Rabbit Anti-Human PTF1A monoclonal antibody, clone 23I5M5

Beta cell development is sensitive to glucocorticoid levels. Although direct effects of glucocorticoids on pancreatic precursors have been shown to control beta cell mass expansion, indirect effects of these hormones on pancreatic development remain unexplored. This issue was addressed in mice lacking the glucocorticoid receptor (GR) in the whole organism. The pancreatic phenotype of GR(null/null) mice was studied at fetal ages (embryonic day [E]) E15.5 and E18 by immunohistochemistry and beta cell fraction measurements. To distinguish between direct and indirect effects, mutant E15.5 fetal pancreata were grafted under the kidney capsule of immunodeficient mice and analysed after 1 week. E18 GR(null/null) fetuses had smaller digestive tracts and tiny pancreata. Massive pancreatic disorganisation and apoptosis were observed despite the presence of all cell types. E15.5 GR(null/null) mutants were indistinguishable from wild-type regarding pancreatic size, tissue structure and organisation, beta cell fraction and production of exocrine transcription factor Ptf1a, neurogenin 3 and Pdx-1. Grafting E15.5 GR(null/null) pancreata into a GR-expressing environment rescued the increased apoptosis and mature islets were observed, suggesting that GR(null/null) pancreatic cell death can be attributed to indirect effects of glucocorticoids on this tissue. Heterozygous GR(+/null) mutants with reduced GR numbers showed no apoptosis but increased beta cell fraction at E18 and the adult age, strengthening the importance of an accurate GR dosage on beta cell mass expansion. Our results provide evidence for GR involvement in pancreatic tissue organisation and survival through indirect effects. GR does not appear necessary for early phases, but its accurate dosage is critical to modulate beta cell mass expansion at later fetal stages, presumably through direct effects.

The critical pancreatic transcription factor Pdx1 is expressed throughout the pancreas early but enriched in insulin-producing P cells postnatally. Previous studies showed that the 5 ' conserved promoter regions areas I and II (Pdx1(PB)) direct endocrine cell expression, while an adjacent region (pdr1(XB)) containing conserved area III directs transient P-cell expression. In this study, we used Cre-mediated lineage tracing to track cells that activated these regions. Pdx(PB)Cre mediated only endocrine cell recombination, while Pdx1(XB)Cre directed broad and early recombination in the developing pancreas. Also, a reporter transgene containing areas I, II, and III was expressed throughout the embryonic day 10.5 (E10.5) pancreas and gradually became P cell enriched, similar to endogenous Pdx1. These data suggested that sequences within area III mediate early pancreas-wide Pdx1 expression. Area III contains a binding site for PTF1, a transcription factor complex essential for pancreas development. This site contributed to area III-dependent reporter gene expression in the acinar AR42J cell line, while PTF1 specifically trans-activated area III-containing reporter expression in a nonpancreatic cell line. Importantly, Ptf1a occupied sequences spanning the endogenous PTF1 site in area III of E11.5 pancreatic buds. These data strongly suggest that PTF1 is an important early activator of Pdx(1) in acinar and endocrine progenitor cells during pancreas development.