Biomedical subjects
Publications and source records attributed to J Naylor.
The human genome holds an extraordinary trove of information about human development, physiology, medicine and evolution. Here we report the results of an international collaboration to produce and make freely available a draft sequence of the human genome. We also present an initial analysis of the data, describing some of the insights that can be gleaned from the sequence.
OBJECTIVES: To provide optimum health care to indigenous people with diabetes, to prevent diabetes, and to monitor the epidemiology of diabetes and selected complications. The purposes of this paper are to describe the program and to present data that highlights the major problems and successes. STUDY DESIGN: Descriptive epidemiology report of diabetes and population service program based on yearly chart review data. METHODS: Almost half of Alaska Natives with diabetes have no direct access to physicians or hospitals. Health care delivery is now managed by the tribes themselves. Program emphases include maintenance of a population-based registry, formal training for village health aides, physical activity programs, patient education, primary prevention activities and adherence to standards of care to prevent complications. A centralized registry is maintained to assure that epidemiological data is available and patients are not lost to follow-up. Each year a random sample of charts at each major facility is audited against nationally standardized care guidelines. RESULTS: The prevalence of diabetes among Alaska Natives increased 80% over the 13 years from 1985 to 1998 (15.7/1000 to 28.3/1000, age adjusted to U.S. 1980 population). For the years 1986-1998 the incidence rates of lower extremity amputation and end stage renal disease were 6.1/1000 and 2.0/1000 respectively. The level of care provided to Alaska Native patients is comparable to that provided to the general diabetic patient population seen in Alaskan urban clinics. CONCLUSIONS: In spite of logistic challenges, care provided to Alaska Native people with diabetes compares favorably to that provided in other settings. Incidence rates of lower extremity amputation and end stage renal disease also remain comparable to or lower than those in other U.S. populations. Many aspects of our system could be extended to other chronic disease programs serving isolated indigenous populations. Primary prevention of diabetes remains a major challenge as life styles change.
In order to evaluate the potential of TCR as vaccines for immunomodulation, the immunogenicity of soluble versions of D10 TCR has been investigated in mice. Soluble D10 TCR containing the extracellular domains were produced either as dual chain (dc) TCR lacking transmembrane and cytoplasmic regions or as a TCR-IgG1 chimeric protein. Soluble single chain (sc) D10 TCR contained only the Valpha and Vbeta segments joined by a peptide linker. Syngeneic D10 dcTCR or D10 TCR-IgG1 immunizations of AKR mice induced antibody responses to D10 clonotypic epitopes and to constant region epitopes that are not exposed on D10 cells. Only clonotypic antibodies were produced after D10 scTCR immunizations. Immunization of AKR mice with D10 dcTCR and D10 TCR-IgG1 primed I-Ak- and I-Ek-restricted CD4+ T cells recognizing constant region epitopes, but there was no detectable response to the variable region. Comparison of the in vitro proliferative responses of CD4+ T cells from D10 scTCR-primed H-2 congenic mice revealed that H-2u was a responder haplotype for the variable region. How the immunogenicity of particular regions of the TCR appears to be shaped by tolerance induction in vivo and the implications for immunotherapy with soluble TCR vaccinations are discussed.
Maternal transfer of TCR clonotypic Ab protected young NOD mice against the adoptive transfer of diabetes by the BDC 2.5 T cell clone. The effect of maternal anti-TCR Vbeta-8 Ab on T cell development and function has now been investigated. SJL/J mice, which lack TCR Vbeta-8, were immunized with soluble, chimeric D10 TCR-IgG1 containing Vbeta-8.2. The (SJL/J x AKR/J) F1 offspring of immunized female SJL/J mice were severely depleted of peripheral T cells bearing Vbeta-8 until 11 to 17 wk of age. The loss of Vbeta-8 expression did not appear to be due to modulation of cell surface TCR. Since the Vbeta-8+ T cell population was unperturbed in the (AKR/J x SJL/J) F1 offspring of D10 TCR-IgG1-immunized AKR/J mothers making D10 clonotypic Ab, the effect was immunologically specific. The deletion of Vbeta-8+ T cells had functional consequences. In the in vitro response to the superantigen, staphylococcal enterotoxin B, the usually observed participation of Vbeta-8.2+ T cells was largely suppressed, whereas the recruitment of Vbeta-3+ T cells remained unaltered. In control mice, T cell responses to the 134- to 146-residue peptide of conalbumin (pCA(134-146)) were biased toward use of Valpha-2/Vbeta-8.2 TCR. In D10 TCR-IgG1 maternally immunized (SJL x AKR/J) F1 mice, the T cell responses to pCA(134-146) were suppressed, and T cell lines derived from these in vitro were devoid of Vbeta-8.2 expression. With an increased understanding of TCR V gene usage in autoimmune diseases, similar strategies for the depletion of autoreactive T cells may become feasible in humans.
