Background The COVID-19 pandemic created a unique situation in which researchers repurposed human models of varying complexity to investigate SARS-CoV-2, providing the opportunity to analyse their contribution across organs. Methods We conducted a systematic review of 558 studies, divided into pandemic and post-pandemic periods, to assess how human models were applied to investigate host factors, viral replication of SARS-CoV-2, immune responses, and to evaluate reporting quality. Findings Our analysis revealed substantial limitations that were only partially alleviated in post-pandemic studies. These limitations included heterogeneity of outcome measures, incomplete documentation of model characteristics and experimental procedures, and variable reporting quality, which were associated with reduced cross-study comparability, more difficult risk-of-bias assessment, and limited reliability and interpretability of evidence synthesis and meta-analytical results. Interpretation Strengthening community-driven reporting standards and methodological transparency is essential to enable robust evidence synthesis and to fully realise the potential of human organ models in future pandemics and translational research. Funding The study was supported by Einstein Foundation Berlin; Federal Ministry of Research, Technology and Space; European Union; Else Kröner-Fresenius-Stiftung; Volkswagen Foundation; Charité 3R; Foundation Charité.

The microtubule-stabilizing drug paclitaxel remains the standard of care for various solid malignancies but frequently leads to chemotherapy-induced peripheral neuropathy (CIPN). CIPN is a leading cause for premature treatment termination and a significantly reduced quality of life in long-term cancer survivors. The molecular mechanisms of neuro-axonal degeneration, neuroinflammation, and pain in patients treated with paclitaxel remain incompletely understood, and there are currently no predictive biomarkers or preventive treatments. We used human iPSC-derived sensory neurons exposed to paclitaxel to comprehensively model the pathophysiology of CIPN. Neurotoxicity was assessed over time using viability assays and sequential RNA sequencing, as well as deep proteome and lipidomic analyses. We observed a time and dose-dependent decline of cell viability at clinically relevant paclitaxel doses. Sequential RNA sequencing defined JUN as an early immediate gene, followed by the overexpression of genes of the neuronal stress response (e.g., ARID5A, WEE1, DUSP16, GADD45A), neuronal injury and apoptotic pathways (e.g., ATF3, HRK, BBC3 [PUMA], BCL2L11 [BIM], CASP3), neuroinflammation and nociception (CALCB, MMP10, IL31RA, CYSLTR2, C3AR1, TNFRSF12A) and neuronal transduction (e.g., CAMK2A, STOML3, PIRT), while key enzymes of lipid biosynthesis were markedly downregulated (e.g., LSS, HMGCS1, HMGCR, DHCR24). Deep proteome analyses following 48 h of exposure to 100 nM paclitaxel revealed a strong correlation of differentially expressed RNA with proteins, and a marked degradation of essential axonal transport proteins such as kinesins, stathmins, and scaffold proteins. Consistent with the downregulation of rate-limiting enzymes of lipid biosynthesis, lipidome analysis confirmed deregulation of neuronal lipid homeostasis. In summary, paclitaxel induces transcriptomic and proteomic signatures of the neuronal stress response, neuroinflammation, nociception, and disturbed metabolism. These may explain, in part, the clinical phenotype of sensory loss, hypersensitivity, and neuropathic pain frequently observed in patients suffering from CIPN, but constitute pharmacologically addressable targets.

Lung cancer, the leading cause of cancer-related mortality, presents major challenges for both standard therapies and chimeric antigen receptor (CAR) T cell therapy due to tumour heterogeneity and resistance. Preclinical models that capture patient-specific factors are essential for personalizing treatment decisions. Here we show that matched lung tumouroids and healthy lung organoids derived from patients provide a robust platform for studying therapy responses. The tumouroids faithfully retained the molecular and histological identity of the original tumours, as confirmed by genomic, epigenomic and proteomic analyses, and accurately replicated individual patient responses to standard-of-care therapies. Importantly, the platform also revealed patient-specific CAR T cell responses, uncovering a complex interplay between target antigen density and broader, tumour-intrinsic resistance programmes. By capturing these individualized factors, our model supports rational patient selection for CAR T cell therapy in lung cancer and provides a framework for designing CAR T cells tailored to overcome resistance mechanisms in solid tumours. A platform using matched patient-derived lung tumouroids and healthy lung organoids enables accurate examination of patient responses to CAR T therapy and offers a faithful framework for improved CAR T design.

