. In the IDDM sera, GAD65 and GAD67 antibodies were concordant in 7% (6 of 81) and GAD65 antibodies and ICA in 89% (72 of 81) without a correlation between the autoantibody levels. Autoantibodies to recombinant human islet GAD65 are specific and sensitive markers for childhood IDDM in this immunoassay with in vitro synthesized 35S-methionine-labelled recombinant GAD.
Adolescent↗
Correct sequence of the A*3001 allele obtained by PCR-SSP typing and automated nucleotide sequencing.
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Alleles↗
The plantar plate of the lesser toes: an anatomical study in human cadavers.
The purpose of this study was to evaluate the anatomic structure and biochemical composition of the plantar plate of the lesser toes. Fresh frozen-human cadaveric feet were used to study 20 metatarsophalangeal and proximal interphalangeal plantar plates. The observations of foot dissections were compared with the finger volar plate. The plantar plate of the toe is a rectangular structure with a stout distal insertion and relatively flimsy proximal origin. The anatomic relationships to adjacent structures and composition are similar between the volar plates of the fingers and plantar plates of the toes. The plantar plate is known to experience extension forces that the volar plate does not experience. The weightbearing nature of the foot and forces imposed by toe-off may create chronic hyperextension of the metatarsophalangeal joint and predispose the plantar plate to attenuation or rupture, thus leading to instability of the metatarsophalangeal joint. These findings may explain in part the clinical condition of spontaneous metatarsophalangeal joint dislocation, most commonly found in the second toe.
Collateral Ligaments↗