Biomedical subjects

S M Smith

Publications and source records attributed to S M Smith.

At least 73 records · Page 4Linked to original sources

PubMed2000

Human CD8+ CTL specific for the mycobacterial major secreted antigen 85A.

The role of CD8(+) CTL in protection against tuberculosis in human disease is unclear. In this study, we stimulated the peripheral blood mononuclear cells of bacillus Calmette-Guérin (BCG)-vaccinated individuals with live Mycobacterium bovis BCG bacilli to establish short-term cell lines and then purified the CD8(+) T cells. A highly sensitive enzyme-linked immunospot (ELISPOT) assay for single cell IFN-gamma release was used to screen CD8(+) T cells with overlapping peptides spanning the mycobacterial major secreted protein, Ag85A. Three peptides consistently induced a high frequency of IFN-gamma responsive CD8(+) T cells, and two HLA-A*0201 binding motifs, P(48-56) and P(242-250), were revealed within the core sequences. CD8(+) T cells responding to the 9-mer epitopes were visualized within fresh blood by ELISPOT using free peptide or by binding of HLA-A*0201 tetrameric complexes. The class I-restricted CD8(+) T cells were potent CTL effector cells that efficiently lysed an HLA-A2-matched monocyte cell line pulsed with peptide as well as autologous macrophages infected with Mycobacterium tuberculosis or recombinant vaccinia virus expressing the whole Ag85A protein. Tetramer assays revealed a 6-fold higher frequency of peptide-specific T cells than IFN-gamma ELISPOT assays, indicating functional heterogeneity within the CD8(+) T cell population. These results demonstrate a previously unrecognized, MHC class I-restricted, CD8(+) CTL response to a major secreted Ag of mycobacteria and supports the use of Ag85A as a candidate vaccine against tuberculosis.

Acyltransferases↗

PubMed2000

Estrogen protects against vaginal transmission of simian immunodeficiency virus.

Postmenopausal women and women who use injectable, progestin-based contraceptives are at increased risk of human immunodeficiency virus (HIV) infection, suggesting that progesterone and estrogen affect HIV-1 vaginal transmission. To evaluate the individual roles of these sex hormones in vaginal transmission, ovariectomized female macaques were treated with either progesterone or estrogen followed by intravaginal inoculation with SIVmac. All 6 untreated control macaques and 5 (83%) of 6 progesterone-treated animals became infected following intravaginal SIV inoculation. Conversely, none of 6 estrogen-treated macaques was infected. Vaginal subepithelial inoculation of estrogen-treated animals resulted in infection, which shows that the block occurred at the vaginal epithelium and/or lumen. These data suggest that estrogen-deficient women are at increased risk of HIV infection, because their vaginal microenvironments are rendered more susceptible. Moreover, topical vaginal estrogen therapy may be an effective means of reducing HIV vaginal transmission in these high-risk groups.

Animals↗

PubMed2000

Postgerminative growth and lipid catabolism in oilseeds lacking the glyoxylate cycle.

The glyoxylate cycle is regarded as essential for postgerminative growth and seedling establishment in oilseed plants. We have identified two allelic Arabidopsis mutants, icl-1 and icl-2, which lack the glyoxylate cycle because of the absence of the key enzyme isocitrate lyase. These mutants demonstrate that the glyoxylate cycle is not essential for germination. Furthermore, photosynthesis can compensate for the absence of the glyoxylate cycle during postgerminative growth, and only when light intensity or day length is decreased does seedling establishment become compromised. The provision of exogenous sugars can overcome this growth deficiency. The icl mutants also demonstrate that the glyoxylate cycle is important for seedling survival and recovery after prolonged dark conditions that approximate growth in nature. Surprisingly, despite their inability to catalyze the net conversion of acetate to carbohydrate, mutant seedlings are able to break down storage lipids. Results suggest that lipids can be used as a source of carbon for respiration in germinating oilseeds and that products of fatty acid catabolism can pass from the peroxisome to the mitochondrion independently of the glyoxylate cycle. However, an additional anaplerotic source of carbon is required for lipid breakdown and seedling establishment. This source can be provided by the glyoxylate cycle or, in its absence, by exogenous sucrose or photosynthesis.

