PT  - JOURNAL ARTICLE
AU  - Sevim Kahraman
AU  - Kimitaka Shibue
AU  - Dario F. De Jesus
AU  - Jiang Hu
AU  - Debasish Manna
AU  - Bridget K. Wagner
AU  - Amit Choudhary
AU  - Rohit N. Kulkarni
TI  - Fluorescein-based sensors to purify human α-cells for functional and transcriptomic analyses
AID  - 10.1101/2022.11.27.518097
DP  - 2022 Jan 01
TA  - bioRxiv
PG  - 2022.11.27.518097
4099  - http://biorxiv.org/content/early/2022/11/28/2022.11.27.518097.short
4100  - http://biorxiv.org/content/early/2022/11/28/2022.11.27.518097.full
AB  - Pancreatic α-cells secrete glucagon, an insulin counter-regulatory peptide hormone critical for the maintenance of glucose homeostasis. Investigation of the function of human α-cells remains a challenge due to the lack of cost-effective purification methods to isolate high-quality α-cells from islets. Here, we use the reaction-based probe diacetylated Zinpyr1 (DA-ZP1) to introduce a novel and simple method for enriching live α-cells from dissociated human islet cells with > 97% purity. The α-cells, confirmed by sorting and immunostaining for glucagon, were cultured up to 10 days to form α-pseudoislets. The α-pseudoislets could be maintained in culture without significant loss of viability, and responded to glucose challenge by secreting appropriate levels of glucagon. RNA-sequencing analyses (RNA-seq) revealed that expression levels of key α-cell identity genes were sustained in culture while some of the genes such as DLK1, GSN, SMIM24 were altered in α-pseudoislets in a time-dependent manner. In conclusion, we report a method to sort human primary α-cells with high purity that can be used for downstream analyses such as functional and transcriptional studies.Competing Interest StatementThe authors have declared no competing interest.