Block copolymer nanopatterns affect cell spreading: Stem versus cancer bone cells.

Bone healing after a tumor removal can be promoted by biomaterials that enhance the bone regeneration and prevent the tumor relapse. Herein, we obtained several nanopatterns by self-assembly of polystyrene-block-poly-(2-vinylpyridine) (PS-b-P2VP) with different molecular weights and investigated the adhesion and morphology of human bone marrow mesenchymal stem cells (BMMSC) and osteosarcoma cell line (SaOS-2) on these patterns aiming to identify topography and chemistry that promote bone healing. We analyzed > 2000 cells per experimental condition using imaging software and different morphometric descriptors, namely area, perimeter, aspect ratio, circularity, surface/area, and fractal dimension of cellular contour (FDC). The obtained data were used as inputs for principal component analysis, which showed distinct response of BMMSC and SaOS-2 to the surface topography and chemistry. Among the studied substrates, micellar nanopatterns assembled from the copolymer with high molecular weight promote the adhesion and spreading of BMMSC and have an opposite effect on SaOS-2. This nanopattern is thus beneficial for bone regeneration after injury or pathology, e.g. bone fracture or tumor removal.

Antithrombotic and hemocompatible properties of nanostructured coatings assembled from block copolymers.

We describe the antithrombotic properties of nanopatterned coatings created by self-assembly of poly(styrene-block-2-vinylpyridine) (PS-b-P2VP) with different molecular weights. By changing the assembly conditions, we obtained nanopatterns that differ by their morphology (size and shape of the nanopattern) and chemistry. The surface exposition of P2VP block allowed quaternization, i.e. introduction of positive surface charge and following electrostatic deposition of heparin. Proteins (albumin and fibrinogen) adsorption, platelet adhesion and activation, cytocompatibility, and reendothelization capacity of the coatings were assessed and discussed in a function of the nanopattern morphology and chemistry. We found that quaternization results in excellent antithrombotic and hemocompatible properties comparable to heparinization by hampering the fibrinogen adhesion and platelet activation. In the case of quaternization, this effect depends on the size of the polymer blocks, while all heparinized patterns had similar performance showing that heparin surface coverage of 40 % is enough to improve substantially the hemocompatibility.

Bactericidal nanopatterns generated by block copolymer self-assembly.

We describe the bactericidal capacity of nanopatterned surfaces created by self-assembly of block copolymers. Distinct nanotopographies were generated by spin-coating with polystyrene-block-poly(2-vinylpyridine) (PS-b-P2VP) followed by solvent vapor annealing. We demonstrate that the bactericidal efficiency of the developed coatings depends on the morphology and the chemistry of the surface: cylindrical nanotopographies presenting both blocks at the surface have stronger bactericidal effect on Escherichia coli than micellar patterns with only PS exposed at the surface. The identified mechanism of bacterial death is a mechanical stress exerted by the nanostructures on the cell-wall. Moreover, the developed nanopatterns are not cytotoxic, which makes them an excellent option for coating of implantable materials and devices. The proposed approach represents an efficient tool in the fight against bacteria, which acts via compromising the bacterial wall integrity. STATEMENT OF SIGNIFICANCE: Bacterial infections represent an important risk during biomaterial implantation in surgeries due to the increase of antibiotic resistance. Bactericidal surfaces are a promising solution to avoid the use of antibiotics, but most of those systems do not allow mammalian cell survival. Nanopatterned silicon surfaces have demonstrated to be simultaneously bactericidal and allow mammalian cell culture but are made by physical methods (e.g. plasma etching) applicable to few materials and small surfaces. In this article we show that block copolymer self-assembly can be used to develop surfaces that kill bacteria (E. coli) but do not harm mammalian cells. Block copolymer self-assembly has the advantage of being applicable to many different types of substrates and large surface areas.

An aquaporin 4 antisense oligonucleotide loaded, brain targeted nanoparticulate system design.

