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Background: Atrial fibrillation has an estimated prevalence of 1.5-2%, making it the most common cardiac arrhythmia. The processes that cause and sustain the disease are still not completely understood. An association between atrial fibrillation and systemic, as well as local, inflammatory processes has been reported. However, the exact mechanisms underlying this association have not been established. While it is understood that inflammatory macrophages can influence cardiac electrophysiology, a direct, causative relationship to atrial fibrillation has not been described. This study investigated the pro-arrhythmic effects of activated M1 macrophages on human induced pluripotent stem cell (hiPSC)-derived atrial cardiomyocytes, to propose a mechanistic link between inflammation and atrial fibrillation., Methods: Two hiPSC lines from healthy individuals were differentiated to atrial cardiomyocytes and M1 macrophages and integrated in an isogenic, pacing-free, atrial fibrillation-like coculture model. Electrophysiology characteristics of cocultures were analysed for beat rate irregularity, electrogram amplitude and conduction velocity using multi electrode arrays. Cocultures were additionally treated using glucocorticoids to suppress M1 inflammation. Bulk RNA sequencing was performed on coculture-isolated atrial cardiomyocytes and compared to meta-analyses of atrial fibrillation patient transcriptomes., Results: Multi electrode array recordings revealed M1 to cause irregular beating and reduced electrogram amplitude. Conduction analysis further showed significantly lowered conduction homogeneity in M1 cocultures. Transcriptome sequencing revealed reduced expression of key cardiac genes such as SCN5A, KCNA5, ATP1A1, and GJA5 in the atrial cardiomyocytes. Meta-analysis of atrial fibrillation patient transcriptomes showed high correlation to the in vitro model. Treatment of the coculture with glucocorticoids showed reversal of phenotypes, including reduced beat irregularity, improved conduction, and reversed RNA expression profiles., Conclusions: This study establishes a causal relationship between M1 activation and the development of subsequent atrial arrhythmia, documented as irregularity in spontaneous electrical activation in atrial cardiomyocytes cocultured with activated macrophages. Further, beat rate irregularity could be alleviated using glucocorticoids. Overall, these results point at macrophage-mediated inflammation as a potential AF induction mechanism and offer new targets for therapeutic development. The findings strongly support the relevance of the proposed hiPSC-derived coculture model and present it as a first of its kind disease model., (© 2024. The Author(s).)

Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disease leading to motor neuron loss. Currently mutations in > 40 genes have been linked to ALS, but the contribution of many genes and genetic mutations to the ALS pathogenic process remains poorly understood. Therefore, we first performed comparative interactome analyses of five recently discovered ALS-associated proteins (C21ORF2, KIF5A, NEK1, TBK1, and TUBA4A) which highlighted many novel binding partners, and both unique and shared interactors. The analysis further identified C21ORF2 as a strongly connected protein. The role of C21ORF2 in neurons and in the nervous system, and of ALS-associated C21ORF2 variants is largely unknown. Therefore, we combined human iPSC-derived motor neurons with other models and different molecular cell biological approaches to characterize the potential pathogenic effects of C21ORF2 mutations in ALS. First, our data show C21ORF2 expression in ALS-relevant mouse and human neurons, such as spinal and cortical motor neurons. Further, the prominent ALS-associated variant C21ORF2-V58L caused increased apoptosis in mouse neurons and movement defects in zebrafish embryos. iPSC-derived motor neurons from C21ORF2-V58L-ALS patients, but not isogenic controls, show increased apoptosis, and changes in DNA damage response, mitochondria and neuronal excitability. In addition, C21ORF2-V58L induced post-transcriptional downregulation of NEK1, an ALS-associated protein implicated in apoptosis and DDR. In all, our study defines the pathogenic molecular and cellular effects of ALS-associated C21ORF2 mutations and implicates impaired post-transcriptional regulation of NEK1 downstream of mutant C21ORF72 in ALS., (© 2024. The Author(s).)

Background: Reduction of adult hippocampal neurogenesis is an early critical event in Alzheimer's disease (AD), contributing to progressive memory loss and cognitive decline. Reduced levels of the nucleoporin 153 (Nup153), a key epigenetic regulator of NSC stemness, characterize the neural stem cells isolated from a mouse model of AD (3×Tg) (AD-NSCs) and determine their altered plasticity and gene expression., Methods: Nup153-regulated mechanisms contributing to NSC function were investigated: (1) in cultured NSCs isolated from AD and wild type (WT) mice by proteomics; (2) in vivo by lentiviral-mediated delivery of Nup153 or GFP in the hippocampus of AD and control mice analyzing neurogenesis and cognitive function; (3) in human iPSC-derived brain organoids obtained from AD patients and control subjects as a model of neurodevelopment., Results: Proteomic approach identified Nup153 interactors in WT- and AD-NSCs potentially implicated in neurogenesis regulation. Gene ontology (GO) analysis showed that Nup153-bound proteins in WT-NSCs were involved in RNA metabolism, nuclear import and epigenetic mechanisms. Nup153-bound proteins in AD-NSCs were involved in pathways of neurodegeneration, mitochondrial dysfunction, proteasomal processing and RNA degradation. Furthermore, recovery of Nup153 levels in AD-NSCs reduced the levels of oxidative stress markers and recovered proteasomal activity. Lentiviral-mediated delivery of Nup153 in the hippocampal niche of AD mice increased the proliferation of early progenitors, marked by BrdU/DCX and BrdU/PSANCAM positivity and, later, the integration of differentiating neurons in the cell granule layer (BrdU/NeuN + cells) compared with GFP-injected AD mice. Consistently, Nup153-injected AD mice showed an improvement of cognitive performance in comparison to AD-GFP mice at 1 month after virus delivery assessed by Morris Water Maze. To validate the role of Nup153 in neurogenesis we took advantage of brain organoids derived from AD-iPSCs characterized by fewer neuroepithelial progenitor loops and reduced differentiation areas. The upregulation of Nup153 in AD organoids recovered the formation of neural-like tubes and differentiation., Conclusions: Our data suggest that the positive effect of Nup153 on neurogenesis is based on a complex regulatory network orchestrated by Nup153 and that this protein is a valuable disease target., (© 2024. The Author(s).)

