The Role of ARX in Human Pancreatic Endocrine Specification

Abstract

The in vitro differentiation of human embryonic stem cells (hESCs) offers a model system to explore human development. Humans with mutations in the transcription factor Aristaless Related Homeobox (ARX) often suffer from the syndrome X-linked lissencephaly with ambiguous genitalia (XLAG), affecting many cell types including those of the pancreas. Indeed, XLAG pancreatic islets lack glucagon and pancreatic polypeptide-positive cells but retain somatostatin, insulin, and ghrelin-positive cells. To further examine the role of ARX in human pancreatic endocrine development, we utilized genomic editing in hESCs to generate deletions in ARX. ARX knockout hESCs retained pancreatic differentiation capacity and ARX knockout endocrine cells were biased toward somatostatin-positive cells (94% of endocrine cells) with reduced pancreatic polypeptide (rarely detected), glucagon (90% reduced) and insulin-positive (65% reduced) lineages. ARX knockout somatostatin-positive cells shared expression patterns with human fetal and adult δ-cells. Differentiated ARX knockout cells upregulated PAX4, NKX2.2, ISL1, HHEX, PCSK1, PCSK2 expression while downregulating PAX6 and IRX2. Re-expression of ARX in ARX knockout pancreatic progenitors reduced HHEX and increased PAX6 and insulin expression following differentiation. Taken together these data suggest that ARX plays a key role in pancreatic endocrine fate specification of pancreatic polypeptide, somatostatin, glucagon and insulin positive cells from hESCs.

Converting Adult Pancreatic Islet α Cells into β Cells by Targeting Both Dnmt1 and Arx

“…However the molecular or regulatory features underlying development of these abnormal Glucagon + cells in T1D has not been described, reflecting inherent difficulties of pancreas procurement in humans with specific diseases. The finding of ARX and DNMT1 loss or reduction in Glucagon + Insulin + cells from two subjects with T1D corroborates prior studies in mice and human cell lines suggesting that Arx or Dnmt1 establish and/or maintain α-cell fate and function (Collombat et al, 2003; Collombat et al, 2007; Collombat et al, 2009; Avrahami et al, 2015; Gage et al, 2015). Moreover, the abnormal expression of non α-cell factors such as NKX6.1 and PDX1 in a subset of Glucagon + cells is consistent with prior reports of impaired α-cell function in T1D (Cryer et al, 2003; Pietropaolo et al, 2013).…”

Mutant Mice With Calcium-Sensing Receptor Activation Have Hyperglycemia That Is Rectified by Calcilytic Therapy

“…5(i) and 5(j) ]. qRT-PCR analysis utilizing RNA from isolated Casr +/+ and Casr Nuf/Nuf islets revealed that these changes in β -cell and α -cell proliferation were not associated with significant alterations in the expression of genes regulating islet mass such as Foxo1 , Foxm1 , Ngn3 , and Tcf7l2 ( 31 – 34 ), which promote β -cell proliferation, or in the expression of genes such as Arx and Irx2 ( 35 ), which influence α -cell proliferation ( ).…”

Present status and expectation of aristaless-related homeobox (ARX) in endocrine pancreas

“…ARX inactivation is expected to induce stem cell or other mature pancreatic endocrine cell in to insulin-producing cell as an essential factor (Pearl and Horb, 2008). Surprisingly, the experiment in vitro suggests that ARX inhibition does not have obvious effective on the trandifferentiation into b-cells in differentiated human embryonic stem cells using 33-day and7satges protocol or expanded a-cells treated with a combination of solube factors (Gage et al, 2015). However, misactivation of ARX inhibits the redifferentation of ex-vivo expansion of b-cells, elevates insulin mRNA levels and increases the productivity of insulin-positive cells, which suggests ARX blocking could be an effective approach of facilitate the generation of abundant b-cells under defined conditions (Friedman-Mazursky et al, 2016).…”