Poly(ester-urethane) scaffolds: effect of structure on properties and osteogenic activity of stem cells

The present study aimed to investigate the effect of structure (design and porosity) on the matrix stiffness and osteogenic activity of stem cells cultured on poly(ester-urethane) (PEU) scaffolds. Different three-dimensional (3D) forms of scaffold were prepared from lysine-based PEU using traditional salt-leaching and advanced bioplotting techniques. The resulting scaffolds were characterized by differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), scanning electron microscopy (SEM), mercury porosimetry and mechanical testing. The scaffolds had various pore sizes with different designs, and all were thermally stable up to 300â °C. In vitrotests, carried out using rat bone marrow stem cells (BMSCs) for bone tissue engineering, demonstrated better viability and higher cell proliferation on bioplotted scaffolds compared to salt-leached ones, most probably due to their larger and interconnected pores and stiffer nature, as shown by higher compressive moduli, which were measured by compression testing. Similarly, SEM, von Kossa staining and EDX analyses indicated higher amounts of calcium deposition on bioplotted scaffolds during cell culture. It was concluded that the design with larger interconnected porosity and stiffness has an effect on the osteogenic activity of the stem cells.

An evolutionary approach to the use of Petri net based models: from parallel controllers to HW/SW co-design

"A workshop within the 19th International Conference on Applications and Theory of Petri Nets - ICATPN’1998"

Diseño y caracterización de plataformas de biorreconocimiento basadas en el uso de nanoestructuras, biomoléculas y polímeros. Aplicaciones para el desarrollo de (bio)sensores electroquímicos. Design and characterization of biorecognition platforms based on the use of nanostructures, biomolecules and polymers. Applications for the development of electrochemical (bio)sensors.

Los requerimientos de métodos analíticos que permitan realizar determinaciones más eficientes en diversas ramas de la Química, así como el gran desarrollo logrado por la Nanobiotecnología, impulsaron la investigación de nuevas alternativas de análisis. Hoy, el campo de los Biosensores concita gran atención en el primer mundo, sin embargo, en nuestro país es todavía un área de vacancia, como lo es también la de la Nanotecnología. El objetivo de este proyecto es diseñar y caracterizar nuevos electrodos especialmente basados en el uso de nanoestructuras y estudiar aspectos básicos de la inmovilización de enzimas, ADN, aptámeros, polisacáridos y otros polímeros sobre dichos electrodos a fin de crear nuevas plataformas de biorreconocimiento para la construcción de (bio)sensores electroquímicos dirigidos a la cuantificación de analitos de interés clínico, farmaco-toxicológico y ambiental.Se estudiarán las propiedades de electrodos de C vítreo, Au, "screen printed" y compósitos de C modificados con nanotubos de C (CNT) y/o nanopartículas (NP) de oro y/o nanoalambres empleando diversas estrategias. Se investigarán nuevas alternativas de inmovilización de las biomoléculas antes mencionadas sobre dichos electrodos, se caracterizarán las plataformas resultantes y se evaluarán sus posibles aplicaciones analíticas al desarrollo de biosensores con enzimas y ADNs como elementos de biorreconocimiento. Se funcionalizarán CNT con polímeros comerciales y sintetizados en nuestro laboratorio modificados con moléculas bioactivas. Se diseñarán y caracterizarán nuevas arquitecturas supramoleculares basadas en el autoensamblado de policationes, enzimas y ADNs sobre Au. Se evaluarán las propiedades catalíticas de NP de magnetita y de perovskitas de Mn y su aplicación al desarrollo de biosensores enzimáticos. Se diseñarán biosensores que permitan la detección altamente sensible y selectiva de secuencias específicas de ADNs de interés clínico. Se estudiará la interacción de genotóxicos con ADN (en solución e inmovilizado) y se desarrollarán biosensores que permitan su cuantificación. Se construirán biosensores enzimáticos para la cuantificación de bioanalitos, especialmente glucosa, fenoles y catecoles, y sensores electroquímicos para la determinación de neurotransmisores, ácido úrico y ácido ascórbico. Se diseñarán nuevos aptasensores electroquímicos para la cuantificación de biomarcadores, comenzando por lisozima y trombina y continuando con otros de interés regional/nacional.Se emplearán las siguientes técnicas: voltamperometrías cíclica (CV), de pulso diferencial (DPV) y de onda cuadrada (SWV); "stripping" potenciométrico a corriente constante (PSA); elipsometría; microbalanza de cristal de cuarzo con cálculo de pérdida de energía por disipación (QCM-D); resonancia de plasmón superficial con detección dual (E-SPR); espectroscopía de impedancia electroquímica (EIE); microscopías de barrido electroquímico (SECM), de barrido electrónico (SEM), de transmisión (TEM) y de fuerzas atómicas (AFM); espectrofotometría UV-visible; espectroscopías IR, Raman, de masas, RMN.Se espera que la inclusión de los CNT y/o de las NP metálicas y/o de los nanoalambres en los diferentes electrodos permita una mejor transferencia de carga de diversos analitos y por ende una detección más sensible y selectiva de bioanalitos empleando enzimas, ADN y aptámeros como elementos de biorreconocimiento. Se espera una mayor eficiencia en los aptasensores respecto de los inmunosensores, lo que permitirá la determinacion selectiva de diversos biomarcadores. La modificación de electrodos con nanoestructuras posibilitará la detección altamente sensible y selectiva del evento de hibridación. La respuesta obtenida luego de la interacción de genotóxicos con ADN permitirá un mejor conocimiento de la asociación establecida, de la cinética y de las constantes termodinámicas. Los neurotransmisores podrán ser determinados a niveles nanomolares aún en muestras complejas.

