Investigating Internalization of Reporter-Protein-Functionalized Polyhedrin Particles by Brain Immune Cells.

Achieving sustained drug delivery to the central nervous system (CNS) is a major challenge for neurological injury and disease, and various delivery vehicles are being developed to achieve this. Self-assembling polyhedrin crystals (POlyhedrin Delivery System; PODS) are being exploited for the delivery of therapeutic protein cargo, with demonstrated efficacy in vivo. However, to establish the utility of PODS for neural applications, their handling by neural immune cells (microglia) must be documented, as these cells process and degrade many biomaterials, often preventing therapeutic efficacy. Here, primary mouse cortical microglia were cultured with a GFP-functionalized PODS for 24 h. Cell counts, cell morphology and Iba1 expression were all unaltered in treated cultures, indicating a lack of acute toxicity or microglial activation. Microglia exhibited internalisation of the PODS, with both cytosolic and perinuclear localisation. No evidence of adverse effects on cellular morphology was observed. Overall, 20-40% of microglia exhibited uptake of the PODS, but extracellular/non-internalised PODS were routinely present after 24 h, suggesting that extracellular drug delivery may persist for at least 24 h.

Seasonal activity of Trechnites insidiosus (Hymenoptera: Encyrtidae) and its host Cacopsylla pyricola (Hemiptera: Psyllidae) in pear.

Cacopsylla pyricola (Förster) (Hemiptera: Psyllidae) is the most expensive and challenging insect pest of commercial pear trees in the Pacific Northwest. Integrated pest management (IPM) programs are working toward relying more heavily on natural enemies to reduce insecticide use. Trechnites insidiosus (Crawford) (Hymenoptera: Encyrtidae) is the main parasitoid of C. pyricola, but little is known about its biology in the region. Developing sampling tools is important for the deployment of IPM programs, including monitoring of natural enemies. In this study, we examined 2 conventional monitoring methods: beat trays and yellow sticky cards, in addition to screened sticky cards and 3D-printed cylinder traps. Additionally, we tested an overwintering trap for the collection of parasitized C. pyricola. The trapping methods were tested in orchards in Oregon and Washington. Unscreened cards caught the most T. insidiosus and C. pyricola, followed by screened cards, cylinder traps, and then beat trays. Beat trays sometimes failed to catch any T. insidiosus, even when it was found in abundance via other methods. Screened cards and cylinder traps reduced bycatch and increased ease of identifying T. insidiosus. Specimens from the cylinder traps were also more suitable for use in molecular analysis. The overwintering traps were effective at capturing parasitized C. pyricola, but were highly variable year to year. The ideal trapping method will vary based on research needs (e.g., DNA preservation, reducing bycatch, catching higher numbers), but both screened sticky cards and cylinder traps were viable methods for monitoring T. insidiosus and its host.

Erythritol sprays reduce Drosophila suzukii infestation without impacting honey bee visitation nor fruit quality.

BACKGROUND: Spotted-wing drosophila, Drosophila suzukii, is an economic pest of small fruits and cherries. Insecticides primarily control this pest while alternative controls are in development. Laboratory studies show that erythritol is insecticidal to D. suzukii and other pests while approved for human consumption. Moreover, erythritol combined with sucrose or non-caloric sucralose can stimulate feeding and quicken mortality. Before growers can use erythritol, the impact on crop protection, non-target insects, and fruit quality need evaluation. RESULTS: In three blueberry and cherry field cage trials, oviposition on fruit sprayed with erythritol:sucrose or erythritol:sucralose formulations was lowered by 59%-81% compared with unsprayed controls. Fly infestation (larval or adult counts from fruit) was 90% lower in a greenhouse blueberry trial, and 49% lower in an open field blueberry trial with 2 m erythritol : 0.5 m sucrose. Infestation was also 57% lower in an open field cherry trial with 1.5 m erythritol:0.5 m sucrose. Other field trials with very low pest pressure or frequent rains revealed no differences from controls. Field trials consistently revealed that honey bees did not preferentially visit plants sprayed with either erythritol formulation, although yellow jackets visited plants sprayed with erythritol:sucrose more frequently. Erythritol formulations consistently led to more leaf spotting, but there was no reduction in the quality of treated blueberries or cherries in terms of mold development, firmness, diameter, epidermal penetration force, and Brix° (total soluble solids) at harvest. CONCLUSION: Eleven trials conducted over four years show that erythritol formulations can reduce D. suzukii pressure without attracting foraging honey bees nor negatively impacting fruit quality. © 2023 Society of Chemical Industry. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

Robotic-assisted total hip arthroplasty: an economic analysis.