Background: Atrial fibrillation (AF) is a stroke risk factor that often remains undetected at hospital admission. Long-term Holter monitoring helps to identify patients with previously unrecognized AF. Asymmetric (ADMA) and symmetric dimethylarginine (SDMA) are elevated in AF in cross-sectional studies. We analyzed ADMA, SDMA, and other L-arginine metabolites to assess their association with AF in the Find-AF trial., Methods and Results: We included 280 patients presenting with acute cerebral ischemia. Patients presenting in sinus rhythm received 7-day Holter-ECG. Biomarkers were quantified by ultra-performance liquid chromatography-tandem mass spectrometry. We also analyzed protein methylation and L-arginine-related metabolites in human induced pluripotent stem cell-derived cardiomyocytes in vitro. ADMA and SDMA were elevated in 44 patients who presented with AF. SDMA, but not ADMA, was significantly elevated in patients newly diagnosed with AF in Holter-ECG as compared with those in sinus rhythm. SDMA plasma concentration >0.571 μmol/L significantly predicted presence of AF in Holter-ECG (area under the curve=0.676 [0.530-0.822]; P =0.029; sensitivity 0.786, specificity 0.572). SDMA levels further increased in patients with AF during the first 24 hours in hospital, and ADMA levels remained stable. In vitro, induced pluripotent stem cell-derived cardiomyocytes showed increased symmetric protein methylation and elevated SDMA during rapid pacing (2.0 Hz versus 0.5 Hz), whereas asymmetric protein methylation and ADMA were unchanged., Conclusions: SDMA at admission was significantly elevated in stroke patients presenting with AF and showed a moderate but significant prospective association with previously unrecognized AF. Thus, stroke patients with elevated SDMA concentration at admission may specifically benefit from extended Holter-ECG monitoring.

The synchronization of the electrical and mechanical coupling assures the physiological pump function of the heart, but life-threatening pathologies may jeopardize this equilibrium. Recently, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have emerged as a model for personalized investigation because they can recapitulate human diseased traits, such as compromised electrical capacity or mechanical circuit disruption. This research avails the model of hiPSC-CMs and showcases innovative techniques to study the electrical and mechanical properties as well as their modulation due to inherited cardiomyopathies. In this work, hiPSC-CMs carrying either Brugada syndrome (BRU) or dilated cardiomyopathy (DCM), were organized in a bilayer configuration to first validate the experimental methods and second mimic the physiological environment. High-density CMOS-based microelectrode arrays (HD-MEA) have been employed to study the electrical activity. Furthermore, mechanical function was investigated via quantitative video-based evaluation, upon stimulation with a β-adrenergic agonist. This study introduces two experimental methods. First, high-throughput mechanical measurements in the hiPSC-CM layers (xy-inspection) are obtained using both a recently developed optical tracker (OPT) and confocal reference-free traction force microscopy (cTFM) aimed to quantify cardiac kinematics. Second, atomic force microscopy (AFM) with FluidFM probes, combined with the xy-inspection methods, supplemented a three-dimensional understanding of cell-cell mechanical coupling (xyz-inspection). This particular combination represents a multi-technique approach to detecting electrical and mechanical latency among the cell layers, examining differences and possible implications following inherited cardiomyopathies. It can not only detect disease characteristics in the proposed in vitro model but also quantitatively assess its response to drugs, thereby demonstrating its feasibility as a scalable tool for clinical and pharmacological studies.

Background: Variants of the SCN5A gene, which encodes the Na V 1.5 cardiac sodium channel, have been linked to arrhythmic disorders associated with dilated cardiomyopathy (DCM). However, the precise pathological mechanisms remain elusive. The present study aimed to elucidate the pathophysiological consequences of the DCM-linked Nav1.5/R219H variant, which is known to generate a gating pore current, using patient-specific human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) cultured in monolayers., Methods: Ventricular- and atrial-like hiPSC-CM monolayers were generated from DCM patients carrying the R219H SCN5A variant as well as from healthy control individuals. CRISPR-corrected hiPSC-CMs served as isogenic controls. Simultaneous optical mapping of action potentials (APs) and calcium transients (CaTs) was employed to measure conduction velocities (CVs) and AP durations (APDs) and served as markers of electrical excitability. Calcium handling was evaluated by assessing CaT uptake (half-time to peak), recapture (tau of decay), and durations (TD 50 and TD 80 ). A multi-electrode array (MEA) analysis was conducted on hiPSC-CM monolayers to measure field potential (FP) parameters, including corrected Fridericia FP durations (FPDc)., Results: Our results revealed that CVs were significantly reduced by more than 50 % in both ventricular- and atrial-like hiPSC-CM monolayers carrying the R219H variant compared to the control group. APDs were also prolonged in the R219H group compared to the control and CRISPR-corrected groups. CaT uptake, reuptake, and duration were also markedly delayed in the R219H group compared to the control and CRISPR-corrected groups in both the ventricular- and the atrial-like hiPSC-CM monolayers. Lastly, the MEA data revealed a notably prolonged FPDc in the ventricular- and atrial-like hiPSC-CMs carrying the R219H variant compared to the control and isogenic control groups., Conclusions: These findings highlight the impact of the gating pore current on AP propagation and calcium homeostasis within a functional syncytium environment and offer valuable insights into the potential mechanisms underlying DCM pathophysiology., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Mohamed Chahine reports was provided by Laval University. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Charcot-Marie-Tooth disease type 1A (CMT1A) is the most common inherited peripheral neuropathy caused by a 1.5 Mb tandem duplication of chromosome 17 harbouring the PMP22 gene. This dose-dependent overexpression of PMP22 results in disrupted Schwann cell myelination of peripheral nerves. To obtain better insights into the underlying pathogenic mechanisms in CMT1A, we investigated the role of PMP22 duplication in cellular homeostasis in CMT1A mouse models and in patient-derived induced pluripotent stem cells differentiated into Schwann cell precursors (iPSC-SCPs). We performed lipidomic profiling and bulk RNA sequencing (RNA-seq) on sciatic nerves of two developing CMT1A mouse models and on CMT1A patient-derived iPSC-SCPs. For the sciatic nerves of the CMT1A mice, cholesterol and lipid metabolism was downregulated in a dose-dependent manner throughout development. For the CMT1A iPSC-SCPs, transcriptional analysis unveiled a strong suppression of genes related to autophagy and lipid metabolism. Gene ontology enrichment analysis identified disturbances in pathways related to plasma membrane components and cell receptor signalling. Lipidomic analysis confirmed the severe dysregulation in plasma membrane lipids, particularly sphingolipids, in CMT1A iPSC-SCPs. Furthermore, we identified reduced lipid raft dynamics, disturbed plasma membrane fluidity and impaired cholesterol incorporation and storage, all of which could result from altered lipid storage homeostasis in the patient-derived CMT1A iPSC-SCPs. Importantly, this phenotype could be rescued by stimulating autophagy and lipolysis. We conclude that PMP22 duplication disturbs intracellular lipid storage and leads to a more disordered plasma membrane owing to an alteration in the lipid composition, which might ultimately lead to impaired axo-glial interactions. Moreover, targeting lipid handling and metabolism could hold promise for the treatment of patients with CMT1A., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Epigenetic modifications (methylation, acetylation, etc.) of core histones play a key role in regulation of gene expression. Thus, the epigenome changes strongly during various biological processes such as cell differentiation and dedifferentiation. Classical methods of analysis of epigenetic modifications such as mass-spectrometry and chromatin immuno-precipitation, work with fixed cells only. Here we present a genetically encoded fluorescent probe, MPP8-Green, for detecting H3K9me3, a histone modification associated with inactive chromatin. This probe, based on the chromodomain of MPP8, allows for visualization of H3K9me3 epigenetic landscapes in single living cells. We used this probe to track changes in H3K9me3 landscapes during the differentiation of induced pluripotent stem cells (iPSCs) into induced neurons. Our findings revealed two major waves of global H3K9me3 reorganization during 4-day differentiation, namely on the first and third days, whereas nearly no changes occurred on the second and fourth days. The proposed method LiveMIEL (Live-cell Microscopic Imaging of Epigenetic Landscapes), which combines genetically encoded epigenetic probes and machine learning approaches, enables classification of multiparametric epigenetic signatures of single cells during stem cell differentiation and potentially in other biological models., (© 2024. The Author(s).)

