A significant correlation between C - reactive protein levels in blood monocytes derived macrophages versus content in carotid atherosclerotic lesions.

BACKGROUND: Atherosclerosis is a complex disease involving different cell types, including macrophages that play a major role in the inflammatory events occurring in atherogenesis. C-Reactive Protein (CRP) is a sensitive systemic marker of inflammation and was identified as a biomarker of cardiovascular diseases. Histological studies demonstrate CRP presence in human atherosclerotic lesions, and we have previously shown that macrophages express CRP mRNA. CRP could be locally secreted in the atherosclerotic lesion by arterial macrophages and local regulation of CRP could affect its pro-atherogenic effects. Moreover, human blood derived macrophages (HMDM) expression of CRP could reflect atherosclerotic lesion secretion of CRP. METHODS: Ten type 2 diabetic patients and ten non-diabetic patients scheduled to undergo carotid endarterectomy were enrolled in this study, and their blood samples were used for serum CRP, lipid determination, and for preparation of HMDM further analyzed for their CRP mRNA expression and CRP content. Carotid lesions obtained from the patients were analyzed for their CRP and interleukin 6 (IL-6) content by immunohistochemistry. RESULTS: Lesions from diabetic patients showed substantially higher CRP levels by 62% (p = 0.05) than lesions from non diabetic patients, and CRP staining that co-localized with arterial macrophages. CRP carotid lesion levels positively correlated with CRP mRNA expression (r2 = 0.661) and with CRP content (r2 = 0.611) in the patient's HMDM. CONCLUSIONS: Diabetes up-regulated carotid plaques CRP levels and CRP measurements in HMDM could reflect atherosclerotic lesion macrophages secretion of CRP. Understanding the regulation of locally produced macrophage CRP in the arterial wall during atherogenesis could be of major importance in identifying the underlying mechanisms of inflammatory response pathways during atherogenesis.

Niacin administration significantly reduces oxidative stress in patients with hypercholesterolemia and low levels of high-density lipoprotein cholesterol.

Oxidative stress has been implicated in the pathogenesis of cardiovascular disorders, including atherosclerosis. In pharmacological doses, niacin (vitamin B3) was proven to reduce total cholesterol, triglyceride, very-low-density lipoprotein, and low-density lipoprotein levels, and to increase high-density lipoprotein (HDL) levels. The aim of this study was to evaluate the effect of niacin treatment in patients with low levels of HDL cholesterol (HDL-C; <40 mg%) on their lipid profile and oxidative stress status. Seventeen patients with hypercholesterolemia and low HDL-C and 8 healthy control subjects were enrolled in the study. The patients were treated with niacin for 12 weeks. Lipid profile, oxidative stress and C-reactive protein (CRP) levels were determined at the time of enrollment, and 2 and 12 weeks after initiation of niacin treatment. Subjects with lower HDL-C levels exhibited higher oxidative stress compared with subjects with normal HDL-C levels. Niacin treatment in hypercholesterolemic patients caused a significant increase in HDL-C and apolipoprotein A1 levels, and a decrease in triglyceride levels. Niacin also significantly reduced oxidative stress, as measured by a significant decrease in the serum content of thiobarbituric acid reactive substances, lipid peroxides and paraoxonase activity, compared with the levels before treatment. Although serum CRP levels were not affected by niacin treatment, a correlation between CRP and HDL levels was obtained when computing the results. Niacin treatment in hypercholesterolemic patients with low HDL levels caused a significant decrease in their oxidative stress status. These results indicate an additional beneficial effect of niacin beyond its ability to affect the lipid profile.

The antioxidant HDL-associated paraoxonase-1 (PON1) attenuates diabetes development and stimulates ß-cell insulin release.

