Résumé
Abstract only Aim: Leukotrienes are pro-inflammatory mediators that are locally produced in coronary atherosclerotic plaques. This study aimed to elucidate cysteinyl leukotriene (CysLT) signaling in vascular smooth muscle cells (SMCs), and the effects of inflammation on this process. Hypothesis: We tested the hypotheses that a) the CysLT 1 receptor is upregulated in vascular SMCs in the context of atherosclerosis, and that b) CysLT 1 receptor activation is coupled to nuclear calcium signaling, which c) leads to alterations in gene expression. Methods & Results: Immunohistochemical analyses revealed that CysLT 1 receptor expression co-localized with α-smooth muscle actin in human carotid endarterectomy samples, and that the CysLT 1 receptor exhibited a perinuclear expression in human coronary artery SMCs. Real time quantitative PCR demonstrated that lipopolysaccharide (LPS) significantly upregulated CysLT 1 receptor mRNA levels in a time-dependent fashion. Fluorescent calcium-signaling experiments showed that LPS significantly enhanced the changes in intracellular calcium induced by leukotriene C 4 (LTC 4 ) in these cells. LTC 4 stimulation predominantly enhanced nuclear calcium increase, and this response was inhibited by the CysLT 1 receptor-antagonist MK-571. Microarray analysis revealed that after 24h stimulation of human coronary artery SMCs with LTC 4 , there was a 5-fold increase mRNA levels for Plasminogen Activator Inhibitor (PAI)-2, among a number of significantly upregulated genes. The LTC 4 -induced increase in PAI-2 expression was confirmed by real time quantitative PCR and ELISA, and was inhibited both by MK-571 and by calcium chelators. Conclusions: Pro-inflammatory stimulation of vascular SMCs upregulated CysLT 1 receptor expression with perinuclear localization. CysLT 1 receptor expression was coupled to nuclear calcium signaling and changes in gene expression, such as upregulation of PAI-2. Together, these findings suggest a role of nuclear CysLT 1 receptor signaling in vascular SMCs including gene expression patterns associated with atherosclerosis.