Elsevier

Atherosclerosis

Volume 143, Issue 1, March 1999, Pages 115-123
Atherosclerosis

Monocyte chemotactic protein-1 gene and protein expression in atherogenesis of hypercholesterolemic rabbits

https://doi.org/10.1016/S0021-9150(98)00285-8Get rights and content

Abstract

Monocyte adherence to the endothelium and subsequent migration into the subendothelial space are important early events in atherogenesis. Monocyte chemotactic protein-1 (MCP-1) has been shown to be highly expressed in both human atheroma and advanced atherosclerotic lesions of experimental animals. To establish the temporal correlation between MCP-1 expression and plaque development, we examined the expression of MCP-1 during atherogenesis of hypercholesterolemic rabbits using Northern blot analysis, in situ hybridization, and immunohistochemistry. New Zealand White rabbits were fed with 2% cholesterol-containing diet for 1 day, 3 days, 1 week, 3 weeks or 6 weeks. The plasma levels of total cholesterol were significantly increased 3 days after cholesterol feeding and continued to increase during the entire cholesterol-feeding period. Northern blot analysis showed that MCP-1 mRNA levels remained unchanged following cholesterol feeding for up to 1 week, were higher than control levels at 3-week and increased even higher at 6-week. In situ hybridization showed that after 3 weeks of cholesterol feeding, MCP-1 mRNA expression was up-regulated in newly-formed fatty streaks and parts of tunica media in the presence or absence of fatty streaks. At 6-week, pronounced MCP-1 mRNA expression was detected with similar distribution. In contrast, MCP-1 mRNA was detected only in a few endothelial cells and adventitia in control and experimental groups feeding cholesterol up to 1-week. Immunostaining of serial sections indicated that MCP-1 was expressed by macrophages and smooth muscle cells in rabbits fed with cholesterol for 3 or 6 weeks. No MCP-1 was detected in intima or media in all other groups. These results show that a lag period exists between serum cholesterol increase and upregulation of MCP-1 expression, suggesting that cholesterol modifications (e.g. oxidation) are required to stimulate MCP-1 expression. In addition, MCP-1 expressed by both macrophages and smooth muscle cells during the initial stages of atherosclerosis is likely to contribute to the development of fatty streaks in hypercholesterolemic rabbits.

Section snippets

Experimental animals

Male New Zealand white rabbits (2 kg body weight) were fed with regular feed (control group) or regular feed containing 2% (wt/wt) cholesterol (Purina Mills, USA) for 1 day, 3 days, 1 week, 3 weeks or 6 weeks to induce hypercholesterolemia. During and after 6 weeks of feeding, serum total cholesterol and triglycerides were measured in blood samples collected from the ear vein. Four sets of cholesterol-feeding experiments were performed. In each set of experiments, each group consisted of two to

Plasma cholesterol and triglyceride levels

In the control group, the plasma cholesterol and triglyceride concentrations before the experiment was 0.70±0.064 g/l and 0.84±0.107 g/l, respectively (n=4). The concentrations did not change significantly during the 6-week feeding period (0.64±0.044 g/l and 0.83±0.046 g/l at 6-weeks for cholesterol and triglycerides, respectively, n=4). In the hyperlipidemic group, the plasma cholesterol concentration increased immediately 1 day after feeding cholesterol-containing chow from 0.73±0.072 g/l (n

Discussion

Hyperlipidemic rabbits fed with high-cholesterol diet are commonly used as experimental animals to examine the pathogenesis and mechanisms of atherosclerosis [33]. The atherosclerotic lesions observed in hyperlipidaemic rabbits are mainly composed of fatty streaks, similar to the type II lesions of human atherosclerosis [34]. In contrast to human atherosclerotic plaques which usually take decades to develop, the development of atherosclerosis in this animal model can be easily evaluated at

Acknowledgements

We thank Ya-Ting Chen and Shu-Feng Tsai for their assistance in taking photographs and W.L. Cheong for plotting figures. This work was supported in part by research grants from the National Science Council (NSC85-2331-B006-109M26 and NSC86-2314-B006-021M26) of ROC.

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