Genetic diversity of bovine Picobirnavirus, Brazil.

Picobirnaviruses (PBVs) are emerging and opportunistic viruses with possible zoonotic potential. In this study, we present the detection, molecular characterization, and genotypic differentiation of PBVs from genogroup I in bovine stool samples from different Brazilian regions. A high proportion of PCR-positive samples (23.4%) was detected in a total of 77 analyzed. Nucleotide identity, alignment, and phylogenetic analyses revealed high diversity among the studied sequences. The results obtained indicate, for the first time, the circulation of bovine PBVs belonging to genogroup I in different Brazilian states, with heterogeneous phylogenetic-clustering profiles.

Mechanistic insights into functional characteristics of native crotamine.

The chemical composition of snake venoms is a complex mixture of proteins and peptides that can be pharmacologically active. Crotamine, a cell-penetrating peptide, has been described to have antimicrobial properties and it exerts its effects by interacting selectively with different structures, inducing changes in the ion flow pattern and cellular responses. However, its real therapeutic potential is not yet fully known. Bearing in mind that crotamine is a promising molecule in therapeutics, this study investigated the action of purified molecule in three aspects: I) antibacterial action on different species of clinical interest, II) the effect of two different concentrations of the molecule on platelet aggregation, and III) its effects on isolated mitochondria. Crotamine was purified to homogeneity in a single step procedure using Heparin Sepharose. The molecular mass of the purified enzyme was 4881.4 Da, as determined by mass spectrometry. To assess antibacterial action, changes in the parameters of bacterial oxidative stress were determined. The peptide showed antibacterial activity on Escherichia coli (MIC: 2.0 µg/µL), Staphylococcus aureus (MIC: 8-16 µg/µL) and methicillin-resistant Staphylococcus aureus (MIC: 4.0-8.0 µg/µL), inducing bacterial death by lipid peroxidation and oxidation of target proteins, determined by thiobarbituric acid reactive substances and sulfhydryl groups, respectively. Crotamine induced increased platelet aggregation (IPA) at the two concentrations analyzed (0.1 and 1.4 µg/µL) compared to ADP-induced aggregation of PRP. Mitochondrial respiratory parameters and organelle structure assays were used to elucidate the action of the compound in this organelle. The exposure of mitochondria to crotamine caused a decrease in oxidative phosphorylation and changes in mitochondrial permeability, without causing damage in the mitochondrial redox state. Together, these results support the hypothesis that, besides the antimicrobial potential, crotamine acts on different molecular targets, inducing platelet aggregation and mitochondrial dysfunction.

Aislamiento e identificación de células madre adultas a partir de la grasa infrapatelar de Hoffa / Isolation and identification of stem cells derived of infrapatelar fat pad

Introducción: La grasa infrapatelar de Hoffa es una inclusión infraarticular presente en la rodilla y existen evidencias que células multipotentes están presentes en tejido adiposo del humano adulto. Objetivo: El propósito de este trabajo es aislar e identificar células mesenquimales a partir de muestras de la grasa de Hoffa tomadas de pacientes a los que se les practicó cirugía de rodilla que expongan este tejido. Metodología: Una vez obtenido el consentimiento informado de un total de 12 pacientes, se obtuvo la muestra de tejido graso. La grasa fue disgregada incubándola en colagenasa en PBS a 37ºC y con agitación, luego se inactivó con suero fetal bovino, se separaron mediante centrifugación los adipocitos maduros de las células multipotentes y se obtuvieron las muestras para sembrar en medio Mesencult. Una porción de muestra fue utilizada para identificación mediante citometría de flujo y otra parte para ser coloreada con azul de metileno. Resultados y Discusión: Tanto en las tinciones con azul de metileno como en los cultivos celulares se han apreciado células con la morfología propia de una célula mesenquimal en forma de estrella y que fueron corroborados mediante citometría de flujo para los marcadores CD13, CD29, CD59 y CD105; igualmente hemos podido observar la efectividad del tratamiento enzimático. Conclusiones: En la actualidad nos encontramos consolidando los cultivos primarios para posteriormente diferenciarlos en líneas celulares específicas que puedan ser utilizadas en estudios de patologías, tales como obesidad, diabetes y trastornos articulares...

Introduction: The infrapatellar fat pad (sometimes know Hoffa's pad) is a soft tissue that lies beneath the patella (kneecap) separating it from the femoral condyle. There is evidence suggesting the presence of stem cells on adipose tissue in the adult human. Objective: The purpose of this work is to isolate and identify grown stem cells from Hoffa's fat samples obtained from patients undergoing surgeries exposing this tissue. Materials and Methods: After obtaining informed consent, the biopsy of Hoff's fat pad was obtained. The samples were incubated with collagenase and PBS a 37ºC and agitated, then it was inactivated with fetal bovine serum and centrifuged, washed twice with PBS, the pellet was resuspended and one part was cultivated on Mesencult medium the other part was used for flow cytometry and stained with methylene blue for morphologic analysis. Also before and after the enzyme digestion, the samples were added to 10% formaldehyde to evaluate the collagenase treatment. Results and analysis: The results show the effectiveness of the enzymatic treatment, the architecture of the adipose tissue was lost. The star shape stem cells morphology was appreciated with methylene blue in the cultures, it was corroborated by flow cytometry with CD13, CD29, CD59 and CD 105 markers. Conclusions: Primary cultures are consolidating, the next aim will be to obtain differentiated specific cell types that can be use in the study of obesity, diabetes and articular illnesses...