Although human pluripotent stem cells (hPSCs) can generate all tissues of the body, hPSCs in vitro frequently exhibit differentiation biases or failure that pose substantial challenges for disease modeling and regenerative medicine. The origins of these biases remain incompletely understood and extend beyond reprogramming artifacts. Here we show that loss of default neural differentiation capacity and failure to form brain organoids are linked to erosion of bivalent chromatin marks at developmental gene loci, independent of DNA methylation, driving acquisition of a posterior epiblast-like state and premature developmental gene expression. We develop a chemical chromatin restoration (CHR) approach that rescues this differentiation bias by reinstating transcriptional programs and chromatin landscapes characteristic of the competent anterior epiblast-like state, restoring broad differentiation potential. These findings establish locus-specific patterns of repressive and activating histone post-translational modifications as a tractable and experimentally targetable determinant of hPSC fate competency, and offer an effective route to rescue differentiation-compromised hPSC lines for applications in disease modeling and regenerative medicine.

We report on the generation of human induced pluripotent stem cell (iPSC) lines, BIHi005-A-86 and BIHi005-A-87, carrying the KIT D816V mutation associated with Indolent Systemic Mastocytosis (ISM). To overcome the confounding genetic backgrounds of existing leukemic models, we introduced this gain-of-function mutation into the healthy BIHi005-A line using CRISPR/Cas9 editing. The resulting clones were validated via whole-genome sequencing (WGS) to confirm specific on-target editing and lack of predicted or disease-relevant off-target effects, while maintaining genomic stability. Together with the parental line, this resource provides an isogenic controlled platform for investigating KIT D816V-driven pathogenesis

Thyroid hormones (THs) are essential regulators of human brain development, and disrupted TH availability during pregnancy or early life is linked to adverse neurodevelopmental outcomes. Concerns that environmental chemicals interfere with TH signalling have increased the need for human-relevant in vitro systems to identify thyroid hormone system-disrupting chemicals (THSDCs) for risk assessment. Here, we compared two human-induced pluripotent stem cell (hiPSC)-derived brain organoid models for THSDC assessment: (i) human cortical organoids (COs) generated by unguided differentiation, offering higher architectural complexity but lower throughput; and (ii) neural stem cell-derived organoids (NSCOs), designed for scalability with reduced cellular diversity. Both models expressed key TH handling components, including the transporter SLC16A2 (MCT8) and the inactivating enzyme DIO3. Using LC–MS/MS, we show that exogenous T3 is depleted from culture media and metabolized to 3,3′-T2 and 3′-T1 in both models, alongside upregulation of T3-responsive genes (HR, KLF9, DIO3, SEMA3C). Pulse and chronic co-exposures to reference disruptors iopanoic acid (IA, deiodinase inhibitor) and silychristin (SC, MCT8 inhibitor) altered T3 metabolism and modulated T3-responsive transcriptional endpoints. In NSCOs, high-content imaging revealed treatment-associated changes in cell composition, with chronic T3 reducing the SOX2-positive progenitor pool and THSDCs blocking this effect. Together, these findings provide a framework for organoid qualification—linking TH handling, transcriptomic responsiveness, and scalable phenotypic readouts—as a necessary step toward model validation and implementation of brain organoids in THSDC risk assessment pipelines.

