Phages are unrecognized players in the ecology of the oral pathogen Porphyromonas gingivalis.

BACKGROUND: Porphyromonas gingivalis (hereafter "Pg") is an oral pathogen that has been hypothesized to act as a keystone driver of inflammation and periodontal disease. Although Pg is most readily recovered from individuals with actively progressing periodontal disease, healthy individuals and those with stable non-progressing disease are also colonized by Pg. Insights into the factors shaping the striking strain-level variation in Pg, and its variable associations with disease, are needed to achieve a more mechanistic understanding of periodontal disease and its progression. One of the key forces often shaping strain-level diversity in microbial communities is infection of bacteria by their viral (phage) predators and symbionts. Surprisingly, although Pg has been the subject of study for over 40 years, essentially nothing is known of its phages, and the prevailing paradigm is that phages are not important in the ecology of Pg. RESULTS: Here we systematically addressed the question of whether Pg are infected by phages-and we found that they are. We found that prophages are common in Pg, they are genomically diverse, and they encode genes that have the potential to alter Pg physiology and interactions. We found that phages represent unrecognized targets of the prevalent CRISPR-Cas defense systems in Pg, and that Pg strains encode numerous additional mechanistically diverse candidate anti-phage defense systems. We also found that phages and candidate anti-phage defense system elements together are major contributors to strain-level diversity and the species pangenome of this oral pathogen. Finally, we demonstrate that prophages harbored by a model Pg strain are active in culture, producing extracellular viral particles in broth cultures. CONCLUSION: This work definitively establishes that phages are a major unrecognized force shaping the ecology and intra-species strain-level diversity of the well-studied oral pathogen Pg. The foundational phage sequence datasets and model systems that we establish here add to the rich context of all that is already known about Pg, and point to numerous avenues of future inquiry that promise to shed new light on fundamental features of phage impacts on human health and disease broadly. Video Abstract.

Endothelial mechanisms for inactivation of inflammation-induced hyperpermeability.

Microvascular hyperpermeability is a hallmark of inflammation. Many negative effects of hyperpermeability are due to its persistence beyond what is required for preserving organ function. Therefore, we propose that targeted therapeutic approaches focusing on mechanisms that terminate hyperpermeability would avoid the negative effects of prolonged hyperpermeability while retaining its short-term beneficial effects. We tested the hypothesis that inflammatory agonist signaling leads to hyperpermeability and initiates a delayed cascade of cAMP-dependent pathways that causes inactivation of hyperpermeability. We applied platelet-activating factor (PAF) and vascular endothelial growth factor (VEGF) to induce hyperpermeability. We used an Epac1 agonist to selectively stimulate exchange protein activated by cAMP (Epac1) and promote inactivation of hyperpermeability. Stimulation of Epac1 inactivated agonist-induced hyperpermeability in the mouse cremaster muscle and in human microvascular endothelial cells (HMVECs). PAF induced nitric oxide (NO) production and hyperpermeability within 1 min and NO-dependent increased cAMP concentration in about 15-20 min in HMVECs. PAF triggered phosphorylation of vasodilator-stimulated phosphoprotein (VASP) in a NO-dependent manner. Epac1 stimulation promoted cytosol-to-membrane eNOS translocation in HMVECs and in myocardial microvascular endothelial (MyEnd) cells from wild-type mice, but not in MyEnd cells from VASP knockout mice. We demonstrate that PAF and VEGF cause hyperpermeability and stimulate the cAMP/Epac1 pathway to inactivate agonist-induced endothelial/microvascular hyperpermeability. Inactivation involves VASP-assisted translocation of eNOS from the cytosol to the endothelial cell membrane. We demonstrate that hyperpermeability is a self-limiting process, whose timed inactivation is an intrinsic property of the microvascular endothelium that maintains vascular homeostasis in response to inflammatory conditions.NEW & NOTEWORTHY Termination of microvascular hyperpermeability has been so far accepted to be a passive result of the removal of the applied proinflammatory agonists. We provide in vivo and in vitro evidence that 1) inactivation of hyperpermeability is an actively regulated process, 2) proinflammatory agonists (PAF and VEGF) stimulate microvascular hyperpermeability and initiate endothelial mechanisms that terminate hyperpermeability, and 3) eNOS location-translocation is critical in the activation-inactivation cascade of endothelial hyperpermeability.

