Comparative analysis of mammalian Y chromosomes illuminates ancestral structure and lineage-specific evolution

Although more than thirty mammalian genomes have been sequenced to draft quality, very few of these include the Y chromosome. This has limited our understanding of the evolutionary dynamics of gene persistence and loss, our ability to identify conserved regulatory elements, as well our knowledge of the extent to which different types of selection act to maintain genes within this unique genomic environment. Here, we present the first MSY (male-specific region of the Y chromosome) sequences from two carnivores, the domestic dog and cat. By combining these with other available MSY data, our multiordinal comparison allows for the first accounting of levels of selection constraining the evolution of eutherian Y chromosomes. Despite gene gain and loss across the phylogeny, we show the eutherian ancestor retained a core set of 17 MSY genes, most being constrained by negative selection for nearly 100 million years. The X-degenerate and ampliconic gene classes are partitioned into distinct chromosomal domains in most mammals, but were radically restructured on the human lineage. We identified multiple conserved noncoding elements that potentially regulate eutherian MSY genes. The acquisition of novel ampliconic gene families was accompanied by signatures of positive selection and has differentially impacted the degeneration and expansion of MSY gene repertoires in different species.Y chromosomes have arisen independently in divergent evolutionary lineages across the eukaryotic tree of life (Rice 1996; Marin and Baker 1998; Liu et al. 2004; Graves 2006; Koerich et al. 2008; Carvalho et al. 2009; Kaiser and Bachtrog 2010). While many genes are known to be Y-linked, the actual number of Y chromosomes that have been sequenced is extremely small (Skaletsky et al. 2003; Hughes et al. 2005; Kuroki et al. 2006; Clark et al. 2007; Koerich et al. 2008; Carvalho et al. 2009; Hughes et al. 2010, 2012) in relation to the rapid rate at which whole genomes are presently being sequenced. This reduced emphasis on sequencing Y chromosomes can be primarily attributed to their presumed low gene content and large amounts of repetitive DNA, that is often arrayed into long stretches of nearly identical sequence which precludes the use of shotgun sequencing approaches to assemble Y chromosome sequence into large, contiguous scaffolds. These assembly problems are further exacerbated when applying short-read next-generation sequence methods (Alkan et al. 2011).The most completely sequenced and annotated Y chromosomes are from three recently diverged catarrhine primates: human, chimpanzee, and rhesus macaque (Skaletsky et al. 2003; Hughes et al. 2005, 2010, 2012; Kuroki et al. 2006 ). Comparisons between these species offer an important glimpse into the immense structural variation and complexity that can emerge on the Y within a very short period of evolutionary time. However, these three primate species last shared a common ancestor ∼21 million years ago (Mya) (Meredith et al. 2011) and thus offer limited comparative breadth to discriminate characteristics present across most mammalian Y chromosomes from idiosyncratic features reflective of a small evolutionary sample. This lack of phylogenetic scope has (1) hampered identification of the ancestral properties of eutherian Y chromosomes, (2) obscured broader patterns of evolutionary constraint and selection in a nonrecombining environment, and (3) hidden the frequency with which novel genes arise and/or acquire new functions in species with diverse phenotypes and reproductive strategies.To expand our knowledge of eutherian Y chromosome structure and gene function, we generated the first extensive MSY (male-specific Y chromosome) sequence from two members of the Carnivora: the domestic cat, Felis silvestris catus, and domestic dog, Canis lupus familiaris. These two carnivore genomes provide a phylogenetically distinct vantage point with which to interpret the evolutionary patterns observed in the primate MSY comparisons, diverging from each other ∼55 Mya, and from primates ∼92 Mya (Meredith et al. 2011). Our combined analysis of two carnivore and three primate MSY sequences, together with physical mapping and functional sequence data from the mouse MSY, represent the first multiordinal comparison assessing deeper levels of evolutionary constraint. We also present a complete analysis of patterns of negative and positive selection to assess their effects on MSY degeneration and expansion.     

