Keratinized squamous epithelium and down-regulated RARβ gene expression in the lung of cigarette smoke-exposed ferret after β-carotene supplementation

To determine if high-dose β-carotene (β-C) result in enhanced cell proliferation and down-regulated RARβ gene expression in the lung epithelium, we carried out an in vivo study in smoke-exposed ferrets. Male ferrets were divided into four groups: Control (C); Smoke-exposed (SM); Smoke-exposed plus β-C supplementation (SM+βC); and β-C supplementation alone (βC), and were killed after six months of treatment. The results showed that both plasma and lung β-C decreased significantly in the SM+βC group vs. the BC group. The incubation in vitro of β-C with lung post-nuclear fraction from either the SM or the C group ferret showed that smoke-exposure enhances significantly the excentric cleavage of β-C in ferret lung tissue, producing increased amounts of β-C oxidative metabolites. The pathologic and immunohistochemistry analysis showed enhanced proliferation in alveolar cells and keratinized squamous metaplasia in the SM+βC group as compared to the other three groups, which correlated with increased proliferating cellular nuclear antigen expression. Furthermore, RARβ gene expression, but not RARα, was down-regulated in the SM+βC group ferrets. These observations suggest that high-dose β-C with additional smoke-exposure causes down-regulation of RARβ expression in ferret lung which may be one of the mechanisms for enhancement of lung tumorigenesis after high-dose β-C supplementation and smoking.