Effects of Oral Health Interventions in People with Oropharyngeal Dysphagia: A Systematic Review.

People with oropharyngeal dysphagia (OD) are at risk of developing aspiration pneumonia. However, there is no "best practice" for oral health interventions to improve swallowing-related outcomes, the incidence of aspiration pneumonia, and oral health in people with OD. Systematic literature searches were conducted for oral health interventions in OD in PubMed, Embase, CINAHL, and PsycINFO until July 2021. Original articles published in English and reporting pre- and post-intervention measurements were included. The methodology and reporting were guided by the PRISMA checklist. The methodological quality of the eight included studies was rated using the QualSyst critical appraisal tool. The oral health interventions in people with OD were diverse. This study shows little evidence that regular oral care and the free water protocol or oral disinfection reduced the incidence of aspiration pneumonia in people with OD. Oral cleaning, twice a day with an antibacterial toothpaste in combination with intraoral cleaning or the free water protocol, proved to be the most promising intervention to improve oral health. The effect of improved oral health status on swallowing-related outcomes could not be established. Increasing awareness of the importance of oral health and implementing practical oral care guidelines for people involved in the daily care of people with OD are recommended.

Draft Genome Sequence of Schaalia odontolytica Strain ORNL0103, a Basibiont of "Candidatus Saccharibacteria" HMT352.

Here, we report the draft, nearly complete genome sequence of the human oral actinobacterium Schaalia odontolytica strain ORNL0103, which was isolated in association with "Candidatus Saccharibacteria" HMT352 strain ORNL0105. The genome was sequenced using a combination of Pacific Biosciences and Illumina platforms and encodes 1,948 proteins and 60 RNAs.

TNF-α-activated eNOS signaling increases leukocyte adhesion through the S-nitrosylation pathway.

Nitric oxide (NO) is a key factor in inflammation. Endothelial nitric oxide synthase (eNOS), whose activity increases after stimulation with proinflammatory cytokines, produces NO in endothelium. NO activates two pathways: 1) soluble guanylate cyclase-protein kinase G and 2) S-nitrosylation (NO-induced modification of free-thiol cysteines in proteins). S-nitrosylation affects phosphorylation, localization, and protein interactions. NO is classically described as a negative regulator of leukocyte adhesion to endothelial cells. However, agonists activating NO production induce a fast leukocyte adhesion, which suggests that NO might positively regulate leukocyte adhesion. We tested the hypothesis that eNOS-induced NO promotes leukocyte adhesion through the S-nitrosylation pathway. We stimulated leukocyte adhesion to endothelium in vitro and in vivo using tumor necrosis factor-α (TNF-α) as proinflammatory agonist. ICAM-1 changes were evaluated by immunofluorescence, subcellular fractionation, immunoprecipitation, and fluorescence recovery after photobleaching (FRAP). Protein kinase Cζ (PKCζ) activity and S-nitrosylation were evaluated by Western blot analysis and biotin switch method, respectively. TNF-α, at short times of stimulation, activated the eNOS S-nitrosylation pathway and caused leukocyte adhesion to endothelial cells in vivo and in vitro. TNF-α-induced NO led to changes in ICAM-1 at the cell surface, which are characteristic of clustering. TNF-α-induced NO also produced S-nitrosylation and phosphorylation of PKCζ, association of PKCζ with ICAM-1, and ICAM-1 phosphorylation. The inhibition of PKCζ blocked leukocyte adhesion induced by TNF-α. Mass spectrometry analysis of purified PKCζ identified cysteine 503 as the only S-nitrosylated residue in the kinase domain of the protein. Our results reveal a new eNOS S-nitrosylation-dependent mechanism that induces leukocyte adhesion and suggests that S-nitrosylation of PKCζ may be an important regulatory step in early leukocyte adhesion in inflammation.NEW & NOTEWORTHY Contrary to the well-established inhibitory role of NO in leukocyte adhesion, we demonstrate a positive role of nitric oxide in this process. We demonstrate that NO induced by eNOS after TNF-α treatment induces early leukocyte adhesion activating the S-nitrosylation pathway. Our data suggest that PKCζ S-nitrosylation may be a key step in this process.

Degradation of host translational machinery drives tRNA acquisition in viruses.