Design of biped robot walking based on non-linear periodical oscillations

Magdeburg, Univ., Fak. für Elektrotechnik und Informationstechnik, Diss., 2012

Process design based on CO 2-expanded liquids as solvents

Magdeburg, Univ., Fak. für Verfahrens- und Systemtechnik, Diss., 2014

Bioluminescent sensor proteins for point-of-care therapeutic drug monitoring.

For many drugs, finding the balance between efficacy and toxicity requires monitoring their concentrations in the patient's blood. Quantifying drug levels at the bedside or at home would have advantages in terms of therapeutic outcome and convenience, but current techniques require the setting of a diagnostic laboratory. We have developed semisynthetic bioluminescent sensors that permit precise measurements of drug concentrations in patient samples by spotting minimal volumes on paper and recording the signal using a simple point-and-shoot camera. Our sensors have a modular design consisting of a protein-based and a synthetic part and can be engineered to selectively recognize a wide range of drugs, including immunosuppressants, antiepileptics, anticancer agents and antiarrhythmics. This low-cost point-of-care method could make therapies safer, increase the convenience of doctors and patients and make therapeutic drug monitoring available in regions with poor infrastructure.

Single-cell gene-expression profiling reveals qualitatively distinct CD8 T cells elicited by different gene-based vaccines.

CD8 T cells play a key role in mediating protective immunity against selected pathogens after vaccination. Understanding the mechanism of this protection is dependent upon definition of the heterogeneity and complexity of cellular immune responses generated by different vaccines. Here, we identify previously unrecognized subsets of CD8 T cells based upon analysis of gene-expression patterns within single cells and show that they are differentially induced by different vaccines. Three prime-boost vector combinations encoding HIV Env stimulated antigen-specific CD8 T-cell populations of similar magnitude, phenotype, and functionality. Remarkably, however, analysis of single-cell gene-expression profiles enabled discrimination of a majority of central memory (CM) and effector memory (EM) CD8 T cells elicited by the three vaccines. Subsets of T cells could be defined based on their expression of Eomes, Cxcr3, and Ccr7, or Klrk1, Klrg1, and Ccr5 in CM and EM cells, respectively. Of CM cells elicited by DNA prime-recombinant adenoviral (rAd) boost vectors, 67% were Eomes(-) Ccr7(+) Cxcr3(-), in contrast to only 7% and 2% stimulated by rAd5-rAd5 or rAd-LCMV, respectively. Of EM cells elicited by DNA-rAd, 74% were Klrk1(-) Klrg1(-)Ccr5(-) compared with only 26% and 20% for rAd5-rAd5 or rAd5-LCMV. Definition by single-cell gene profiling of specific CM and EM CD8 T-cell subsets that are differentially induced by different gene-based vaccines will facilitate the design and evaluation of vaccines, as well as enable our understanding of mechanisms of protective immunity.

