DHEA modulates the expression of the FoxP3 transcription factor in coinfected individuals. A and B.

<p>Percentages of Treg cells (defined as CD4+FoxP3+CD25+) in PBMC from <b>A.</b> HIV-TB and <b>B.</b> HD individuals stimulated with <i>M. tuberculosis</i> antigen in the presence or absence of DHEA at the indicated concentrations for 3 days. Bars indicate the mean ± SEM for each experimental condition. <b>C and D.</b> Relative FoxP3 Median florescence intensity (MFI) in Treg lymphocytes from <b>C.</b> HIV-TB patients and <b>D.</b> HD volunteers after culturing PBMC as detailed in A. Bars indicate the mean ± SEM for each treatment. Data are representative of four different experiments. *: <i>p</i> < 0.05; **: <i>p</i> < 0.01. <b>E.</b> Representative flow cytometry graphs depicting the results obtained from culturing PBMC from HIV-TB and HD individuals as indicated above.</p

Epidemiological characteristics of the subjects enrolled.

<p>BCG, Bacillus Calmette-Guerin. IQR, Interquartile range.</p

Modulation of <i>M. tuberculosis</i>-induced IFN-γ production by adrenal hormones.

<p><b>A.</b> Absolute numbers of IFN-γ producer cells from PBMC of HIV+, HIV-TB, IRIS, HIV-LTB and HD patients, which have been stimulated with <i>M. tuberculosis</i> antigen for 16 hours. Horizontal lines indicate the mean and comparisons between groups and statistically significant differences are shown. SFU, Spots forming units. <b>B.</b> Percentage of IFN-γ producer cells relative to <i>M. tuberculosis</i> in PBMC of HIV-TB patients (n = 12). PBMC (10⁵ cells/well) were stimulated in the presence of <i>M. tuberculosis</i> with or without addition of DHEA and/or cortisol at the indicated concentrations. Each bar illustrates the mean ± SEM of the percentage for IFN-γ producer cells relative to <i>M. tuberculosis</i> for the each group, calculated as follows: % of IFN-γ relative to <i>Mtb</i> = ([(<i>Mtb</i> hormone-Media)-(<i>Mtb</i>-Media)]/(<i>Mtb</i>-Media))×100. Asterisks indicate comparisons between each condition against <i>Mtb</i> specific response. *: <i>p</i> < 0.05; **: <i>p</i> < 0.01.</p

Cortisol, dehidroepindrosterone-sulfate (DHEA-s), DHEA levels and Cortisol/DHEA ratio in HIV, HIV-TB, IRIS, HIV-LTB and HD.

<p><b>A.</b> DHEA, <b>B.</b> DHEA-s, <b>C.</b> Cortisol and <b>D.</b> Cortisol/DHEA (ratio) levels in plasma of HIV+, HIV-TB, IRIS, HIV-LTB, and HD individuals. Bars indicate the mean ± SEM for each group. Horizontal lines indicate comparisons between groups and statistically significant differences. DHEA was measured by radioimmunoassay, DHEA-s by immunochemoluminiscence tests and Cortisol by electrochemiluminescence. HIV+ individuals n = 10, HIV-TB n = 21, IRIS n = 6, HIV-LTB n = 5, and HD n = 16. *: <i>p</i> < 0.05; **: <i>p</i> < 0.01; ***: <i>p</i> < 0.001.</p

Expanded “non-conventional” Treg in HIV-TB and IRIS patients.

<p>Analysis of CD4+FoxP3+CD25+ and CD4+FoxP3+CD25- populations within Treg in <b>A;</b> HIV-TB (n = 11), <b>B;</b> IRIS (n = 5), <b>C;</b> HIV-LTB (n = 4), <b>D;</b> HD (n = 10) and <b>E;</b> HIV (n = 7) individuals. Horizontal lines inside the boxes indicate the means ± SEM of each group. Asterisks denote comparisons between each sub-population. *: p < 0.05; **: p < 0.01; ***: p< 0.001. <b>F.</b> Spearman correlation analysis between plasma DHEA and the percentage of CD4+FoxP3+CD25- lymphocytes from all grouped patients (HIV+, HIV-TB, IRIS, HIV-LTB and HD). Results of statistical analysis are shown in the graphic.</p