Ex-vivo systems for neuromodulation: A comparison of ex-vivo and in-vivo large animal nerve electrophysiology.

BACKGROUND: Little research exists on extending ex-vivo systems to large animal nerves, and to the best of our knowledge, there has yet to be a study comparing these against in-vivo data. This paper details the first ex-vivo system for large animal peripheral nerves to be compared with in-vivo results. NEW METHOD: Detailed ex-vivo and in-vivo closed-loop neuromodulation experiments were conducted on pig ulnar nerves. Temperatures from 20 °C to 37 °C were evaluated for the ex-vivo system. The data were analysed in the time and velocity domains, and a regression analysis established how evoked compound action potential amplitude and modal conduction velocity (CV) varied with temperature and time after explantation. MAIN RESULTS: Pig ulnar nerves were sustained ex-vivo up to 5 h post-explantation. CV distributions of ex-vivo and in-vivo data were compared, showing closer correspondence at 37 °C. Regression analysis results also demonstrated that modal CV and time since explantation were negatively correlated, whereas modal CV and temperature were positively correlated. COMPARISON WITH EXISTING METHODS: Previous ex-vivo systems were primarily aimed at small animal nerves, and we are not aware of an ex-vivo system to be directly compared with in-vivo data. This new approach provides a route to understand how ex-vivo systems for large animal nerves can be developed and compared with in-vivo data. CONCLUSION: The proposed ex-vivo system results were compared with those seen in-vivo, providing new insights into large animal nerve activity post-explantation. Such a system is crucial for complementing in-vivo experiments, maximising collected experimental data, and accelerating neural interface development.

Denoising and decoding spontaneous vagus nerve recordings with machine learning.

Neural interfaces that electrically stimulate the peripheral nervous system have been shown to successfully improve symptom management for several conditions, such as epilepsy and depression. A crucial part for closing the loop and improving the efficacy of implantable neuromodulation devices is the efficient extraction of meaningful information from nerve recordings, which can have a low Signal-to-Noise ratio (SNR) and non-stationary noise. In recent years, machine learning (ML) models have shown outstanding performance in regression and classification problems, but it is often unclear how to translate and assess these for novel tasks in biomedical engineering. This paper aims to adapt existing ML algorithms to carry out unsupervised denoising of neural recordings instead. This is achieved by applying bandpass filtering and two novel ML algorithms to in-vivo spontaneous, low-SNR vagus nerve recordings. The performance of each approach is compared using the task of extracting respiratory afferent activity and validated using cross-correlation, MSE, and accuracy in terms of extracting the true respiratory rate. A variational autoencoder (VAE) model in particular produces results that show better correlation with respiratory activity compared to bandpass filtering, highlighting that these models have the potential to preserve relevant features in complex neural recordings.

From data to insight: Exploring contaminants in different food groups with literature mining and machine learning techniques.

Food remains a major source of human exposure to chemical contaminants that are unintentionally present in commodities globally, despite strict regulation. Scientific literature is a valuable source of quantification data on those contaminants in various foods, but manually summarizing the information is not practicable. In this review, literature mining and machine learning techniques were applied in 72 foods to obtain relevant information on 96 contaminants, including heavy metals, polychlorinated biphenyls, dioxins, furans, polycyclic aromatic hydrocarbons (PAHs), pesticides, mycotoxins, and heterocyclic aromatic amines (HAAs). The 11,723 data points collected from 254 papers from the last two decades were then used to identify the patterns of contaminants distribution. Considering contaminant categories, metals were the most studied globally, followed by PAHs, mycotoxins, pesticides, and HAAs. As for geographical region, the distribution was uneven, with Europe and Asia having the highest number of studies, followed by North and South America, Africa and Oceania. Regarding food groups, all contained metals, while PAHs were found in seven out of 12 groups. Mycotoxins were found in six groups, and pesticides in almost all except meat, eggs, and vegetable oils. HAAs appeared in only three food groups, with fish and seafood reporting the highest levels. The median concentrations of contaminants varied across food groups, with citrinin having the highest median value. The information gathered is highly relevant to explore, establish connections, and identify patterns between diverse datasets, aiming at a comprehensive view of food contamination.

Intracellular Antioxidant and Anti-Inflammatory Effects and Bioactive Profiles of Coffee Cascara and Black Tea Kombucha Beverages.

