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Background: Background: Cortical stimulation (CS) as a part of presurgical investigations in patients undergoing implantation of depth electrodes (SEEG) is a growing practice in some Comprehensive Epilepsy Centers. After-discharges (AD) are useful to determine epileptogenic tissue within or outside the epileptogenic network. Classification of afterdischarges was proposed by Blume using subdural recordings(1); its utility in SEEG is unknown. Methods: Methods: Single center, retrospective study that included patients with SEEG that underwent CS in the Epilepsy Monitoring Unit. Demographic characteristics were explored and Blume’s proposed AD classification was used to determine whether or not the CS changed surgical outcomes. Results: Results: From January 2015 to June 2021, a total of 177 patients were implanted with SEEG and analyzed. 95 patients had CS and 91 had AD. Morphologies found were: Rhythmic waves in 4 (0.04%), Rhythmic waves evolving into spikes in18 (19%), Polyspike bursts in 14 (15%), Spike-waves in 28 (30%), and sequential spikes in 18 (19%). 12/14 (86%) patients with Spike-waves had Engel I outcome; Engel class IV patients were more likely to have an evolution of morphology and frequency of ADs in 3 patients(75%). Conclusions: Conclusions: The most frequent morphology of ADs seen was spike-waves. AD morphology and duration may predict post-operative seizure outcomes.

Background: Background: Language determination is a pivotal part of presurgical investigations. Presurgical cortical stimulation (CS) with language mapping (LM) in patients with intracranial recordings (SEEG) is a growing practice in some Comprehensive Epilepsy Centers. Methods: Methods: This retrospective, single center study included patients implanted with SEEG that underwent CS for LM in our Epilepsy Monitoring Unit. We describe frequencies, demographic characteristics of these patients and whether or not CS with LM was useful. Results: Results: From January 2015 to June2021, a total of 177 patients were implanted with SEEG and analyzed. 95 patients had CS and 44 of these had CS with LM. The mean age was 33 (ranging from 15-70). During LM, anomia was induced in 26 (58%), speech arrest in 22 (49%), paraphasic errors in 13 (29%), and hesitation in 9 (20%). LM results were recorded as influencing surgical decision in 7 (16%) patients, 4 (9%) did not undergo surgery due to expected language deficits and 3 (7%) proceeded with surgery due to an acceptable risk of language deficit. Conclusions: Conclusions: Cortical stimulation language mapping is useful for decision-making in presurgical evaluation and should be encouraged whenever involvement of language is suspected when determining the epileptogenic zone.

ARXis a paired-type homeobox gene located on the X chromosome that contains five exons with four polyalanine (PolyA) tracts, a homeodomain, and a conserved C-terminal aristaless domain. Studies in humans have demonstrated remarkable pleiotropy: malformation phenotypes are associated with protein truncation mutations and missense mutations in the homeobox; nonmalformation phenotypes, including X-linked infantile spasms (ISS), are associated with missense mutations outside of the homeobox and expansion of the PolyA tracts.

Chemical sensors utilizing immobilized enzymes and proteins are important for monitoring chemical processes and biological systems. In this study, calcium-cross-linked alginate hydrogel microspheres were fabricated as enzyme carriers by an emulsification technique. Glucose oxidase (GOx) was encapsulated in alginate microspheres using three different methods:  physical entrapment (emulsion), chemical conjugation (conjugation), and a combination of physical entrapment and chemical conjugation (emulsion−conjugation). Nano-organized coatings were applied on alginate/GOx microspheres using the layer-by-layer self-assembly technique in order to stabilize the hydrogel/enzyme system under biological environment. The encapsulation of GOx and formation of nanofilm coating on alginate microspheres were verified with FTIR spectral analysis, ζ-potential analysis, and confocal laser scanning microscopy. To compare both the immobilization properties of enzyme encapsulation techniques and the influence of nanofilms with uncoated microspheres, the relationship between enzyme loading, release, and effective GOx activity (enzyme activity per unit protein loading) were studied over a period of four weeks. The results produced four key findings:  (1) the emulsion−conjugation technique improved the stability of GOx in alginate microspheres compared to the emulsion technique, reducing the GOx leaching from microsphere from 50% to 17%; (2) the polyelectrolyte nanofilm coatings increased the GOx stability over time, but also reduced the effective GOx activity; (3) the effective GOx activity for the emulsion−conjugation technique (about 3.5 × 10^-5 AU μg^-1 s^-1) was higher than that for other methods, and did not change significantly over four weeks; and (4) the GOx concentration, when compared after one week for microspheres with three bilayers of poly(allylamine hydrochloride)/sodium poly(styrene sulfonate) ({PAH/PSS}) coating, was highest for the emulsion−conjugation technique. As a result, the comparison of these three techniques showed the emulsion−conjugation technique to be a potentially effective and practical way to fabricate alginate/GOx microspheres for implantable glucose biosensor application.

