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Biomedical subjects

M A Shaw

Publications and source records attributed to M A Shaw.

At least 37 records · Page 2Linked to original sources
PubMed1995

Evidence for exclusion of a mutation in NRAMP as the cause of familial disseminated atypical mycobacterial infection in a Maltese kindred.

In mice, susceptibility to intracellular infections in inbred strains is controlled by a single locus, Lsh/Ity/Bcg, and the gene responsible has been cloned and designated Nramp (Natural resistance associated macrophage protein). We have identified a group of related children who appear to have a single gene defect, inherited recessively, which results in increased susceptibility to myocabacterial infection. The immunological defect observed in the affected children resembles that in mice homozygous for the Lsh/Ity/Bcg susceptible allele. To test the hypothesis that a mutation in NRAMP is responsible for the immunodeficiency observed in the affected children, we have typed eight markers in the region of human 2q34-q37 where NRAMP, the human homologue of Nramp, maps. We have shown discordance with the defect in one family and the chromosomes in the three affected children have different haplotypes making it unlikely that inheritance of an ancestral mutation in the NRAMP gene is the cause of increased mycobacterial susceptibility in this group of children.

Animals↗
PubMed1995

Human genetic susceptibility and infection with Leishmania peruviana.

Racial differences, familial clustering, and murine studies are suggestive of host genetic control of Leishmania infections. Complex segregation analysis has been carried out by use of the programs POINTER and COMDS and data from a total population survey, comprising 636 nuclear families, from an L. peruviana endemic area. The data support genetic components controlling susceptibility to clinical leishmaniasis, influencing severity of disease and resistance to disease among healthy individuals. A multifactorial model is favored over a sporadic model. Two-locus models provided the best fit to the data, the optimal model being a recessive gene (frequency .57) plus a modifier locus. Individuals infected at an early age and with recurrent lesions are genetically more susceptible than those infected with a single episode of disease at a later age. Among people with no lesions, those with a positive skin-test response are genetically less susceptible than those with a negative response. The possibility of the involvement of more than one gene together with environmental effects has implications for the design of future linkage studies.

Adolescent↗
PubMed1995

Genomic organization and sequence of the human NRAMP gene: identification and mapping of a promoter region polymorphism.

BACKGROUND: Murine Nramp is a candidate for the macrophage resistance gene Ity/Lsh/Bcg. Sequence analysis of human NRAMP was undertaken to determine its role in man. MATERIALS AND METHODS: A yeast artificial chromosome carrying NRAMP was subcloned and positive clones sequenced. The transcriptional start site was mapped using 5' RACE PCR. Polymorphic variants were amplified by PCR. Linkage analysis was used to map NRAMP. RESULTS: NRAMP spans 12kb and has 15 exons encoding a 550 amino acid protein showing 85% identity (92% similarity) with Nramp. Two conserved PKC sites occur in exon 2 encoding the Pro/Ser rich SH3 binding domain, and in exon 3. Striking sequence similarities (57 and 53%) were observed with yeast mitochondrial proteins, SMF1 and SMF2, especially within putative functional domains: exon 6 encoding the second transmembrane spanning domain, site of the murine susceptibility mutation; and exon 11 encoding a conserved transport motif. No mutations comparable to the murine susceptibility mutation were found. The transcriptional initiation site mapped 148 bp 5' of the translational initiation codon. 440bp of 5' flanking sequence contained putative promoter region elements: 6 interferon-gamma response elements, 3 W-elements, 3 NF kappa B binding sites and 1 AP-1 site. Nine purine-rich GGAA core motifs for the myeloid-specific PU.1 transcription factor were identified, two combining with imperfect AP1-like sites to create PEA3 motifs. TATA, GC and CCAAT boxes were absent. A possible enhancer element containing the Z-DNA forming dinucleotide repeat t(gt),ac(gt),ac(gt),g was polymorphic (4 alleles; n = 4,9,10,11), and was used to map NRAMP to 2q35. CONCLUSIONS: This analysis provides important resources to study the role of NRAMP in human disease.

