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BackgroundTuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) in HIV-TB co-infected patients receiving antiretroviral therapy (ART) has been linked to neutrophil activation. Anti-neutrophil cytoplasmic antibodies (ANCAs) are also associated with neutrophil activation. Since ANCAs are reportedly skewed in TB and HIV infections, we investigated plasma levels of 7 ANCAs in TB-IRIS patients.MethodsWe retrospectively compared 17 HIV-TB patients who developed TB-IRIS with controls of similar CD4 count, age and gender who did not (HIV+TB+ n = 17), HIV-infected patients without TB (HIV+TB-, n = 17) and 10 HIV-negative (HIV-TB-) controls. Frozen plasma was collected before ART, at 3 and 9 months of ART, and examined by ELISA for levels of 7 ANCAs directed against; Proteinase 3 (PR3), Myeloperoxidase (MPO), Permeability-increasing protein (BPI), Elastase, Cathepsin, Lysozyme, and Lactoferrin.ResultsCompared to HIV+TB+ controls, pre-ART anti-elastase levels were lower in TB-IRIS patients (p = 0.026) and HIV-TB- controls (p = 0.044), whereas other ANCAs did not show significant differences between groups at any time point. A significant decrease over time could be observed in TB-IRIS patients during ART for anti -PR3 (p = 0.027), -lysozyme (p = 0.011), and -lactoferrin (p = 0.019). Conversely, HIV+TB+ controls showed a significant decrease over time for anti -MPO (p = 0.002), -lyzosyme (p = 0.002) and -elastase (p < 0.001).ConclusionThe lack of elevated anti-elastase levels in TB-IRIS patients as opposed to HIV+TB+ controls correspond to previous findings of lowered immune capacity in patients that will develop TB-IRIS. This may suggest a specific role for anti-elastase, elastase or even matrix-metalloproteinases in TB-IRIS. The precise dynamics of neutrophil activation in HIV-TB merits further investigation and could provide more insight in the early mechanisms leading up to TB-IRIS.

BackgroundTuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) is an inflammatory complication in HIV-TB co-infected patients receiving antiretroviral therapy (ART). The exact contribution of T cells, natural killer (NK) cells, and monocytes to TB-IRIS development remains unclear. Here, we studied the expression of exhaustion markers on lymphocytes at different intervals during ART.MethodsWe compared 13 HIV-TB patients who developed TB-IRIS with 13 patients who did not (HIV+TB+), 13 HIV-patients without TB (HIV+TB-) and 9 HIV/TB-negative controls (HIV-TB-). Patients did not differ in age, gender, or CD4-count prior to ART. Frozen peripheral blood mononuclear cells, collected before ART and during 3 months and 9 months of ART, were analysed using flow cytometry. We examined expression of KLRG1, PD-1 and IL-27R on CD4+ and CD8hi T cells, as well as CD3-negative CD8lo lymphocytes as an approximate subset of NK cells. In addition, expression of TLR2, TLR4, IL1RL1, and TRAILR on CD14+ monocytes were investigated.ResultsPrior to ART, TB-IRIS patients had higher percentages of CD8hi T cells that are KLRG1+PD-1+ compared to each control group (p≤0.034). Though PD-1 expression decreased during ART in all groups (p≤0.026), the percentage KLRG1+PD-1+CD8hi T cells remained higher in TB-IRIS patients after 3 months of ART (p≤0.013). Though these patterns were less pronounced in CD3-CD8lo lymphocytes, the percentage of KLRG1+ cells was higher in TB-IRIS patients prior to ART (p≤0.043). In contrast, no clear differences could be observed for CD4+ T cells or monocytes.ConclusionTB-IRIS is preceded by a high level of exhausted (KLRG1+PD-1+) CD8hi T cells, which persists during 3 months of ART. This trait is potentially mirrored in a subpopulation of NK cells, but not CD4+ T cells. Since a dysfunctional CD8+ lymphocyte compartment could predispose patients to TB-IRIS, the functional role of these cells prior to TB-IRIS development should be further explored.

Tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) is an inflammatory complication in HIV-TB co-infected patients receiving antiretroviral therapy (ART). The role of disturbed T cell reconstitution in TB-IRIS is not well understood. We investigated T cell activation and maturation profiles in patients who developed TB-IRIS at different intervals during ART.Twenty-two HIV-TB patients who developed early-onset TB-IRIS and 10 who developed late-onset TB-IRIS were matched for age, sex and CD4 count to equal numbers of HIV-TB patients who did not develop TB-IRIS. Flow cytometry analysis was performed on fresh blood, drawn before and after ART initiation and during TB-IRIS events. T cell activation and maturation was measured on CD4+ and CD8+ T cells using CD45RO, CD38, HLA-DR, CCR7 and CD27 antibodies.CD8+ T cell activation before ART was decreased in both early-onset (77% vs. 82%, p = 0.014) and late-onset (71% vs. 83%, p = 0.012) TB-IRIS patients compared to non-IRIS controls. After ART initiation, the observed differences in T cell activation disappeared. During late-onset, but not early-onset TB-IRIS, we observed a skewing from memory to terminal effector CD4+ and CD8+ T cell populations (p≤0.028).Our data provide evidence of reduced CD8+ T cell activation before ART as a common predisposing factor of early- and late-onset TB-IRIS. The occurrence of TB-IRIS itself was not marked by an over-activated CD8+ T cell compartment. Late- but not early-onset TB-IRIS was characterized by a more terminally differentiated T cell phenotype.