Kombucha is a functional beverage obtained through fermentation of sweetened Camellia sinensis infusion by a symbiotic culture of bacteria and yeasts that exerts many beneficial biological effects, mostly related to its antioxidant and anti-inflammatory effects. Alternative raw materials have been used to create new kombucha or kombucha-like products. Coffee is the most important food commodity worldwide and generates large amounts of by-products during harvest and post-harvest processing. The main coffee by-product is the dried fruit skin and pulp, popularly known as cascara. To date, no studies have evaluated the potential bioactivity of coffee cascara kombucha. In this study, we aimed to measure and compare the effects of infusions and kombuchas made with arabica coffee cascaras (n = 2) and black tea leaves (n = 1), fermented for 0, 3, 6, and 9 days on the intracellular production of Reactive Oxygen Species (ROS) and Nitric Oxide (NO) in model cells. Oxidative stress was induced in HK-2 cells with indoxyl sulfate (IS) and high glucose (G). Inflammation was induced with lipopolysaccharide (LPS) in RAW 264.7 macrophage. The contents of phenolic compounds, caffeine, and other physicochemical parameters were evaluated. To the best of our knowledge, this is the first study providing information on the bioactive profile and on the potential biological effects of coffee cascara kombucha. Fermentation caused the release of bound phenolic compounds from the infusions, especially total chlorogenic acids, with an average increase from 5.4 to 10.7 mg/100 mL (98%) and 2.6-3.4 mg/100 mL (30%) in coffee cascara and black tea kombucha, respectively, up to day 9. All evaluated beverages reduced (p < 0.0001) similarly the intracellular ROS (41% reduction, on average) and uric acid (10-55%) concentrations in HK-2 model cells, reversing the induced oxidative stress. All beverages also reduced (p < 0.0001, 81-90%) NO formation in LPS-induced macrophages, exhibiting an anti-inflammatory effect. These potential health benefits may be mostly attributed to polyphenols and caffeine, whose contents were comparable in all beverages. Coffee cascara showed similar potential to C. sinensis to produce healthy beverages and support sustainable coffee production.

The assessment list for trustworthy artificial intelligence: A review and recommendations.

In July 2020, the European Commission's High-Level Expert Group on AI (HLEG-AI) published the Assessment List for Trustworthy Artificial Intelligence (ALTAI) tool, enabling organizations to perform self-assessments of the fit of their AI systems and surrounding governance to the "7 Principles for Trustworthy AI." Prior research on ALTAI has focused primarily on specific application areas, but there has yet to be a comprehensive analysis and broader recommendations aimed at proto-regulators and industry practitioners. This paper therefore starts with an overview of this tool, including an assessment of its strengths and limitations. The authors then consider the success by which the ALTAI tool is likely to be of utility to industry in improving understanding of the risks inherent in AI systems and best practices to mitigate such risks. It is highlighted how research and practices from fields such as Environmental Sustainability, Social Justice, and Corporate Governance (ESG) can be of benefit for addressing similar challenges in ethical AI development and deployment. Also explored is the extent to which the tool is likely to be successful in being taken up by industry, considering various factors pertaining to its likely adoption. Finally, the authors also propose recommendations applicable internationally to similar bodies to the HLEG-AI regarding the gaps needing to be addressed between high-level principles and practical support for those on the front-line developing or commercializing AI tools. In all, this work provides a comprehensive analysis of the ALTAI tool, as well as recommendations to relevant stakeholders, with the broader aim of promoting more widespread adoption of such a tool in industry.

Delphinidin-3-rutinoside from Blackcurrant Berries (Ribes nigrum): In Vitro Antiproliferative Activity and Interactions with Other Phenolic Compounds.