Alleles↗
PubMed1994

Genetic and physical mapping of 2q35 in the region of the NRAMP and IL8R genes: identification of a polymorphic repeat in exon 2 of NRAMP.

Recent interest has focused on the region of conserved synteny between mouse chromosome 1 and human 2q33-q37, particularly over the region encoding the murine macrophage resistance gene Ity/Lsh/Bcg (candidate Nramp) and members of the Il8r interleukin-8 (IL8) receptor gene cluster. In this paper, identification of a restriction fragment length polymorphism in the IL8RB gene in 35 pedigrees previously typed for markers in the 2q33-q37 interval provided evidence (lod scores > 3) for linkage between IL8RB and the 2q34-q35 markers FN1, TNP1, VIL1, and DES. Physical mapping, using yeast artificial chromosomes isolated with VIL1, confirmed that IL8RA, IL8RB, and the IL8RB pseudogene map within the NRAMP-VIL1 interval, with the physical distance (155 kb) from 5' LSH to 3' VIL1 representing approximately 3-fold that observed in the mouse. Partial sequencing of NRAMP confirmed the presence of the N-terminal proline/serine-rich putative SH3 binding domain in exon 2 of the human gene. Further analysis of Brazilian leprosy and visceral leishmaniasis pedigrees identified a rare second allele varying in a 9-nucleotide repeat motif of the exon 2 sequence but segregating independently of the disease phenotype.

Animals↗
PubMed1994

Genetic regulation of leishmanial and mycobacterial infections: the Lsh/Ity/Bcg gene story continues.

A common basis to genetic regulation of leishmanial and mycobacterial infections is provided by the action of the murine Lsh/Ity/Bcg gene in controlling the priming/activation of macrophages for antimicrobial activity. This relies on the TNF-alpha-dependent sustained expression of the inducible nitric oxide synthase (iNOS) gene responsible for the generation of large amounts of toxic nitric oxide (NO). The Lsh/Ity/Bcg gene has many pleiotropic effects, including differential expression of the early response gene KC following stimulation of macrophages with bacterial lipopolysaccharide (LPS) and mycobacterial lipoarabinomannan (LAM). The major signal transduction pathway involved in KC induction requires the generation of low levels of NO via constitutive nitric oxide synthase (cNOS) activity, leading to activation of guanylate cyclase and the cGMP-dependent kinase pathway. NO therefore appears to provide a common link between the early influence of Lsh in regulating the expression of genes which mediate many pleiotropic effects, and the later production of NO as the final effector mechanism for kill. The recently cloned candidate for Lsh/Ity/Bcg, designated Nramp for Natural resistance associated macrophage protein, encodes a polytopic integral membrane protein that has structural features common to prokaryotic and eukaryotic transporters and includes a conserved binding-protein-dependent transport motif which may be involved in interaction with peripheral ATP-binding subunits. The N-terminal sequence also carries a proline/serine rich putative SH3 binding domain, consistent with a role for tyrosine kinases in regulating Nramp function. (ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1994

Immune response in healthy volunteers vaccinated with BCG plus killed leishmanial promastigotes: antibody responses to mycobacterial and leishmanial antigens.

