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J W Thomas

Publications and source records attributed to J W Thomas.

At least 55 records · Page 3Linked to original sources
PubMed1997

T cell-independent response to Brucella-insulin identifies a preimmune repertoire for insulin.

An early or preimmune repertoire for anti-insulin B cells was examined using the T cell-independent (TI) response to insulin conjugated to Brucella abortus (BA-ins). mAbs from the BA-ins response reflect a repertoire present 7 to 10 days before the first Ab-forming cells (AFC) are detected in primary T cell-dependent (TD) responses to insulin. Although 4 of 6 BA-ins mAb express IgG2 isotype, evidence for somatic mutation is limited. A total of 9 of 12 V regions are identical with known V(H) or Vkappa genes, and consensus sequences suggest two other V genes may be in germ-line configuration. The relative avidities (50% inhibition of binding) of TI anti-insulins cover a broad range and are consistent with a germ-line anti-insulin repertoire that is functionally diverse. The V(H)s of 5 mAb are from two subsets of J558 genes (205.1 and 186.3) that dominate the B cell pool of adult mice and are different from the V(H)s used by anti-insulin mAb in primary TD responses. One IgM anti-insulin (mAb 301) uses V(H)-J606 and Vkappa1, and this mAb binds beta cells. Other TI mAb use either Vkappa5 or Vkappa19.3 genes and are similar to Vkappa genes used by anti-insulin mAb from TD responses. The data show that mutations in germ-line genes are not required for measurable insulin binding by monospecific mAb from adult mice. The recurrent use of Vkappa genes in both early (TI response) and late (TD responses) suggest that these structures are important in insulin binding.

Amino Acid Sequence↗
PubMed1997

Restricted V gene repertoire in the secondary response to insulin in young BALB/c mice.

Previous data on the genetic origin of insulin Abs in BALB/c mice are derived from immune responses following a single or primary immunization. This primary response, however, may not represent the repertoire established by chronic insulin administration, which is used in the treatment of diabetes mellitus and in protocols to alter beta cell destruction in pre-insulin-dependent (type 1) diabetes mellitus. In this study, the genetic composition and structure of 16 anti-insulin mAbs from the secondary response of a single BALB/c mouse were determined. In contrast to studies on other protein Ags, the V genes in the secondary response to insulin show only a low level of somatic mutation and the majority of the response is represented by restricted sets of V(H) and Vkappa genes. The restriction of V genes is not chiefly due to oligoclonal expansion but reflects recurrent usage of structurally similar V regions of independent origin. Secondary anti-insulin V(H) genes in 12 of 16 mAbs are derived from related J558 subsets, and 8 of 16 mAbs used the Vkappa19.34 gene without mutations in CDRLs. The CDHR3s of half of the mAbs are generated from an uncommon second reading frame of DSP2, suggesting that the amino acid motif STMIT may contact insulin and contribute to selection of these B cells. A total of 14 of 15 mAbs bind autologous rodent insulin with 40 to 100% of the activity on human insulin, and all mAbs bind proinsulin. These data show that a preferred set of V(H), D, and Vkappa gene segments dominate secondary responses to insulin and suggest that specific structural interactions with epitopes that are shared with autologous insulin shape this secondary response.

Age Factors↗
PubMed1997

Identification and partial characterization of ectoATPase expressed by immortalized B lymphocytes.

EctoATPases are extracellular membrane-bound enzymes that catalyze the hydrolysis of the gamma phosphate from ATP. EctoATPase is expressed by activated and immortalized Epstein-Barr virus-transformed human peripheral blood B lymphocytes and murine B cell hybridomas. By contrast, ectoATPase activity is not expressed on nontransformed human peripheral blood B lymphocytes, murine spleen cells, or murine myeloma cells. The K(m) for ATP for the B cell ectoATPases ranged from 5 to 77 microM; the Vmax ranged from 48 to 129 pmol/ min/10(4) cells. The enzyme required Mg2+ for maximal activity with little dependence on Ca2+. ADP and purine and pyrimidine nucleoside triphosphates were competitive inhibitors of the catalytic reaction. A putative ectoATPase protein has been identified by Western blot analysis of membrane proteins from the immortalized B cells. Under reducing conditions, antiectoATPase antibodies cross-reacted with a 66-kDa protein from murine B cell hybridoma membranes. By contrast a 200-kDa protein from the B cell hybridoma membranes cross-reacted with the antibodies under nonreducing conditions, suggesting a disulfide-linked trimer. The antibodies also cross-reacted with a 66-kDa protein from human B cell membranes under reducing conditions, but did not cross-react with membrane proteins under nonreducing conditions. This suggests that the antibody epitope(s) recognized on the reduced human protein is masked under nonreducing conditions. Thus, this work demonstrates: (1) that ectoATPase may serve as a marker for B cell activation; and (2) mammalian and avian ectoATPases have conserved interspecies immunological epitopes and kinetic properties.

