CDX2 hox gene product in a rat model of esophageal cancer

In the homeobox gene family, the caudal-related CDX2 gene encodes for an intestine-specific transcription factor involved in both cell turnover and intestinal differentiation [1]. Nuclear immunostain for Cdx2 is restricted to the native intestinal epithelia and its de novo expression is considered as suitable marker of a newly achieved intestinal commitment [2, 3].

Barrett's esophagus (BE) is defined as replacement of the native esophageal squamous epithelium by columnar (intestinalized) mucosa [4‐6]. Longstanding exposure of the squamous esophageal epithelium to gastric reflux is a primary risk factor for columnar metaplasia, which is consistently considered as precursor of esophageal adenocarcinoma (Ac) [7, 8].

Esophageal Ac is the final step in a sequence of phenotypic changes that include long-standing esophagitis, columnar cell metaplasia, and non-invasive neoplasia (NiN). The molecular derangements occurring in each of these phenotypic changes are largely unknown and they involve both genetic and chromosomal instability [9, 10]. More information on such molecular changes is crucial in any strategy of primary prevention of Barrett's Ac [11‐14].

In humans, both practical and ethical limitations prevent any sequential exploration of the cascade of Barrett's Ac, so experimental models are used to characterize the biological alterations leading to neoplastic transformation [15‐31].

In this experimental study, the expression of Cdx2 protein was tested over the whole spectrum of phenotypic lesions detected in a surgical murine model of esophago-gastroduodenal anastomosis (EGDA) resulting in longstanding esophageal reflux of gastro-duodenal contents [19, 21‐24, 29].

Gastro-esophageal reflux is generally considered the main promoter of esophageal columnar metaplasia and adenocarcinoma.

Cdx2 is a transcription factor that regulates the expression of differentiation-related molecules and it is specifically involved in intestinal cells commitment. Based on this rationale, Cdx2 immunohistochemical expression was explored in a rat model of EGDA.

As in previous studies, de novo Cdx2 expression was documented in the whole spectrum of phenotypic changes induced by experimental EGDA. The prevalence of Cdx2 expression increased significantly with time (i.e. the prevalence of IM and MLE was higher in Groups B and C than in Group A), suggesting a time-dependent relationship between the "chemical" injury and the severity of the lesions.

Cdx2 expression in full-blown metaplastic transformation was expected. This study, however, also showed that de novo Cdx2 expression is an early event among the morphological changes caused by the refluxate. The early deregulation of Cdx2 expression has already been demonstrated by Pera et al. [28], who described Cdx2 immunostaining in the basal cell layer close to esophageal ulcers 16 weeks after surgery. More recently, however, in a study using a similar EGDA model, Xiaoxin Chen et al. [17] considered Cdx2 over-expression as a late marker of the metaplastic cascade.

Our study provides evidence that "protometaplastic" changes (in both the squamous stem cell and MLE) could be revealed by Cdx2 immunostaining even before the IM becomes histologically assessable. It is worth noting that MLE (which can also be a feature of normal rat mucosa) might be considered as a "partially-committed" cell population, prone to a chimeric intestinal differentiation under critical conditions (such as those produced by EGDA). Such speculations might also apply to the staminal cells compartment of the native esophageal mucosa: in cultured esophageal epithelia, in fact, chemical injuries (acid and/or bile components) may result in Cdx2 promoter demethylation/activation [33]. These hypotheses are further supported by the finding that no Cdx2 expression was detected in squamous epithelia (far from esophageal ulcers/metaplastic changes), nor in any of the 4 cases of SCC.

Together with Cdx2, also other intestine-specific transcription factors have been described as involved in Barrett's epithelium development [34‐36]. In a similar rat model, Kazumori et al. [36] showed, that a de novo expression of Cdx1 (another member of the caudal-related homeobox gene family) significantly antecedes Cdx2 expression [35, 36]. Further studies are needed to investigate on the interplay of these two genes in the morphogenesis of Barrett's mucosa.

The SCC cases detected in this study prompts us to hypothesize that the environmental conditions resulting from EGDA may also result into the derangement of cell regulatory mechanisms involving both multilayered and squamous epithelia. Previous studies documented that several transcription factors (p63, among others) are over-expressed in squamous esophageal epithelia after EGDA. Such an observation could explain, at least in part, the high prevalence of SCC documented in this and other studies.