Micro y nanocanales integrados: nuevas herramientas para control de movimiento molecular

This paper presents a review of work on the fabrication and use of nanochannels in silicon and polymers for the control of molecular transport. The method of Sacrifi cial Layer Lithography is reviewed and demonstrated for silicon and polymers. A novel technique for the productions of conical nanopores through a polymer membrane is also reviewed. Nanochannels and nanopores have many potential applications for drug delivery, immunoprotection of cell implants, blocking of globular proteins from biosensor surfaces, and diagnostic devices. All of these applications benefi t from the more direct interactions of devices with biomolecules.El presente trabajo presenta una revisión literaria sobre los métodos de fabricación de nanocanales en silicio y diferentes materiales poliméricos; y su uso en control de transporte molecular. Se describe el método “Sacrifi cial Layer Lithography” para silicio y polímeros. Adicionalmente, una novedosa técnica para la producción de nanoporos cónicos a través de una membrana polimérica es descrita. Los nanocanales y los nanoporos poseen diversas aplicaciones potenciales en la liberación de drogas, en la inmunoprotección de implantes celulares, el bloqueo de proteínas globulares en la superfi cie de biosensores, y en dispositivos para diagnóstico. Todas estas aplicaciones se benefi cian de la interacción directa entre los dispositivos y las biomoléculas

Acoustic sensor with PVDF micro pillars and patterned electrodes

ABSTRACT This paper addresses the design and theoretical analysis of a new type of millimeter-size acoustic sensor that uses Polyvinylidene Fluoride (PVDF) micro-pillars and patterned electrodes. The sensor has the potential to achieve 100x the sensitivity of existing commercial sensors in combination with a dynamic range of 181dB and a frequency bandwidth of at least 100 kHz. A constrained optimization algorithm has been developed as a function of geometric parameters (sensor footprint, diameter and height of the micro-pillars, gap between pillar edges, number of pillars) and electrical parameters of the sensor and conditioning amplifier

Novel clone selection technique reveals heterogeneity among HEK293T cells engineered to produce therapeutic extracellular vesicles

HEK293T cells have been engineered to produce extracellular vesicles (EVs) that deliver miR-199a-3p to CD44+ hepatocellular carcinoma cells. Restoration of this miRNA has been shown to slow cancer progression in-vitro. Isolation and analysis of EVs from cell culture media containing selection agent revealed that the number of miRNA-199a-3p copies was less than the number of cells in culture suggesting that not all cells produce therapeutic EVs. Therefore, therapeutic EV production can be significantly increased by selecting the HEK293T clones that produce the most therapeutic EVs. While clone selection is traditionally accomplished by cell analysis techniques such as fluorescence activated cell sorting (FACS), detection of therapeutic EVs poses a unique challenge in that cellular expression of miRNA-199a-3p does not necessarily correlate to the amount of exosomal miRNA-199a-3p. In response to this challenge, a fibrous microwell array was developed to screen thousands of clones for therapeutic EV productivity (figure 1). The fibrous microwell system is able to evaluate cell growth rate under fluid shear stress, EV productivity and EV characterization using fluorescently labeled antibodies or cationic lipoplex nanoparticles (detect presence of miRNA-199a-3p inside captured EVs produced by single clones). The most productive clones can be released from the microwells and grown in large scale cell culture to significantly increase therapeutic EV production. Please click Additional Files below to see the full abstract

Over-the-counter orlistat: early experiences, views and attitudes of community pharmacists in Great Britain.

Objectives of the study To describe community pharmacists early experiences, views and attitudes with over-the-counter orlistat, 9 months post legal re-classification from November 2009 to January 2010. Setting 13,200 (81%) randomly selected registered community pharmacies across Great Britain out of a potential 16,200. Methods A cross-sectional postal questionnaire survey of the main pharmacist with greatest responsibility for over-the-counter (OTC) supply. Main outcome measures Pharmacists early experiences, views and attitudes of orlistat supply, demographic data of respondents and personal opinions with the supply of orlistat. Results Questionnaires were returned by 32.4% (n = 4,026) of pharmacists surveyed. Just over half (51.9%, n = 2,091) reported no sales of orlistat in the previous 4 weeks with only 5.1% (n = 203) reporting frequent (5.1%) or very frequent supply in the same time frame. Two thirds (66.5%, n = 2,676) agreed or strongly agreed that the sale of orlistat was a good opportunity to extend their role as a healthcare professional and 92% (n = 3,712) felt confident in their ability to supply this product. Over half (57.9%, n = 2,334) admitted that customers frequently complained about the cost of the product and 47.8% (n = 1,926) agreed that customers could misuse the product. Conclusion Despite community pharmacists welcoming orlistat re-classification to increase medicines availability as an opportunity to extend their healthcare professional role there were concerns about poor public uptake, high cost and the potential for misuse. Exploratory studies collecting the views and experiences of the general public about the access and provision of weight management services through community pharmacies are warranted