Antibody (IgG) responses to mycobacterial (BCG; PPD; Mycobacterium leprae soluble antigen, MLSA) and leishmanial (Leishmania mexicana LV4) antigens were measured in 208 initially PPD and leishmanin skin-test negative volunteers divided into four vaccine groups as follows: 68 received BCG plus killed promastigotes (group A), 47 received BCG alone (group B), 47 received killed promastigotes alone (group C), and 46 formed the diluent control (placebo, group D). Three vaccine doses were administered at 8-12 week intervals. Vaccinees were bled immediately prior to each vaccination, and again at 3- and 12-month follow-up. Skin tests were performed prevaccination, and again at the 3- and 12-month follow-up. Anti-BCG, anti-PPD and anti-MLSA IgG levels increased significantly in groups A and B receiving BCG. The presence of leishmanial antigen (with BCG) in the inoculum suppressed the IgG response to Mycobacterium tuberculosis/Mycobacterium bovis-related (PPD and BCG), but not M. leprae-related (MLSA)-related, antigens. A small but significant increase (relative to prevaccination level) in response to MLSA, but not to BCG or PPD was observed in the non-BCG-vaccinated groups. The background level of response to mycobacterial and leishmanial antigens was higher in the Venezuelan vaccinees than in non-endemic (British) volunteers. Responses to leishmanial antigen were not enhanced in the two vaccine groups receiving killed promastigotes (with/without BCG) compared with the BCG alone and placebo groups. Instead, all vaccine groups showed a pattern of response consistent with either (i) a response to the skin-test antigen or, more likely, (ii) seasonal endemic exposure to leishmanial antigen. Interestingly, this endemic response to leishmanial antigen was enhanced in the vaccine groups receiving BCG, despite the fact that group B received no leishmanial antigen in the vaccine inoculum. When prevaccination IgG levels (mean + 3 standard deviations) were used to determine a negative cut-off, a low percentage (< 38%) of vaccinees converted to responder status for either anti-mycobacterial or anti-leishmanial responses, and those who did would be classified as 'low-responder' status compared with titres observed in severe forms of disease. Hence, although there was evidence for a background endemic response to both leishmanial and mycobacterial antigens, there was no evidence that vaccination per se led to a potentially disease exacerbatory level of TH2-associated antibody response especially with respect to the anti-leishmanial response.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMed1993

An RFLP map for 2q33-q37 from multicase mycobacterial and leishmanial disease families: no evidence for an Lsh/Ity/Bcg gene homologue influencing susceptibility to leprosy.

The mycobacterial diseases leprosy and tuberculosis (TB) and the leishmaniases are characterized by a wide spectrum of disease phenotypes, and by the fact that the majority of individuals exposed to the causative organisms Mycobacterium leprae, M. tuberculosis and Leishmania sp. become infected but do not present with clinical disease. In order to determine whether a human homologue to the murine macrophage resistance gene Lsh/Ity/Bcg influences susceptibility to human disease, multicase families for all three diseases have been collected, and linkage analysis performed using a panel of markers in the region of human chromosome 2q33-q37 known to be conserved with the Lsh/Ity/Bcg-containing region of murine chromosome 1. Because of the paucity of available polymorphic markers/linkage information for 2q33-q37, data from 35 multicase leprosy, TB and visceral leishmaniasis families (310 individuals) were first pooled to produce a detailed RFLP map of the region. Peak LOD scores well in excess of 3 were observed for linkage between adjacent pairs of a more proximal (2q33-q35) set of markers CRYGP1, MAP2, FN1, TNP1, VIL1 and DES, and between adjacent pairs of a more distal (2q35-q37) set COL6A3, D2S55 and D2S3. These peak LOD scores and the corresponding values for theta were used in the MAP92 program to generate a multiple two-point map with gene order/map intervals (cM) of: CRYGP1-4.65-MAP2-3.45-FN1-5.95-TNP1-3.41-VIL1-3. 01- DES-20.14-COL6A-10.91-D2S55-3.67-D2S3. Although local support for the placement of loci in this order was weak (LOD < 2, except for DES-COL6A3 where LOD = 6.02), the map is consistent with the gene order for those loci (Cryg, Fn-1, Tp-1, Vil, Des, Col6a3) previously mapped in the mouse. Data from 17 multicase leprosy families (149 individuals) were further analysed for linkage between a putative disease susceptibility locus (DSL) controlling susceptibility to leprosy per se and each of the marker loci. Assuming 100% penetrance for the susceptibility allele, no positive LOD score was obtained for linkage between the DSL and any of the marker genes. Instead, the data provide convincing evidence (LOD scores < -2) that a DSL does not fall within 10-20 cM of CRYGP1, MAP2, TNP1, VIL1, DES or D2S55, or within 5-10 cM of FN1, COL6A3 or D2S3. This effectively excludes a putative DSL controlling susceptibility to leprosy per se from the entire region 2q33-q37.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Transforming growth factor-beta is the major mediator of natural suppressor cells derived from normal bone marrow.

