Microfluidic multipoles: theory and applications

Microfluidic multipoles (MFMs) have been realized experimentally and hold promise for "open-space" biological and chemical surface processing. Whereas convective flow can readily be predicted using hydraulic-electrical analogies, the design of advanced MFMs is constrained by the lack of simple, accurate models to predict mass transport within them. In this work, we introduce the first exact solutions to mass transport in multipolar microfluidics based on the iterative conformal mapping of 2D advection-diffusion around a simple edge into dipoles and multipolar geometries, revealing a rich landscape of transport modes. The models were validated experimentally with a library of 3D printed MFM devices and found in excellent agreement. Following a theory-guided design approach, we further ideated and fabricated two new classes of spatiotemporally reconfigurable MFM devices that are used for processing surfaces with time-varying reagent streams, and to realize a multistep automated immunoassay. Overall, the results set the foundations for exploring, developing, and applying open-space MFMs.Comment: 16 pages, 5 figure

An environmental signature for 323 microbial genomes based on codon adaptation indices

BACKGROUND: Codon adaptation indices (CAIs) represent an evolutionary strategy to modulate gene expression and have widely been used to predict potentially highly expressed genes within microbial genomes. Here, we evaluate and compare two very different methods for estimating CAI values, one corresponding to translational codon usage bias and the second obtained mathematically by searching for the most dominant codon bias. RESULTS: The level of correlation between these two CAI methods is a simple and intuitive measure of the degree of translational bias in an organism, and from this we confirm that fast replicating bacteria are more likely to have a dominant translational codon usage bias than are slow replicating bacteria, and that this translational codon usage bias may be used for prediction of highly expressed genes. By analyzing more than 300 bacterial genomes, as well as five fungal genomes, we show that codon usage preference provides an environmental signature by which it is possible to group bacteria according to their lifestyle, for instance soil bacteria and soil symbionts, spore formers, enteric bacteria, aquatic bacteria, and intercellular and extracellular pathogens. CONCLUSION: The results and the approach described here may be used to acquire new knowledge regarding species lifestyle and to elucidate relationships between organisms that are far apart evolutionarily

Fiber-based tissue engineering: Progress, challenges, and opportunities.

Tissue engineering aims to improve the function of diseased or damaged organs by creating biological substitutes. To fabricate a functional tissue, the engineered construct should mimic the physiological environment including its structural, topographical, and mechanical properties. Moreover, the construct should facilitate nutrients and oxygen diffusion as well as removal of metabolic waste during tissue regeneration. In the last decade, fiber-based techniques such as weaving, knitting, braiding, as well as electrospinning, and direct writing have emerged as promising platforms for making 3D tissue constructs that can address the abovementioned challenges. Here, we critically review the techniques used to form cell-free and cell-laden fibers and to assemble them into scaffolds. We compare their mechanical properties, morphological features and biological activity. We discuss current challenges and future opportunities of fiber-based tissue engineering (FBTE) for use in research and clinical practice

Nutritional values of forage-legume-based silages and protein concentrates for growing pigs

