2,177 research outputs found

    Scalable, biofunctional, ultra-stable nano- bio- composite materials containing living cells

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    Three-dimensional encapsulation of cells within nanostructured silica gels or matrices enables applications as diverse as biosensors, microbial fuel cells, artificial organs, and vaccines. It also allows study of individual cell behaviors. Recent progress has improved the performance and flexibility of cellular encapsulation, yet there remains a need for robust scalable processes for large format production of cell-encapsulating materials. Here, we detail two novel techniques, that enable the large-scale production of functional Nano-Bio-Composites (NBCs) containing living cells within ordered 3-D lipid/silica nanostructures: 1) thick-casting and 2) spray drying. Furthermore, we detail a third technique for material scaling in which aqueous, silicate-based gel monoliths encapsulate biofunctional yeast or bacteria. Both dry processes are demonstrated to work with multiple cell types and result in dry powders exhibiting a unique combination of properties including: highly ordered 3-D nanostructure, extended lipid fluidity, tunable macro-morphologies and aerodynamic diameters, and unexpectedly high physical strength. Nanoindentation of the encasing nanostructure revealed Young’s modulus and hardness of 13 and 1.4 GPa respectively, which was unexpected considering the low processing conditions. We hypothesized and confirmed that NBC-encapsulated cells would remain viable for extended periods of time under elevated aging conditions. We attribute this due to the high material strength as observed with nanoindentation, which would prevent cell growth and force bacteria into viable but not culturable (VBNC) states. In concordance with the VBNC state, cellular ATP levels remained elevated even over eight months confirming temperature stable, viable cells. However, their ability to undergo resuscitation and enter growth phase greatly decreased with time in the VBNC state. A quantitative method of determining resuscitation frequencies was developed and showed that, after 36 weeks in an NBC-induced VBNC state, less than 1 in 10,000 cells underwent resuscitation. We verify the VBNC phenotype in gel-encapsulated cells by studying cellular RNA expression levels. These latent behaviors are further demonstrated with an in-vivo immunological study in which mice, immunized with NBCs containing the vaccine Bacillus Calmette-Guérin, were observed to be immunized against a latent form of Tuberculosis. This finding is, in our understanding, the first demonstration of a latent disease-specific live cell immunotherapy. The NBC platform production of industrially scalable quantities of VBNC cells is of interest for research in bacterial persistence and screening of drugs targeting such cells. NBC’s may also enable long-term preservation of living cells for applications in cell-based sensing and the packaging and delivery of live-cell vaccines. Moreover, our methodology represents a novel process for preparing formulations of latent cells in-silico, which could find application in basic cellular research and for the development of a latent-specific vaccine

    Diffusion measurements to understand dynamics and structuring in solutions involving a homologous series of ionic liquids

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    The self-diffusion coefficients of each of the components in mixtures containing pyridine and each of the homologous series 1-alkyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imides in acetonitrile were determined using NMR diffusometry (i. e., Pulsed Gradient Spin Echo). The nature of solvation was found to change significantly with the proportion of salt in the mixtures. Increased diffusion coefficients (when corrected for viscosity) for the molecular components were observed with increasing proportion of ionic liquid and with increasing alkyl chain length on the cation. Comparison of the molecular solvents suggests increased interactions in solution of the pyridine with other components of the mixture, consistent with the proposed interactions shown previously to drive changes in reaction kinetics. Discontinuities were seen in the diffusion data for each species in solution across different ionic liquids between the hexyl and octyl derivatives, suggesting a change in the structuring in solution as the alkyl chain on the cation changes and demonstrating the importance of such when considering homologous series

    Medically relevant ElectroNeedle technology development.

