Cyclooxygenase (COX)-2-derived prostanoids can influence several processes that are linked to carcinogenesis. We aimed to address the hypothesis that platelets contribute to aberrant COX-2 expression ...in HT29 colon carcinoma cells and to reveal the role of platelet-induced COX-2 on the expression of proteins involved in malignancy and marker genes of epithelial-mesenchymal transition (EMT). Human platelets cocultured with HT29 cells rapidly adhered to cancer cells and induced COX-2 mRNA expression, but not protein synthesis, which required the late release of platelet-derived growth factor and COX-2 mRNA stabilization. Platelet-induced COX-2-dependent prostaglandin E2 (PGE2) synthesis in HT29 cells was involved in the downregulation of p21(WAF1/CIP1) and the upregulation of cyclinB1 since these effects were prevented by rofecoxib (a selective COX-2 inhibitor) and rescued by exogenous PGE2. Galectin-3, which is highly expressed in HT29 cells, is unique among galectins because it contains a collagen-like domain. Thus, we studied the role of galectin-3 and platelet collagen receptors in platelet-induced COX-2 overexpression. Inhibitors of galectin-3 function (β-lactose, a dominant-negative form of galectin-3, Gal-3C, and anti-galectin-3 antibody M3/38) or collagen receptor-mediated platelet adhesion (revacept, a dimeric platelet collagen receptor GPVI-Fc) prevented aberrant COX-2 expression. Inhibition of platelet-cancer cell interaction by revacept was more effective than rofecoxib in preventing platelet-induced mRNA changes of EMT markers, suggesting that direct cell-cell contact and aberrant COX-2 expression synergistically induced gene expression modifications associated with EMT. In conclusion, our findings provide the rationale for testing blockers of collagen binding sites, such as revacept, and galectin-3 inhibitors in the prevention of colon cancer metastasis in animal models, followed by studies in patients.
3-(5-Chlorofuran-2-yl)-5-methyl-4-phenylisoxazole (P6), a known selective cyclooxygenase-1 (COX-1) inhibitor, was used to design a new series of 3,4-diarylisoxazoles in order to improve its ...biochemical COX-1 selectivity and antiplatelet efficacy. Structure–activity relationships were studied using human whole blood assays for COX-1 and COX-2 inhibition in vitro, and results showed that the simultaneous presence of 5-methyl (or -CF3), 4-phenyl, and 5-chloro(-bromo or -methyl)furan-2-yl groups on the isoxazole core was essential for their selectivity toward COX-1. 3g, 3s, 3d were potent and selective COX-1 inhibitors that affected platelet aggregation in vitro through the inhibition of COX-1-dependent thromboxane (TX) A2. Moreover, we characterized their kinetics of COX-1 inhibition. 3g, 3s, and 3d were more potent inhibitors of platelet COX-1 and aggregation than P6 (named 6) for their tighter binding to the enzyme. The pharmacological results were supported by docking simulations. The oral administration of 3d to mice translated into preferential inhibition of platelet-derived TXA2 over protective vascular-derived prostacyclin (PGI2).
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•In treated HIV, cardiovascular disease is a leading cause of morbidity and mortality, the prevention of which is not fully known.•Activated platelets are causal in atherosclerotic ...cardiovascular disease, and platelets in HIV exhibit heightened activity.•In this study, clopidogrel, as opposed to aspirin, reduced platelet activation, and platelet-induced endothelial inflammation in persons with HIV.•The use of clopidogrel for the primary prevention of cardiovascular disease in HIV should be evaluated in more extensive clinical trials.
