Suchergebnisse

Trypanosoma cruzi, the protozoan parasite responsible for Chagas' disease, causes severe myocarditis often resulting in death. Here, we report that Slamf1-/- mice, which lack the hematopoietic cell surface receptor Slamf1, are completely protected from an acute lethal parasite challenge. Cardiac damage was reduced in Slamf1-/- mice compared to wild type mice, infected with the same doses of parasites, as a result of a decrease of the number of parasites in the heart even the parasitemia was only marginally less. Both in vivo and in vitro experiments reveal that Slamf1-defIcient myeloid cells are impaired in their ability to replicate the parasite and show altered production of cytokines. Importantly, IFN-γ production in the heart of Slamf1 deficient mice was much lower than in the heart of wt mice even though the number of infiltrating dendritic cells, macrophages, CD4 and CD8 T lymphocytes were comparable. Administration of an anti-Slamf1 monoclonal antibody also reduced the number of parasites and IFN-γ in the heart. These observations not only explain the reduced susceptibility to in vivo infection by the parasite, but they also suggest human Slamf1 as a potential target for therapeutic target against T. cruzi infection. ; Fondo de Investigación Sanitaria (PI040993); Ministerio de Ciencia e Innovación (SAF2005-02220, SAF2007-61716 and SAF2010-18733); European Union (Eicosanox and ChagasEpiNet); CSIC-CONICET; BSCH/UAM; Comunidad de Madrid S2010/BMD-2332; RED RECAVA RD06/0014/1013; RED RICET RD06/0021/0016; Fundacion Ramon Areces; Government of Panama ; Peer reviewed

Background Leishmania major cutaneous leishmaniasis is an infectious zoonotic disease. It is produced by a digenetic parasite, which resides in the phagolysosomal compartment of different mammalian macrophage populations. There is an urgent need to develop new therapies (drugs) against this neglected disease that hits developing countries. The main goal of this work is to establish an easier and cheaper tool of choice for real-time monitoring of the establishment and progression of this pathology either in BALB/c mice or in vitro assays. To validate this new technique we vaccinated mice with an attenuated Δhsp70-II strain of Leishmania to assess protection against this disease. ; Methodology We engineered a transgenic L. major strain expressing the mCherry red-fluorescent protein for real-time monitoring of the parasitic load. This is achieved via measurement of fluorescence emission, allowing a weekly record of the footpads over eight weeks after the inoculation of BALB/c mice. ; Results In vitro results show a linear correlation between the number of parasites and fluorescence emission over a range of four logs. The minimum number of parasites (amastigote isolated from lesion) detected by their fluorescent phenotype was 10,000. The effect of antileishmanial drugs against mCherry+L. major infecting peritoneal macrophages were evaluated by direct assay of fluorescence emission, with IC50 values of 0.12, 0.56 and 9.20 µM for amphotericin B, miltefosine and paromomycin, respectively. An experimental vaccination trial based on the protection conferred by an attenuated Δhsp70-II mutant of Leishmania was used to validate the suitability of this technique in vivo. ; Conclusions A Leishmania major strain expressing mCherry red-fluorescent protein enables the monitoring of parasitic load via measurement of fluorescence emission. This approach allows a simpler, faster, non-invasive and cost-effective technique to assess the clinical progression of the infection after drug or vaccine therapy. ; This research was supported by Ministerio de Ciencia y Tecnología (grants AGL2010 16078/GAN), Instituto de Salud Carlos III (grant PI09/0448 and the Network of Tropical Diseases RICET RD06/0021/1004). RAV, CFP and ECA are pre-doctoral fellows granted by RICET (ISCIII), Junta de Castilla y León (ESF; European Social Founding) and University of León, respectively to RMR. Instituto de Salud Carlos III (Network of Tropical Disasese RICET RD06/0021/0016), Ministerio de Ciencia e Innovación (SAF2010-17833), ChagasEpiNet 223034 European Union Seventh Framework Programme and Fundación Ramón Areces to MF. Funding by ISCIII-RETIC RD06/0021/0008-FEDER to JMR and ISCIII-RETIC RD06/0021/0006-FEDER to LR is also acknowledged. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. ; Peer reviewed

