Effect of chondroitin sulphate in a rabbit model of atherosclerosis aggravated by chronic arthritis

Abstract

Background and purpose: Among the agents employed to manage osteoarthritis, chondroitin sulphate (CS) is a natural glycosaminoglycan with an anti-inflammatory effect on joint cells. CS might also influence the inflammatory component of atherosclerosis. Our aim was to examine the effect of CS administration on vascular injury and on markers of systemic inflammation in a rabbit model of atherosclerosis aggravated by systemic inflammation provoked by chronic antigen-induced arthritis.

Experimental approach: Atherosclerosis was induced in rabbits by maintaining them on a hyperlipidaemic diet after producing an endothelial lesion in the femoral arteries. Simultaneously, chronic arthritis was induced in these animals by repeated intraarticular injections of ovalbumin in previously immunized rabbits. A group of these rabbits were treated prophylactically with CS (100 mg kg(-1)day(-1)) and when the animals were killed, serum and peripheral blood mononuclear cells (PBMC) were isolated. Furthermore, femoral arteries and thoracic aorta were used for gene expression studies and histological examination.

Key results: CS administration reduced the concentration of the proinflammatory molecules C-reactive protein and IL-6 in serum. Likewise, CS inhibited the expression of CCL2/monocyte chemoattractant protein (MCP)-1 and cyclooxygenase (COX)-2 in PBMC, and reduced the nuclear translocation of nuclear factor-kappaB. In the femoral lesion, CS also diminished the expression of CCL2 and COX-2, as well as the ratio of the intima/media thickness. Moreover, CS decreased the percentage of rabbits with atherosclerosis and chronic arthritis that developed vascular lesions in the aorta.

Conclusions and implications: These findings suggest that CS treatment may to some extent impede the progression of atherosclerosis.

Figure 1

Schematic representation of the experimental model.

Figure 2

Effect of chondroitin sulphate (CS) administration to rabbits with atherosclerosis plus chronic arthritis on IL-6 (open columns) and C-reactive protein (solid columns) concentration in serum. ^*P<0.01 vs healthy controls; ^#P<0.05 vs NT rabbits.

Figure 3

Effect of CS on peripheral blood mononuclear cells (PBMC). COX-2 (a) and CCL2 (b) gene expression in the PBMC cells extracted at the time of death from all the experimental animals. (c) Left panel, a representative example of an electrophoretic mobility shift assay of radioactive NF-κB bound to the nuclear proteins extracted from the PBMC; right panel, densitometric analysis of EMSA experiments. ^*P<0.05 vs healthy controls; ^#P<0.05 vs NT rabbits.

Figure 4

Analysis of the femoral vascular lesions. Top, photomicrographs of representative haematoxylin-eosin stained arterial sections of femoral arteries. Representative femoral sections of NT and CS treated rabbits are shown. (a) NT rabbit; (b) CS treated rabbit (magnification × 100). (c) Intima/media thickness ratio in the arterial sections of all the groups studied. ^*P<0.05 vs healthy controls; ^#P<0.05 vs NT rabbits.

Figure 5

Macrophage detection by immunohistochemistry in sections of femoral arteries. Top, photomicrographs of arterial sections stained with the RAM11 antibody, specific for rabbit macrophages. Representative sections of NT- and CS-treated rabbits are shown: (a) NT rabbit; (b) CS-treated rabbit (magnification × 200). (c) Quantification of positive staining in the neointimal area in all the rabbits studied. ^*P<0.05 vs healthy controls.

Figure 6

COX-2 protein (a) and mRNA (b) expression in the femoral artery in the rabbits studied. (a) Top, a representative western blot of COX-2 in femoral arteries. (a) Bottom, densitometric analysis of western blot studies. (b and c) Analysis of COX-2 (b) and CCL2 (c) mRNA expression measured by real-time PCR method. ^*P<0.05 vs healthy controls; ^#P<0.05 vs NT rabbits.

Figure 7

Presence of atherosclerotic lesions in the aorta of the animals. All the aortae were examined as detailed in the methods section. (a) Haematoxylin-eosin section from a NT rabbit (magnification × 200). A complete section of the vessel is shown in each inset ( × 40). (b) Orcein staining of an aortic lesion from a NT rabbit, magnification × 100. (c) Percentage of rabbits with an atherosclerotic plaque in the aorta. ^*P<0.05 vs healthy controls; ^#P< 0.05 vs NT rabbits.