The blood supply in the retina ensures photoreceptor function and maintains regular vision. Leber's hereditary optic neuropathy (LHON), caused by the mitochondrial DNA mutations that deteriorate complex I activity, is characterized by progressive vision loss. Although some reports indicated retinal vasculature abnormalities as one of the comorbidities in LHON, the paracrine influence of LHON-affected retinal ganglion cells (RGCs) on vascular endothelial cell physiology remains unclear. To address this, we established an in vitro model of mitochondrial complex I deficiency using induced pluripotent stem cell-derived RGCs (iPSC-RGCs) treated with a mitochondrial complex I inhibitor rotenone (Rot) to recapitulate LHON pathologies. The secretomes from Rot-treated iPSC-RGCs (Rot-iPSC-RGCs) were collected, and their treatment effect on human umbilical vein endothelial cells (HUVECs) was studied. Rot induced LHON-like characteristics in iPSC-RGCs, including decreased mitochondrial complex I activity and membrane potential, and increased mitochondrial reactive oxygen species (ROS) and apoptosis, leading to mitochondrial dysfunction. When HUVECs were exposed to conditioned media (CM) from Rot-iPSC-RGCs, the angiogenesis of HUVECs was suppressed compared to those treated with CM from control iPSC-RGCs (Ctrl-iPSC-RGCs). Angiogenesis-related proteins were altered in the secretomes from Rot-iPSC-RGC-derived CM, particularly angiopoietin, MMP-9, uPA, collagen XVIII, and VEGF were reduced. Notably, GeneMANIA analysis indicated that VEGFA emerged as the pivotal angiogenesis-related protein among the identified proteins secreted by health iPSC-RGCs but reduced in the secretomes from Rot-iPSC-RGCs. Quantitative real-time PCR and western blots confirmed the reduction of VEGFA at both transcription and translation levels, respectively. Our study reveals that Rot-iPSC-RGCs establish a microenvironment to diminish the angiogenic potential of vascular cells nearby, shedding light on the paracrine regulation of LHON-affected RGCs on retinal vasculature., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

The dysfunction of TAR DNA-binding protein 43 (TDP-43) is implicated in various neurodegenerative diseases, though the specific contributions of its toxic gain-of-function versus loss-of-function effects remain unclear. This study investigates the impact of TARDBP loss on cellular metabolism and viability using human-induced pluripotent stem cell-derived motor neurons and HeLa cells. TARDBP silencing led to reduced metabolic activity and cell growth, accompanied by neurite degeneration and decreased oxygen consumption rates in both cell types. Notably, TARDBP depletion induced a metabolic shift, impairing ATP production, increasing metabolic inflexibility, and elevating free radical production, indicating a critical role for TDP-43 in maintaining cellular bioenergetics. Furthermore, TARDBP loss triggered non-apoptotic cell death, increased ACSL4 expression, and reprogrammed lipid metabolism towards lipid droplet accumulation, while paradoxically enhancing resilience to ferroptosis inducers. Overall, our findings highlight those essential cellular traits such as ATP production, metabolic activity, oxygen consumption, and cell survival are highly dependent on TARDBP function., Competing Interests: Declaration of competing interest To whom it may be of interest. On behalf of all the authors, I hereby confirm that the authors of "Loss of TDP-43 Induces ALS-Related Collapse of Cellular Homeostasis" have not declared or known conflicts of interest. And to confirm that I sign the present in Lleida, at June 4th, 2024., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

We report the generation of a gene-edited human induced pluripotent stem cell (iPSC) line from an Alzheimer's disease patient-derived iPSC line harbouring the PSEN1 H163R mutation. This line demonstrates pluripotent stem cell morphology, expression of pluripotency markers, and maintains a normal karyotype., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

We investigated the impact of the human-specific gene CHRFAM7A on the function of α7 nicotinic acetylcholine receptors (α7 nAChRs) in two different types of neurons: human-induced pluripotent stem cell (hiPSC)-derived cortical neurons, and superior cervical ganglion (SCG) neurons, taken from transgenic mice expressing CHRFAM7A. dupα7, the gene product of CHRFAM7A, which lacks a major part of the extracellular N-terminal ligand-binding domain, co-assembles with α7, the gene product of CHRNA7. We assessed the receptor function in hiPSC-derived cortical and SCG neurons with Fura-2 calcium imaging and three different α7-specific ligands: PNU282987, choline, and 4BP-TQS. Given the short-lived open state of α7 receptors, we combined the two orthosteric agonists PNU282987 and choline with the type-2 positive allosteric modulator (PAM II) PNU120596. In line with different cellular models used previously, we demonstrate that CHRFAM7A has a major impact on nicotinic α7 nAChRs by reducing calcium transients in response to all three agonists., (© 2024 The Author(s). European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Chromosome 15q11.2-13.1 duplication (Dup15q) syndrome is one of the most common autism spectrum disorders (ASDs) associated with copy number variants (CNVs). For the analysis of CNV-relevant pathological cellular phenotypes, a CNV-corrected isogenic cell line is useful for excluding the influence of genetic background. Here, we devised a strategy to remove the isodicentric chromosome 15 by inserting a puro-ΔTK selection cassette into the extra chromosome using the CRISPR-Cas9 system, followed by a subsequent two-step drug selection. A series of assays, including qPCR-based copy number analysis and karyotype analysis, confirmed the elimination of the extra chromosome. Furthermore, cerebral organoids were generated from the parental Dup15q iPSCs and their isogenic iPSCs. scRNA-seq analysis revealed the alteration of expression levels in ion-channel-related genes and synapse-related genes in glutamatergic and GABAergic neurons in Dup15q organoids, respectively. The established isogenic cell line is a valuable resource for unraveling cellular and molecular alterations associated with Dup15q syndrome., Competing Interests: Declaration of Competing Interest This work was supported by Takeda Pharmaceutical Company Limited. H.Y. is an employee of Takeda Pharmaceutical Company Limited. No other potential conflicts of interest relevant to this article were reported., (Copyright © 2024. Published by Elsevier GmbH.)

