Deficiency of Leishmania phosphoglycans influences the magnitude but does not affect the quality of secondary (memory) anti-Leishmania immunity

Despite inducing very low IFN-γ response and highly attenuated in vivo, infection of mice with phosphoglycan (PG) deficient Leishmania major (lpg2-) induces protection against virulent L. major challenge. Here, we show that mice infected with lpg2- L. major generate Leishmania-specific memory T cells. However, in vitro and in vivo proliferation, IL-10 and IFN-γ production by lpg2- induced memory cells were impaired in comparison to those induced by wild type (WT) parasites. Interestingly, TNF recall response was comparable to WT infected mice. Despite the impaired proliferation and IFN-γ response, lpg2- infected mice were protected against virulent L. major challenge and their T cells mediated efficient infection-induced immunity. In vivo depletion and neutralization studies with mAbs demonstrated that lpg2- L. major-induced resistance was strongly dependent on IFN-γ, but independent of TNF and CD8(+) T cells. Collectively, these data show that the effectiveness of secondary anti-Leishmania immunity depends on the quality (and not the magnitude) of IFN-γ response. These observations provide further support for consideration of lpg2- L. major as a live-attenuated candidate for leishmanization in humans since it protects strongly against virulent challenge, without inducing pathology in infected animals

Regulatory T cells enhance susceptibility to experimental Trypanosoma congolense infection independent of mouse genetic background.

BACKGROUND: BALB/c mice are highly susceptible while C57BL/6 are relatively resistant to experimental Trypanosoma congolense infection. Although regulatory T cells (Tregs) have been shown to regulate the pathogenesis of experimental T. congolense infection, their exact role remains controversial. We wished to determine whether Tregs contribute to distinct phenotypic outcomes in BALB/c and C57BL/6 mice and if so how they operate with respect to control of parasitemia and production of disease-exacerbating proinflammatory cytokines. METHODOLOGY/FINDINGS: BALB/c and C57BL/6 mice were infected intraperitoneally (i.p) with 10(3)T. congolense clone TC13 and both the kinetics of Tregs expansion and intracellular cytokine profiles in the spleens and livers were monitored directly ex vivo by flow cytometry. In some experiments, mice were injected with anti-CD25 mAb prior or post T. congolense infection or adoptively (by intravenous route) given highly enriched naïve CD25(+) T lymphocytes prior to T. congolense infection and the inflammatory cytokine/chemokine levels and survival were monitored. In contrast to a transient and non significant increase in the percentages and absolute numbers of CD4(+)CD25(+)Foxp3(+) T cells (Tregs) in C57BL/6 mouse spleens and livers, a significant increase in the percentage and absolute numbers of Tregs was observed in spleens of infected BALB/c mice. Ablation or increasing the number of CD25(+) cells in the relatively resistant C57BL/6 mice by anti-CD25 mAb treatment or by adoptive transfer of CD25(+) T cells, respectively, ameliorates or exacerbates parasitemia and production of proinflammatory cytokines. CONCLUSION: Collectively, our results show that regulatory T cells contribute to susceptibility in experimental murine trypanosomiasis in both the highly susceptible BALB/c and relatively resistant C57BL/6 mice

B7H1 Expression and Epithelial-To-Mesenchymal Transition Phenotypes on Colorectal Cancer Stem-Like Cells.

Cancer stem cells (CSCs) can invade and metastasize by epithelial-to-mesenchymal transition (EMT). However, how they escape immune surveillance is unclear. B7H1 is crucial negative co-stimulatory molecule but little information about whether it works in CSCs. Therefore, we determined the expression of B7H1 and EMT-associated markers in colorectal cancer stem-like cells to investigate a possible immunoevasion way of CSCs. We enriched CD133+ colorectal cancer cells which manifested the CSCs-like properties such as higher levels of other stem cell markers Oct-4 and Sox-2, tumor sphere forming ability and more tumorigenic in NOD/SCID mice. These CD133+ cells possess EMT gene expression profile including higher level of Snail, Twist, vimentin, fibronectin and lower level of E-cadherin. Moreover, CD133+ cells in both cell line and colorectal cancer tissues expressed high level of negative co-stimulate molecule B7H1. Furthermore, some B7H1+ cancer cells also showed the characteristic of EMT, indicating EMT cells could escape immune attack during metastasis. B7H1 expression and EMT phenotypes on CSCs indicates a possible immunoevasion way

CD8⁺ T cells Are Preferentially Activated during Primary Low Dose <i>Leishmania major</i> Infection but Are Completely Dispensable during Secondary Anti-<i>Leishmania</i> Immunity

