Antigen presentation by HLA-DR15+ MN from individual HIV-infected and uninfected subjects.

<p>MN (5×10⁴/well) were incubated with soluble HEL or peptide and T cell hybridoma (1×10⁵/well) for 24 hrs. Viremic HIV-infected individuals with VL>1000 are High VL and VL<1000 are Low VL. A. Control MN and soluble HEL, B. HIV+ MN and soluble HEL, C. Combined data on soluble HEL presentation, D. Control MN and peptide, E. HIV+ MN and peptide, F. Combined data on peptide presentation.</p

Antigen presentation by HLA-DR1+ MN from individual HIV-infected and uninfected subjects.

<p>In all four panels, MN (5×10⁴/well) were incubated with soluble antigen or peptide and T cell hybridoma (1×10⁵/well) for 24 hrs. Viremic HIV-infected individuals with VL>1000 are High VL and VL<1000 are Low VL. A. Control MN and soluble RT, B. HIV+ MN and soluble RT, C. Combined data on soluble RT presentation, D. Control MN and peptide, E. HIV+ MN and peptide, F. Combined data on peptide presentation.</p

Relationship between HLA-DR level and antigen presentation.

<p>Data from all 38 subjects in the study were normalized. The individual's ratio of presentation was calculated by dividing their IL-2 level for the middle concentration of RT, HEL or peptide by the mean IL-2 level from the control subjects at the same middle concentration of antigen or peptide. Open red diamonds are data from HIV-infected individuals and closed black diamonds for uninfected controls.</p

Characterization of T cell hybridoma clone 15HEL.

<p>MN (5×10⁴/well) were incubated with antigen (intact HEL for A and C, HEL peptides for B) and 15HEL (1×10⁵/well) for 24 hrs. Supernatants were harvested, and IL-2 measured by ELISA. A. Effect of anti-HLA-DR or isotype on response. B. Identification of HEL peptide epitope recognized by 15HEL. C. Effect of anti-CD80 and CD86 antibodies together (5 µg/ml of each) or isotype control (10 µg/ml) on responses. Results of A and C represent triplicate wells and error bars SEM.</p

Immune Activation at Sites of HIV/TB Co-Infection Contributes to the Pathogenesis of HIV-1 Disease

<div><p>Systemic immune activation is critical to the pathogenesis of HIV-1 disease, and is accentuated in HIV/TB co-infected patients. The contribution of immune activation at sites of HIV/TB co-infection to viral activity, CD4 T cell count, and productive HIV-1 infection remain unclear. In this study, we measured markers of immune activation both in pleural fluid and plasma, and in T cells in pleural fluid mononuclear cell (PFMC) and peripheral blood mononuclear cell (PBMC) in HIV/TB co-infected subjects. The relationship between soluble and T cell activation markers with viral load in pleural fluid and blood CD4 T cell count were assessed. The T cell phenotype and activation status of HIV-1 p24 + T cells in PFMC and PBMC from HIV/TB patients were determined. We found that T cell and macrophage-specific and non-specific soluble markers of immune activation, sCD27, sCD163, IL1Ra, and sCD14, were higher in pleural fluid as compared to plasma from HIV/TB co-infected subjects, and higher as compared to pleural fluid from TB mono-infected subjects. Intestinal fatty acid-binding protein, a marker of intestinal tract damage, in plasma from HIV/TB co-infected patients was not different than that in HIV+ subjects. Expression of HLADR and CD38 double positive (HLADR/CD38) on CD4 T cells, and CD69+ on CD8 T cells correlated with pleural fluid viral load, and inversely with blood CD4 T cell count. Higher expression of HLADR/CD38 and CCR5 on CD4 T cells, and HLADR/CD38 and CD69 on CD8 T cells in PFMC were limited to effector memory populations. HIV-1 p24+ CD8 negative (includes CD4 + and double negative T cells) effector memory T cells in PFMC had higher expression of HLADR/CD38, Ki67, and CCR5 compared to HIV-1 p24- CD8 negative PFMC. Cumulatively, these data indicate that sites of HIV/TB co-infection are the source of intense immune activation.</p></div

Soluble Markers of immune activation at sites of HIV/TB co-infection.

<p>Plasma (PL) and Pleural fluid (PF) from HIV/TB co-infected subjects were assessed for soluble activation markers and compared to PL from HIV-1 infected and PF from TB mono-infected subjects. (<b>A</b>) sCD14 levels in PL and PF, (<b>B</b>) sCD14 correlation between PF and PL, (<b>C</b>) sCD163 and (<b>D</b>) sCD27 levels in PL and PF. **, p <b><</b> 0.01 and ***, p < 0.001</p

Expression of markers of immune activation on central and effector memory CD4 and CD8 T cells in PFMC.

<p>Expression of activation markers (HLADR/CD38, CCR5, CD69) was assessed on central memory (Tcm) (<b>A</b> and <b>C</b>) and effector memory (Tem) (<b>B</b> and <b>D</b>) for CD4 (<b>A</b> and <b>B</b>) and CD8 (<b>C</b> and <b>D</b>) in PBMC and PFMC T cells from HIV/TB co-infected subjects. **, p <b><</b> 0.01 and ***, p < 0.001</p

Expression of markers of immune activation and proliferation on CD4 and CD8 T cells at sites of HIV/TB, and association with HIV viral load in pleural fluid and CD4 T cell counts.

<p>Markers of immune activation and proliferation were assessed on PFMC T cells as compared to PBMC from HIV/TB co-infected subjects, and between PFMC from HIV/TB and TB mono-infected subjects. Analysis of CD4+ (<b>A</b>) and CD8+ (<b>B</b>)T cells are shown. Association of HLADR/CD38 on PFMC CD4 (<b>C and E</b>) and CD69 on PFMC CD8 (<b>D and F</b>) T cells with HIV viral load in pleural fluid (PF) <b>(C and D)</b> and blood CD4 T cell count (<b>E and F</b>). *, p< 0.05; **, p< 0.01 and ***, p<b><</b> 0.001.</p

IFABP in HIV/TB subjects with pleural TB.

<p>IFABP levels were measured in pleural fluid (PF) and plasma (PL) from HIV/TB co-infected patients, and in PL from CD4-matched HIV-1 infected (HIV) and HIV-1 un-infected healthy (HC) subjects.</p

Activation and differentiation state of PFMC T cells with productive HIV-1infection.

<p>T cells were gated by expression of intracellular HIV-1 p24 and differentiation and activation markers (CD45RO, CCR7, HLADR, CD38), proliferation marker (Ki67) and CCR5. <b>(A)</b> A representative analysis of PFMC p24+ (lower panel) and p24- (upper panel) CD8- T cells is shown. <b>(B):</b> Comparative analysis of frequency of naïve, central memory (TCM), effector memory (TEM) and terminally differentiated (TEMRA) HIV-1 p24 positive and negative CD8- T cells. <b>(C)</b>: Analysis of HLADR/CD38, Ki67 and CCR5 on HIV-1 p24 positive (black bar) and negative CD8- T cells (white bar) (n = 8) are shown. Association of HIV-1 p24 +CD8—T cells with viral load in pleural fluid <b>(D</b>) and HLADR+/CD38+ dual positive CD4 T cells <b>(E)</b>. *, p<0.05, **, p<0.01 and ***, p<0.001.</p