Natural transmission of Leishmania infantum through experimentally infected Phlebotomus perniciosus highlights the virulence of Leishmania parasites circulating in the human visceral leishmaniasis outbreak in Madrid, Spain.

A human leishmaniasis outbreak is occurring in the Madrid region, Spain, with the parasite and vector involved being Leishmania infantum and Phlebotomus perniciosus respectively. The aim of this study was to investigate the virulence of L. infantum isolates from the focus using a natural transmission model. Hamsters were infected by intraperitoneal inoculation (IP) or by bites of sand flies experimentally infected with L. infantum isolates obtained from P. perniciosus collected in the outbreak area (IPER/ES/2012/BOS1FL1 and IPER/ES/2012/POL2FL6) and a well characterized L. infantum strain JPCM5 (MCAN/ES/98/LLM-877). Hamster infections were monitored by clinical examination, serology, culture, parasite burden, Giemsa-stained imprints, PCR, histopathology and xenodiagnostic studies. Establishment of infection of L. infantum was achieved with the JPCM5 strain and outbreak isolates by both P. perniciosus infective bites or IP route. However, high virulence of BOS1FL1 and POL2FL6 isolates was highlighted by the clinical outcome of disease, high parasite detection in spleen and liver, high parasitic loads and positivity of Leishmania serology. Transmission by bite of POL2FL6 infected flies generated a slower progression of clinical disease than IP infection, but both groups were infective to P. perniciosus by xenodiagnosis at 2 months post-infection. Conversely, hamsters inoculated with JPCM5 were not infective to sand flies. Histopathology studies confirmed the wide spread of POL2FL6 parasites to several organs. A visceral leishmaniasis model that mimics the natural transmission in nature allowed us to highlight the high virulence of isolates that are circulating in the focus. These findings contribute to a better understanding of the outbreak epidemiology.

Urokinase receptor-deficient mice mount an innate immune response to and clarify respiratory viruses as efficiently as wild-type mice.

The plasminogen activator receptor (uPAR) is required for lung infiltration by innate immune cells in respiratory bacterial infections. In order to verify if this held true for respiratory viruses, wild type (WT) and uPAR knockout (uPAR(-/-)) mice were inoculated intranasally with the human respiratory syncytial virus (HRSV) and the influenza A virus. At several days post-infection (dpi), viral titers in the lungs were determined while cell infiltrates in the bronchoalveolar lavage (BAL) were analyzed by flow cytometry. In the case of influenza A, body weight loss and mortality were also monitored. Only minor differences were observed between infected WT and uPAR(-/-) mice, primarily in influenza virus replication and pathology. These results indicate that uPAR does not play a major role in limiting virus replication or in orchestrating the innate immune response against HRSV or influenza infections in mice. This suggests that there are fundamental differences in the immune control of the viral infections studied here and those caused by bacteria.

Aß increases neural stem cell activity in senescence-accelerated SAMP8 mice.

Neurogenesis persists in the adult brain as a form of plasticity due to the existence of neural stem cells (NSCs). Alterations in neurogenesis have been found in transgenic Alzheimer's disease (AD) mouse models, but NSC activity and neurogenesis in sporadic AD models remains to be examined. We herein describe a remarkable increase in NSC proliferation in the forebrain of SAMP8, a non-transgenic mouse strain that recapitulates the transition from healthy aging to AD. The increase in proliferation is transient, precedes AD-like symptoms such as amyloid beta 1-42 [Aß(1-42)] increase or gliosis, and is followed by a steep decline at later stages. Interestingly, in vitro studies indicate that secreted Aß(1-42) and PI3K signaling may account for the early boost in NSC proliferation. Our results highlight the role of soluble Aß(1-42) peptide and PI3K in the autocrine regulation of NSCs, and further suggest that over-proliferation of NSCs before the appearance of AD pathology may underlie neurogenic failure during the age-related progression of the disease. These findings have implications for therapeutic approaches based on neurogenesis in AD.

Complement regulatory protein Crry/p65 costimulation expands natural treg cells with enhanced suppressive properties in proteoglycan-induced arthritis.

OBJECTIVE: To investigate the costimulatory role of Crry/p65 (Crry), a membrane complement regulatory protein, on the expansion and function of natural Treg cells and their ability to ameliorate proteoglycan-induced arthritis (PGIA), an animal model of inflammatory arthritis in which the role of natural Treg cells is not well established. METHODS: CD4+CD25+ natural Treg cells from BALB/c mice were activated in vitro and costimulated by Crry. The expanded cells were phenotypically characterized, and their suppressive effect on T cell proliferation was assayed in vitro. The potential prophylactic and therapeutic effects of this population versus those of natural Treg cells in PGIA were studied. The clinical score, histology, the antigen-specific isotype antibody pattern, in vitro T cell responses, and the presence of Treg cells in the paws were studied. RESULTS: Crry costimulation enhanced the in vitro expansion of natural Treg cells while maintaining their phenotypic and suppressive properties. Crry-expanded Treg cells had stronger suppressive properties in vivo and a longer ameliorating effect in the PGIA model than did natural Treg cells. Crry-expanded Treg cells suppressed T cell- and B cell-dependent responses in PGIA, changing the pathogenic antibody isotype pattern and decreasing antigen-dependent secretion of cytokines, including interferon-γ, interleukin-12 (IL-12), and IL-17. Increased FoxP3 expression was detected in the paws of mice transferred with Crry-expanded Treg cells. CONCLUSION: Crry-mediated costimulation facilitates in vitro expansion of natural Treg cells while maintaining their suppressive properties in vitro and in vivo in the PGIA model. These results highlight the potential of the complement regulatory protein Crry to costimulate and expand natural Treg cells capable of suppressing disease in an animal model of chronic inflammatory arthritis.