Aquaporins (AQPs), members of the water-channel protein family, are highly expressed in brain tissue especially in astrocytic end-feet. They are important players for water hemostasis during development of cytotoxic as well as vasogenic edema. Increased expression of AQPs is important in pathophysiology of neurological diseases such as neuroinflammation and ischemia. Unfortunately, there are a few pharmacological inhibitors of AQP4 with several side effects limiting their translation as a drug for use in clinical conditions. Another therapeutic approach is using antisense oligonucleotides (ASOs) to block AQP4 activity. These are short, synthetic, modified nucleic acids that bind RNA to modulate its function. However, they cannot pass the blood brain barrier (BBB). To overcome this obstacle we designed a nanoparticulate system made up of chitosan nanoparticles surface modified with PEG and conjugated with monoclonal anti transferrin receptor-1 antibody via streptavidin-biotin binding. The nanocarrier system could be targeted to the transferrin receptor-1 at the brain endothelial capillaries through monoclonal antibodies. It is hypothesized that the nanoparticles could pass the BBB via receptor mediated transcytosis and reach brain parenchyma. Particle size, zeta potential, loading capacity and release profiles of nanoparticles were investigated. It was observed that all types of chitosau (CS) nanoparticles had positive zeta potential values and nanoparticle particle size distribution varied between 100 and 800 nm. The association efficiency of ASOs into the nanoparticles was between 80-97% and the release profiles of the nanoparticles exhibited an initial burst effect followed by a controlled release. The results showed that the designed chitosan based nanocarriers could be a promising carrier system to transport nucleic acid based drugs to brain parenchyma.

¿Se realiza correctamente la punción lumbar en pediatría? Revisión de las recomendaciones actuales y análisis de la realidad / Do paediatricians perform lumbar puncture correctly? Review of recommendations and analysis the technique in Spain

Introducción: La punción lumbar (PL) es una técnica ampliamente utilizada en pediatría. La realización adecuada de la técnica puede evitar la mayoría de las complicaciones asociadas. Objetivo: Evaluar si en España los pediatras y los residentes de pediatría siguen las recomendaciones de la técnica de la PL. Material y métodos: Estudio transversal basado en un cuestionario escrito distribuido por correo electrónico a través de la Sociedad Española de Urgencias Pediátricas, que recogía datos epidemiológicos y preguntas de respuesta múltiple sobre la realización de la PL. Resultados: Se analizaron 206 cuestionarios, 143 (69,5%) respondidos por pediatras y 63 (30,5%) por residentes de pediatría. Hacen la PL sin los padres presentes 128 médicos (62,1%), aplican sedoanalgesia 198 (96,1%); 84 (42%) sólo analgesia local. Colocan al paciente sentado 108 (53,7%). La mayoría utiliza aguja tipo Quincke (126; 62,7%), orienta bien el trocar al hacer la punción 22 (36,1%) residentes y 21 pediatras (15,1%), diferencia estadísticamente significativa (p=0,001). En los neonatos, 63 (46%) pediatras y 19 (30,2%) residentes usan aguja sin fiador tipo «palomilla», diferencia también significativa (p=0,035). Reintroducen el estilete para redirigir el trocar 190 (92,2%) encuestados y para extraerlo 186 (93%). Recomiendan reposo tras la PL 195 (94,7%) médicos. Conclusiones: La mayoría de los pediatras orientan mal el trocar cuando hacen una PL y siguen utilizando aguja tipo «palomilla» en los neonatos a pesar de que está desaconsejado. Los residentes de pediatría y los pediatras con menor experiencia siguen las recomendaciones con mayor frecuencia(AU)

Introduction: Lumbar puncture (LP) is a commonly performed procedure in paediatrics. Performing this technique properly can avoid the most common associated complications. Objective: To assess whether paediatricians and paediatric residents in Spain follow current recommendations for the LP technique. Material and methods: A cross-sectional study was conducted by sending a questionnaire by mail through the Spanish Society of Paediatric Emergencies, collecting demographic information and responses to multiple choice questions about LP technique. Results: A total of 206 questionnaires were analysed, of which 143 (69.5%) were answered by paediatricians, and 63 (30.5%) by paediatric residents. The majority (128; 62.1%) of physicians did not allow parents to be present during LP, 198 (96.1%) routinely use analgesia and sedation; 84 (42%) only used local anaesthesia. The majority of respondents used standard Quincke needles (126; 62.7%). The bevel was correctly positioned when puncturing the dura mater by 22 residents (36.1%) and 21 paediatricians (15.1%), a variation that was statistically significant (P=.001). For neonatal lumbar punctures, 63 paediatricians (46%) and 19 paediatric residents used a butterfly needle which did not contain a stylet, and this difference was also statistically significant (P=.035). Of those surveyed, 190 (92.2%) re-inserted the stylet when re-orientating the needle, and 186 (93%) re-oriented this when removing it. The recommendation of bed rest was made by 195 (94.7%) physicians. Conclusions: The majority of paediatricians orient the bevel wrongly when inserting the needle during LP, and still use "butterfly" needles in newborns, despite warnings to the contrary. Paediatric residents and less experienced paediatricians follow the recommendations more frequently(AU)

[Do paediatricians perform lumbar puncture correctly? Review of recommendations and analysis the technique in Spain]. / ¿Se realiza correctamente la punción lumbar en pediatría? Revisión de las recomendaciones actuales y análisis de la realidad.

