The study aimed to determine whether tuberculosis patients who suffer relapse have different immune responses to mycobacteria in vitro compared to patients who remain cured for two years, as currently no tools to accurately predict... more
The study aimed to determine whether tuberculosis patients who suffer relapse have different immune responses to mycobacteria in vitro compared to patients who remain cured for two years, as currently no tools to accurately predict tuberculosis relapse exist. First episode pulmonary tuberculosis patients were recruited in South Africa. Diluted blood, collected at diagnosis and after two and four weeks of treatment, was cultured with live Mycobacterium tuberculosis for six days and cellular RNA frozen. Gene expression in ten patients who subsequently relapsed, confirmed by strain genotyping, was compared to those who remained cured using microarrays. At diagnosis, expression of 668 genes was significantly different in relapse patients' samples compared to successfully cured patients: these differences persisted for at least four weeks. Gene Ontology and biological pathways analyses revealed significant up-regulation of genes involved in cytotoxic cell-mediated killing. Results...
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Th17 cells play an important role in immunity to fungal and bacterial pathogens, although their role in the female genital tract (FGT), where exposure to these pathogens is common, is not well understood. We investigated the relationship... more
Th17 cells play an important role in immunity to fungal and bacterial pathogens, although their role in the female genital tract (FGT), where exposure to these pathogens is common, is not well understood. We investigated the relationship between FGT infections, cervicovaginal interleukin (IL)-17 concentrations and Th17 cell frequencies. Forty-two cytokines were measured in cervicovaginal lavages (CVLs) from HIV-uninfected and HIV-infected women. Frequencies of Th17 cells (CD3+CD4+IL-17a+) were evaluated in cervical cytobrushes and blood by flow cytometry. Women were screened for Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, Trichomonas vaginalis and HSV-2 by PCR, and candidal infections and bacterial vaginosis by Gram stain. Women with bacterial sexually transmitted infections (STIs), specifically chlamydia and gonorrhoea, had higher genital IL-17 concentrations than women with no STI, while women with candidal pseudohyphae/spores had lower IL-17 concentrations compared to women without candidal infections. Viral STIs (HSV-2 and HIV) were not associated with significant changes in genital IL-17 concentrations. Genital IL-17 concentrations correlated strongly with other inflammatory cytokines and growth factors. Although Th17 cells were depleted from blood during HIV infection, cervical Th17 cell frequencies were similar in HIV-uninfected and infected women. Cervical Th17 cell frequencies were also not associated with STIs or candida, although few women had a STI. These findings suggest that IL-17 production in the FGT is induced in response to bacterial but not viral STIs. Decreased IL-17 associated with candidal infections suggests that candida may actively suppress IL-17 production or women with dampened IL-17 responses may be more susceptible to candidal outgrowth. This article is protected by copyright. All rights reserved.
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The synthetic progestins, medroxyprogesterone acetate (MPA) and norethisterone acetate (NET-EN or NET-A), are widely used as female contraceptive agents and in hormone replacement therapy (HRT). Competitive binding revealed that MPA... more
The synthetic progestins, medroxyprogesterone acetate (MPA) and norethisterone acetate (NET-EN or NET-A), are widely used as female contraceptive agents and in hormone replacement therapy (HRT). Competitive binding revealed that MPA displays a higher relative binding affinity than NET-A and progesterone (prog) for the human GR (Kd of 4.2 nM for dexamethasone (dex) and Ki's of 10.8, 270 and 215 nM for MPA, NET-A and prog, respectively). Furthermore, MPA displays much greater glucocorticoid (GC) transactivation agonist potency than NET-A or prog (EC50s of 1.1, 7.2, >1000 and 280 nM for dex, MPA, NET-A and prog, respectively) and much greater GC agonist potency for transrepression than NET-A or prog (EC50s of 0.21, 2.7, >100 and 26 nM for dex, MPA, NET-A and prog, respectively). In addition, MPA induces phosphorylation of the GR at Ser 211 to a much greater extent than NET-A or prog and protects the GR from partial trypsin digestion in vitro to a much greater extent than NET-A or prog at saturating concentrations. Together these results suggest that the differences in biological activity of the progestins are not merely due to differences in their affinity for the GR but also due to the induction of different conformational changes in the liganded-GR. MPA and NET-A therefore display very different GC-like properties compared to each other and to prog, and are likely to exhibit different side effects via the GR.
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A central question in glucocorticoid mechanism of action via the glucocorticoid receptor (GR) is what determines ligand-selective transcriptional responses. Using a panel of 12 GR ligands, we show that the extent of GR phosphorylation at... more
A central question in glucocorticoid mechanism of action via the glucocorticoid receptor (GR) is what determines ligand-selective transcriptional responses. Using a panel of 12 GR ligands, we show that the extent of GR phosphorylation at S226 and S211, GR half-life and transcriptional response, occur in a ligand-selective manner. While GR phosphorylation at S226 was shown to inhibit maximal transcription efficacy, phosphorylation at S211 is required for maximal transactivation, but not for transrepression efficacy. Both ligand-selective GR phosphorylation and half-life correlated with efficacy for transactivation and transrepression. For both expressed and endogenous GR, in two different cell lines, agonists resulted in the greatest extent of phosphorylation and the greatest extent of GR downregulation, suggesting a link between these functions. However, using phosphorylation-deficient GR mutants we established that phosphorylation of the GR at S226 or S211 does not determine the rank order of ligand-selective GR transactivation. These results are consistent with a model whereby ligand-selective GR phosphorylation and half-life are a consequence of upstream events, such as ligand-specific GR conformations, which are maintained in the phosphorylation mutants.