Biomedical subjects

Matthew J Thomas

Publications and source records attributed to Matthew J Thomas.

15 recordsLinked to original sources

PubMed2006

The cellular protein P58IPK regulates influenza virus mRNA translation and replication through a PKR-mediated mechanism.

We previously hypothesized that efficient translation of influenza virus mRNA requires the recruitment of P58(IPK), the cellular inhibitor of PKR, an interferon-induced kinase that targets the eukaryotic translation initiation factor eIF2alpha. P58(IPK) also inhibits PERK, an eIF2alpha kinase that is localized in the endoplasmic reticulum (ER) and induced during ER stress. The ability of P58(IPK) to interact with and inhibit multiple eIF2alpha kinases suggests it is a critical regulator of both cellular and viral mRNA translation. In this study, we sought to definitively define the role of P58(IPK) during viral infection of mammalian cells. Using mouse embryo fibroblasts from P58(IPK-/-) mice, we demonstrated that the absence of P58(IPK) led to an increase in eIF2alpha phosphorylation and decreased influenza virus mRNA translation. The absence of P58(IPK) also resulted in decreased vesicular stomatitis virus replication but enhanced reovirus yields. In cells lacking the P58(IPK) target, PKR, the trends were reversed-eIF2alpha phosphorylation was decreased, and influenza virus mRNA translation was increased. Although P58(IPK) also inhibits PERK, the presence or absence of this kinase had little effect on influenza virus mRNA translation, despite reduced levels of eIF2alpha phosphorylation in cells lacking PERK. Finally, we showed that influenza virus protein synthesis and viral mRNA levels decrease in cells that express a constitutively active, nonphosphorylatable eIF2alpha. Taken together, our results support a model in which P58(IPK) regulates influenza virus mRNA translation and infection through a PKR-mediated mechanism which is independent of PERK.

Animals↗

PubMed2006

Genomic analysis of increased host immune and cell death responses induced by 1918 influenza virus.

The influenza pandemic of 1918-19 was responsible for about 50 million deaths worldwide. Modern histopathological analysis of autopsy samples from human influenza cases from 1918 revealed significant damage to the lungs with acute, focal bronchitis and alveolitis associated with massive pulmonary oedema, haemorrhage and rapid destruction of the respiratory epithelium. The contribution of the host immune response leading to this severe pathology remains largely unknown. Here we show, in a comprehensive analysis of the global host response induced by the 1918 influenza virus, that mice infected with the reconstructed 1918 influenza virus displayed an increased and accelerated activation of host immune response genes associated with severe pulmonary pathology. We found that mice infected with a virus containing all eight genes from the pandemic virus showed marked activation of pro-inflammatory and cell-death pathways by 24 h after infection that remained unabated until death on day 5. This was in contrast with smaller host immune responses as measured at the genomic level, accompanied by less severe disease pathology and delays in death in mice infected with influenza viruses containing only subsets of 1918 genes. The results indicate a cooperative interaction between the 1918 influenza genes and show that study of the virulence of the 1918 influenza virus requires the use of the fully reconstructed virus. With recent concerns about the introduction of highly pathogenic avian influenza viruses into humans and their potential to cause a worldwide pandemic with disastrous health and economic consequences, a comprehensive understanding of the global host response to the 1918 virus is crucial. Moreover, understanding the contribution of host immune responses to virulent influenza virus infections is an important starting point for the identification of prognostic indicators and the development of novel antiviral therapies.

Animals↗

PubMed2006

PR interval: a comparison of electrical and mechanical methods in the fetus.

