In silico study of the mechanisms of hypoxia and contractile dysfunction during ischemia and reperfusion of hiPSC cardiomyocytes.

Interconnected mechanisms of ischemia and reperfusion (IR) has increased the interest in IR in vitro experiments using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). We developed a whole-cell computational model of hiPSC-CMs including the electromechanics, a metabolite-sensitive sarcoplasmic reticulum Ca2+-ATPase (SERCA) and an oxygen dynamics formulation to investigate IR mechanisms. Moreover, we simulated the effect and action mechanism of levosimendan, which recently showed promising anti-arrhythmic effects in hiPSC-CMs in hypoxia. The model was validated using hiPSC-CM and in vitro animal data. The role of SERCA in causing relaxation dysfunction in IR was anticipated to be comparable to its function in sepsis-induced heart failure. Drug simulations showed that levosimendan counteracts the relaxation dysfunction by utilizing a particular Ca2+-sensitizing mechanism involving Ca2+-bound troponin C and Ca2+ flux to the myofilament, rather than inhibiting SERCA phosphorylation. The model demonstrates extensive characterization and promise for drug development, making it suitable for evaluating IR therapy strategies based on the changing levels of cardiac metabolites, oxygen and molecular pathways.

Simulation-based feasibility study of monitoring of intracerebral hemorrhages and detection of secondary hemorrhages using electrical impedance tomography.

Purpose: We present a simulation-based feasibility study of electrical impedance tomography (EIT) for continuous bedside monitoring of intracerebral hemorrhages (ICH) and detection of secondary hemorrhages. Approach: We simulated EIT measurements for six different hemorrhage sizes at two different hemorrhage locations using an anatomically detailed computational head model. Using this dataset, we test the ICH monitoring and detection performance of our tailor-made, patient-specific stroke-monitoring algorithm that utilizes a novel combination of nonlinear region-of-interest difference imaging, parallel level sets regularization and a prior-conditioned least squares algorithm. We compare the results of our algorithm to the results of two reference algorithms, a total variation regularized absolute imaging algorithm and a linear difference imaging algorithm. Results: The tailor-made stroke-monitoring algorithm is capable of indicating smaller changes in the simulated hemorrhages than either of the reference algorithms, indicating better monitoring and detection performance. Conclusions: Our simulation results from the anatomically detailed head model indicate that EIT equipped with a patient-specific stroke-monitoring algorithm is a promising technology for the unmet clinical need of having a technology for continuous bedside monitoring of brain status of acute stroke patients.

Ascorbic Acid 2-Phosphate Releasing Supercritically Foamed Porous Poly-L-Lactide-Co-ε-Caprolactone Scaffold Enhances the Collagen Production of Human Vaginal Stromal Cells: A New Approach for Vaginal Tissue Engineering.

BACKGROUND: The reconstructive surgery of vaginal defects is highly demanding and susceptible to complications, especially in larger defects requiring nonvaginal tissue grafts. Thus, tissue engineering-based solutions could provide a potential approach to the reconstruction of vaginal defects. METHODS: Here, we evaluated a novel porous ascorbic acid 2-phosphate (A2P)-releasing supercritical carbon dioxide foamed poly-L-lactide-co-ε-caprolactone (scPLCLA2P) scaffold for vaginal reconstruction with vaginal epithelial (EC) and stromal (SC) cells. The viability, proliferation, and phenotype of ECs and SCs were evaluated in monocultures and in cocultures on d 1, d 7 and d 14. Furthermore, the collagen production of SCs on scPLCLA2P was compared to that on scPLCL without A2P on d 14. RESULTS: Both ECs and SCs maintained their viability on the scPLCLA2P scaffold in mono- and coculture conditions, and the cells maintained their typical morphology during the 14-d culture period. Most importantly, the scPLCLA2P scaffolds supported the collagen production of SCs superior to plain scPLCL based on total collagen amount, collagen I and III gene expression results and collagen immunostaining results. CONCLUSION: This is the first study evaluating the effect of A2P on vaginal tissue engineering, and the results are highly encouraging, indicating that scPLCLA2P has potential as a scaffold for vaginal tissue engineering.

