Type 1 helper T cells and FoxP3-positive T cells in HIV-tuberculosis-associated immune reconstitution inflammatory syndrome

Abstract

Rationale: Tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) induced by combination antiretroviral therapy (cART) has been attributed to dysregulated expansion of tuberculin PPD-specific IFN-gamma-secreting CD4(+) T cells.

Objectives: To investigate the role of type 1 helper T cell expansions and regulatory T cells in HIV-TB IRIS.

Methods: Longitudinal and cross-sectional studies of Mycobacterium tuberculosis-specific IFN-gamma enzyme-linked immunospot responses and flow cytometric analysis of blood cells from a total of 129 adults with HIV-1-associated tuberculosis, 98 of whom were prescribed cART.

Measurements and main results: In cross-sectional analysis the frequency of IFN-gamma-secreting T cells recognizing early secretory antigenic target (ESAT)-6, alpha-crystallins 1 and 2, and PPD of M. tuberculosis was higher in patients with TB-IRIS than in similar patients treated for both HIV-1 and tuberculosis who did not develop IRIS (non-IRIS; P <or= 0.03). The biggest difference was in the recognition of alpha-crystallin molecules: peptide mapping indicated a polyclonal response. Flow cytometric analysis indicated equal proportions of CD4(+) and CD8(+) cells positive for activation markers HLA-DR and CD71 in both patients with TB-IRIS and non-IRIS patients. The percentage of CD4(+) cells positive for FoxP3 (Forkhead box P3) was low in both groups (TB-IRIS, 5.3 +/- 4.5; non-IRIS, 2.46 +/- 2.46; P = 0.13). Eight weeks of longitudinal analysis of patients with tuberculosis who were starting cART showed dynamic changes in antigen-specific IFN-gamma-secreting T cells in both the TB-IRIS and non-IRIS groups: the only significant trend was an increased response to PPD in the TB-IRIS group (P = 0.041).

Conclusions: There is an association between helper T-cell type 1 expansions and TB-IRIS, but the occurrence of similar expansions in non-IRIS brings into question whether these are causal. The defect in immune regulation responsible for TB-IRIS remains to be fully elucidated.

Figure 1.

Frequency of Mycobacterium tuberculosis antigen–specific IFN-γ spot-forming cells (SFCs) in HIV-infected patients with tuberculosis-associated immune reconstitution inflammatory syndrome (TB–IRIS). Two comparator groups were patients with HIV-associated tuberculosis before treatment and not receiving combination antiretroviral therapy (cART) (untreated HIV+TB), and patients with treated HIV-associated TB also receiving cART for a similar duration to the patients with TB–IRIS (non-IRIS). The frequency of IFN-γ SFCs was higher in patients with TB–IRIS than in non-IRIS patients (P ≤ 0.03), with the exception of the response to 38-kD cell wall antigen. The frequency of IFN-γ SFCs was also higher in patients with TB–IRIS than in patients with untreated HIV+TB in every instance (P ≤ 0.004). By contrast, the frequency of IFN-γ SFCs did not differ between the non-IRIS and untreated HIV+TB groups in response to any antigen.

Figure 2.

FoxP3 and T-cell activation markers in peripheral blood mononuclear cells (PBMCs) of patients with tuberculosis-associated immune reconstitution inflammatory syndrome (TB–IRIS). PBMCs from 11 TB–IRIS and 8 non-IRIS patients were analyzed. Although the proportion of activated T cells (both CD4⁺ and CD8⁺) was high in both groups, there were no significant differences between clinical groups. For FoxP3 analysis cells were either unstimulated (unstim) or restimulated (stim) with PPD for 24 hours. Again, there were no significant differences between TB–IRIS and non-IRIS groups. Error bars show the SD.

Figure 3.

Longitudinal analysis of the frequency of M. tuberculosis antigen–specific IFN-γ spot-forming cells in response to five antigens and C-reactive protein (CRP) in HIV-TB patients receiving combination antiretroviral therapy (cART). An open triangle in the TB–IRIS group indicates the point at which tuberculosis-associated immune reconstitution inflammatory syndrome (TB–IRIS) was diagnosed. Repeated measures analysis demonstrated a significant increase in PPD response only in the TB–IRIS group (P = 0.041). Within-category analysis confirmed the increase in median PPD response in the TB–IRIS group between Weeks 0 and 8 to be significant (P = 0.008). In addition, the increase in PPD response in the non-IRIS group between Week 0 and both Weeks 2 and 4 was significant (P ≤ 0.04). When comparing IRIS and non-IRIS groups the only significant difference was between the 8-week response to PPD (P = 0.001). The increase in median CRP response in the TB–IRIS group from between Weeks 0 and 2 was significant (P = 0.031). In addition, the increase in CRP in the non-IRIS group between Week 0 and both Weeks 2 and 4 (P ≤ 0.006) was significant. When comparing IRIS and non-IRIS groups significant differences in CRP were found between the 2-week response in the non-IRIS group when compared with those who developed TB–IRIS (P = 0.027).