Macrophage Polarization in Leprosy-HIV Co-infected Patients.

In HIV-infected individuals, a paradoxical clinical deterioration may occur in preexisting leprosy when highly active antiretroviral therapy (HAART)-associated reversal reaction (RR) develops. Leprosy-HIV co-infected patients during HAART may present a more severe form of the disease (RR/HIV), but the immune mechanisms related to the pathogenesis of leprosy-HIV co-infection remain unknown. Although the adaptive immune responses have been extensively studied in leprosy-HIV co-infected individuals, recent studies have described that innate immune cells may drive the overall immune responses to mycobacterial antigens. Monocytes are critical to the innate immune system and play an important role in several inflammatory conditions associated with chronic infections. In leprosy, different tissue macrophage phenotypes have been associated with the different clinical forms of the disease, but it is not clear how HIV infection modulates the phenotype of innate immune cells (monocytes or macrophages) during leprosy. In the present study, we investigated the phenotype of monocytes and macrophages in leprosy-HIV co-infected individuals, with or without RR. We did not observe differences between the monocyte profiles in the studied groups; however, analysis of gene expression within the skin lesion cells revealed that the RR/HIV group presents a higher expression of macrophage scavenger receptor 1 (MRS1), CD209 molecule (CD209), vascular endothelial growth factor (VEGF), arginase 2 (ARG2), and peroxisome proliferator-activated receptor gamma (PPARG) when compared with the RR group. Our data suggest that different phenotypes of tissue macrophages found in the skin from RR and RR/HIV patients could differentially contribute to the progression of leprosy.

Role of CD8(+) T cells in triggering reversal reaction in HIV/leprosy patients.

It has been reported that the initiation of highly active anti-retroviral therapy (HAART) is associated with the development of reversal reaction (RR) in co-infected HIV/leprosy patients. Nevertheless, the impact of HIV and HAART on the cellular immune response to Mycobacterium leprae (ML) remains unknown. In the present study, we observed that ex vivo peripheral blood mononuclear cells (PBMCs) of both RR and RR/HIV patients presented increased percentages of activated CD4(+) T cells when compared with the healthy individuals (HC) group. The frequency of CD8(+)  CD38(+) cells increased in the PBMCs of RR/HIV patients but not in RR patients when compared with the HC group. Both RR and RR/HIV skin lesion cells presented similar percentages of activated CD4(+) cells, but the numbers of activated CD8(+) cells were higher in RR/HIV in comparison to the RR group. The frequency of interferon-γ-producing cells was high in response to ML regardless of HIV co-infection. In ML-stimulated cells, there was an increase in central memory CD4(+) T-cell frequencies in the RR and RR/HIV groups, but an increase in central memory CD8(+) T-cell frequency was only observed in the RR/HIV group. ML increased granzyme B(+) effector memory CD8(+) T-cell frequencies in the RR/HIV PBMCs, but not in the HC and RR groups. Our data suggest that the increased expression of effector memory CD8(+) T cells, together with greater perforin/granzyme B production, could be an additional mechanism leading to the advent of RR in co-infected patients. Moreoever, this increased expression may explain the severity of RR occurring in these patients.

Evaluation of cellular phenotypes implicated in immunopathogenesis and monitoring immune reconstitution inflammatory syndrome in HIV/leprosy cases.

BACKGROUND: It is now evident that HAART-associated immunological improvement often leads to a variety of new clinical manifestations, collectively termed immune reconstitution inflammatory syndrome, or IRIS. This phenomenon has already been described in cases of HIV coinfection with Mycobacterium leprae, most of them belonging to the tuberculoid spectrum of leprosy disease, as observed in leprosy reversal reaction (RR). However, the events related to the pathogenesis of this association need to be clarified. This study investigated the immunological profile of HIV/leprosy patients, with special attention to the cellular activation status, to better understand the mechanisms related to IRIS/RR immunopathogenesis, identifying any potential biomarkers for IRIS/RR intercurrence. METHODS/PRINCIPAL FINDINGS: Eighty-five individuals were assessed in this study: HIV/leprosy and HIV-monoinfected patients, grouped according to HIV-viral load levels, leprosy patients without HIV coinfection, and healthy controls. Phenotypes were evaluated by flow cytometry for T cell subsets and immune differentiation/activation markers. As expected, absolute counts of the CD4+ and CD8+ T cells from the HIV-infected individuals changed in relation to those of the leprosy patients and controls. However, there were no significant differences among the groups, whether in the expression of cellular differentiation phenotypes or cellular activation, as reflected by the expression of CD38 and HLA-DR. Six HIV/leprosy patients identified as IRIS/RR were analyzed during IRIS/RR episodes and after prednisone treatment. These patients presented high cellular activation levels regarding the expression of CD38 in CD8+ cells T during IRIS/RR (median: 77,15%), dropping significantly (p<0,05) during post-IRIS/RR moments (median: 29,7%). Furthermore, an increase of cellular activation seems to occur prior to IRIS/RR. CONCLUSION/SIGNIFICANCE: These data suggest CD38 expression in CD8+ T cells interesting tool identifying HIV/leprosy individuals at risk for IRIS/RR. So, a comparative investigation to leprosy patients at RR should be conducted.