Myelin basic protein (MBP) is a major component of the myelin sheaths of oligodendrocytes in the central nervous system and Schwann cells of the peripheral nervous system. Here we generated heterozygous fluorescent reporter of MBP gene in human induced pluripotent stem cells (hiPSCs). CRISPR/Cas9 genome editing technology was employed to knock in fused tdTomato fluorescent protein and EF1 alpha promoter-driven Bleomycin (Zeocin) resistance gene to the translational MBP C-terminal region. The resulting line, MBP-TEZ, showed tdTomato fluorescence upon oligodendrocyte differentiation. This reporter hiPSC line provides a precedential opportunity for monitoring human myelin formation and degeneration and purifying MBP-expressing cell lineages., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Apolipoprotein E (APOE)is the gene with greatest genetic risk for Alzheimer's disease (AD). We successfully established a human induced pluripotent stem cell(iPSC) line from a woman mutated by APOE gene. The cell line was isolated from this woman's peripheral blood mononuclear cells using a non-integrated Sendai virus, which retained the original genotype, showed a normal karyotype, highly expressed pluripotent markers and could differentiate into three germ layers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

A better understanding of the underlying pathomechanisms of diastolic dysfunction is crucial for the development of targeted therapeutic options with the aim to increase the patients' quality of life. In order to shed light on the processes involved, suitable models are required. Here, effects of endothelin-1 (ET-1) treatment on cardiomyocytes derived from human induced pluripotent stem cells (hiPSCs) were investigated. While it is well established, that ET-1 treatment induces hypertrophy in cardiomyocytes, resulting changes in cell mechanics and contractile behavior with focus on relaxation have not been examined before. Cardiomyocytes were treated with 10 nM of ET-1 for 24 h and 48 h, respectively. Hypertrophy was confirmed by real-time deformability cytometry (RT-DC) which was also used to assess the mechanical properties of cardiomyocytes. For investigation of the contractile behavior, 24 h phase contrast video microscopy was applied. To get a deeper insight into changes on the molecular biological level, gene expression analysis was performed using the NanoString nCounter® cardiovascular disease panel. Besides an increased cell size, ET-1 treated cardiomyocytes are stiffer and show an impaired relaxation. Gene expression patterns in ET-1 treated hiPSC derived cardiomyocytes showed that pathways associated with cardiovascular diseases, cardiac hypertrophy and extracellular matrix were upregulated while those associated with fatty acid metabolism were downregulated. We conclude that alterations in cardiomyocytes after ET-1 treatment go far beyond hypertrophy and represent a useful model for diastolic dysfunction., Competing Interests: Declaration of competing interest Oliver Otto is co-founder of Zellmechanik Dresden commercializing real-time deformability cytometry. All other authors declare that they have no conflict of interest. The authors did not use generative AI or AI-assisted technologies in the development of this manuscript., (Copyright © 2024. Published by Elsevier Ltd.)

Usher syndrome (USH) is the most common cause of inherited deaf-blindness. Here, we produced the LEIi020-A and LEIi020-B induced pluripotent stem cell (iPSC) lines from dermal fibroblasts derived from a patient with USH1B caused by inheritance of homozygous c.496del variants in MYO7A using episomal plasmids encoding OCT4, SOX2, KLF4, L-MYC, LIN28, mir302/367 microRNA and shRNA for TP53. Both iPSC lines expressed pluripotency markers, demonstrated trilineage differentiation potential and displayed a 46,XY karyotype. These cell lines represent a valuable resource for the production of retinal and otic tissues to support research into the pathogenesis and treatment of USH1B., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The transcription factor WT1 plays a critical role in several embryonic developmental processes such as gonadogenesis, nephrogenesis, and cardiac development. We generated a homozygous (MCRIi031-A-3) WT1 knockout induced pluripotent stem cell (iPSC) line from human fibroblasts using a one-step protocol for CRISPR/Cas9 gene-editing and episomal-based reprogramming. The cells exhibit a normal karyotype and morphology, express pluripotency markers, and have the capacity to differentiate into the three embryonic germ layers. These cell lines will allow us to further explore the role of WT1 in critical developmental processes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Disease-related cells differentiated from patient-derived iPSCs are useful for elucidating the pathophysiological mechanisms underlying these diseases. In this study, four iPSC lines were established from independent patients with sensorineural hearing loss and a mutation in EYA4. These iPSCs showed pluripotency, the capacity to differentiate into three germ layers, and normal karyotypes, suggesting that these lines are useful for the pathological study of sensorineural hearing loss and drug screening for ear disorders., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Epilepsy is a chronic neurological disease. Here we describe the generation of induced pluripotent stem cells (iPSCs) from a patient diagnosed as epilepsy caused by ATP1A2 gene mutation. Induced pluripotent stem cells (iPSCs) were developed using non-integrating episomal vectors containing OCT4, SOX2, KLF4, BCL-XL and C-MYC. The established iPSC line (SDCHi007-A) displayed pluripotent cell morphology, high expression levels of pluripotency markers, differentiation potential in vitro, normal karyotype, and remaining the original ATP1A2 gene mutation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Age-related macular degeneration (AMD) is characterized by visual impairment observed in elderly population. Two forms of the disease are generally described, the atrophic (AMDa) and exudative forms (AMDe). Up until now, no curative treatment is available for this disease. The retinal pigment epithelium (RPE) plays a central role in the pathogenesis of age-related macular degeneration. Here, involvement of RPE dysfunction in AMD onset and progression was analyzed by a comparison of transcriptome profiles of hiPSC-RPE derived from healthy individuals or individuals affected by AMDa or AMDe. The analysis highlighted almost 1000 genes differentially expressed between the three comparison groups. Among these genes, 33 genes were already known to be involved in AMD pathogenesis. To establish an AMD genetic signature, we focused on genes differentially expressed in both AMDa/e cell lines compared to control cells and focused on the three genes (ABCA1, RPN2, RB1CC1) that were related to lipidic homeostasis. Differences in level expression of these three genes are found not only in control and AMDa/e cell lines, but also between AMDa and AMDe populations., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Patients with JAK2V617F-positive myeloproliferative neoplasms (MPNs) and clonal hematopoiesis of indeterminate potential face a significantly elevated risk of cardiovascular diseases. Endothelial cells carrying the JAK2V617F mutation have been detected in many patients with MPN. In this study, we investigated the molecular basis for the high incidence of cardiovascular complications in patients with MPN., Methods: We investigated the impact of endothelial JAK2V617F mutation on cardiovascular disease development using both transgenic murine models and MPN patient-derived induced pluripotent stem cell lines., Results: Our investigations revealed that JAK2V617F mutant endothelial cells promote cardiovascular diseases under stress, which is associated with endothelial-to-mesenchymal transition and endothelial dysfunction. Importantly, we discovered that inhibiting the endothelial TPO (thrombopoietin) receptor MPL (myeloproliferative leukemia virus oncogene) suppressed JAK2V617F-induced endothelial-to-mesenchymal transition and prevented cardiovascular dysfunction caused by mutant endothelial cells. Notably, the endothelial MPL receptor is not essential for the normal physiological regulation of blood cell counts and cardiac function., Conclusions: JAK2V617F mutant endothelial cells play a critical role in the development of cardiovascular diseases in JAK2V617F-positive MPNs, and endothelial MPL could be a promising therapeutic target for preventing or ameliorating cardiovascular complications in these patients., Competing Interests: None.

