Aims/hypothesis  

We evaluated whether hyperinsulinaemia stimulates the expression of transcription factor CCAAT/enhancer binding protein (C/EBP)-β and C/EBP-δ and leads to the induction of the chemokine (C-C motif) ligand 2 gene (Ccl2, also known as MCP-1) expression in aortas.

Methods  

Hyperinsulinaemia was induced by feeding rats a high-fructose diet. CCL2 production was analysed by ELISA. The expression of Ccl2, Cebpb and Cebpd mRNAs was investigated by quantitative RT-PCR. The binding of C/EBP-β to Ccl2 was assessed by chromatin immunoprecipitation (ChIP) assay.

Results  

Insulin at a concentration of 10 nmol/l significantly stimulated the expression of Cebpb, Cebpd and Ccl2 mRNAs, depending on activation of phosphatidylinositol 3-kinase (PI3K) in cultured vascular smooth muscle cells. The knock-down of C/EBP-β with siRNA abolished the insulin-induced Ccl2 mRNA expression. In the aortas from fructose-fed rats, the levels of phosphorylation of Akt/protein kinase B, a downstream effector of PI3K, were also increased. The expression of Cebpb, Cebpd and Ccl2 mRNAs in the aortas from fructose-fed rats were significantly elevated, by 330, 300 and 300%, respectively, compared with those of control-fed rats. The induction Ccl2 mRNA expression in the aortas was significantly correlated with the expression of Cebpb and Cebpd mRNAs in the aortas. Furthermore, the ChIP assay showed elevated binding of C/EBP-β to the 5′ upstream region of Ccl2 in the aortas from fructose-fed rats.

Conclusions/interpretation  

These findings clearly indicate the role of C/EBPs in the mechanism of upregulation of CCL2, an inflammation-related protein, observed in the hyperinsulinaemic state, which may initiate the process of atherosclerosis.