Schwann cells producing matrix metalloproteinases under Mycobacterium leprae stimulation may play a role in the outcome of leprous neuropathy.

Matrix metalloproteinases (MMPs) mediate demyelination and breakdown of the blood-nerve barrier in peripheral neuropathies. Matrix metalloproteinases and tissue inhibitor of metalloproteinase 1 gene expression and secretion were studied in cells of the human Schwann cell line ST88-14 stimulated with Mycobacterium leprae and tumor necrosis factor (TNF) and in nerve biopsies from patients with neural leprosy (n = 21) and nonleprous controls (n = 3). Mycobacterium leprae and TNF induced upregulation of MMP-2 and MMP-9 and increased secretion of these enzymes in cultured ST88-14 cells. The effects of TNF and M. leprae were synergistic, and anti-TNF antibody blockage partially inhibited this synergistic effect. Nerves with inflammatory infiltrates and fibrosis displayed higher TNF, MMP-2, and MMP-9 mRNA than controls. Leprous nerve biopsies with no inflammatory alterations also exhibited higher MMP-2 and MMP-9; tissue inhibitor of metalloproteinase 1 was significantly higher in biopsies with fibrosis and inflammation. Immunohistochemical double labeling of the nerves demonstrated that the MMPs were mainly expressed by macrophages and Schwann cells. The biopsies with endoneurial inflammatory infiltrates and epithelioid granulomas had the highest levels of MMP-2 and MMP-9 mRNA detected. Together, these results suggest that M. leprae and TNF may directly induce Schwann cells to upregulate and secrete MMPs regardless of the extent of inflammation in leprous neuropathy.

MMP-2 e MMP-9 na patogênese da lesão neural da hanseníase / MMP-2 and MMP-9 in the patogenesis of neural injury in leprosy

As metaloproteinases de matriz (MMPs) formam uma família de endopeptidades cálcio-dependentes capazes de degradar componentes da matriz extracelular, influenciando em processos como migração e interações célula-célula. Além disso, atuam em condições patológicas e em danos teciduais participando de doenças do sistema nervoso e doenças infecciosas, principalmente quando em desiquilíbrio com seu inibidor tecidual específico, os TIMPs. A hanseníase é caracterizada por lesões de pele e nervo, esta última, conseqüência da interação do M.leprae (ML) com a Célula de Schwann (CS). A expressão de MMP-2 e MMP-9 encontra-se aumentada em lesões de nervo periférico de pacientes com hanseníase sugerindo uma participação dessas MMPs nesta doença. O objetivo deste trabalho é elucidar se as CS são capazes de produzir MMP-2 e MMP-9 quando infectadas com M.leprae e se o TNF, uma importante citocina proinflamatória na hanseníase, está contribuindo com essa produção. Para isso, CS humanas da linhagem ST88-14 e CS primárias foram cultivadas e estimuladas com ML e/ou TNF. Em seguida, o RNA total dessas células foi extraído para avaliar a expressão de MMPs e TIMP-1 por RT-PCR em tempo real. Para verificar o perfil de secreção das MMPs e TIMP-1 as células foram cultivadas por 24 horas nas mesmas condições descritas anteriormente e os sobrenadantes foram então analisados por ELISA. Além disso, os mesmos sobrenadantes foram submetidos à técnica de zimografia para avaliar a atividade proteolítica de MMP-2 e MMP-9. A expressão de mRNA de MMPs foi detectada constitutivamente nessas células. No entanto, quando as culturas foram estimuladas com ML, os níveis de mRNA e de proteína de MMP-2 e MMP-9 aumentaram em relação ao controle enquanto que os níveis de TIMP-1 não foram alterados. Quando a razão entre MMP-9 e TIMP-1 foi avaliada um desiquilíbrio favorecendo MMP-9 foi observado. O TNF associado ao ML também aumentou a expressão e secreção de MMP-9, porém não aumentou a secreção...fibrose.