Embryoid body-based differentiation of human-induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotype
Objective The transparency of the cornea is determined by the extracellular matrix, which is secreted by corneal stromal keratocytes (CSKs). Human-induced pluripotent stem cell (hiPSC)-derived keratocytes (hiPSC-CSKs) can be used in cell-based therapy for treating corneal blindness. Our goal was to...
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BMJ Publishing Group
2024-12-01
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| Series: | BMJ Open Ophthalmology |
| Online Access: | https://bmjophth.bmj.com/content/9/1/e001828.full |
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| author | Qian Li Jie Chen Yifan Liu Qingjian Ou Tingting Cui Huimin Yang Jiancen Tang Lixia Lu Guotong Xu Hongping Cui Caixia Jin |
| author_facet | Qian Li Jie Chen Yifan Liu Qingjian Ou Tingting Cui Huimin Yang Jiancen Tang Lixia Lu Guotong Xu Hongping Cui Caixia Jin |
| author_sort | Qian Li |
| collection | DOAJ |
| description | Objective The transparency of the cornea is determined by the extracellular matrix, which is secreted by corneal stromal keratocytes (CSKs). Human-induced pluripotent stem cell (hiPSC)-derived keratocytes (hiPSC-CSKs) can be used in cell-based therapy for treating corneal blindness. Our goal was to develop an effective small molecule-based technique for differentiating hiPSCs into keratocytes.Methods and analysis hiPSCs were cultured in chemically defined medium, and embryoid bodies (EBs) were generated; these EBs were induced into CSKs using keratocyte-differentiated medium. The expression of keratocyte-specific markers was assessed using quantitative RT-PCR, immunostaining and Western blotting.Results We found that the expression of genes encoding keratocyte markers, including aldehyde dehydrogenase 1 family member A1 (ALDH1A1), lumican and keratocan, was upregulated. Immunostaining showed positive staining for ALDH1A1 and keratocan in the hiPSC-CSK samples. Similarly, western blot analysis indicated that ALDH1A1 and keratocan expression levels were significantly greater in the hiPSC-CSKs than in the control cells. In addition, hiPSC-CSKs were not transformed into fibroblasts or myofibroblasts.Conclusion We established an innovative and effective method to generate CSKs via the EB-based differentiation of hiPSCs, which might be employed for cell-based therapy of corneal stromal opacities. |
| format | Article |
| id | doaj-art-e8a751dcdbcc4d97b4208bfb1197b52d |
| institution | Kabale University |
| issn | 2397-3269 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | BMJ Publishing Group |
| record_format | Article |
| series | BMJ Open Ophthalmology |
| spelling | doaj-art-e8a751dcdbcc4d97b4208bfb1197b52d2024-12-14T23:15:08ZengBMJ Publishing GroupBMJ Open Ophthalmology2397-32692024-12-019110.1136/bmjophth-2024-001828Embryoid body-based differentiation of human-induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotypeQian Li0Jie Chen1Yifan Liu2Qingjian Ou3Tingting Cui4Huimin Yang5Jiancen Tang6Lixia Lu7Guotong Xu8Hongping Cui9Caixia Jin10Department of Ophthalmology, Shanghai East Hospital, School of Medicine, Tongji University, Shanghai 200120, ChinaDepartment of Ophthalmology, Shanghai East Hospital, School of Medicine, Tongji University, Shanghai 200120, ChinaDepartment of Ophthalmology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, ChinaDepartment of Ophthalmology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, ChinaDepartment of Ophthalmology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, ChinaDepartment of Ophthalmology, Ren Ji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, ChinaDepartment of Ophthalmology, Shanghai East Hospital, School of Medicine, Tongji University, Shanghai 200120, ChinaDepartment of Ophthalmology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, ChinaDepartment of Ophthalmology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, ChinaDepartment of Ophthalmology, Shanghai East Hospital, School of Medicine, Tongji University, Shanghai 200120, ChinaDepartment of Ophthalmology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, ChinaObjective The transparency of the cornea is determined by the extracellular matrix, which is secreted by corneal stromal keratocytes (CSKs). Human-induced pluripotent stem cell (hiPSC)-derived keratocytes (hiPSC-CSKs) can be used in cell-based therapy for treating corneal blindness. Our goal was to develop an effective small molecule-based technique for differentiating hiPSCs into keratocytes.Methods and analysis hiPSCs were cultured in chemically defined medium, and embryoid bodies (EBs) were generated; these EBs were induced into CSKs using keratocyte-differentiated medium. The expression of keratocyte-specific markers was assessed using quantitative RT-PCR, immunostaining and Western blotting.Results We found that the expression of genes encoding keratocyte markers, including aldehyde dehydrogenase 1 family member A1 (ALDH1A1), lumican and keratocan, was upregulated. Immunostaining showed positive staining for ALDH1A1 and keratocan in the hiPSC-CSK samples. Similarly, western blot analysis indicated that ALDH1A1 and keratocan expression levels were significantly greater in the hiPSC-CSKs than in the control cells. In addition, hiPSC-CSKs were not transformed into fibroblasts or myofibroblasts.Conclusion We established an innovative and effective method to generate CSKs via the EB-based differentiation of hiPSCs, which might be employed for cell-based therapy of corneal stromal opacities.https://bmjophth.bmj.com/content/9/1/e001828.full |
| spellingShingle | Qian Li Jie Chen Yifan Liu Qingjian Ou Tingting Cui Huimin Yang Jiancen Tang Lixia Lu Guotong Xu Hongping Cui Caixia Jin Embryoid body-based differentiation of human-induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotype BMJ Open Ophthalmology |
| title | Embryoid body-based differentiation of human-induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotype |
| title_full | Embryoid body-based differentiation of human-induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotype |
| title_fullStr | Embryoid body-based differentiation of human-induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotype |
| title_full_unstemmed | Embryoid body-based differentiation of human-induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotype |
| title_short | Embryoid body-based differentiation of human-induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotype |
| title_sort | embryoid body based differentiation of human induced pluripotent stem cells into cells with a corneal stromal keratocyte phenotype |
| url | https://bmjophth.bmj.com/content/9/1/e001828.full |
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