Expression and regulation of cyclooxygenase-2 in normal and neoplastic canine keratinocytes

Squamous cell carcinoma (SCC) is one of the most common cancers in dogs, yet relatively little is known about the molecular events involved in its development. Increasing evidence implicates cyclooxygenase-2 (COX-2) in the pathogenesis of various cancers in humans and animals. COX-2 overexpression has recently been demonstrated in canine SCCs. The objective of our study was to characterize the expression and regulation of COX-2 in normal and neoplastic canine keratinocytes (CKs) to provide an in vitro system to investigate the implication of COX-2 in SCC oncogenesis in dogs. Cell lines derived from normal CKs and neoplastic CKs (SCCs) were cultured in the absence or presence of agonists, and immunoblots, immunocytochemistry, radioimmunoassays and a cell proliferation assay were used to characterize COX-2 expression and action. Results showed that neoplastic keratinocytes had a higher basal COX-2 expression than normal keratinocytes. In both cell lines, stimulation with the tumour promoter phorbol 12-myristate 13-acetate induced a time-dependent increase in COX-2 protein, with COX-2 induction being stronger in cancerous SCC than in normal CK cells. Moreover, SCC cells produced significantly more PGE(2) than CK cells, under both baseline and stimulated conditions (P < 0.05). NS-398, a selective COX-2 inhibitor, inhibited prostaglandin (PG)E(2) synthesis and decreased proliferation of CK and SCC cells (P < 0.05). Collectively, our results indicate that the canine neoplastic keratinocyte SCC cell line expresses more COX-2 and produces more PGE(2) than the normal keratinocyte CK cell line, thus providing an in vitro system to study the molecular basis of elevated COX-2 expression in SCCs in dogs.