Abstracts
2000 Digestive Disease Week

# 2287 pbx-1, a Homeodomain Protein Transcriptional Regulator Is a Marker for Early Pancreatic Ducts and a Necessary Factor in Exocrine Differentiation.
Thomas S. Maldonado, Chris A. Crisera, Alan S. Kadison, Michael T. Longaker, George K. Gittes, Kansas City, MO, New York, NY

While various patterning control genes important in pancreatic endocrine differentiation have been identified, no such control gene has been found for pancreatic ducts. However, a homeotic regulator, pbx-1, has been shown to divert pancreatic cell lines toward exocrine phenotype, rather than endocrine. Given the important role of homeodomain proteins and, more recently, the new family of homeotic regulators, such as pbx, we hypothesized that pbx-1 may play a role in normal pancreatic growth and differentiation during organogenesis. Thus, in order to better understand its role in vivo, we wished to determine the expression of pbx-1 in developing embryonic pancreas. Next, we blocked pbx-1 expression in vitro to determine whether it was necessary for promoting normal exocrine differentiation. Mouse embryonic pancreas from gestational ages 12-18 days (E12.5 to E18.5), as well as adult pancreas, were dissected and processed for immunohistochemistry. Immunostaining for PBX-1 was performed on all specimens. Next, E11.5 whole pancreas was grown for 7 days in media containing antisense oligonucleotides in order to block transcription of pbx-1 in vitro. Mis-sense (random) oligonucleotides were used as controls. Specimens were then immunostained for glucagon, insulin, and carbonic anhydrase II (a duct-specific marker in the pancreas). Immunostaining for pbx-1 on adult and embryonic pancreases localized this transcription factor specifically to the ducts. Interestingly, in E12.5 pancreas, pbx-1 was present in the mesenchyme but not in the epithelium. Moreover, pbx-1 staining was predominantly cytoplasmic in mature ducts, but nuclear in mesenchymal cells, suggesting that the transcriptional activity of pbx was only present in the mesenchyme. E11.5 whole pancreas treated with antisense oligonucleotides to pbx-1 failed to undergo any exocrine differentiation, and stained positively for insulin and glucagon as a pure cluster of endocrine cells. These data have shown, for the first time, a patterning gene that is a duct-specific marker. In addition, the absence of any exocrine differentiation when pbx-1 expression was blocked is consistent with the notion that this homeotic regulator is essential for normal duct development. Finally, our finding of pbx-1 in early embryonic mesenchyme, localizing to cell nuclei, may explain the previous observation that mesenchyme is critical to exocrine duct development.