A simple and high-efficiency approach to loading macromolecules into microscale carriers is presented. Calcium-cross-linked alginate hydrogel microspheres were fabricated by an emulsification technique and then used as negatively charged templates to form polyelectrolyte multilayer coatings. A calcium ion chelator, EDTA, was used to free the Ca²⁺-cross-linked alginate hydrogel within {poly(allylamine hydrochloride)/poly (styrene sulfonate)}4 ({PAH/PSS}4) coating, allowing partial release of alginate. The retention of alginate in {PAH/PSS}4 microcapsule was confirmed by FTIR spectroscopy and confocal microscopy. Real-time confocal microscopy was used to investigate the loading process of positively charged macromolecules (dextran-amino, and peroxidase) into alginate-templated microcapsules, which showed the loading occurred in <2 min for dextran-amino and <10 min for peroxidase, respectively. A high loading efficiency of 25 μg peroxidase in ~1.0 × 10⁷ microcapsules (2.5 pg POx/capsule) was achieved with a low concentration of peroxidase loading solution (10 μg/mL). This spontaneous loading technique for encapsulating positively charged molecules in alginate-templated polyelectrolyte microcapsules shows strong potential for biosensor and drug delivery applications.

Inorganic colloids are usually prepared via precipitation reactions, which involve two sequential steps:  nucleation and growth of the nuclei. On the basis of this theory, preformed manganese carbonate nanoseeds were prepared by mixing diluted ammonium hydrogen carbonate with manganese sulfate solution to serve as nuclei for production of monodisperse manganese carbonate microparticles. It was found that the average particle diameter can be finely tuned from 1 to 10 μm by adding a different volume of nanoseed solution. It was further verified that the manganese carbonate microparticles can be used as templates for efficient formation of uniform, smooth multilayer polyelectrolyte microcapsules, which are promising systems for biomedical and other applications, such as biosensors, bioreactors, and drug-delivery devices.

A stable enzyme encapsulation technique based on the conversion of weak interactions between diazo resin/poly(styrene sulfonate) to covalent bonds was explored. Photosensitive diazoresin-based polyelectrolyte microcapsules were prepared via layer-by-layer electrostatic self-assembly of poly(styrene sulfonate) and diazoresin on MnCO3 templates. UV−vis and ζ-potential measurements confirmed the alternate deposition of {PSS/DAR} multilayers on the micrometer-sized dissolvable templates. The DAR-based microcapsules were demonstrated to be permeable to enzymes prior to UV irradiation, while the permeability of the multilayer wall was changed substantially after photo-cross-linking. Encapsulated molecules were stably entrapped after UV irradiation, as shown by confocal microscopy and atomic force microscopy images. Activity assays revealed that encapsulated glucose oxidase possessed 52.8% of the catalytic activity exhibited by the same amount of free enzyme, proving the preservation of native conformation and accessibility of substrate. This encapsulation technique is promising for many biomedical and biotechnological applications, particularly enzyme biosensors, which require stable immobilization of functional components while allowing sufficient transport rates for substrate molecules.

Methods for producing protein patterns with defined spatial arrangement and micro- and nanoscale features are important for studying cellular-level interactions, including basic cell−cell communications, cell signaling, and mechanisms of drug action. Toward this end, a straightforward, versatile procedure for fabricating micropatterns of bioactive nanofilm coatings as multifunctional biological testbeds is demonstrated. The method, based on a combination of photolithography and layer-by-layer self-assembly (LbL), allows for precise construction of nanocomposite films of potentially complex architecture, and patterning of these films on substrates using a modified lift-off (LO) procedure. As a first step in evaluating nanostructures made with this process, “comparison chips,” comprising two coexisting regions of square patterns with relevant proteins/polypeptides on a single substrate, were fabricated with poly(diallyldimethylammonium chloride) (PDDA) as a cell-repellent background. Using neuronal cells as a model biological system, comparison chips were produced with secreted phospholipase A2 (sPLA2), a known membrane-active enzyme for neurons, for direct comparison with gelatin, poly-l-lysine (PLL), or bovine serum albumin (BSA). Fluorescence microscopy, surface profilometry, and atomic force microscopy techniques were used to evaluate the structural properties of the patterns on these chips and show that the patterning technique was successful. Preliminary cell culture studies show that neurons respond and bind specifically to the sPLA2 enzyme embedded in the polyelectrolyte thin films and present as the outermost layer. These findings point to the potential for this method to be applied in developing test substrates for a broad array of studies aimed at identifying important biological structure−function relationships.

