Screening of factors inducing alveolar type 1 epithelial cells using human pluripotent stem cells.

ヒトiPS細胞由来肺胞スフェロイドの"on-gel培養法"による化合物スクリーニング --I型肺胞上皮細胞の分化を促進するシグナル経路を同定--. 京都大学プレスリリース. 2024-03-29.Constructing "on-gel" alveolar organoids as a new screening platform. 京都大学プレスリリース. 2024-03-29.Alveolar type 2 (AT2) epithelial cells are tissue stem cells capable of differentiating into alveolar type 1 (AT1) cells for injury repair and maintenance of lung homeostasis. However, the factors involved in human AT2-to-AT1 cell differentiation are not fully understood. Here, we established SFTPCGFP and AGERmCherry-HiBiT dual-reporter induced pluripotent stem cells (iPSCs), which detected AT2-to-AT1 cell differentiation with high sensitivity and identified factors inducing AT1 cell differentiation from AT2 and their progenitor cells. We also established an "on-gel" alveolar epithelial spheroid culture suitable for medium-throughput screening. Among the 274 chemical compounds, several single compounds, including LATS-IN-1, converted AT1 cells from AT2 and their progenitor cells. Moreover, YAP/TAZ signaling activation and AKT signaling suppression synergistically recapitulated the induction of transcriptomic, morphological, and functionally mature AT1 cells. Our findings provide novel insights into human lung development and lung regenerative medicine

Disease modeling of pulmonary fibrosis using human pluripotent stem cell-derived alveolar organoids

iPS細胞を用いて作製した肺胞オルガノイドで間質性肺炎の病態再現に成功 --治療満足度の低い間質性肺炎の治療薬開発に向けて前進--. 京都大学プレスリリース. 2021-11-19.Although alveolar epithelial cells play a critical role in the pathogenesis of pulmonary fibrosis, few practical in vitro models exist to study them. Here, we established a novel in vitro pulmonary fibrosis model using alveolar organoids consisting of human pluripotent stem cell-derived alveolar epithelial cells and primary human lung fibroblasts. In this human model, bleomycin treatment induced phenotypes such as epithelial cell-mediated fibroblast activation, cellular senescence, and presence of alveolar epithelial cells in abnormal differentiation states. Chemical screening performed to target these abnormalities showed that inhibition of ALK5 or blocking of integrin αVβ6 ameliorated the fibrogenic changes in the alveolar organoids. Furthermore, organoid contraction and extracellular matrix accumulation in the model recapitulated the pathological changes observed in pulmonary fibrosis. This human model may therefore accelerate the development of highly effective therapeutic agents for otherwise incurable pulmonary fibrosis by targeting alveolar epithelial cells and epithelial-mesenchymal interactions

ACE2 knockout hinders SARS-CoV-2 propagation in iPS cell-derived airway and alveolar epithelial cells

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, continues to spread around the world with serious cases and deaths. It has also been suggested that different genetic variants in the human genome affect both the susceptibility to infection and severity of disease in COVID-19 patients. Angiotensin-converting enzyme 2 (ACE2) has been identified as a cell surface receptor for SARS-CoV and SARS-CoV-2 entry into cells. The construction of an experimental model system using human iPS cells would enable further studies of the association between viral characteristics and genetic variants. Airway and alveolar epithelial cells are cell types of the lung that express high levels of ACE2 and are suitable for in vitro infection experiments. Here, we show that human iPS cell-derived airway and alveolar epithelial cells are highly susceptible to viral infection of SARS-CoV-2. Using gene knockout with CRISPR-Cas9 in human iPS cells we demonstrate that ACE2 plays an essential role in the airway and alveolar epithelial cell entry of SARS-CoV-2 in vitro. Replication of SARS-CoV-2 was strongly suppressed in ACE2 knockout (KO) lung cells. Our model system based on human iPS cell-derived lung cells may be applied to understand the molecular biology regulating viral respiratory infection leading to potential therapeutic developments for COVID-19 and the prevention of future pandemics

Cryptotanshinone is a candidate therapeutic agent for interstitial lung disease associated with a BRICHOS-domain mutation of SFTPC

Summary: Interstitial lung disease (ILD) represents a large group of diseases characterized by chronic inflammation and fibrosis of the lungs, for which therapeutic options are limited. Among several causative genes of familial ILD with autosomal dominant inheritance, the mutations in the BRICHOS domain of SFTPC cause protein accumulation and endoplasmic reticulum stress by misfolding its proprotein. Through a screening system using these two phenotypes in HEK293 cells and evaluation using alveolar epithelial type 2 (AT2) cells differentiated from patient-derived induced pluripotent stem cells (iPSCs), we identified Cryptotanshinone (CPT) as a potential therapeutic agent for ILD. CPT decreased cell death induced by mutant SFTPC overexpression in A549 and HEK293 cells and ameliorated the bleomycin-induced contraction of the matrix in fibroblast-dependent alveolar organoids derived from iPSCs with SFTPC mutation. CPT and this screening strategy can apply to abnormal protein-folding-associated ILD and other protein-misfolding diseases