Aim: To evaluate 90-day episode-of-care (EOC) resource consumption in robotic-assisted total hip arthroplasty (RATHA) versus manual total hip arthroplasty (mTHA). Methods: THA procedures were identified in Medicare 100% data. After propensity score matching 1:5, 938 RATHA and 4,670 mTHA cases were included. 90-day EOC cost, index costs, length of stay and post-index rehabilitation utilization were assessed. Results: RATHA patients were significantly less likely to have post-index inpatient rehabilitation or skilled nursing facility admissions and used fewer home health agency visits, compared with mTHA patients. Total 90-day EOC costs for RATHA patients were found to be US$785 less than those of mTHA patients (p = 0.0095). Conclusion: RATHA was associated with an overall lower 90-day EOC cost when compared with mTHA. The savings associated with RATHA were driven by reduced utilization and cost of post-index rehabilitation services.

Implementation of a Skull-Conformal Phased Array for Transcranial Focused Ultrasound Therapy.

OBJECTIVE: To implement a skull-conformal phased array for ultrasound-guided transcranial focused ultrasound therapy with improved patient comfort. METHODS: Using patient-specific computed tomography and MRI neuroimaging data, tightly-conforming helmet scaffolds were designed computationally. The helmet scaffolds were designed to hold reusable transducer modules at near-normal incidence in an optimal configuration for the treatment location(s) of interest. Numerical simulations of trans-skull ultrasound propagation were performed to evaluate different conformal array designs and to compare with hemispherical arrays similar to those employed clinically. A 4096-element phased array was constructed by 3D printing a helmet scaffold optimised for an ex vivo human skullcap, and its performance was evaluated via benchtop and in vivo experiments. RESULTS: Acoustic field measurements confirmed the system's ability to focus through human skull bone using simulation-based transcranial aberration corrections. Preliminary in vivo testing demonstrated safe trans-human skull blood-brain barrier (BBB) opening in rodents. CONCLUSION: Patient-specific conformal ultrasound phased arrays appear to be a feasible and safe approach for conducting transcranial BBB opening procedures. SIGNIFICANCE: Skull-conformal phased arrays stand to improve patient comfort and have the potential to accelerate the adoption of transcranial FUS therapy by improving access to the technology.

Enhancing the regenerative potential of stem cell-laden, clinical-grade implants through laminin engineering.

Protected delivery of neural stem cells (NSCs; a major transplant population) within bioscaffolds has the potential to improve regenerative outcomes in sites of spinal cord injury. Emergent research has indicated clinical grade bioscaffolds (e.g. those used as surgical sealants) may be repurposed for this strategy, bypassing the long approval processes and difficulties in scale-up faced by laboratory grade materials. While promising, clinical scaffolds are often not inherently regenerative. Extracellular molecule biofunctionalisation of scaffolds can enhance regenerative features such as encapsulated cell survival/distribution, cell differentiation into desired cell types and nerve fibre growth. However, this strategy is yet to be tested for clinical grade scaffolds. Here, we show for the first time that Hemopatch™, a widely used, clinically approved surgical matrix, supports NSC growth. Further, functionalisation of Hemopatch™ with laminin promoted homogenous distribution of NSCs and their daughter cells within the matrix, a key regenerative criterion for transplant cells.

HIFU Power Monitoring Using Combined Instantaneous Current and Voltage Measurement.

During high-intensity focused ultrasound (HIFU) therapy, it is important that the electrical power delivered to the transducer is monitored to avoid underexposure or overexposure, ensure patient safety, and to protect the transducer itself. Due to ease of measurement, the transducer's potential difference may be as an indicator of power delivery. However, even when a transducer's complex impedance is well characterized at small amplitudes and matching networks are used, voltage-only (VO) monitoring cannot account for the presence of drive waveform distortion, changes to the acoustic path, or damage to the transducer. In this study, combined current and voltage (CCV) is proposed as a magnetic resonance imaging (MRI)-compatible, miniature alternative to bidirectional power couplers, which is compatible with switched amplifiers. For CCV power measurement, current probe data were multiplied by the voltage waveform and integrated in the frequency domain. Transducer efficiency was taken into account to predict acoustic power. The technique was validated with a radiation force balance (RFB). When using a typical HIFU transducer and amplifier, VO predictions and acoustic power had a maximum difference of 20%. However, under the same conditions, CCV only had a maximum difference of 5%. The technique was applied to several lesioning experiments and it was shown that when VO was used as a control between two amplifiers, there was up to a 38% difference in lesion area. This greatly reduced to a maximum of 5% once CCV was used instead. These results demonstrate that CCV can accurately predict real-time electrical power delivery, leading to safer HIFU treatments.

