Inhibition Of4NQO-induced Tumorigenesis In The Oral Squamous Epithelium By Dietary Calcium

Objective:Calcium is not only essential for the formation and maintenance of bones, but also plays an important role in the regulation of muscle contraction, glands secretion and cell proliferation and differentiation. We have previously demonstrated that calcium promotes keratinocyte differentiation via E-cadherin-β-catenin-p120complex mediated phospholipase C-yl (PLC-yl) activation in the plasma membrane. However, it is unknown whether dietary calcium affects tumorigenesis of keratinocytes in the squamous epithelium. To address this issue, we examined effects of dietary calcium on chemically-induced tumor formation in the squamous epithelium in mice.Methods:Sixty-six mice (C57BL/6) at four weeks old were fed with drinking water containing4-nitroquinoline1-oxide (4NQO) to induce tumor formation in the oral squamous epithelium. They were also fed with different calcium diets (the diet containing0.01%calcium,1.3%calcium or2.0%calcium) for28weeks. We first examined rates of tumors in the oral squamous epithelium. We then examined the expression levels of proliferation and differentiation of these tumors and levels of E-cadherin,β-catenin, p120, epidermal growth factor receptor (EGFR), phosphatidylinositol3-kinase enhancer (PIKE) and PLC-γ1by immunohistochemical staining, calcium levels in oral squamous epithelium by ion-capture cytochemistry, serum calcium levels by an ion-specific electrode, and intact parathyroid hormone (iPTH) by chemiluminescence.Results:The calcium level in the oral squamous epithelium was increased significantly in mice on the high calcium diet and reduced significantly in mice on the low calcium diet. Rates of oral tumors was increased significantly in mice on the low calcium diet and reduced significantly in mice on the high calcium diet. Mice on the high calcium diet had lower levels expression of proliferation and PLC-γ1and higher levels of differentiation, E-cadherin, β-catenin, p120and EGFR in the normal oral squamous epithelium. In contrast, mice on the low calcium diet had higher expression levels of proliferation and PLC-γ1and lower levels of differentiation, E-cadherin, β-catenin, p120and EGFR in the normal oral squamous epithelium. There were no differences in the levels of proliferation, differentiation, E-cadherin,(3-catenin, p120, EGFR, and PLC-γ1in the oral tumor epithelium including the epithelium of papilloma and squamous cell carcinoma as well as the expression level of PIKE in normal and tumor epithelium, in mice on different calcium diets. There were no differences in the level of serum calcium and iPTH in mice on different calcium diets. Conclusion:Dietary calcium increases calcium in oral epithelium, suppresses4NQO-induced tumor formation in the oral squamous epithelium, and the effect is likely to be mediated by inhibiting proliferation and promoting differentiation of oral squamous epithelium. Increased levels of E-cadherin,β-catenin and EGFR and decreased levels of PLC-yl may play a role in the inhibition of tumorigenesis in the oral squamous epithelium by dietary calcium.