The Effect of Substrate Stiffness on Elastic Force Transmission in the Epithelial Monolayers over Short Timescales.

Purpose: The importance of mechanical forces and microenvironment in guiding cellular behavior has been widely accepted. Together with the extracellular matrix (ECM), epithelial cells form a highly connected mechanical system subjected to various mechanical cues from their environment, such as ECM stiffness, and tensile and compressive forces. ECM stiffness has been linked to many pathologies, including tumor formation. However, our understanding of the effect of ECM stiffness and its heterogeneities on rapid force transduction in multicellular systems has not been fully addressed. Methods: We used experimental and computational methods. Epithelial cells were cultured on elastic hydrogels with fluorescent nanoparticles. Single cells were moved by a micromanipulator, and epithelium and substrate deformation were recorded. We developed a computational model to replicate our experiments and quantify the force distribution in the epithelium. Our model further enabled simulations with local stiffness gradients. Results: We found that substrate stiffness affects the force transduction and the cellular deformation following an external force. Also, our results indicate that the heterogeneities, e.g., gradients, in the stiffness can substantially influence the strain redistribution in the cell monolayers. Furthermore, we found that the cells' apico-basal elasticity provides a level of mechanical isolation between the apical cell-cell junctions and the basal focal adhesions. Conclusions: Our simulation results show that increased ECM stiffness, e.g., due to a tumor, can mechanically isolate cells and modulate rapid mechanical signaling between cells over distances. Furthermore, the developed model has the potential to facilitate future studies on the interactions between epithelial monolayers and elastic substrates. Supplementary Information: The online version of this article (10.1007/s12195-023-00772-0) contains supplementary material, which is available to authorized users.

Multifocal optical projection microscopy enables label-free 3D measurement of cardiomyocyte cluster contractility.

Human induced pluripotent stem cell (hiPSC)-derived cardiomyocyte (CM) models have become an attractive tool for in vitro cardiac disease modeling and drug studies. These models are moving towards more complex three-dimensional microphysiological organ-on-chip systems. Label-free imaging-based techniques capable of quantifying contractility in 3D are needed, as traditional two-dimensional methods are ill-suited for 3D applications. Here, we developed multifocal (MF) optical projection microscopy (OPM) by integrating an electrically tunable lens to our in-house built optical projection tomography setup for extended depth of field brightfield imaging in CM clusters. We quantified cluster biomechanics by implementing our previously developed optical flow-based CM video analysis for MF-OPM. To demonstrate, we acquired and analyzed multiangle and multifocal projection videos of beating hiPSC-CM clusters in 3D hydrogel. We further quantified cluster contractility response to temperature and adrenaline and observed changes to beating rate and relaxation. Challenges emerge from light penetration and overlaying textures in larger clusters. However, our findings indicate that MF-OPM is suitable for contractility studies of 3D clusters. Thus, for the first time, MF-OPM is used in CM studies and hiPSC-CM 3D cluster contraction is quantified in multiple orientations and imaging planes.

Improvements in Maturity and Stability of 3D iPSC-Derived Hepatocyte-like Cell Cultures.

Induced pluripotent stem cell (iPSC) technology enables differentiation of human hepatocytes or hepatocyte-like cells (iPSC-HLCs). Advances in 3D culturing platforms enable the development of more in vivo-like liver models that recapitulate the complex liver architecture and functionality better than traditional 2D monocultures. Moreover, within the liver, non-parenchymal cells (NPCs) are critically involved in the regulation and maintenance of hepatocyte metabolic function. Thus, models combining 3D culture and co-culturing of various cell types potentially create more functional in vitro liver models than 2D monocultures. Here, we report the establishment of 3D cultures of iPSC-HLCs alone and in co-culture with human umbilical vein endothelial cells (HUVECs) and adipose tissue-derived mesenchymal stem/stromal cells (hASCs). The 3D cultures were performed as spheroids or on microfluidic chips utilizing various biomaterials. Our results show that both 3D spheroid and on-chip culture enhance the expression of mature liver marker genes and proteins compared to 2D. Among the spheroid models, we saw the best functionality in iPSC-HLC monoculture spheroids. On the contrary, in the chip system, the multilineage model outperformed the monoculture chip model. Additionally, the optical projection tomography (OPT) and electrical impedance tomography (EIT) system revealed changes in spheroid size and electrical conductivity during spheroid culture, suggesting changes in cell-cell connections. Altogether, the present study demonstrates that iPSC-HLCs can successfully be cultured in 3D as spheroids and on microfluidic chips, and co-culturing iPSC-HLCs with NPCs enhances their functionality. These 3D in vitro liver systems are promising human-derived platforms usable in various liver-related studies, specifically when using patient-specific iPSCs.

