Figure 3.

Loss of PDX1 in EndoC-βH1cells impairs insulin secretion and calcium flux and results in aberrant regulation of β-cell-specific genes. (A) EndoC-βH1 cells PDX1 knockout show impaired insulin secretion and calcium flux as measured by luciferase secretion and GFP intensity of GCaMP6s in a perfusion system compared with wild-type cells in Fig. 1C. (B) EndoC-βH1 cells were transduced with lentivirus carrying CRISPR–CAS9 and gRNA specific for PDX1 gene; EndoC-βH1 transduced cells were then sorted into distinct PDX1+/+ and PDX1–/– populations by intracellular FACS. (C) Quantification of RNA expression by qRT-PCR comparing PDX1+/+ and PDX1–/– cell populations. Dysregulation of β-cell-specific genes is seen including downregulation of INS, MAFA, and NKX6.1 and upregulation of GCG. Unpaired 2-tailed Student’s t-tests were performed for each gene (n = 3, *P < .05).