Blackcurrant berries (Rigrum L.) are of great interest for food scientists/technologists as a source of delphinidin-3-rutinoside (D3R). This is an uncommon phenolic compound in diets that unveils potent antiproliferative activity besides its colour. Other phenolic compounds, such as chlorogenic acid (CA) and epicatechin (EC), also known by their antiproliferative effects, are abundant in foods and beverages. To design smart food/supplements combinations containing blackcurrant and improved anticancer properties at the gastrointestinal level, there is the need for more data concerning the combined effects of those molecules. In this work, synergistic, additive, or antagonistic effects against gastric and intestinal cancers of D3R, CA, and EC were assessed in vitro. The antiproliferative activity of D3R, CA, and EC, alone and in binary combinations (D3R+CA, D3R+EC, and CA+EC) on NCI-N87 (gastric) and Caco-2 (intestinal) cells, was assessed following the Chou-Talalay theorem at equipotent contributions (i.e., (IC50)1/(IC50)2). D3R presented the strongest antiproliferative activity of the single molecules tested, with IC50 values of 24.9 µM and 102.5 µM on NCI-N87 and Caco-2 cells, respectively. The combinations D3R+CA and CA+EC were synergic against NCI-N87 until IC50 and IC75, respectively, while D3R+EC shifted from slight antagonism to synergism at higher doses. On Caco-2 cells, antagonism at low doses and synergism at high doses was observed. Therefore, the synergisms observed on the gastric cancer model at low doses occurred on the colon model only at high doses. Data herein described is vital to the targeted smart design of foods and supplements, as it is foreseen that the same combination of phenolic compounds causes different interactions/effects depending on the dose and gastrointestinal compartment.

Pig Ulnar Nerve Recording with Sinusoidal and Temporal Interference Stimulation.

Temporal interference stimulation has been suggested as a method to reach deep targets during transcutaneous electrical stimulation. Despite its growing use in transcutaneous stimulation therapies, the mechanism of its operation is not fully understood. Recent efforts to fill that gap have focused on computational modelling, in vitro and in vivo experiments relying on physical observations - e.g., sensation or movement. This paper expands the current range of experimental methods by demonstrating in vivo extraneural recordings from the ulnar nerve of a pig while applying temporal interference stimulation at a location targeting a distal part of the nerve. The main aim of the experiment was to compare neural activation using sinusoidal stimulation (100 Hz, 2 kHz, 4 kHz) and temporal interference stimulation (2 kHz and 4 kHz). The recordings showed a significant increase in the magnitude of stimulation artefacts at higher frequencies. While those artefacts could be removed and provided an indication of the depth of modulation, they resulted in the saturation of the amplifiers, limiting the stimulation currents and amplifier gains used. The results of the 100 Hz sine wave stimulation showed clear neural activity correlated to the stimulation waveform. However, this was not observed with temporal interference stimulation. The results suggest that, despite its greater penetration, higher currents might be required to observe a neural response with temporal interference stimulation, and more complex artefact rejection techniques may be required to validate the method.

An in-vitro system for closed loop neuromodulation of peripheral nerves.

Current neuromodulation research relies heavily on in-vivo animal experiments for developing novel devices and paradigms, which can be costly, time-consuming, and ethically contentious. As an alternative to this, in-vitro systems are being developed for examining explanted tissue in a controlled environment. However, these systems are typically tailored for cellular studies. Thus, this paper describes the development of an in-vitro system for electrically recording and stimulating large animal nerves. This is demonstrated experimentally using explanted pig ulnar nerves, which show evoked compound action potentials (eCAPs) when stimulated. These eCAPs were examined both in the time and velocity domain at a baseline temperature of 20° C, and at temperatures increasing up to those seen in-vivo (37°C). The results highlight that as the temperature is increased within the in-vitro system, faster conduction velocities (CVs) similar to those present in-vivo can be observed. To our knowledge, this is the first time an in-vitro peripheral nerve system has been validated against in-vivo data, which is crucial for promoting more widespread adoption of such systems for the optimisation of neural interfaces.

Adaptation of the Two-CAP Method for Conduction Velocity Distribution Estimation in Multi-Channel Recordings.

Closed-loop neural interfaces capable of both stimulating and recording from peripheral nerves have the potential to enhance the long-term efficacy of neural implants. One challenge associated with closed loop interfaces is the accurate estimation of the distribution of active fibre conduction velocities (DCV) when recording the immediate effect of stimulation. DCV estimation has been performed in monopolar surface recordings using the Two-CAP method. This work extends the Two-CAP method and demonstrates its application to bipolar in-vivo recordings made with multiple-electrode arrays. A sensitivity analysis was conducted using simulated data with ground truth to ascertain the stability and limits of the algorithm before experimental data was examined. The sensitivity analysis highlighted that recording distance shows a considerable impact on the performance of this extended Two-CAP method, as well as the velocity interval chosen when creating the model. The in-vivo data was also compared against an equivalent simulated model, and a relatively low mean squared error was obtained when comparing the two distributions.

Nutritional Characterization of Strychnos madagascariensis Fruit Flour Produced by Mozambican Communities and Evaluation of Its Contribution to Nutrient Adequacy.