CTNNB1 encodes beta-catenin, which plays a crucial role in Wnt signaling pathway. Mutations in CTNNB1 involve in tumor developing, Primary Aldosteronism, Neurodevelopmental disorders (NDDs), etc. NDDs is a class of disorders that impact brain development and function, manifesting symptom including autism spectrum disorder (ASD), intellectual disability (ID), schizophrenia (SCZ), and epilepsy. Here, we generated an iPSC line (CTUi005-A) from a patient diagnosed with NDDs, carrying a heterozygous mutation of the CTNNB1 gene. CTUi005-A exhibits typical iPSC characteristics, and holds potential as a cellular tool for investigating the pathogenic mechanisms underlying NDDs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Pluripotent stem cells were generated through the electroporation of episomal plasmids, containing crucial reprogramming factors, into skin fibroblasts extracted from a female Alzheimer's patient harboring the PSEN1 709 T > C (p.Phe237Leu) heterozygous mutation. The pluripotent stem cells exhibit a normal karyotype and express pivotal stem cell markers including TRA-1-60, Nanog, SOX2, and OCT4. Furthermore, their capacity to differentiate into the three germ layers in in vivo teratoma experiments has been substantiated. The pluripotent stem cell line can serve as a cellular model for Alzheimer's disease, offering significant value in elucidating the pathogenesis and therapeutic strategies of the disease., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Jiansheng Kang reports financial support was provided by National Natural Science Foundation of China. None reports a relationship with none that includes:. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Vascular Ehlers-Danlos Syndrome (vEDS) is an inherited connective tissue disorder caused by COL3A1 gene, mutations that encodes type III collagen, a crucial component of blood vessels. vEDS can be life-threatening as these patients can have severe internal bleeding due to arterial rupture. Here, we generated induced pluripotent stem cell (iPSC) lines from two vEDS patients carrying a missense mutation in the COL3A1 (c.226A > G, p.Asn76Asp) gene. These lines exhibited typical iPSC characteristics including morphology, expression of pluripotency markers, and could differentiate to all three germ layer. These iPSC lines can serve as valuable tools for elucidating the pathophysiology underlying vEDS., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Marfan Syndrome, a connective tissue disorder caused by Fibrillin-1 (FBN1) gene mutations, induces disease in the ocular, musculoskeletal, and cardiovascular systems and increases aortic vulnerability to rupture associated with high mortality rates. We describe an induced pluripotent stem cell line (HFD1) generated from patient-derived human dermal fibroblasts harboring a heterozygous c.3338-2A>C intronic splice acceptor site variant preceding Exon 28 of FBN1. The clonal line, which produces abnormal FBN1 splice variants, has a normal karyotype, expresses appropriate stemness markers, and maintains trilineage differentiation potential. This line represents a valuable resource for studying how abnormal splicing variants contribute to Marfan Syndrome., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

ALDH7A1 encodes for the enzyme catalyzing the third step of the lysine degradation pathway. Biallelic pathogenic variants in ALDH7A1 are associated with pyridoxine dependent epilepsy (PDE), of which the c.1279G>C (p.Glu427Gln) variant is the most commonly reported variant and is carried by 30% of PDE patients with European ancestry. In this study, hiPSC lines derived from four PDE patients carrying the c.1279G>C variant in homozygosis in ALDH7A1 were generated and fully characterized. These hiPSC lines can contribute to better understand the molecular mechanism of disease underlying PDE as well as serving as a model system to evaluate new therapeutic strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Friedreich's ataxia (FRDA) is a rare neurodegenerative disease caused by an expansion of a GAA repeat sequence within the Frataxin (FXN) gene. Prominent regions of neurodegeneration include sensory neurons within the dorsal root ganglia. Here we present a set of genetically modified FRDA induced pluripotent stem cell (iPSC) lines that carry an inducible neurogenin-2 (NGN2) expression cassette. Exogenous expression of NGN2 in iPSC derived neural crest progenitors efficiently generates functionally mature sensory neurons. These cell lines will provide a streamlined source of FRDA iPSC sensory neurons for studying both disease mechanism and screening potential therapeutics., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: M Dottori collaborates with Dmitry A. Ovchinnikov, who is an editor for Stem Cell Research. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

GCDH encodes for the enzyme catalyzing the sixth step of the lysine degradation pathway. Autosomal recessive variants in GCDH are associated with glutaric aciduria type I (GA1), of which a wide genotypic spectrum of pathogenic variants have been described. In this study, hiPSC lines derived from four GA1 patients with different genotypes were generated and fully characterized. Two patients carry compound heterozygous variants in GCDH, while the other two patients carry a variant in homozygosis. These hiPSC lines can significantly contribute to better understand the molecular mechanism underlying GA1 and provide excellent models for the development of new therapeutic strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