Population Normalization with Ammonium in Wastewater-Based Epidemiology: Application to Illicit Drug Monitoring

Fluctuations in ammonium (NH4+), measured as NH4-N loads using an ion-selective electrode installed at the inlet of a sewage treatment plant, showed a distinctive pattern which was associated to weekly (i.e., commuters) and seasonal (i.e., holidays) fluctuations of the population. Moreover, population size estimates based on NH4-N loads were lower compared to census data. Diurnal profiles of benzoylecgonine (BE) and 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (THC-COOH) were shown to be strongly correlated to NH4-N. Characteristic patterns, which reflect the prolonged nocturnal activity of people during the weekend, could be observed for BE, cocaine, and a major metabolite of MDMA (i.e., 4-hydroxy-3-methoxymethamphetamine). Additional 24 h composite samples were collected between February and September 2013. Per-capita loads (i.e., grams per day per 1000 inhabitants) were computed using census data and NH4-N measurements. Normalization with NH4-N did not modify the overall pattern, suggesting that the magnitude of fluctuations in the size of the population is negligible compared to those of illicit drug loads. Results show that fluctuations in the size of the population over longer periods of time or during major events can be monitored using NH4-N loads: either using raw NH4-N loads or population size estimates based on NH4-N loads, if information about site-specific NH4-N population equivalents is available.

Low-frequency drug-resistant HIV-1 and risk of virological failure to first-line NNRTI-based ART: a multicohort European case-control study using centralized ultrasensitive 454 pyrosequencing.

OBJECTIVES: It is still debated if pre-existing minority drug-resistant HIV-1 variants (MVs) affect the virological outcomes of first-line NNRTI-containing ART. METHODS: This Europe-wide case-control study included ART-naive subjects infected with drug-susceptible HIV-1 as revealed by population sequencing, who achieved virological suppression on first-line ART including one NNRTI. Cases experienced virological failure and controls were subjects from the same cohort whose viraemia remained suppressed at a matched time since initiation of ART. Blinded, centralized 454 pyrosequencing with parallel bioinformatic analysis in two laboratories was used to identify MVs in the 1%-25% frequency range. ORs of virological failure according to MV detection were estimated by logistic regression. RESULTS: Two hundred and sixty samples (76 cases and 184 controls), mostly subtype B (73.5%), were used for the analysis. Identical MVs were detected in the two laboratories. 31.6% of cases and 16.8% of controls harboured pre-existing MVs. Detection of at least one MV versus no MVs was associated with an increased risk of virological failure (OR = 2.75, 95% CI = 1.35-5.60, P = 0.005); similar associations were observed for at least one MV versus no NRTI MVs (OR = 2.27, 95% CI = 0.76-6.77, P = 0.140) and at least one MV versus no NNRTI MVs (OR = 2.41, 95% CI = 1.12-5.18, P = 0.024). A dose-effect relationship between virological failure and mutational load was found. CONCLUSIONS: Pre-existing MVs more than double the risk of virological failure to first-line NNRTI-based ART.

Long-term effects of cannabis on brain structure.

The dose-dependent toxicity of the main psychoactive component of cannabis in brain regions rich in cannabinoid CB1 receptors is well known in animal studies. However, research in humans does not show common findings across studies regarding the brain regions that are affected after long-term exposure to cannabis. In the present study, we investigate (using Voxel-based Morphometry) gray matter changes in a group of regular cannabis smokers in comparison with a group of occasional smokers matched by the years of cannabis use. We provide evidence that regular cannabis use is associated with gray matter volume reduction in the medial temporal cortex, temporal pole, parahippocampal gyrus, insula, and orbitofrontal cortex; these regions are rich in cannabinoid CB1 receptors and functionally associated with motivational, emotional, and affective processing. Furthermore, these changes correlate with the frequency of cannabis use in the 3 months before inclusion in the study. The age of onset of drug use also influences the magnitude of these changes. Significant gray matter volume reduction could result either from heavy consumption unrelated to the age of onset or instead from recreational cannabis use initiated at an adolescent age. In contrast, the larger gray matter volume detected in the cerebellum of regular smokers without any correlation with the monthly consumption of cannabis may be related to developmental (ontogenic) processes that occur in adolescence.

Desenvolupament de vacunes peptídiques y dendrímers basats en polietilenglicol per al tractament del càncer