The BDC 2.5 T cell clone is specific for pancreatic beta-cell antigen presented by I-Ag7, and greatly accelerates diabetes when injected into 10-21-d-old nonobese diabetic (NOD) mice. The BDC 2.5 T cell receptor (TCR) has been solubilized as a TCR-IgG1 chimeric protein. All NOD mice immunized against BDC 2.5 TCR-IgG1 produced antibodies recognizing TCR C alpha/C beta epitopes that were inaccessible on the T cell surface. 56% of the mice produced antibodies against the BDC 2.5 clonotype that specifically blocked antigen activation of BDC 2.5 cells. We have used the adoptive transfer model of diabetes to demonstrate that maternal immunization with soluble TCR protects young mice from diabetes induced by the BDC 2.5 T cell clone.
Analysis of mRNA in two haemophilic monozygotic twins offers novel information on the organisation of expressed sequences distal to the coagulation factor VIII gene. These patients show an inversion that, in contrast to the common inversions responsible for 1/5 of all haemophilia A, affects the first rather than intron 22 of the gene. This displaces the most telomeric of the factor VIII exons (exon 1) by approximately 100 kb towards the telomere, and close to the region of the C6.1A gene. This novel inversion creates two hybrid transcription units: one formed by the promoter and first exon of the factor VIII gene followed by a widely expressed sequence; the other by the promoter and coding region of the C6.1A gene plus most of the factor VIII gene (part of intron 1 and exons 2-26). Investigation of this transcription unit reveals that the C6.1A gene has an unsuspected intron in the region coding for the previously described 3'-untranslated tail of the message. Furthermore, exons located beyond the known C6.1A sequence and present in normal transcripts precede exons 2-26 of the factor VIII gene in the hybrid mRNA of the haemophilic twins. The factor VIII sequences in this hybrid mRNA are not expected to be expressed because they lack the first exon, encoding the prepeptide, and follow a translation stop in the C6.1A gene. Leukaemia-related translocations in the C6.1A region suggest that this region may be somewhat unstable.
Freshwater crayfish, Austropotamobius pallipes, caught in the winter when water temperature was 5 °C, were acclimated in the laboratory to 5 °C for 1 week and to 1 and 12 °C for 1 month before haemolymph and tissue samples were taken for acid­base analysis. In vivo, haemolymph (extracellular) pH increased by 0.14 units between 5 and 1 °C but remained unchanged between 5 and 12 °C, giving an overall deltapHe/deltat value of -0.009 pH units °C-1 and an [H+] value of +3.3 nmol l-1. Haemolymph withdrawn from crayfish acclimated to 1 °C and warmed anaerobically in vitro to 12 °C had a deltapHe/deltat value of -0.010 pH units °C-1. This was depressed compared with the values of -0.015 and -0.016 pH units °C-1 obtained from haemolymph withdrawn from crayfish held at 5 and 12 °C, respectively, but changes in H+ concentration were similar in all three groups. There was little change in intracellular pH with rise in temperature in either claw muscle (deltapHi/deltat=-0.003 pH units °C-1) or hepatopancreas (deltapHi/deltat=-0.006 pH units °C-1). In the heart, deltapHi/deltat paralleled that in the haemolymph (-0.010 pH units °C-1), but the largest change was found in the abdominal muscle (-0.025 pH units °C-1). In all tissues apart from claw muscle, there was a metabolic acidosis, with an increase in metabolic acid load as temperature rose from 1 to 12 °C. The depressed in vivo deltapHe/deltat value and the differences in deltapHi/deltat between the tissues were attributed to active regulation of pHi, possibly to control enzyme function in the intracellular compartments. Tissues that are functionally important in the winter months, such as abdominal muscle, followed alphastat regulation, whereas tissues that are less active in the winter, such as claw muscle and hepatopancreas, maintained pHi irrespective of temperature, resulting in a relatively acidotic pHi, which may serve to depress the activity of metabolic enzymes.