Adenosine Triphosphatases↗
PubMed1997

Preference for IgG mAb binding insulin in solution or on surfaces is related to immunoglobulin variable region structures.

Studies on insulin autoantibodies often show a lack of correlation between enzyme-linked immunosorbent assays (ELISAs) and fluid phase radioimmunoassays (RIAs). Similarly, a set of IgG anti-insulin monoclonal antibodies (mAb) from BALB/c mice are found to differ in their binding in ELISAs and RIAs. To understand the structural basis for differential insulin binding, soluble and complexed biotinylated insulin is used to confirm binding properties independent of insulin-coated plastic and radioiodination. The binding properties of intact mAb are also present in Fab fragments, indicating ligand preference is not related to valence or to the Fc portion of Ig. Analysis of binding to soluble or bound ligand in relationship to antibody variable (V) region structures indicates that differential binding in the two assays is a property of heavy chain variable region structure. Studies also show that limited amino acid replacements arising during maturation of the immune response may change the binding preference for an individual mAb. These findings indicate that differences in detection of insulin binding in solid phase and fluid phase are not artefactual but reflect intrinsic structural features of immunoglobulin interaction with insulin.

Amino Acid Sequence↗
PubMed1997

Is patient length of stay related to quality of care?

In this article, we investigate the relationship between hospital length-of-stay (LOS) and quality of care. We use hospital claims records from Medicare beneficiaries in Michigan to estimate condition-specific models for predicting patients' LOSs. With these models and a data set provided by Michigan Peer Review Organization, Inc. (MPRO), each patient's risk-adjusted LOS is then linked to a quality-of-care judgment (good care, poor care) from physician peer reviewers. LOS is a widely used indicator of hospital performance. Most commonly, it is viewed as an indicator of hospital efficiency and as a surrogate measure for costs, with hospitals having long average LOSs considered relatively inefficient in the use of resources and those with low LOSs considered to be efficient. Sometimes, however, LOS is assumed to relate to quality. For example, if hospitals were to respond to the financial incentives of prospective payment by attempting to lower costs by prematurely discharging patients, LOSs significantly lower than expected might be considered indicative of poor quality care. On the other hand, if poor quality of care causes complications, it would tend to extend LOSs. Under this assumption, longer than expected LOSs could be viewed as indicative of poor quality care. This article shows that in every one of the 13 clinical conditions examined, cases that received poor quality care had significantly longer risk-adjusted LOSs than cases whose care was of acceptable quality.

Health Services Research↗
PubMed1996

CD11b+CD28-CD4+ human T cells: activation requirements and association with HLA-DR alleles.

Engagement of CD28 induces a major costimulatory pathway required by many CD4+ T cells in addition to activation via the TCR. In the absence of signals provided by CD28, ligation of the TCR alone can induce anergy or apoptosis in CD28+ cells. However, we report here characterization of a distinct subset of CD4+ T cells that are CD28-. Three autoreactive CD4+ human T cell clones that could be activated to produce IL-2 and proliferate by anti-CD3 alone were found to lack expression of CD28. CD28- clones that were activated with anti-CD3 alone were not anergic to restimulation via CD3. The presence of CD28-CD4+ T cells was verified in peripheral blood, and their frequency ranged from 0% to >22% of CD4+ T cells in different individuals. The percentage of CD28-CD4+ T cells in the peripheral blood of 57 individuals was significantly correlated with specific class II MHC alleles. Persons with HLA-DRB1*0401 and DR1 alleles had significantly higher numbers of CD28- T cells, while individuals with HLA-DR2(15) had significantly fewer CD28-CD4+ T cells than the mean. Like the CD28- clones, CD28-CD4+ T cells isolated from peripheral blood proliferated upon CD3 cross-linking in the absence of costimulation. The finding that CD28-CD4+ T cells resist induction of anergy following engagement of the TCR in the absence of conventional costimulation demonstrates one mechanism by which autoreactive T cells can escape processes that censor self-reactivity. The MHC associations observed suggest a relationship with autoimmunity and loss of self-tolerance.