Patients with HIV exhibit platelet activation and increased risk of cardiovascular disease, the prevention of which is not fully known. Fifty-five HIV-positive patients were randomized to clopidogrel, aspirin, or no-treatment for 14 days, and the platelet phenotype and ability to induce endothelial inflammation assessed. Clopidogrel as opposed to aspirin and no-treatment reduced platelet activation (P-selectin and PAC-1 expression). Compared with baseline, platelet-induced proinflammatory transcript expression of cultured endothelial cells were reduced in those assigned to clopidogrel, with no change in the aspirin and no-treatment arms. In HIV, clinical trials of clopidogrel to prevent cardiovascular disease are warranted. (Antiplatelet Therapy in HIV; NCT02559414)
Cardiovascular risk factors contribute to enhanced oxidative stress which leads to endothelial dysfunction. These events trigger platelet activation and their interaction with leukocytes and ...endothelial cells, thus contributing to the induction of chronic inflammatory processes at the vascular wall and to the development of atherosclerotic lesions and atherothrombosis. In this scenario, endogenous antioxidant pathways are induced to restrain the development of vascular disease. In the present paper, we will discuss the role of heme oxygenase (HO)-1 which is an enzyme of the heme catabolism and cleaves heme to form biliverdin and carbon monoxide (CO). Biliverdin is reduced enzymatically to the potent antioxidant bilirubin. Recent evidence supports the involvement of HO-1 in the antioxidant and antiinflammatory effect of cyclooxygenase(COX)-2-dependent prostacyclin in the vasculature. Moreover, the role of HO-1 in estrogen vasoprotection is emerging. Finally, possible strategies to develop novel therapeutics against cardiovascular disease by targeting the induction of HO-1 will be discussed.
Systemic lupus erythematosus (SLE) is associated with the premature development of cardiovascular disease. The platelet-endothelium interaction is important in the pathogenesis of cardiovascular ...disease. In this study, we investigated the platelet phenotype from patients with SLE and matched controls, and their effect on endothelial cells.
Platelet aggregability was measured in 54 SLE subjects off antiplatelet therapy (mean age 40.1±12.8 years; 82% female; 37% white) with age- and sex-matched controls. Platelets were coincubated with human umbilical vein endothelial cells (HUVECs) and changes to gene expression assessed by an RNA array and quantitative reverse transcription polymerase chain reaction. SLE disease activity index ranged from 0 to 22 (mean 5.1±3.9). Compared with controls, patients with SLE had significantly increased monocyte and leukocyte-platelet aggregation and platelet aggregation in response to submaximal agonist stimulation. An agnostic microarray of HUVECs cocultured with SLE platelets found a platelet-mediated effect on endothelial gene pathways involved in cell activation. Sera from SLE versus control subjects significantly increased (1) activation of control platelets; (2) platelet adhesion to HUVECs; (3) platelet-induced HUVEC gene expression of interleukin-8, and intercellular adhesion molecule 1; and (4) proinflammatory gene expression in HUVECs, mediated by interleukin-1β-dependent pathway. Incubation of SLE-activated platelets with an interleukin-1β-neutralizing antibody or HUVECs pretreated with interleukin-1 receptor antibodies attenuated the platelet-mediated activation of endothelial cells.
Platelet activity measurements and subsequent interleukin-1β-dependent activation of the endothelium are increased in subjects with SLE. Platelet-endothelial interactions may play a role in the pathogenesis of cardiovascular disease in patients with SLE.
Patients with HIV exhibit platelet activation and increased risk of cardiovascular disease, the prevention of which is not fully known. Fifty-five HIV-positive patients were randomized to ...clopidogrel, aspirin, or no-treatment for 14 days, and the platelet phenotype and ability to induce endothelial inflammation assessed. Clopidogrel as opposed to aspirin and no-treatment reduced platelet activation (P-selectin and PAC-1 expression). Compared with baseline, platelet-induced proinflammatory transcript expression of cultured endothelial cells were reduced in those assigned to clopidogrel, with no change in the aspirin and no-treatment arms. In HIV, clinical trials of clopidogrel to prevent cardiovascular disease are warranted. (Antiplatelet Therapy in HIV; NCT02559414).
Background and Purpose
Hyperglycaemic memory describes the progression of diabetic complications during subsequent periods of improved glycaemia. We addressed the hypothesis that transient ...hyperglycaemia causes aberrant COX‐2 expression in HUVEC in response to IL‐1β through the induction of long‐lasting epigenetic changes involving microRNA‐16 (miR‐16), a post‐transcriptional modulator of COX‐2 expression.
Experimental Approach Studies were performed on HUVEC collected from women with gestational diabetes mellitus (GDM) (dHUVEC) and normal women (nHUVEC).
