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Hepatocellular carcinoma (HCC) is an aggressive disease with poor prognosis, necessitating preclinical models for evaluating novel therapies. Large-animal models are particularly valuable for assessing locoregional therapies, which are widely employed across HCC stages. This study aimed to develop a large-animal HCC model with tailored tumor mutations. The Oncopig, a genetically engineered pig with inducible TP53R167H and KRASG12D, was used in the study. Hepatocytes were isolated from Oncopigs and exposed to Cre recombinase in vitro to create HCC cells, and additional mutations were introduced by CRISPR/Cas9 knockout of PTEN and CDKN2A. These edits increased Oncopig HCC cell proliferation and migration. Autologous HCC cells with these CRISPR edits were implanted into Oncopigs using two approaches: ultrasound-guided percutaneous liver injections, which resulted in the development of localized intrahepatic masses, and portal vein injections, which led to multifocal tumors that regressed over time. Tumors developed by both approaches harbored PTEN and CDKN2A knockout mutations. This study demonstrates the feasibility of developing genetically tailored HCC tumors in Oncopigs using somatic cell CRISPR editing and autologous implantation, providing a valuable large-animal model for in vivo therapeutic assessment., Competing Interests: Competing interests L.E. reports grants from United States National Institutes of Health and the Society of Interventional Radiology. M.J.C. and L.R.J. report working for Sus Clinicals, Inc. L.B.S. and K.M.S. report grants from United States National Institutes of Health, United States Department of Agriculture, Society of Interventional Radiology, Guerbet USA LLC, Janssen Research & Development LLC, TriSalus Life Sciences, and Earli Inc.; work for Sus Clinicals, Inc.; and pending patents for Modeling Oncology on Demand, United States Provisional Patent Application No. 62/813,307, and Patch for Targeted Delivery of an Oncogenic Cargo to a Tissue, United States Patent Application No. 63/306,449. R.C.G. reports grants from United States National Institutes of Health, Guerbet USA LLC, Janssen Research & Development LLC, NeoTherma Oncology, TriSalus Life Sciences, and Sus Clinicals, Inc.; consulting fees from Boston Consulting Group; pending patent for Modeling Oncology on Demand, United States Provisional Patent Application No. 62/813,307; participation on Sus Clinicals, Inc. Scientific Advisory Board; participation on Fluidx Medical Technology Advisory board; participation on Kaveri University Advisory board; and stock or stock options in Sus Clinicals, Inc. The remaining authors have no conflicts of interest to declare., (© 2025. Published by The Company of Biologists.)

Introduction: Bladder cancer is a common neoplasia of the urinary tract that holds the highest cost of lifelong treatment per patient, highlighting the need for a continuous search for new therapies for the disease. Current bladder cancer models are either imperfect in their ability to translate results to clinical practice (mouse models), or rare and not inducible (canine models). Swine models are an attractive alternative to model the disease due to their similarities with humans on several levels. The Oncopig Cancer Model has been shown to develop tumors that closely resemble human tumors. However, urothelial carcinoma has not yet been studied in this platform., Methods: We aimed to develop novel Oncopig bladder cancer cell line (BCCL) and investigate whether these urothelial swine cells mimic human bladder cancer cell line (5637 and T24) treatment-responses to cisplatin, doxorubicin, and gemcitabine in vitro., Results: Results demonstrated consistent treatment responses between Oncopig and human cells in most concentrations tested (p>0.05). Overall, Oncopig cells were more predictive of T24 than 5637 cell therapeutic responses. Microarray analysis also demonstrated similar alterations in expression of apoptotic (GADD45B and TP53INP1) and cytoskeleton-related genes (ZMYM6 and RND1) following gemcitabine exposure between 5637 (human) and Oncopig BCCL cells, indicating apoptosis may be triggered through similar signaling pathways. Molecular docking results indicated that swine and humans had similar Dg values between the chemotherapeutics and their target proteins., Discussion: Taken together, these results suggest the Oncopig could be an attractive animal to model urothelial carcinoma due to similarities in in vitro therapeutic responses compared to human cells., Competing Interests: LBS, LJ, CB, and KS work for Sus Clinicals, which provides the Oncopig and other pig-based preclinical testing services to customers. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Segatto, Simões, Bender, Sousa, Oliveira, Paschoal, Pacheco, Lopes, Seixas, Qazi, Thomas, Chaki, Robertson, Newsom, Patel, Rund, Jordan, Bolt, Schachtschneider, Schook and Collares.)

Melanoides tuberculata sensu lato (Thiaridae) are polymorphic female-clonal snails of Asian and African origins that have invaded freshwaters worldwide, including those in Florida. Although the snails have been documented in Florida for at least 70 years, no studies have investigated whether the observed distribution is due to a single introduction or multiple independent invasions. Here, cytochrome oxidase I was used to measure genetic diversity within and among sites in Florida and compare genetic diversity between Florida and other regions of the world. We also examined the relationship between shell morphology and haplotype diversity to determine if shell morphs can serve as a proxy for haplotypes. In total, we recovered 8 haplotypes randomly distributed across populations in Florida. Phylogenetic reconstruction supported the hypothesis of multiple invasions by diverse representatives of the M. tuberculata species complex. In contrast, shell morphology was not found to be a useful phylogeographic character, with divergent haplotypes represented by similar shell forms. These results suggest that the observed invasion patterns in Florida are best explained by serial introductions, and that shell morphology cannot be used to predict haplotypes or reconstruct invasion history of Melanoides tuberculata s.l. and that extensive taxonomic revisions are needed to investigate invasion dynamics., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Tolley-Jordan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Suids, both domesticated and wild, are found on all continents except for Antarctica and provide valuable food resources for humans in addition to serving as important models for biomedical research. Continuing advances in genome sequencing have allowed researchers to compare the genomes from diverse populations of suids helping to clarify their evolution and dispersal. Further analysis of these samples may provide clues to improve disease resistance/resilience and productivity in domestic suids as well as better ways of classifying and conserving genetic diversity within wild and captive suids. Collecting samples from diverse populations of suids is resource intensive and may negatively impact endangered populations. Here we catalog extensive tissue and DNA samples from suids in collections in both Europe and North America. We include samples that have previously been used for whole genome sequencing, targeted DNA sequencing, RNA sequencing, and reduced representation bisulfite sequencing (RRBS). This work provides an important centralized resource for researchers who wish to access published databases., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Analytical characterization of small biological particles, such as extracellular vesicles (EVs), is complicated by their extreme heterogeneity in size, lipid, membrane protein, and cargo composition. Analysis of individual particles is essential for illuminating particle property distributions that are obscured by ensemble measurements. To enable high-throughput analysis of individual particles, liftoff nanocontact printing (LNCP) is used to define hexagonal antibody and toxin arrays that have a 425 nm dot size, on average, and 700 nm periodicity. The LNCP process is rapid, simple, and does not require access to specialized nanofabrication tools. These densely packed, highly ordered arrays are used to capture liposomes and bacterial outer membrane vesicles on the basis of their surface biomarkers, with a maximum of one particle per array dot, resulting in densely packed arrays of particles. Despite the high particle density, the underlying antibody or toxin array ensured that neighboring individual particles are optically resolvable. Provided target particle biomarkers and suitable capture molecules are identified, this approach can be used to generate high density arrays of a wide variety of small biological particles, including other types of EVs like exosomes., (© 2021 Wiley-VCH GmbH.)