Viruses are traditionally thought to be under selective pressure to maintain compact genomes and thus depend on host cell translational machinery for reproduction. However, some viruses encode abundant tRNA and other translation-related genes, potentially optimizing for codon usage differences between phage and host. Here, we systematically interrogate selective advantages that carrying 18 tRNAs may convey to a T4-like Vibriophage. Host DNA and RNA degrade upon infection, including host tRNAs, which are replaced by those of the phage. These tRNAs are expressed at levels slightly better adapted to phage codon usage, especially that of late genes. The phage is unlikely to randomly acquire as diverse an array of tRNAs as observed (p = 0.0017). Together, our results support that the main driver behind phage tRNA acquisition is pressure to sustain translation as host machinery degrades, a process resulting in a dynamically adapted codon usage strategy during the course of infection.

Role of NO and S-nitrosylation in the Expression of Endothelial Adhesion Proteins That Regulate Leukocyte and Tumor Cell Adhesion.

Leukocyte recruitment is one of the most important cellular responses to tissue damage. Leukocyte extravasation is exquisitely regulated by mechanisms of selective leukocyte-endothelium recognition through adhesion proteins in the endothelial cell surface that recognize specific integrins in the activated leukocytes. A similar mechanism is used by tumor cells during metastasis to extravasate and form a secondary tumor. Nitric oxide (NO) has been classically described as an anti-inflammatory molecule that inhibits leukocyte adhesion. However, the evidence available shows also a positive role of NO in leukocyte adhesion. These apparent discrepancies might be explained by the different NO concentrations reached during the inflammatory response, which are highly modulated by the expression of different nitric oxide synthases, along the inflammatory response and by changes in their subcellular locations.

Subregistro de los suicidios en el Boletín Estadístico de Defunción Judicial en Málaga / Notification of suicides in the Statistical Bulletin of Judicial Death in Malaga (Spain)

El objetivo de este trabajo es confirmar, a partir de las Defunciones con Intervención Judicial (DIJ), si la causa de defunción fue un suicidio, según la información de la familia e identificar el porcentaje de los casos señalados como suicidio en el Boletín Estadístico de Defunción Judicial (BEDJ). Para ello se seleccionaron las DIJ de la base de datos del Parque Cementerio de Málaga en el año 2017. Se recabó información de las familias de las personas fallecidas a través del Gabinete Psicológico del Cementerio para confirmar o descartar el suicidio, y posteriormente se consultó el BEDJ para comprobar si los casos estaban identificados como suicidios. A través de los familiares se confirmaron 65 suicidios, de los que solo un 27% están identificados así en la sección correspondiente del BEDJ, documento que sirve de fuente de información para las estadísticas oficiales de suicidios. Se concluye que la familia puede ofrecer información complementaria que ayudaría a mejorar las estadísticas de suicidios

The objective of this study is to confirm from Deaths with Judicial Intervention (DIJ), whether cause of death was suicide, through family information and to establish the percentage of those identified as such in the Statistical Death Bulletin of Judicial Court (BEDJ). For this purpose, the DIJ were selected from the database of the Cementary Park of Málaga in 2017. Information was collected from the family of the deceased, through the Cementary's Psychological Cabinet to confirm or rule out suicide and the BEDJ was subsequently consulted to confirm whether these cases were identified as such. Relatives confirmed 65 suicides, of which only 27% are identified as such in the relevant section of the BEDJ, a document that serves as a source of information for official suicide statistics. From this study we concluded that the family can offer complementary information that would help improve suicide statistics

[Notification of suicides in the Statistical Bulletin of Judicial Death in Malaga (Spain)]. / Subregistro de los suicidios en el Boletín Estadístico de Defunción Judicial en Málaga.

The objective of this study is to confirm from Deaths with Judicial Intervention (DIJ), whether cause of death was suicide, through family information and to establish the percentage of those identified as such in the Statistical Death Bulletin of Judicial Court (BEDJ). For this purpose, the DIJ were selected from the database of the Cementary Park of Málaga in 2017. Information was collected from the family of the deceased, through the Cementary's Psychological Cabinet to confirm or rule out suicide and the BEDJ was subsequently consulted to confirm whether these cases were identified as such. Relatives confirmed 65 suicides, of which only 27% are identified as such in the relevant section of the BEDJ, a document that serves as a source of information for official suicide statistics. From this study we concluded that the family can offer complementary information that would help improve suicide statistics.