Surprisingly half of all severe haemophilia A patients have no mutation in the promoter, coding sequences and normal RNA processing signals of the factor VIII gene. Instead they manifest a unique mRNA defect that prevents the amplification of the message across the boundary between exon 22 and 23. This locates the defect to internal regions of intron 22. Novel sequences 3' to exon 22 were isolated from the 9 available patients with the above abnormality by combining RACE and vectorette amplifications on trace amounts of mRNA. This showed that exons 1-22 of the factor VIII mRNA had become part of a hybrid message containing new multi exonic sequences expressed in normal cells. The novel sequences were not located in a YAC covering the whole factor VIII gene. Southern blots from patients probed by novel sequences and clones covering intron 22 showed no obvious abnormalities. This suggested inversions involving intron 22 repeated sequences. Screening of 3 YAC libraries with the novel sequences located them at least 200 kb telomeric (5') to factor VIII and pulsed field gel analysis detected abnormal bands in patients. This demonstrates that the mutations in the patients are inversions of long DNA regions possibly involving the repeated sequences and occurring at the surprising rate of approximately 4 x 10(-6) per gene per gamete per generation.
AIM: To analyse the configuration of the antigen receptor genes in Hodgkin's disease. METHODS: DNA extracted from 45 samples of Hodgkin's disease was analysed using Southern blotting and DNA hybridisation, using probes to the joining region of the immunoglobulin heavy chain gene, the constant region of kappa immunoglobulin light chain gene, and the constant region of the beta chain of the T cell receptor gene. RESULTS: A single case of nodular sclerosing disease showed clonal rearrangement of the immunoglobulin heavy and light chain genes, all other samples having germline immunoglobulin genes. The nature of the clonal population in the diseased tissue is uncertain, because the intensity of the rearranged bands did not correlate with the percentage of Reed-Sternberg cells present. The T cell receptor genes were in germline configuration in all the samples. CONCLUSIONS: Antigen receptor gene rearrangement is a rare finding in unselected cases of Hodgkin's disease.
We studied an immunotoxin consisting of recombinant ricin A chain (rRA) conjugated to 454A12 MoAb, a monoclonal antibody which recognizes an epitope on the human transferrin receptor, and compared the ability of 454A12 MoAb-rRA immunotoxin to inhibit the growth of erythroid burst-forming units (BFU-e) and myeloid colony-forming units (CFU-c) with unconjugated 454A12 MoAb. A significant reduction in BFU-e colony growth was observed at 0.001 microgram/ml of 454A12 MoAb-rRA versus 0.1 microgram/ml of unconjugated 454A12 MoAb (p = 0.005). Comparison of the effects of 454A12 MoAb-rRA and 454A12 MoAb on myeloid colony development gave markedly different results. Unconjugated antibody had no effect on CFU-c colony growth; in contrast, 0.01 microgram/ml of 454A12 MoAb-rRA reduced the number of colonies from 139 per 1 X 10(5) to 75 per 1 X 10(5) cells plated (p = 0.0005). No myeloid progenitor colonies developed at 0.1 microgram/ml of immunotoxin. These observations suggest that 454A12 MoAb-rRA inhibits growth by a potent, ricin A chain-mediated toxic effect on any proliferating cells expressing transferrin receptors, whereas the 454A12 MoAb exerts a selective inhibitory effect primarily on erythroid progenitors by perturbing the transferrin cycle. While growth factor receptors expressed on hematopoietic cells represent promising targets for immunotoxin therapy, our data indicate that an immunotoxin could inhibit cellular proliferation by a different mechanism than the corresponding unconjugated MoAb. Depending on the antibody used, these differences may be important in trials using immunotoxins for in vivo treatment or in vitro purging of malignant hematopoietic cells.
