Objective: Homocysteine (Hcy) is known as an independent risk factor for atherosclerosis. C-reactive protein (CRP) directly participates in initiation and progression of atherosclerosis. However, there is no direct evidence to demonstrate pro-inflammatory effect of Hcy on vascular smooth muscle cells (VSMCs) through CRP. In the present study, we examined the effect of Hcy on CRP expression and investigated the related mechanism in VSMCs.
Methods and results: Protein expression and secretion were detected by Western blot and ELISA, respectively. mRNA expression was detected by RT-PCR. Superoxide anion was detected by lucigenin chemiluminometry and the immunofluorescence staining was observed by a fluorescence microscope. The results revealed that Hcy significantly induced mRNA and protein expressions of CRP in VSMCs both in vitro and in vivo, and anti-IL-1β or anti-IL-6 neutralizing antibody alone or in combination partially reduced Hcy-induced CRP expression. Hcy increased the expression of NR1 subunit of N-methyl-d-aspartate receptor (NMDAr), and MK-801 alleviated Hcy-induced CRP expression in VSMCs. Further studies showed that Hcy-stimulated superoxide anion generation in VSMCs. Nevertheless, pretreatment of the cells with MK-801, TTFA and DPI significantly reduced Hcy-stimulated superoxide anion generation, and antioxidant NAC decreased Hcy-induced CRP expression in VSMCs. Additionally, PD98059, SB205380 or PDTC antagonized Hcy-induced CRP expression, and MK-801, NAC, PD98059 or SB205380 inhibited Hcy-activated phosphorylations of ERK1/2 and p38.
Conclusion: The present study demonstrates that Hcy is able to initiate an inflammatory response in VSMCs by stimulating CRP production, which is mediated through NMDAr-ROS-ERK1/2/p38-NF-κB signal pathway. These findings provide new evidence for a role of Hcy in pathogenesis of atherosclerosis.
Keywords: Atherosclerosis; C-reactive protein; Homocysteine; Inflammation; Vascular smooth muscle cells.
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