Avaliação da resposta imune a antígenos de fase ativa, de fase latente (DOSR) e de fase de reativação (RPF) de Mycobacterium tuberculosis em pacientes com tuberculose e contatos com diagnóstico de infecção latente

Abstract

Tuberculosis (TB) remains a major public health problem in the world. The identification of new Mycobacterium tuberculosis antigens and infection biomarkers capable of assisting in the diagnosis of active disease and latent infection are important targets for TB control. In the present study, TB patients before and after receiving treatment and contacts TB diagnosed with latent infection (PPD +), were studied. We investigated the levels of serum IgG and IgG1 and the production of cytokines and chemokines against the following antigens: 1) antigens from the active phase of infection (ESAT-6/CFP-10, Rv0717 and Rv3353); 2) latent infection antigens or DosR (Rv1733, Rv2628, Rv2029 and Rv1737); and 3) antigens from the reactivation phase of infection or Rpf (Rv0867 and Rv2389). The production of IFN-ꝩ, TNF-α, IL-17, IL-10 and IL-4 was evaluated in peripheral blood mononuclear cell (PBMC) cultures stimulated with M. tuberculosis antigens, by using ELISA and/or CBA-cytometry. Serum levels of CCL-2, CXCL-8, CXCL-9 and CXCL-10 were measured in TB patients and controls. Our results show elevated serum levels of IgG1 antibodies against ESAT6/CFP-10, Rv0717, Rv3353, Rv1733, Rv2628, Rv2029 and Rv0867 in patients with active TB, in comparison with the control group (p<0.001). After completion of the chemotherapy, the IgG1 response to ESAT-6/CFP-10, Rv3353, Rv2628, Rv2029 and Rv0867 was reduced to control levels (p <0.01). The ROC curve analysis confirmed the good performance of Rv0717, Rv3353, Rv1733, Rv2628, Rv2029 and Rv0867, and antigens in the diagnosis of TB. Interestingly, Rv0717 and Rv1733 induced an IgG1 peak response after 1-3 months of chemotherapy (p<0.01). In this patient group (1-3m), a high IFN-ꝩ and TNF-α response was observed after stimulation of PBMC with ESAT-6/CFP-10, Rv1733 and Rv2029. The groups did not differ in IL-17, IL-10 and IL4, indicating a predominant Th1 response in the patients studied. High levels of CXCL8, CXCL-9 and CXCL-10 were detected in the serum of patients with active TB. After 6 months of treatment, levels of both CXCL-9 and CXCL-10 were reduced to control levels. Elevated serum levels of IgG and IgG1 specific for both ESAT-6/CFP-10 and Rv1733 were observed in patients with active TB and in TB contacts, in comparison with the control group (p<0.05). Similarly, TB patients and TB contacts also did not differ in the production of IFN-ꝩ in response to ESAT-6/CFP-10, indicating that the

discrimination between patients with active TB and individuals with latent TB is difficult. Interestingly, in response to the DosR Rv1733 antigen, high levels of IFN-ꝩ were observed in PBMC cultures of TB contacts in comparison to both active TB and healthy control groups (p<0.05). Together, these results suggest the existence of tuberculosis biomarkers represented by: 1) elevated serum levels of IgG1 antibodies against ESAT6/CFP-10, Rv0717, Rv3353, Rv1733, Rv2628, Rv2029 and Rv0867; 2) high IFN-ꝩ and TNF-α production in response to different M. tuberculosis antigens; and 3) increased serum levels of CXCL-9 and CXCL-10. This profile of immune response may assist in the diagnosis of TB and monitoring drug therapy. Furthermore, detection of high levels of IFN-ꝩ in response to the Rv1733 antigen may contribute to the diagnosis of TB disease in contacts presenting asymptomatic tuberculosis.