Alleles↗
PubMed1996

Kinetics of electron and proton transfer during the reaction of wild type and helix VI mutants of cytochrome bo3 with oxygen.

Site-directed mutagenesis was used to investigate the mechanism of electron and proton transfer in the ubiquinol oxidase, cytochrome bo3, from Escherichia coli. The reaction between the fully reduced form of the enzyme and dioxygen was studied using the flow--flash method. After rapid mixing of CO-bound enzyme with an O2-containing solution, CO was photodissociated, and the subsequent electron- and proton-transfer reactions were measured spectrophotometrically, the latter using a pH-indicator dye. In the wild-type, pure bo3 enzyme, without bound quinones, we observed a single kinetic phase with a rate constant of about 2.4 x 10(4) s-1, associated with formation of the ferry1 oxygen intermediate, followed by proton uptake from solution with a rate constant of about 1.2 x 10(4) s-1. Enzyme in which heme o instead of heme b was incorporated into the low-spin site displayed a slower ferry1 formation with a rate constant of about 3.6 x 10(3) s-1. Upon replacement of the acidic residue glutamate 286 in helix VI of subunit I with a nonprotonatable residue, electron transfer was slightly accelerated, and proton uptake was impaired. Mutations of other residues in the vicinity of E286 also resulted in a dramatic decrease of proton uptake, suggesting that the environment of this residue is important for efficient proton transfer. In the closely related cytochrome aa3 from P. denitrificans, the corresponding residue (E278) has been suggested to be part of a proton-transfer pathway [Iwata, S., Ostermeier, C., Ludwig, B., & Michel, H. (1995) Nature 376, 660-669]. The results are discussed in terms of a model for electron-proton coupling during dioxygen reduction.

Carbon Monoxide↗
PubMed1996

Somatically mutated B cell pool provides precursors for insulin antibodies.

Antibodies to insulin are products of autoreactive B lymphocytes that escape inactivation or clonal deletion and are examples of "clonal ignorance." To understand the genetic origin of Abs from clonally ignorant B cells, the roles of somatic mutation and germ-line V(H) structures were examined for two murine IgG1 mAb that bind human and rodent insulin. Engineered mAb constructs that express germ-line or mutated V(H) genes show that somatic mutations introducing aspartic acid in or adjacent to CDRH2 play a key role in insulin binding. When either of the two anti-insulin V(H) regions is returned to its germ-line (unmutated) sequence, neither mAb binds insulin and the germ-line-encoded mAb are not polyreactive. Reconstruction of the somatic evolution of insulin binding in both mAbs shows that a single mutation in CDRH2 is sufficient to generate anti-insulin activity from a nonbinding precursor. When the role of somatic mutation in the binding of rodent insulin is examined, autoreactivity is associated with single mutations in both Abs. Together these findings indicate that, despite a low mutation frequency, IgG insulin Abs may not be derived directly from germ-line (unmutated) precursors. The requirement for somatic mutation as a prerequisite for measurable insulin binding suggests these Abs have their origin in a previously mutated B cell pool as a consequence of the individual's immune history. Low avidity interaction with endogenous insulin may play a role in selection of these B cells and contribute to the origin of clonal ignorance.

Amino Acid Sequence↗
PubMed1996

Expression and functional expansion of fibroblast growth factor receptor T cells in rheumatoid synovium and peripheral blood of patients with rheumatoid arthritis.

