Biswas C.,University of Sydney |
Zuo X.,University of Sydney |
Chen S.C.-A.,University of Sydney |
Chen S.C.-A.,Westmead Hospital |
And 5 more authors.
Fungal Genetics and Biology | Year: 2014
Miltefosine (MI) is a novel, potential antifungal agent with activity against some yeast and filamentous fungal pathogens. We previously demonstrated in the model yeast, Saccharomyces cerevisiae, that MI causes disruption of mitochondrial membrane potential and apoptosis-like cell death via interaction with the Cox9p sub-unit of cytochrome c oxidase (COX). To identify additional mechanisms of antifungal action, MI resistance was induced in S. cerevisiae by exposure to the mutagen, ethyl methanesulfonate, and gene mutation(s) responsible for resistance were investigated. An MI-resistant haploid strain (H-C101) was created. Resistance was retained in the diploid strain (D-C101) following mating, confirming dominant inheritance. Phenotypic assessment of individual D-C101 tetrads revealed that only one mutant gene contributed to the MI-resistance phenotype. To identify this gene, the genome of H-C101 was sequenced and 17 mutated genes, including metacaspase-encoding MCA1, were identified. The MCA1 mutation resulted in substitution of asparagine (N) with aspartic acid (D) at position 164 (MCA1N164D). MI resistance was found to be primarily due to MCA1N164D, as single-copy episomal expression of MCA1N164D, but not two other mutated genes (FAS1T1417I and BCK2T104A), resulted in MI resistance in the wild-type strain. Furthermore, an MCA1 deletion mutant (mca1δ) was MI-resistant. MI treatment led to accumulation of reactive oxygen species (ROS) in MI-resistant (MCA1N164D-expressing and mca1δ) strains and MI-susceptible (MCA1-expressing) strains, but failed to activate Mca1 in the MI-resistant strains, demonstrating that ROS accumulation does not contribute to the fungicidal effect of MI. In conclusion, functional disruption of Mca1, leads to MI resistance and inability to mediate MI-induced apoptotic effects. Mca1-mediated apoptosis is therefore a major mechanism of MI-induced antifungal action. © 2014 Elsevier Inc.
Rajasuriar R.,Monash University |
Rajasuriar R.,University of Malaya |
Booth D.,Institute for Immunology and Allergy Research |
Solomon A.,Monash University |
And 16 more authors.
Journal of Infectious Diseases | Year: 2010
Background. Multiple host factors may influence CD4+ T cell reconstitution in human immunodeficiency virus (HIV)-infected patients after suppressive antiretroviral therapy (ART). We hypothesized that residual immune activation and polymorphisms in the interleukin 7 (IL-7) receptor α (IL-7Rα) gene were important for immune recovery. Methods. We examined HIV-infected patients receiving suppressive ART (n = 96) for their IL-7Rα haplotypes and measured levels of lipopolysaccharide (LPS), soluble CD14, and IL-7 in plasma samples collected before and after ART initiation. Levels of soluble IL-7Rα were measured in HIV-infected patients with IL-7Rα haplotype 2 (n = 11) and those without IL-7Rα haplotype 2 (n = 22). Multivariate analysis was used to identify variables associated with faster recovery to CD4+ T cell counts of >500 and >200 cells/μL. Results. Both LPS and soluble CD14 levels were significantly decreased with ART (P < .001 respectively) but remained elevated compared with uninfected controls. In a multivariate analysis, faster recovery to a CD4+ T cell count of >500 cells/μL was significantly associated with higher baseline CD4+ T cell count, younger age, lower pre-ART LPS level, higher pre-ART soluble CD14 level, lower pre-ART IL-7 level, and IL-7Rα haplotype 2 (hazard ratio, 1.50; 95% confidence interval, 1.03-2.19; P = .034). HIV-infected patients with haplotype 2 had significantly lower soluble IL-7Rα levels compared with those of patients without haplotype 2 (P < .001). Conclusion. Both the extent of immune depletion prior to ART and IL-7Rα haplotype 2 are important determinants of time to CD4+ T cell recovery to counts of 1500 cells/μL. © 2010 by the Infectious Diseases Society of America. All rights reserved.
Sedger L.M.,University of Technology, Sydney |
Sedger L.M.,Institute for Immunology and Allergy Research |
Katewa A.,University of Technology, Sydney |
Katewa A.,Institute for Immunology and Allergy Research |
And 7 more authors.