<div><p>We previously showed that CD8⁺ T cells are required for optimal primary immunity to low dose <i>Leishmania major</i> infection. However, it is not known whether immunity induced by low dose infection is durable and whether CD8⁺ T cells contribute to secondary immunity following recovery from low dose infection. Here, we compared primary and secondary immunity to low and high dose <i>L. major</i> infections and assessed the influence of infectious dose on the quality and magnitude of secondary anti-<i>Leishmania</i> immunity. In addition, we investigated the contribution of CD8⁺ T cells in secondary anti-<i>Leishmania</i> immunity following recovery from low and high dose infections. We found that the early immune response to low and high dose infections were strikingly different: while low dose infection preferentially induced proliferation and effector cytokine production by CD8⁺ T cells, high dose infection predominantly induced proliferation and cytokine production by CD4⁺ T cells. This differential activation of CD4⁺ and CD8⁺ T cells by high and low dose infections respectively, was imprinted during <i>in vitro</i> and <i>in vivo</i> recall responses in healed mice. Both low and high dose-infected mice displayed strong infection-induced immunity and were protected against secondary <i>L. major</i> challenge. While depletion of CD4⁺ cells in mice that healed low and high dose infections abolished resistance to secondary challenge, depletion of CD8⁺ cells had no effect. Collectively, our results show that although CD8⁺ T cells are preferentially activated and may contribute to optimal primary anti-<i>Leishmania</i> immunity following low dose infection, they are completely dispensable during secondary immunity.</p></div

Protection in <i>lpg2-</i>infected mice is dependent on IFN-γ but is independent of CD8⁺ T cells.

<p>C57BL/6 mice were infected in the right footpad with WT and <i>lpg2− L. major</i> and after 16 weeks, challenged with virulent <i>L. major</i> in the left footpads. Some mice were treated with anti-IFN-γ or anti-CD8 mAb (1 mg/ml) 24 hr prior to challenge and once weekly for additional 3 weeks. At 3 weeks post-challenge, lesion sizes (<b>A, C</b>) were determined and mice were sacrificed and parasite burdens (<b>B, D</b>) in the challenged footpads were determined. Data presented are representative of 2–3 independent experiments (n = 3–5 mice per group) with similar results.</p

Deficiency of p110δ Isoform of the Phosphoinositide 3 Kinase Leads to Enhanced Resistance to <i>Leishmania donovani</i>

<div><p>Background</p><p>Visceral leishmaniasis is the most clinically relevant and dangerous form of human leishmaniasis. Most traditional drugs for treatment of leishmaniasis are toxic, possess many adverse reactions and drug resistance is emerging. Therefore, there is urgent need for identification of new therapeutic targets. Recently, we found that mice with an inactivating knock-in mutation in the p110δ isoform of pi3k, (p110δ^d910a) are hyper resistant to <i>L. major</i>, develop minimal cutaneous lesion and rapidly clear their parasite. Here, we investigated whether pi3k signaling also regulates resistance to <i>L. donovani</i>, one of the causative agents of visceral leishmaniasis.</p><p>Methodology/Principal Findings</p><p>WT and p110δ^D910A mice (on a BALB/c background) were infected with <i>L. donovani</i>. At different time points, parasite burden and granuloma formation were assessed. T and B cell responses in the liver and spleen were determined. In addition, Tregs were expanded <i>in vivo</i> and its impact on resistance was assessed. We found that p110δ^D910A mice had significantly reduced splenomegaly and hepatomegaly and these organs harbored significantly fewer parasites than those of WT mice. Interestingly, infected p110δ^D910A mice liver contains fewer and less organized granulomas than their infected WT counterparts. Cells from p110δ^D910A mice were significantly impaired in their ability to produce cytokines compared to WT mice. The percentage and absolute numbers of Tregs in infected p110δ^D910A mice were lower than those in WT mice throughout the course of infection. <i>In vivo</i> expansion of Tregs in infected p110δ^D910A mice abolished their enhanced resistance to <i>L. donovani</i> infection.</p><p>Conclusions/Significance</p><p>Our results indicate that the enhanced resistance of p110δ^D910A mice to <i>L. donovani</i> infection is due to impaired activities of Tregs. They further show that resistance to <i>Leishmania</i> in the absence of p110δ signaling is independent of parasite species, suggesting that targeting the PI3K signaling pathway may be useful for treatment of both visceral and cutaneous leishmaniasis.</p></div

The differential expansion of CD4⁺ and CD8⁺ T cells by high and low dose infections, respectively is sustained during recall response.

<p>C57BL/6 mice infected with low dose (1×10³) and high dose (2×10⁶) <i>L. major</i> and allowed to completely resolve (heal) their lesion (>12 wks.). Healed mice were sacrificed and dLN cells were labelled with CFSE dye and co-cultured with <i>L. major</i>-infected BMDCs at a DC:dLN cell ratio of 1∶100. After 4 days, the percentages of proliferating (CFSE^lo) CD4⁺ and CD8⁺ T cells were determined by flow cytometry (A and B). The percentages of IFN-γ (upper panel) and TNF (lower panel) -producing CD4⁺ (C and D) and CD8⁺ (E and F)) T cells within the proliferating (CFSE^lo) population were also assessed. Shown are the representative histogram (A) and dot (C and E) plots of individual animals within the group and bar graphs (B, D and F) showing the means +/− SE of all the animals (3–4 mice) within the group. The absolute numbers of CD11c⁺MHCII⁺ dendritic cells in the draining lymph nodes of high and low dose infected mice were also determined by flow cytometry following Collagenase/Dispase digestion (G). Results presented are representative of 3 independent experiments (n = 3–4 mice/group) with similar results. *, p<0.05; **, p<0.01; ***, p<0.001.</p