Ex Vivo Host and Parasite Response to Antileishmanial Drugs and Immunomodulators

Abstract

BackgroundTherapeutic response in infectious disease involves host as well as microbial determinants. Because the immune and inflammatory response to Leishmania (Viannia) species defines the outcome of infection and efficacy of treatment, immunomodulation is considered a promising therapeutic strategy. However, since Leishmania infection and antileishmanial drugs can themselves modulate drug transport, metabolism and/or immune responses, immunotherapeutic approaches require integrated assessment of host and parasite responses.MethodologyTo achieve an integrated assessment of current and innovative therapeutic strategies, we determined host and parasite responses to miltefosine and meglumine antimoniate alone and in combination with pentoxifylline or CpG 2006 in peripheral blood mononuclear cells (PBMCs) of cutaneous leishmaniasis patients. Parasite survival and secretion of TNF-α, IFN-γ, IL-10 and IL-13 were evaluated concomitantly in PBMCs infected with Luc-L. (V.) panamensis exposed to meglumine antimoniate (4, 8, 16, 32 and 64 μg SbV/mL) or miltefosine (2, 4, 8, 16 and 32 μM HePC). Concentrations of 4 μM of miltefosine and 8 μg SbV/mL were selected for evaluation in combination with immunomodulators based on the high but partial reduction of parasite burden by these antileishmanial concentrations without affecting cytokine secretion of infected PBMCs. Intracellular parasite survival was determined by luminometry and cytokine secretion measured by ELISA and multiplex assays.Principal FindingsAnti- and pro-inflammatory cytokines characteristic of L. (V.) panamensis infection were evaluable concomitantly with viability of Leishmania within monocyte-derived macrophages present in PBMC cultures. Both antileishmanial drugs reduced the parasite load of macrophages; miltefosine also suppressed IL-10 and IL-13 secretion in a dose dependent manner. Pentoxifylline did not affect parasite survival or alter antileishmanial effects of miltefosine or meglumine antimoniate. However, pentoxifylline diminished secretion of TNF-α, IFN-γ and IL-13, cytokines associated with the outcome of infection by species of the Viannia subgenus. Exposure to CpG diminished the leishmanicidal effect of meglumine antimoniate, but not miltefosine, and significantly reduced secretion of IL -10, alone and in combination with either antileishmanial drug. IL-13 increased in response to CpG plus miltefosine.Conclusions and SignificanceHuman PBMCs allow integrated ex vivo assessment of antileishmanial treatments, providing information on host and parasite determinants of therapeutic response that may be used to tailor therapeutic strategies to optimize clinical resolution.

The Src kinases Hck, Fgr, and Lyn activate Abl2/Arg to facilitate IgG-mediated phagocytosis andLeishmaniainfection

“…Background levels were assessed using unstimulated cell supernatants. Levels of the cytokines and chemokines IL-1β, IL-1α, IP-10, MIP-1α (CCL3), MIP-2 (CXCL2), RANTES (CCL5), and MIP-1β (CCL4) were profiled by using the custom multiplex Luminex Platform (R&D Systems, Inc, Minneapolis, MN) through the UT Southwestern Microarray Core as described previously (Gonzalez-Fajardo et al, 2015;Navas et al, 2014). CXCL9 levels were assessed by ELISA (R&D Systems, Inc).…”

Glucantime reduces mechanical hyperalgesia in cutaneous leishmaniasis and complete Freund's adjuvant models of chronic inflammatory pain

“…However, other studies have demonstrated that proinflammatory cytokines may be decreased during antimonial therapy in leishmaniasis . These data, in turn, demonstrated that immunomodulatory activity of Glucantime is dependent on the amount and time of infection as well as dose and time of treatment with Glucantime suggesting that leishmanicidal properties occurs through several mechanisms and not solely by the immunomodulatory effect …”

Profiles of Local and Systemic Inflammation in the Outcome of Treatment of Human Cutaneous Leishmaniasis Caused byLeishmania(Viannia)

“…Our parallel expression profiling of PBMCs and lesion biopsy specimens from CL patients supports and extends these observations, demonstrating the concurrence of some immune response parameters in the skin and blood cells during in vivo exposure to antimonials and also underscoring the importance of direct evaluation of lesion samples to describe and identify the determinants of pathogenesis and healing. Nevertheless, the feasibility and potential to elicit and screen cell-or mediator-specific responses in PBMCs support the usefulness of PBMCs as biologically relevant models to explore mechanisms of modulation and/or intervention of immune and inflammatory pathways (34).…”