In organic pig production systems, one of the main challenges is to meet the demand for resources rich in protein. Among the resources available, temperate green plants, such as forage legumes, are potential sources of energy and protein. The aim of the study was to determine the nutritional value of silages (S) from the whole plant of lucerne (L) and red clover (R) and protein pastes (PPs) obtained from L and R leaves. In a first trial, 30 pigs were used in a factorial design to determine the total tract digestibility (TTD) of dietary nutrients and energy in five dietary treatments. The control group was fed a control diet (C1). The lucerne silage (LS) and red clover silage (RS) groups were fed a 78%:22% mixture (on a DM basis) of the C1 diet and LS or RS. The lucerne protein paste (LPP) and the red clover protein paste (RPP) groups were fed an 81%:19% mixture (on a DM basis) of the C1 diet and LPP or RPP. In the second trial, five pigs were used in a 5 × 5 Latin square design to evaluate the standardised ileal digestibility (SID) of amino acids (AAs) in the four legume products. The control diet (C2) was formulated with casein as the sole protein source. The LS and RS groups were fed an 85%:15% mixture (on a DM basis) of the C2 diet and LS or RS. The LPP and RPP groups were fed an 80%:20% mixture (on a DM basis) of the C2 diet and LPP or RPP. Regardless of the plant species, silages obtained from L and R leaves contained less AA and more fibre than protein pastes. While the fresh forages contained the same percentage of protein N in total N (63.6%), lucerne lost more protein N during ensiling than red clover (−75.5 vs −33.8%). The calculated TTD coefficient of energy was higher in silages than in protein pastes and lower in R than in L products (72.8, 71.5, 67.7, and 61.3 for LS, RS, LPP and RPP, respectively). The SID of total essential AA was higher in LPP than in RPP (87.2 vs 79.2%) whereas it was lower in LS than in RS (33.2 vs 56.8%). The lower SID values in silages were explained by the protein degradation during the ensiling process and a high proportion of AA linked to the NDF fraction. The results of the present study show that protein pastes obtained from lucerne and red clover are valuable protein sources for pig. In contrast, legume silages have to be considered as an energy source rather than a protein source.publishedVersio

Two-Aperture Microfluidic Probes as Flow Dipoles: Theory and Applications

A microfluidic probe (MFP) is a mobile channel-less microfluidic system under which a fluid is injected from an aperture into an open space, hydrodynamically confined by a surrounding fluid, and entirely re-aspirated into a second aperture. Various MFPs have been developed, and have been used for applications ranging from surface patterning of photoresists to local perfusion of organotypic tissue slices. However, the hydrodynamic and mass transfer properties of the flow under the MFP have not been analyzed, and the flow parameters are adjusted empirically. Here, we present an analytical model describing the key transport properties in MFP operation, including the dimensions of the hydrodynamic flow confinement (HFC) area, diffusion broadening, and shear stress as a function of: (i) probe geometry (ii) aspiration-to-injection flow rate ratio (iii) gap between MFP and substrate and (iv) reagent diffusivity. Analytical results and scaling laws were validated against numerical simulations and experimental results from published data. These results will be useful to guide future MFP design and operation, notably to control the MFP "brush stroke" while preserving shear-sensitive cells and tissues

Protein Co-Enrichment Analysis of Extracellular Vesicles

Extracellular Vesicles (EVs) carry cell-derived proteins that confer functionality and selective cell uptake. However, whether proteins are packaged stochastically or co-enriched within individual EVs, and whether co-enrichment fluctuates under homeostasis and disease, has not been measured. EV abundance and protein global relative expression have been qualified by bulk analysis. Meanwhile, co-enrichment is not directly accessible via bulk measurement and has not been reported for single EV analysis. Here, we introduce the normalized index of co-enrichment (NICE) to measure protein co-enrichment. NICE was derived by (i) capturing EVs based on the expression of a membrane-bound protein, (ii) probing for the co-expression of a second protein at the population level - EV integrity underwrites the detection of single EV co-expression without the need to resolve single EVs - and (iii) normalizing measured values using two universal normalization probes. Axiomatically, NICE = 1 for stochastic inclusion or no overall co-enrichment, while for positive and negative co-enrichment NICE > 1 or < 1, respectively. We quantified the NICE of tetraspanins, growth factor receptors and integrins in EVs of eight breast cancer cell lines of varying metastatic potential and organotropism, combinatorially mapping up to 104 protein pairs. Our analysis revealed protein enrichment and co-expression patterns consistent with previous findings. For the organotropic cell lines, most protein pairs were co-enriched on EVs, with the majority of NICE values between 0.2 to 11.5, and extending from 0.037 to 80.4. Median NICE were either negative, neutral or positive depending on the cells. NICE analysis is easily multiplexed and is compatible with microarrays, bead-based and single EV assays. Additional studies are needed to deepen our understanding of the potential and significance of NICE for research and clinical uses

The viral transcription group determines the HLA class I cellular immune response against human respiratory syncytial virus