BACKGROUND: Mechanical surrogates are used to assess fetal cardiac electrical activity. AIMS: To compare electrical PR interval measured using non-invasive fetal electrocardiography (fECG) with mechanical atrioventricular (AV) interval using Doppler. STUDY DESIGN AND SUBJECTS: Prospective study of 55 recordings made in 50 human fetuses. Those with structural heart defects, second degree or complete heart block were excluded. OUTCOME MEASURES: Mechanical AV interval was measured from the onset of mitral A wave to onset of aortic ejection. Electrical PR interval was measured from a coherent averaged signal obtained using non-invasive fECG recorded from the maternal abdomen. Wilcoxon signed rank test was used to compare both methods. Agreement between AV and PR intervals was assessed using linear regression and by Bland-Altman plots. Bland-Altman analysis assessed inter-observer and intra-observer variability. RESULTS: There was no significant difference in the heart rates of the 55 paired traces measured consecutively using both methods (p<0.35). AV interval was longer than PR (median [range] 116 [96-169] vs. 102 [75-143] ms; p<0.001), with mean difference -16.47 ms (95% Confidence Interval -43.43, 10.44), reflecting the increased proportion of the cardiac cycle measured. Using fECG, PR inter-observer and intra-observer mean differences were 0.4 ms (CI -7.29, 8.09) and 0.7 ms (CI -3.22, 4.62) respectively. R values for inter and intra-observer studies were 0.95 and 0.99 respectively. Using Doppler methods, AV inter-observer and intra-observer mean differences were -2.69 ms, (CI -15.33, 9.95) and 0.92 ms, (CI -9.41, 11.26) respectively. R values for AV measurements were 0.93 for inter-observer and 0.96 for intra-observer variation. CONCLUSIONS: Non-invasive fECG is a robust tool to measure the PR interval with narrow limits of agreement.

Adult↗

PubMed2006

Identification of a specific gene expression pattern associated with HCV-induced pathogenesis in HCV- and HCV/HIV-infected individuals.

Gene expression profiling was performed on liver biopsies from 28 patients (12 HCV and 16 HCV/HIV infected) in an attempt to understand the mechanisms of HCV liver disease in the presence and absence of HIV coinfection. The data were compared with clinical observations and a gene expression database obtained for transplant HCV-infected samples. This is the first report of functional genomics being used to compare intrahepatic gene expression profiles of HCV- and HCV/HIV-infected individuals. Significantly, the intrahepatic global gene expression profiles do not differ between HCV- and HCV/HIV-infected individuals. However, a subset of patients was identified who share a specific pattern of gene expression, termed the enhanced gene expression (EGE) pattern. Specifically, the EGE (+) patients show a dramatic decreased expression of multiple genes associated with the FAS-apoptosis pathway and increased expression of lymphocyte adhesion molecules and lymphocyte-specific genes. The EGE (+) patients also have partially impaired Type I and II IFN-mediated antiviral responses, including a lack of induction of the anti-fibrogenic cytokine IFN-gamma. Importantly, the pattern of gene expression observed in EGE (+) patients has similarities to patients who developed fibrosis within 1 year of receiving a liver transplant.

Female↗

PubMed2006

Functional gene analysis of individual response to challenge of SIVmac239 in M. mulatta PBMC culture.

It has previously been shown in macaques that individual animals exhibit varying responses to challenge with the same strain of SIV. We attempted to elucidate these differences using functional genomics and correlate them to biological response. Unfractionated PBMC from three rhesus macaques were isolated, activated, and infected with SIVmac239. Interestingly, one of the three animals used for these experiments exhibited a completely unique response to infection relative to the other two. After repeated attempts to infect the PBMC from this animal, little or no infectivity was seen across the time points considered, and corresponding to this apparent lack of infection, few genes were seen to be differentially expressed when compared to mock-infected cells. For the remaining two animals, gene expression analysis showed that while they exhibited responses for the same groups of pathways, these responses included differences specific to the individual animal at the gene level. In instances where the patterns of differential gene expression differed between these animals, the genes being differentially expressed were associated with the same categories of biological process, mainly immune response and cell signaling. At the pathway level, these animals again exhibited similar responses that could be predicted based on the experimental conditions. Even in these expected results, the degree of response and the specific genes being regulated differed greatly from animal to animal. The differences in gene expression on an individual level have the potential to be used as markers in identification of animals suitable for lentiviral infection experiments. Our results highlight the importance of individual variation in response to viral challenge.

Animals↗

PubMed2006

Suppression of proinflammatory signal transduction and gene expression by the dual nucleic acid binding domains of the vaccinia virus E3L proteins.

Cells have evolved elaborate mechanisms to counteract the onslaught of viral infections. To activate these defenses, the viral threat must be recognized. Danger signals, or pathogen-associated molecular patterns, that are induced by pathogens include double-stranded RNA (dsRNA), viral single-stranded RNA, glycolipids, and CpG DNA. Understanding the signal transduction pathways activated and host gene expression induced by these danger signals is vital to understanding virus-host interactions. The vaccinia virus E3L protein is involved in blocking the host antiviral response and increasing pathogenesis, functions that map to separate C-terminal dsRNA- and N-terminal Z-DNA-binding domains. Viruses containing mutations in these domains allow modeling of the role of dsRNA and Z-form nucleic acid in the host response to virus infection. Deletions in the Z-DNA- or dsRNA-binding domains led to activation of signal transduction cascades and up-regulation of host gene expression, with many genes involved in the inflammatory response. These data suggest that poxviruses actively inhibit cellular recognition of viral danger signals and the subsequent cellular response to the viral threat.