Substrate microtopographies induce cellular alignment and affect nuclear force transduction.

Cellular physiology has been mainly studied by using two-dimensional cell culture substrates which lack in vivo-mimicking extracellular environment and interactions. Thus, there is a growing need for more complex model systems in life sciences. Micro-engineered scaffolds have been proven to be a promising tool in understanding the role of physical cues in the co-regulation of cellular functions. These tools allow, for example, probing cell morphology and migration in response to changes in chemo-physical properties of their microenvironment. In order to understand how microtopographical features, what cells encounter in vivo, affect cytoskeletal organization and nuclear mechanics, we used direct laser writing via two-photon polymerization (TPP) to fabricate substrates which contain different surface microtopographies. By combining with advanced high-resolution spectral imaging, we describe how the constructed grid and vertical line microtopographies influence cellular alignment, nuclear morphology and mechanics. Specifically, we found that growing cells on grids larger than 10 × 20 µm2 and on vertical lines increased 3D actin cytoskeleton orientation along the walls of microtopographies and abolished basal actin stress fibers. In concert, the nuclei of these cells were also more aligned, elongated, deformed and less flattened, indicating changes in nuclear force transduction. Importantly, by using fluorescence lifetime imaging microscopy for measuring Förster resonance energy transfer for a genetically encoded nesprin-2 molecular tension sensor, we show that growing cells on these microtopographic substrates induce lower mechanical tension at the nuclear envelope. To conclude, here used substrate microtopographies modulated the cellular mechanics, and affected actin organization and nuclear force transduction.

Transcriptional Landscape of 3D vs. 2D Ovarian Cancer Cell Models.

Three-dimensional (3D) cancer models are revolutionising research, allowing for the recapitulation of an in vivo-like response through the use of an in vitro system, which is more complex and physiologically relevant than traditional monolayer cultures. Cancers such as ovarian (OvCa) are prone to developing resistance, are often lethal, and stand to benefit greatly from the enhanced modelling emulated by 3D cultures. However, the current models often fall short of the predicted response, where reproducibility is limited owing to the lack of standardised methodology and established protocols. This meta-analysis aims to assess the current scope of 3D OvCa models and the differences in the genetic profiles presented by a vast array of 3D cultures. An analysis of the literature (Pubmed.gov) spanning 2012-2022 was used to identify studies with paired data of 3D and 2D monolayer counterparts in addition to RNA sequencing and microarray data. From the data, 19 cell lines were found to show differential regulation in their gene expression profiles depending on the bio-scaffold (i.e., agarose, collagen, or Matrigel) compared to 2D cell cultures. The top genes differentially expressed in 2D vs. 3D included C3, CXCL1, 2, and 8, IL1B, SLP1, FN1, IL6, DDIT4, PI3, LAMC2, CCL20, MMP1, IFI27, CFB, and ANGPTL4. The top enriched gene sets for 2D vs. 3D included IFN-α and IFN-γ response, TNF-α signalling, IL-6-JAK-STAT3 signalling, angiogenesis, hedgehog signalling, apoptosis, epithelial-mesenchymal transition, hypoxia, and inflammatory response. Our transversal comparison of numerous scaffolds allowed us to highlight the variability that can be induced by these scaffolds in the transcriptional landscape and identify key genes and biological processes that are hallmarks of cancer cells grown in 3D cultures. Future studies are needed to identify which is the most appropriate in vitro/preclinical model to study tumour microenvironments.

Iterative immunostaining combined with expansion microscopy and image processing reveals nanoscopic network organization of nuclear lamina.

