ORIGINAL ARTICLE Visualizing expression patterns of Shh and Foxf1 genes in the foregut and lung buds by optical projection tomography Hideaki Sato Paula Murphy Shay Giles John Bannigan Hajime Takayasu Prem Puri Published online: 26 October 2007 Ó Springer-Verlag 2007 Abstract Congenital malformations of the foregut are common in humans. The respiratory and digestive tubes are both derived by division of the foregut primordium. Sonic hedgehog (Shh) and Fork head box F1 (Foxf1) genes encode regulatory molecules that play a pivotal role in gut and lung morphogenesis and are therefore impor- tant candidate genes to be examined in models of foregut developmental disruption. Optical projection tomography (OPT) is a new, rapid and non-invasive technique for three-dimensional (3D) imaging of small biological tissue specimens that allows visualization of the tissue distri- bution of RNA in developing organs while also recording morphology. To explore the application of OPT in this context, we visualized Shh and Foxf1 gene expression patterns in the mouse foregut and lung buds at several stages of development. Time-mated CBA/Ca mice were harvested on embryonic days 9–12. The embryos were stained following whole mount in situ hybridization with labelled RNA probes to detect Shh and Foxf1 transcripts at each stage. The embryos were scanned by OPT to obtain 3D representations of gene expression domains in the context of the changing morphology of the embryo. OPT analysis of Shh and Foxf1 expression in the foregut and lung buds revealed extra details of the patterns not previously reported, particularly in the case of Foxf1 where gene expression was revealed in a changing pattern in the mesenchyme around the developing lung. Shh expression was also revealed in the epithelium of the lung bud itself. Both genes were detected in complementary patterns in the developing bronchi as late as E12, showing successful penetration of molecular probes and imaging at later stages. OPT is a valuable tool for revealing gene expression in an anatomical context even in internal tis- sues like the foregut and lung buds across stages of development, at least until E12. This provides the possi- bility of visualizing altered gene expression in an in vivo context in genetic or teratogenic models of congenital malformations. Keywords Oesophageal atresia Á Foregut Á Sonic hedgehog Á Foxf1 Á Optical projection tomography Introduction Congenital malformations of the foregut such as the oesophageal atresia/tracheo-oesophageal fistula (OA/TOF) are common in humans. Until recently, little was known about the pathogenesis of these anomalies and there was little opportunity to study them. The accidental finding that the anthracycline antibiotic Adriamycin has teratogenic effects on rats, producing tracheo-oesophageal malforma- tions has provided a reproducible model [1]. This Adriamycin rat model has contributed to investigating the mechanisms of OA/TOF congenital anomalies [2–4]. The mouse is the foremost mammalian model of development, offering an expanding wealth of genetic and molecular H. Sato Á H. Takayasu Á P. Puri (&) Children’s Research Centre, Our Lady’s Children’s Hospital, Crumlin, Dublin, Ireland e-mail: prem.puri@ucd.ie P. Murphy Department of Zoology, University of Dublin, Trinity College, Dublin 2, Ireland S. Giles Á J. Bannigan School of Medicine and Medical Science, University College Dublin, Belfield, Dublin, Ireland 123 Pediatr Surg Int (2008) 24:3–11 DOI 10.1007/s00383-007-2036-1