DNA-Binding Proteins↗

PubMed2006

Global suppression of the host antiviral response by Ebola- and Marburgviruses: increased antagonism of the type I interferon response is associated with enhanced virulence.

We studied the effect of filovirus infection on host cell gene expression by characterizing the regulation of gene expression responses in human liver cells infected with Zaire Ebolavirus (ZEBOV), Reston Ebolavirus (REBOV), and Marburgvirus (MARV), using transcriptional profiling and bioinformatics. Expression microarray analysis demonstrated that filovirus infection resulted in the up-regulation of immune-related genes and the down-regulation of many coagulation and acute-phase proteins. These studies further revealed that a common feature of filovirus virulence is suppression of key cellular antiviral responses, including TLR-, interferon (IFN) regulatory factor 3-, and PKR-related pathways. We further showed that ZEBOV and MARV were more potent antagonists of the IFN response and inhibited the expression of most of the IFN-stimulated genes (ISGs) observed in mock-infected IFN-alpha-2b treated cells, compared to REBOV infection, which activated more than 20% of these ISGs. Finally, we examined IFN-related gene expression in filovirus-infected cells treated with IFN-alpha-2b. These experiments revealed that a majority of genes induced in mock-infected cells treated with type I IFN were antagonized in treated ZEBOV- and MARV-infected cells, while in contrast, REBOV infection resulted in a significant increase in ISG expression. Analysis of STAT1 and -2 phosphorylation following IFN treatment showed a significant reduction of STAT phosphorylation for MARV but not for ZEBOV and REBOV, indicating that different mechanisms might be involved in antagonizing IFN signaling pathways by the different filovirus species. Taken together, these studies showed a correlation between antagonism of type I IFN responses and filovirus virulence.

Animals↗

PubMed2006

Use of human nasal cannulas during bronchoscopy procedures as a simple method for maintaining adequate oxygen saturation in pigtailed macaques (Macaca nemestrina).

Rising concerns over respiratory illnesses caused by agents such as avian influenza viruses and SARS coronavirus have prompted intensive research efforts and the resurgence of nonhuman primates as models for these human diseases. In the context of studying influenza infection and vaccine development, serial bronchoscopic procedures, including bronchial brush biopsies and bronchoalveolar lavage, were performed in pigtailed macaques (Macaca nemestrina). The possible need for oxygen supplementation during these procedures was anticipated because of the size of the animals relative to the 5-mm bronchoscope. We therefore monitored oxyhemoglobin saturation, a measure of arterial blood oxygen content, before and after insertion of the bronchoscope, during bronchoalveolar lavage, and after initiation of oxygen supplementation. Although more data are required to draw definitive conclusions, our findings suggested the need for oxygen supplementation during such procedures in nonhuman primates, despite the fact that human patients undergoing bronchoscopy and lavage do not routinely get oxygen unless they are already compromised. Our data also suggested that the need for supplementation could not be predicted from simple parameters such as size of the animal, presence of respiratory clinical signs, or experimental treatment. Finally, we show a simple and cost-effective method of using human nasal cannulas for delivering oxygen to pigtailed macaques during bronchoscopic procedures, and we believe that, after further testing, this method could be used safely and effectively in other nonhuman primate species.

Animals↗

PubMed2005

Analysis of the Macaca mulatta transcriptome and the sequence divergence between Macaca and human.

We report the initial sequencing and comparative analysis of the Macaca mulatta transcriptome. Cloned sequences from 11 tissues, nine animals, and three species (M. mulatta, M. fascicularis, and M. nemestrina) were sampled, resulting in the generation of 48,642 sequence reads. These data represent an initial sampling of the putative rhesus orthologs for 6,216 human genes. Mean nucleotide diversity within M. mulatta and sequence divergence among M. fascicularis, M. nemestrina, and M. mulatta are also reported.

Animals↗

PubMed2005

Airway inflammation: chemokine-induced neutrophilia and the class I phosphoinositide 3-kinases.

