Preparation of exosomes for siRNA delivery to cancer cells

Extracellular vesicles, in particular exosomes, have recently gained interest as novel drug delivery vectors due to their biological origin, abundance, and intrinsic capability in intercellular delivery of various biomolecules. This work establishes an isolation protocol to achieve high yield and high purity of exosomes for siRNA delivery. Human Embryonic Kidney cells (HEK-293 cells) are cultured in bioreactor flasks and the culture supernatant (hereon referred to as conditioned medium) is harvested on a weekly basis to allow for enrichment of HEK-293 exosomes. The conditioned medium (CM) is pre-cleared of dead cells and cellular debris by differential centrifugation and is subjected to ultracentrifugation onto a sucrose cushion followed by a washing step, to collect the exosomes. Isolated HEK-293 exosomes are characterized for yield, morphology and exosomal marker expression by nanoparticle tracking analysis, protein quantification, electron microscopy and flow cytometry, respectively. Small interfering RNA (siRNA), fluorescently labeled with Atto655, is loaded into exosomes by electroporation and excess siRNA is removed by gel filtration. Cell uptake in PANC-1 cancer cells, after 24 h incubation at 37 °C, is confirmed by flow cytometry. HEK-293 exosomes are 107.0 ± 8.2 nm in diameter. The exosome yield and particle-to-protein ratio (P:P) ratio are 6.99 ± 0.22 × 1012 particle/mL and 8.3 ± 1.7 × 1010 particle/µg, respectively. The encapsulation efficiency of siRNA in exosomes is ~ 10-20%. Forty percent of the cells show positive signals for Atto655 at 24 h post-incubation. In conclusion, exosome isolation by ultracentrifugation onto sucrose cushion offers a combination of good yield and purity. siRNA could be successfully loaded into exosomes by electroporation and subsequently delivered into cancer cells in vitro. This protocol offers a standard procedure for developing siRNA-loaded exosomes for efficient delivery to cancer cells

The chromosome content and genotype of two wheat cell lines and of their somatic fusion product with oat

Somatic hybridization seeks to genetically combine phylogenetically distant parents. An effective system has been established in bread wheat (Triticum aestivum L.) involving protoplasts from a non-totipotent cell line adapted to in vitro culture (T1) in combination with totipotent protoplasts harvested from embryogenic calli (T2). Here, we report the karyotype and genotype of T1 and T2. Line T1 carries nine A (A-genome of wheat), seven B (B-genome of wheat) and eight D (D-genome of wheat) genome chromosomes, while T2 cells have 12 A, 10 B and 12 D genome chromosomes. Rates of chromosome aberration in the B- and D-genomes were more than 25%, higher than in the A-genome. DNA deletion rates were 55.6% in T1 and 19.4% in T2, and DNA variation rates were 8.3% in T1 and 13.9% in T2. Rate of DNA elimination was B- > D- > A-genome in both T1 and T2. The same set of cytological and genetic assays was applied to a derivative of the somatic fusion between protoplasts of T1, T2 and oat (Avena sativa L.). The regenerant plants were near euploid with respect to their wheat complement. Six wheat chromosome arms—4AL, 3BS, 4BL, 3DS, 6DL and 7DL—carried small introgressed segments of oat chromatin. A genotypic analysis of the hybrid largely confirmed this cytologically-based diagnosis

Acute rejection is associated with antibodies to non-Gal antigens in baboons using Gal-knockout pig kidneys

We transplanted kidneys from α1,3-galactosyltransferase knockout (GalT-KO) pigs into six baboons using two different immunosuppressive regimens, but most of the baboons died from severe acute humoral xenograft rejection. Circulating induced antibodies to non-Gal antigens were markedly elevated at rejection, which mediated strong complement-dependent cytotoxicity against GalT-KO porcine target cells. These data suggest that antibodies to non-Gal antigens will present an additional barrier to transplantation of organs from GalT-KO pigs to humans. © 2005 Nature Publishing Group

Active dielectric antenna on chip for spatial light modulation

Integrated photonic resonators are widely used to manipulate light propagation in an evanescently-coupled waveguide. While the evanescent coupling scheme works well for planar optical systems that are naturally waveguide based, many optical applications are free-space based, such as imaging, display, holographics, metrology and remote sensing. Here we demonstrate an active dielectric antenna as the interface device that allows the large-scale integration capability of silicon photonics to serve the free-space applications. We show a novel perturbation-base diffractive coupling scheme that allows a high-Q planer resonator to directly interact with and manipulate free-space waves. Using a silicon-based photonic crystal cavity whose resonance can be rapidly tuned with a p-i-n junction, a compact spatial light modulator with an extinction ratio of 9.5 dB and a modulation speed of 150 MHz is demonstrated. Method to improve the modulation speed is discussed.Air Force Office of Scientific Research (AFOSR grant FA9550-12-1-0261

A high density genetic map of tobacco (Nicotiana tabacum L.) obtained from large scale microsatellite marker development

Tobacco (Nicotiana tabacum L.) is a species in the large family of the Solanaceae and is important as an agronomic crop and as a model system in plant biotechnology. Despite its importance, only limited molecular marker resources are available that can be used for genome analysis, genetic mapping and breeding. We report here on the development and characterization of 5,119 new and functional microsatellite markers and on the generation of a high-resolution genetic map for the tetraploid tobacco genome. The genetic map was generated using an F2 mapping population derived from the intervarietal cross of Hicks Broadleaf × Red Russian and merges the polymorphic markers from this new set with those from a smaller set previously used to produce a lower density map. The genetic map described here contains 2,317 microsatellite markers and 2,363 loci, resulting in an average distance between mapped microsatellite markers which is less than 2 million base pairs or 1.5 cM. With this new and expanded marker resource, a sufficient number of markers are now available for multiple applications ranging from tobacco breeding to comparative genome analysis. The genetic map of tobacco is now comparable in marker density and resolution with the best characterized genomes of the Solanaceae: tomato and potato

