Bioactivated gellan gum hydrogels affect cellular rearrangement and cell response in vascular co-culture and subcutaneous implant models.

Hydrogels are suitable soft tissue mimics and capable of creating pre-vascularized tissues, that are useful for in vitro tissue engineering and in vivo regenerative medicine. The polysaccharide gellan gum (GG) offers an intriguing matrix material but requires bioactivation in order to support cell attachment and transfer of biomechanical cues. Here, four versatile modifications were investigated: Purified NaGG; avidin-modified NaGG combined with biotinylated fibronectin (NaGG-avd); oxidized GG (GGox) covalently modified with carbohydrazide-modified gelatin (gelaCDH) or adipic hydrazide-modified gelatin (gelaADH). All materials were subjected to rheological analysis to assess their viscoelastic properties, using a time sweep for gelation analysis, and subsequent amplitude sweep of the formed hydrogels. The sweeps show that NaGG and NaGG-avd are rather brittle, while gelatin-based hydrogels are more elastic. The degradation of preformed hydrogels in cell culture medium was analyzed with an amplitude sweep and show that gelatin-containing hydrogels degrade more dramatically. A co-culture of GFP-tagged HUVEC and hASC was performed to induce vascular network formation in 3D for up to 14 days. Immunofluorescence staining of the αSMA+ network showed increased cell response to gelatin-GG networks, while the NaGG-based hydrogels did not allow for the elongation of cells. Preformed, 3D hydrogels disks were implanted to subcutaneous rat skin pockets to evaluate biological in vivo response. As visible from the hematoxylin and eosin-stained tissue slices, all materials are biocompatible, however gelatin-GG hydrogels produced a stronger host response. This work indicates, that besides the biochemical cues added to the GG hydrogels, also their viscoelasticity greatly influences the biological response.

Ultrastructural analysis of olfactory ensheathing cells derived from olfactory bulb and nerve of neonatal and juvenile rats.

Olfactory nerve derived and olfactory bulb derived olfactory ensheathing cells (OECs) have the ability to promote axonal regeneration and remyelination, both of which are essential in a successful cell transplant. Thus, morphological identification of OECs is a key aspect to develop an applicable cell therapy for injuries to the nervous system. However, there is no clear definition regarding which developmental stage or anatomical origin of OECs is more adequate for neural repair. In the present study, an ultrastructural comparison was made between OECs recovered from primary cultures of olfactory nerve and bulb in two developmental stages. The most notorious difference between cells obtained from olfactory nerve and bulb was the presence of indented nuclei in bulb derived OECs, suggesting a greater ability for possible chemotaxis. In neonatal OECs abundant mitochondria, lipid vacuoles, and smooth endoplasmic reticulum were detected, suggesting an active lipid metabolism, probably involved in synthesis of myelin. Our results suggest that neonatal OECs obtained from olfactory bulb have microscopic properties that could make them more suitable for neural repair.

[Pathogenesis of spinal cord injuries and mechanisms of repair induced by olfactory ensheathing cells]. / Patogenia de la lesion medular y mecanismos de reparacion inducidos por las celulas de la glia envolvente olfatoria.

INTRODUCTION: Spinal cord injury is a catastrophic event with permanent consequences during the all life. Treatment research has been based in the development of therapies that reduce the discapacity, but since the nineties there has been an important advance and several cellular transplants have been tested in spinal cord animal models, like Schwann cells, astrocytes and olfactory and olfactory ensheathing cells (OEC). AIM: Detailed account of spinal cord injury pathogeny, primary and secondary, and the OEC mechanisms for the regeneration effects that have been described in the literature. DEVELOPMENT: After the trauma, spinal cord injury develops in two phases, the primary injury with characteristics compression lesions, and the secondary produce for several factors that occur in parallel and include: vascular, cellular and molecular factors, and glial scar formation. The most of spinal cord models and OEC transplants have been reported functional recovery, remielinization and axonal regeneration. These cells exert their action in a direct way by producing grow factors and in an indirect way inducing directly neuronal an axonal regeneration and remielinization. CONCLUSIONS: OEC are a therapeutic option in patients with spinal cord injury, because they induce in a direct or indirect way, neuronal and axonal regeneration, remielinization, decrease the glial scar and produce other effects that conduce a functional recovery.