INTRODUCTION: Lumbar puncture (LP) is a commonly performed procedure in paediatrics. Performing this technique properly can avoid the most common associated complications. OBJECTIVE: To assess whether paediatricians and paediatric residents in Spain follow current recommendations for the LP technique. MATERIAL AND METHODS: A cross-sectional study was conducted by sending a questionnaire by mail through the Spanish Society of Paediatric Emergencies, collecting demographic information and responses to multiple choice questions about LP technique. RESULTS: A total of 206 questionnaires were analysed, of which 143 (69.5%) were answered by paediatricians, and 63 (30.5%) by paediatric residents. The majority (128; 62.1%) of physicians did not allow parents to be present during LP, 198 (96.1%) routinely use analgesia and sedation; 84 (42%) only used local anaesthesia. The majority of respondents used standard Quincke needles (126; 62.7%). The bevel was correctly positioned when puncturing the dura mater by 22 residents (36.1%) and 21 paediatricians (15.1%), a variation that was statistically significant (P=.001). For neonatal lumbar punctures, 63 paediatricians (46%) and 19 paediatric residents used a butterfly needle which did not contain a stylet, and this difference was also statistically significant (P=.035). Of those surveyed, 190 (92.2%) re-inserted the stylet when re-orientating the needle, and 186 (93%) re-oriented this when removing it. The recommendation of bed rest was made by 195 (94.7%) physicians. CONCLUSIONS: The majority of paediatricians orient the bevel wrongly when inserting the needle during LP, and still use "butterfly" needles in newborns, despite warnings to the contrary. Paediatric residents and less experienced paediatricians follow the recommendations more frequently.

Complicaciones del cateterismo vesical realizado en un servicio de urgencias para obtener una muestra de orina / Complications of bladder catheterization to obtain a urine sample in the emergency department

Introducción: El cateterismo vesical (CV) es una técnica utilizada con frecuencia para diagnosticar infecciones urinarias (ITU) (infección bacteriana invasora más frecuente de los lactantes) en los niños no continentes. El objetivo principal de este estudio es conocer el rendimiento y las complicaciones del CV transitorio para obtener una muestra de orina. Material y mé todos: Se ha realizado un estudio prospectivo de los pacientes sometidos a CV entre el 1 de noviembre de 2009 y el 15 de diciembre de 2009 en el servicio de urgencias (SU) de un hospital terciario. Además de recoger los datos epidemiológicos y el motivo de consulta que motivó el CV, se contactó por teléfono con la familia al menos un mes después, para conocer las complicaciones. Resultados: Se realizaron 126 procedimientos en 124 pacientes, 45 varones (35,7%) y 81 mujeres (64,3%). Se obtuvo muestra de orina en todos los casos. La mediana de edad fue de 7 meses (5 días-35 meses). De los 126 urocultivos recogidos, resultaron positivos 34 (26,98%), siendo el patógeno más frecuente la Escherichia coli (27; 79,41%). Se consideró contaminado 1 cultivo (0,8%). La fiebre (76,2%) fue el principal motivo de consulta que estableció la indicación del CV. Las complicaciones de los 116 pacientes con los que se contactó por teléfono fueron: dolor referido al área genital (3), hematuria no confirmada (1) e ITU 12 días después (1). Conclusiones: El CV transitorio para obtener una muestra de orina en los niños no continentes es efectivo y seguro. El CV es una técnica invasora que necesita controles de calidad (AU)

Introduction: Bladder catheterization (BC) is frequently performed to diagnose urinary tract infection (UTI) (the most common serious bacterial infection among young children). The purposes of this study were to evaluate performance and complications of BC to obtain a urine specimen. Material and methods: This was a prospective study conducted in an urban childrenŒs hospital on patients who had a urine specimen obtained in the emergency department (ED) by BC between November 1, 2009 and December 15, 2009. Epidemiological data and the medical reasons why the BC was performed were reviewed. Finally, to determine the complications we contacted the family by telephone at least one month after the BC. Results: A total of 126 BC were performed in 124 patients, 45 males (35.7%) and 81 female (64.3%). A urine sample was obtained in all cases. The median age was 7 months (5 days-35 months). Of the 126 urine cultures collected, 34 were positive (26.98%) and 1 was considered contaminated (0.8%). Escherichia coli was the most common pathogen (27, 79.41%). Fever (76.2%) was the main reason why BC was performed in children. Of the 116 patients contacted by telephone, only 5 mentioned complications: genital pain (3), unconfirmed hematuria (1) and UTI 12 days after (1). Conclusions: BC performed in ED to obtain a urine sample in non toilet-trained children is effective and safe. BC is an invasive method which needs to be quality controlled (AU)

[Complications of bladder catheterization to obtain a urine sample in the emergency department]. / Complicaciones del cateterismo vesical realizado en un servicio de urgencias para obtener una muestra de orina.