We previously reported that murine bone marrow cells activated by interleukin-3 (IL-3) or granulocyte-macrophage colony-stimulating factor (GM-CSF) had potent nonspecific natural suppressor (NS) cell activity. In the present study, we demonstrated that these activated NS cells released a soluble factor (or factors) capable of nonspecifically inhibiting T cell mitogenic responses. Consistent with the properties of transforming growth factor-beta (TGF-beta), treatment of the NS supernates with heat failed to denature the factor, and in fact significantly increased its suppressive activity. The NS suppressor factor strongly inhibited proliferation of the TGF-beta-sensitive tumor cell line, A549. Cytokine activation of suppressive activity correlated with the production of a 10- to 13-kDa protein, consistent with the size of TGF-beta and rIL-3 induced a sevenfold increase in TGF-beta transcription. Finally, neutralizing anti-TGF-beta antibody inhibited the suppressive activity of the supernates, indicating that TGF-beta was responsible for most, if not all, of the suppression expressed by these bone marrow NS cells.

Animals↗
PubMed1992

Contemporary (1979-1988) and inferred historical status of headwater lakes in North Central Ontario, Canada.

Fifty-six headwater Canadian Shield lakes were repetitively sampled from 1979 to 88 to determine their response to changes in acidic deposition of the period. Annual wet sulphate loadings varied between 38 and 83 meq m(-2), with highest deposition in the late 1970s followed by somewhat lower but variable deposition in the 1980s. Median pH of the lakes increased 0.42 pH units from 1979 to 1985 and decreased by 0.15 units between 1985 and 1988. Short water renewal times (x=1.1 y) promoted rapid equilibration. Since lake were so responsive to changes in SO4(2-) inputs, they were at or near steady state at all times. Comparison of predicted original pH and ANC with 1979 data indicate a median decline of 0.45 pH units and a loss of 34 microeq litre(-1). ANC. Four of 9 lakes were found to be historically fishless, based on the continued presence of Chaoborus americanus in sediment cores. The remaining five lakes historically had fish populations, but fish were not collected in 1979 when pH ranged betwen 4.6 and 5.3. By 1987, fish species were found in five of these lakes where pH had increased on average by 0.9 pH units. Our data indicate that water quality improvements could allow for the reinvasion or resumption of recruitment for a significant number of Ontario lakes.

Journal Article↗
PubMed1992

Inoculated mouse model of Pneumocystis carinii infection.

A transtracheally inoculated mouse model of Pneumocystis carinii has been developed using BALB/c mice. The advantage of this strain of mice include that they are widely available, inexpensive, and were not infected with Pneumocystis before inoculation. Inoculated mice that were not treated had a mean infectivity score of 4.1 compared with inoculated mice treated with the effective anti-Pneumocystis drug combination of trimethoprim plus sulfamethoxazole, which had a mean infectivity score of 0.1, an approximately 4 log difference. The inoculated BALB/c mouse provides a model to serve as a valuable addition to rat models currently used, providing a source of organisms from a different host for cross-species comparisons and for studies of drug efficacy for therapy and prophylaxis. The inoculated mouse is especially cost effective and allows testing of compounds in short supply.

Aminoquinolines↗
PubMed1992

Serum growth hormone (GH)-binding protein/receptor: an important determinant of GH responsiveness.

Individual growth rates (or responses to GH therapy) and adult heights vary over a wide range. The reasons for this variation are poorly understood. Based on the reciprocal relationship between GH production and serum GH-binding protein/receptor (GH-BP), we hypothesized that genetic growth potential was achieved by a specific combination of GH-BP/receptor and GH production in each individual. To address the question whether GH production regulates GH-BP, or vice versa, we studied GH-deficient children, where one of the parameters, GH exposure, could be controlled through exogenous administration. Forty-three untreated prepubertal GH-deficient children were studied before and after 6 and 12 months of GH replacement therapy (0.18 mg/kg.week). Growth velocity, height, bone age, weight and their respective Z scores, serum GH-BP, and serum insulin-like growth factor I (IGF-I) were measured at each time point. The patients responded with significant increases in serum IGF-I, age-adjusted growth velocity, and height (P < 10(-6) for all). Before therapy, GH-BP correlated directly with chronologic and bone age (P < 10(-4), but not with either growth velocity or IGF-I. In contrast, GH-BP correlated strongly with the response to therapy whether assessed as the incremental change in IGF-I (P < 10(-6)) or as the increase in growth velocity (P approximately 0.003). GH treatment had no consistent effect on GH-BP/receptor levels. These findings support the concept that the GH-BP/receptor endowment is characteristic for an individual and plays a pivotal role in somatic growth. The GH-BP/receptor system and its ontogeny appears relatively independent of regulation by GH. Differences in individual GH-BP/GH receptor complement account for some of the variability in the response to GH, and GH-BP levels may serve as a predictor for the degree of response. The reciprocal relationship between GH production and GH-BP in normal subjects probably results from adjustment of GH secretion to accommodate the prevailing GH-BP/receptor environment.