Class I phosphoinositide 3-kinases (PI3K) are known to play a significant role in neutrophil chemotaxis. However, the relative contributions of different PI3K isoforms, and how these impact on lung inflammation, have not been addressed. In vitro studies using wild-type and PI3Kgamma knockout neutrophils demonstrated the major role of the gamma isoform in chemotactic but not chemokinetic events. This was confirmed by a model of direct chemokine instillation into the airways in vivo. Within all studies, a low yet significant degree of neutrophil movement in the absence of PI3Kgamma could be observed. No role for the delta isoform was demonstrated both in vitro and in vivo using PI3Kdelta kinase-dead knock-in mice. Moreover, further studies using the broad-spectrum PI3K inhibitors wortmannin or LY294002 showed no other class I PI3K isoforms to be involved in these chemotactic processes. Here, we identify a contributory PI3K-independent mechanism of neutrophil movement, yet demonstrate PI3Kgamma as the pivotal mediator through which the majority of neutrophils migrate into the lung in response to chemokines. These data resolve the complexities of chemokine-induced neutrophilia and PI3K signaling and define the gamma isoform as a promising target for new therapeutics to treat airway inflammatory diseases.

Animals↗

PubMed2005

The Thr92Ala deiodinase type 2 (DIO2) variant is not associated with type 2 diabetes or indices of insulin resistance in the old order of Amish.

A common polymorphism of the type 2 deiodinase gene (Thr92Ala DIO2) was found to be associated with insulin resistance in a mixed Caucasian population. The aim of this study was to investigate the association of the Thr92Ala DIO2 variant to indices of insulin resistance in the Old Order Amish. A genotype-phenotype association study was performed at the research clinic in Strasburg, Pennsylvania, and the molecular genetics laboratory at the University of Maryland, Baltimore, Maryland, and the National Institutes of Health, Bethesda, Maryland. A total of 1,268 subjects participated in the Amish Family Diabetes Study. An association among the Thr92Ala DIO2 variant and type 2 diabetes, indices of insulin resistance (HOMA-IR), insulin secretion, free thyroid hormones, and thyrotropin (TSH) was found. No association was found among the Thr92Ala DIO2 variant and type 2 diabetes, impaired glucose tolerance, or body mass index (BMI) in the Amish. In nondiabetics (n = 747), the Ala92 allele tended to be associated with decreased rather then increased insulin secretion. No differences were observed in thyroid hormones or TSH. Contrary to prior findings, the Thr92Ala DIO2 variant tends to be associated with increased rather then decreased insulin sensitivity in the Amish. These findings could be secondary to a different genetic background or to environmental factors specific for this population.

Adult↗

PubMed2005

Non-invasive intrapartum fetal ECG: preliminary report.

OBJECTIVES: To obtain fetal heart rate, detailed fetal electrocardiography (fECG) signals and uterine contractions during labour using a single device. DESIGN: Prospective observational study. SETTING: Delivery suite at a tertiary referral hospital, London, UK. POPULATION: Fifteen patients at median gestation of 39 weeks (range 24-41) were recruited at median cervical dilatation of 4.0 cm (range 0-10) of whom 8/15 (53%) had intact amniotic membranes. METHODS: Using 12 abdominally sited electrodes, we recorded the composite abdominal signal in pregnancies intrapartum. The recorded data were analysed off-line using a blind signal separation technique. MAIN OUTCOME MEASURES: Success of signal separation and fECG time intervals. RESULTS: Successful fECG signal acquisition was achieved in 12/15 (80%) patients and an averaged fECG waveform acquired. In these patients, P and QRS waves were seen in all cases, and T waves in 11/12 (92%). True beat-to-beat heart rate (HR) was displayed and measures of its variability obtained. The mother's ECG and uterine electrical activity, shown to match tocographically recorded uterine contractions, were also separated and displayed. Failure to acquire fECG in three cases was attributed to excessive abdominal muscular activity and electrical interference. CONCLUSIONS: This study demonstrates a non-invasive technique that displays detailed intrapartum fECG waveforms, HR variability, maternal ECG and uterine contractions simultaneously, all in a single device and which avoids the potential risks of invasive monitoring with a fetal scalp electrode.

Cardiotocography↗

PubMed2004

The contribution of expectations to motion sickness symptoms and gastric activity.