Enrichment of prostate cancer cells from blood cells with a hybrid dielectrophoresis and immunocapture microfluidic system

The isolation of circulating tumor cells (CTCs) from cancer patient blood is a technical challenge that is often addressed by microfluidic approaches. Two of the most prominent techniques for rare cancer cell separation, immunocapture and dielectrophoresis (DEP), are currently limited by a performance tradeoff between high efficiency and high purity. The development of a platform capable of these two performance criteria can potentially be facilitated by incorporating both DEP and immunocapture. We present a hybrid DEP-immunocapture system to characterize how DEP controls the shear-dependent capture of a prostate cancer cell line, LNCaP, and the nonspecific adhesion of peripheral blood mononuclear cells (PBMCs). Characterization of cell adhesion with and without DEP effects was performed in a Hele-Shaw flow cell that was functionalized with the prostate-specific monoclonal antibody, J591. In this model system designed to make nonspecific PBMC adhesion readily apparent, we demonstrate LNCaP enrichment from PBMCs by precisely tuning the applied AC electric field frequency to enhance immunocapture of LNCaPs and reduce nonspecific adhesion of PBMCs with positive and negative DEP, respectively. Our work shows that DEP and immunocapture techniques can work synergistically to improve cancer cell capture performance, and it informs the design of future hybrid DEP-immunocapture systems with improved CTC capture performance to facilitate research on cancer metastasis and drug therapies.     

Culture of primary rat hippocampal neurons: design, analysis, and optimization of a microfluidic device for cell seeding, coherent growth, and solute delivery

We present the design, analysis, construction, and culture results of a microfluidic device for the segregation and chemical stimulation of primary rat hippocampal neurons. Our device is designed to achieve spatio-temporal solute delivery to discrete sections of neurons with mitigated mechanical stress. We implement a geometric guidance technique to direct axonal processes of the neurons into specific areas of the device to achieve solute segregation along routed cells. Using physicochemical modeling, we predict flows, concentration profiles, and mechanical stresses within pertiment sections of the device. We demonstrate cell viability and growth within the closed device over a period of 11 days. Additionally, our modeling methodology may be generalized and applied to other device geometries.     

Circulating pro-inflammatory cytokines are elevated and peak power output correlates with 25-hydroxyvitamin D in vitamin D insufficient adults

The purpose of this study was to identify circulating cytokines, skeletal muscle strength, and peak power output in young adults with contrasting serum 25-hydroxyvitamin D (25(OH)D) concentrations. Serum 25(OH)D, inflammatory cytokines, muscle strength, and peak power output were, therefore, measured in young adults (25–42 years). Data were collected during the winter to avoid the seasonal influence on serum 25(OH)D. After serum 25(OH)D concentration measurements, subjects were separated into one of two groups: (1) vitamin D insufficient [serum 25(OH)D ≤32 ng/mL, n = 14], or (2) vitamin D sufficient [serum 25(OH)D >32 ng/mL, n = 14]. Following group allocation, serum 25(OH)D concentrations were significantly (p < 0.05) lower and pro-inflammatory cytokines [interleukin (IL)-2, IL-1β, tumor necrosis factor-α, and interferon-γ] were significantly (all p < 0.05) greater in vitamin D insufficient adults. An anti-inflammatory cytokine (i.e., IL-10; p > 0.05), peak isometric forces (p > 0.05), and peak power outputs (p > 0.05) were not significantly different between vitamin D groups. However, peak power outputs correlated with serum 25(OH)D concentrations in vitamin D insufficient (r = 0.55, p < 0.05) but not in vitamin D sufficient adults (r = ?0.27, p = 0.36). Based on these data, we conclude that vitamin D insufficiency, in part, could result in pro-inflammatory stress without altering muscular strength or function in young adults. Future research investigating the causality of the correlation between low-serum 25(OH)D and peak power output in young adults is required.     

