Fibroblast growth factor signaling in the developing tracheoesophageal fistula.

BACKGROUND/PURPOSE: The Adriamycin-induced rat model of esophageal atresia and tracheoesophageal fistula (EA/TEF) provides a reliable system for the study of EA/TEF pathogenesis. The authors previously hypothesized that faulty branching lung morphogenesis pathways were a critical component of its pathogenesis. The authors have found evidence for faulty fibroblast growth factor (FGF) signaling related to epithelial-mesenchymal interactions in the fistula tract. To better define FGF signaling, the differential expression of FGF ligands and their receptors between lung, fistula tract, and esophagus are described. METHODS: Time-dated pregnant, Sprague-Dawley rats were injected with Adriamycin (2 mg/kg intraperitoneally) on days 6 through 9 of gestation. Tissues were processed for histology and reverse transcriptase polymerase chain reaction. FGF-1, -7 and -10 were measured from whole lung, fistula tract, and esophagus of TEF or normal embryos. Expression of FGF2RIIIb and FGF2RIIIc receptors was measured in isolated epithelium and mesenchyme of lung and fistula tract of TEF embryos as well as lung and esophagus from normal controls. RESULTS: FGF-1 mRNA was present in the fistula tract and normal and Adriamycin-exposed lung but absent from whole esophagus. Interestingly, FGF-7 mRNA was present only in normal lung. FGF-10 was present in all tissues examined. FGF2RIIIb mRNA was absent in fistula mesenchyme but present in all other tissues examined. However, the splice variant FGF2RIIIc mRNA was present in all tissues examined. CONCLUSIONS: These findings support defective FGF signaling in the rat model of EA/TEF. Absence of FGF-7 mRNA in Adriamycin-exposed tissues suggests the primary effect of Adriamycin may be to inhibit FGF-7 expression. Moreover, absence of FGF2RIIIb in fistula mesenchyme may be caused by loss of positive feedback from FGF-7, its normal obligate ligand. Understanding these specific defects in FGF signaling may provide insight into faulty mechanisms of EA/TEF.

Glucagon is required for early insulin-positive differentiation in the developing mouse pancreas.

The embryonic pancreas is thought to develop from pluripotent endodermal cells that give rise to endocrine and exocrine cells. A key guidance mechanism for pancreatic development has previously been found to be epithelial-mesenchymal interaction. Interactions within the epithelium, however, have not been well studied. Glucagon is the earliest peptide hormone present at appreciable levels in the developing pancreatic epithelium (embryonic day [E]-9.5 in mouse). Insulin accumulation begins slightly later (E11 in mouse), followed by a rapid accumulation during the "second wave" of insulin differentiation ( approximately E15). Here we found that blocking early expression and function of glucagon, but not GLP-1, an alternate gene product of preproglucagon mRNA, prevented insulin-positive differentiation in early embryonic (E11) pancreas. These results suggest a novel concept and a key role for glucagon in the paracrine induction of differentiation of other pancreatic components in the early embryonic pancreas.

Complete discontinuity of the distal fistula tract from the developing gut: direct histologic evidence for the mechanism of tracheoesophageal fistula formation.

The embryogenesis of tracheoesophageal anomalies remains controversial. The purpose of this study was to better define the embryogenesis of developing esophageal atresia with tracheoesophageal fistula (EA/TEF), with specific attention to the controversial issue of whether a discontinuity exists in the foregut during its development of EA/TEF. Pregnant outbred rats were injected with adriamycin (2 mg/kg i.p.) on days 6-9 of gestation (E6-E9). At E12.5 and 13.5, microdissection of the entire foregut was performed. Foreguts were examined by phase microscopy, and serial, precisely transverse sections were created for hematoxylin and eosin (H&E) staining. Gross microdissection of the developing foregut at E12.5 (n = 9) revealed a blind-ending, bulbous fistula tract arising from the middle branch of the tracheal trifurcation (as seen by direct and phase microscopy). No connection with the gut could be appreciated at E12.5, but by E13.5 (n = 10) there was an obvious connection between the fistula and the stomach. Serial H&E transverse sections also demonstrated a blind-ending fistula tract arising from the trachea at E12.5. This fistula tract was clearly discontinuous from the developing stomach, which appeared much further caudal to the end of the fistula tract. These results strongly support a model of experimental TEF wherein the fistula tract arises from a trifurcation of the trachea, and (only during a specific gestational window between days 12.5 and 13.5) there is discontinuity between the fistula tract and the stomach. By day 13.5, the fistula joins with the stomach anlage. These observations in the developing EA/TEF should help to resolve the controversy about the mechanism of EA/TEF formation.