OBJECTIVE: Rheumatoid arthritis (RA) is an inflammatory disorder of the diarthroidial joints, characterized by fibroblast proliferation, angiogenesis, and perivascular CD4+ T cell infiltration. The present study examined the interactions between fibroblast growth factor-1 (FGF-1) and T cells. METHODS: Synovial tissues from patients with RA or noninflammatory arthritis were examined for the expression of FGF-1 and its receptor, FGFR-1, by immunohistology and reverse transcriptase-polymerase chain reaction. Functional assays were used to detect enrichment of FGF-1-responsive peripheral CD4+ T cells in RA. RESULTS: FGF-1 is abundantly expressed by rheumatoid synovium. Enhanced expression of its receptor, FGFR-1, was found in perivascular CD4+ T cells. In addition, T cells that are activated by FGF-1 are increased in the peripheral blood of patients with RA, as compared with other inflammatory conditions. CONCLUSION: The increased frequency of peripheral T cells that respond to FGF-1 in RA is consistent with expansion of FGFR-1-expressing T cells in the rheumatoid synovium.

Arthritis, Rheumatoid↗
PubMed1996

Guide-wire entrapment by inferior vena caval filters: in vitro evaluation.

PURPOSE: To study guide-wire engagement and entrapment by inferior vena caval (IVC) filters. MATERIALS AND METHODS: Patterns of engagement were determined in six IVC filters with four commonly used guide wires in a simulated IVC. Force required to disengage guide wires from filters was measured. RESULTS: Eleven engagement patterns were identified with J-tipped guide wires; straight guide wires never engaged. The Simon-Nitinol filter engaged guide wires with the highest frequency (P < .05). The 15-J wire was engaged most frequently overall (P < .0005) but disengaged with forces not significantly different (P > .05) from those required to open the J. Entrapment occurred with only the Vena-Tech (VT) and 12-F stainless steel Greenfield (12SSG) filters and the 1.5-J and 3-J guide wires. The former guide wire became entrapped regardless of engagement pattern; the latter became entrapped only when engaged in the hole in the apex of the 12SSG and VT filters. CONCLUSION: Guide wires with a J tip 3 mm or less in radius are at risk for entrapment by the 12SSG and VT filters.

Catheterization, Central Venous↗
PubMed1996

Does risk-adjusted readmission rate provide valid information on hospital quality?

The purpose of this study was to determine whether readmission rates, adjusted to account for differences in clinical characteristics of patients treated, provide valid information on hospital quality of care. Twelve clinical conditions were selected. Condition-specific models to predict readmission risk were estimated using Medicare UB-82 claims data. These models then were used to calculate readmission risk probabilities for patients whose medical records had been reviewed for quality of care by a Medicare peer review organization. I hypothesized that patients who received poor quality care during a hospitalization episode would be more likely to be readmitted after discharge than those who received acceptable quality care. My analysis found no support for this hypothesis. In each of the 12 clinical conditions studied, readmission rates of cases that received poor quality care were essentially the same as those whose care was judged acceptable. This was true both for readmission rates that were adjusted for patients' demographic and clinical characteristics, and for unadjusted rates, such as those typically displayed in hospital report cards.

Confounding Factors, Epidemiologic↗
PubMed1995

Costimulation of human CD4+ T cells by fibroblast growth factor-1 (acidic fibroblast growth factor).

T cell infiltration is prevalent in wound healing, atherosclerosis, vascular lesions in chronic allograft rejection, and autoimmune diseases. Whether T cells play a role in the migration and proliferation of vascular smooth muscle cells and endothelial cells in these lesions is not known. We previously reported that some human T cells express FGF-1, a potent growth factor for vascular smooth muscle cells and endothelial cells. In this study, we extend this observation and examine the expression and function of FGF receptors on human T cells. Using reverse transcription-PCR, Northern analysis, and immunohistochemistry, we found that some human T cells also express high affinity FGF receptor 1 (FGFR-1) respond to FGF-1. In the presence of anti-CD3, exogenous FGF-1 functions as a costimulator for these T cells, while FGF-1 alone does not induce T cell proliferation. [3H]Thymidine incorporation is sevenfold higher in T cells costimulated with FGF-1 compared with stimulation with anti-CD3 alone. Using limiting dilution, we demonstrate that FGF-responsive T cells are present in normal peripheral blood at a mean frequency of 1:19780 (95% confidence limits, 1:15100-1:23000), and similar T cells are increased in the peripheral blood of heart transplant recipients (mean frequency, 1:4210; 95% confidence limits, 1:3420-1:6781). In addition, a subline of Jurkat, a human T cell tumor, expresses FGFR-1 receptor. The function of FGFR-1 receptor in Jurkat T cells is demonstrated by the production of IL-2 after stimulation with FGF-1 and anti-CD3. IL-2 levels are sevenfold higher in Jurkat T cells costimulated with FGF-1 compared with those stimulated with anti-CD3 alone. FGF-1 alone has no effect on Jurkat T cells. These findings thus provide evidence that a subset of human T cells expresses a receptor for vascular cell growth factors, and this receptor functions to increase IL-2 production consistent with costimulation. The potential role of FGF-responsive T cells in a variety of vascular and inflammatory lesions is discussed.