We show that photosensitized phospholipid oxidation, initiated by the lipid-conjugated fluorophore TopFluor-PC, causes defects, namely, membrane tubes and vesicle-like structures, in supported lipid bilayers (SLBs). Lipid oxidation is detrimental to the integrity of the lipid molecules; when oxidized, they undergo a conformational expansion, which causes membrane tubes to protrude from the SLB. Lipid oxidation is verified by FT-IR spectroscopy, and area expansion is observed in Langmuir trough experiments. Upon growing to a critical length, the membrane tubes arising from SLBs rapidly undergo transition to vesicle-like structures. We find a correlation between the maximum tube length and the diameter of the resulting vesicle, suggesting the conservation of the surface area between these features. We use geometric modeling and the measured tube length and vesicle radius to calculate the tube radius; our calculated mean tube diameter of 243 nm is comparable to other groups' experimental findings. In the presence of fluid flow, membrane tubes can be extended to tens to hundreds of microns in length. SLBs composed of saturated lipids resist light-induced tubulation, and the inclusion of the lipophilic antioxidant α-tocopherol attenuates the tubulation process and increases the light intensity threshold for tubulation.

In the nervous system, a myelin sheath that originates from oligodendrocytes or Schwann cells wraps around axons to facilitate electrical signal transduction. The interface between an axon and myelin is maintained by a number of biomolecular interactions. Among the interactions are those between GD1a and GT1b gangliosides on the axon and myelin-associated glycoprotein (MAG) on myelin. Interestingly, these interactions can also inhibit neuronal outgrowth. Ganglioside-MAG interactions are often studied in cellular or animal models where their relative concentrations are not easily controlled or in assays where the gangliosides and MAG are not presented as part of fluid lipid bilayers. Here, we present an approach to characterize MAG-ganglioside interactions in real time, where MAG, GD1a, and GT1b contents are controlled and they are in their in vivo orientation within fluid lipid bilayers. Using a quartz crystal microbalance with dissipation monitoring (QCM-D) biosensor functionalized with a supported lipid bilayer (SLB) and MAG, we detect vesicular GD1a and GT1b binding and determine the interaction kinetics as a function of vesicular ganglioside content. MAG-bound vesicles are deformed similarly, regardless of the ganglioside or its mole fraction. We further demonstrate how MAG-ganglioside interactions can be disrupted by antiganglioside antibodies that override MAG-based neuron growth inhibition.

Gangliosides are glycolipids that are enriched on the outer surface of cell membranes. Gangliosides are receptors for a number of signaling molecules and toxins, and therefore are often incorporated into biosensors. Many of these biosensors incorporate gangliosides into supported lipid bilayers which are formed by the spontaneous rupture of unilamellar vesicles on glass or SiO 2 substrates. In this work, we used quartz crystal microbalance with dissipation monitoring (QCM-D) to investigate how the presence of the four major brain gangliosides (GM1, GD1a, GD1b, and GT1b) influences the process of supported lipid bilayer formation on SiO 2 surfaces. We show that the rate of supported bilayer formation is dependent on both the charge and position of sialic acid moieties on ganglioside molecules. Additionally, Ca 2+ can accelerate ganglioside-rich supported bilayer formation, but the degree of acceleration differs for vesicles containing different gangliosides. Fluorescence recovery after photobleaching measurements show that the presence of all gangliosides reduces lipid diffusion coefficients in a concentration-dependent manner, and that Ca 2+ slows lipid diffusion in membranes with and without gangliosides. Finally, we use ganglioside-rich supported bilayers to measure binding constants for a GD1a-binding antibody that has similar properties to antibodies present in a variant of Guillain-Barré syndrome., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Imprime PGG (Imprime) is an i.v. administered, yeast β-1,3/1,6 glucan in clinical development with checkpoint inhibitors. Imprime-mediated innate immune activation requires immune complex formation with naturally occurring IgG anti-β glucan Abs (ABA). We administered Imprime to healthy human volunteers to assess the necessity of ABA for Imprime-mediated immunopharmacodynamic (IPD) changes. Imprime (4 mg/kg) was administered i.v. in single and multiple infusions. Subsets of subjects were premedicated with antihistamine and corticosteroid. Peripheral blood was measured before, during and after Imprime administration for IPD changes (e.g., ABA, circulating immune complexes, complement activation, complete blood counts, cytokine/chemokine, and gene expression changes). IPD changes were analyzed based on pretreatment serum ABA levels: low-ABA (<20 μg/ml), mid-ABA (≥20-50 μg/ml), and high-ABA (≥50 μg/ml). At the end of infusion, free serum ABA levels decreased, circulating immune complex levels increased, and complement activation was observed. At ∼1-4 h after end of infusion, increased expression of cytokines/chemokines, a 1.5-4-fold increase in neutrophil and monocyte counts and a broad activation of innate immune genes were observed. Low-ABA subjects typically showed minimal IPD changes except when ABA levels rose above 20 μg/ml after repeated Imprime dosing. Mild-to-moderate infusion-related reactions occurred in subjects with ABA ≥20 μg/ml. Premedications alleviated some of the infusion-related reactions, but also inhibited cytokine responses. In conclusion, ABA levels, being critical for Imprime-mediated immune activation may provide a plausible, mechanism-based biomarker to identify patients most likely to respond to Imprime-based anticancer immunotherapy., (Copyright © 2019 by The American Association of Immunologists, Inc.)