The transcription factor SOX9 plays a critical role in several embryonic developmental processes such as gonadogenesis, chrondrogenesis, and cardiac development. We generated heterozygous (MCRIi031-A-1) and homozygous (MCRIi031-A-2) SOX9 knockout induced pluripotent stem cell (iPSC) lines from human fibroblasts using a one-step protocol for CRISPR/Cas9 gene-editing and episomal-based reprogramming. Both iPSC lines exhibit a normal karyotype and morphology, express pluripotency markers, and have the capacity to differentiate into the three embryonic germ layers. These cell lines will allow us to further explore the role of SOX9 in critical developmental processes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background Aims: Substrate elasticity may direct cell-fate decisions of stem cells. However, it is largely unclear how matrix stiffness affects the differentiation of induced pluripotent stem cells (iPSCs) and whether this is also reflected by epigenetic modifications., Methods: We cultured iPSCs on tissue culture plastic (TCP) and polydimethylsiloxane (PDMS) with different Young's modulus (0.2 kPa, 16 kPa or 64 kPa) to investigate the sequel on growth and differentiation toward endoderm, mesoderm and ectoderm., Results: Immunofluorescence and gene expression of canonical differentiation markers were hardly affected by the substrates. Notably, when we analyzed DNA methylation profiles of undifferentiated iPSCs or after three-lineage differentiation, we did not see any significant differences on the three different PDMS elasticities. Only when we compared DNA methylation profiles on PDMS-substrates versus TCP we did observe epigenetic differences, particularly on mesodermal differentiation., Conclusions: Stiffness of PDMS substrates did not affect directed differentiation of iPSCs, whereas the moderate epigenetic differences on TCP might also be attributed to other chemical parameters., Competing Interests: Declaration of competing interest WW is a founder of Cygenia GmbH, which provides services for various epigenetic signatures (www.cygenia.com). The medical faculty of RWTH-Aachen has claimed patent application for GermLayerTracker and MEM, KZ and WW are co-applicants for the patent., (Copyright © 2024 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.)

The X-linked lysosomal storage disorder Fabry disease originates from GLA gene mutations causing α-galactosidase A enzyme deficiency. Here we generated the GLA knockout hiPSC line MHHi001-A-15 (GLA-KOhiPSC) as an in vitro Fabry disease model by targeting exon 2 of the GLA gene by CRISPR/Cas9 in the established control hiPSC line MHHi001-A. GLA-KOhiPSCs retained the expression of pluripotency markers, trilineage differentiation potential, as well as normal karyotype and stem cell morphology but lacked α-galactosidase A enzyme activity. The GLA-KOhiPSCs represent a potent resource to not only study the Fabry disease manifestation but also screen for novel treatment options., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Hexanucleotide repeat expansions in the C9orf72 gene are the primary genetic cause for both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two related neurodegenerative diseases. Significant advances in the elucidation of the disease mechanisms responsible for C9orf72 ALS-FTD have revealed both a toxic gain-of-function and a loss-of-function mechanism as possible underlying disease cause. As the differential contribution of both gain and loss of function in C9orf72 ALS-FTD pathogenesis remains debated, we investigated disease mechanisms in motor neurons derived from both authentic human patient C9orf72 ALS-FTD iPSCs as well as a C9orf72 knockout iPSC line. We found that patient neurons presented with less motile and enlarged lysosomes, a decrease in autophagic flux and an increase in SQSTM1/p62 puncta and insoluble TARDBP/TDP-43 species. Importantly, we found that C9orf72 knockout barely has any influence on these phenotypes and mainly results in impaired endosomal maturation. Together, our data suggest that toxic gain-of-function, rather than loss-of-function, mechanisms in C9orf72 ALS-FTD impair the autophagy-lysosome system in neurons.

Calcium indicators are sensitive tools to image neural activity. However, their use in human induced pluripotent stem cells (iPSC)-derived neurons is limited by silencing of the transgene. We generated the iPSC line MSE2336A carrying heterozygous insertion in the safe-harbor locus AAVS1 of the ultrasensitive protein calcium sensor (GCaMP6) under the control of CAG promoter and UCOE to maintain robust transgene expression in differentiated cells. The iPSC exhibited normal cell morphology, expression of pluripotency markers, genome integrity, and the ability to differentiate into the three primary germ layers. This line provides a powerful model to study activity in human neurons., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Samuele Marro is an active editor for Stem Cell Research Lab Resources. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Cadherin 23 (CDH23) is one of the most common genes responsible for hereditary hearing loss; a mutation of CDH23 can cause a wide range of symptoms depending on the variant. In this study, an iPSC line was generated from a patient with late-onset, progressive high frequency hearing loss caused by c.[719C > T];[6085C > T]:p.[P240L];[R2029W] compound heterozygous variants of CDH23. The cells were confirmed to have a normal karyotype, express markers of pluripotency, and have tri-embryonic differentiation potential. This disease-specific iPSC line will further the construction of disease models and the elucidation of the pathophysiology of CDH23 mutations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Here we present the generation of HIMRi006-A and HIMRi007-A Pompe disease (PD) patient derived human induced pluripotent stem cell (hiPSC) lines. HIMRi006-A represents an infantile onset disease (IOPD) phenotype caused by a homozygous c.307 T > G mutation in the GAA gene. HIMRi007-A is characterized by heterozygous mutations c.-32-13 T > G/c.1716C > G and is associated with an adult onset of disease symptoms (LOPD). Both lines are generated via lentiviral expression of OCT4, SOX2, KLF4, and c-MYC. The lines display a typical embryonic stem cell morphology, express pluripotency markers, retain a normal karyotype (46, XX/XY) and have the differentiation capacity in all three germ layers. Altogether, both lines provide a resource tool to the community for future in depth molecular studies of PD pathomechanism., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

A basic assumption underlying induced pluripotent stem cell (iPSC) models of neurodegeneration is that disease-relevant pathologies present in brain tissue are also represented in donor-matched cells differentiated from iPSCs. However, few studies have tested this hypothesis in matched iPSCs and neuropathologically characterized donated brain tissues. To address this, we assessed iPSC-neuron production of β-amyloid (Aβ) Aβ40, Aβ42, and Aβ43 in 24 iPSC lines matched to donor brains with primary neuropathologic diagnoses of sporadic AD (sAD), familial AD (fAD), control, and other neurodegenerative disorders. Our results demonstrate a positive correlation between Aβ43 production by fAD iPSC-neurons and Aβ43 accumulation in matched brain tissues but do not reveal a substantial correlation in soluble Aβ species between control or sAD iPSC-neurons and matched brains. However, we found that the ApoE4 genotype is associated with increased Aβ production by AD iPSC-neurons. Pathologic tau phosphorylation was found to be increased in AD and fAD iPSC-neurons compared to controls and positively correlated with the relative abundance of longer-length Aβ species produced by these cells. Taken together, our results demonstrate that sAD-predisposing genetic factors influence iPSC-neuron phenotypes and that these cells are capturing disease-relevant and patient-specific components of the amyloid cascade., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Association of Neuropathologists, Inc.)