Beta-interferon unbalances the peripheral T cell proinflammatory response in experimental autoimmune encephalomyelitis.

Interferon beta (IFNbeta) is a widespread therapy for multiple sclerosis (MS). We have analyzed some critical features of the T cell activation process in lymph nodes after IFNbeta treatment of experimental autoimmune encephalomyelitis (EAE) in SJL mice. Prevention of clinical signs and drastic reduction of perivascular infiltrates in the central nervous system (CNS) were accompanied by alterations in nuclear DNA binding activity levels of NFkappaB and Stat6 transcription factors in lymph node cells (LNC). A decrease of active NFkappaB subunits in treated animals correlated with lower levels of the cytoplasmic phosphorylated form of IkappaBalpha. Results also showed that nuclear DNA binding activity of Stat6 was increased by IFNbeta treatment, as were the cytoplasmic levels of phosphorilated Stat6 (P-Stat6). These high levels of P-Stat6 in IFNbeta-treated animals were accompanied by an increase of IL-4 expression levels measured by real time PCR. In vitro experiments with the IL-4 producing clone D10.G4.1 indicates that the IFNbeta-mediated IL-4 induction is not an effect exclusive to MBP-reactive cells, and suggest that it could be mediated by mRNA stability enlargement. On the other hand, IFNbeta treatment of EAE produced no significant changes in peripheral IFNgamma expression and a striking decrease of IL-17. These findings suggest that the inhibition of NFkappaB activity, the increase of IL-4 expression and its signaling transduction, and the decrease of IL-17 may cooperate to some of the antiinflammatory effects of IFNbeta on EAE.

Multitissue array review: a chronological description of tissue array techniques, applications and procedures.

We performed a systematic search for literature dealing with tissue microarray technology. During the last two decades, these procedures have developed into a powerful tool for the high-throughput analysis of tissue specimens. This technology offers the following advantages: amplification of a scarce resource, experimental uniformity (tissue of multiple patients are treated in an identical manner), decreased assay volume, preservation of original blocks, amenability to a wide range of techniques and evaluation of tissue from multiple patients on the same slide. Depending on the shape of the tissue sample and the method used to obtain it, multitissue array techniques may be classified into two different groups: rod-shaped tissue techniques and core tissue techniques. These techniques have been used for quality control, diagnosis, and teaching and screening purposes. Some technical aspects should be considered when deciding which technique should be used: the number, size and origin of tissue samples; the quality of paraffin wax, the distance between samples and the depth in the receptor block; antigenicity preservation and block sectioning. This review shows different techniques, the use of which is dependent on the requirements of the arrays and the technical possibilities.

The course of infections and pathology in immunomodulated NOD/LtSz-SCID mice inoculated with Plasmodium falciparum laboratory lines and clinical isolates.

Human chimeras are potentially invaluable models for hemoprotozoan parasites such as Plasmodium falciparum. The work presented assesses the susceptibility of immunomodulated NOD/LtSz-SCID mice to genetically distinct P. falciparum parasites. To this end, mice grafted with human erythrocytes were inoculated with two P. falciparum laboratory lines, 3D7 and Dd2 and four clinical isolates, ISCIII-230, ISCIII-231, ISCIII-381 and ISCIII-399. The results showed that, without a previous period of parasite adaptation, 100% of the inoculated mice developed an infection, generally self-limited, though some mice died. The parasitemias ranged from 0.05 to 8% and lasted an average of 19 days (15-26 days) depending on the line or isolate studied. Sexual forms of different maturity, stage II-IV and mature gametocytes were observed in the peripheral blood of mice in 22, 50, 25, 72 and 80% of the mice infected with Dd2, ISCIII-399, ISCIII-230, ISCIII-231 and ISCIII-381 isolates, respectively. The study of the clinical symptoms, the haematological parameters and the histopathological changes in the infected mice showed that most of the malaria features were present in the infected mice except that the sequestration of infected erythrocytes was absent or at most a minor phenomenon, as also indicated by the presence of mature forms of the parasites in the peripheral blood. This study shows that the human chimeras allow the complete asexual and sexual erythrocytic cycle of different P. falciparum lines and clinical isolates to be observed in vivo. It opens a new way to investigate any parasite population in terms of infectivity, transmission, and drug resistance.

Experimental infection of immunomodulated NOD/LtSz-SCID mice as a new model for Plasmodium falciparum erythrocytic stages.

The main objective of this study was to determine whether a chemical immunomodulation protocol could reduce the resistance of NOD/LtSz-SCID mice to Plasmodium falciparum infection and provide an improved mouse model for screening the antimalarial activity of new compounds. This model was compared with the presently used immunodeficient Beige/Nude/Xid (BNX) mouse model, using the same protocol, in terms of percentage of infected mice, parasite development, leukocyte response and phagocytosis of P. falciparum infected cells in various organs. Our results show that the combination of the chemical immune modulation protocol with the genetic background of NOD/LtSz-SCID mice results in the development of long-lasting P. falciparum infection in a high percentage of mice. A comparison of the results obtained in the histological study for both mouse models suggests that the higher rate of success in NOD/LtSz-SCID mice could be related to the reduced macrophage recruitment developed in different tissues to remove the parasite from blood.