Acetates↗

PubMed2000

A c-fos/Estrogen receptor fusion protein promotes cell cycle progression and proliferation of human cancer cell lines.

c-fos is the prototypic member of a family of transcription factors that regulate many cellular processes, including proliferation. c-fos heterodimerizes with jun family members to form the AP-1 transcription factor complex which binds specific DNA recognition elements in the promoters of many genes. Following rapid induction in response to serum or growth factors, c-fos regulates expression of downstream target genes involved in cellular proliferation. Although much work has focused on activation of cell cycle regulatory genes by c-fos, less is known about negative regulation of gene expression by this transcription factor. The cyclin-dependent kinase (cdk) inhibitor p21(Cip1/WAF1) is a negative regulator of cdk activity, thereby impeding cell cycle progression. By sequence analysis, we identified a putative AP-1 element in the p21(Cip1/WAF1) promoter. To investigate how this site regulated p21(Cip1/WAF1) expression and mitigate external effects on c-fos expression, we used a c-fos/estrogen receptor (c-fosER) fusion construct in which this transcription factor is conditionally activated by estradiol. In the presence of estradiol, c-fosER downregulated p21(Cip1/WAF1) promoter activity. This inhibition was dependent on the putative AP-1 site. Activation of c-fosER induced cell cycle progression and proliferation in a manner similar to serum stimulation. We concluded that activation of c-fosER mediated transcriptional inhibition of p21(Cip1/WAF1) through a previously uncharacterized AP-1 site, revealing an important role for c-fos in negative control of cell cycle regulatory genes.

Cell Cycle↗

PubMed2000

Molecular characterization and developmental expression of the aryl hydrocarbon receptor from the chick embryo.

The aryl hydrocarbon receptor (AhR) was cloned from the chick embryo and its function and developmental expression characterized. Chicken AhR cDNA coded for 858 amino acid protein and 396 bp of 3' UTR. The basic helix loop helix domain exhibited 87-100% amino acid identity to avian, mammalian, and amphibian AhR, and 69-74% to piscine AhR. The PAS (Per-ARNT-Sim) region was slightly less well conserved with (a) 97% identity to other avian sequences, (b) 81-86% to amphibian and mammalian AhR, and (c) 64-69% with piscine AhR. The carboxy terminus diverged the most among species with less than 53% amino acid identity between chicken and any available mammalian and piscine AhR sequences. The chicken AhR RNA and protein were 6.1 kb and 103 kDa, respectively. Chicken AhR dimerized with human AhR nuclear translocator and bound the mammalian dioxin-response element in a ligand-dependent manner. AhR protein was detected in neural ganglia; smooth, cardiac, and skeletal muscle; and epithelium involved in epithelial-to-mesenchymal transformations, such as pituitary, gastrointestinal tract, limb apical-ectodermal ridge, and kidney collecting ducts. AhR mRNA was detected in all tissues expressing protein, except myocardium. Cytochrome P4501A4 mRNA was highly induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in a subset of tissues expressing AhR, including small intestine, liver, kidney, blood vessels, and outflow tract myocardium. In conclusion, the AhR sequence and function is highly conserved between birds and mammals, and although many tissues express AhR during chick embryo development, only a subset are responsive to TCDD induction of CYP1A4.

Amino Acid Sequence↗

PubMed2000

Vaccinia expression of Mycobacterium tuberculosis-secreted proteins: tissue plasminogen activator signal sequence enhances expression and immunogenicity of M. tuberculosis Ag85.

There is increasing evidence to implicate a role for CD8(+) T cells in protective immunity against tuberculosis. Recombinant vaccinia (rVV) expressing Mycobacterium tuberculosis (MTB) proteins can be used both as tools to dissect CD8(+) T-cell responses and, in attenuated form, as candidate vaccines capable of inducing a balanced CD4(+)/CD8(+) T-cell response. A panel of rVV was constructed to express four immunodominant secreted proteins of MTB: 85A, 85B and 85C and ESAT-6. A parallel group of rVV was constructed to include the heterologous eukaryotic tissue plasminogen activator (tPA) signal sequence to assess if this would enhance expression and immunogenicity. Clear expression was obtained for 85A, 85B and ESAT-6 and the addition of tPA resulted in N-glycosylation and a 4-10-fold increase in expression. Female C57BL/6 mice were immunised using the rVV-Ag85 constructs, and interleukin-2 and gamma-interferon were assayed using a co-culture of immune splenocytes and recall antigen. There was a marked increase in cytokine production in mice immunised with the tPA-containing constructs. We report the first data demonstrating enhanced immunogenicity of rVV using a tPA signal sequence, which has significant implications for future vaccine design.

Acyltransferases↗

PubMed2000

Postdictors of eyewitness errors: can false identifications be diagnosed?