BACKGROUND:Dysregulation of microRNA (miRNA) expression in the brain is a common feature of neurodegenerative diseases. Beyond their conventional role in regulating gene expression at the post-transcriptional level, certain miRNAs can act extracellularly as signaling molecules. Our study elucidates the identity of such miRNA species serving as ligands for membrane receptors expressed in central nervous system (CNS) neurons and the impact of such miRNAs on neurons in the context of neurodegenerative disease. METHODS:We combined a machine learning approach with the analysis of disease-associated miRNA databases to predict Alzheimer's disease (AD)-associated miRNAs as potential signaling molecules for single-stranded RNA-sensing Toll-like receptors (TLRs) 7 and 8. TLR-expressing HEK-Blue reporter cells, primary murine microglia, and human THP-1 macrophages were used to validate the AD miRNAs as ligands for human and mouse TLR7 and/or TLR8. Interaction between mouse cortical neurons and extracellularly applied AD miRNAs was analyzed by live cell imaging and confocal microscopy. Transcriptome changes in cortical neurons exposed to AD miRNAs were assessed by RNAseq and RT-qPCR. The extracellular AD miRNAs' effects on CNS neuron structure were investigated in cell cultures of murine primary cortical neurons and iPSC-derived human cortical neurons by immunocytochemistry. We employed a mouse model of intrathecal injection to assess effects of AD miRNAs acting as signaling molecules on neurons in vivo. RESULTS:We identified the AD-associated miRNAs miR-124-5p, miR-92a-1-5p, miR-9-5p, and miR-501-3p as novel endogenous ligands for TLR7 and/or TLR8. These miRNAs being extracellularly stable and active were taken up by murine cortical neurons via endocytosis and induced changes in neuronal inflammation-, proliferation-, and apoptosis-related gene expression. Exposure of both murine and human cortical neurons to the AD-associated miRNAs led to alterations of dendrite and axon structure, synapse protein expression, and cell viability in a sequence-dependent fashion. Extracellular introduction of the AD miRNAs into the cerebrospinal fluid of mice resulted in both changes in neuronal structure and synapses, and neuronal loss in the cerebral cortex. Most of the observed extracellular miRNA-induced effects on cortical neurons involved TLR7/8 signaling. CONCLUSION:Neurodegenerative disease-associated miRNAs in extracellular form act as signaling molecules for CNS neurons including human cortical neurons, thereby modulating their structure and viability.

This protocol describes picking and expansion of many colonies at once (manual clump passaging) and the picking of single colony (positive selection).

T lymphocytes are key contributors to the adaptive immune system. During development, tightly regulated T cell receptor (TCR) gene rearrangement determines antigen specificity and maturation into distinct lineages with pro- or anti-inflammatory functions. From two male donors, we generated four hiPSC lines from isolated T cell lineages (CD4+ conventional helper, CD4+ regulatory, CD8+ cytotoxic). Two lines harbor genetically pre-rearranged TCRs specific to an allogeneic cell line. All lines were generated by integration-free reprogramming using Sendai virus and underwent characterization and quality control. They represent a valuable platform to investigate how a rearranged and pre-selected TCR influences lymphoid differentiation and function.

Neural progenitors drive cellular diversification in the neocortex. Prolongation of their proliferative capacity and increased diversity underpins the evolutionary expansion and morphological complexity of the human neocortex. Here, we investigate the mechanisms that regulate maintenance of the highly proliferative early neural progenitor subtypes and transition to subsequent progenitors of limited proliferative capacity during human and murine neocortical development. We identify DTX4, a Deltex family member, as an evolutionarily conserved molecular determinant of neural progenitor identity in the mammalian neocortex. DTX4 sustains the identity of radial glia, a highly proliferative progenitor subtype. Loss of DTX4, on the other hand, is a prerequisite for the generation of intermediate progenitors which possess low proliferative capacity. Perturbing DTX4 expression in human cerebral organoids and in the murine neocortex, alters progenitor composition, thereby inducing changes in neuronal diversity and cortical morphology. Mechanistically, we reveal that DTX4 controls progenitor identity by regulating the length of the cell cycle. Our findings underscore the critical role of cell cycle dynamics and of DTX4 in determining progenitor identity and thereby defining neuronal outcome during mammalian development. Highlights ### Competing Interest Statement The authors have declared no competing interest.