The Comal River, a spring-fed system in central Texas, was invaded in the 1960s by two Asian aquatic snails (Thiaridae: red-rimmed melania Melanoides tuberculata and quilted melania Tarebia granifera) and subsequently by three of their trematode parasites (the avian eye-fluke Philophthalmus gralli in the 1960s; the gill trematode Centrocestus formosanus in the 1990s; and the intestinal fluke Haplorchis pumilio in the 2000s). Previous snail collections (2001-2002) established that habitat conditions significantly affect the distribution of both snail species. However, the effects of snail size (known to influence infection prevalence) and habitat conditions (known to influence snail size) on trematode infection patterns in this system were not evaluated. In a re-evaluation of this data set, logistic regression analyses with individual snails showed that for both M. tuberculata and T. granifera populations, large snails were more likely to be infected than small snails, and habitat conditions were significantly related to infection in T. granifera. However, only snail size was significant in explaining the probability of infection in M. tuberculata. This result was confirmed by linear regression models, which showed that both infected and noninfected M. tuberculata used similar habitats, as large individuals in both infection categories were found in patches dominated by fine substrates and high levels of aquatic vegetation and detritus. For the large size-class of T. granifera, noninfected individuals were found primarily in habitats with silt/sand substrates and high vegetation and detritus cover, while infected individuals occurred among all available habitats. Using these results, we suggest that targeted sampling of large individuals of M. tuberculata in habitats with high detritus and vegetation and large individuals of T. granifera in any habitat can be used to efficiently ascertain parasite "hot spots" and to evaluate changes in parasite prevalence or detect the invasion of new parasites in these thiarid snails., (© 2018 American Fisheries Society.)

rHIgM22 is a recombinant human monoclonal IgM designed to promote remyelination, and it is currently in Phase I clinical trials in patients with multiple sclerosis (MS). In animal models of demyelination, a single low dose of rHIgM22 stimulates oligodendrocyte maturation, induces remyelination, preserves axons, and slows the decline of locomotor deficits. Natural autoantibodies like rHIgM22 typically bind to multiple antigens with weak affinity. rHIgM22 binds to oligodendrocytes and myelin. Because the antigens for rHIgM22 is prevalent within and exclusive to central nervous system (CNS) myelin, we used CNS myelin particles in combination with surface plasmon resonance to determine the kinetic and affinity constants for the interaction of rHIgM22 to myelin. We found that both the serum and recombinant forms of the antibody bind to myelin with very small dissociation constants in the 100 pM range, which is highly unusual for natural autoantibodies. The extraordinary affinity between rHIgM22 and myelin may explain why such a low effective dose can stimulate CNS repair in animal models of demyelination and underlie the accumulation of rHIgM22 in the CSF in treated MS patients by targeting myelin.

A plug and socket approach for tightening polyelectrolyte multilayers is introduced based on the use pendant β-cyclodextrin groups. Prototypical multilayers derived from poly(sodium 4-styrene sulfonate) and β-cyclodextrin-containing poly(4-vinylbenzyltrimethylammonium chloride) are described. Evidence for tightened multilayers has been obtained from gas permeation, swelling and quartz crystal microbalance with dissipation (QCM-D) measurements.

Importance: Modulating the immune system does not reverse long-term disability in neurologic disorders. Better neuroregenerative and neuroprotective treatment strategies are needed for neuroinflammatory and neurodegenerative diseases., Objective: To review the role of monoclonal, naturally occurring antibodies (NAbs) as novel therapeutic molecules for treatment of neurologic disorders., Evidence Review: Peer-reviewed articles, including case reports, case series, retrospective reviews, prospective randomized clinical trials, and basic science reports, were identified in a PubMed search for articles about NAbs and neurologic disorders that were published from January 1, 1964, through June 30, 2015. We concentrated our review on multiple sclerosis, Parkinson disease, Alzheimer disease, and amyotrophic lateral sclerosis., Findings: Many insults, including trauma, ischemia, infection, inflammation, and neurodegeneration, result in irreversible damage to the central nervous system. Central nervous system injury often results in a pervasive inhibitory microenvironment that hinders regeneration. A common targeted drug development strategy is to identify molecules with high potency in animal models. Many approaches often fail in the clinical setting owing to a lack of efficacy in human diseases (eg, less than the response demonstrated in animal models) or a high incidence of toxic effects. An alternative approach is to identify NAbs in humans because these therapeutic molecules have potential physiologic function without toxic effects. NAbs of the IgG, IgA, or IgM isotype contain germline or close to germline sequences and are reactive to self-components, altered self-components, or foreign antigens. Our investigative group developed recombinant, autoreactive, natural human IgM antibodies directed against oligodendrocytes or neurons with therapeutic potential for central nervous system repair. One such molecule, recombinant HIgM22, directed against myelin and oligodendrocytes completed a successful phase 1 clinical trial without toxic effects with the goal of promoting remyelination in multiple sclerosis., Conclusions and Relevance: Animal studies demonstrate that certain monoclonal NAbs are beneficial as therapeutic agents for neurologic diseases. This class of antibodies represents a unique source from which to develop a new class of disease-modifying therapies.

Amyotrophic lateral sclerosis (ALS) is a devastating, fatal neurological disease that primarily affects spinal cord anterior horn cells and their axons for which there is no treatment. Here we report the use of a recombinant natural human IgM that binds to the surface of neurons and supports neurite extension, rHIgM12, as a therapeutic strategy in murine models of human ALS. A single 200 µg intraperitoneal dose of rHIgM12 increases survival in two independent genetic-based mutant SOD1 mouse strains (SOD1G86R and SOD1G93A) by 8 and 10 days, delays the onset of neurological deficits by 16 days, delays the onset of weight loss by 5 days, and preserves spinal cord axons and anterior horn neurons. Immuno-overlay of thin layer chromatography and surface plasmon resonance show that rHIgM12 binds with high affinity to the complex gangliosides GD1a and GT1b. Addition of rHIgM12 to neurons in culture increases α-tubulin tyrosination levels, suggesting an alteration of microtubule dynamics. We previously reported that a single peripheral dose of rHIgM12 preserved neurological function in a murine model of demyelination with axon loss. Because rHIgM12 improves three different models of neurological disease, we propose that the IgM might act late in the cascade of neuronal stress and/or death by a broad mechanism., (© 2015. Published by The Company of Biologists Ltd.)