Significados de madres y padres mexicanos del mismo género en torno a la crianza / Meanings of Mexican same-sex parents around parenting

Este estudio muestra los significados expresados por tres parejas del mismo género (cuatro mujeres y dos hombres) sobre la crianza de sus hijas e hijos, quienes tenían entre uno y tres años de edad. Desde un enfoque hermenéutico-interpretativo, analizamos entrevistas narrativas, resultando en las siguientes categorías: Importancia de verbalizar y explicar; Reglas, límites y consecuencias; Enseñanza y aprendizaje de valores y habilidades sociales; Mantener una relación cercana; Rasgos/atributos personales valorados (en las y los niños). Encontramos diversas funciones asumidas en su labor como educadores, que se vincularon de forma dinámica con lo que esperaban y valoraban en cuanto a aprendizaje y desarrollo. Fue importante para ellos/as complacer, dar gusto y satisfacer las necesidades de sus niñas y niños, sin que esto significara descuidar una alimentación saludable, establecer y mantener reglas y rutinas, así como enseñar habilidades acordes a su edad. Un significado compartido giró en torno al lenguaje y comunicación, pues para los adultos era importante explicar: desde rutinas, hasta verbalizar sentimientos, o hacer explícito su tipo de familia (tener dos padres o dos madres); siempre considerando lo que puede comprenderse a esa edad. También fue importante para los adultos que sus hijos e hijas fueran alegres y sintieran su amor, manteniendo una relación cercana: cálida, lúdica y afectuosa. Concluimos que estos padres gais y madres lesbianas significaron a la crianza como una tarea amorosa, ardua, permanente y dirigida a formar sujetos con derechos, con quienes se podía negociar y llegar a acuerdos, aun siendo niños y niñas

This study shows the meanings expressed by three Mexican same-gender couples (four women and two men) about the upbringing of their daughters and sons, who were aged between one and three years. From a hermeneutic-interpretative approach, we analysed narrative interviews, resulting in the following categories: Importance of verbalizing and expressing; Rules, limits and consequences; Teaching and learning values and social skills; Maintain a warm relationship; Personal traits/attributes valued (in children). We found that fathers and mothers played different roles in their work as educators, which were linked dynamically with what they expect and value in terms of learning and development. It was important for them to please, to indulge and to satisfy their children's needs, without this neglecting a healthy diet, establishing and maintaining rules and routines, as well as teaching values and skills according to their age. A shared meaning revolved around language and communication: for adults it was important explaining routines, verbalizing feelings, or making explicit their type of family (having two fathers or two mothers); always considering what can be understood at that age. It was also important for our participants that their sons and daughters felt happy and loved through a close, warm, playful and affectionate relationship. We conclude that these gay parents and lesbian mothers meant the upbringing as a loving, arduous and permanent task, committed to forming subjects with rights, with whom they could negotiate and reach agreements

Plasmid pSfr64a and the symbiotic plasmid pSfr64b of Sinorhizobium fredii GR64 control each other's conjugative transfer through quorum-sensing elements.

Rhizobia are nitrogen-fixing symbionts of plants. Their genomes frequently contain large plasmids, some of which are able to perform conjugative transfer. Plasmid pSfr64a from Sinorhizobium fredii GR64 is a conjugative plasmid, whose transfer is regulated by quorum sensing genes encoded by itself (traR64a, traI64a), in the symbiotic plasmid pSfr64b (traR64b, traI64b), and in the chromosome (ngrI). Also, transfer of pSfr64b requires quorum sensing elements encoded in this plasmid (traR64b, traI64b), in pSfr64a (traR64a), and in the chromosome (ngrI). These results demonstrate that pSfr64a and the symbiotic plasmid depend on each other for conjugative transfer. Plasmid pSfr64a from S. fredii GR64 is unable to transfer from the genomic background of Rhizobium etli CFN42. Our results show that the relaxase of pRet42a is able to process the oriT of pSfr64a, and viceversa, underlining their functional similarity and suggesting that in addition to the external signals, the "cytoplasmic environment" may pose a barrier to plasmid dissemination, even if the plasmids are functional in other aspects.

Allopurinol Prevents the Lipogenic Response Induced by an Acute Oral Fructose Challenge in Short-Term Fructose Fed Rats.