Adolescent↗
PubMed1992

The role of body mass in the response to growth hormone therapy.

Obesity is associated with normal or increased growth despite diminished GH secretion compared to lean children. The mechanism by which adequate growth is maintained in the presence of low GH levels is unknown, but is possibly mediated at the GH receptor level. To probe this hypothesis, we examined the relationship between GH responsivity, body mass index (BMI) and plasma GH-binding protein (GH-BP)/receptor level in 43 GH-deficient children during treatment with a fixed dose of GH (0.18 mg/kg.week). Before treatment, BMI [expressed as standard deviation score (SDS) for age (BMI-SDS)] did not correlate with either growth velocity or serum insulin-like growth factor-I (IGF-I). In contrast, after 12 months of GH therapy BMI-SDS correlated directly with plasma IGF-I (P < 10(-5)) and growth velocity (P < 10(-3)). These findings parallel those obtained for GH-BP vs. the response to GH, suggesting that BMI and GH-BP are covariants. The interrelationships among BMI, GH-BP, and response to GH were further probed by multiple regression analysis. Partial correlation coefficients vs. response to GH were consistently stronger for GH-BP than for BMI-SDS, indicating that GH-BP is the dominant factor between these two covariants in determining responsiveness to GH. The data suggest a primary role for GH-BP/receptor levels in determining GH action, with secondary but significant effects of nutrition and degree of adiposity. The latter may be mediated through the impact of nutrition and body mass on GH-BP/receptor levels.

Body Height↗
PubMed1991

Genetic regulation of macrophage priming/activation: the Lsh gene story.

This paper describes functional and genetic studies on the macrophage resistance gene Lsh/Ity/Bcg first described almost two decades ago. Working in vitro with resident peritoneal, liver (Kupffer cells) and bone marrow derived macrophages from congenic B10 (LshS) and B10.L-LshR mice it has been possible to demonstrate that the final effector mechanism for the gene in regulating antileishmanial activity involves production of reactive nitrogen rather than reactive oxygen intermediates. This in turn is dependent upon priming/activation of macrophages for enhanced TNF-alpha release which acts back on the macrophage in an autocrine manner to increase nitric oxide production. The precise point at which Lsh acts to control macrophage priming/activation has not been identified, but studies of early response gene expression show differences in KC mRNA levels at 2 h after LPS stimulation, and in c-fos mRNA as early as 20 min after stimulation with PMA plus ionophore, in peritoneal macrophages from congenic LshS and LshR mice. Data available suggest that both negative and positive signals may be involved in macrophage priming/activation, with LshS macrophages down-regulating their capacity for continued response to the autocrine loop. Work in progress will examine the role of TPA and cAMP response element-binding proteins in regulating gene expression in Lsh congenic mice. A major new initiative has also commenced to clone the Lsh gene by reverse genetics using yeast artificial chromosomes to walk towards Lsh from the closet proximal and distal markers on mouse chromosome 1.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Effects of growth hormone-binding proteins on serum growth hormone measurements.

