An in vivo humanized model to study homing and sequestration of Plasmodium falciparum transmission stages in the bone marrow.

Introduction: Recent evidence suggests that the bone marrow (BM) plays a key role in the diffusion of P. falciparum malaria by providing a "niche" for the maturation of the parasite gametocytes, responsible for human-to-mosquito transmission. Suitable humanized in vivo models to study the mechanisms of the interplay between the parasite and the human BM components are still missing. Methods: We report a novel experimental system based on the infusion of immature P. falciparum gametocytes into immunocompromised mice carrying chimeric ectopic ossicles whose stromal and bone compartments derive from human osteoprogenitor cells. Results: We demonstrate that immature gametocytes home within minutes to the ossicles and reach the extravascular regions, where they are retained in contact with different human BM stromal cell types. Discussion: Our model represents a powerful tool to study BM function and the interplay essential for parasite transmission in P. falciparum malaria and can be extended to study other infections in which the human BM plays a role.

Phagocytosis and activation of bone marrow-derived macrophages by Plasmodium falciparum gametocytes.

BACKGROUND: The innate immune response against various life cycle stages of the malaria parasite plays an important role in protection against the disease and regulation of its severity. Phagocytosis of asexual erythrocytic stages is well documented, but little and contrasting results are available about phagocytic clearance of sexual stages, the gametocytes, which are responsible for the transmission of the parasites from humans to mosquitoes. Similarly, activation of host macrophages by gametocytes has not yet been carefully addressed. METHODS: Phagocytosis of early or late Plasmodium falciparum gametocytes was evaluated through methanol fixed cytospin preparations of immortalized mouse C57Bl/6 bone marrow-derived macrophages treated for 2 h with P. falciparum and stained with Giemsa, and it was confirmed through a standardized bioluminescent method using the transgenic P. falciparum 3D7elo1-pfs16-CBG99 strain. Activation was evaluated by measuring nitric oxide or cytokine levels in the supernatants of immortalized mouse C57Bl/6 bone marrow-derived macrophages treated with early or late gametocytes. RESULTS: The results showed that murine bone marrow-derived macrophages can phagocytose both early and late gametocytes, but only the latter were able to induce the production of inflammatory mediators, specifically nitric oxide and the cytokines tumour necrosis factor and macrophage inflammatory protein 2. CONCLUSIONS: These results support the hypothesis that developing gametocytes interact in different ways with innate immune cells of the host. Moreover, the present study proposes that early and late gametocytes act differently as targets for innate immune responses.

Gametocytes of the Malaria Parasite Plasmodium falciparum Interact With and Stimulate Bone Marrow Mesenchymal Cells to Secrete Angiogenetic Factors.

The gametocytes of Plasmodium falciparum, responsible for the transmission of this malaria parasite from humans to mosquitoes, accumulate and mature preferentially in the human bone marrow. In the 10 day long sexual development of P. falciparum, the immature gametocytes reach and localize in the extravascular compartment of this organ, in contact with several bone marrow stroma cell types, prior to traversing the endothelial lining and re-entering in circulation at maturity. To investigate the host parasite interplay underlying this still obscure process, we developed an in vitro tridimensional co-culture system in a Matrigel scaffold with P. falciparum gametocytes and self-assembling spheroids of human bone marrow mesenchymal cells (hBM-MSCs). Here we show that this co-culture system sustains the full maturation of the gametocytes and that the immature, but not the mature, gametocytes adhere to hBM-MSCs via trypsin-sensitive parasite ligands exposed on the erythrocyte surface. Analysis of a time course of gametocytogenesis in the co-culture system revealed that gametocyte maturation is accompanied by the parasite induced stimulation of hBM-MSCs to secrete a panel of 14 cytokines and growth factors, 13 of which have been described to play a role in angiogenesis. Functional in vitro assays on human bone marrow endothelial cells showed that supernatants from the gametocyte mesenchymal cell co-culture system enhance ability of endothelial cells to form vascular tubes. These results altogether suggest that the interplay between immature gametocytes and hBM-MSCs may induce functional and structural alterations in the endothelial lining of the human bone marrow hosting the P. falciparum transmission stages.

Gut Mesenchymal Stromal Cells in Immunity.