INTRODUCTION: Bladder catheterization (BC) is frequently performed to diagnose urinary tract infection (UTI) (the most common serious bacterial infection among young children). The purposes of this study were to evaluate performance and complications of BC to obtain a urine specimen. MATERIAL AND METHODS: This was a prospective study conducted in an urban children's hospital on patients who had a urine specimen obtained in the emergency department (ED) by BC between November 1, 2009 and December 15, 2009. Epidemiological data and the medical reasons why the BC was performed were reviewed. Finally, to determine the complications we contacted the family by telephone at least one month after the BC. RESULTS: A total of 126 BC were performed in 124 patients, 45 males (35.7%) and 81 female (64.3%). A urine sample was obtained in all cases. The median age was 7 months (5 days-35 months). Of the 126 urine cultures collected, 34 were positive (26.98%) and 1 was considered contaminated (0.8%). Escherichia coli was the most common pathogen (27, 79.41%). Fever (76.2%) was the main reason why BC was performed in children. Of the 116 patients contacted by telephone, only 5 mentioned complications: genital pain (3), unconfirmed hematuria (1) and UTI 12 days after (1). CONCLUSIONS: BC performed in ED to obtain a urine sample in non toilet-trained children is effective and safe. BC is an invasive method which needs to be quality controlled.

Preparation and evaluation of alpha-phenyl-n-tert-butyl nitrone (PBN)-encapsulated chitosan and PEGylated chitosan nanoparticles.

Alpha-phenyl-n-tert-butyl nitrone (PBN) shows its major effect by scavenging free radicals formed in the ischemia and it has the ability to penetrate through the blood brain barrier easily. The in vivo stability of PBN is very low and when administered systemically, it has a mean plasma half life of about three hours. Therefore, formulations which are able to prolong the plasma residence time of PBN are of major interest, because oxygen radicals are usually continuously formed under pathological conditions. In this study, PBN, a nitrone compound having neuroprotective properties, was encapsulated in chitosan (CS) and chitosan-poly(ethylene glycol) (CS-PEG) nanoparticles for treatment of diseases such as stroke, in which sustained free radical production is reported. The nanoparticles were characterized through particle size determination, zeta potential, encapsulation efficiency, surface morphology determinations and in vitro release studies. The surface morphologies were evaluated by transmission electron microscopy (TEM) and nanoparticles having spherical shapes were characterized. The particle size distribution was between approximately 97 nm and approximately 322 nm; and the zeta potentials varied between approximately 9 mV and approximately 33 mV. Size of the nanoparticle formulations was important for the release of PBN from nanoparticles. The quantitative determination of PBN has been evaluated by a validated analytical HPLC method. The presented chitosan-based nanotechnology opens new perspectives for testing antioxidant activity in vivo.

Chitosan-PEG nanocapsules as new carriers for oral peptide delivery. Effect of chitosan pegylation degree.

We have previously reported the ability of chitosan nanocapsules to enhance and prolong the oral absorption of peptides. In the present work, our goal was to design a new type of nanocapsules, using chitosan chemically modified with poly(ethylene glycol) (PEG) (0.5% and 1% pegylation degree) and to investigate the consequences of this modification on the in vitro and in vivo behaviour of the nanocapsules. Chitosan-PEG nanocapsules and the control PEG-coated nanoemulsions were obtained by the solvent displacement technique. Their size was in the range of 160-250 nm. Their zeta potential was greatly affected by the nature of the coating, being positive for chitosan-PEG nanocapsules and negative in the case of PEG-coated nanoemulsions. The presence of PEG, whether alone or grafted to chitosan, improved the stability of the nanocapsules in the gastrointestinal fluids. Using the Caco-2 model cell line it was observed that the pegylation of chitosan reduced the cytotoxicity of the nanocapsules. In addition, these nanocapsules did not cause a significant change in the transepithelial resistance of the monolayer. Finally, the results of the in vivo studies showed the capacity of chitosan-PEG nanocapsules to enhance and prolong the intestinal absorption of salmon calcitonin. Additionally, they indicated that the pegylation degree affected the in vivo performance of the nanocapsules. Therefore, by modulating the pegylation degree of chitosan, it was possible to obtain nanocapsules with a good stability, a low cytotoxicity and with absorption enhancing properties.