Prostaglandins are involved in the regulation of several reproductive processes such as ovulation, luteolysis, and establishment of pregnancy. Prostaglandin E₂ (PGE₂) appears to favor establishment of pregnancy in most mammals studied so far. The primary enzymes involved in the production of PGE₂ from arachidonic acid are cyclooxygenases and prostaglandin E synthases (PGES). Three PGES have been identified in humans, but in the bovine, microsomal PGES2 and cytosolic PGES genes have neither been cloned nor associated to any physiological processes. The present study was undertaken to clone bovine MPGES2 and CPGES and to report on their regulation in the endometrium during the estrous cycle. CPGES mRNA expression declines progressively during the cycle; its protein is not modulated according to a precise pattern. MPGES2 mRNA and protein expression decrease from the beginning of the cycle until Days 13–15 and then increase until ovulation. Immunohistochemical analysis reveals that both enzymes are located in luminal epithelial and glandular epithelial cells and at a lower level in stromal cells. In addition, using the bovine endometrial cell line BEND, where higher accumulation of PGE₂ is observed following treatment with phorbol 12-myristate 13-actetate (PMA) and tumor necrosis factor-α (TNF-α), we have found an associated increase of MPGES1 and COX2 but not CPGES or MPGES2 protein expression. Together, our results suggest that MPGES1 is not the only PGES present in the bovine endometrium but is the main enzyme associated with increased PGE₂ production in vitro.