In two separate studies comparing the handedness of patients suffering from myasthenia gravis with matched controls, no evidence was found to support the Geschwind Behan hypothesis of an association between autoimmune disease and left-handedness. Counter to prediction both studies found marginally lower incidences of left-handedness in myasthenics, and when combined with the similar result of Cosi et al. (Cortex 24, 573-577, 1988) the difference was highly statistically significant. The personality of myasthenics, as assessed by the Eysenck Personality Questionnaire (EPQ), and in particular the psychoticism scale, which has been postulated to be related to androgen levels, was not significantly different from controls. However, assessment of sex-roles using the Bem Sex Role Inventory suggested that female myasthenics were more masculine than controls.
We have identified regions of the major capsid protein, L1, of the human papillomavirus (HPV) type 16 (HPV-16 L1), that are recognized by five monoclonal antibodies (MAbs) raised to a bacterial fusion protein containing residues 172 to 375 of HPV-16 L1. All five MAbs recognized HPV-16-infected tissue sections by immunohistochemistry, but not sections infected with HPV-1a (cutaneous warts), HPV-6b or -11 (genital warts). MAbs 3D1, 5A4 and 1D6 also recognized HPV-2-infected sections (cutaneous warts); MAb 8C4 recognized only sections containing HPV-16. Four MAbs (8C4, 3D1, 1D6 and 5A4) recognized a synthetic peptide corresponding to residues 269 to 284 of HPV-16 L1; within this region a minimum antibody binding site was identified, a tripeptide 276 to 278. However the complete epitope appears to extend beyond these residues and beyond HPV-16 L1 (269 to 284). The fifth MAb, 1C6, recognized bacterial fusion proteins containing HPV-6b L1, -16 L1 or -18 L1 using immunoblots, yet appeared HPV-16-specific when tested on infected tissue sections. This MAb recognized five amino acids within a different region of HPV-16 L1 (residues 299 to 313).
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Spigelian and Richter's hernias, individually, are both moderately rare. To encounter a patient having both of these surgical diseases as a single entity is indeed unique. Such a patient with an acute partial enterocele of cecum (Richter's hernia) strangulated in a lateral ventral musculoaponeurotic defect (Spigelian hernia) was successfully treated with wedge cecectomy and simple herniorraphy.
Three, single-day nutritional surveys at weekly intervals were conducted in the general medical wards of an urban municipal teaching hospital. The techniques of nutritional assessment included anthropometric measures (weight/height, triceps skin fold, arm-muscle circumference, serum albumin, and hematocrit). The prevalence of protein-calorie malnutrition was 44% or greater by these criteria (weight/height, 45%; triceps skin fold, 76%; arm-muscle circumference, 55%; serum albumin, 44%; and hematocrit, 48%). These results were reproducible without significant variation between surveys. In 34% of patients, a lymphopenia of 1,200 cells/cu mm or less was found, a level likely to be associated with diminished cell-mediated immunity. Compared with a similar survey among surgical patients, the medical patients were more depleted calorically (weight/height, triceps skin fold) but had better protein status (arm-muscle circumference, serum albumin). Significant protein-calorie malnutrition occurs commonly in municipal hospitals in both medical and surgical services.
Ribosomal preparations from Vibrio cholerae Ogawa and Inaba are protective immunogens for mice challenged with either serotype. Column chromatography of ribosomal fractions separated protective antigen from the ribosomes. The antigen is a heterogeneous colloid which contains protein, lipid, and carbohydrate in the ratio 3:1:1. Amino acid composition is that of a slightly acidic protein with no unusual complement of amino acids. The lipid component consists of several longchain fatty acids and of phosphatidyl ethanolamine. The major sugars identified were glucose and galactose. The colloid can be dissolved by treatment with chelating agents and sodium dodecyl sulfate. Purification of the solubilized material or of the colloid was unsuccessful.