During vesicle budding or endocytosis, biomembranes undergo a series of lipid- and protein-mediated deformations involving cholesterol-enriched lipid rafts. If lipid rafts of high bending rigidities become confined to the incipient curved membrane topology such as a bud-neck interface, they can be expected to reform as ring-shaped rafts. Here, we report on the observation of a disk-to-ring shape morpho-chemical transition of a model membrane in the absence of geometric constraints. The raft shape transition is triggered by lateral compositional heterogeneity and is accompanied by membrane deformation in the vertical direction, which is detected by height-sensitive fluorescence interference contrast microscopy. Our results suggest that a flat membrane can become curved simply by dynamic changes in local chemical composition and shape transformation of cholesterol-rich domains.

We report on the conformal surface passivation of photonic crystal (PC) laser devices with an ultrathin dielectric layer. Air-bridge-type Γ-point band-edge lasers (BELs) are fabricated by forming a honeycomb lattice two-dimensional PC structure into an InGaAsP multiple-quantum-well epilayer. Atomic layer deposition (ALD) is employed for conformal deposition of a few-nanometer-thick SiO2 layer over the entire device surface, not only on the top and bottom surfaces of the air-bridge membrane but also on the air-hole sidewalls. Despite its extreme thinness, the ALD passivation layer is found to protect the InGaAsP BEL devices from harsh chemicals. In addition, the ALD-SiO2 is compatible with the silane-based surface chemistry, which allows us to use ALD-passivated BEL devices as label-free biosensors. The standard streptavidin-biotin interaction shifts the BEL lasing wavelength by ∼1 nm for the dipole-like Γ-point band-edge mode. A sharp lasing line (<0.2 nm, full width at half-maximum) and a large refractive index sensitivity (∼163 nm per RIU) produce a figure of merit as high as ∼800 for our BEL biosensor, which is at least an order of magnitude higher than those of more common biosensors that rely on a broad resonance peak, showing that our nanolaser structures are suitable for highly sensitive biosensor applications.

During vesicular trafficking and release of enveloped viruses, the budding and fission processes dynamically remodel the donor cell membrane in a protein- or a lipid-mediated manner. In all cases, in addition to the generation or relief of the curvature stress, the buds recruit specific lipids and proteins from the donor membrane through restricted diffusion for the development of a ring-type raft domain of closed topology. Here, by reconstituting the bud topography in a model membrane, we demonstrate the preferential localization of cholesterol- and sphingomyelin-enriched microdomains in the collar band of the bud-neck interfaced with the donor membrane. The geometrical approach to the recapitulation of the dynamic membrane reorganization, resulting from the local radii of curvatures from nanometre-to-micrometre scales, offers important clues for understanding the active roles of the bud topography in the sorting and migration machinery of key signalling proteins involved in membrane budding.

Lipid bilayer membranes form the plasma membranes of cells and define the boundaries of subcellular organelles. In nature, these membranes are heterogeneous mixtures of many types of lipids, contain membrane-bound proteins and are decorated with carbohydrates. In some experiments, it is desirable to decouple the biophysical or biochemical properties of the lipid bilayer from those of the natural membrane. Such cases call for the use of model systems such as giant vesicles, liposomes or supported lipid bilayers (SLBs). Arrays of SLBs are particularly attractive for sensing applications and mimicking cell-cell interactions. Here we describe a new method for forming SLB arrays. Submicron-diameter SiO2 beads are first coated with lipid bilayers to form spherical SLBs (SSLBs). The beads are then deposited into an array of micro-fabricated submicron-diameter microwells. The preparation technique uses a "squeegee" to clean the substrate surface, while leaving behind SSLBs that have settled into microwells. This method requires no chemical modification of the microwell substrate, nor any particular targeting ligands on the SSLB. Microwells are occupied by single beads because the well diameter is tuned to be just larger than the bead diameter. Typically, more 75% of the wells are occupied, while the rest remain empty. In buffer SSLB arrays display long-term stability of greater than one week. Multiple types of SSLBs can be placed in a single array by serial deposition, and the arrays can be used for sensing, which we demonstrate by characterizing the interaction of cholera toxin with ganglioside GM1. We also show that phospholipid vesicles without the bead supports and biomembranes from cellular sources can be arrayed with the same method and cell-specific membrane lipids can be identified.

Characterization of binding kinetics and affinity between a potential drug and its receptor are key steps in the development of new drugs. Among the techniques available to determine binding affinities, surface plasmon resonance has emerged as the gold standard because it can measure binding and dissociation rates in real-time in a label-free fashion. Surface plasmon resonance is now finding applications in the characterization of molecules for treatment of neurodegenerative diseases, characterization of molecules associated with pathogenesis of neurodegenerative diseases and detection of neurodegenerative disease biomarkers. In addition it has been used in the characterization of a new class of natural autoantibodies that have therapeutic potential in a number of neurologic diseases. In this review we will introduce surface plasmon resonance and describe some applications of the technique that pertain to neurodegenerative disorders and their treatment.

Matrix molecules convey biochemical and physical guiding signals to neurons in the central nervous system (CNS) and shape the trajectory of neuronal fibers that constitute neural networks. We have developed recombinant human IgMs that bind to epitopes on neural cells, with the aim of treating neurological diseases. Here we test the hypothesis that recombinant human IgMs (rHIgM) can guide neurite outgrowth of CNS neurons. Microcontact printing was employed to pattern rHIgM12 and rHIgM22, antibodies that were bioengineered to have variable regions capable of binding to neurons or oligodendrocytes, respectively. rHIgM12 promoted neuronal attachment and guided outgrowth of neurites from hippocampal neurons. Processes from spinal neurons followed grid patterns of rHIgM12 and formed a physical network. Comparison between rHIgM12 and rHIgM22 suggested the biochemistry that facilitates anchoring the neuronal surfaces is a prerequisite for the function of IgM, and spatial properties cooperate in guiding the assembly of neuronal networks.