Systems for glucose monitoring based on resonance energy transfer (RET) and competitive binding using Concanavalin A (Con A) are problematic as a result of problems of toxicity, aggregation, and irreversible binding. This paper presents an improved RET assay wherein Con A was replaced by apo-glucose oxidase (apo-GOx). The basic principle for transduction is identical to that used in assays based on Con A−dextran:  a reduction in RET from fluorescein isothiocyanate (FITC) to tetramethyl rhodamine isothiocyanate (TRITC) occurs when FITC−dextran (donor) is displaced from TRITC−apo-GOx (acceptor) as a result of the competition of glucose. Fluorescence measurements confirm that the apo-GOx/dextran complexes are highly sensitive to glucose, measured as an increase in the donor peak relative to acceptor due to stepwise addition of glucose. The solution-phase assay showed strong signals and excellent repeatability, with a sensitivity of 0.0163 (ratio units)/mM over the range of 0−90 mM glucose. If properly encapsulated, these sensors can potentially be used for in vivo sensing without the concern of toxicity associated with Con A.

Organized multilayers of nanoparticles (9-, 20-, and 45-nm-diameter silica or 12-nm magnetite) and glucose oxidase (GOx) were assembled in alternation with oppositely charged polyelectrolytes on 420-nm latex particles. Stepwise growth of the multilayer films on latex was confirmed by microelectrophoresis and transmission electron microscopy. The inclusion of silica layers on latex yields a higher surface area, resulting in greater GOx adsorption and thereby increasing the catalytic activity of the bioreactor. The bioactivity was proportional to the core surface area and also to the number of GOx layers in the shells. Also, the presence of magnetic nanoparticles allows self-stirring of the nanoreactors with a rotating magnetic field and enhances its productivity. The ensemble of GOx and fluorescent dyes in the shells demonstrated the correlation between Ru-bpy fluorescence and glucose concentration in solution.

This paper describes the neuroimaging and neuropathological findings in five cases of severe brain damage after traumatic mid-trimester amniocentesis, all performed between 1986 and 1994. Although fetal injury after amniocentesis has been reported, reports of brain injury are infrequent. Continuous ultrasound monitoring may reduce the risk of fetal injury but follow-up ultrasound scans can be falsely reassuring. Withdrawal of blood-stained fluid, particularly if it contains tissue fragments, should alert the operator to the possibility of fetal damage. Histological examination of such tissue fragments may confirm the nature of the fetal damage. The consequences of fetal brain injury are severe, all five of our cases showed evidence of disruption of brain development compatible with mid-term injury. Obstetricians and their patients should be aware of the small but significant risk of brain damage after mid-term amniocentesis.

An increase in cellular levels of cyclic nucleotides activates serine/threonine-dependent kinases that lead to diverse physiological effects. Recently we reported the activation of the p38 mitogen-activated protein kinase (MAPK) pathway in neutrophils by a cGMP-dependent mechanism. In this study we demonstrated that exogenously supplied nitric oxide leads to activation of p38 MAPK in 293T fibroblasts. Phosphorylation of p38 corresponded with an increase in ATF-2-dependent gene expression. The effect of nitric oxide was mimicked by addition of 8-bromo-cGMP, indicating that activation of soluble guanylyl cyclase was involved. The importance of cGMP-dependent protein kinase in the activation of p38 MAPK by nitric oxide in 293T cells was assessed in a transfection based assay. Overexpression of cGMP-dependent protein kinase-1alpha caused phosphorylation of p38 in these cells and potentiated the effectiveness of cGMP. Overexpression of a catalytically inactive mutant form of this enzyme (T516A) blocked the ability of both nitric oxide and 8-bromo-cGMP to activate p38 as measured by both p38 phosphorylation and ATF-2 driven gene expression. Together, these data demonstrate that nitric oxide stimulates a novel pathway leading to activation of p38 MAPK that requires activation of cGMP-dependent protein kinase.