Chemotherapy-induced peripheral neuropathy (CIPN) affects up to two-thirds of cancer patients undergoing cytotoxic chemotherapy. Here, we used human iPSC-derived sensory neurons (iPSC-DSN) to model CIPN in vitro. Administration of various chemotherapeutic agents (i.e., paclitaxel, vincristine, bortezomib and cisplatin) at clinically applicable concentrations resulted in reduced cell viability, axonal degeneration, electrophysiological dysfunction and increased levels of phosphorylated c-Jun in iPSC-DSN. Transcriptomic analyses revealed that the upregulation of c-Jun strongly correlated with the expression of genes of neuronal injury, apoptosis and inflammatory signatures. To test whether c-Jun plays a central role in the development of CIPN, we applied the small molecule inhibitor of the Jun N-terminal kinase, SP600125, to iPSC-DSN treated with neurotoxic chemotherapy. c-Jun inhibition prevented chemotherapy-induced neurotoxicity by preserving cell viability, axonal integrity and electrophysiological function of iPSC-DSN. These findings identify c-Jun as a key mediator of CIPN pathophysiology across multiple drug types and present preclinical evidence that c-Jun inhibition is an attractive therapeutic target to prevent CIPN.

The X-linked Allan-Herndon-Dudley syndrome (AHDS) is a genetic disorder characterized by severe psychomotor impairment, resulting from mutations in the SLC16A2 gene, which encodes the thyroid hormone transporter MCT8 (monocarboxylate transporter 8). Previously, we established a hiPSC line from a patient carrying the SLC16A2:R401G mutation (BIHi045-A). Using CRISPR/Cas9-mediated gene editing, we targeted exon 3 of SLC16A2 and used single-stranded oligodeoxynucleotides as homology-directed repair templates to correct the R401G missense mutation, generating an isogenic control cell line.

Human induced pluripotent stem cells (iPSCs) hold great promise for regenerative medicine, disease modelling, and drug discovery, but most downstream applications require differentiation into specialised cell types not covered by current quality control assays. Here, we present “SteMClass”, a proof-of-concept DNA methylation-based classifier that standardises iPSC differentiation state identification across protocols with one test. We curated a reference cohort of 15 iPSC lines differentiated into seven distinct states (n = 97), performed array-based DNA methylation profiling, and trained a random forest model to classify the eight distinct differentiation states. In nested cross-validation, SteMClass achieved a Brier score of 0.0264, and on an independent cohort (n = 58) attained 96.5% accuracy (Cohen’s K = 0.959) with a 3% rejection rate. Applied to external data (n = 241), performance was 76.5% accuracy (Cohen’s K = 0.593) with a 16.6% rejection rate and enabled the detection of potentially inefficient differentiations. SteMClass is compatible with all Illumina methylation array versions, and accessible via an interactive web interface that supports classification and exploration of DNA methylation profiles. By providing a harmonised, single-assay framework for iPSC-derived differentiation state characterisation, SteMClass improves reproducibility and comparability across studies, paving the way for robust quality control standards and accelerating clinical translation. ### Competing Interest Statement The authors have declared no competing interest. German Academic Exchange Service, https://ror.org/039djdh30 Deutschen Konsortium für Translationale Krebsforschung, https://ror.org/02pqn3g31

Heritable pulmonary arterial hypertension (HPAH) and underlying pulmonary vascular disease (PVD) are often caused by TBX4 mutations—either loss- or gain-of-function—which are a leading cause of childhood-onset PAH. The clinically heterogeneous TBX4 syndrome can include skeletal anomalies (e.g., small patella syndrome) and developmental lung disease (DEVLD) (Galambos, 2019). TBX4 is expressed in lung mesenchymal cells such as matrix fibroblasts, pericytes, and smooth muscle cells, all contributing to PAH pathogenesis (Karolak, 2023, Maldonado, 2025). Our five patient-derived TBX4-mutant hiPSC lines provide a powerful model to investigate cell-specific mechanisms in HPAH/DEVLD-PH and support precision drug discovery and therapy development targeting TBX4-related abnormalities.