With recent advances in high-throughput proteomics and systems biology, there is a growing demand for new instruments that can precisely quantify a wide range of receptor-ligand binding kinetics in a high-throughput fashion. Here we demonstrate a surface plasmon resonance (SPR) imaging spectroscopy instrument capable of simultaneously extracting binding kinetics and affinities from 50 parallel microfluidic channels. The instrument utilizes large-area (~ cm(2)) metallic nanohole arrays as SPR sensing substrates and combines a broadband light source, a high-resolution imaging spectrometer and a low-noise CCD camera to extract spectral information from every channel in real time with a refractive index resolution of 7.7 × 10(-6) refractive index units. To demonstrate the utility of our instrument for quantifying a wide range of biomolecular interactions, each parallel microfluidic channel is coated with a biomimetic supported lipid membrane containing ganglioside (GM1) receptors. The binding kinetics of cholera toxin b (CTX-b) to GM1 are then measured in a single experiment from 50 channels. By combining the highly parallel microfluidic device with large-area periodic nanohole array chips, our SPR imaging spectrometer system enables high-throughput, label-free, real-time SPR biosensing, and its full-spectral imaging capability combined with nanohole arrays could enable integration of SPR imaging with concurrent surface-enhanced Raman spectroscopy.

The recent explosion of interest in brain-machine interfaces (BMIs) has spurred research into choosing the optimal input signal source for a desired application. The signals with highest bandwidth--single neuron action potentials or spikes--typically are difficult to record for more than a few years after implantation of intracortical electrodes. Fortunately, field potentials recorded within the cortex (local field potentials, LFPs), at its surface (electrocorticograms, ECoG) and at the dural surface (epidural, EFPs) have also been shown to contain significant information about movement. However, the relative performance of these signals has not yet been directly compared. Furthermore, while it is widely postulated, it has not yet been demonstrated that these field potential signals are more durable than spike recordings. The aim of this study was to address both of these questions. We assessed the offline decoding performance of EFPs, LFPs and spikes, recorded sequentially, in primary motor cortex (M1) in terms of their ability to decode the target of reaching movements, as well as the endpoint trajectory. We also examined the decoding performance of LFPs on electrodes that are not recording spikes, compared with the performance when they did record spikes. Spikes were still present on some of the other electrodes throughout this study. We showed that LFPs performed nearly as well as spikes in decoding velocity, and slightly worse in decoding position and in target classification. EFP performance was slightly inferior to that reported for ECoG in humans. We also provided evidence demonstrating that movement-related information in the LFP remains high regardless of the ability to record spikes concurrently on the same electrodes. This is the first study to provide evidence that LFPs retain information about movement in the absence of spikes on the same electrodes. These results suggest that LFPs may indeed remain informative after spike recordings are lost, thereby providing a robust, accurate signal source for BMIs.

Brain-machine interfaces (BMIs) use signals from the brain to control a device such as a computer cursor. Various types of signals have been used as BMI inputs, from single-unit action potentials to scalp potentials. Recently, intermediate-level signals such as subdural field potentials have also shown promise. These different signal types are likely to provide different amounts of information, but we do not yet know what signal types are necessary to enable a particular BMI function, such as identification of reach target location, control of a two-dimensional cursor or the dynamics of limb movement. Here we evaluated the performance of field potentials, measured either intracortically (local field potentials, LFPs) or epidurally (epidural field potential, EFPs), in terms of the ability to decode reach direction. We trained rats to move a joystick with their forepaw to control the motion of a sipper tube to one of the four targets in two dimensions. We decoded the forelimb reach direction from the field potentials using linear discriminant analysis. We achieved a mean accuracy of 69 ± 3% with EFPs and 57 ± 2% with LFPs, both much better than chance. Signal quality remained good up to 13 months after implantation. This suggests that using epidural signals could provide BMI inputs of high quality with less risk to the patient than using intracortical recordings.

The center-out task is a standard paradigm often used to study the neural control of reaching movements in human and non-human primates. However, there are several disadvantages to the use of monkeys, notably costs, infrastructural requirements, and ethical considerations. Here we describe a similar task designed to examine forelimb movements in rats. Rats were trained to grasp a joystick with their forepaw and use it to control the movements of a sipper tube in two dimensions. The rats learned to move the joystick in four directions with at least 70% accuracy after about 45 days of training. In addition, rats were able to learn a reversed mapping between joystick and sipper tube movement. This is a more complicated behavior than has been previously demonstrated for rats, and it could allow more motor behavior studies to be conducted in rodents instead of monkeys. We currently are using this behavior to decode the rats' forelimb movements from their brain signals., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Brain-machine interfaces (BMIs) use signals recorded directly from the brain to control an external device, such as a computer cursor or a prosthetic limb. These control signals have been recorded from different levels of the brain, from field potentials at the scalp or cortical surface to single neuron action potentials. At present, the more invasive recordings have better signal quality, but also lower stability over time. Recently, subdural field potentials have been proposed as a stable, good quality source of control signals, with the potential for higher spatial and temporal bandwidth than EEG. Here we used finite element modeling in rats and humans and spatial spectral analysis in rats to compare the spatial resolution of signals recorded epidurally (outside the dura), with those recorded from subdural and scalp locations. Resolution of epidural and subdural signals was very similar in rats and somewhat less so in human models. Both were substantially better than signals recorded at the scalp. Resolution of epidural and subdural signals in humans was much more similar when the cerebrospinal fluid layer thickness was reduced. This suggests that the less invasive epidural recordings may yield signals of similar quality to subdural recordings, and hence may be more attractive as a source of control signals for BMIs.