We investigated whether short term high fructose intake may induce early hepatic dysfunction in rats and to test whether allopurinol treatment may have beneficial effects. Twenty male Sprague-Dawley rats received 20% fructose in drinking water (10 treated with allopurinol and 10 received vehicle) and 10 control rats received tap water. After 14 days, the hepatic response to an acute fructose load was evaluated, and in fasted animals, respirometry studies in freshly isolated mitochondria were performed. In fasting rats, we did not find differences in systemic or hepatic uric acid and triglyceride concentrations among the groups, but mitochondrial respiratory control rate was significantly decreased by high fructose feeding and correlated with a reduced expression of Complex I, as well as decreased aconitase-2 activity. On the other hand, in fructose fed rats, an acute fructose load increased systemic and hepatic uric acid, triglycerides and oxidative stress. Fructose feeding was also associated with fructokinase and xanthine oxidase overexpression and increased liver de novo lipogenesis program (fatty acid synthase (FAS) and cell death-inducing DFFA-like effector C (CIDEC) overexpression, ATP citrate lyase (ACL) and acetyl coA carboxylase (ACC) overactivity and decreased AMP-activated protein kinase (AMPk) and endothelial nitric oxide synthase (eNOS) activation). Allopurinol treatment prevented hepatic and systemic alterations. These data suggest that early treatment with xanthine oxidase inhibitors might provide a therapeutic advantage by delaying or even halting the progression of non-alcoholic fatty liver disease (NAFLD).

Cuantificación de hormonas 17β - estradiol y progesterona en el caracol rosa Lobatus gigas (Mollusca, Gastropoda)

La cuantificación de las hormonas esteroides en moluscos se lleva a cabo con diferentes técnicas utilizando la hemolinfa o gónadas. Lobatus gigas es un gasterópodo de interés comercial en el Caribe, indexado en CITES como una especie protegida. Los estudios hormonales de esta especie aún no están disponibles. Por lo tanto, el objetivo de este estudio fue determinar la presencia de las hormonas esteroides 17ß - estradiol y progesterona en L. gigas a través de un método no invasivo y comparar dos técnicas para su cuantificación. Cada dos meses en un período de un año, se recolectaron las heces de diez organismos en Xel-Há, Quintana Roo, México. Las muestras se analizaron con cromatografía líquida de alta resolución y EIA. Los valores más altos de ambas hormonas se presentaron de marzo a septiembre y disminuyeron en noviembre y enero. La comparación de las concentraciones obtenidas con HPLC y EIA, mostró que los resultados son similares para el 17β-estradiol (Passing - Bablock r = 0.673; -0.17 ng / ml). En contraste, los resultados de la progesterona con ambas técnicas no mostraron ajuste (Passing - Bablock r = 0.389; -1.43 ng / ml). Nuestros resultados sugieren que la técnica de EIA es adecuada para el estudio de hormonas en esta especie. El conocimiento generado permitirá monitorear y seleccionar organismos reproductores que se encuentren acondicionados en laboratorios y de esta manera no incidir en la recolecta de masas ovígeras silvestres.

Quantification of steroid hormones in molluscs is performed with different techniques, using the hemolymph or gonads. Lobatus gigas is a Caribbean gastropod of commercial interest indexed in CITES as a protected species. Hormonal studies of this species are as yet unavailable. The objective of this study is to determine the presence of the steroid hormones 17ß - estradiol and progesterone in L. gigas using a non-invasive method, and to compare two techniques for their quantification. Every two months over the course of one year, the feces of ten organisms were collected in Xel-Ha park Quintana Roo, México. The samples were analyzed with High resolution liquid chromatography and Enzyme-linked immunosorbent assays. The values of both hormones were highest during the months of March to September then decreased during November and January. Comparison of the concentrations obtained with HPLC and EIA, presented similar results for 17β-estradiol (Passing - Bablock r = 0.673; mean differences -0.17 ng / ml). In contrast, the progesterone results with both techniques showed no adjustment (Passing - Bablock r = 0.389; mean differences -1.43 ng / ml). Our results suggest that the enzyme-linked immunosorbent assay is suitable for the study of hormones in L. gigas. The knowledge generated will allow the monitoring and selection of breeding organisms that are conditioned in laboratories and thus will not affect the collection of wild egg masses.

Interleukin-8 Secreted by Glioblastoma Cells Induces Microvascular Hyperpermeability Through NO Signaling Involving S-Nitrosylation of VE-Cadherin and p120 in Endothelial Cells.

