The molecular mechanism underlying Barrett’s metaplasia, the precursor to oesophageal adenocarcinoma (OAC), remains unknown. Understanding the etiopathogenesis of Barrett’s oesophagus (BO) is important for elucidating candidates for the development of biomarkers and/or novel therapies. Our previous work implicated sonic hedgehog (SHH) signalling in the pathogenesis of BO, with BMP4 and SOX9 as key downstream mediators of this process. Here, we examined the effect of BMP4, SOX9 and CDX2, which has also been implicated in the pathogenesis of BO, on the squamous oesophageal epithelium in an in vivo tissue reconstitution model.
Epithelium reconstituted from oesophageal squamous cells co-cultured with fibroblasts over-expressing BMP4 had a squamous morphology and lacked expression of columnar (CK8, CK18, CK20), gastric (MUC5AC) and intestinal (villin, A33, MUC2, CDX2, Alcian blue stain) markers. In contrast, ectopic expression of SOX9 in squamous oesophageal epithelial cells induced formation of a columnar-like epithelium and expression of CK8/18, and the intestinal specific glycoprotein, A33. Expression of CDX2 alone had no effect on the squamous epithelium. Furthermore, CDX2 did not induce any further changes to those induced by SOX9 alone when CDX2 and SOX9 were co-expressed. We also assessed expression of Mini-SOX9, a protein coding splice variant of SOX9, in BO and OAC cell lines and tissue, and found it to be progressively increased and correlated with SOX9 expression.
Our results support a role for SHH in the development of BO through induction of SOX9, which may represent an early event in the metaplastic process. In contrast, neither BMP4 nor CDX2 alone are sufficient for columnar differentiation. Our results do not rule out CDX2 having a role later in the development of BO. Finally, up-regulation of Mini-SOX9, shown to antagonize tumour suppressive functions of SOX9 in colon cancer, suggests this gene may have additional roles in the progression of BO.