In this study, IMA and FV had a similar gene expression pattern but very different to that seen in OPA.
Atherosclerosis is the consequence of excess lipid accumulation in vessels, which triggers an immune response, and the secretion of inflammatory cytokines that promote its development. The arteries and veins differ in that the atherosclerotic process is accentuated in large arteries where hemodynamic forces are exerted. We found significant differences between these different vessels as result of an advanced atherosclerotic process.
Lesion disruption facilitates the interaction between circulating blood and prothrombotic substances, such as tissue factor (tF) present within the atherosclerotic lesion. However, an increase of tF levels (mRNA and protein expression) was found in OPA with respect to IMA or FV, in agreement with previous studies from coronary arteries .
Our interest in inflammation as a component of CVD led us to study other additional mediators, such as the metalloproteinases (MMP), which form a family of zinc-dependent endopeptidases that degrade vascular extracellular matrix and basement membrane components playing a main role in tissue repair and vascular remodelling. It is widely accepted that plaque rupture plays a crucial role in the pathogenesis of vascular events and that atherosclerotic plaque destabilization is mediated by MMP. In the present study, we found higher MMP2 and MMP9 mRNA in OPA. These findings are consistent with previously published results . MMP2 and MMP9 are regulated by tissue inhibitors, pro-inflammatory cytokines and other factors such as oxLDL or situations of insulin resistance, where the oxidative stress is increased , and they have both pro- and anti-inflammatory actions. These actions are associated with arterial stiffness and essential hypertension . It was shown that MMP inhibitors and eNOS inhibit the smooth muscle cells migration in vitro and neointima formation in vivo. Moreover, NO was shown to attenuate gene expression associated with insulin resistance .
As expected, gene expression levels of TIMP-1 and TIMP-3, regulators of the cytoskeleton, were also increased in OPA with respect to IMA and FV. TIMP-1 and TIMP-3 are endogenous inhibitors regulated by oxLDL in vascular endothelial cells , and are compensating mechanisms to prevent the breakdown of the atheroma plaque.
Cholesterol and LDL concentrations have indubitable value as risk markers for future cardiovascular events. Recent studies have demonstrated that increased levels of oxLDL are markedly associated with MMP-9 activation, and that statins reduce inflammatory responses. The relationship between lipid metabolism and popliteal plaque has been poorly studied. In the present study, NDUFA and LRP1 mRNA expression levels were highly increased in occluded OPA. LRP1 is upregulated by cardiovascular risk factors such as hypercholesterolemia  and hypertension . Aditionally, this receptor contributes to the uptake of aggregated LDL , one of the main modifications of LDL in the arterial intima. The increase of LRP1 expression in OPA (mRNA and protein expression) suggest that LRP1 may play a crucial role in atherosclerosis progression, as previously demonstrated in other studies [20, 21].
Delivery of free fatty acids excess to peripheral tissues can worsen insulin resistance and may play a role in activating inflammatory processes through activation of toll-like receptors . TLRs are a family of pattern recognition receptors found in various inflammatory cells . TLR2 showed increased levels in popliteal artery compared with vessels without atheroma in our study.
Different biomarkers were studied in the same environment, i.e., OPA and FV biopsies from the same diabetic patient. For this reason, it is very interesting to study what happens with the biomarkers of different metabolic pathways when the patient has good or poor glycemic control. It is likely that hyperglycemia-induced intra- and extra-cellular changes lead to alterations in signal transduction pathways, affecting gene expression and protein function and causing cell dysfunction and damage. However, in patients with type 2 diabetes mellitus, pathways involved in the diffuse vasculopathy present in non atherosclerotic arterial tissue and mRNA-alterations are already established .
Our results demonstrated that patients with good glycemic control had greater SREBP2 expression levels in FV. Hyperinsulinemia is related to an up-regulation of SREBPs , which could conflict with our results, but we previously showed that SREBP2 controls the expression of some LDL receptor genes, such as CD36 gene expression. A strong relation between SREBP2 and CD36 was found in FV but not in OPA . Sampson et al.  showed that diabetic subjects with good glycemic control had higher CD36 expression, which could reflect a post-transcriptional efficiency of this receptor and thus there would be greater metabolism of oxLDL in these patients.
Our patients with good glycemic control showed an increased expression from genes involved in protection against apoptosis and cell turnover. Our data agree with the results recently published by Redondo et al. (2011), who demonstrated that there is a link between inflammation (COX-2) and apoptosis resistance (BCL2) in the vessels of diabetic patients .
Some drugs (e.g., metformin, thiazolidinediones and statins) used in the treatment of diabetes and atherosclerosis are able to up-regulate both processes. These may exert their protective effects through activation of AMPK which has potentially beneficial anti-atherosclerotic effects, such as reducing adhesion of inflammatory cells, lipid accumulation and the proliferation of inflammatory cells [29–31]. Recently, it has also been shown that adiponectin receptors ADIPOR1 and ADIPOR2, through the AMPK, may modify the risk of CVD in individuals with IGT, possibly through alterations in the mRNA expression levels . Note that in this year, AMPK has been proposed as a therapeutic target for diabetic vascular disease .
Results from the present study show that glycemic control only exerts a significant effect on MMP-9 expession in OPA. These results are in agreement with previous studies showing that strict glycemic control does not improve cardiovascular disease progression in situations of advanced atherosclerosis [34, 35].
Our study has certain limitations. The study population consists of diabetic patients with advanced atherosclerosis. Therefore, results from the present study may not be extrapolated to other population types. In addition, the sample size was small, although the significant differences found, contribute to the strength of the results.
In conclusion, when we compared occluded arteries with vessels without atheromas in diabetic and atherosclerotic patients we found significant differences in biomarkers involved in inflammation, lipid metabolism and apoptotic pathways. In addition, compensatory mechanisms could exist that prevent the rupture of the atheromatous plaque in peripheral arterial occlusive disease. On the other hand, when the atherosclerotic process is studied in terms of good or poor glycemic control into the context of diabetes, we observed that the expression of genes involved in inflammation and apoptosis protection was increased in veins from patients with good diabetic control. In contrast to veins, in arteries with advanced thrombosis, like OPA, where lumen is almost completely occluded, the glycemic control did not seem to exert any effect on gene expression profile.