CD4-Positive T-Lymphocytes↗
PubMed1995

The highly conserved methionine of subunit I of the heme-copper oxidases is not at the heme-copper dinuclear center: mutagenesis of M110 in subunit I of cytochrome bo3-type ubiquinol oxidase from Escherichia coli.

A common feature within the heme-copper oxidase superfamily is the dinuclear heme-copper center. Analysis via extended X-ray absorption fine structure (EXAFS) has led to the proposal that sulfur may be bound to CuB, a component of the dinuclear center, and a highly conserved methionine (M110 in the E. coli oxidase) in subunit I has been proposed as the ligand. Recent models of subunit I, however, suggest that this residue is unlikely to be near CuB, but is predicted to be near the low spin heme component of the heme-copper oxidases. In this paper, the role of M110 is examined by spectroscopic analyses of site-directed mutants of the bo3-type oxidase from Escherichia coli. The results show that M110 is a non-essential residue and suggest that it is probably not near the heme-copper dinuclear center.

Amino Acid Sequence↗
PubMed1995

Physical localization of eed: a region of mouse chromosome 7 required for gastrulation.

In the mouse, the embryonic ectoderm development (eed) region is defined by deletions encompassing the albino (c) locus of chromosome 7. The region is located 1-2 cM distal to the c locus and was of undetermined size. Embryos homozygous for deletions removing eed display defects in axial organization during gastrulation. Two loci, identified by chemical mutagenesis, are known to map within the eed interval. One, l7Rn5, probably represents the gene required for gastrulation. The second, l7Rn6, is required for survival after birth. fit1, a third locus identified by chemical mutagenesis, maps distal to the eed interval and is also required for survival after birth. A 900-kb YAC contig has been constructed, and deletion breakpoints defining the limits of the regions containing these loci have been localized. Their positions place the eed region within a maximum 150-kb interval at the proximal end of the contig, while fit1 maps to a 360-kb interval within the middle of the contig. Several clusters of rare-cutting restriction sites map within these regions and represent potential locations of candidate genes.

Animals↗
PubMed1995

Potent interleukin 3 receptor agonist with selectively enhanced hematopoietic activity relative to recombinant human interleukin 3.

A systematic evaluation of structure-activity information led to the construction of genetically engineered interleukin 3 (IL-3) receptor agonists (synthokines) with enhanced hematopoietic potency. SC-55494, the most extensively characterized member of this series, exhibits 10- to 20-fold greater biological activity than recombinant human IL-3 (rhIL-3) in human hematopoietic cell proliferation and marrow colony-forming-unit assays. In contrast, SC-55494 is only twice as active as rhIL-3 in priming the synthesis of inflammatory mediators such as leukotriene C4 and triggering the release of histamine from peripheral blood leukocytes. The enhanced hematopoietic activity of SC-55494 correlates with a 60-fold increase in IL-3 alpha-subunit binding affinity and a 20-fold greater affinity for binding to alpha/beta receptor complexes on intact cells relative to rhIL-3. SC-55494 demonstrates a 5- to 10-fold enhanced hematopoietic response relative to its ability to activate the priming and release of inflammatory mediators. Therefore, SC-55494 may ameliorate the myeloablation of cancer therapeutic regimens while minimizing dose-limiting inflammatory side effects.

Amino Acid Sequence↗