Impaired skeletal muscle mitochondrial pyruvate uptake rewires glucose metabolism to drive whole-body leanness.

Metabolic cycles are a fundamental element of cellular and organismal function. Among the most critical in higher organisms is the Cori Cycle, the systemic cycling between lactate and glucose. Here, skeletal muscle-specific Mitochondrial Pyruvate Carrier (MPC) deletion in mice diverted pyruvate into circulating lactate. This switch disinhibited muscle fatty acid oxidation and drove Cori Cycling that contributed to increased energy expenditure. Loss of muscle MPC activity led to strikingly decreased adiposity with complete muscle mass and strength retention. Notably, despite decreasing muscle glucose oxidation, muscle MPC disruption increased muscle glucose uptake and whole-body insulin sensitivity. Furthermore, chronic and acute muscle MPC deletion accelerated fat mass loss on a normal diet after high fat diet-induced obesity. Our results illuminate the role of the skeletal muscle MPC as a whole-body carbon flux control point. They highlight the potential utility of modulating muscle pyruvate utilization to ameliorate obesity and type 2 diabetes.

Humility, Relational Spirituality, and Well-being among Religious Leaders: A Moderated Mediation Model.

Prior research has demonstrated positive associations between general humility and well-being, and posited a protective effect for intellectual humility against maladjustment among religious leaders. We tested a model that extended findings on general humility to include intellectual humility among religious leaders (N = 258; M age = 42.31; 43% female; 63.7% White; 91.9% Christian affiliation). We observed a positive general humility-well-being association. Contrary to expectations, we observed risk effects for religion-specific intellectual humility. Our findings also point to the possibility that these risk effects might be attenuated by the integration of high levels of general and intellectual humility.

HIFU Drive System Miniaturization Using Harmonic Reduced Pulsewidth Modulation.

Switched excitation has the potential to improve on the cost, efficiency, and size of the linear amplifier circuitry currently used in high-intensity focused ultrasound (HIFU) systems. Existing switching schemes are impaired by high harmonic distortion or lack array apodisation capability, so require adjustable supplies and/or large power filters to be useful. A multilevel pulsewidth modulation (PWM) topology could address both of these issues but the switching-speed limitations of transistors mean that there are a limited number of pulses available in each waveform cycle. In this study, harmonic reduction PWM (HRPWM) is proposed as an algorithmic solution to the design of switched waveforms. Its appropriateness for HIFU was assessed by design of a high power five-level unfiltered amplifier and subsequent thermal-only lesioning of ex vivo chicken breast. Three switched waveforms of different electrical powers (16, 26, 35 W) were generated using the HRPWM algorithm. Lesion sizes were measured and compared with those made at the same electrical power using a linear amplifier and bi-level excitation. HRPWM produced symmetric, thermal-only lesions that were the same size as their linear amplifier equivalents ( ). At 16 W, bi-level excitation produced smaller lesions but at higher power levels large transients in the acoustic waveform nucleated undesired cavitation. These results demonstrate that HRPWM can minimize HIFU drive circuity size without the need for filters to remove harmonics or adjustable power supplies to achieve array apodisation.

Nanoelectronic Platform for Ultrasensitive Detection of Protein Biomarkers in Serum using DNA Amplification.

Silicon nanowire field effect transistors (NWFETs) are low noise, low power, ultrasensitive biosensors that are highly amenable to integration. However, using NWFETs to achieve direct protein detection in physiological buffers such as blood serum remains difficult due to Debye screening, nonspecific binding, and stringent functionalization requirements. In this work, we performed an indirect sandwich immunoassay in serum combined with exponential DNA amplification and pH measurement by ultrasensitive NWFET sensors. Measurements of model cytokine interleukin-2 concentrations from <20 fM to >200 pM were demonstrated, surpassing the conventional NWFET urease-based readout. Our approach paves way for future development of universal, highly sensitive, miniaturized, and integrated nanoelectronic devices that can be applied to a wide variety of analytes.

An Adaptive Array Excitation Scheme for the Unidirectional Enhancement of Guided Waves.