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    ElectroNeedles technology was developed as part of an earlier Grand Challenge effort on Bio-Micro Fuel Cell project. During this earlier work, the fabrication of the ElectroNeedles was accomplished along with proof-of-concept work on several electrochemically active analytes such as glucose, quinone and ferricyanide. Additionally, earlier work demonstrated technology potential in the field of immunosensors by specifically detecting Troponin, a cardiac biomarker. The current work focused upon fabrication process reproducibility of the ElectroNeedles and then using the devices to sensitively detect p-cresol, a biomarker for kidney failure or nephrotoxicity. Valuable lessons were learned regarding fabrication assurance and quality. The detection of p-cresol was accomplished by electrochemistry as well as using fluorescence to benchmark ElectroNeedles performance. Results from these studies will serve as a guide for the future fabrication processes involving ElectroNeedles as well as provide the groundwork necessary to expand technology applications. One paper has been accepted for publication acknowledging LDRD funding (K. E. Achyuthan et al, Comb. Chem. & HTS, 2008). We are exploring the scope for a second paper describing the applications potential of this technology

    Nature versus nurture in cellular behavior and disease.

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    Augmentation of Recipient Adaptive Alloimmunity by Donor Passenger Lymphocytes within the Transplant.

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    Chronic rejection of solid organ allografts remains the major cause of transplant failure. Donor-derived tissue-resident lymphocytes are transferred to the recipient during transplantation, but their impact on alloimmunity is unknown. Using mouse cardiac transplant models, we show that graft-versus-host recognition by passenger donor CD4 T cells markedly augments recipient cellular and humoral alloimmunity, resulting in more severe allograft vasculopathy and early graft failure. This augmentation is enhanced when donors were pre-sensitized to the recipient, is dependent upon avoidance of host NK cell recognition, and is partly due to provision of cognate help for allo-specific B cells from donor CD4 T cells recognizing B cell MHC class II in a peptide-degenerate manner. Passenger donor lymphocytes may therefore influence recipient alloimmune responses and represent a therapeutic target in solid organ transplantation.This work was supported by a British Heart Foundation project grant, the National Institute of Health Research Cambridge Biomedical Research Centre and the National Institute of Health Research Blood and Transplant Research Unit. IGH and JMA were supported by a Wellcome Trust Clinical Research Training Fellowship and Raymond and Beverly Sackler Scholarship. KSP was supported by an Academy of Medical Sciences / Wellcome Trust starter grant.This is the final version of the article. It first appeared from Elsevier via https://doi.org/10.1016/j.celrep.2016.04.00

    Effect of multi-planar CT image reformatting on surgeon diagnostic performance for localizing thoracolumbar disc extrusions in dogs

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    Accurate pre-operative localization and removal of disc material are important for minimizing morbidity in dogs with thoracolumbar disc extrusions. Computed tomography (CT) is an established technique for localizing disc extrusions in dogs, however the effect of multi-planar reformatting (MPR) on surgeon diagnostic performance has not been previously described. The purpose of this study was to test the effect of MPR CT on surgeon diagnostic accuracy, certainty and agreement for localizing thoracolumbar disc extrusions in dogs. Two veterinary surgeons and one veterinary neurologist who were unaware of surgical findings independently reviewed randomized sets of two-dimensional (2D) and MPR CT images from 111 dogs with confirmed thoracolumbar disc extrusions. For each set of images, readers recorded their localizations for extruded disc material and their diagnostic certainty. For MPR images, readers also recorded views they considered most helpful. Diagnostic accuracy estimates, mean diagnostic certainty scores and inter-observer agreement were compared using surgery as the gold standard. Frequencies were compared for MPR views rated most helpful. Diagnostic accuracy estimates were significantly greater for MPR vs. 2D CT images in one reader. Mean diagnostic certainty scores were significantly greater for MPR images in two readers. The change in agreement between 2D and MPR images differed from zero for all analyses (site, side, number affected) among all three readers. Multi-planar views rated most helpful with the highest frequency were oblique transverse and curved dorsal planar MPR views. Findings from this study indicate that multi-planar CT can improve surgeon diagnostic performance for localizing canine thoracolumbar disc extrusions
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