Elucidating the role of nitrogen plasma composition in the low-temperature self-limiting growth of indium nitride thin films

In this work, we have studied the role varying nitrogen plasma compositions play in the low-temperature plasma-assisted growth of indium nitride (InN) thin films. Films were deposited on Si (100) substrates using a plasma-enhanced atomic layer deposition (PE-ALD) reactor featuring a capacitively-coupled hollow-cathode plasma source. Trimethylindium (TMI) and variants of nitrogen plasma (N₂-only, Ar/N₂, and Ar/N₂/H₂) were used as the metal precursor and nitrogen co-reactant, respectively. In situ ellipsometry was employed to observe individual ligand exchange and plasma-assisted ligand removal events in real-time during the growth process. Only the samples grown under hydrogen-free nitrogen plasmas (Ar/N₂ or N₂-only) resulted in nearly stoichiometric single-phase crystalline hexagonal InN (h-InN) films at substrate temperatures higher than 200 °C under 100 W rf-plasma power. Varying the plasma gas composition by adding H₂ led to rather drastic microstructural changes resulting in a cubic phase oxide (c-In₂O₃) film. Combining the in situ measured growth evolution with ex situ materials characterization, we propose a simplified model describing the possible surface reactions/groups during a unit PE-ALD cycle, which depicts the highly efficient oxygen incorporation in the presence of hydrogen radicals. Further structural, chemical, and optical characterization have been carried out on the optimal InN films grown with Ar/N₂ plasma to extract film properties. Samples grown at lower substrate temperature (160 °C) and reduced/elevated rf-plasma power levels (50/150 W) displayed similar amorphous character, which is attributed to either insufficient surface energy or plasma-induced crystal damage. InN samples grown at 240 °C under 100 W rf-plasma showed clear polycrystalline h-InN layers with ∼20 nm average-sized single crystal domains exhibiting hexagonal symmetry.

Hollow-cathode plasma-generated hydrogen radicals induce crystal phase transformation from h-InN to c-In₂O₃ during plasma-enhanced atomic layer deposition using trimethyl-indium and Ar/N₂ plasma.     

Infobuttons for Genomic Medicine: Requirements and Barriers

Objectives  The study aimed to understand potential barriers to the adoption of health information technology projects that are released as free and open source software (FOSS). Methods  We conducted a survey of research consortia participants engaged in genomic medicine implementation to assess perceived institutional barriers to the adoption of three systems: ClinGen electronic health record (EHR) Toolkit, DocUBuild, and MyResults.org. The survey included eight barriers from the Consolidated Framework for Implementation Research (CFIR), with additional barriers identified from a qualitative analysis of open-ended responses. Results  We analyzed responses from 24 research consortia participants from 18 institutions. In total, 14 categories of perceived barriers were evaluated, which were consistent with other observed barriers to FOSS adoption. The most frequent perceived barriers included lack of adaptability of the system, lack of institutional priority to implement, lack of trialability, lack of advantage of alternative systems, and complexity. Conclusion  In addition to understanding potential barriers, we recommend some strategies to address them (where possible), including considerations for genomic medicine. Overall, FOSS developers need to ensure systems are easy to trial and implement and need to clearly articulate benefits of their systems, especially when alternatives exist. Institutional champions will remain a critical component to prioritizing genomic medicine projects.     

Evaluation of the potential eye hazard at visible wavelengths of the supercontinuum generated by an ultrafast NIR laser in water

Lasers with ultrashort pulse durations have become ubiquitous in various applications, including ocular surgery. Therefore, we need to consider the role of nonlinear optical effects, such as supercontinuum generation during propagation within the ocular media, when evaluating their potential hazard. We used a NIR femtosecond laser to generate a supercontinuum within an artificial eye. We recorded the visible spectra of the supercontinuum generated and calculated the energy contained within the visible band. Our results indicate that for wavelengths between 1350 nm and 1450 nm the energy contained within the visible band of the generated white light supercontinuum may surpass current safety exposure limits, and pose a risk of injury to the retina.