Expression of COX-2 in platelet-monocyte interactions occurs via combinatorial regulation involving adhesion and cytokine signaling

Abstract

Tight regulation of COX-2 expression is a key feature controlling eicosanoid production in atherosclerosis and other inflammatory syndromes. Adhesive interactions between platelets and monocytes occur in these conditions and deliver specific signals that trigger inflammatory gene expression. Using a cellular model of monocyte signaling induced by activated human platelets, we identified the central posttranscriptional mechanisms that regulate timing and magnitude of COX-2 expression. Tethering of monocytes to platelets and to purified P-selectin, a key adhesion molecule displayed by activated platelets, induces NF-κB activation and COX-2 promoter activity. Nevertheless, COX-2 mRNA is rapidly degraded, leading to aborted protein synthesis. Timedependent signaling of monocytes induces a second phase of transcript accumulation accompanied by COX-2 enzyme synthesis and eicosanoid production. Here, generation of IL-1β, a proinflammatory cytokine, promoted stabilization of COX-2 mRNA by silencing of the AU-rich mRNA decay element (ARE) in the 3′-untranslated region (3'UTR) of the mRNA. Consistent with observed mRNA stabilization, activated platelets or IL-1β treatment induced cytoplasmic accumulation and enhanced ARE binding of the mRNA stability factor HuR in monocytes. These findings demonstrate that activated platelets induce COX-2 synthesis in monocytes by combinatorial signaling to transcriptional and posttranscriptional checkpoints. These checkpoints may be altered in disease and therefore useful as targets for antiinflammatory intervention.

Pharmacological Inhibition of Platelet-Tumor Cell Cross-Talk Prevents Platelet-Induced Overexpression of Cyclooxygenase-2 in HT29 Human Colon Carcinoma Cells

“…A possible mechanism to explain the discrepancy between mRNA accumulation and protein synthesis of COX-2 is that platelets switched on a stabilization pathway of COX-2 mRNA (Dixon et al, 2006). This hypothesis was addressed by comparing COX-2 mRNA levels in HT29 cells cocultured with platelets for 20 hours versus 4 hours or in HT29 cells cultured alone in the presence of actinomycin D to stop transcription.…”

“…HT29 cells (0.5 Â10 6 cells) were cultured alone or with platelets (0.5 Â 10 8 ) for 2, 4, and 20 hours; then culture media were collected and cells were washed twice with PBS and fixed in cold acetone/methanol solution (40:60) for 20 minutes at room temperature. COX-2, COX-1, HuR, and GAPDH immunostaining were detected as previously reported (Dixon et al, 2006), and a detailed description is reported in Supplemental Methods.…”

“…COX-2 mediates these effects by enhancing the generation of prostaglandin E 2 (PGE 2 ), which triggers tumorigenic signals in target cells by coupling to four subtypes of receptors, classified as EP1, EP2, EP3, and EP4 (E-series prostanoid receptors), expressed on the plasma membrane and/or the nuclear envelope (Bhattacharya et al, 1998;Regan, 2003;Cha and DuBois, 2007). COX-2 overexpression, in both cancer and inflammation, is associated with altered expression and cytoplasmic accumulation of transacting factors that bind to adenylate-uridylate-rich elements of COX-2 mRNA and influence its stability, such as the mRNA-stability factor human antigen R (HuR) (Dixon et al, 2006;Young et al, 2009).…”

Mannan and peptidoglycan induce COX‐2 protein in human PMN via the mammalian target of rapamycin

“…Interestingly, a recent report using a preparation of platelets and monocytes has shown that efficient induction of COX-2 protein requires combination of signals involving activation of jB-driven transcriptional activation and stabilization of COX-2 mRNA by silencing the AU-rich mRNA element [51]. Whereas cells with a lasting life span might need combination of mechanisms for a timely regulation of COX-2 protein expression, the existence of rapid regulatory mechanisms in short-lived cells such as PMN seems most appropriate.…”

Platelet-mediated adhesion facilitates leukocyte sequestration in hypoxia-reoxygenated microvessels

“…Such circumstances can trigger the conformational changes of integrins and further augment the adhesive capacity of leukocytes (da Costa Martins et al, 2006;Piccardoni et al, 2001). The binding of P-selectin to PSGL-1 could also induce a series of inflammatory responses, including nuclear translocation of NF-B and the gene expression of various inflammatory cytokines, which is essential for the inflammatory phenotype acquisition of leukocytes (Dixon et al, 2006;Weyrich et al, 1996). The processes of platelet interaction with ECs and leukocytes are complicated and involve a great deal of conformational and functional alterations, which may contribute to the augmented leukocyte adhesion to ECs.…”