Endothelial cells (ECs) are crucial in maintaining vascular homeostasis. Endothelial dysfunction was involved in many cardiovascular diseases (CVDs). Recently, antimalarial medicine artemisinin and its derivatives including dihydroartemisinin (DHA) were found to be beneficial in some diseases including CVDs. Prostaglandin (PG) E₂ is a known inflammatory mediator and plays important roles in cardiovascular system. This study was to investigate the role of DHA in regulating cyclooxygenase (COX)/PGE synthase (PGES)/PGE₂ cascade and inflammation in ECs. After DHA treatment, the mRNA and protein levels of COX-2 were strikingly upregulated in time- and dose-dependent manners. In contrast, COX-1 was significantly downregulated. As expected, inhibition of COX-1 or COX-2 further reduced PGE₂ production after DHA treatment. Moreover, DHA enhanced microsomal PGE₂ synthase (mPGES)-2 and moderately modulated cytosolic PGE₂ synthase (cPGES) with no effect on mPGES-1 expression. Importantly, DHA significantly reduced PGE₂ levels in line with the upregulation of 15-hydroxyprostaglandin dehydrogenase (15-PGDH, a key enzyme for prostaglandin degradation). Lastly, we observed that DHA not only reduced the PGE₂ levels in tumor necrosis factor-α (TNF-α)-treated ECs but also blunted the upregulation of inflammatory cytokines of interleukin (IL)-6 and IL-1β induced by TNF-α or PGE₂. These findings demonstrated an important role of DHA in regulating PGE₂ synthesis cascade and inflammation in ECs, suggesting a potential of DHA for the treatment of inflammatory vascular diseases.