Blood | Year: 2010
To delineate the relative roles of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and Fas ligand in lymphocyte biology and lymphoproliferative disease, we generated mice defective in both molecules. B6.GT mice develop severe polyclonal lymphoproliferative disease because of accumulating CD3+CD4- CD8-B220+ T cells, CD4+ and CD8+ T cells, and follicular B cells, and mice die prematurely from extreme lymphocytosis, thrombocytopenia, and hemorrhage. Accumulating lymphocytes resembled antigen-experienced lymphocytes, consistent with the maximal resistance of B6.GT CD4+ and CD8 + T cell to activation-induced cell death. More specifically, we show that TRAIL contributes to Fas ligandmediated activation-induced cell death and controls lymphocyte apoptosis in the presence of interferon-γ once antigen stimulation is removed. Furthermore, dysregulated lymphocyte homeostasis results in the production of anti-DNA and rheumatoid factor autoantibodies, as well as antiplatelet IgM and IgG causing thrombocytopenia. Thus, B6.GT mice reveal new roles for TRAIL in lymphocyte homeostasis and autoimmune lymphoproliferative syndromes and are a model of spontaneous idiopathic thrombocytopenia purpura secondary to lymphoproliferative disease. © 2010 by The American Society of Hematology.
Parnell G.P.,University of Sydney |
McLean A.S.,University of Sydney |
Booth D.R.,Institute for Immunology and Allergy Research |
Armstrong N.J.,Garvan Institute of Medical Research |
And 8 more authors.
Critical Care | Year: 2012
Introduction: Diagnosis of severe influenza pneumonia remains challenging because of a lack of correlation between the presence of influenza virus and clinical status. We conducted gene-expression profiling in the whole blood of critically ill patients to identify a gene signature that would allow clinicians to distinguish influenza infection from other causes of severe respiratory failure, such as bacterial pneumonia, and noninfective systemic inflammatory response syndrome.Methods: Whole-blood samples were collected from critically ill individuals and assayed on Illumina HT-12 gene-expression beadarrays. Differentially expressed genes were determined by linear mixed-model analysis and overrepresented biological pathways determined by using GeneGo MetaCore.Results: The gene-expression profile of H1N1 influenza A pneumonia was distinctly different from those of bacterial pneumonia and systemic inflammatory response syndrome. The influenza gene-expression profile is characterized by upregulation of genes from cell-cycle regulation, apoptosis, and DNA-damage-response pathways. In contrast, no distinctive gene-expression signature was found in patients with bacterial pneumonia or systemic inflammatory response syndrome. The gene-expression profile of influenza infection persisted through 5 days of follow-up. Furthermore, in patients with primary H1N1 influenza A infection in whom bacterial co-infection subsequently developed, the influenza gene-expression signature remained unaltered, despite the presence of a superimposed bacterial infection.Conclusions: The whole-blood expression-profiling data indicate that the host response to influenza pneumonia is distinctly different from that caused by bacterial pathogens. This information may speed the identification of the cause of infection in patients presenting with severe respiratory failure, allowing appropriate patient care to be undertaken more rapidly. © 2012 Parnell et al.; licensee BioMed Central Ltd.
Parnell G.P.,University of Sydney |
Parnell G.P.,Institute for Immunology and Allergy Research |
Tang B.M.,University of Sydney |
Nalos M.,University of Sydney |
And 4 more authors.
Shock | Year: 2013
There is currently no reliable tool available to measure immune dysfunction in septic patients in the clinical setting. This proof-of-concept study assesses the potential of gene expression profiling of whole blood as a tool to monitor immune dysfunction in critically ill septic patients. Whole-blood samples were collected daily for up to 5 days from patients admitted to the intensive care unit with sepsis. RNA isolated from whole-blood samples was assayed on Illumina HT-12 gene expression microarrays consisting of 48,804 probes. Microarray analysis identified 3,677 genes as differentially expressed across 5 days between septic patients and healthy controls. Of the 3,677 genes, biological pathway analysis identified 86 genes significantly downregulated in the sepsis patients were present in pathways relating to immune response. These 86 genes correspond to known immune pathways implicated in sepsis, including lymphocyte depletion, reduced T-lymphocyte activation, and deficient antigen presentation. Furthermore, expression levels of these genes correlated with clinical severity, with a significantly greater degree of downregulation found in nonsurvivors compared with survivors. The results show that whole-blood gene expression analysis can capture systemic immune dysfunctions in septic patients. Our study provides an experimental basis to support further study on the use of a gene expression-based assay, to assess immunosuppression, and to guide immunotherapy in future clinical trials. © 2013 by the Shock Society.