Background: Psoriasis is a frequent inflammatory skin disease that is mainly mediated by IL-23, IL-1β, and IL-17 cytokines. Although psoriasis is a hyperproliferative skin disorder, the possible role of amino acid transporters has remained unexplored. Objective: We sought to investigate the role of the essential amino acid transporter L-type amino acid transporter (LAT) 1 (SLC7A5) in psoriasis. Methods: LAT1 floxed mice were crossed to Cre-expressing mouse strains under the control of keratin 5, CD4, and retinoic acid receptor–related orphan receptor γ. We produced models of skin inflammation induced by imiquimod (IMQ) and IL-23 and tested the effect of inhibiting LAT1 (JPH203) and mammalian target of rapamycin (mTOR [rapamycin]). Results: LAT1 expression is increased in keratinocytes and skin-infiltrating lymphocytes of psoriatic lesions in human subjects and mice. LAT1 deletion in keratinocytes does not dampen the inflammatory response or their proliferation, which could be maintained by increased expression of the alternative amino acid transporters LAT2 and LAT3. Specific deletion of LAT1 in γδ and CD4 T cells controls the inflammatory response induced by IMQ. LAT1 deletion or inhibition blocks expansion of IL-17–secreting γ4+δ4+ and CD4 T cells and dampens the release of IL-1β, IL-17, and IL-22 in the IMQ-induced model. Moreover, inhibition of LAT1 blocks expansion of human γδ T cells and IL-17 secretion by human CD4 T cells. IL-23 and IL-1β stimulation upregulates LAT1 expression and induces mTOR activation in IL-17+ γδ and TH17 cells. Deletion or inhibition of LAT1 efficiently controls IL-23– and IL-1β–induced phosphatidylinositol 3-kinase/AKT/mTOR activation independent of T-cell receptor signaling. Conclusion: Targeting LAT1-mediated amino acid uptake is a potentially useful immunosuppressive strategy to control skin inflammation mediated by the IL-23/IL-1β/IL-17 axis. ; Supported by grants SAF 2017-82886-R (to F.S.-M.), PI17/01972 (to E.D.), and SAF 2013-42850-R (to M.F.) from the Spanish Ministry of Economy and Competitiveness; CAM (S2017/BMD-3671-INFLAMUNE-CM) from the Comunidad de Madrid (to F.S.-M.); and CIBERCV, BIOIMID PIE13/041 from Instituto de Salud Carlos III and Fundación La Marat o TV3 (20152330 31). The project leading to these results has also received funding from FUNDACIóN BBVA A EQUIPOS DE INVESTIGACIÓN CIENTÍFICA 2018 and from ''la Caixa'' Banking Foundation under the project code HR17-00016 (to F.S.-M.) and from Agencia Estatal de Investigación, Fondo Europeo de Desarrollo Regional, European Union

