Identification of Sinorhizobium (Ensifer) medicae based on a specific genomic sequence unveiled by M13-PCR fingerprinting

A collection of nodule isolates from Medicago polymorpha obtained from southern and central Portugal was evaluated by M13-PCR fingerprinting and hierarchical cluster analysis. Several genomic clusters were obtained which, by 16S rRNA gene sequencing of selected representatives, were shown to be associated with particular taxonomic groups of rhizobia and other soil bacteria. The method provided a clear separation between rhizobia and co-isolated non-symbiotic soil contaminants. Ten M13-PCR groups were assigned to Sinorhizobium (Ensifer) medicae and included all isolates responsible for the formation of nitrogen-fixing nodules upon re-inoculation of M. polymorpha test-plants. In addition, enterobacterial repetitive intergenic consensus (ERIC)-PCR fingerprinting indicated a high genomic heterogeneity within the major M13-PCR clusters of S. medicae isolates. Based on nucleotide sequence data of an M13-PCR amplicon of ca. 1500 bp, observed only in S. medicae isolates and spanning locus Smed_3707 to Smed_3709 from the pSMED01 plasmid sequence of S. medicae WSM419 genome’s sequence, a pair of PCR primers was designed and used for direct PCR amplification of a 1399-bp sequence within this fragment. Additional in silico and in vitro experiments, as well as phylogenetic analysis, confirmed the specificity of this primer combination and therefore the reliability of this approach in the prompt identification of S. medicae isolates and their distinction from other soil bacteria. [Int Microbiol 2009; 12(4):215-225

Influence of migration on the thought process of individuals at ultra-high risk for psychosis

OBJECTIVE To assess the influence of migration on the psychopathological presentation of individuals at ultra-high risk for psychosis (UHR) in São Paulo, Brazil. METHODS This study is part of the Subclinical Symptoms and Prodromal Psychosis (SSAPP) project, a cohort study in São Paulo, Brazil, designed to follow individuals at UHR. After screening with the Prodromal Questionnaire (PQ) and a clinical interview, the Global Assessment of Functioning (GAF) was administered, a neuropsychological assessment was performed, sociodemographic and migration data were obtained. We then analyzed UHR individuals who had migration data to see if migration had any effect on their cognition and psychopathology. Chi-square tests were used for categorical variables, and Student's t test or analysis of variance (ANOVA) were used for nonparametric and parametric distributions, respectively. RESULTS The sample was composed of 42 at-risk subjects, of whom 5 had a migration history in the past two generations. Those with migration history showed significantly more formal thought disturbances (p = 0.012) and sleeping problems (p = 0.033) compared to those without. CONCLUSIONS Our data reinforce migration as a risk factor for psychosis in developing countries as well, and highlights the importance of studying the specific effect of this factor in UHR psychopathology

Development and Characterization of Aloe vera Mucilaginous-Based Hydrogels for Psoriasis Treatment

The Aloe vera (L.) Burman f. pulp extract (AE), obtained from the inner parts of Aloe vera leaves, is rich in polysaccharides, including glucomannans, acemannans, pectic compounds, cellulose, and hemicelluloses; acemannan and glucomannan are considered the two main components responsible for most of the plant’s therapeutical properties. Besides having anti-inflammatory activity, these polysaccharides accelerate wound healing and promote skin regeneration, thus they can be utilized in healing products. The objective of this study was to develop Aloe vera mucilaginous-based hydrogels for topical use in psoriasis treatment. The hydrogels were prepared with 80% w/w of A. vera mucilaginous gel, evaluating two distinct polymers as the gelling agent: 1% carbopol 940 (FC1 and FC2) or 2% hydroxyethylcellulose (FH3 and FH4). FC1, FC2, FH3 and FH4 were evaluated for their organoleptic characteristics, rheological properties, pH and glucomannan content. Polysaccharide fractions (PFs) were extracted from the AE and used as a group of chemical markers and characterized by infrared (IR) spectroscopy and 1H nuclear magnetic resonance (H NMR). The quantification of these markers in the raw material (AE) and in the hydrogels was carried out using spectrophotometric techniques in the UV-VIS region. The hydrogels-based hydroxyethylcellulose (FH3 and FH4) had glucomannan contents of 6.76 and 4.01 mg/g, respectively. Formulations with carbopol, FC1 and FC2, had glucomannan contents of 8.69 and 9.17 mg/g, respectively, an ideal pH for application on psoriasis, in addition to good spreadability and pseudoplastic and thixotropic behavior. Considering these results, hydrogel FC1 was evaluated for its keratolytic activity in a murine model of hyperkeratinization. For that, 0.5 mL of test formulations FC1 and FPC (0.05% clobetasol propionate cream) were topically applied to the proximal region of adult rats daily for 13 days. After euthanasia, approximately 2.5 cm of the proximal portion of each animal’s tail was cut and placed in 10% buffered formalin. Then, each tail fragment was processed and stained with hematoxylin and eosin (HE), and the results obtained from the histological sections indicated a 61% reduction in stratum corneum for animals treated with the A. vera hydrogel (FC1G) and 66% for animals treated with clobetasol propionate (PCG), compared to the group of animals that did not receive treatment (WTG). This study led to the conclusion that compared to the classic treatment (clobetasol propionate), the 80% A. vera hydrogel showed no significant difference, being effective in controlling hyperkeratinization