Investigation of nuclear lamina architecture relies on superresolved microscopy. However, epitope accessibility, labeling density, and detection precision of individual molecules pose challenges within the molecularly crowded nucleus. We developed iterative indirect immunofluorescence (IT-IF) staining approach combined with expansion microscopy (ExM) and structured illumination microscopy to improve superresolution microscopy of subnuclear nanostructures like lamins. We prove that ExM is applicable in analyzing highly compacted nuclear multiprotein complexes such as viral capsids and provide technical improvements to ExM method including three-dimensional-printed gel casting equipment. We show that in comparison with conventional immunostaining, IT-IF results in a higher signal-to-background ratio and a mean fluorescence intensity by improving the labeling density. Moreover, we present a signal-processing pipeline for noise estimation, denoising, and deblurring to aid in quantitative image analyses and provide this platform for the microscopy imaging community. Finally, we show the potential of signal-resolved IT-IF in quantitative superresolution ExM imaging of nuclear lamina and reveal nanoscopic details of the lamin network organization-a prerequisite for studying intranuclear structural coregulation of cell function and fate.

Altered contractility in mutation-specific hypertrophic cardiomyopathy: A mechano-energetic in silico study with pharmacological insights.

Introduction: Mavacamten (MAVA), Blebbistatin (BLEB), and Omecamtiv mecarbil (OM) are promising drugs directly targeting sarcomere dynamics, with demonstrated efficacy against hypertrophic cardiomyopathy (HCM) in (pre)clinical trials. However, the molecular mechanism affecting cardiac contractility regulation, and the diseased cell mechano-energetics are not fully understood yet. Methods: We present a new metabolite-sensitive computational model of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) electromechanics to investigate the pathology of R403Q HCM mutation and the effect of MAVA, BLEB, and OM on the cell mechano-energetics. Results: We offer a mechano-energetic HCM calibration of the model, capturing the prolonged contractile relaxation due to R403Q mutation (∼33%), without assuming any further modifications such as an additional Ca2+ flux to the thin filaments. The HCM model variant correctly predicts the negligible alteration in ATPase activity in R403Q HCM condition compared to normal hiPSC-CMs. The simulated inotropic effects of MAVA, OM, and BLEB, along with the ATPase activities in the control and HCM model variant agree with in vitro results from different labs. The proposed model recapitulates the tension-Ca2+ relationship and action potential duration change due to 1 µM OM and 5 µM BLEB, consistently with in vitro data. Finally, our model replicates the experimental dose-dependent effect of OM and BLEB on the normalized isometric tension. Conclusion: This work is a step toward deep-phenotyping the mutation-specific HCM pathophysiology, manifesting as altered interfilament kinetics. Accordingly, the modeling efforts lend original insights into the MAVA, BLEB, and OM contributions to a new interfilament balance resulting in a cardioprotective effect.

Validation of the X-ray microtomography in the assessment of duodenal morphometry and surface area in celiac disease.

Background: Duodenal histology remains the diagnostic reference standard in celiac disease. However, traditional methods have suboptimal sensitivity and reproducibility for early mucosal changes and research purposes. We validated a recently introduced micro-CT imaging method for an accurate digital evaluation of duodenal histomorphometry and mucosal surface areas. Methods: Endoscopic biopsies from 58 individuals were utilized for the micro-CT imaging, selecting histological changes ranging from normal to severely damaged mucosa. The imaging protocol was optimized for practicability and resolution. The Bland-Altman method was applied to test intra- and interobserver variations in the blinded measurements. Results: The 3D micro-CT reconstructions enabled easy and precise digital cutting with optimal orientation and computer-assisted measurement of the surface area. Intraobserver analysis of morphological measurements showed a mean difference of 0.011 with limits of agreement (LA) from -0.397 to 0.375 and a standard deviation (SD) of 0.197. The corresponding figures for interobserver analysis were 0.080, from -0.719 to 0.537 and 0.320, respectively. The intraclass correlation coefficients (ICC) for the intraobserver and interobserver variations were 0.981 and 0.954, respectively. Intraobserver surface area analysis yielded a mean difference of 0.010, LA from -0.764 to 0.785 and an SD of 0.395, and an interobserver analysis mean difference of 0.028, LA from -0.642 to 0.698 and SD of 0.342. The respective ICCs for the intra- and interobserver variations were 0.963 and 0.972. Conclusions: Micro-CT showed excellent accuracy and reproducibility in the evaluation of mucosal morphometry and surface areas. The improved sensitivity for histological changes is a powerful tool for the diagnosis of celiac disease and for clinical and pharmacological studies.

