The present study demonstrates that glucose enhanced, in a dose-dependent manner, the expression of the DC scavenger receptors SR-A, CD36 and LOX-1, both at the mRNA and protein levels. This phenomenon was suppressed by a p38 MAPK inhibitor, an NF-κB inhibitor and an antioxidant. Our observation that high glucose enhanced DC ROS generation provides strong evidence for a crucial role of ROS in high glucose-induced scavenger receptor upregulation. We also found that the antioxidant and p38 MAPK inhibitor blocked glucose-induced p38 MAPK activation. Taken together, these results indicate that the increased production of intracellular ROS and the activation of p38 MAPK pathways are initial signalling events in the regulation of scavenger receptor expression by glucose that are required for subsequent activation of NF-κB. We also found that glucose promoted DCs’ ability to take up oxLDL. This capability was partially blocked by neutralising antibodies against CD36 and SR-A, but not LOX-1. Moreover, high glucose induced a proinflammatory cytokine profile in human DCs and triggered DC maturation. To our knowledge, this is the first study showing these important effects of glucose on DCs.
Cardiovascular diseases in patients with type 2 diabetes are a large and increasing health problem. Unfortunately, the exactly mechanisms leading to atherosclerosis in diabetes are only partially understood. Quite recently, Skov V et al.  reported that dysregulated gene interactions and pathways in the cells of the arterial wall in diabetes may play important roles in the arterial response to injury and the sequential inflammation. Insulin resistance and hyperglycaemia in type 2 diabetes mellitus are associated with a systemic proinflammatory state that promotes the development of atherosclerosis . Our recent in vitro studies have shown that advanced glycosylation end products (AGEs)  and hyperinsulinaemia  enhance DC maturation and induce antigen-specific T-cell activation. DCs and oxLDL accumulate in progressive atherosclerotic plaque. In the subendothelial space, DCs phagocytose antigens such as oxLDL. Then, DCs can either accumulate in the arterial wall and/or migrate to draining lymph nodes for antigen presentation . Lipid and oxLDL uptake might result in the enhanced presentation of lipid and peptide antigens to NKT and T-cells. Furthermore, some foam cells can derive from DCs, and the oxLDL uptake capacity of DCs is reflected by the expression of SR-A, CD36 and LOX-1 . The engulfment of lipid by resident intimal DCs and differentiation into foam cells is possibly one of the earliest steps of atherogenesis . In addition, CD36 mediates oxLDL-induced TLR4/TLR6 activation . oxLDL stimulates DCs by binding to CD36 and TLR4, leading to DC activation, which can be accompanied by enhanced cytokine production . The present study shows that high glucose can mediate upregulation of SR-A, CD36 and LOX-1, which could be related to initiation and progression of atherosclerosis in diabetes patients.
High glucose induced an increased endocytic function of DCs, which was determined by a semiquantitative method using a fluorescence-labelled ligand (DiI-OxLDL). This effect was partially but significantly inhibited by an anti-CD36 or anti-SR-A neutralising antibody, suggesting some contribution of CD36 and SR-A to atherosclerosis formation by human DCs under diabetic conditions. However, the anti-LOX-1-neutralising antibody did not show significant inhibition of the endocytosis of DiI-oxLDL by high glucose-treated DCs. The reason may be that LOX-1 is mainly an endothelial-specific scavenger receptor, and although it is also expressed on DCs, macrophages, smooth muscle cells and platelets, its expression level is relatively low in these cell types [23–26]. Therefore, it is likely that LOX-1 is not a major receptor responsible for the uptake of oxLDL in DCs. Further studies are needed to determine the exact contribution of LOX-1 to high glucose-induced upregulation of DCs in vitro. The relative contributions of CD36, SR-A and LOX-1 to the diabetes-induced enhancement of foam cell and atherosclerosis formation by DCs has to be clarified by future studies.
The effects of several inhibitors on high glucose-enhanced scavenger receptor expression were examined to determine the signalling pathway between glucose stimulation and scavenger receptor expression. High glucose-enhanced scavenger receptor expression was suppressed by the p38 MAPK inhibitor, NF-κB inhibitor and antioxidant. Hyperglycaemia-induced oxidative stress is implicated in the pathogenesis of chronic diabetic complications. Recent studies [12–14] found that ROS generated by glucose in macrophages may be key intermediates in the regulation of SR-A, CD36 and LOX-1 expression by this metabolic factor. Moreover, Farhangkhoee H et al.  found that high glucose-induced upregulation of CD36 may be involved in increased oxidative stress in microvascular endothelial cell. In this study, we found that high glucose can also enhanced ROS generation in DCs. Evidence linking glucose-induced oxidative stress with activation of p38 MAPK in macrophages supports a role for this kinase in the control of scavenger receptor expression by hyperglycaemia [4, 28]. In line with these hypotheses, we found that antioxidant and p38 MAPK inhibitor abolished glucose-induced scavenger receptor expression, implicating ROS and kinases as signalling molecules mediating this effect of glucose. Our data also indicate a role for oxidative stress in p38 MAPK activation. Finally, we found that NF-κB inhibitor suppressed glucose-induced scavenger receptor expression. So we propose that high glucose induces DCs to generate ROS and activate p38 MAPK signalling, which leads to the subsequent activation of NF-κB and finally upregulates scavenger receptor expression. But Takaki KF et al.  reported that p38 MAPK inhibitor have no effect on high glucose-induced SR-A expression in macrophages, which is different from our present study. We do not know the reason for the discrepancy at present. One possible reason is that Takaki KF et al. used monocyte-derived macrophages whereas we used monocyte-derived DCs. The difference in cell types might cause different results.
In this study, we focussed not only on the impact of glucose on scavenger receptors but also on DC activation. In doing so, we investigated the influence of glucose on DC maturation and cytokine production. CD86 and CD83 are mature markers of DCs. Co-stimulation by the ligand CD86 and its receptor CD28 is required for efficient T-cell stimulation . We found that glucose induced the upregulation of the co-stimulatory receptors CD86 and CD83, which supports the hypothesis that T-cells are activated by DCs in plaque lesions . Consistent with this study, our recent in vivo study  also showed that the expression of CD86 was significantly increased in diabetes patients with unstable angina pectoris(UAP), which indicated that the functional status of DCs in diabetic patients with UAP were more mature and activated than none diabetic patients with UAP. The inflammatory activation of DCs is reflected by the differentially expressed cytokine-chemokine spectrum induced by glucose. We discovered that the pro-atherosclerotic chemokines IL-6 and IL-12 are induced by high glucose, while the release of anti-atherosclerotic IL-10 is reduced. IL-10 may have therapeutic potential in various inflammatory diseases, including atherosclerosis, as it can inhibit oxLDL-induced foam cell formation and apoptosis in macrophages and endothelial cells. The inhibitory effect of IL-10 on oxLDL-induced apoptosis was partially dependent on reduced p38 MAPK phosphorylation . IL-6 is a pro-atherogenic cytokine that is locally produced by plaque-infiltrating inflammatory cells . IL-12 enhances the ability of activated human monocytes to oxidise LDL and is able to modify the chemokine production of human vascular smooth muscle cells . IL-12 also seems to be important for both Th1 differentiation within the plaque and for overall T-cell recruitment into the plaque . Considering that DCs are most frequently observed in atherosclerotic lesions enriched with T-cells, the increased expression of costimulatory molecules on DCs and cytokine secretion induced by high glucose suggest that activation of DCs in diabetes patients is an important mediator in immuno-inflammatory process, as DCs are responsible for T-cell activation, and this cellular interaction plays a role in plaque instability and vulnerability, leading to rupture.