Unlabelled: A novel low-stiffness extensively porous-coated total hip femoral component was designed to achieve stable skeletal fixation, structural durability, and reduced periprosthetic femoral stress shielding. In short- to intermediate-term clinical review, this implant achieved secure biologic fixation and preserved periprosthetic bone. We retrospectively reviewed all 102 prospectively followed patients (106 implants) with this implant to document the longer-term implant survivorship, clinical function, fixation quality, and periprosthetic bone preservation. Ninety-seven patients with 101 implants had current followup or were followed to patient death (range, 1-14 years; average, 10 years). Eighty-six living patients were followed for an average implant survivorship of 10 years. There were no known femoral implant removals. The average Harris hip score at 10-year followup was 98. Radiographs demonstrated secure implant fixation and maintenance of periprosthetic bone. These data suggest this implant design provided long-term function characterized by extensive fixation, structural durability, and radiographic appearance of maintained periprosthetic cortical thickness and density., Level of Evidence: Level I, therapeutic study. See Guidelines for Authors for a complete description of levels of evidence.

Background: Discriminating taxa with the nuclear marker, amplified fragment length polymorphism (AFLP) has been accomplished for various organisms in economic, ecological, and evolutionary studies. The protocol available for AFLP generation does not require prior knowledge of the genome; however, it is often extensively modified to fit the needs of the researcher. Modification of this protocol for new labs is intimidating and time-consuming, particularly for taxa in which AFLP have not been previously developed. Furthermore, determining what constitutes quality output during different stages of fragment generation is not well defined and this may further hinder the use AFLP by new researchers., Findings: We present a step-by-step AFLP protocol, using flourophore-labeled primers for use with automated sequencers, including examples of both successful and unsuccessful results. We sufficiently normalized peak intensity and standardized allele calling across all samples for each primer combination. Repeatability was assessed with a phylogenetic tree in which replicate samples clustered together using the minimum evolution procedure. We found differences greater than 10% in allele position among replicated samples would cause replicates to no longer cluster. To minimize offset allele positions, we suggest that researchers analyze different primer combinations at the same time using multiple dyes with the automated sequencer to minimize mismatched alleles across replicates., Conclusion: For researchers wanting to use AFLP, this molecular technique is difficult and time-consuming to develop. Clarifying what constitutes quality output for each step in AFLP generation will help to reduce redundant trials in protocol development and, in turn, advance the discipline of population genetics.

A growing number of brain-machine interfaces have now been developed that allow movements of an external device to be controlled using recordings from the brain. This work has been undertaken with a number of different animal models, as well as several human patients with quadriplegia. The resulting movements, whether of computer cursors or robotic limbs, remain quite slow and unstable compared to normal limb movements. It is an open question, how much of this instability is the result of the limited forward control path, and how much has to do with the total lack of normal proprioceptive feedback. We have begun preliminary studies of the effectiveness of electrical stimulation in the proprioceptive area of the primary somatosensory cortex (area 3a) as a potential means to deliver an artificial sense of proprioception to a monkey. We have demonstrated that it is possible for the monkey to detect brief stimulus trains at relatively low current levels, and to discriminate between trains of different frequencies. These observations need to be expanded to include more complex, time-varying waveforms that could potentially convey information about the state of the limb.

Brain-machine interfaces (BMIs) have the potential to improve quality of life for thousands of motor-impaired individuals. Many different signal sources have been investigated for use in controlling a BMI, including scalp EEG, field potentials from inside and the surface of the cerebral cortex, and single-neuron action potentials. A relatively unexplored region for recording signals is the epidural space. This study attempts to help determine the optimal spatial resolution of epidural and subdural electrode arrays using both a mathematical model and spatial spectral analysis. For rats, optimal spacing for both epidural and subdural electrodes was approximately 0.7 mm.

Since its introduction in the early 1980s, the concept of a "preferred direction" for neuronal discharge has proven to be a powerful means of studying motor areas of the brain. In the current paper, we introduce the concept of a "muscle-space"-preferred direction (PD(M)) that is analogous to the familiar hand-space-preferred direction (PD(H)). PD(M) reflects the similarity between the discharge of a given neuron and the activity of each muscle in much the way that PD(H) reflects the similarity of discharge with motion along each of the three Cartesian coordinate axes. We used PD(M) to analyze the data recorded from neurons in the primary motor cortex (M1) of three different monkeys. The monkeys performed center-out movements within two different cubical workspaces centered either to the left or right of the monkey's shoulder while we simultaneously recorded neuronal discharge, muscle activity, and limb orientation. We calculated preferred directions in both hand space and muscle space, and computed the angles between these vectors under a variety of conditions. PDs for different neurons were broadly distributed throughout both hand space and muscle space, but the muscle-space vectors appeared to form clusters of functionally similar neurons. In general, repeated estimates of PD(M) were more stable over time than were similar estimates of PD(H). Likewise, there was less change in PD(M) than in PD(H) for data recorded from the two different workspaces. However, although a majority of neurons had this muscle-like property, a significant minority was more stable in Cartesian hand space, reflecting a heterogeneity of function within M1.

Unlabelled: Enthusiasm for metal-backed patella has waned because of the high incidence of complications. Considering that perhaps all metal-backed patellae may not be the same, 256 primary consecutive metal-backed mobile bearing TKAs done between May 1985 and January 1989 were retrospectively reviewed to evaluate the results of a unique mobile bearing metal-backed patella. There were four complications (1.6%). Three patella were revised for polyethylene complications and one well-functioning component was removed at the time of revision of the tibial polyethylene to facilitate range of motion and wound closure. No patella was revised for loosening, subluxation, or dislocation. Our results show that all metal-backed patella are not the same and that compared with the high incidence of failure of fixed bearing metal-backed patellae, the use of the anatomic mobile bearing metal-backed patella can produce excellent, durable long-term clinical and radiographic results with a low incidence (1.6%) of complications. Life table survivorship using revision for any reason as the end point was 97% (95% confidence interval, 93%-100%) at a maximum of 19 years., Level of Evidence: Therapeutic study, Level IV (case series--no, or historical control group). See the Guidelines for Authors for a complete description of levels of evidence.

