Interdependencies between Toll-like receptors in Leishmania infection.

Multiple pathogen-associated molecular patterns (PAMPs) on a pathogen's surface imply their simultaneous recognition by the host cell membrane-located multiple PAMP-specific Toll-like receptors (TLRs). The TLRs on endosomes recognize internalized pathogen-derived nucleic acids and trigger anti-pathogen immune responses aimed at eliminating the intracellular pathogen. Whether the TLRs influence each other's expression and effector responses-termed TLR interdependency-remains unknown. Herein, we first probed the existence of TLR interdependencies and next determined how targeting TLR interdependencies might determine the outcome of Leishmania infection. We observed that TLRs selectively altered expression of their own and of other TLRs revealing novel TLR interdependencies. Leishmania major-an intra-macrophage parasite inflicting the disease cutaneous leishmaniasis in 88 countries-altered this TLR interdependency unfolding a unique immune evasion mechanism. We targeted this TLR interdependency by selective silencing of rationally chosen TLRs and by stimulation with selective TLR ligands working out a novel phase-specific treatment regimen. Targeting the TLR interdependency elicited a host-protective anti-leishmanial immune response and reduced parasite burden. To test whether this observation could be used as a scientific rationale for treating a potentially fatal L. donovani infection, which causes visceral leishmaniasis, we targeted the inter-TLR dependency adopting the same treatment regimen. We observed reduced splenic Leishman-Donovan units accompanied by host-protective immune response in susceptible BALB/c mice. The TLR interdependency optimizes TLR-induced immune response by a novel immunoregulatory framework and scientifically rationalizes targeting TLRs in tandem and in sequence for redirecting immune responses against an intracellular pathogen.

Tumor Arrests DN2 to DN3 Pro T Cell Transition and Promotes Its Conversion to Thymic Dendritic Cells by Reciprocally Regulating Notch1 and Ikaros Signaling.

Tumor progression in the host leads to severe impairment of intrathymic T-cell differentiation/maturation, leading to the paralysis of cellular anti-tumor immunity. Such suppression manifests the erosion of CD4+CD8+ double-positive (DP) immature thymocytes and a gradual increase in CD4-CD8- double negative (DN) early T-cell progenitors. The impact of such changes on the T-cell progenitor pool in the context of cancer remains poorly investigated. Here, we show that tumor progression blocks the transition of Lin-Thy1.2+CD25+CD44+c-KitlowDN2b to Lin-Thy1.2+CD25+CD44-c-Kit-DN3 in T-cell maturation, instead leading to DN2-T-cell differentiation into dendritic cells (DC). We observed that thymic IL-10 expression is upregulated, particularly at cortico-medullary junctions (CMJ), under conditions of progressive disease, resulting in the termination of IL-10Rhigh DN2-T-cell maturation due to dysregulated expression of Notch1 and its target, CCR7 (thus restricting these cells to the CMJ). Intrathymic differentiation of T-cell precursors in IL-10-/- mice and in vitro fetal thymic organ cultures revealed that IL-10 promotes the interaction between thymic stromal cells and Notch1low DN2-T cells, thus facilitating these DN2-T cells to differentiate toward CD45+CD11c+MHC-II+ thymic DCs as a consequence of activating the Ikaros/IRF8 signaling axis. We conclude that a novel function of thymically-expressed IL-10 in the tumor-bearing host diverts T-cell differentiation toward a DC pathway, thus limiting the protective adaptive immune repertoire.

Leishmania species-dependent functional duality of toll-like receptor 2.

Toll-like receptors (TLRs) are a subset of pattern recognition receptors (PRR) in innate immunity and act as a connecting link between innate and adaptive immune systems. During Leishmania infection, the activation of TLRs influences the pathogen-specific immune responses, which may play a decisive role in determining the outcome of infection, toward elimination or survival of the pathogen. Antigen-presenting cells (APCs) of the innate immune system such as macrophages, dendritic cells (DCs), neutrophils, natural killer (NK) cells, and NKT cells express TLR2, which plays a crucial role in the parasite recognition and elicitation of immune responses in Leishmania infection. Depending on the infecting Leishmania species, the TLR2 pathways may result in a host-protective or a disease-exacerbating response. While Leishmania major and Leishmania donovani infections trigger TLR2-related host-protective and non-protective immune responses, Leishmania mexicana and Leishmania infantum infections are reported to elicit TLR2-mediated host-protective responses and Leishmania amazonensis and Leishmania braziliensis infections are reported to evoke a disease-exacerbating response. These findings illustrate that TLR2-related effector functions are diverse and may be exerted in a species- or strain-dependent manner. TLR2 agonists or antagonists may have therapeutic potentials to trigger the desired immune response during leishmaniasis. In this review, we discuss the TLR2-related immune responses during leishmaniasis and highlight the novel insights into the possible role of TLR2-driven resistance or susceptibility to Leishmania.

