Human Embryonic Stem Cell-derived Lung Organoids-based Cell Therapy For Lung Injury In Mouse Models

Pulmonary fibrosis,especially idiopathic pulmonary fibrosis(IPF),is a chronic,progressive,and fibrotic lung disease.There is currently no effective treatment.Human embryonic stem cell-derived lung organoids(HLOs)can better simulate the lung development process in vivo and differentiate into multiple lung cell types including proximal tracheal cells and distal alveolar cells,which opens up the possibility of cell therapy for IPF.Based on the HLOs platform that has been established in this laboratory,this topic investigates its application and effect in treating acute lung injury mice.First,using six cytokines or small molecule compounds to induce human embryonic stem cells to differentiate into lung organoids by day 25(at this time,HLOs highly express lung progenitor cell marker NKX2.1),and a typical spherical structure of HLOs was observed,QRT-PCR detected HLOs overexpressing NKX2.1;Studies have shown that most of the cells expressing CD47 in the lungs(CD47H)are mostly NKX2.1 positive cells.Therefore,CD47 is used as a marker for sorting NKX2.1+lung progenitor cells to obtain donor cells suitable for transplantation.After the establishment of mice acute lung injury models(40-48 hours after NA injury,8 days after bleomycin injury),different seed cells(divided into normal group,DPBS group,POOL group and CD47H group)were transplanted through the trachea.for lung injury.Immunofluorescence staining experiments showed that donor cells can survive,differentiate and form specific lung epithelial structures in mouse lungs 12 weeks after tracheal transplantation.Living lung function test experiments 6 weeks after cell transplantation showed that lung function such as respiratory rate(f),tidal volume(TVb),and expiratory flow-50(EF50)of the treatment group mice were alleviated to a certain extent;Blood gas exchange analysis after 12 weeks of cell transplantation showed that in the NA injury model,the blood gas function of the POOL transplantation group mice was restored to a certain extent;while in the BLE injury model,the CD47H transplantation group seemed to have a certain effect.HE staining and Masson staining of the lung lobes further support the above-mentioned blood gas exchange results.In summary,this study used tracheal transplantation of HLOs lung cells to treat acute lung injury model mice.The results show that the cells can survive in the recipient and form specific lung epithelial structures,and can alleviate mouse lung injury in many aspects such as living lung function,blood gas exchange function,and pulmonary fibrosis.It provides a reference for treating human lung diseases such as IPF.