Two hundred fifty-six primary cementless meniscal-bearing total knee arthroplasties were performed between May 1985 and January 1989. All knees were replaced with a low contact stress metal-backed anatomic mobile patella. The average patient follow-up was 11.5 years. No patellae were revised for failure of fixation and no dislocations or subluxations were reported. One patella was revised for polyethylene wear, and one well-functioning component was removed at the time of revision to facilitate range of motion and wound closure. The survival estimate at 12 years was 99.5%. Compared to the high incidence of failure of metal-backed fixed-bearing patellae components, the anatomic rotating patella provides durable long-term results with a low incidence of complications.

Using T2-weighted Magnetic Resonance Imaging (MRI) in a pyrithiamin-treated, thiamin deficient (TD) rat model of Wernicke's encephalopathy (WE), we have observed hyperintensity in the thalamus, hypothalamus, collicular bodies and hippocampus which was enhanced 40 min after a glucose load. Hyperintensity was not evident in these structures in thiamin replete rats receiving glucose nor was it enhanced in TD rats administered 2-deoxyglucose. Residual hyperintensity was still evident in the hippocampus as long as 30 days after thiamin administration and was increased by repeat glucose challenge at that time. These data indicate that the hippocampus is as vulnerable as the thalamus to some persistent pathological change when glucose is metabolised in a state of thiamin deficiency., (Copyright 1998 Elsevier Science B.V.)

Four hundred seventy-three consecutive cementless cruciate retaining meniscal bearing primary total knee replacements were done on 375 patients from May 1985 to February 1991. These were observed for a 10-year period (average, 5 years). Seventeen (3.6%) required change of components because of mechanical failure. There were 12 polyethylene fractures or dislocations. There were 5 tibial subluxations secondary to ligamentous instability occurring at an average of 21 months postoperatively. There were 2 component loosenings secondary to bone graft resorption (1 femoral, 1 tibial). There were 5 infections (4 Staphylococcus aureus, 1 Pseudomonas). Significantly, with the exception of the 2 knees with bone graft resorption, there was no component (femoral, tibial, or patellar) loosening. Kaplan-Meier survival estimates, using as an endpoint of revision surgery for any mechanical reason (polyethylene breakage, polyethylene dislocation, or ligamentous instability), showed a survivorship of 94.6% at the 8-year interval. Survivorship related to mechanical loosening of fixation of any component at the 8-year interval was 99%.

Biphasic vasodilatory responses to adenosine and 5'-N-ethylcarboxamidoadenosine (NECA) were observed in the coronary vasculature of K(+)-arrested perfused rat hearts. Dose-response data for both agonists were best represented by two-site models. For adenosine, two sites with negative log ED50 (pED50) values of 8.1 +/- 0.1 (mean +/- S.E.M) and 5.2 +/- 0.1 were obtained, mediating 31 +/- 2% and 69 +/- 2% of the total response. In the presence of 8-phenyltheophylline, the vasodilatory response to adenosine remained best fitted to a two-site model with pED50 values of 7.0 +/- 0.2 and 5.4 +/- 0.2. The relative contribution of each site to the total response remained unchanged. For NECA, pED50 values of 9.6 +/- 0.1 and 6.8 +/- 0.2 were obtained, representing 48 +/- 3% and 52 +/- 3% of the sites, respectively. In contrast, ATP produced a monophasic response with a pED50 value of 8.8 +/- 0.1. These results provide evidence of adenosine receptor and response heterogeneity in the in situ coronary vasculature.

Exogenous adenosine causes a monophasic dilation of the coronary vessels in paced, perfused rat heart preparations. Because levels of endogenous adenosine in paced hearts may mask the presence of high potency adenosine receptors, we have developed a method to measure coronary vascular responses in a potassium-arrested heart. Hearts from adult male, Wistar rats were perfused at a constant flow rate of 10 mL/min in the nonrecirculating, Langendorff mode, using Krebs-Henseleit buffer. After 30 min, coronary perfusion pressure was 44 +/- 1 mmHg (mean +/- SEM). Hearts were then perfused with a modified Krebs-Henseleit buffer containing 35 mM potassium. Coronary perfusion pressure increased by 84 +/- 3 mmHg. Adenosine-induced reductions in coronary perfusion pressure were expressed as a percentage of the maximal increase in pressure produced by modified Krebs-Henseleit buffer from the equilibration level. A concentration-response curve for adenosine (n = 6) was biphasic and best described by the presence of two adenosine receptors, with negative log EC50 values of 8.8 +/- 0.3 and 4.3 +/- 0.1, representing 29 +/- 3 and 71 +/- 3%, respectively, of the observed response. Interstitial adenosine sampled by microdialysis during potassium arrest was 25% of the concentration found in paced hearts. Endogenous adenosine in nonarrested hearts may obscure the biphasic response of the coronary vessels to adenosine.

Inorganic phosphate (Pi) accumulates extremely rapidly in ischaemic heart muscle and intracellular binding of this metabolite may account for the precipitous loss of function seen at the onset of severe ischaemia. We have used 31P-NMR spectroscopy to measure the free cytosolic [Pi] and chemical assay techniques to measure total tissue Pi at 0, 1, 2, 3, 4, 5, and 12 min of complete global ischaemia in the isolated isovolumic rat heart. At zero time, the Pi assayed chemically was 30.77 +/- 5.52 mumol/g dry wt (mean +/- SD, n = 7) whilst Pi assayed by NMR was 3.39 +/- 1.21 mumol/g dry wt (n = 15). Thus, 27.38 mumol/g dry wt of Pi was bound at a cytosolic [Pi] of 0.82 mM. After 12 min of ischaemia, 49.88 mumol/g dry wt of Pi was bound at a cytosolic [Pi] of 4.11 mM. When all data were fitted, using a non-linear, least squares fit (p < 0.05), to the binding isotherm: Bound Pi = Bmax'. [Pi]/Kd'+[Pi], the apparent binding parameters Kd' and Bmax' were estimated to be 1.1 +/- 0.6 mM and 64.0 +/- 10.2 mumol/g dry wt respectively. During the first minute of global ischaemia when the rate-pressure product had decreased by 79% of its pre-ischaemic value, bound Pi had increased by 58% and free cytosolic [Pi] by 162%. When functional and metabolite changes were expressed as a fraction of the total change which occurred during the 12-min ischaemic period, bound Pi had the profile most similar to the rate-pressure product. Both the amount of bound Pi and free cytosolic [Pi] correlated with loss of contractile function as the ischaemic period progressed. The results show that during ischaemia, Pi is bound progressively as free cytosolic [Pi] is increased as the result of high energy phosphate hydrolysis. While these results are consistent with the possibility that Pi binding may contribute to ischaemic contractile failure, no molecular explanation for the possible effect of bound Pi on contraction has been proposed.