Objective Supervised Machine Learning-Based Classification and Inference of Biological Neuronal Networks.

The classification of biological neuron types and networks poses challenges to the full understanding of the human brain's organisation and functioning. In this paper, we develop a novel objective classification model of biological neuronal morphology and electrical types and their networks, based on the attributes of neuronal communication using supervised machine learning solutions. This presents advantages compared to the existing approaches in neuroinformatics since the data related to mutual information or delay between neurons obtained from spike trains are more abundant than conventional morphological data. We constructed two open-access computational platforms of various neuronal circuits from the Blue Brain Project realistic models, named Neurpy and Neurgen. Then, we investigated how we could perform network tomography with cortical neuronal circuits for the morphological, topological and electrical classification of neurons. We extracted the simulated data of 10,000 network topology combinations with five layers, 25 morphological type (m-type) cells, and 14 electrical type (e-type) cells. We applied the data to several different classifiers (including Support Vector Machine (SVM), Decision Trees, Random Forest, and Artificial Neural Networks). We achieved accuracies of up to 70%, and the inference of biological network structures using network tomography reached up to 65% of accuracy. Objective classification of biological networks can be achieved with cascaded machine learning methods using neuron communication data. SVM methods seem to perform better amongst used techniques. Our research not only contributes to existing classification efforts but sets the road-map for future usage of brain-machine interfaces towards an in vivo objective classification of neurons as a sensing mechanism of the brain's structure.

A method for evaluating sensitivity of electromagnetic localization systems for wireless capsule endoscopes.

This paper studies the use of electromagnetic induction in localization of wireless capsule endoscopes (WECs). There is still currently a need for an accurate localization system to enable localizing possible findings in the gastrointestinal tract, and to develop an active steering system for the capsule. Developing an optimal localization system requires the sensitivity of the system to be analyzed. In this paper, three different coil geometries are modelled with a computer simulation platform, and their sensitivities and target responses are compared. In order to do that, a formulation for the sensitivity based on the dipole model approximation is presented. The first coil array is based on literature and is used as a reference. The second array presents how having more transmit-receive channels in the array effects the sensitivity. The third coil array simulates the effect of increasing the field excitation intensity in different directions by using a three-axial Helmholtz array. In addition, both proposed coil arrays utilize larger coils than the reference. As a result, it seems that both increasing the coil size and the number of field projections interrogating the target increase the overall sensitivity in the region of interest and the target response. The findings suggest that an optimal coil array could utilize both large coils and multiple transmit-receive channels to increase the number of independent fields incident onto the target.

Corrections to "Sensitivity Analysis Highlights the Importance of Accurate Head Models for Electrical Impedance Tomography Monitoring of Intracerebral Hemorrhagic Stroke".

In the above paper [1] there are two errors which we correct here. An important reference was omitted.

Effect of fall direction on the lower hip fracture risk in athletes with different loading histories: A finite element modeling study in multiple sideways fall configurations.

