Islet Cell and Regenerative Biology, Joslin Diabetes Center, Boston, United States.
Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, United States.
Elife. 2023 Sep 21;12:e85056. doi: 10.7554/eLife.85056.
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 ~95% 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 , , 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.
胰岛的α细胞分泌胰高血糖素，这是一种胰岛素的反向调节肽激素，对维持血糖稳态至关重要。由于缺乏经济有效的纯化方法从胰岛中分离出高质量的α细胞，因此研究人类α细胞的功能仍然是一个挑战。在这里，我们使用基于反应的探针二乙酰化 Zinpyr1（DA-ZP1），引入了一种新颖而简单的方法，可从分离的人胰岛细胞中富集活的α细胞，纯度高达 95%。通过分选和免疫染色鉴定为胰高血糖素的α细胞可培养长达 10 天形成α细胞假胰岛。α细胞假胰岛在培养中可保持活力，且对葡萄糖刺激的反应可分泌适当水平的胰高血糖素。RNA 测序分析（RNA-seq）表明，在培养过程中关键的α细胞特征基因的表达水平得以维持，而一些基因如 、 、 则随着时间的推移在α细胞假胰岛中发生改变。总之，我们报告了一种可以高纯度分选人原代α细胞的方法，可用于下游分析，如功能和转录研究。