The cytotoxic T-lymphocyte-mediated killing of virus-infected cells requires previous recognition of short viral antigenic peptides bound to human leukocyte antigen class I molecules that are exposed on the surface of infected cells. The cytotoxic T-lymphocyte response is critical for the clearance of human respiratory syncytial virus infection. In this study, naturally processed viral human leukocyte antigen class I ligands were identified with mass spectrometry analysis of complex human leukocyte antigen-bound peptide pools isolated from large amounts of human respiratory syncytial virus-infected cells. Acute antiviral T-cell response characterization showed that viral transcription determines both the immunoprevalence and immunodominance of the human leukocyte antigen class I response to human respiratory syncytial virus. These findings have clear implications for antiviral vaccine design.This work was supported by the Spanish Ministry of Economy grants BIO2011-25636 to D.L. and to A. A. from the ISF 916/05. The funding agencies had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. We have no conflicting financial interestsS

Substrate Micropatterning as a New in Vitro Cell Culture System to Study Myelination

Artículo de publicación ISIMyelination is a highly regulated developmental process whereby oligodendrocytes in the central nervous system and Schwann cells in the peripheral nervous system ensheathe axons with a multilayered concentric membrane. Axonal myelination increases the velocity of nerve impulse propagation. In this work, we present a novel in vitro system for coculturing primary dorsal root ganglia neurons along with myelinating cells on a highly restrictive and micropatterned substrate. In this new coculture system, neurons survive for several weeks, extending long axons on defined Matrigel tracks. On these axons, myelinating cells can achieve robust myelination, as demonstrated by the distribution of compact myelin and nodal markers. Under these conditions, neurites and associated myelinating cells are easily accessible for studies on the mechanisms of myelin formation and on the effects of axonal damage on the myelin sheath.Regenerative Medicine and Nanomedicine Initiative of the Canadian Institutes of Health Research (CIHR) RMF-7028 FONDECYT 1080252 CIHR Ministry of Industry of Canada Rio Tinto Alcan Molson Foundatio

In vitro analysis of the effects on wound healing of high- and low-molecular weight chains of hyaluronan and their hybrid H-HA/L-HA complexes

Abstract Background: Recent studies have reported the roles of Hyaluronic acid (HA) chains of diverse length in wound repair, especially considering the simultaneous occurrence in vivo of both high- (H-HA) and low-molecular weight (L-HA) hyaluronan at an injury site. It has been shown that HA fragments (5 ≤ MW ≤ 20 kDa) usually trigger an inflammatory response that, on one hand, is the first signal in the activation of a repair mechanism but on the other, when it’s overexpressed, it may promote unwanted side effects. The present experimental research has aimed to investigate H-HA, L-HA and of a newly developed complex of the two (H-HA/L-HA) for stability (e.g. hyaluronidases digestion), for their ability to promote wound healing of human keratinocytes in vitro and for their effect on cellular biomarker expression trends. Results: Time-lapse video microscopy studies proved that the diverse HA was capable of restoring the monolayer integrity of HaCat. The H-HA/L-HA complex (0.1 and 1%w/v) proved faster in regeneration also in co-culture scratch test where wound closure was achieved in half the time of H-HA stimulated cells and 2.5-fold faster than the control. Gene expression was evaluated for transformation growth factor beta 1 (TGF-β1) proving that L-HA alone increased its expression at 4 h followed by restoration of similar trends for all the stimuli. Depending on the diverse stimulation (H-HA, L-HA or the complex), metalloproteinases (MMP-2, -9, -13) were also modulated differently. Furthermore, type I collagen expression and production were evaluated. Compared to the others, persistence of a significant higher expression level at 24 h for the H-HA/L-HA complex was found. Conclusions: The outcomes of this research showed that, both at high and low concentrations, hybrid complexes proved to perform better than HA alone thus suggesting their potential as medical devices in aesthetic and regenerative medicine. Keywords: Wound healing, Hyaluronan, MMPs, Hybrid complexe