Peripheral blood mononuclear cells (PBMCs) were obtained from a patient diagnosed with Familial Hemiplegic Migraine Type 3, who carried a heterozygous A > C mutation in the SCN1A gene and reprogrammed using CytoTune TM -iPS 2.0 Sendai Reprogramming Kit. The iPSC line maintained the mutation while expressing markers of pluripotency. Additionally, it exhibited a normal karyotype and demonstrated potential for in vitro differentiation into cells representing all three embryonic germ layers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Hereditary spastic paraplegias (HSPs) a group of rare, clinically, and genetically heterogeneous disorders characterized by progressive degeneration of the corticospinal tract. Among these HSPs, SPG31 is due to autosomal dominant mutations in the receptor expression-enhancing protein 1 (REEP1) gene. Over 80 genes have been associated with HSPs, and the list is constantly growing as research progresses. This study is aimed to create a patient-derived human induced pluripotent stem cell (hiPSC) line with a specific nonsense mutation to better characterize the etiopathogenesis of the disease., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Autism spectrum disorder (ASD) is a complex developmental disorder characterized by challenges with social interactions and restricted/repetitive behaviors. Here, we recruited nine Qatari children of Arab ethnicity (males, aged 2-4 years), including six ASD subjects (n = 3 mild-to-moderate ASD and n = 3 severe ASD) and three control subjects. We generated induced pluripotent stem cell (iPSC) lines from PBMC samples of these subjects using non-integrating Sendai viral vectors. These iPSC lines were fully characterized and exhibited pluripotency characteristics, normal karyotypes, and trilineage differentiation potential. These iPSC lines provide valuable cell models for understanding ASD pathophysiology and developing new therapeutics for ASD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Oxaliplatin, a platinum-based chemotherapeutic agent, frequently causes acute and chronic peripheral sensory neuropathy, for which no effective treatment has been established. In particular, chronic neuropathy can persist for years even after treatment completion, thus worsening patients' quality of life. To avoid the development of intractable adverse effects, a predictive biomarker early in treatment is awaited. In this study, we explored extracellular long non-coding RNAs (lncRNAs) released from primary sensory neurons as biomarker candidates for oxaliplatin-induced peripheral neuropathy. Because many human-specific lncRNA genes exist, we induced peripheral sensory neurons from human induced pluripotent stem cells. Oxaliplatin treatment changed the levels of many lncRNAs in extracellular vesicles (EVs) released from cultured primary sensory neurons. Among them, the levels of release of lncRNAs that were considered to be selectively expressed in dorsal root ganglia were correlated with those of lncRNAs in plasma EV obtained from healthy individuals. Several lncRNAs in plasma EVs early after the initiation of treatment showed greater changes in patients who did not develop chronic neuropathy that persisted for more than 1 year than in those who did. Therefore, these extracellular lncRNAs in plasma EVs may represent predictive biomarkers for the development of chronic peripheral neuropathy induced by oxaliplatin., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Ataxia Telangiectasia (AT) is a rare disorder caused by mutations in the ATM gene and results in progressive neurodegeneration for reasons that remain poorly understood. In addition to its central role in nuclear DNA repair, ATM operates outside the nucleus to regulate metabolism, redox homeostasis and mitochondrial function. However, a systematic investigation into how and when loss of ATM affects these parameters in relevant human neuronal models of AT was lacking. We therefore used cortical neurons and brain organoids from AT-patient iPSC and gene corrected isogenic controls to reveal levels of mitochondrial dysfunction, oxidative stress, and senescence that vary with developmental maturity. Transcriptome analyses identified disruptions in regulatory networks related to mitochondrial function and maintenance, including alterations in the PARP/SIRT signalling axis and dysregulation of key mitophagy and mitochondrial fission-fusion processes. We further show that antioxidants reduce ROS and restore neurite branching in AT neuronal cultures, and ameliorate impaired neuronal activity in AT brain organoids. We conclude that progressive mitochondrial dysfunction and aberrant ROS production are important contributors to neurodegeneration in AT and are strongly linked to ATM's role in mitochondrial homeostasis regulation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Proarrhythmic cardiotoxicity remains a substantial barrier to drug development as well as a major global health challenge. In vitro human pluripotent stem cell-based new approach methodologies have been increasingly proposed and employed as alternatives to existing in vitro and in vivo models that do not accurately recapitulate human cardiac electrophysiology or cardiotoxicity risk. In this study, we expanded the capacity of our previously established 3D human cardiac microtissue model to perform quantitative risk assessment by combining it with a physiologically based pharmacokinetic model, allowing a direct comparison of potentially harmful concentrations predicted in vitro to in vivo therapeutic levels. This approach enabled the measurement of concentration responses and margins of exposure for 2 physiologically relevant metrics of proarrhythmic risk (i.e. action potential duration and triangulation assessed by optical mapping) across concentrations spanning 3 orders of magnitude. The combination of both metrics enabled accurate proarrhythmic risk assessment of 4 compounds with a range of known proarrhythmic risk profiles (i.e. quinidine, cisapride, ranolazine, and verapamil) and demonstrated close agreement with their known clinical effects. Action potential triangulation was found to be a more sensitive metric for predicting proarrhythmic risk associated with the primary mechanism of concern for pharmaceutical-induced fatal ventricular arrhythmias, delayed cardiac repolarization due to inhibition of the rapid delayed rectifier potassium channel, or hERG channel. This study advances human-induced pluripotent stem cell-based 3D cardiac tissue models as new approach methodologies that enable in vitro proarrhythmic risk assessment with high precision of quantitative metrics for understanding clinically relevant cardiotoxicity., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Diamond-Blackfan anemia syndrome (DBAS) is an inherited bone marrow failure disorder that typically presents in infancy as hypoplastic anemia and developmental abnormalities in approximately 50% of cases. DBAS is caused by haploinsufficiency in one of 24 ribosomal protein genes, with RPS19 mutations accounting for 25% of cases. We generated iPSC lines from two patients with different heterozygous RPS19 mutations (c.191T > C and c.184C > T) and isogenic lines in which the mutations were corrected by Cas9-mediated homology directed repair., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Mitchell Weiss reports a relationship with Fulcrum Therapeutics, Inc. that includes: consulting or advisory. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Dietary supplements containing usnic acid have been increasingly marketed for weight loss over the past decades, even though incidences of severe hepatotoxicity and acute liver failure due to their overuse have been reported. To date, the toxic mechanism of usnic acid-induced liver injury at the molecular level still remains to be fully elucidated. Here, we conducted a transcriptomic study on usnic acid using a novel in vitro hepatotoxicity model employing human induced pluripotent stem cell (iPSC)-derived hepatocytes. Treatment with 20 μM usnic acid for 24 h caused 4272 differentially expressed genes (DEGs) in the cells. Ingenuity Pathway Analysis (IPA) based on the DEGs and gene set enrichment analysis (GSEA) using the whole transcriptome expression data concordantly revealed several signaling pathways and biological processes that, when taken together, suggest that usnic acid caused oxidative stress and DNA damage in the cells, which further led to cell cycle arrest and eventually resulted in cell death through apoptosis. These transcriptomic findings were subsequently corroborated by a variety of cellular assays, including reactive oxygen species (ROS) generation and glutathione (GSH) depletion, DNA damage (pH2AX detection and 8-hydroxy-2'-deoxyguanosine [8-OH-dg] assay), cell cycle analysis, and caspase 3/7 activity. Collectively, the results of the current study accord with previous in vivo and in vitro findings, provide further evidence that oxidative stress-caused DNA damage contributes to usnic acid-induced hepatotoxicity, shed new light on molecular mechanisms of usnic acid-induced hepatotoxicity, and demonstrate the usefulness of iPSC-derived hepatocytes as an in vitro model for hepatotoxicity testing and prediction., (© 2024 The Authors. Journal of Applied Toxicology published by John Wiley & Sons Ltd.)