Physical loading makes bones stronger through structural adaptation. Finding effective modes of exercise to improve proximal femur strength has the potential to decrease hip fracture risk. Previous proximal femur finite element (FE) modeling studies have indicated that the loading history comprising impact exercises is associated with substantially higher fracture load. However, those results were limited only to one specified fall direction. It remains thus unclear whether exercise-induced higher fracture load depends on the fall direction. To address this, using magnetic resonance images of proximal femora from 91 female athletes (mean age 24.7 years with >8 years competitive career) and their 20 non-athletic but physically active controls (mean age 23.7 years), proximal femur FE models were created in 12 different sideways fall configurations. The athletes were divided into five groups by typical loading patterns of their sports: high-impact (H-I: 9 triple- and 10 high-jumpers), odd-impact (O-I: 9 soccer and 10 squash players), high-magnitude (H-M: 17 powerlifters), repetitive-impact (R-I: 18 endurance runners), and repetitive non-impact (R-NI: 18 swimmers). Compared to the controls, the FE models showed that the H-I and R-I groups had significantly (p < 0.05) higher fracture loads, 11-17% and 22-28% respectively, in all fall directions while the O-I group had significantly 10-11% higher fracture loads in four fall directions. The H-M and R-NI groups did not show significant benefit in any direction. Also, the analyses of the minimum fall strength (MFS) among these multiple fall configurations confirmed significantly 15%, 11%, and 14% higher MFSs in these impact groups, respectively, compared to the controls. These results suggest that the lower hip fracture risk indicated by higher fracture loads in athletes engaged in high impact or repetitive impact sports is independent of fall direction whereas the lower fracture risk attributed to odd-impact exercise is more modest and specific to the fall direction. Moreover, in concordance with the literature, the present study also confirmed that the fracture risk increases if the impact is imposed on the more posterolateral aspect of the hip. The present results highlight the importance of engaging in the impact exercises to prevent hip fractures and call for retrospective studies to investigate whether specific impact exercise history in adolescence and young adulthood is also associated with lower incidence of hip fractures in later life.

Humanin vivoliver and tumor bioimpedance measured with biopsy needle.

Objective.Liver biopsy is an essential procedure in cancer diagnostics but targeting the biopsy to the actual tumor tissue is challenging. Aim of this study was to evaluate the clinical feasibility of a novel bioimpedance biopsy needle system in liver biopsy and simultaneously to gatherin vivobioimpedance data from human liver and tumor tissues.Approach.We measured human liver and tumor impedance datain vivofrom 26 patients who underwent diagnostic ultrasound-guided liver biopsy. Our novel 18 G core biopsy needle tip forms a bipolar electrode that was used to measure bioimpedance during the biopsy in real-time with frequencies from 1 kHz to 349 kHz. The needle tip location was determined by ultrasound. Also, the sampled tissue type was determined histologically.Main results.The bioimpedance values showed substantial variation between individual cases, and liver and tumor data overlapped each other. However, Mann-Whitney U test showed that the median bioimpedance values of liver and tumor tissue are significantly (p < 0.05) different concerning the impedance magnitude at frequencies below 25 kHz and the phase angle at frequencies below 3 kHz and above 30 kHz.Significance.This study uniquely employed a real-time bioimpedance biopsy needle in clinical liver biopsies and reported the measured humanin vivoliver and tumor impedance data. Impedance is always device-dependent and therefore not directly comparable to measurements with other devices. Although the variation in tumor types prevented coherent tumor identification, our study provides preliminary evidence that tumor tissue differs from liver tissuein vivo,and this association is frequency-dependent.

Sensitivity Analysis Highlights the Importance of Accurate Head Models for Electrical Impedance Tomography Monitoring of Intracerebral Hemorrhagic Stroke.

OBJECTIVE: Electrical impedance tomography (EIT) has been proposed as a novel tool for diagnosing stroke. However, so far, the clinical feasibility is unresolved. In this study, we aim to investigate the need for accurate head modeling in EIT and how the inhomogeneities of the head contribute to the EIT measurement and affect its feasibility in monitoring the progression of a hemorrhagic stroke. METHODS: We compared anatomically detailed six- and three-layer finite element models of a human head and computed the resulting scalp electrode potentials and the lead fields of selected electrode configurations. We visualized the resulting EIT measurement sensitivity distributions, computed the scalp electrode potentials, and examined the inverse imaging with selected cases. The effect of accurate tissue geometry and conductivity values on the EIT measurement is assessed with multiple different hemorrhagic perturbation locations and sizes. RESULTS: Our results show that accurate tissue geometries and conductivity values inside the cranial cavity, especially the highly conductive cerebrospinal fluid, significantly affect EIT measurement sensitivity distribution and measured potentials. CONCLUSIONS: We can conclude that the three-layer head models commonly used in EIT literature cannot depict the current paths correctly in the head. Thus, our study highlights the need to consider the detailed geometry of the cerebrospinal fluid (CSF) in EIT. SIGNIFICANCE: The results clearly show that the CSF should be considered in the head EIT calculations.