Eyewitness researchers have "postdicted" identification accuracy using witness confidence (S. L. Sporer, S. Penrod, D. Read, & B. Cutler, 1995), response latency (S. L. Sporer, 1993, 1994), and endorsement of statements consistent with using relative versus absolute judgment strategies (D. Dunning & L. B. Stern, 1994; R. C. L. Lindsay & K. Bellinger, 1999). All of these measures were collected from 321 introductory psychology students who had viewed a staged crime and completed a lineup identification task. Some participants received feedback after identification (G. L. Wells & A. L. Bradfield, 1998). Lineup fairness was also used as a postdictor of eyewitness accuracy. Discriminant function analysis indicated that 75.2% of choosers and 63.0% of nonchoosers were correctly classified. Decision time and lineup fairness were the best postdictors of accuracy. The implications for postdicting real eyewitness decisions are discussed.

Crime↗

PubMed2000

Estimating quantitative genetic parameters in haplodiploid organisms.

Haplodiploid organisms (diploid females and haploid males) are genetically distinct in that they have asymmetrical genetic segregation, gene dosage compensation in the haploid male, maternal effects, and a general propensity to inbreed. Taking these characteristics into account, we demonstrate how conventional experimental designs and statistics derived to measure quantitative genetic parameters for diploid organisms can be applied to haplodiploid ones. First, the C matrix of the joint-scaling test is modified to test the additive-dominance model and then the calculations for fractions in the C matrices (for males and females) are shown for an infinite number of generations with both random and brother-sister mating. Second, analytical adjustments are outlined for the derivation of covariance between relatives using populations at either Hardy-Weinberg equilibrium or those that prefer to inbreed. Finally, four of the conventional designs (sib-analysis, offspring-parental regression, North Carolina III (NC III), and diallel cross) are modified and then compared according to their ability to deal with the experimental conditions encountered in haplodiploid systems. Although all designs can be used with caution, our analysis suggests that the NC III design is the most broadly applicable because it consistently meets the theoretical assumptions. The parameters we derive here for inbred populations are recommended for analysing the typically skewed genotypic distributions found in many natural populations of haplodiploid organisms. The theoretical and applied aspects of our work as well as the difference between our work and that for X-linked genes are discussed.

Animals↗

PubMed2000

Monosaccharide/proton symporter AtSTP1 plays a major role in uptake and response of Arabidopsis seeds and seedlings to sugars.

The aim of this study was to investigate the in vivo properties and function of the high-affinity monosaccharide/proton symporter AtSTP1 of Arabidopsis. We isolated an Atstp1 knock-out mutant and found that this plant grows and develops normally. The AtSTP1 gene is expressed in germinating seeds and seedlings, with AtSTP1 activity found mainly in the seedling root. The rate of uptake of [(14)C]-3-O-methylglucose and [(14)C]-D-glucose is 60% less in Atstp1 seedlings than in the wild type, showing that AtSTP1 is the major monosaccharide transporter in Arabidopsis seedlings. Transport of D-galactose and D-mannose is also up to 60% less in Atstp1 seedlings compared to wild type, but transport of D-fructose, L-arabinose and sucrose is not reduced. Germination of Atstp1 seed shows reduced sensitivity to D-mannose, demonstrating that AtSTP1 is active before germination. Atstp1 seedlings grow effectively on concentrations of D-galactose that inhibit wild-type growth, even at up to 100 mM D-galactose, indicating that active transport by AtSTP1 plays a major role at very high concentrations of exogenous sugar. These findings provide insight into the physiological function of AtSTP1 and clearly establish its importance in the uptake of extracellular sugars by the embryo and in seedlings.

Arabidopsis↗

PubMed2000

Role of CD8 T cells in mycobacterial infections.

The interaction between IFN-gamma-secreting CD4+ T cells and macrophages has long been established as integral in the protective immune response against tuberculosis. More recently, evidence from murine experiments and human studies has begun to demonstrate an essential role for MHC class I restricted CD8+ T cells in this immune response. CD8+ T cells can produce the protective cytokines IFN-gamma and TNF-alpha in addition to their classical cytolytic functions. However, the exact protective mechanisms and antigens recognized by these important cells remain poorly characterized.

Animals↗

PubMed2000

Characterisation of human adenylyl cyclase IX reveals inhibition by Ca(2+)/Calcineurin and differential mRNA plyadenylation.

The functional diversity of adenylyl cyclases provides for different modes of cyclic AMP signalling in mammals. This study reports the cloning and functional characterisation of a cDNA encoding human adenylyl cyclase IX (ACIX). The data show that human ACIX is a Ca(2+)/calcineurin-inhibited adenylyl cyclase prominently expressed in vital organs, including brain, heart, and pancreas. ACIX mRNA was detected in several brain regions, including neocortex, hippocampus, striatum, and cerebellum. By in situ hybridisation, ACIX mRNA was localised to pyramidal and granule cells of the hippocampus, indicating that it is expressed predominantly in nerve cells. Further analysis of ACIX mRNA expression revealed two major forms of ACIX mRNA that arose through tissue-specific differential mRNA polyadenylation. Taken together, the data show that (a) human ACIX is under inhibitory control by Ca(2+) through calcineurin, (b) ACIX may be involved in higher brain functions, and (c) post-transcriptional regulation of ACIX gene expression is a species-specific control mechanism that may enhance the versatility of cyclic AMP signalling in humans.