One hundred knees that underwent primary total knee arthroplasties were divided into 2 groups: the first 50 consecutive knees were assigned retrospectively to Group I (control), and the following 50 knees were assigned prospectively to Group II (early flexion). All procedures were cementless meniscal-bearing total knee arthroplasties and were performed by the same surgeon. Maximum early flexion (Group II) resulted in decreased length of stay, decreased hospital costs, and increased range of motion at 1 year. In light of current government interest in hospital cost reduction, this method should be considered as an attractive alternative to continuous passive motion.

In the present study we examined the action of native and oxidized low-density lipoproteins (LDL) on coronary vascular and cardiac function and ultrastructure in rat hearts perfused isovolumically in the Langendorff mode. Responses of the coronary resistance vessels to the endothelium-dependent vasodilator, histamine, and the endothelium-independent vasodilator, NaNO2, were measured together with contractile function (rate-pressure product) before and after perfusion for 20 min with native - or oxidized-LDL at a concentration of 100 mu g protein/ml. Ultrastructural damage was assessed via electron microscopy of perfusion-fixed heart specimens. When compared to findings in untreated, control hearts, both native and oxidized LDL significantly reduced the responsiveness of the coronary resistance vessels to histamine and NaNO2, by about 50%. The rate-pressure product was decreased more by oxidized-LDL (41%) than by native-LDL (26%). Electron microscopy showed no ultrastructural abnormalities in the vasculature or myocytes of control hearts. The administration of both native- and oxidized-LDL caused distortion of endothelial cells, increased levels of pinocytotic vesicles in both endothelial and smooth muscle cells, detachment of blood vessels from surrounding tissue, and some regions of myocyte injury with evidence of mitochondrial injury and fluid accumulation. Our results show that both native- and oxidized-LDL are toxic to the isolated heart preparation. They inhibit coronary vascular responsiveness to vasodilators, reduce contractile function, and produce damage to cardiac ultrastructure.

Objective: To examine the effect of hydrogen peroxide on the function and ultrastructure of cardiac muscle and the coronary vasculature in an isovolumic rat heart preparation perfused at constant flow., Design: Ventricular function was monitored via a balloon placed in the left ventricle and the response of the coronary vessels to vasodilators was assessed in hearts arrested with 35 mM potassium and treated with 5 microM phenylephrine to contract the coronary resistance vessels. Changes in coronary perfusion pressure reflect changes in resistance vessel tone. SETTING/ANIMALS: This experimental study consisted of 14 heart preparations, six control and eight treated hearts., Interventions: Hydrogen peroxide was included in the perfusate at a final concentration of 250 microM for 20 mins., Main Results: Hydrogen peroxide reduced rate-pressure product by 42%, caused a fivefold increase in end-diastolic pressure and increased coronary perfusion pressure by 33%. Also, the response of the coronary vasculature to the endothelium-dependent vasodilator, histamine, and endothelium-independent vasodilator, sodium nitrite, was decreased by 55% and 53%, respectively. Electron microscopy of hydrogen peroxide-treated hearts showed damage to both capillaries and arterioles. Endothelial cells were distorted and contained pinocytotic vesicles, endothelial cell junctions were disrupted and blood vessels were detached from surrounding tissue. A comparatively small amount of injury was seen in the myocyte population., Conclusions: The greater amount of ultrastructural damage seen in blood vessels compared with cardiac muscle suggests that the smooth muscle and endothelial cells of the vasculature are more susceptible to oxidant injury than the myocytes.

A prospective study was conducted to determine the rate of puncture of the inner glove when various double-gloving protocols were used for elective total joint procedures. Group 1 consisted of 22 cases in which two pairs of regular latex gloves were used. Group 2 consisted of 25 cases in which outer "orthopaedic" gloves were used over regular latex gloves. Group 3 consisted of 24 cases in which Repel gloves were used between two regular latex gloves. The patients were randomly assigned to one of the three groups. All gloves, both inner and outer, were tested for holes at the end of the procedure. All gloves that were changed for presumed intraoperative holes were also tested. The type and duration of the procedure was also documented. Of the inner holes, there were significant differences between each group. Group 1 (double latex) and Group 2 (outer orthopaedic/inner regular) had significantly more holes than Group 3 (Repel between two regular gloves). There were no holes in Group 3, and Group 2 offered significant protection over Group 1.

Vitamin E is an endogenous antioxidant and is known to afford protection against lipid peroxidation. If lipid peroxidation was an important factor in the pathogenesis of reoxygenation injury in heart, then both the extent of lipid peroxidation and cell injury would be expected to be exacerbated in vitamin E-deficient hearts. To study reoxygenation injury in the present experiments, rat hearts were perfused in the Langendorff mode with a modified Krebs-Henseleit buffer under anoxic conditions for 60 min before resuming normoxic perfusion for 20 min. Creatine phosphokinase (CPK) activity and malonyldialdehyde (MDA), a product of lipid peroxidation, were assayed in the perfusate effluent from hearts during reoxygenation injury. Also, myocardial MDA and vitamin E contents were measured in extracts of freeze-clamped heart tissue obtained immediately before and 2 min after reoxygenation. Experiments were performed on hearts from groups of weanling rats fed either a vitamin E-deficient or vitamin E-supplemented diet (50 I.U. vitamin E/kg) for 5 to 6 weeks. After 5 weeks, the myocardial vitamin-E content was 103.8 +/- 5.3 (n = 5) and 11.5 +/- 1.6 (n = 4) ng/mg protein (mean +/- SEM) in the vitamin E-supplemented and vitamin E-deficient groups respectively. Perfused hearts from both dietary groups showed a peak of enzyme release 2 to 3 min after the reintroduction of oxygen, and enzyme release from vitamin E-deficient hearts was two-fold greater than enzyme release from vitamin E-supplemented hearts.(ABSTRACT TRUNCATED AT 250 WORDS)