Glioblastoma is a highly aggressive brain tumor, characterized by the formation of dysfunctional blood vessels and a permeable endothelial barrier. S-nitrosylation, a post-translational modification, has been identified as a regulator of endothelial function. In this work we explored whether S-nitrosylation induced by glioblastoma tumors regulates the endothelial function. As proof of concept, we observed that S-nitrosylation is present in the tumoral microenvironment of glioblastoma in two different animal models. Subsequently, we measured S nitrosylation and microvascular permeability in EAhy296 endothelial cells and in cremaster muscle. In vitro, conditioned medium from the human glioblastoma cell line U87 activates endothelial nitric oxide synthase, causes VE-cadherin- S-nitrosylation and induces hyperpermeability. Blocking Interleukin-8 (IL-8) in the conditioned medium inhibited S-nitrosylation of VE-cadherin and hyperpermeability. Recombinant IL-8 increased endothelial permeability by activating eNOS, S-nitrosylation of VE-cadherin and p120, internalization of VE-cadherin and disassembly of adherens junctions. In vivo, IL-8 induced S-nitrosylation of VE-cadherin and p120 and conditioned medium from U87 cells caused hyperpermeability in the mouse cremaster muscle. We conclude that eNOS signaling induced by glioma cells-secreted IL-8 regulates endothelial barrier function in the context of glioblastoma involving S-nitrosylation of VE-cadherin and p120. Our results suggest that inhibiting S-nitrosylation may be an effective way to control and/or block damage to the endothelial barrier and prevent cancer progression.

Coagulation Factor Xa Promotes Solid Tumor Growth, Experimental Metastasis and Endothelial Cell Activation.

Hypercoagulable state is linked to cancer progression; however, the precise role of the coagulation cascade is poorly described. Herein, we examined the contribution of a hypercoagulative state through the administration of intravenous Coagulation Factor Xa (FXa), on the growth of solid human tumors and the experimental metastasis of the B16F10 melanoma in mouse models. FXa increased solid tumor volume and lung, liver, kidney and lymph node metastasis of tail-vein injected B16F10 cells. Concentrating on the metastasis model, upon coadministration of the anticoagulant Dalteparin, lung metastasis was significantly reduced, and no metastasis was observed in other organs. FXa did not directly alter proliferation, migration or invasion of cancer cells in vitro. Alternatively, FXa upon endothelial cells promoted cytoskeleton contraction, disrupted membrane VE-Cadherin pattern, heightened endothelial-hyperpermeability, increased inflammatory adhesion molecules and enhanced B16F10 adhesion under flow conditions. Microarray analysis of endothelial cells treated with FXa demonstrated elevated expression of inflammatory transcripts. Accordingly, FXa treatment increased immune cell infiltration in mouse lungs, an effect reduced by dalteparin. Taken together, our results suggest that FXa increases B16F10 metastasis via endothelial cell activation and enhanced cancer cell-endothelium adhesion advocating that the coagulation system is not merely a bystander in the process of cancer metastasis.

S-nitrosylation and its role in breast cancer angiogenesis and metastasis.

S-nitrosylation, the modification by nitric oxide of free sulfhydryl groups in cysteines, has become an important regulatory mechanism in carcinogenesis and metastasis. S-nitrosylation of targets in tumor cells contributes to metastasis regulating epithelial to mesenchymal transition, migration and invasion. In the tumor environment, the role of S-nitrosylation in endothelium has not been addressed; however, the evidence points out that S-nitrosylation of endothelial proteins may regulate angiogenesis, adhesion of tumor cells to the endothelium, intra and extravasation of tumor cells and contribute to metastasis.

Galectin-8 induces endothelial hyperpermeability through the eNOS pathway involving S-nitrosylation-mediated adherens junction disassembly.