El principal problema de les teràpies actuals contra el càncer es la baixa especificitat envers les cèl•lules tumorals, cosa que comporta gran quantitat d’efectes secundaris. Per això es important el desenvolupament de nous tipus de teràpies i sistemes d’alliberament efectius per als fàrmacs ja existents al mercat. En la immunoteràpia contra el càncer es pretén estimular el sistema immunològic per a eliminar les cèl•lules canceroses de manera selectiva. En aquest projecte s’han sintetitzat derivats de l’antigen peptídic de melanoma NY-ESO1 i s’ha estudiat la seva capacitat per a estimular el sistema immunològic com a vacunes contra el càncer. També s’han encapsulat el antígens peptídics en liposomes com a adjuvants i sistemes d’alliberament. De totes les variants peptídiques la que resultà més immunogènica fou la que contenia el grup palmitoil i el fragment toxoide tetànic en la seva estructura. La utilització de liposomes com a sistema adjuvant sembla una estratègia interessant per al disseny de vacunes contra el càncer donat que l’encapsulació del pèptid en liposomes va augmentar notablement la resposta immunològica de l’antigen. Per altra banda, s’han desenvolupat dendrímers basats en polietilenglicol com a sistemes alliberadors de fàrmacs per al tractament de tumors. El polietilenglicol és àmpliament utilitzat com a sistema d’alliberament de fàrmacs degut a les seves interessants propietats, augment de la solubilitat i dels temps de residència en plasma, entre d’altres. La metodologia química descrita permet la diferenciació controlada de varies posicions en la superfície del dendrímer a més del creixement del dendrímer fins a una segona generació. S’ha sintetitzat la primera generació del dendrímer contenint el fàrmac antitumoral 5-fluorouracil i s’han realitzat estudis de citotoxicitat comprovant que l’activitat del nanoconjugat és del mateix ordre de magnitud que el 5-fluorouracil sense conjugar.

Suivi de l'adhésion thérapeutique des patients hypertendus ou dyslipi- démiques par les cercles de qualité médecins-pharmaciens [Monitoring drug adherence for hypertensive and dyslipidemic patients in networks of community-based pharmacists and physicians].

Patient adherence is often poor for hypertension and dyslipidaemia. A monitoring of drug adherence might improve these risk factors control, but little is known in ambulatory care. We conducted a randomised controlled study in networks of community-based pharmacists and physicians in the canton of Fribourg to examine whether monitoring drug adherence with an electronic monitor (MEMS) would improve risk factor control among treated, but uncontrolled hypertensive and dyslipidemic patients. The results indicate that MEMS achieve a better blood pressure control and lipid profile, although its implementation requires considerable resources. The study also shows the value of collaboration between physicians and pharmacists in the field of patient adherence to improve ambulatory care of patients with cardiovascular risk factors.

Mapping HIV-related behavioural surveillance among injecting drug users in Europe, 2008.

The systematic collection of behavioural information is an important component of second-generation HIV surveillance. The extent of behavioural surveillance among injecting drug users (IDUs) in Europe was examined using data collected through a questionnaire sent to all 31 countries of the European Union and European Free Trade Association as part of a European-wide behavioural surveillance mapping study on HIV and other sexually transmitted infections. The questionnaire was returned by 28 countries during August to September 2008: 16 reported behavioural surveillance studies (two provided no further details). A total of 12 countries used repeated surveys for behavioural surveillance and five used their Treatment Demand Indicator system (three used both approaches). The data collected focused on drug use, injecting practices, testing for HIV and hepatitis C virus and access to healthcare. Eight countries had set national indicators: three indicators were each reported by five countries: the sharing any injecting equipment, uptake of HIV testing and uptake of hepatitis C virus testing. The recall periods used varied. Seven countries reported conducting one-off behavioural surveys (in one country without a repeated survey, these resulted an informal surveillance structure). All countries used convenience sampling, with service-based recruitment being the most common approach. Four countries had used respondent-driven sampling. Three fifths of the countries responding (18/28) reported behavioural surveillance activities among IDUs; however, harmonisation of behavioural surveillance indicators is needed.

Population structure of the malaria vector Anopheles darlingi in Rondônia, Brazilian Amazon, based on mitochondrial DNA

Anopheles darlingi is the most important Brazilian malaria vector, with a widespread distribution in the Amazon forest. Effective strategies for vector control could be better developed through knowledge of its genetic structure and gene flow among populations, to assess the vector diversity and competence in transmitting Plasmodium. The aim of this study was to assess the genetic diversity of An. darlingi collected at four locations in Porto Velho, by sequencing a fragment of the ND4 mitochondrial gene. From 218 individual mosquitoes, we obtained 20 different haplotypes with a diversity index of 0.756, equivalent to that found in other neotropical anophelines. The analysis did not demonstrate significant population structure. However, haplotype diversity within some populations seems to be over-represented, suggesting the presence of sub-populations, but the presence of highly represented haplotypes complicates this analysis. There was no clear correlation among genetic and geographical distance and there were differences in relation to seasonality, which is important for malarial epidemiology.