GH-binding proteins (GH-BPs) in human blood theoretically may interfere with measurements of immunoreactive GH by forming complexes with GH and competing with antibody reagents for ligand. Indeed, results of serum GH obtained by immunoassays are known to differ markedly depending on the assay employed. To assess the potential role of circulating GH-BPs in this phenomenon, we systematically examined their effect on GH measurement in four RIAs and two immunoradiometric assays. In all except one RIA, the effect of the BPs on tracer binding to antibody was mildly inhibitory. In both immunoradiometric assays, BPs increased the nonspecific association of the tracer with the solid phase reagent. However, these effects were minor at the BP concentrations realistically encountered in practice. Furthermore, the impact of BPs on GH standard curves and final results was negligible because BP effects fall into the category of nonspecific or zero dose counts, which are subtracted during data reduction. We conclude that GH-BPs are only a minor disturbance in GH immunoassays, which is completely compensated for by conventional assay design. Disparities among results yielded by different assays are probably not due to BP interference.

Binding, Competitive↗
PubMed1991

Short stature and decreased serum growth hormone-binding protein in the Mountain Ok people of Papua New Guinea.

Two circulating GH-binding proteins (GH-BP), one of which is related to the GH receptor, have been described. To assess their possible role in or link with determining statural growth, we measured their activity/level in the serum of 25 adult subjects from a short-statured population from the highlands of Papua New Guinea (Mountain Ok people, similar in stature to African pygmies) and in 25 normal-statured North American control subjects. The Mountain Ok people have normal levels of GH, insulin-like growth factor-I (IGF-I), IGF-II, and serum albumin and prealbumin in association with short stature, making them a unique study population. Levels of the high affinity (GH receptor-related) GH-BP were significantly lower in the Mountain Ok subjects than in the taller controls (5.2 +/- 3.0% vs. 10.6 +/- 3.9% GH bound/160 microL, respectively; mean +/- SD; P less than 0.001). In contrast, the low affinity GH-BP was comparable in Mountain Ok and control subjects. Because of the structural similarity between the GH receptor and GH-BP, these data suggest that a limitation in GH receptor/GH-BP endowment may be associated with short stature despite normal circulating IGF-I levels. Alternatively, it is possible that the GH-BP itself plays an important role in growth promotion. Thus, the GH receptor (and/or GH-BP) complement may be a determinant of the genetically programmed height achieved.

Adult↗
PubMed1991

Growth hormone-binding protein activity is inversely related to 24-hour growth hormone release in normal boys.

To investigate the physiological relationship between serum GH-binding proteins and 24-h GH release, we compared the 24-h GH pulse attributes in serum samples obtained at 20-min intervals to the serum GH-binding protein activity (GH-BP) from 38 normal boys between 7 5/12 and 18 4/12 yr of age. GH-BP was determined in a serum sample from each study (containing less than 1.0 micrograms/L GH) using a standardized GH-BP assay. GH-BP results are expressed as the percentage of [125I]human GH bound to the high affinity GH-BP complex (peak II) per 160 microL serum. There were significant inverse relationships between the high affinity (receptor-related) GH-BP and several characteristics of 24-h GH release. Specifically, GH-BP was significantly (P less than 0.005 for all), but negatively, correlated with mean 24-h GH concentration (r = -0.62), sum of the GH pulse amplitudes (r = -0.57), sum of the GH pulse areas (r = -0.55), interpulse mean GH concentration (r = -0.53), and number of GH pulses per 24 h (r = -0.53). In addition, GH-BP correlated positively with the mean time interval between pulses (r = 0.59). There was also a significant positive correlation (r = 0.75; P less than 0.001) between GH-BP and the subject's age-adjusted body mass index SD score (BMI-SDS). Each characteristic of 24-h GH release correlating inversely with GH-BP also correlated inversely with BMI-SDS (P less than 0.01 for all comparisons). GH-BP did not, however, correlate with plasma insulin-like growth factor-I levels, serum testosterone concentrations, or height SDS. Binding to the low affinity GH-BP (peak I) did not correlate significantly with any of the examined GH pulse attributes, BMI-SDS, or the degree of binding to the high affinity GH-BP (peak II). We conclude that an inverse relationship exists between the high affinity serum GH-BP and 24-h GH release in boys under normal physiological conditions. We speculate that abnormalities in this relationship probably also exist and may underlie some disorders of growth.

Adolescent↗