Age-dependent human β cell proliferation induced by glucagon-like peptide 1 and calcineurin signaling
Chunhua Dai, Yan Hang, Alena Shostak, Greg Poffenberger, Nathaniel Hart, Nripesh Prasad, Neil Phillips, Shawn E. Levy, Dale L. Greiner, Leonard D. Shultz, Rita Bottino, Seung K. Kim, Alvin C. Powers
Chunhua Dai, Yan Hang, Alena Shostak, Greg Poffenberger, Nathaniel Hart, Nripesh Prasad, Neil Phillips, Shawn E. Levy, Dale L. Greiner, Leonard D. Shultz, Rita Bottino, Seung K. Kim, Alvin C. Powers
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Research Article Endocrinology

Age-dependent human β cell proliferation induced by glucagon-like peptide 1 and calcineurin signaling

Abstract

Inadequate pancreatic β cell function underlies type 1 and type 2 diabetes mellitus. Strategies to expand functional cells have focused on discovering and controlling mechanisms that limit the proliferation of human β cells. Here, we developed an engraftment strategy to examine age-associated human islet cell replication competence and reveal mechanisms underlying age-dependent decline of β cell proliferation in human islets. We found that exendin-4 (Ex-4), an agonist of the glucagon-like peptide 1 receptor (GLP-1R), stimulates human β cell proliferation in juvenile but not adult islets. This age-dependent responsiveness does not reflect loss of GLP-1R signaling in adult islets, since Ex-4 treatment stimulated insulin secretion by both juvenile and adult human β cells. We show that the mitogenic effect of Ex-4 requires calcineurin/nuclear factor of activated T cells (NFAT) signaling. In juvenile islets, Ex-4 induced expression of calcineurin/NFAT signaling components as well as target genes for proliferation-promoting factors, including NFATC1, FOXM1, and CCNA1. By contrast, expression of these factors in adult islet β cells was not affected by Ex-4 exposure. These studies reveal age-dependent signaling mechanisms regulating human β cell proliferation, and identify elements that could be adapted for therapeutic expansion of human β cells.

Authors

Chunhua Dai, Yan Hang, Alena Shostak, Greg Poffenberger, Nathaniel Hart, Nripesh Prasad, Neil Phillips, Shawn E. Levy, Dale L. Greiner, Leonard D. Shultz, Rita Bottino, Seung K. Kim, Alvin C. Powers

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Figure 3

Gene expression in human islets and juvenile grafts after treatment with Ex-4 and/or FK506.

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Gene expression in human islets and juvenile grafts after treatment with...
(A and B) RNA-Seq analysis of PBS- or Ex-4–treated juvenile grafts (n = 3 grafts per treatment, 6-year-old donor). (A) Number of genes regulated by Ex-4 (≥1.5-fold) in juvenile grafts (Ex-4 vs. PBS); 1,115 genes are upregulated and 325 are downregulated by Ex-4 treatment. (B) Signaling pathways upregulated by Ex-4. (C) Genes of the NFATC family are expressed more highly in juvenile islets (n = 5 donors, age: 0.5, 1.2, 1.7, 3, and 4 years) than adult (n = 5 donors, age: 29, 43, 50, 53, and 60 years). The average expression level in juvenile islets is defined as 1.00 for each gene. (D) Schematic of transplantation and treatment with Ex-4 plus FK506. Two weeks after transplantation, a pump with PBS or Ex-4 was implanted, and a second pump with saline or FK506 was implanted in the last 2 weeks. Grafts were removed at 4 weeks. (E–G) Juvenile graft gene expression measured by qPCR. Ex-4 upregulates genes of the NFAT family (E), cell cycle regulators (F), and transcription factors (G), and the effects were diminished by FK506 (n = 5 graft samples per treatment from 0.2- and 6-year-old donors). The average expression level in PBS+saline–treated islets is defined as 1.00 for each gene. Error bars represent SEM. *P < 0.05, **P < 0.01, ***P < 0.001 vs. PBS+saline; †P < 0.05, ††P < 0.01, †††P < 0.001 vs. Ex-4+saline; no significant differences between PBS+FK506 and Ex-4+FK506. Unpaired 2-tailed Student’s t test or 1-way ANOVA followed by Newman-Keuls multiple-comparisons test (E–G) was used for analysis of statistical significance. See also Supplemental Figures 4 and 5.