Lactoferrin-modified Betulinic Acid-loaded PLGA nanoparticles are strong anti-leishmanials.

Visceral Leishmaniasis (VL), caused by the protozoan parasite Leishmania donovani, is a potentially fatal disease. The only orally bioavailable drug miltefosine is toxic and the effective liposomal Amphotericin B (AmBisome) is limited by its prohibitive cost and requirement for parenteral administration. Therefore, finding a new potential drug candidate and an alternative delivery system is imperative. We report that Betulinic acid (BA), a pentacyclic triterpenoid from Betula alba bark, was loaded onto uniformly spherical PLGA nanoparticles (BANPs; diameter 187.5 ±â€¯5.60 nm) coated with Lactoferrin (Lf-BANPs). The amastigotes count in macrophages was more effectively reduced by Lf-BANP than BA and BANP. Lf-BANPs reduced the pro-parasitic, anti-inflammatory cytokine IL-10, but increased nitric oxide (NO), production in L. donovani-infected macrophages indicating that Lf-BANP possesses a significant anti-leishmanial activity.

TLR11 or TLR12 silencing reduces Leishmania major infection.

Toll-like receptors (TLRs) recognize the pathogen-associated molecular patterns (PAMPs) and induce host-protective immune response. The role of the profilin-recognizing TLR11/TLR12 in Leishmania infection is unknown. Herein, we report that TLR11/ TLR12 expression increases in virulent L. major-infected macrophages but is prevented by miltefosine, an anti-leishmanial drug. While lipohosphoglycan (LPG) increases, LPG or TLR2 blockade prevents, the heightened TLR11/TLR12 expression. LPG-TLR2 interaction triggers MyD88- and TIRAP-mediated signaling enhancing ERK-1/2 activation and increased production of IL-10 that promotes TLR11/TLR12 expression. Profilin expression was higher in the virulent L. major and L. donovani parasites than that observed in the avirulent parasites. TLR11 or TLR12 silencing reduces parasite burden and increases IFN-γ, but reduces IL-4, production indicating that TLR11 and TLR12 play a pro-leishmanial role.

Pegylated bisacycloxypropylcysteine, a diacylated lipopeptide ligand of TLR6, plays a host-protective role against experimental Leishmania major infection.

TLRs recognize pathogen-expressed Ags and elicit host-protective immune response. Although TLR2 forms heterodimers with TLR1 or TLR6, recognizing different ligands, differences in the functions of these heterodimers remain unknown. In this study, we report that in Leishmania major-infected macrophages, the expression of TLR1 and TLR2, but not TLR6, increased; TLR2-TLR2 association increased, but TLR2-TLR6 association diminished. Lentivirus-expressed TLR1-short hairpin RNA (shRNA) or TLR2-shRNA administration reduced, but TLR6-shRNA increased L. major infection in BALB/c mice. Corroboratively, Pam3CSK4 (TLR1-TLR2 ligand) and peptidoglycan (TLR2 ligand) increased L. major infection but reduced TLR9 expression, whereas pegylated bisacycloxypropylcysteine (BPPcysMPEG; TLR2-TLR6 ligand) reduced L. major number in L. major-infected macrophages, accompanied by increased TLR9 expression, higher IL-12 production, and inducible NO synthase expression. Whereas MyD88, Toll/IL-1R adaptor protein, and TNFR-α-associated factor 6 recruitments to TLR2 were not different in Pam3CSK4-, peptidoglycan-, or BPPcysMPEG-treated macrophages, only BPPcysMPEG enhanced p38MAPK and activating transcription factor 2 activation. BPPcysMPEG conferred antileishmanial functions to L. major-infected BALB/c-derived T cells in a macrophage-T cell coculture and in BALB/c mice; the protection was TLR6 dependent and IL-12 dependent, and it was accompanied by reduced regulatory T cell number. BPPcysMPEG administration during the priming with fixed L. major protected BALB/c mice against challenge L. major infection; the protection was accompanied by low IL-4 and IL-10, but high IFN-γ productions and reduced regulatory T cells. Thus, BPPcysMPEG, a novel diacylated lipopeptide ligand for TLR2-TLR6 heterodimer, induces IL-12-dependent, inducible NO synthase-dependent, T-reg-sensitive antileishmanial protection. The data reveal a novel dimerization partner-dependent duality in TLR2 function.