The X-linked Allan-Herndon-Dudley syndrome (AHDS) is a genetic disorder characterized by severe psychomotor impairment, resulting from mutations in the SLC16A2 gene, which encodes the thyroid hormone transporter MCT8 (monocarboxylate transporter 8). Previously, we established a hiPSC line from a patient carrying the SLC16A2:R401G mutation (BIHi045-A). Using CRISPR/Cas9-mediated gene editing, we targeted exon 3 of SLC16A2 and used single-stranded oligodeoxynucleotides as homology-directed repair templates to correct the R401G missense mutation, generating an isogenic control cell line.

We generated the human induced pluripotent stem cell (iPSC) line BIHi261-A from dermal fibroblasts of a patient with severe early-onset obesity caused by a homozygous truncating mutation in the POMC gene (W84X). Reprogramming was performed using a non-integrating, RNA-based vector expressing key pluripotency factors. The resulting iPSC line exhibited typical morphology, expressed markers of undifferentiated cells, maintained a normal karyotype, and demonstrated the capacity to differentiate into cell types of all three germ layers. BIHi261-A provides a valuable tool for studying the molecular mechanisms of POMC-related obesity and for developing potential therapeutic strategies.

Allan-Herndon-Dudley syndrome (AHDS) is an X-linked disorder characterized by profound psychomotor impairment. It is caused by mutations in the SLC16A2 gene, which encodes monocarboxylate transporter 8 (MCT8), a crucial thyroid hormone transporter. Here we report generation of two male patient-derived iPSC lines harboring either SLC16A2:G401R or SLC16A2:H192R.

Immunocompromised patients, such as those undergoing hematopoietic stem cell or solid organ transplantation, are highly susceptible to viral complications. Given the limitations and side effects of available antiviral therapies, adoptive transfer of antiviral T cells offers a promising alternative by restoring immune defense. However, existing models for evaluating antiviral T cell therapies lack physiological relevance, limiting accurate predictions of efficacy and safety. There is a critical need for in vitro human infection platforms that support personalized assessment of therapeutic responses. To address this, we developed antiviral T cell products (TCPs) targeting Influenza A virus (IAV)-infected cells, alongside an autologous human induced pluripotent stem cell (iPSC)-derived 3D lung organoid infection platform. This model recapitulates key immunological responses and is compatible with a new 3D high-throughput, high-content imaging pipeline. Our study provides the first proof-of-concept for assessing T cell-mediated cytotoxicity in a 3D in vitro lung infection model, advancing personalized antiviral immunotherapy development.

Understanding how fluctuations propagate across spatial scales is central to our understanding of inanimate matter from turbulence to critical phenomena. In contrast to physical systems, biological systems are organized into a hierarchy of processes on a discrete set of spatial scales: they are compartmentalized. Here, we show that dynamic compartmentalization of stochastic systems leads to emergent, quasi-particle-like kinetics which are used by cells to perform key biological functions. Specifically, we derive a general theory that predicts the emergence of a single degree of freedom irrespective of system specifics. We obtain equations of motion and response characterising its unique kinetic properties. We experimentally demonstrate the biological relevance of quasi-particle kinetics in the decision of cells to commit suicide (apoptosis). Using fluorescent microscopy, we show that the response of cells to apoptotic stimuli exhibits quasi-particle like kinetics which establish a low-pass filter for cellular stress signals. By highlighting that cells manipulate how noise and signals propagate across spatial scales, our work reveals a new mechanism of cell fate decision-making.

The role of the nuclear thyroid hormone receptor TRα1 as a mediator of thyroid hormone action on target gene expression is well understood. However, the function of the TRα2 splicing isoform, which does not bind thyroid hormones, remains unexplored. As no reliable antibodies are available to investigate TRα1 and TRα2 specifically, we introduced small fusion tags into the THRA locus of the male healthy donor iPSC lines BIHi001-B and BIHi005-A by CRISPR/Cas9-mediated genome editing. Consequently, the modified lines express C-terminally tagged TRα1-2xHA or TRα2-3xFLAG. These genome-edited lines facilitate the investigation of isoform-specific actions of TRα in different cell types.