BACKGROUND:Tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) remains a poorly understood complication in HIV-TB patients receiving antiretroviral therapy (ART). TB-IRIS could be associated with an exaggerated immune response to TB-antigens. We compared the recovery of IFNγ responses to recall and TB-antigens and explored in vitro innate cytokine production in TB-IRIS patients. METHODS:In a prospective cohort study of HIV-TB co-infected patients treated for TB before ART initiation, we compared 18 patients who developed TB-IRIS with 18 non-IRIS controls matched for age, sex and CD4 count. We analyzed IFNγ ELISpot responses to CMV, influenza, TB and LPS before ART and during TB-IRIS. CMV and LPS stimulated ELISpot supernatants were subsequently evaluated for production of IL-12p70, IL-6, TNFα and IL-10 by Luminex. RESULTS:Before ART, all responses were similar between TB-IRIS patients and non-IRIS controls. During TB-IRIS, IFNγ responses to TB and influenza antigens were comparable between TB-IRIS patients and non-IRIS controls, but responses to CMV and LPS remained significantly lower in TB-IRIS patients. Production of innate cytokines was similar between TB-IRIS patients and non-IRIS controls. However, upon LPS stimulation, IL-6/IL-10 and TNFα/IL-10 ratios were increased in TB-IRIS patients compared to non-IRIS controls. CONCLUSION:TB-IRIS patients did not display excessive IFNγ responses to TB-antigens. In contrast, the reconstitution of CMV and LPS responses was delayed in the TB-IRIS group. For LPS, this was linked with a pro-inflammatory shift in the innate cytokine balance. These data are in support of a prominent role of the innate immune system in TB-IRIS.

In this study, we retrospectively analysed a total of 605 clinical isolates from six West or Central African countries (Benin, Cameroon, Central African Republic, Guinea-Conakry, Niger and Senegal). Besides spoligotyping to assign isolates to ancient and modern mycobacterial lineages, we conducted phenotypic drug-susceptibility-testing for each isolate for the four first-line drugs. We showed that phylogenetically modern Mycobacterium tuberculosis strains are more likely associated with drug resistance than ancient strains and predict that the currently ongoing replacement of the endemic ancient by a modern mycobacterial population in West/Central Africa might result in increased drug resistance in the sub-region.

BACKGROUND: Tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) remains a poorly understood complication in HIV-TB co-infected patients initiating antiretroviral therapy (ART). The role of the innate immune system in TB-IRIS is becoming increasingly apparent, however the potential involvement in TB-IRIS of a leaky gut and proteins that interfere with TLR stimulation by binding PAMPs has not been investigated before. Here we aimed to investigate the innate nature of the cytokine response in TB-IRIS and to identify novel potential biomarkers. METHODS: From a large prospective cohort of HIV-TB co-infected patients receiving TB treatment, we compared 40 patients who developed TB-IRIS during the first month of ART with 40 patients matched for age, sex and baseline CD4 count who did not. We analyzed plasma levels of lipopolysaccharide (LPS)-binding protein (LBP), LPS, sCD14, endotoxin-core antibody, intestinal fatty acid-binding protein (I-FABP) and 18 pro-and anti-inflammatory cytokines before and during ART. RESULTS: We observed lower baseline levels of IL-6 (p = 0.041), GCSF (p = 0.036) and LBP (p = 0.016) in TB-IRIS patients. At IRIS event, we detected higher levels of LBP, IL-1RA, IL-4, IL-6, IL-7, IL-8, G-CSF (p ≤ 0.032) and lower I-FABP levels (p = 0.013) compared to HIV-TB co-infected controls. Only IL-6 showed an independent effect in multivariate models containing significant cytokines from pre-ART (p = 0.039) and during TB-IRIS (p = 0.034). CONCLUSION: We report pre-ART IL-6 and LBP levels as well as IL-6, LBP and I-FABP levels during IRIS-event as potential biomarkers in TB-IRIS. Our results show no evidence of the possible contribution of a leaky gut to TB-IRIS and indicate that IL-6 holds a distinct role in the disturbed innate cytokine profile before and during TB-IRIS. Future clinical studies should investigate the importance and clinical relevance of these markers for the diagnosis and treatment of TB-IRIS.

BACKGROUND: High concentrations of pro-inflammatory cytokines have been previously observed in the genital fluids of women enrolled in microbicide trials and may explain observed increased HIV transmission in some of these trials. Although the longitudinal nature of these studies allows within-subject comparisons of post-product levels to baseline levels, the fact that the physiologic variations of these cytokines and other markers of immune activation are not fully defined in different populations, makes it difficult to assess changes that can be directly attributed to microbicide use as opposed to other biological and behavioural factors. METHODS: Cervicovaginal lavage samples were collected from 30 healthy Caucasian and assayed for concentrations of ten cytokines/chemokines, total protein content and two antimicrobial proteins using a multiplex immunoassay and ELISA. Cellular markers were characterized by flow cytometry on mononuclear cells collected from the endocervix using flocked swabs. Bacterial quantification was performed using quantitative PCR. RESULTS: Ectopy, menstrual cycle phase, prostate-specific antigen and presence of leucocytes in endocervical cells' supernatant were associated with the concentrations of cyto-/chemokines in cervicovaginal secretions. Approximately 3% of endocervical cells collected were monocytes of which a median of 52% (SD = 17) expressed both CD4 and CCR5 markers. Approximately 1% of the total cells were T-cells with a median of 61% (SD = 10) CD4 and CCR5 expression. Around 5% of the monocytes and 16% of the T-cells expressed the immune activation marker HLA-DR. Higher percentages of T-cells were associated with greater quantities of IL-1RA, GM-CSF and elafin. CONCLUSION: We demonstrate the presence of selected soluble and cellular immune activation markers and identify their predictors in the female genital tract of healthy women. Future clinical trials should consider ectopy, sexual activity, menstrual cycle phase and presence of bacterial species as possible confounders when evaluating the possible inflammatory effects of microbicide compounds.