3' Untranslated Regions↗

PubMed2000

Monocyte-derived macrophage cytokine responses induced by M. bovis BCG.

SETTING: One important aspect of macrophage function is the production of inflammatory and anti-inflammatory cytokines, which in turn affect the survival of intracellular organisms such as mycobacteria. OBJECTIVE: To determine the relationship between phagocytosis of mycobacteria and expression of intracellular cytokines. DESIGN: Phagocytosis and cytokine production were studied simultaneously within human monocyte-derived macrophages (MDMs) from healthy donors using fluorescent labelling of M. bovis BCG and flow cytometry. RESULTS: At a range of infection ratios (5:1, 1:1, 0.2:1) TNF- alpha, IL-10, IL-6 and IL-12 were all produced in a dose-dependent manner. At an infection ratio representative of the in vivo situation (1:1), cytokine production was induced in both MDMs containing intracellular M. bovis BCG and in uninfected bystander MDMs. Phagocytosis increased over time, but there was considerable donor variation: the proportions of cells containing one or more mycobacterium were 15.4+/-14.8% (mean+/-SD) at 4 h and 32.7+/-21.1% at 24 h (n=19). Analysis of cytokine production by MDMs not containing mycobacteria (bystander cells) at 4 h revealed that these uninfected cells produced 79+/-6.6% of the TNF- alpha, 53.9+/-40.0% of the IL-10 and 64.2+/-12.4% of the IL-12. By 20 h these proportions had decreased to 57+/-13.5%, 30.9+/-7.4% and 45.5+/-13.3% respectively. CONCLUSION: Both infected and bystander MDMs can be stimulated to produce cytokines in response to M. bovis BCG, indicating that the ability of MDMs to produce cytokines is not necessarily dependent on the ability to phagocytose mycobacteria.

Adult↗

PubMed2000

Human CD8(+) T cells specific for Mycobacterium tuberculosis secreted antigens in tuberculosis patients and healthy BCG-vaccinated controls in The Gambia.

Intracellular flow cytometry analysis of perforin production by CD8(+) T cells showed levels were greatly reduced in tuberculosis (TB) patients compared to healthy controls. Reduced cytotoxic-T-lymphocyte activity was also obtained with CD8(+) T cells from TB patients compared to healthy controls in The Gambia. A change in antigen recognition was noted between the two groups of donors: in addition to recognition of Ag85A and Ag85B, as seen in healthy donors, a prominent ESAT-6 response was found in TB patients.

Adult↗

PubMed2000

Hand/wrist disorders among sign language communicators.

The study assessed the frequency of self-reported hand/wrist problems among sign language communicators, including interpreters, deaf or hard-of-hearing individuals, and educators. Comparisons were also made between sign language communicators and the general population regarding the frequency of medically diagnosed carpal tunnel syndrome. A survey was mailed to attendees of a national conference that focused on the quality of postsecondary educational opportunities for students with hearing deficits. Fifty-nine percent of 184 respondents reported experiencing hand/wrist problems. Twenty-six percent reported experiencing hand/wrist problems severe enough to limit the ability to work, and 18% reported a medical diagnosis of wrist tendinitis, carpal tunnel syndrome, or both. The frequency of self-reported, medically diagnosed carpal tunnel syndrome for the surveyed sign language communicators was five times the rate experienced by working Americans between ages 18 and 60 years (Tanaka, Wild, Seigman, Halperin, Behrens, & Putz-Anderson, 1995). The study supports increased emphasis on and availability of hand/wrist disorder risk reduction programs for sign language users.

Adolescent↗

PubMed2000

Category structure and created memories.

Cued recall of categorized lists was used to examine effects of category structure on the creation of false memories. In three experiments, category members that had not been presented on studied categorized lists were nonetheless recalled by participants. Delaying the category cued recall test (Experiment 1) and priming category members that had been omitted from target lists (Experiment 3) increased the frequency of false recall. All three experiments showed that nonpresented category members that were higher in output dominance were more frequent intrusions. The typicality of category members, however, did not uniquely contribute to the predictability of false recall (or accurate recall) once the contribution of output dominance was taken into account, suggesting that item accessibility (related to output dominance) may be more instrumental than item distinctiveness (related to typicality) in causing certain types of false recall. The results show that created memories in category cued recall are strongly biased by prior category knowledge and can be predicted by the graded structure of categories, particularly in terms of the output dominance of category instances.

