Successful Treatment of Human Visceral Leishmaniasis Restores Antigen-Specific IFN-γ, but not IL-10 Production

Abstract

One of the key immunological characteristics of active visceral leishmaniasis (VL) is a profound immunosuppression and impaired production of Interferon-γ (IFN-γ). However, recent studies from Bihar in India showed using a whole blood assay, that whole blood cells have maintained the capacity to produce IFN-γ. Here we tested the hypothesis that a population of low-density granulocytes (LDG) might contribute to T cell responses hyporesponsiveness via the release of arginase. Our results show that this population is affected by the anticoagulant used to collect blood: the frequency of LDGs is significantly lower when the blood is collected with heparin as compared to EDTA; however, the anticoagulant does not impact on the levels of arginase released. Next, we assessed the capacity of whole blood cells from patients with active VL to produce IFN-γ and IL-10 in response to antigen-specific and polyclonal activation. Our results show that whole blood cells produce low or levels below detection limit of IFN-γ and IL-10, however, after successful treatment of VL patients, these cells gradually regain their capacity to produce IFN-γ, but not IL-10, in response to activation. These results suggest that in contrast to VL patients from Bihar, India, whole blood cells from VL patients from Gondar, Ethiopia, have lost their ability to produce IFN-γ during active VL and that active disease is not associated with sustained levels of IL-10 production following stimulation.

Visceral Leishmaniasis Patients Display Altered Composition and Maturity of Neutrophils as well as Impaired Neutrophil Effector Functions

“…These results show that despite the “clinical cure,” the immune system of these patients is still impaired at the time they are released from hospital. This is in line with our previous study that showed that the capacity of whole blood cells to respond to antigenic and polyclonal stimulation with IFN-γ production was not restored in treated patients (7). This is of relevance in Ethiopia, where a new treatment is used, a combination of PM and SSG, which reduced the hospitalization of VL patients from 28 to 17 days.…”

“…Recently, this notion was challenged by studies performed in India (5, 6): using a whole blood assay, the authors showed that CD4 + T cells produce similar levels of IFN-γ that can limit Leishmania parasite replication during active VL. In contrast, our recent work performed in Ethiopia showed that whole blood cells produce no or low IFN-γ, suggesting that whole blood cells from VL patients in Ethiopia are hyporesponsive (7). …”

Whole Blood Stimulation Assay as a Treatment Outcome Monitoring Tool for VL Patients in Ethiopia: A Pilot Evaluation

“…The latter reported on an incremental increase in IFN-γ levels in cured individuals living in Northwest Ethiopia as compared to active patients but could not detect differences in IL-10 levels. Similar findings were reported in Turkish, Indian, Iranian, Spanish, and Bangladeshi VL patients [20,21,23,32,33], indicating a typical cellular hyporesponsiveness during active disease that restores after successful therapy, as was also reported in studies using peripheral blood mononuclear cell (PBMC). This was also reflected in the gradual increase in nonspecific PHA stimulation which suggests a broad mechanism of immunosuppression that merits further investigation.…”

“…This was also reflected in the gradual increase in nonspecific PHA stimulation which suggests a broad mechanism of immunosuppression that merits further investigation. Studies in Spain and Ethiopia demonstrated that a long-lasting memory response remains in cured patients and increased levels of IFN-γ and IFN-γ-inducible protein 10 (IP-10) could be detected in almost all patients at 6 and 12 months after treatment [21,23]. This is, however, the first time that a longitudinal study in diseased individuals was performed with an early measurement at one week of treatment.…”

“…Therefore, techniques such as the whole blood stimulation assay (WBA) which measures the in vitro cell-mediated immune response after stimulation with soluble Leishmania antigens have been proposed to monitor disease recovery [19,20]. Recent findings in cross-sectional studies of Ethiopia, Spain, and Bangladesh further enforced the idea that gradual increases in IFN-γ or in IFN-γ-induced protein 10 (IP-10 or CXCL10), monokine induced by IFN-γ (MIG or CXCL9), tumor necrosis factor-alpha (TNF-α), and monocyte chemoattractant protein 1 (MCP-1 or CCL2) levels in soluble Leishmania antigen-(SLA-) stimulated whole blood could indicate that treatment has been successful and could be targeted as biomarkers of clinical cure in VL [20][21][22][23]. The search for a robust marker is still ongoing, as different cytokines and chemokines have been proposed, but vary by geographical area, Leishmania strain, and outcome identification (asymptomatic, active, cured).…”