Imbalances in serum angiopoietin concentrations are early predictors of septic shock development in patients with post chemotherapy febrile neutropenia

Background: Febrile neutropenia carries a high risk of sepsis complications, and the identification of biomarkers capable to identify high risk patients is a great challenge. Angiopoietins (Ang -) are cytokines involved in the control microvascular permeability. It is accepted that Ang-1 expression maintains endothelial barrier integrity, and that Ang-2 acts as an antagonizing cytokine with barrier-disrupting functions in inflammatory situations. Ang-2 levels have been recently correlated with sepsis mortality in intensive care units. Methods: We prospectively evaluated concentrations of Ang-1 and Ang-2 at different time-points during febrile neutropenia, and explored the diagnostic accuracy of these mediators as potential predictors of poor outcome in this clinical setting before the development of sepsis complications. Results: Patients that evolved with septic shock (n = 10) presented higher levels of Ang-2 measured 48 hours after fever onset, and of the Ang-2/Ang-1 ratio at the time of fever onset compared to patients with non-complicated sepsis (n = 31). These levels correlated with sepsis severity scores. Conclusions: Our data suggest that imbalances in the concentrations of Ang-1 and Ang-2 are independent and early markers of the risk of developing septic shock and of sepsis mortality in febrile neutropenia, and larger studies are warranted to validate their clinical usefulness. Therapeutic strategies that manipulate this Ang-2/Ang-1 imbalance can potentially offer new and promising treatments for sepsis in febrile neutropenia

Constitutively active GSK3 beta as a means to bolster dendritic cell functionality in the face of tumor-mediated immune suppression

In patients with cancer, the functionality of Dendritic Cells (DC) is hampered by high levels of tumor-derived suppressive cytokines, which interfere with DC development and maturation. Poor DC development can limit the efficacy of immune checkpoint blockade and in vivo vaccination approaches. Interference in intracellular signaling cascades downstream from the receptors of major tumor-associated suppressive cytokines like IL-10 and IL-6, might improve DC development and activation, and thus enhance immunotherapy efficacy. We performed exploratory functional screens on arrays consisting of >1000 human kinase peptide substrates to identify pathways involved in DC development and its inhibition by IL-10 or IL-6. The resulting alterations in phosphorylation of the kinome substrate profile pointed to glycogen-synthase kinase-3 beta (GSK3 beta) as a pivotal kinase in both DC development and suppression. GSK3 beta inhibition blocked human DC differentiation in vitro, which was accompanied by decreased levels of IL-12p70 secretion, and a reduced capacity for T cell priming. More importantly, adenoviral transduction of monocytes with a constitutively active form of GSK3 beta induced resistance to the suppressive effects of IL-10 and melanoma-derived supernatants alike, resulting in improved DC development, accompanied by up-regulation of co-stimulatory markers, an increase in CD83 expression levels in mature DC, and diminished release of IL-10. Moreover, adenovirus-mediated intratumoral manipulation of this pathway in an in vivo melanoma model resulted in DC activation and recruitment, and in improved immune surveillance and tumor control. We propose the induction of constitutive GSK3 beta activity as a novel therapeutic means to bolster DC functionality in the tumor microenvironment.Peer reviewe