Control over the direction of wave propagation allows an engineer to spatially locate defects. When imaging with longitudinal waves, time delays can be applied to each element of a phased array transducer to steer a beam. Because of the highly dispersive nature of guided waves (GWs), this beamsteering approach is suboptimal. More appropriate time delays can be chosen to direct a GW if the dispersion relation of the material is known. Existing techniques, however, need a priori knowledge of material thickness and acoustic velocity, which change as a function of temperature and strain. The scheme presented here does not require prior knowledge of the dispersion relation or properties of the specimen to direct a GW. Initially, a GW is generated using a single element of an array transducer. The acquired waveforms from the remaining elements are then processed and retransmitted, constructively interfering with the wave as it travels across the spatial influence of the transducer. The scheme intrinsically compensates for the dispersion of the waves, and thus can adapt to changes in material thickness and acoustic velocity. The proposed technique is demonstrated in simulation and experimentally. Dispersion curves from either side of the array are acquired to demonstrate the scheme's ability to direct a GW in an aluminum plate. The results show that unidirectional enhancement is possible without a priori knowledge of the specimen using an arbitrary pitch array transducer. The experimental results show a 34-dB enhancement in one direction compared with the other.

Metal-coated microfluidic channels: An approach to eliminate streaming potential effects in nano biosensors.

We report a method to suppress streaming potential using an Ag-coated microfluidic channel on a p-type silicon nanowire (SiNW) array measured by a multiplexed electrical readout. The metal layer sets a constant electrical potential along the microfluidic channel for a given reference electrode voltage regardless of the flow velocity. Without the Ag layer, the magnitude and sign of the surface potential change on the SiNW depends on the flow velocity, width of the microfluidic channel and the device's location inside the microfluidic channel with respect to the reference electrode. Noise analysis of the SiNW array with and without the Ag coating in the fluidic channel shows that noise frequency peaks, resulting from the operation of a piezoelectric micropump, are eliminated using the Ag layer with two reference electrodes located at inlet and outlet. This strategy presents a simple platform to eliminate the streaming potential and can become a powerful tool for nanoscale potentiometric biosensors.

Magnetic nanoparticle mediated transfection of neural stem cell suspension cultures is enhanced by applied oscillating magnetic fields.

Safe genetic modification of neural stem cell (NSC) transplant populations is a key goal for regenerative neurology. We describe a technically simple and safe method to increase transfection in NSCs propagated in the neurosphere (suspension culture) model, using magnetic nanoparticles deployed with applied oscillating magnetic fields ('magnetofection technology'). We show that transfection efficiency was enhanced over two-fold by oscillating magnetic fields (frequency=4 Hz). The protocols had no effect on cell viability, cell number, stem cell marker expression and differentiation profiles of 'magnetofected' cultures, highlighting the safety of the technique. As far as we are aware, this is the first successful application of magnetofection technology to suspension cultures of neural cells. The procedures described offer a means to augment the therapeutic potential of NSCs propagated as neurospheres - a culture model of high clinical translational relevance - by safe genetic manipulation, with further potential for incorporation into 'magneto-multifection' (repeat transfection) protocols. FROM THE CLINICAL EDITOR: This team of investigators describe a simple and safe method to increase transfection in neural stem cells using magnetic nanoparticles deployed with oscillating magnetic fields, demonstrating a greater than two-fold transfection efficiency increase by applying low frequency magnetic oscillation.

The effect of estrogen in a man with Parkinson's disease and a review of its therapeutic potential.

Estrogen has been implicated in controlling the pathogenesis and symptoms of Parkinson's disease (PD) in women. Here, we report a 53-year-old male with PD who underwent estrogen therapy with estradiol (E2). Within a month, he exhibited increased dyskinesias. His medication was reduced by 35% from a levodopa equivalent dose (LED) of 820-535 over three months, which overall improved his motor fluctuations and dyskinesias. Therefore, E2 therapy could have therapeutic potential in males with PD.

Amyloid fibrils composed of hexameric peptides attenuate neuroinflammation.

The amyloid-forming proteins tau, αB crystallin, and amyloid P protein are all found in lesions of multiple sclerosis (MS). Our previous work established that amyloidogenic peptides from the small heat shock protein αB crystallin (HspB5) and from amyloid ß fibrils, characteristic of Alzheimer's disease, were therapeutic in experimental autoimmune encephalomyelitis (EAE), reflecting aspects of the pathology of MS. To understand the molecular basis for the therapeutic effect, we showed a set of amyloidogenic peptides composed of six amino acids, including those from tau, amyloid ß A4, major prion protein (PrP), HspB5, amylin, serum amyloid P, and insulin B chain, to be anti-inflammatory and capable of reducing serological levels of interleukin-6 and attenuating paralysis in EAE. The chaperone function of the fibrils correlates with the therapeutic outcome. Fibrils composed of tau 623-628 precipitated 49 plasma proteins, including apolipoprotein B-100, clusterin, transthyretin, and complement C3, supporting the hypothesis that the fibrils are active biological agents. Amyloid fibrils thus may provide benefit in MS and other neuroinflammatory disorders.