The indigenous fruit Strychnos madagascariensis is usually processed to flour, called nfuma, being highly consumed during staple food shortage. This study aimed to evaluate the nutritional composition of nfuma and its nutrient adequacy. Flours from four districts of Mozambique were analyzed using AOAC methods for proximate composition, HPLC for sugar, amino acids (AA), vitamin E and carotenoids and ICP-MS and FAAS for minerals. The results showed that nfuma stands out for its high content of fat (26.3-27.8%), mainly oleic acid, fiber (>6%), vitamin E (6.7 to 8.0 mg/100 g) and carotenes (2.2 to 2.6 mg/100 g). The main amino acids of nfuma protein were Arg, Asp and Glu, and Lys was the limiting one. The mineral composition reveals K (~1200 to 1700 mg/100 g) as the main macromineral followed by Mg > Ca > Na. The main trace element was Mn (~4 mg/100 g) followed by Fe > Zn > Cu > Cr > Co. Aluminum (~3 mg/100 g) was the main non-essential element and Rb, Ni, Sr, Ba, V, Cd were also quantified. Assuming the daily consumption of 50 g, nfuma provides 82% of Vitamin A dietary reference value for toddlers, while the consumption of 100 g contributes to 132% and 60% of Mn and vitamin A DRV for adults, respectively. Despite the nutritional advantages of nfuma, this flour can be a source of Ni, highlighting the importance of the study of good practices in its preparation to decrease the exposure to non-essential elements.

Microfluidics Integration into Low-Noise Multi-Electrode Arrays.

Organ-on-Chip technology is commonly used as a tool to replace animal testing in drug development. Cells or tissues are cultured on a microchip to replicate organ-level functions, where measurements of the electrical activity can be taken to understand how the cell populations react to different drugs. Microfluidic structures are integrated in these devices to replicate more closely an in vivo microenvironment. Research has provided proof of principle that more accurate replications of the microenvironment result in better micro-physiological behaviour, which in turn results in a higher predictive power. This work shows a transition from a no-flow (static) multi-electrode array (MEA) to a continuous-flow (dynamic) MEA, assuring a continuous and homogeneous transfer of an electrolyte solution across the measurement chamber. The process through which the microfluidic system was designed, simulated, and fabricated is described, and electrical characterisation of the whole structure under static solution and a continuous flow rate of 80 µL/min was performed. The latter reveals minimal background disturbance, with a background noise below 30 µVpp for all flow rates and areas. This microfluidic MEA, therefore, opens new avenues for more accurate and long-term recordings in Organ-on-Chip systems.

Human Breast Cancer Cells Demonstrate Electrical Excitability.

Breast cancer is one of the most prevalent types of cancers worldwide and yet, its pathophysiology is poorly understood. Single-cell electrophysiological studies have provided evidence that membrane depolarization is implicated in the proliferation and metastasis of breast cancer. However, metastatic breast cancer cells are highly dynamic microscopic systems with complexities beyond a single-cell level. There is an urgent need for electrophysiological studies and technologies capable of decoding the intercellular signaling pathways and networks that control proliferation and metastasis, particularly at a population level. Hence, we present for the first time non-invasive in vitro electrical recordings of strongly metastatic MDA-MB-231 and weakly/non-metastatic MCF-7 breast cancer cell lines. To accomplish this, we fabricated an ultra-low noise sensor that exploits large-area electrodes, of 2 mm2, which maximizes the double-layer capacitance and concomitant detection sensitivity. We show that the current recorded after adherence of the cells is dominated by the opening of voltage-gated sodium channels (VGSCs), confirmed by application of the highly specific inhibitor, tetrodotoxin (TTX). The electrical activity of MDA-MB-231 cells surpasses that of the MCF-7 cells, suggesting a link between the cells' bioelectricity and invasiveness. We also recorded an activity pattern with characteristics similar to that of Random Telegraph Signal (RTS) noise. RTS patterns were less frequent than the asynchronous VGSC signals. The RTS noise power spectral density showed a Lorentzian shape, which revealed the presence of a low-frequency signal across MDA-MB-231 cell populations with propagation speeds of the same order as those reported for intercellular Ca2+ waves. Our recording platform paves the way for real-time investigations of the bioelectricity of cancer cells, their ionic/pharmacological properties and relationship to metastatic potential.