An association of gallbladder carcinoma with cholelithiasis suggests that chronic inflammation may modulate tumorigenesis and/or progression of the carcinoma. An enhanced expression of cyclooxygenase-2 (COX-2) is observed frequently in advanced carcinomas of gastrointestinal tracts, which in turn suggests that potentiated arachidonate metabolism may play a crucial role in tumor biology. In the present study, the expression levels of COX-2 and prostaglandin E receptor subtypes were determined in 16 cases of gallbladder carcinomas of different depths of invasion (pT(1) 3, pT(2) 2, pT(3) 4, and pT(4) 7) to determine the role of arachidonate metabolism in tumor growth and progression. The mRNA levels of COX-2 were increased significantly in pT(3) and pT(4) carcinomas compared with the levels in pT(1) and pT(2) carcinomas. Immunohistochemistry and in situ hybridization revealed the existence of COX-2 mRNA and protein in both the cancerous epithelia and adjacent stroma of pT(1)-pT(4) carcinomas. Only in pT(3) and pT(4) carcinomas was intense expression of COX-2 observed in the adjacent stroma. The tissue concentration of PGE(2) was significantly increased in pT(3) and pT(4) carcinomas. The mRNAs of PGE receptor subtypes EP(2), EP(3), and EP(4) were amplified in pT(1)-pT(4) gallbladder carcinomas, in which their mRNAs and EP(4) protein were expressed mostly in the cancerous epithelia. Treatment with a specific EP(4) agonist, as well as PGE(2) but not EP(2) and EP(3) agonists, up-regulated the expression of c-fos, an induced growth response gene, and increased colony formation. In advanced gallbladder carcinoma, enhanced expression of COX-2 is observed in the adjacent stroma rather than in the cancerous epithelia, and the stroma is a potent source of PG synthesis. In epithelial-stromal interactions, the increased PGE(2) synthesis in the adjacent stroma and its biological effect via EP(4) on the carcinoma cells may contribute to tumor growth and progression of gallbladder carcinoma.