TITLE: Patogenia de la lesion medular y mecanismos de reparacion inducidos por las celulas de la glia envolvente olfatoria.Introduccion. La lesion medular es un evento catastrofico, cuyas consecuencias persisten durante toda la vida del paciente. La investigacion en tratamiento se ha basado principalmente en el desarrollo de terapias que reduzcan la discapacidad, pero desde los anos noventa hay un avance significativo y se han probado varios trasplantes celulares en modelos animales de lesion medular, celulas de Schwann, astrocitos y celulas de la glia envolvente olfatoria (CGEO). Objetivo. Hacer un recuento detallado de la patogenia de la lesion medular primaria y secundaria y de los mecanismos por los cuales las CGEO inducirian sus posibles efectos regenerativos descritos en la bibliografia. Desarrollo. Despues del traumatismo, la lesion se desarrolla en dos fases, la primaria se caracteriza por las lesiones de compresion y la secundaria se produce por una serie de factores que se dan en paralelo y que incluyen factores vasculares, celulares, moleculares y formacion de cicatriz glial. La mayoria de los modelos de lesion medular y trasplante con CGEO han comunicado recuperacion funcional, remielinizacion y regeneracion axonal. Estas celulas ejercen su accion de manera indirecta a traves de la produccion de factores de crecimiento y de manera directa induciendo regeneracion neuronal, axonal y remielinizacion. Conclusiones. Las CGEO son una opcion terapeutica en pacientes con lesion medular debido a que inducen de modo directo o indirecto regeneracion neuronal, axonal, remielinizacion de axones, disminucion de cicatriz glial y otros efectos que conducen a la recuperacion funcional.

Patogenia de la lesión medular y mecanismos de reparación inducidos por las células de la glía envolvente olfatoria / Pathogenesis of spinal cord injuries and mechanisms of repair induced by olfactory ensheathing cells

Introducción. La lesión medular es un evento catastrófico, cuyas consecuencias persisten durante toda la vida del paciente. La investigación en tratamiento se ha basado principalmente en el desarrollo de terapias que reduzcan la discapacidad, pero desde los años noventa hay un avance significativo y se han probado varios trasplantes celulares en modelos animales de lesión medular, células de Schwann, astrocitos y células de la glía envolvente olfatoria (CGEO). Objetivo. Hacer un recuento detallado de la patogenia de la lesión medular primaria y secundaria y de los mecanismos por los cuales las CGEO inducirían sus posibles efectos regenerativos descritos en la bibliografía. Desarrollo. Después del traumatismo, la lesión se desarrolla en dos fases, la primaria se caracteriza por las lesiones de compresión y la secundaria se produce por una serie de factores que se dan en paralelo y que incluyen factores vasculares, celulares, moleculares y formación de cicatriz glial. La mayoría de los modelos de lesión medular y trasplante con CGEO han comunicado recuperación funcional, remielinización y regeneración axonal. Estas células ejercen su acción de manera indirecta a través de la producción de factores de crecimiento y de manera directa induciendo regeneración neuronal, axonal y remielinización. Conclusiones. Las CGEO son una opción terapéutica en pacientes con lesión medular debido a que inducen de modo directo o indirecto regeneración neuronal, axonal, remielinización de axones, disminución de cicatriz glial y otros efectos que conducen a la recuperación funcional (AU)

Introduction. Spinal cord injury is a catastrophic event with permanent consequences during the all life. Treatment research has been based in the development of therapies that reduce the discapacity, but since the nineties there hasbeen an important advance and several cellular transplants have been tested in spinal cord animal models, like Schwann cells, astrocytes and olfactory and olfactory ensheathing cells (OEC). Aim. Detailed account of spinal cord injury pathogeny, primary and secondary, and the OEC mechanisms for the regeneration effects that have been described in the literature. Development. After the trauma, spinal cord injury develops in two phases, the primary injury with characteristics compression lesions, and the secondary produce for several factors that occur in parallel and include: vascular, celular and molecular factors, and glial scar formation. The most of spinal cord models and OEC transplants have been reported functional recovery, remielinization and axonal regeneration. These cells exert their action in a direct way by producing grow factors and in an indirect way inducing directly neuronal an axonal regeneration and remielinization. Conclusions. OEC are a therapeutic option in patients with spinal cord injury, because they induce in a direct or indirect way, neuronal and axonal regeneration, remielinization, decrease the glial scar and produce other effects that conduce a functional recovery (AU)

Ultrastructural Comparison of the Nasal Epithelia of Healthy and Naturally Affected Rabbits with Pasteurella multocida A.