Isolation of human bone marrow mesenchymal stem cells and evaluation of their osteogenic potential

Las células madre mesenquimatosas de médula ósea humana (abreviadas hBMSCs) constituyen una fuente de células auto-renovables con alto potencial de diferenciación, comúnmente aisladas a partir de los aspirados medulares en huesos largos. Su diferenciación hacia el linaje osteogénico, por ejemplo, ha sido ampliamente utilizada para la evaluación biológica de biomateriales o matrices con aplicaciones en la ingeniería de tejidos óseos. El objetivo de este trabajo consistió en aislar hBMSCs a partir de la cabeza femoral de pacientes sometidos a artroplastia total de cadera, así como evaluar su potencial osteogénico. Brevemente, se extrajo el hueso esponjoso y se disgregó mecánicamente; las células desprendidas se cultivaron y las células no adherentes se eliminaron luego de 4 días. El potencial osteogénico se evaluó en la quinta generación de cultivo, mediante ensayos de diferenciación a 14 y 20 días donde se compararon cultivos con y sin suplementos osteogénicos. La evaluación se realizó mediante tinción con Alizarina Roja y la cuantificación de los niveles de expresión génica de los marcadores osteogénicos colágeno tipo I, osteonectinca y sialoprotiena ósea mediante RT-PCR en tiempo real. Las hBMSCs obtenidas presentaron un fenotipo no-diferenciado estable, así como la capacidad de mineralizar la matriz extracelular y expresar un fenotipo similar al osteoblasto durante la inducción osteogénica. Los tres marcadores evaluados se sobre-expresaron en los cultivos en condiciones osteogénicas, y se encontró que cambios hasta de 2X en sus niveles de expresión son relevantes para el desarrollo del proceso de diferenciación. El modelo de hBMSCS presentado podría ser utilizado para la evaluación in vitro de la osteoinductividad de diferentes biomateriales, moléculas bioactivas o matrices para ingeniería de tejidos.Human bone marrow mesenchymal stem cells (hBMSCs) comprise a cell population capable of self-renewal and multilineage differentiation commonly isolated from bone marrow aspirates of large bones. Their osteogenic potential has been extensively exploited for the biological evaluation of scaffolds or biomaterials with applications in bone tissue engineering. This work aimed to isolate hBMSCs from femoral heads of patients undergoing total hip arthroplasty and to evaluate their osteogenic potential. Briefly, the trabecular bone was extracted and mechanically disaggregated; the released cells were cultured and non-adherent cells were removed after 4 days. The osteogenic potential was evaluated at the fifth passage after 14 and 20 days of induction, comparing cultures with and without osteogenic supplements, via Alizarin red staining and the quantification of the gene expression levels of the osteogenic markers collagen type I, osteonectin and bone sialoprotein through real-time RT-PCR. The obtained hBMSCs presented a stable undifferentiated phenotype after prolonged cell culture, matrix mineralization capabilities and expression of osteoblast phenotype upon osteogenic induction. The three markers were up-regulated in cultures under osteogenic conditions and 2 fold differences in their expression levels were found to be significant for the onset of the differentiation process. The obtained hBMSCs may have applications on the in vitro evaluation of the osteoinductivity of different biomaterials, bioactive molecules or tissue engineering scaffolds

A versatile cancer cell trapping and 1D migration assay in a microfluidic device

Highly migratory cancer cells often lead to metastasis and recurrence and are responsible for the high mortality rates in many cancers despite aggressive treatment. Recently, the migratory behavior of patient-derived glioblastoma multiforme cells on microtracks has shown potential in predicting the likelihood of recurrence, while at the same time, antimetastasis drugs have been developed which require simple yet relevant high-throughput screening systems. However, robust in vitro platforms which can reliably seed single cells and measure their migration while mimicking the physiological tumor microenvironment have not been demonstrated. In this study, we demonstrate a microfluidic device which hydrodynamically seeds single cancer cells onto stamped or femtosecond laser ablated polystyrene microtracks, promoting 1D migratory behavior due to the cells' tendency to follow topographical cues. Using time-lapse microscopy, we found that single U87 glioblastoma multiforme cells migrated more slowly on laser ablated microtracks compared to stamped microtracks of equal width and spacing (p < 0.05) and exhibited greater directional persistence on both 1D patterns compared to flat polystyrene (p < 0.05). Single-cell morphologies also differed significantly between flat and 1D patterns, with cells on 1D substrates exhibiting higher aspect ratios and less circularity (p < 0.05). This microfluidic platform could lead to automated quantification of single-cell migratory behavior due to the high predictability of hydrodynamic seeding and guided 1D migration, an important step to realizing the potential of microfluidic migration assays for drug screening and individualized medicine. Published under license by AIP Publishing

Quantum confined peptide assemblies with tunable visible to near-infrared spectral range