Ten-Eleven Translocation methylcytosine dioxygenase 1 (TET1) is known to play a broad tumor suppressor role through demethylating and activating tumor suppressor genes. TET1 missense mutations are previously reported in many types of leukemia. Here, the human induced pluripotent stem cell line MURAi001-A was generated from skin fibroblasts derived from a 56-year-old female patient carrying the TET1 gene mutation c.4404A > G (p.I1468M), who had a history of ovarian germ cell tumor. The MURAi001-A cell line demonstrated embryonic-like characteristics as it expressed specific stemness markers, differentiated into the three germ layers, and retained normal karyotyping., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Cardiovascular disease (CVD) is the leading cause of death worldwide. To this end, human cardiac organoids (hCOs) have been developed for improved organotypic CVD modeling over conventional in vivo animal models. Utilizing human cells, hCOs hold great promise to bridge key gaps in CVD research pertaining to human-specific conditions. hCOs are multicellular 3D models which resemble heart structure and function. Varying hCOs fabrication techniques leads to functional and phenotypic differences. To investigate heterogeneity across hCO platforms, we performed a transcriptomic analysis utilizing bulk RNA-sequencing from four previously published unique hCO studies. We further compared selected hCOs to 2D and 3D hiPSC-derived cardiomyocytes (hiPSC-CMs), as well as fetal and adult human myocardium bulk RNA-sequencing samples. Upon investigation utilizing Principal Component Analysis, K-means clustering analysis of key genes, and further downstream analyses such as Gene Set Enrichment (GSEA), Gene Set Variation (GSVA), and GO term enrichment, we found that hCO fabrication method influences maturity and cellular heterogeneity across models. Thus, we propose that adjustment of fabrication method will result in an hCO with a defined maturity and transcriptomic profile to facilitate its specified applications, in turn maximizing its modeling potential., (© 2024. The Author(s).)

The growth factor Neuregulin-1 (NRG1) has pleiotropic roles in proliferation and differentiation of the stem cell niche in different tissues. It has been implicated in gut, brain and muscle development and repair. Six isoform classes of NRG1 and over 28 protein isoforms have been previously described. Here we report a new class of NRG1, designated NRG1-VII to denote that these NRG1 isoforms arise from a myeloid-specific transcriptional start site (TSS) previously uncharacterized. Long-read sequencing was used to identify eight high-confidence NRG1-VII transcripts. These transcripts presented major structural differences from one another, through the use of cassette exons and alternative stop codons. Expression of NRG1-VII was confirmed in primary human monocytes and tissue resident macrophages and induced pluripotent stem cell-derived macrophages (iPSC-derived macrophages). Isoform switching via cassette exon usage and alternate polyadenylation was apparent during monocyte maturation and macrophage differentiation. NRG1-VII is the major class expressed by the myeloid lineage, including tissue-resident macrophages. Analysis of public gene expression data indicates that monocytes and macrophages are a primary source of NRG1. The size and structure of class VII isoforms suggests that they may be more diffusible through tissues than other NRG1 classes. However, the specific roles of class VII variants in tissue homeostasis and repair have not yet been determined., (© 2024. The Author(s).)

Intermediate-length repeat expansions in ATAXIN-2 (ATXN2) are the strongest genetic risk factor for amyotrophic lateral sclerosis (ALS). At the molecular level, ATXN2 intermediate expansions enhance TDP-43 toxicity and pathology. However, whether this triggers ALS pathogenesis at the cellular and functional level remains unknown. Here, we combine patient-derived and mouse models to dissect the effects of ATXN2 intermediate expansions in an ALS background. iPSC-derived motor neurons from ATXN2-ALS patients show altered stress granules, neurite damage and abnormal electrophysiological properties compared to healthy control and other familial ALS mutations. In TDP-43 Tg -ALS mice, ATXN2-Q33 causes reduced motor function, NMJ alterations, neuron degeneration and altered in vitro stress granule dynamics. Furthermore, gene expression changes related to mitochondrial function and inflammatory response are detected and confirmed at the cellular level in mice and human neuron and organoid models. Together, these results define pathogenic defects underlying ATXN2-ALS and provide a framework for future research into ATXN2-dependent pathogenesis and therapy., (© 2024. The Author(s).)

In fetal development, tissue interaction such as the interplay between blood vessel (BV) and epithelial tissue is crucial for organogenesis. Here we recapitulate the spatial arrangement between liver epithelial tissue and the portal vein to observe the formation of intrahepatic bile ducts (BDs) from human induced pluripotent stem cells (hiPSC). We co-culture hiPSC-liver progenitors on the artificial BV consisting of immature smooth muscle cells and endothelial cells, both derived from hiPSCs. After 3 weeks, liver progenitors within hiPSC-BV-incorporated liver organoids (BVLO) differentiate to cholangiocytes and acquire epithelial characteristics, including intercellular junctions, microvilli on the apical membrane, and secretory functions. Furthermore, liver surface transplanted-BVLO temporarily attenuates cholestatic injury symptoms. Single cell RNA sequence analysis suggests that BD interact with the BV in BVLO through TGFβ and Notch pathways. Knocking out JAG1 in hiPSC-BV significantly attenuates bile duct formation, highlighting BVLO potential as a model for Alagille syndrome, a congenital biliary disease. Overall, we develop a novel 3D co-culture method that successfully establishes functional human BDs by emulating liver epithelial-BV interaction., (© 2024. The Author(s).)