Ramanathan S.,Westmead Hospital |
Ramanathan S.,University of Sydney |
Lenton K.,Institute for Immunology and Allergy Research |
Burke T.,Institute for Immunology and Allergy Research |
And 6 more authors.
Journal of Clinical Neuroscience | Year: 2013
The ability to predict disability development in multiple sclerosis (MS) is limited. While abnormalities of evoked potentials (EP) have been associated with disability, the prognosticating utility of EP in MS remains to be fully elucidated. The present study assessed the utility of multimodal EP as a prognostic biomarker of disability in a cohort of clinically heterogeneous MS patients. Median and tibial nerve somatosensory, visual, and brainstem auditory EP were performed at initial assessment on 63 MS patients (53 relapsing-remitting and 10 secondary progressive) who were followed for an average of 2 years. A combined EP score (CEPS) was calculated consisting of the total number of abnormal EP tests, and was correlated with the Expanded Disability Status Scale (EDSS) at baseline and follow-up. There was a significant correlation between multimodal EP and baseline and follow-up EDSS. Specifically, tibial nerve P37 latencies correlated with EDSS (R BASELINE = 0.49, p < 0.01; RFOLLOW-UP = 0.47, p < 0.01), as did the median nerve N13 (RBASELINE = 0.40, p < 0.01; RFOLLOW-UP = 0.35, p < 0.05) and N20 latencies (R BASELINE = 0.43, p < 0.01; RFOLLOW-UP = 0.47, p < 0.01), and P100 full-field (RBASELINE = 0.50, p < 0.001; R FOLLOW-UP = 0.45, p < 0.001) and central field latencies (R BASELINE = 0.60, p < 0.001; RFOLLOW-UP = 0.50, p < 0.001). In addition, there was a significant correlation between the CEPS with baseline (R = 0.65, p < 0.001) and follow-up (R = 0.57, p < 0.01) EDSS. In contrast, white matter disease burden, as measured by T2 lesion load, exhibited a weaker correlation with EDSS (RBASELINE = 0.28, p < 0.05). In conclusion, these findings suggest that abnormalities of EP, as quantified by the novel CEPS, may be a useful biomarker for prognosticating clinical disability in MS, and may aid in the quantification of MS disease severity and in guiding therapeutic decisions. © 2013 Elsevier Ltd. All rights reserved.
McKay F.C.,Institute for Immunology and Allergy Research |
McKay F.C.,University of Sydney |
Hoe E.,Institute for Immunology and Allergy Research |
Hoe E.,University of Sydney |
And 10 more authors.
PLoS ONE | Year: 2013
The IL7Rα gene is unequivocally associated with susceptibility to multiple sclerosis (MS). Haplotype 2 (Hap 2) confers protection from MS, and T cells and dendritic cells (DCs) of Hap 2 exhibit reduced splicing of exon 6, resulting in production of relatively less soluble receptor, and potentially more response to ligand. We have previously shown in CD4 T cells that IL7Rα haplotypes 1 and 2, but not 4, respond to interferon beta (IFNβ), the most commonly used immunomodulatory drug in MS, and that haplotype 4 (Hap 4) homozygotes have the highest risk of developing MS. We now show that IL7R expression increases in myeloid cells in response to IFNβ, but that the response is haplotype-dependent, with cells from homozygotes for Hap 4 again showing no response. This was shown using freshly derived monocytes, in vitro cultured immature and mature monocyte-derived dendritic cells, and by comparing homozygotes for the common haplotypes, and relative expression of alleles in heterozygotes (Hap 4 vs not Hap 4). As for T cells, in all myeloid cell subsets examined, Hap 2 homozygotes showed a trend for reduced splicing of exon 6 compared to the other haplotypes, significantly so in most conditions. These data are consistent with increased signaling being protective from MS, constitutively and in response to IFNβ. We also demonstrate significant regulation of immune response, chemokine activity and cytokine biosynthesis pathways by IL7Rα signaling in IFNβ -treated myeloid subsets. IFNβ-responsive genes are over-represented amongst genes associated with MS susceptibility. IL7Rα haplotype may contribute to MS susceptibility through reduced capacity for IL7Rα signalling in myeloid cells, especially in the presence of IFNβ, and is currently under investigation as a predictor of therapeutic response. © 2013 McKay et al.