Quantum confined materials have been extensively studied for photoluminescent applications. Due to intrinsic limitations of low biocompatibility and challenging modulation, the utilization of conventional inorganic quantum confined photoluminescent materials in bio-imaging and bio-machine interface faces critical restrictions. Here, we present aromatic cyclo-dipeptides that dimerize into quantum dots, which serve as building blocks to further self-assemble into quantum confined supramolecular structures with diverse morphologies and photoluminescence properties. Especially, the emission can be tuned from the visible region to the near-infrared region (420 nm to 820 nm) by modulating the self-assembly process. Moreover, no obvious cytotoxic effect is observed for these nanostructures, and their utilization for in vivo imaging and as phosphors for light-emitting diodes is demonstrated. The data reveal that the morphologies and optical properties of the aromatic cyclo-dipeptide self-assemblies can be tuned, making them potential candidates for supramolecular quantum confined materials providing biocompatible alternatives for broad biomedical and opto-electric applications

Investigation of hospital discharge cases and SARS-CoV-2 introduction into Lothian care homes

Background The first epidemic wave of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in Scotland resulted in high case numbers and mortality in care homes. In Lothian, over one-third of care homes reported an outbreak, while there was limited testing of hospital patients discharged to care homes. Aim To investigate patients discharged from hospitals as a source of SARS-CoV-2 introduction into care homes during the first epidemic wave. Methods A clinical review was performed for all patients discharges from hospitals to care homes from 1st March 2020 to 31st May 2020. Episodes were ruled out based on coronavirus disease 2019 (COVID-19) test history, clinical assessment at discharge, whole-genome sequencing (WGS) data and an infectious period of 14 days. Clinical samples were processed for WGS, and consensus genomes generated were used for analysis using Cluster Investigation and Virus Epidemiological Tool software. Patient timelines were obtained using electronic hospital records. Findings In total, 787 patients discharged from hospitals to care homes were identified. Of these, 776 (99%) were ruled out for subsequent introduction of SARS-CoV-2 into care homes. However, for 10 episodes, the results were inconclusive as there was low genomic diversity in consensus genomes or no sequencing data were available. Only one discharge episode had a genomic, time and location link to positive cases during hospital admission, leading to 10 positive cases in their care home. Conclusion The majority of patients discharged from hospitals were ruled out for introduction of SARS-CoV-2 into care homes, highlighting the importance of screening all new admissions when faced with a novel emerging virus and no available vaccine

SARS-CoV-2 Omicron is an immune escape variant with an altered cell entry pathway

Vaccines based on the spike protein of SARS-CoV-2 are a cornerstone of the public health response to COVID-19. The emergence of hypermutated, increasingly transmissible variants of concern (VOCs) threaten this strategy. Omicron (B.1.1.529), the fifth VOC to be described, harbours multiple amino acid mutations in spike, half of which lie within the receptor-binding domain. Here we demonstrate substantial evasion of neutralization by Omicron BA.1 and BA.2 variants in vitro using sera from individuals vaccinated with ChAdOx1, BNT162b2 and mRNA-1273. These data were mirrored by a substantial reduction in real-world vaccine effectiveness that was partially restored by booster vaccination. The Omicron variants BA.1 and BA.2 did not induce cell syncytia in vitro and favoured a TMPRSS2-independent endosomal entry pathway, these phenotypes mapping to distinct regions of the spike protein. Impaired cell fusion was determined by the receptor-binding domain, while endosomal entry mapped to the S2 domain. Such marked changes in antigenicity and replicative biology may underlie the rapid global spread and altered pathogenicity of the Omicron variant

Microfabricated nanochannels: New tools for molecular motion control

This paper presents a review of work on the fabrication and use of nanochannels in silicon and polymers for the control of molecular transport. The method of Sacrificial Layer Lithography is reviewed and demonstrated for silicon and polymers. A novel technique for the productions of conical nanopores through a polymer membrane is also reviewed. Nanochannels and nanopores have many potential applications for drug delivery, immunoprotection of cell implants, blocking of globular proteins from biosensor surfaces, and diagnostic devices. All of these applications benefit from the more direct interactions of devices with biomolecules.El presente trabajo presenta una revisión literaria sobre los métodos de fabricación de nanocanales en silicio y diferentes materiales poliméricos; y su uso en control de transporte molecular. Se describe el método "Sacrificial Layer Lithography" para silicio y polímeros. Adicionalmente, una novedosa técnica para la producción de nanoporos cónicos a través de una membrana polimérica es descrita. Los nanocanales y los nanoporos poseen diversas aplicaciones potenciales en la liberación de drogas, en la inmunoprotección de implantes celulares, el bloqueo de proteínas globulares en la superficie de biosensores, y en dispositivos para diagnóstico. Todas estas aplicaciones se benefician de la interacción directa entre los dispositivos y las biomoléculas