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Understanding how chemical stress perturbs human lung physiology requires models that capture dynamic molecular responses in real time. Here, we established a CRISPR/Cas9-engineered human induced pluripotent stem cell (hiPSC)-derived lung organoid expressing endogenous G3BP1–mCherry, enabling live, non-destructive visualization of stress granule (SG) formation under toxicant exposure. The organoids recapitulated airway and alveolar epithelial diversity and displayed lamellar body-like ultrastructures, indicating advanced maturation. Time-lapse imaging revealed rapid and reversible SG dynamics across chemically distinct stressors, while cytotoxicity assays showed that these organoids are significantly more sensitive than conventional 2D or cancer-derived lung models. Importantly, SG dynamics were linked to exposure duration–dependent changes in epithelial barrier integrity, indicating that SG formation precedes overt epithelial injury and serves as an early indicator of toxicant-induced cellular stress. Integration with high-content screening enabled quantitative, image-based analysis of cellular stress phenotypes, greatly enhancing throughput and mechanistic insight, thereby provided next-generation New Approach Methodologies for lung toxicity assessment. Together, this hiPSC-derived lung organoid SG reporter platform links early molecular stress adaptation to tissue-level responses, offering a predictive and mechanistically informative framework for human-relevant lung toxicity evaluation.

Craniosynostosis is a multigenic congenital condition in which one or more calvarial sutures have prematurely fused during the development of the fetus. Pathogenic variants in FGFR2 are associated with the development of syndromic craniosynostosis, such as Crouzon, Apert and Pfeifer syndromes. Investigation of FGFR2 -linked craniosynostosis is hindered by the lack of appropriate in vitro models. Patient-derived human induced pluripotent stem cell (hiPSC) in vitro disease models provide the opportunity to investigate the disease, identify molecular targets for pharmaceutical treatments, and enable the generation of autologous pluripotent stem cell catalogues. Here, we report three patient-derived hiPSC lines carrying the C342Y, S252W or E565G FGFR2 pathogenic variant. The patient hiPSC lines express characteristic pluripotency markers and display distinct phosphorylation profiles under unstimulated conditions. FGFR2 C342Y showed autophosphorylation in the absence of bFGF ligand, although downstream docking proteins PLCγ and FRS2α were not phosphorylated. FGFR2 S252W and FGFR2 E565G hiPSCs showed increased phosphorylation of docking proteins PLCγ and FRS2α, whereas FGFR2 was not phosphorylated. These patient hiPSC lines provide molecular and cellular options to investigate FGFR2 -linked craniosynostosis in the patient-specific genomic context and develop therapeutic modalities.

Neurological damage caused by peripheral nerve injury can be devastating and is a common neurological disorder that, along with muscle disorders, reduces the quality of life. Neural crest cells (NCCs) are a transient cell population that occurs during embryogenesis, can differentiate into various cells upon transplantation, and has potential therapeutic effects on neurological diseases. However, there are limitations to cell therapy, such as uncontrolled differentiation and tumor formation. Extracellular vesicles (EVs), which are non-cellular potential therapeutic candidates, are nanosized membrane-bound vesicles. Studies have been reported using starch cells derived from neural cells, such as neural stem cells, because they are involved in cell-to-cell communication and carry a variety of bioactive molecules. We investigated the effects of EVs isolated from NCCs on neuronal cell death and inflammation. Additionally, we overexpressed the nerve growth factor (NGF), which is involved in neural cell growth and proliferation, in NCCs to further investigate the effects of EVs containing NGF. NCCoe-NGF-EVs showed neuroprotective and regenerative properties by modulating inflammatory pathway, promoting Schwann cell activation, and enhancing remyelination. In vitro studies on NCCoe-NGF-EVs suppressed pro-inflammatory cytokines and reduced oxidative stress-induced neuronal apoptosis through NF-?B pathway inhibition and ERK, AKT signal activation. We also evaluated the effect of EVs on neuropathy by performing in vivo study. Our results suggest that NCCoe-NGF-EV had neuroprotective effects by reducing neuronal apoptosis and promoting neuronal proliferation based on neurite outgrowth and anti-inflammation effects treated with NCCoe-NGF-EVs. In addition, NCCoe-NGF-EVs were protected muscle loss caused by peripheral nerve injury. NCCoe-NGF-EV induced regeneration of damaged nerves and inhibited cell death through anti-inflammatory effects. This study suggests the potential of NGF-enriched EVs as non-cellular therapeutic platform for peripheral neuropathies and other neuroinflammatory disorders.