Key Results
In dHUVEC treated with IL‐1β, the expression of COX‐2 mRNA and protein was enhanced and generation of prostanoids increased (the most abundant was the promitogenic PGF2α). COX‐2 mRNA was more stable in dHUVEC and this was associated with miR‐16 down‐regulation and c‐Myc induction (a suppressor of miR expression). dHUVEC showed increased proliferation in response to IL‐1β, which was prevented by a COX‐2 inhibitor and PGF2α receptor antagonist. Comparable changes in COX‐2 mRNA, miR‐16 and c‐Myc detected in dHUVEC were produced in nHUVEC exposed to transient high glucose and then stimulated with IL‐1β under physiological glucose levels; superoxide anion production was enhanced under these experimental conditions.
Conclusions and Implications
Our results describe a possible mechanism operating in GDM that links the enhanced superoxide anion production and epigenetic changes, associated with hyperglycaemic memory, to endothelial dysfunction through dysregulated post‐transcriptional control of COX‐2 gene expression in response to inflammatory stimuli. The association of conventional therapy for glycaemic control with agents affecting inflammatory responses and oxidative stress might lead to a more effective prevention of the complications associated with GDM.
► Premenopausal women are less susceptible to cardiovascular disease than males. ► Estrogen modulates PGI2 generation via “non-genomic” and “genomic” mechanisms. ► HO-1 exerts antioxidant and ...anti-inflammatory actions through its products. ► 17β-Estradiol may restrain oxidant stress in the endothelium by inducing HO-1. ► HO-1 induction by 17β-estradiol did not involve PGI2 signaling.
We studied the effects of 17β-estradiol (E2) (10, 40nM) on 2 vasoprotective pathways, i.e. cyclooxygenase-2 (COX-2)-dependent prostanoids and the antioxidant heme oxygenase-1 (HO-1), in human umbilical vein endothelial cells (HUVEC) exposed for 6h to steady laminar shear stress (LSS, 10dyn/cm2), characteristic of atherosclerotic lesion-protected areas. COX-2 was induced by LSS versus static condition (SC). E2 did not significantly affect COX-2 expression in HUVEC cultured in SC or exposed to LSS. Prostacyclin (PGI2) and prostaglandin (PG)E2 were induced while PGF2α was reduced by LSS. E2 caused no effect or a small reduction of prostanoid biosynthesis. In HUVEC cultured in SC or exposed to LSS, E210nM caused a comparable HO-1 induction (35–45%) while E2 40nM was 5-fold more potent in LSS-exposed HUVEC than in SC (290% and 58%, respectively). PGI2 receptor antagonist RO3244794 did not affect HO-1 induction by E2. In conclusion, E2 may restrain oxidant stress in the endothelium through HO-1 induction by a mechanism independent on PGI2 signaling.
•HUVEC exposed to a GDM milieu in vivo has reduced migratory capacity.•Dysregulated expression of TGF-β receptors is detected.•Human fetal endothelium of GDM women responds abnormally to ...TGF-β1.•Several genes involved in development, cell movement and migration were downregulated.•Altered transcriptome may affect cardiac morphogenesis and placental development.
Enhanced biosynthesis of several cytokines, such as, transforming growth factor-β1 (TGF-β1), is detected in gestational diabetes mellitus (GDM). In this study, we addressed the question of whether the exposure to the abnormal milieu of GDM in vivo affects gene expression pattern of human umbilical vein endothelial cells (HUVEC) in response to TGF-β1. We found that HUVEC isolated from GDM (dHUVEC) had reduced migratory capacity versus those of healthy women (nHUVEC) and this quiescent phenotype was associated with higher expression levels of the TGF-βtype I receptor ALK5 and a slight increase in the endogenous production of TGF-β1 (mainly in its latent form). Moreover, we performed transcriptome analysis, using microarray technology, of dHUVEC versus nHUVEC, after 3h treatment with exogenous TGF-β1 (10ng/ml). The treatment of dHUVEC with TGF-β1 caused downregulation of the transcription of multiple genes involved in development, cell movement and migration of cells versus TGF-β1-treated nHUVEC. These changes in transcriptome profile might contribute to GDM-dependent alterations in cardiac morphogenesis and placental development.
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