OBJECTIVE: The goal of this investigation was to determine in healthy adults the effect of expectation manipulations on the development of motion sickness, as indicated by abnormal gastric myoelectric activity and subjective reports of symptoms of motion sickness. METHOD: Eighty participants, moderately susceptible to motion sickness, experienced one of four conditions created from a two-variable (Expectation, Drum), two-factor model (High/Low expectation for sickness; Rotating/Stable Drum). The electrogastrogram (EGG) was recorded 6 min prior to the expectation manipulation; 6 min following the expectation manipulation; 6 min before drum activation; and 16 min during drum activation. Self-report questionnaires indicating expectation for sickness (MSEx) and motion sickness symptoms (Nausea Profile [NP]) were obtained following the expectation manipulation and exposure to the drum, respectively. RESULTS: No significant differences were observed among expectation groups for retrospective reports of motion sickness (NP); however, significant differences in EGG responses to drum rotation were obtained. The unexpected results of a univariate analysis of variance (ANOVA) revealed significantly greater gastric tachyarrhythmia and less normal activity, an indication of motion sickness, in the low expectation for sickness conditions. CONCLUSION: These results suggest that inducing a high expectation for sickness in healthy individuals about to be exposed to provocative motion results in a protective effect from motion sickness following exposure to the stimulus, while low expectations may induce abnormal gastric activity.

Adolescent↗

PubMed2003

A classifier for the SNP-based inference of ancestry.

Ancestral inference from DNA could serve as an important adjunct for both standard and future human identity testing procedures. However, current STR methods for the inference of ancestral affiliation have inherent statistical and technical limitations. In an effort to identify bi-allelic markers that can be used to infer ancestral affiliation from DNA, we screened 211 SNPs in the human pigmentation and xenobiotic metabolism genes. Allele frequencies of 56 SNPs (most from pigmentation genes) were dramatically different between groups of unrelated individuals of Asian, African, and European descent, and both observed and simulated log likelihood ratios revealed that the markers were of exceptional value for ancestral inference. Log likelihood ratios of the multilocus estimates of biological ancestry (EAE/EBA) ranged from 7 to 10, which are on par with the best of the STR batteries yet described. A linear classification method was developed for incorporating these SNPs into a classifier model that was 99, 98, and 100% accurate for identifying individuals of European, African, and Asian descent, respectively. The methods and markers we describe are therefore an important first step for the development of a practical multiplex test for the inference of ancestry in a forensics setting.

Algorithms↗

PubMed2002

CD8 T cells inhibit IgE via dendritic cell IL-12 induction that promotes Th1 T cell counter-regulation.

Th1 and Th2 cells are counterinhibitory; their balance determines allergic sensitization. We show here that CD8 T cell subsets break these rules as both T cytotoxic (Tc)1 and Tc2 cells promote Th1 over Th2 immunity. Using IL-12(-/-), IFN-gamma(-/-), and OVA(257-264)-specific Valpha2Vbeta5 TCR-transgenic mice, we have identified the key steps involved. OVA-specific IFN-gamma(-/-) CD8 T cells inhibited IgE responses equivalent to wild-type CD8 T cells (up to 98% suppression), indicating that CD8 T cell-derived IFN-gamma was not required. However, OVA-specific CD8 T cells could not inhibit IgE in IFN-gamma(-/-) recipients unless reconstituted with naive, wild-type CD4 T cells, suggesting that CD4 T cell-derived IFN-gamma did play a role. Transfer of either Tc1 or Tc2 Valpha2Vbeta5 TCR-transgenic CD8 T cells inhibited IgE and OVA-specific Th2 cells while promoting OVA-specific Th1 cell responses, suggesting a potential role for a type 1 inducing cytokine such as IL-12. CD8 T cells were shown to induce IL-12 in OVA(257-264)-pulsed dendritic cells (DC) in vitro. Furthermore, CD8 T cells were unable to inhibit IgE responses in IL-12(-/-) recipients without the addition of naive, wild-type DC, thus demonstrating a pivotal role for IL-12 in this mechanism. These data reveal a mechanism of IgE regulation in which CD8 T cells induce DC IL-12 by an IFN-gamma-independent process that subsequently induces Th1 and inhibits Th2 cells. Th1 cell IFN-gamma is the final step that inhibits B cell IgE class switching. This demonstrates a novel regulatory network through which CD8 T cells inhibit allergic sensitization.

Adoptive Transfer↗