Therapeutic effects of systemic administration of chaperone αB-crystallin associated with binding proinflammatory plasma proteins.

The therapeutic benefit of the small heat shock protein αB-crystallin (HspB5) in animal models of multiple sclerosis and ischemia is proposed to arise from its increased capacity to bind proinflammatory proteins at the elevated temperatures within inflammatory foci. By mass spectral analysis, a common set of ∼70 ligands was precipitated by HspB5 from plasma from patients with multiple sclerosis, rheumatoid arthritis, and amyloidosis and mice with experimental allergic encephalomyelitis. These proteins were distinguished from other precipitated molecules because they were enriched in the precipitate as compared with their plasma concentrations, and they exhibited temperature-dependent binding. More than half of these ligands were acute phase proteins or members of the complement or coagulation cascades. Consistent with this proposal, plasma levels of HspB5 were increased in patients with multiple sclerosis as compared with normal individuals. The combination of the thermal sensitivity of the HspB5 combined with the high local concentration of these ligands at the site of inflammation is proposed to explain the paradox of how a protein believed to exhibit nonspecific binding can bind with some relative apparent selectivity to proinflammatory proteins and thereby modulate inflammation.

Neural precursor cell-expressed developmentally down-regulated protein 4-2 (Nedd4-2) regulation by 14-3-3 protein binding at canonical serum and glucocorticoid kinase 1 (SGK1) phosphorylation sites.

Regulation of epithelial Na(+) channel (ENaC)-mediated transport in the distal nephron is a critical determinant of blood pressure in humans. Aldosterone via serum and glucocorticoid kinase 1 (SGK1) stimulates ENaC by phosphorylation of the E3 ubiquitin ligase Nedd4-2, which induces interaction with 14-3-3 proteins. However, the mechanisms of SGK1- and 14-3-3-mediated regulation of Nedd4-2 are unclear. There are three canonical SGK1 target sites on Nedd4-2 that overlap phosphorylation-dependent 14-3-3 interaction motifs. Two of these are termed "minor," and one is termed "major," based on weak or strong binding to 14-3-3 proteins, respectively. By mass spectrometry, we found that aldosterone significantly stimulates phosphorylation of a minor, relative to the major, 14-3-3 binding site on Nedd4-2. Phosphorylation-deficient minor site Nedd4-2 mutants bound less 14-3-3 than did wild-type (WT) Nedd4-2, and minor site Nedd4-2 mutations were sufficient to inhibit SGK1 stimulation of ENaC cell surface expression. As measured by pulse-chase and cycloheximide chase assays, a major binding site Nedd4-2 mutant had a shorter cellular half-life than WT Nedd4-2, but this property was not dependent on binding to 14-3-3. Additionally, a dimerization-deficient 14-3-3ε mutant failed to bind Nedd4-2. We conclude that whereas phosphorylation at the Nedd4-2 major site is important for interaction with 14-3-3 dimers, minor site phosphorylation by SGK1 may be the relevant molecular switch that stabilizes Nedd4-2 interaction with 14-3-3 and thus promotes ENaC cell surface expression. We also propose that major site phosphorylation promotes cellular Nedd4-2 protein stability, which potentially represents a novel form of regulation for turnover of E3 ubiquitin ligases.

Narcolepsy, REM sleep behavior disorder, and supranuclear gaze palsy associated with Ma1 and Ma2 antibodies and tonsillar carcinoma.

OBJECTIVE: To describe a patient with diencephalic and mesencephalic presentation of a Ma1 and Ma2 antibody-associated paraneoplastic neurological disorder. DESIGN: Case report. SETTING: The Colorado Neurological Institute Movement Disorders Center in Englewood, Colorado, and the Mayo Clinic in Rochester, Minnesota. PATIENT: A 55-year-old man with a paraneoplastic neurological disorder characterized by rapid eye movement sleep behavior disorder, narcolepsy, and a progressive supranuclear palsy-like syndrome in the setting of tonsillar carcinoma. INTERVENTION: Immunotherapy for paraneoplastic neurological disorder, surgery and radiotherapy for cancer, and symptomatic treatment for parkinsonism and sleep disorders. MAIN OUTCOME MEASURES: Polysomnography, multiple sleep latency test, and neurological examination. RESULTS: The cancer was detected at a limited stage and treatable. After oncological therapy and immunotherapy, symptoms stabilized. Treatment with modafinil improved daytime somnolence. CONCLUSIONS: Rapid onset and progression of multifocal deficits may be a clue to paraneoplastic etiology. Early treatment of a limited stage cancer (with or without immunotherapy) may possibly slow progression of neurological symptoms. Symptomatic treatment may be beneficial.