Single-Cell Transcriptomics of the Human Endocrine Pancreas

Abstract

Human pancreatic islets consist of multiple endocrine cell types. To facilitate the detection of rare cellular states and uncover population heterogeneity, we performed single-cell RNA sequencing (RNA-seq) on islets from multiple deceased organ donors, including children, healthy adults, and individuals with type 1 or type 2 diabetes. We developed a robust computational biology framework for cell type annotation. Using this framework, we show that α- and β-cells from children exhibit less well-defined gene signatures than those in adults. Remarkably, α- and β-cells from donors with type 2 diabetes have expression profiles with features seen in children, indicating a partial dedifferentiation process. We also examined a naturally proliferating α-cell from a healthy adult, for which pathway analysis indicated activation of the cell cycle and repression of checkpoint control pathways. Importantly, this replicating α-cell exhibited activated Sonic hedgehog signaling, a pathway not previously known to contribute to human α-cell proliferation. Our study highlights the power of single-cell RNA-seq and provides a stepping stone for future explorations of cellular heterogeneity in pancreatic endocrine cells.

Single-cell transcriptomes identify human islet cell signatures and reveal cell-type–specific expression changes in type 2 diabetes

“…Approximately one-third (340/978) of cells expressed more than one marker gene; these were removed from subsequent analysis due to concerns that these represent two vertically stacked cells in a given capture site (for details, see Methods). Similar ratios of potential stacked cells have been reported in other studies using the Fluidigm C1 platform to capture mouse (Xin et al 2016) and human islet cells (Wang et al 2016). Collectively, these single-cell data capture transcriptome profiles representing each of the major and minor pancreatic endocrine and exocrine cell types.…”

“…We therefore sought to validate our observed celltype-specific differences in T2D islets using these independent data sets (Wang et al 2016;Segerstolpe et al 2016). We found that 54/77 genes and 32/171 were also significantly up-and down-regulated, respectively, in T2D beta cells in these studies (P < 0.05, two-sided Wilcoxon rank-sum test) (Supplemental Fig.…”

“…Five islet endocrine cell types have been assigned based on exclusive and robust expression of the peptide hormone genes INS (beta), GCG (alpha), SST (delta), PPY (PP/gamma), and GHRL (epsilon) (Baetens et al 1979;Nussey and Whitehead 2001;Zhao et al 2008;Li et al 2016;Xin et al 2016;Wang et al 2016). The pancreas also contains three exocrine cell types-acinar, stellate, and ductal-that critically support digestion through synthesis and transport of digestive enzymes (Pandol 2011;Reichert and Rustgi 2011).…”

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

“…GLP1R mRNA levels were similar in juvenile and adult human islets (Figure 2A), suggesting that human β cell GLP-1R expression is not age-dependent. Moreover, in FACSisolated islet cell subsets, GLP1R mRNA was principally expressed in β cells and was very low, or not detectable, in α cells (Supplemental Figure 2, A-C), consistent with prior findings (10,(31)(32)(33)(34)(35).…”

PAX6 maintains β cell identity by repressing genes of alternative islet cell types

“…Haploinsufficiency of PAX6 in humans does interfere with β cell function and leads to abnormal glucose homeostasis, suggesting that even partial effects on PAX6 expression may have glycemic consequences (27,28). We note, however, that in expression profiles obtained from type 2 diabetic donors using both whole islets and single β cells, no significant reduction in PAX6 expression levels was observed compared with levels detected in nondiabetic donors (7,63,64). This may be attributed to euglycemia in those patients.…”