Mesenchymal stromal cells (MSCs), first found in bone marrow (BM), are the structural architects of all organs, participating in most biological functions. MSCs possess tissue-specific signatures that allow their discrimination according to their origin and location. Among their multiple functions, MSCs closely interact with immune cells, orchestrating their activity to maintain overall homeostasis. The phenotype of tissue MSCs residing in the bowel overlaps with myofibroblasts, lining the bottom walls of intestinal crypts (pericryptal) or interspersed within intestinal submucosa (intercryptal). In Crohn's disease, intestinal MSCs are tightly stacked in a chronic inflammatory milieu, which causes their enforced expression of Class II major histocompatibility complex (MHC). The absence of Class II MHC is a hallmark for immune-modulator and tolerogenic properties of normal MSCs and, vice versa, the expression of HLA-DR is peculiar to antigen presenting cells, that is, immune-activator cells. Interferon gamma (IFNγ) is responsible for induction of Class II MHC expression on intestinal MSCs. The reversal of myofibroblasts/MSCs from an immune-modulator to an activator phenotype in Crohn's disease results in the formation of a fibrotic tube subverting the intestinal structure. Epithelial metaplastic areas in this context can progress to dysplasia and cancer.

CCp5A Protein from Toxoplasma gondii as a Serological Marker of Oocyst-driven Infections in Humans and Domestic Animals.

Considering that the current immunoassays are not able to distinguish the infective forms that cause Toxoplasma gondii infection, the present study was carried out to evaluate the reactivity of two recombinant proteins (CCp5A and OWP1) from oocyst/sporozoite, in order to differentiate infections occurring by ingestion of oocysts or tissue cysts. The reactivity of the recombinant proteins was assessed against panels of serum samples from animals (chickens, pigs, and mice) that were naturally or experimentally infected by different infective stages of the parasite. Also, we tested sera from humans who have been infected by oocysts during a well-characterized toxoplasmosis outbreak, as well as sera from pregnant women tested IgM(+)/IgG(+) for T. gondii, which source of infection was unknown. Only the sporozoite-specific CCp5A protein was able to differentiate the parasite stage that infected chickens, pigs and mice, with specific reactivity for oocyst-infected animals. Furthermore, the CCp5A showed preferential reactivity for recent infection by oocyst/sporozoite in pigs and mice. In humans, CCp5A showed higher reactivity with serum samples from the outbreak, compared with serum from pregnant women. Altogether, these findings demonstrate the usefulness of the CCp5A protein as a new tool to identify the parasite stage of T. gondii infection, allowing its application for diagnosis and epidemiological investigations in animals and humans. The identification of parasite infective stage can help to design effective strategies to minimize severe complications in immunocompromised people and, particularly, in pregnant women to prevent congenital infection.

Sam68 regulates translation of target mRNAs in male germ cells, necessary for mouse spermatogenesis.

Sam68 is a KH-type RNA-binding protein involved in several steps of RNA metabolism with potential implications in cell differentiation and cancer. However, its physiological roles are still poorly understood. Herein, we show that Sam68(-/-) male mice are infertile and display several defects in spermatogenesis, demonstrating an essential role for Sam68 in male fertility. Sam68(-/-) mice produce few spermatozoa, which display dramatic motility defects and are unable to fertilize eggs. Expression of a subset of messenger mRNAs (mRNAs) is affected in the testis of knockout mice. Interestingly, Sam68 is associated with polyadenylated mRNAs in the cytoplasm during the meiotic divisions and in round spermatids, when it interacts with the translational machinery. We show that Sam68 is required for polysomal recruitment of specific mRNAs and for accumulation of the corresponding proteins in germ cells and in a heterologous system. These observations demonstrate a novel role for Sam68 in mRNA translation and highlight its essential requirement for the development of a functional male gamete.

Riesgo infantil por exposición a carcinógenos en un área de alto tránsito vehicular / Child risk exposure to carcinogens in a high traffic area

La contaminación del aire es una amenaza aguda, acumulativa y crónica para la salud y el ambiente. Las personas están expuestas a contaminantes de aire en exteriores e interiores. Las exposiciones a los mismos puede generar o agravar afecciones respiratorias, cardíacas y otras. La presente experiencia nace como la necesidad de conocer la calidad del aire en áreas de alto tránsito vehicular, con potencial riesgo para la salud de la población infantil. Experiencia que se desarrolla desde la gestión de la Dirección de Salud Ambiental, con una visión epidemiológica, con la participación de Centros de Investigación-Universidades Nacionales y recursos municipales.Sobresale en esta investigación la caracterización de escenarios de riesgo por los valores alto de benceno; con su correspondiente índice de peligro, según peso corporal del niño. También por el alto porcentaje de emisiones vehicular de material particulado. Se ha detectado la presencia en el 21 % de las muestras benzo[a]pireno, asociados con las partículas finas y se ha encontrado mayor valor de mutagenicidad (ensayo de Ames), en una de las zonas bajo estudio. Detección de tóxicos críticosque estarían presentes, durante la ventana de vulnerabilidad crítica del desarrollo del niño, llevando a intensificar las medidas correctivas y los estudios en la población vulnerable.