Which are the driving forces ford(-)3-hydroxybutyrate (HB) transport in rat renal brush border membranes (RBB)? Sodium, even in the absence of gradients, accelerates the unidirectional (1–5 s) flux of HB into rat RBB vesicles. Valinomycin (andKi=Ko) does not significantly alter the NaCl gradient driven HB influx. Thus, the Na-dependent HB influx is driven by the chemical Na+ gradient but it is not driven by changes in the transmembrane electrical potential. Indeed, in valinomycin-treated membranes, vesicle-inside more negative potentials (K-gluconatein-Na-gluconateout) sufficient to accelerate Na-glucose cotransport, did not stimulate HB influx, in the presence of inwardly directed Na+ gradients, and did not significantly inhibit when in the absence of Na+. Thus, cotransport of HB with Na in rat RBB membranes does not involve the net transfer of positive charge and the passive conductance of this membrane for HB- is not large. However, vesicle inside more negative potentials (induced by inwardly directed NaNO3 gradients or by outwardly directed K+ gradients and valinomycin in the presence of inwardly directed Na+ gradients) inhibited HB influx, suggesting that another potential sensitive mechanism, perhaps redistribution of intramembrane charges, may influence HB influx. Acidification (pHi=pHo=6.4 vs. 7.4) or inwardly directed H+ gradients (pHo/pHi=6.4/7.4) did not alter HB influx, in the absence of Na+. Thus there is no evidence for a H+ driven HB influx. HB influx is significantly inhibited by high (100 mEq/l) trans concentration of Na+. Also, influx of 2.25 mM14C-HB was significantly increased by 5–10 mM intravesicular HB under Na-equilibrated conditions. Thus, the rate of translocation of the free carrier appears to limit HB influx through the cotransport system.

Implantable luminescent sensors are being developed for on-demand monitoring of blood glucose levels. For these sensors to be deployed in vivo, a matched external hardware system is needed. In this paper, we designed a compact, low-cost optical system with highly efficient photon delivery and collection using advanced optical modeling software. Compared to interrogation with a fiber bundle, the new system was predicted to improve interrogation efficiency by a factor of 200 for native sensors; an improvement of 37 times was predicted for sensors implanted at a depth of 1 mm in a skin-simulating phantom. A physical prototype was tested using silicone-based skin phantoms developed specifically to mimic the scattering and absorbing properties of human skin. The experimental evaluations revealed that the prototype device performed in agreement with expectations from simulation results, resulting in an overall improvement of over 2000 times. This efficient system enables use of a low-cost commercial spectrometer for recording sensor emission, which was not possible using only fiber-optic delivery and collection, and will be used as a tool for in vivo studies with animal models or human subjects. [ABSTRACT FROM AUTHOR]

Each year thousands of patients are added to the waiting list for liver transplants. The first 7-10 days after transplant have proven to be the most critical in patient recovery and it is hypothesized that monitoring organ vital signals in this period can increase patient and graft survival rates. An implantable sensor to monitor the organ perfusion and oxygenation signals following surgery is being developed by our group. The sensor operates based on measuring diffuse reflection from three light emitting diodes (735, 805 and 940 nm). In this work the optimal source detector spacing to maximize oxygenation signal level is investigated for a portal vein model. Monte Carlo simulations provided signal levels and corresponding penetration depths as a function of separation between a point optical source and detector. The modeling results indicated a rapid decay in the optical signal with increasing distance. Through further analysis, it was found that there exists an optimal range of point source to detector spacing, between roughly 1 and 2 mm, in which the blood signal from the simulated portal vein was maximized. Overall, these results are being used to guide the placement and configuration of our probe for in vivo animal studies.

Fazio-Londe disease is a label sometimes applied to a degenerative disease of the motor neurons characterized by progressive bulbar paralysis in children. It is very rare with only 22 case reports describing 24 children including four sibling pairs. In two reports mothers and sons were affected. The neuropathology is described in only four cases. Previous authors have recognized that the condition is very heterogeneous. The clinical features of five children with this type of progressive bulbar paralysis, diagnosed at this hospital between 1969 and 1989, are reviewed, and in two cases neuropathological findings are detailed. Based on this experience, suggested criteria for diagnosis include clinical features of a pure motor neuronopathy affecting the bulbar nuclei, exclusion of other causes of progressive bulbar paralysis and positive support for the diagnosis from electromyography and/or pathological examination. A review of the literature, combined with the present series, suggests that there are at least three distinct subtypes: a very rare autosomal dominant form (as described by Fazio) and two variants with probable autosomal recessive inheritance either with early onset of respiratory symptoms and rapid progression to death or later onset, less prominent respiratory symptoms and protracted clinical course. There is strong concordance for each clinical pattern within families.