A differentiation protocol for generating pancreatic delta cells from human pluripotent stem cells

oleh: Tongran Zhang, Tongran Zhang, Nannan Wang, Nannan Wang, Zhiying Liao, Zhiying Liao, Jingyi Chen, Jingyi Chen, Hao Meng, Hao Meng, Haopeng Lin, Tao Xu, Tao Xu, Lihua Chen, Lihua Chen, Ling-Qiang Zhu, Huisheng Liu, Huisheng Liu, Huisheng Liu, Huisheng Liu

Format: Article
Diterbitkan: Frontiers Media S.A. 2024-10-01

Deskripsi

In this protocol, we detail a seven-stage differentiation methodology for generating pancreatic delta cells (SC-delta cells) from human pluripotent stem cells (hPSCs). In the first step, definitive endoderm is generated by activin A and CHIR99021, followed by induction of primitive gut tube and posterior foregut by treatment with FGF7, SANT1, LDN193189, PdBU, and retinoic acid (RA). The subsequent endocrine generation and directed SC-delta cell induction is achieved by a combined treatment of the FGF7 with FGF2 during stage 4 and 5, together with RA, XXI, ALK5 inhibitor II, SANT1, Betacellulin and LDN193189. The planar cultivation is converted to a suspended system after stage 5, allowing cells to aggregate into delta cell-containing spheroids. The differentiation takes approximately 4-5 weeks for delta cell generation and an additional 1-2 weeks for cell expansion and evaluation. We believe that this amenable and simplified protocol can provide a stable source of SC-delta cells from efficient differentiation, facilitating further investigation of the physiological role of delta cells as well as refinement of islet cell therapeutic strategies.