An ultrastructural comparison between the nasal cavities of healthy rabbits and those suffering from two forms of spontaneous infection with Pasteurella multocida was undertaken. Twelve commercially produced rabbits of different ages and respiratory health status were divided into four groups: healthy from 0 to 21 days (G1, n = 2); healthy from 23 to 49 days (G2, n = 2); healthy from 51 to 69 days (G3, n = 2); diseased rabbits with septicemia and the rhinitic form of P. multocida infection (G4, n = 3). The main ultrastructural changes observed were a widening of the interepithelial spaces, increased activity and number of goblet cells, the formation of two types of vacuoles in epithelial cells, the degranulation and migration of heterophils between the epithelial cells, and the association of this migration with some of the other changes. No bacteria were observed adhering to the epithelium, and very few were observed free in the mucus. Scant inter-epithelial spaces were found in healthy rabbits, but they were not as large and numerous as those found in diseased animals. We discuss the origin and meaning of these changes but, we focus on the significance of the inter-epithelial spaces and goblet cells for the defense of the upper respiratory airways against the bacterium and its lipopolysaccharide.

Evaluation of the acute and subchronic oral toxicity of ethanol extract from Valeriana pavonii species in Wistar rats / Evaluación de la toxicidad oral aguda y sub-crónica del extracto etanólico de la especie Valeriana pavonii en ratas Wistar

Introduction: One of the most frequent problems found in medicinal plants is the absence of clinical, toxicological, and pharmacological studies. Valeriana pavonii is one of the species used in Colombia as an anxiolytic. Further study of this specie is rendered to add information in the toxicological area. Objective: The acute and subchronic oral toxicity of V. pavonii ethanolic extract was evaluated in Wistar rats of both sexes. Materials and methods: The rats were distributed into four groups: the control group received the vehicle (0.5 mL/100 g of corporal weight) and the other three groups received increasing levels of the dosage for 90 days to evaluate characteristics like physical exam, laboratory test (blood chemistry and haematology), and anatomopathological findings. Results: This study reveals that there were no signs of toxicity, mortality, or significant alterations attributable to the ethanolic extract of V. pavonii. Conclusions: The Not Observed Adverse Effect Levels (NOAEL) of V. pavonii ethanolic extract were 2000 and 1000 mg/kg of body weight for the acute and subchronic toxicity studies, respectively.

Introducción: Uno de los problemas más frecuentes asociados con el uso de plantas medicinales es la ausencia de evidencias farmacológicas, toxicológicas y clínicas. Valeriana pavonii es una de las especies más utilizadas popularmente en Colombia con fines ansiolíticos. Es necesario avanzar en el estudio de esta especie para aportar información en el campo toxicológico. Objetivos: Evaluar la toxicidad oral aguda y sub-crónica del extracto etanólico de V. pavonii en ratas Wistar de ambos sexos. Materiales y métodos: En cada uno de los estudios se distribuyeron ratas en cuatro grupos; un grupo control que recibió únicamente vehículo (0.5 ml/100 g de peso corporal) y tres grupos correspondientes a niveles crecientes de dosis, así: para el estudio de toxicidad aguda se administraron en dosis única 20, 200 y 2000 mg/kg con un período de observación de 14 días y para el de toxicidad sub-crónica, dosis diarias de 250, 500 y 1000 mg/kg durante 90 días. Se evaluaron los parámetros de examen físico, los exámenes de laboratorio (química sanguínea y hematología) y el estudio anatomopatológico. Resultados: No se presentaron signos de toxicidad, letalidad ni alteraciones significativas atribuibles al consumo del extracto etanólico de V. pavonii, según el examen físico, el examen anatomopatológico y el análisis de las pruebas de química sanguínea y hematología. Conclusiones: Los valores de nivel sin efectos adversos observados (NOAEL) del extracto etanólico de V. pavonii, fueron 2000 y 1000 mg/kg de peso corporal para los estudios de toxicidad aguda y sub-crónica, respectivamente. No se encontraron valores de nivel más bajo de efecto adverso observado (LOAEL).

Bufo marinus oocytes as a model for ion channel protein expression and functional characterization for electrophysiological studies.

Oocytes from Xenopus laevis are commonly used as an expression system for ion channel proteins. The aim of this study was to determine whether oocytes from the Colombian native toad, Bufo marinus, could be used as an alternative expression system for ion channel protein expression and functional characterization using the two-microelectrode voltage clamp method. B. marinus oocytes and X. laevis were isolated and cultured in similar conditions. The mean resting membrane potential of B. marinus oocytes was similar to that of X. laevis oocytes as well as the whole-cell basal currents. The potassium ion channel Kv1.1 was successfully expressed in B. marinus oocytes and showed a typical outward rectifying current. Potassium channel blockers reduced these currents. The similarities on electrical properties and expression of ion channel proteins show that B. marinus oocytes can be used effectively to express these proteins, making these cells a viable heterologous system for the expression of ion channel proteins and their electrophysiological characterization.