[Background]: Psoriasis is a frequent inflammatory skin disease that is mainly mediated by IL-23, IL-1β, and IL-17 cytokines. Although psoriasis is a hyperproliferative skin disorder, the possible role of amino acid transporters has remained unexplored. ; [Objective]: We sought to investigate the role of the essential amino acid transporter L-type amino acid transporter (LAT) 1 (SLC7A5) in psoriasis. ; [Methods]: LAT1 floxed mice were crossed to Cre-expressing mouse strains under the control of keratin 5, CD4, and retinoic acid receptor–related orphan receptor γ. We produced models of skin inflammation induced by imiquimod (IMQ) and IL-23 and tested the effect of inhibiting LAT1 (JPH203) and mammalian target of rapamycin (mTOR [rapamycin]). ; [Results]: LAT1 expression is increased in keratinocytes and skin-infiltrating lymphocytes of psoriatic lesions in human subjects and mice. LAT1 deletion in keratinocytes does not dampen the inflammatory response or their proliferation, which could be maintained by increased expression of the alternative amino acid transporters LAT2 and LAT3. Specific deletion of LAT1 in γδ and CD4 T cells controls the inflammatory response induced by IMQ. LAT1 deletion or inhibition blocks expansion of IL-17–secreting γ4+δ4+ and CD4 T cells and dampens the release of IL-1β, IL-17, and IL-22 in the IMQ-induced model. Moreover, inhibition of LAT1 blocks expansion of human γδ T cells and IL-17 secretion by human CD4 T cells. IL-23 and IL-1β stimulation upregulates LAT1 expression and induces mTOR activation in IL-17+ γδ and TH17 cells. Deletion or inhibition of LAT1 efficiently controls IL-23– and IL-1β–induced phosphatidylinositol 3-kinase/AKT/mTOR activation independent of T-cell receptor signaling. ; [Conclusion]: Targeting LAT1-mediated amino acid uptake is a potentially useful immunosuppressive strategy to control skin inflammation mediated by the IL-23/IL-1β/IL-17 axis. ; Supported by grants SAF2017-82886-R (to F.S.-M.), PI17/01972 (to E.D.), and SAF2013-42850-R (to M.F.) from the Spanish Ministry of Economy and Competitiveness; CAM (S2017/BMD-3671-INFLAMUNE-CM) from the Comunidad de Madrid (to F.S.-M.); and CIBERCV, BIOIMID PIE13/041 from Instituto de Salud Carlos III and Fundacion La Marato TV3 (20152330 31). The project leading to these results has also received funding from FUNDACION BBVA A EQUIPOS DE INVESTIGACION CIENTIFICA 2018 and from ''la Caixa'' Banking Foundation under the project code HR17-00016 (to F.S.-M.) and from Agencia Estatal de Investigacion, Fondo Europeo de Desarrollo Regional, European Union. ; Peer reviewed

Clinical cases of neonatal listeriosis are associated with brain disease and fetal loss due to complications in early or late pregnancy, which suggests that microglial function is altered. This is believed to be the first study to link microglial apoptosis with neonatal listeriosis and listeriosis-associated brain disease, and to propose a new nanovaccine formulation that reverses all effects of listeriosis and confers Listeria monocytogenes (LM)-specific immunity. We examined clinical cases of neonatal listeriosis in 2013–2015 and defined two useful prognostic immune biomarkers to design listeriosis vaccines: high anti-GAPDH1-22 titres and tumor necrosis factor (TNF)/ interleukin (IL)-6 ratios. Therefore, we developed a nanovaccine with gold glyconanoparticles conjugated to LM peptide 1-22 of GAPDH (Lmo2459), GNP-GAPDH1-22 nanovaccines formulated with a pro-inflammatory Toll-like receptor 2/4-targeted adjuvant. Neonates born to non-vaccinated pregnant mice with listeriosis, showed brain and vascular diseases and significant microglial dysfunction by induction of TNF-α-mediated apoptosis. This programmed TNF-mediated suicide explains LM dissemination in brains and livers and blocks production of early pro-inflammatory cytokines such as IL-1β and interferon-α/β. In contrast, neonates born to GNPGAPDH 1–22-vaccinated mothers before LM infection, did not develop listeriosis or brain diseases and had functional microglia. In nanovaccinated mothers, immune responses shifted towards Th1/IL-12 pro-inflammatory cytokine profiles and high production of anti-GAPDH1–22 antibodies, suggesting good induction of LM-specific memory ; This study was supported by IDIVAL grants API2010/03/SAF2009-08695, AIP13/2014/SAF2012- 34203 and INNVAL15/01 (to C. A-D and J.F), the Spanish Government through Plan Nacional (MINECO) (SAF2012-34203), FIPSE (ISCIII) (00-00002755-16) and CIBER-BNN (CIB16-NM009) (to C. A-D and I. G), RED RICET RD12/0018/0004 and BIOMID (ISCIII), SAF2013-42850-R (MINECO), HOMIN-317057- FP7-PEOPLE-2012-ITN (EU) (to M.F) and by the Spanish Government through Plan Nacional (MINECO) (CTQ2011-27268) (to S.P).