The permeability of endothelial cells is regulated by the stability of the adherens junctions, which is highly sensitive to kinase-mediated phosphorylation and endothelial nitric oxide synthase (eNOS)-mediated S-nitrosylation of its protein components. Solid tumors can produce a variety of factors that stimulate these signaling pathways leading to endothelial cell hyperpermeability. This generates stromal conditions that facilitate tumoral growth and dissemination. Galectin-8 (Gal-8) is overexpressed in several carcinomas and has a variety of cellular effects that can contribute to tumor pathogenicity, including angiogenesis. Here we explored whether Gal-8 has also a role in endothelial permeability. We show that recombinant Gal-8 activates eNOS, induces S-nitrosylation of p120-catenin (p120) and dissociation of adherens junction, leading to hyperpermeability of the human endothelial cell line EAhy926. This pathway involves focal-adhesion kinase (FAK) activation downstream of eNOS as a requirement for eNOS-mediated p120 S-nitrosylation. This suggests a reciprocal, yet little understood, regulation of phosphorylation and S-nitrosylation events acting upon adherens junction permeability. In addition, glutathione S-transferase (GST)-Gal-8 pull-down experiments and function-blocking ß1-integrin antibodies point to ß1-integrins as cell surface components involved in Gal-8-induced hyperpermeability. Endogenous Gal-8 secreted from the breast cancer cell line MCF-7 has similar hyperpermeability and signaling effects. Furthermore, the mouse cremaster model system showed that Gal-8 also activates eNOS, induces S-nitrosylation of adherens junction components and is an effective hyperpermeability agent in vivo. These results add endothelial permeability regulation by S-nitrosylation as a new function of Gal-8 that can potentially contribute to the pathogenicity of tumors overexpressing this lectin.

Extracellular adenosine promotes cell migration/invasion of Glioblastoma Stem-like Cells through A3 Adenosine Receptor activation under hypoxia.

Glioblastoma (GBM) is the brain tumor with the worst prognosis composed of a cell subpopulation called Glioblastoma Stem-like Cells (GSCs) responsible for tumor recurrence mediated by cell invasion. GSCs persist in a hypoxic microenvironment which promotes extracellular adenosine production and activation of the A3 Adenosine Receptor (A3AR), therefore, the aim of this study was to determine the role of extracellular adenosine and A3AR on GSCs invasion under hypoxia. GSCs were obtained from a U87MG cell line and primary cultures of GBM patients, and then incubated under normoxia or hypoxia. Gene expression was evaluated by RNAseq, RT-qPCR, and western blot. Cell migration was measured by spreading and transwell boyden chamber assays; cell invasion was evaluated by Matrigel-coated transwell, ex vivo brain slice, and in vivo xenograft assays. The contribution of A3AR on cell migration/invasion was evaluated using the A3AR antagonist, MRS1220. Extracellular adenosine production was higher under hypoxia than normoxia, mainly by the catalytic action of the prostatic acid phosphatase (PAP), promoting cell migration/invasion in a HIF-2-dependent process. A3AR blockade decreased cell migration/invasion and the expression of Epithelial-Mesenchymal Transition markers. In conclusion, high levels of extracellular adenosine production enhance cell migration/invasion of GSCs, through HIF-2/PAP-dependent activation of A3AR under hypoxia.

First report of autochthonous canine leishmaniasis caused by Leishmania (L.) mexicana in Sinaloa, Mexico.

Leishmaniasis is a neglected tropical disease caused by different species of protozoan parasites of the genus Leishmania. Dogs have been proven as primary hosts of the parasite. Cases of cutaneous leishmaniasis in humans caused by Leishmania mexicana have been reported in Sinaloa; however, the vectors and hosts involved in the epidemiology of the parasite in northwestern Mexico are still unknown. Given the public health implications of this parasite's domestic hosts regarding the permanence and transmission of the disease to humans, the objective of the present study was to detect and determine the species of Leishmania that caused the first three cases of autochthonous canine leishmaniasis in the state of Sinaloa, Mexico. Three domestic dogs showing symptoms similar to canine leishmaniasis were identified, including chronic eye inflammation, corneal opacity, ocular exudate, emaciation and hyporexia. DNA was extracted from venous blood of the infected animals using a commercial kit. The internal transcribed spacer (ITS-1) of ribosomal DNA (rDNA) was amplified by specific primers for Leishmania from the extracted DNA, and the PCR products were digested with the restriction enzyme HaeIII. In addition, PCR products were subjected to automated sequencing. Molecular analysis showed that the infecting species was L. mexicana. This is the first report of autochthonous canine leishmaniasis caused by L. mexicana in Sinaloa, Mexico. Further studies are required to identify the species that serve as vectors and other wild and domestic hosts of the parasite, as well as to determine if there are more species of Leishmania circulating in Sinaloa.