Dual-gated bilayer graphene hot electron bolometer

Detection of infrared light is central to diverse applications in security, medicine, astronomy, materials science, and biology. Often different materials and detection mechanisms are employed to optimize performance in different spectral ranges. Graphene is a unique material with strong, nearly frequency-independent light-matter interaction from far infrared to ultraviolet, with potential for broadband photonics applications. Moreover, graphene's small electron-phonon coupling suggests that hot-electron effects may be exploited at relatively high temperatures for fast and highly sensitive detectors in which light energy heats only the small-specific-heat electronic system. Here we demonstrate such a hot-electron bolometer using bilayer graphene that is dual-gated to create a tunable bandgap and electron-temperature-dependent conductivity. The measured large electron-phonon heat resistance is in good agreement with theoretical estimates in magnitude and temperature dependence, and enables our graphene bolometer operating at a temperature of 5 K to have a low noise equivalent power (33 fW/Hz1/2). We employ a pump-probe technique to directly measure the intrinsic speed of our device, >1 GHz at 10 K.Comment: 5 figure

UDBMS : Road to Unification for Multi-model Data Management

One of the greatest challenges in big data management is the “Variety” of the data. The data may be presented in various types and formats: structured, semi-structured and unstructured. For instance, data can be modeled as relational, key-value, and graph models. Having a single data platform for managing both well-structured data and NoSQL data is beneficial to users; this approach reduces significantly integration, migration, development, maintenance, and operational issues. Therefore, a challenging research work is how to develop an efficient consolidated single data management platform covering both NoSQL and relational data to reduce integration issues, simplify operations, and eliminate migration issues. In this paper, we envision novel principles and technologies to handle multiple models of data in one unified database system, including model-agnostic storage, unified query processing and indexes, in-memory structures and multi-model transactions. We discuss our visions as well as present research challenges that we need to address.Peer reviewe

A Pair of Dopamine Neurons Target the D1-Like Dopamine Receptor DopR in the Central Complex to Promote Ethanol-Stimulated Locomotion in Drosophila

Dopamine is a mediator of the stimulant properties of drugs of abuse, including ethanol, in mammals and in the fruit fly Drosophila. The neural substrates for the stimulant actions of ethanol in flies are not known. We show that a subset of dopamine neurons and their targets, through the action of the D1-like dopamine receptor DopR, promote locomotor activation in response to acute ethanol exposure. A bilateral pair of dopaminergic neurons in the fly brain mediates the enhanced locomotor activity induced by ethanol exposure, and promotes locomotion when directly activated. These neurons project to the central complex ellipsoid body, a structure implicated in regulating motor behaviors. Ellipsoid body neurons are required for ethanol-induced locomotor activity and they express DopR. Elimination of DopR blunts the locomotor activating effects of ethanol, and this behavior can be restored by selective expression of DopR in the ellipsoid body. These data tie the activity of defined dopamine neurons to D1-like DopR-expressing neurons to form a neural circuit that governs acute responding to ethanol

Graphene Photonics and Optoelectronics

The richness of optical and electronic properties of graphene attracts enormous interest. Graphene has high mobility and optical transparency, in addition to flexibility, robustness and environmental stability. So far, the main focus has been on fundamental physics and electronic devices. However, we believe its true potential to be in photonics and optoelectronics, where the combination of its unique optical and electronic properties can be fully exploited, even in the absence of a bandgap, and the linear dispersion of the Dirac electrons enables ultra-wide-band tunability. The rise of graphene in photonics and optoelectronics is shown by several recent results, ranging from solar cells and light emitting devices, to touch screens, photodetectors and ultrafast lasers. Here we review the state of the art in this emerging field.Comment: Review Nature Photonics, in pres

Maternal care in rural China: a case study from Anhui province

<p>Abstract</p> <p>Background</p> <p>Studies on prenatal care in China have focused on the timing and frequency of prenatal care and relatively little information can be found on how maternal care has been organized and funded or on the actual content of the visits, especially in the less developed rural areas. This study explored maternal care in a rural county from Anhui province in terms of care organization, provision and utilization.</p> <p>Methods</p> <p>A total of 699 mothers of infants under one year of age were interviewed with structured questionnaires; the county health bureau officials and managers of township hospitals (n = 10) and county level hospitals (n = 2) were interviewed; the process of the maternal care services was observed by the researchers. In addition, statistics from the local government were used.</p> <p>Results</p> <p>The county level hospitals were well staffed and equipped and served as a referral centre for women with a high-risk pregnancy. Township hospitals had, on average, 1.7 midwives serving an average population of 15,000 people. Only 10–20% of the current costs in county level hospitals and township hospitals were funded by the local government, and women paid for delivery care. There was no systematic organized prenatal care and referrals were not mandatory. About half of the women had their first prenatal visit before the 13th gestational week, 36% had fewer than 5 prenatal visits, and about 9% had no prenatal visits. A major reason for not having prenatal care visits was that women considered it unnecessary. Most women (87%) gave birth in public health facilities, and the rest in a private clinic or at home. A total of 8% of births were delivered by caesarean section. Very few women had any postnatal visits. About half of the women received the recommended number of prenatal blood pressure and haemoglobin measurements.</p> <p>Conclusion</p> <p>Delivery care was better provided than both prenatal and postnatal care in the study area. Reliance on user fees gave the hospitals an incentive to put more emphasis on revenue generating activities such as delivery care instead of prenatal and postnatal care.</p