Adult↗

PubMed2000

The phenomenology of real and illusory tip-of-the-tongue states.

The tip-of-the-tongue state (TOT) is the phenomenological experience that a word is on the verge of being recalled. Most research has been directed at TOT etiology and at retrieval processes occurring during a TOT. In this study, TOT phenomenology was examined. In Experiment 1, strong TOTs were more likely than weak TOTs to be followed by correct recognition, and resolution (later recall) of TOTs was higher for strong than for weak TOTs, but only for commission errors. In Experiment 2, emotional TOTs were more likely to be resolved and recognized than nonemotional TOTs. In Experiment 3, imminence was defined as the feeling that retrieval is about to occur. Imminent TOTs were more likely to be followed by resolution and recognition than were nonimminent TOTs. Illusory TOTs (TOTs for unanswerable questions) tended to be weaker, less emotional, and less imminent than TOTs for answerable questions.

Adult↗

PubMed2000

Genetic and developmental modulation of cardiac deficits in prenatal alcohol exposure.

BACKGROUND: Increasing evidence demonstrates that genetic background is an important modulator of alcohol's effects on the developing fetus. Such effects are separable from maternal ethanol metabolism. Here, we study ethanol's effects on cardiogenesis in an avian model that shows strong cell death within neuronal and neural crest precursors following ethanol exposure. METHODS: The study design tested the hypothesis that ethanol-induced losses of cardiac neural crest populations would disrupt outflow tract development and thus contribute to the valvuloseptal deficits observed in prenatal alcohol exposure. Three chick strains were exposed to alcohol at gestational windows between gastrulation and early heart septation (day 3 incubation), and then hearts were examined at the completion of morphogenesis (day 10 incubation). RESULTS: Ethanol's impact on cardiac development was influenced by fetal genetics. The B300 x Hampshire Red cross exhibited pronounced cell death within cardiac neural crest populations but had normal development of the heart and aortic arches. Neural crest migration and differentiation into the distal outflow tract were also normal in these embryos, which suggested a capacity to repair earlier losses. The DeKalb White x Hampshire Red cross also did not exhibit cardiac defects. Hearts of the B300 strain had a unique phenotype with respect to ethanol exposure and exhibited a thin ventricular compact layer, dilatation, and reduced myosin/deoxyribonucleic acid and myosin/protein content, a phenotype that indicates disrupted myocardial maturation and inductive cues. The deficit was only observed when ethanol exposure occurred at stages 15 or 18 and apparently was independent of neural crest cell death. Such ventricular thinning might go undetected in the absence of extensive screening. CONCLUSIONS: Results add to the increasing evidence that genetic background strongly modulates the effects of prenatal alcohol exposure. The results also suggest that embryos have a varying capacity to repair and recover from earlier neural crest losses.

Animals↗

PubMed2000

Avian genetic background modulates the neural crest apoptosis induced by ethanol exposure.

BACKGROUND: Ethanol-induced neural crest apoptosis likely contributes to the distinctive craniofacial phenotype that results from prenatal alcohol exposure. The mechanism responsible for this apoptosis is incompletely understood. A serendipitous change in poultry production flocks led to the discovery that, in chick, the embryo's genetic background modulates its susceptibility to ethanol-induced apoptosis. METHODS: We examined the level of ethanol-induced neural crest apoptosis in 11 chick layer strains or crosses, using acridine orange uptake. RESULTS: Holding the ethanol dose and exposure stage constant, strains were classified into very sensitive (Babcock ISA, HyLine W98, Babcock B300/Hampshire Red cross [BxHR]), moderately sensitive (Spafas, HyLine W36, Babcock B300), and nonresponsive (DeKalb White and Black, Shaver White and 2000, DcKalb White/Hampshire Red cross). Detailed examination of two susceptible strains (W98, BxHR) and a resistant strain (DcKalb White) revealed that the DeKalb's nonresponse was not caused by a shift in timing of apoptosis, or to a lower alcohol exposure at either time of injection or time of death. Strains had identical stage distributions at the time of injection and at apoptosis; housing and diet were held constant. CONCLUSIONS: Factors within the embryo and/or egg environment can affect the susceptibility to ethanol-induced apoptosis. These sensitive and resistant strains will be important tools to dissect the molecular mechanism of ethanol-induced apoptosis, and for understanding how these losses affect subsequent development.

Animals↗