Murine cerebral organoids develop network of functional neurons and hippocampal brain region identity.

Brain organoids are in vitro three-dimensional (3D) self-organized neural structures, which can enable disease modeling and drug screening. However, their use for standardized large-scale drug screening studies is limited by their high batch-to-batch variability, long differentiation time (10-20 weeks), and high production costs. This is particularly relevant when brain organoids are obtained from human induced pluripotent stem cells (iPSCs). Here, we developed, for the first time, a highly standardized, reproducible, and fast (5 weeks) murine brain organoid model starting from embryonic neural stem cells. We obtained brain organoids, which progressively differentiated and self-organized into 3D networks of functional neurons with dorsal forebrain phenotype. Furthermore, by adding the morphogen WNT3a, we generated brain organoids with specific hippocampal region identity. Overall, our results showed the establishment of a fast, robust and reproducible murine 3D in vitro brain model that may represent a useful tool for high-throughput drug screening and disease modeling.

Astrocytes Exhibit a Protective Role in Neuronal Firing Patterns under Chemically Induced Seizures in Neuron-Astrocyte Co-Cultures.

Astrocytes and neurons respond to each other by releasing transmitters, such as γ-aminobutyric acid (GABA) and glutamate, that modulate the synaptic transmission and electrochemical behavior of both cell types. Astrocytes also maintain neuronal homeostasis by clearing neurotransmitters from the extracellular space. These astrocytic actions are altered in diseases involving malfunction of neurons, e.g., in epilepsy, Alzheimer's disease, and Parkinson's disease. Convulsant drugs such as 4-aminopyridine (4-AP) and gabazine are commonly used to study epilepsy in vitro. In this study, we aim to assess the modulatory roles of astrocytes during epileptic-like conditions and in compensating drug-elicited hyperactivity. We plated rat cortical neurons and astrocytes with different ratios on microelectrode arrays, induced seizures with 4-AP and gabazine, and recorded the evoked neuronal activity. Our results indicated that astrocytes effectively counteracted the effect of 4-AP during stimulation. Gabazine, instead, induced neuronal hyperactivity and synchronicity in all cultures. Furthermore, our results showed that the response time to the drugs increased with an increasing number of astrocytes in the co-cultures. To the best of our knowledge, our study is the first that shows the critical modulatory role of astrocytes in 4-AP and gabazine-induced discharges and highlights the importance of considering different proportions of cells in the cultures.

A mathematical model of hiPSC cardiomyocytes electromechanics.

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are becoming instrumental in cardiac research, human-based cell level cardiotoxicity tests, and developing patient-specific care. As one of the principal functional readouts is contractility, we propose a novel electromechanical hiPSC-CM computational model named the hiPSC-CM-CE. This model comprises a reparametrized version of contractile element (CE) by Rice et al., 2008, with a new passive force formulation, integrated into a hiPSC-CM electrophysiology formalism by Paci et al. in 2020. Our simulated results were validated against in vitro data reported for hiPSC-CMs at matching conditions from different labs. Specifically, key action potential (AP) and calcium transient (CaT) biomarkers simulated by the hiPSC-CM-CE model were within the experimental ranges. On the mechanical side, simulated cell shortening, contraction-relaxation kinetic indices (RT50 and RT25 ), and the amplitude of tension fell within the experimental intervals. Markedly, as an inter-scale analysis, correct classification of the inotropic effects due to non-cardiomyocytes in hiPSC-CM tissues was predicted on account of the passive force expression introduced to the CE. Finally, the physiological inotropic effects caused by Verapamil and Bay-K 8644 and the aftercontractions due to the early afterdepolarizations (EADs) were simulated and validated against experimental data. In the future, the presented model can be readily expanded to take in pharmacological trials and genetic mutations, such as those involved in hypertrophic cardiomyopathy, and study arrhythmia trigger mechanisms.