Altered ratio of IFN-γ/IL-10 in patients with drug resistant Mycobacterium tuberculosis and HIV-tuberculosis immune reconstitution inflammatory syndrome

We have described a clinical relationship between HIV-Tuberculosis Immune Reconstitution Inflammatory Syndrome (TB-IRIS) and anti-tubercular drug resistance. Here we studied the immune response of TB-IRIS patients from whom a drug-resistant (n = 11) or drug-susceptible (n = 25) Mycobacterium tuberculosis (MTB) strain was isolated after presenting with TB-IRIS. ELISpot analysis and multiplex cytokine analysis of the supernatant collected from peripheral blood mononuclear cells stimulated overnight with the heat-killed H37Rv MTB laboratory strain was used. Although there was no statistical difference in IFN-gamma ELISpot responses between the two groups, the results point towards higher bacterial load in the drug-resistant patients, possibly due to failed therapy. The ratio between secreted IFN-gamma/IL-10 and IL-2/IL-10 was significantly lower in TB-IRIS patients in whom the cause of TB was a drug-resistant strain compared to those with a fully sensitive strain (p = 0.02). Since host immune responses are dependent on the bacterial load, we hypothesise that the impaired cytokine balance is likely to be caused by the poorly controlled bacterial growth in these patients

The ratio of secreted cytokines measured by luminex analysis in the supernatants of peripheral blood mononuclear cells (PBMC) stimulated with heat killed H37Rv (MOI = 1∶1) for 24 hours.

<p><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0046481#s3" target="_blank">Results</a> from TB-IRIS patients with multi-drug resistant (MDR) TB are shown in black full circles (n = 7), rifampicin mono-resistant (RM) cases are shown as a cross (n = 3), and fully sensitive (FS) TB cases are shown as open circles (n = 10).</p

Individual patient's data on hkH37Rv stimulated release of IFN-γ, IL-2, IL-10, ratios of IFN-gamma/IL-10, IL-2/IL-10 and ELISpot responses to ESAT-6 and hkH37Rv.

<p>Individual patient's data on hkH37Rv stimulated release of IFN-γ, IL-2, IL-10, ratios of IFN-gamma/IL-10, IL-2/IL-10 and ELISpot responses to ESAT-6 and hkH37Rv.</p

Multiplex cytokine analysis (corrected for multiple comparisons).

(1)<p>p value comparing FS unstimulated with FS stimulated.</p>(2)<p>p value comparing MDR unstimulated with MDR stimulated.</p>(3)<p>p value comparing FS stimulated with MDR stimulated.</p><p>N/A: comparison not applicable due to very low values.</p

IFN-gamma ELISpot response of peripheral blood mononuclear cells (PBMC) stimulated with a range of <i>Mycobacterium tuberculosis</i> antigens, expressed as IFN-gamma spot forming cells per million (SFC/10⁶) PBMC.

<p>TB-IRIS patients with multi-drug resistant (MDR) TB are shown in black full circles, rifampicin mono-resistant (RM) cases are shown as a cross, and fully sensitive (FS) TB cases are shown as open circles.</p

Role of the Interleukin 10 Family of Cytokines in Patients With Immune Reconstitution Inflammatory Syndrome Associated With HIV Infection and Tuberculosis

Background.  The interleukin 10 (IL-10) family comprises cytokines structurally related to IL-10 that share signaling receptors that have conserved signaling cascades. The immunopathogenesis of immune reconstitution inflammatory syndrome (IRIS) in patients with human immunodeficiency virus (HIV) infection and tuberculosis remains incompletely understood. We hypothesized that a deficiency of IL-10 and its homologs may contribute to the immunopathology of IRIS in these patients. Methods. We performed a case-control analysis involving patients with HIV infection and tuberculosis who had IRIS at clinical presentation (tuberculosis-IRIS) and similar patients with HIV infection and tuberculosis who did not develop tuberculosis-IRIS (non-IRIS). Peripheral blood mononuclear cells (PBMCs) were cultured in the presence or absence of heat-killed Mycobacterium tuberculosis for 6 and 24 hours. Messenger RNA was analyzed by quantitative reverse transcription polymerase chain reaction analysis. Cytokine concentrations in serum were also determined. Results. Cultures of PBMCs stimulated with M. tuberculosis for 24 hours yielded higher IL-10 and interleukin 22 (IL-22) transcript levels for tuberculosis-IRIS patients, compared with non-IRIS patients. Analysis of corresponding serum samples showed significantly higher concentrations of IL-10 and IL-22 in tuberculosis-IRIS patients, compared with non-IRIS patients. Conclusions. IL-10 and IL-22 were differentially induced in PBMCs from tuberculosis-IRIS patients after in vitro stimulation, and higher concentrations of their corresponding proteins were detected in serum (in vivo). The higher levels of IL-10 observed in this study may represent a compensatory antiinflammatory response during tuberculosis-IRIS. The elevated levels of IL-22 suggest an association between this cytokine and immunopathology during tuberculosis-IRIS