OBJECTIVE: To analyze the direct effects of paraoxonase-1 (PON1) on diabetes development and on ß-cell insulin release. METHODS AND RESULTS: Injection of rePON1 to mice, prior to STZ-induced diabetes, resulted in reduced incidence of diabetes, as well as, in higher serum insulin levels. Incubation of ß-cells with PON1 also dose-dependently increased insulin secretion and its cellular content. PON1 increased cell survival under high glucose levels, but not under high STZ concentrations. The addition of the PON1 carrier in the circulation - HDL, to ßTC3 cell line, had an additive effect on PON1-induced insulin secretion. PON1 administration to mice or incubation with ß-cells was associated with a substantial decreased oxidative stress. Just like PON1, the dietary anti-oxidants, pomegranate juice, punicalagin (major polyphenol in pomegranate) or vitamin E, also increased insulin release from ßTC3, but unlike PON1, failed to increase insulin cellular content, suggesting a possible role for PON1 in insulin biosynthesis, separately from PON1 antioxidative effect. Both, PON1 catalytic activity and PON1 association to HDL, were not required for PON1 stimulation of insulin release from ß-cells. However, the PON1 free sulfhydryl group was shown to be essential for insulin release by PON1, as blocking the PON1 SH group, abolished PON1 stimulatory effect on insulin secretion. CONCLUSION: PON1 is a potent anti-diabetic enzyme that exerts this protection against diabetes through its antioxidative, as well as via its insulin stimulation properties on ß-cells.

Insulin increases macrophage triglyceride accumulation under diabetic conditions through the down regulation of hormone sensitive lipase and adipose triglyceride lipase.

Diabetes mellitus (DM) is a major risk factor for the development of atherosclerosis, and high-serum levels of insulin are strongly associated with type 2 DM. Atherosclerosis is characterized by lipid-laden macrophage foam cell formations, which contain substantial amount of cholesterol and triglycerides (TG). This study analyzed for the first time, the effects of insulin on TG metabolism in macrophages under normal and diabetic conditions. Mouse peritoneal macrophages from C57BL6 mice were cultured under normal (5 mM) or high (diabetic condition, 25 mM) glucose concentration, with or without insulin, followed by the assessment of TGs metabolism in these cells. Under diabetic condition, insulin increased TG accumulation in macrophages by 100%, decreased cellular TG degradation by 21%, and increased C-reactive protein levels in macrophages by 83%. Insulin decreased hormone-sensitive lipase mRNA and protein expression by 28 and 60%, respectively, and adipose TG lipase (ATGL) protein expression by 36%, with no significant reduction in ATGL mRNA levels. The inhibition of insulin-mediated phosphorylation, and the addition of cyclic adenosine 3'5'-monoposphate, abolished the insulin-mediated inhibition of TGs degradation in cells. Insulin increases macrophage TGs accumulation only under diabetic conditions, suggesting that impaired glycemic control in diabetic patients treated with insulin may contribute to foam cell formations and enhanced inflammation in macrophages.

Paraoxonase 2 (PON2) decreases high glucose-induced macrophage triglycerides (TG) accumulation, via inhibition of NADPH-oxidase and DGAT1 activity: studies in PON2-deficient mice.

OBJECTIVE: The present study investigates the role of paraoxonase 2 (PON2) in the attenuation of macrophage triglycerides (TG) biosynthesis, and oxidative stress, under diabetic conditions. METHODS: Peritoneal macrophages (MPM) from PON2-deficient and from C57BL/6 control mice were harvested and cultured under normal (5mM) or high glucose concentration (30mM), and evaluated for cellular TG metabolism as well as for their oxidative stress. RESULTS: In PON2-deficient MPM vs. control MPM, under diabetic conditions (high glucose concentration), we observed substantial increment in TG accumulation (3 fold), TG biosynthesis (2.6 fold) and microsomal diacylglycerol acyltransferase1 (DGAT1) activity (+60%). Furthermore, in these cells we have demonstrated increased oxidative stress, as expressed by significant increment in cellular oxidative stress (+25%), macrophage-mediated LDL oxidation (+41%) and expression of the receptor for advanced glycation end products - RAGE (+18%). Apocynin, an NADPH-oxidase inhibitor, abolished the increment in MPM TG accumulation, MPM TG biosynthesis, and microsomal DGAT1 activity, as a result of PON2-deficiency, under diabetic conditions. CONCLUSION: We conclude that PON2 has a significant protective role against macrophage triglyceride accumulation, macrophage TG biosynthesis, microsomal DGAT1 activity and macrophage oxidative stress, under high glucose concentrations. We suggest that this protective effect may be mediated by PON2 through the attenuation of NADPH-oxidase activity. The use of appropriate means to increase macrophage PON2 expression can lead to attenuation in macrophage TG accumulation and in cellular oxidative stress, under diabetic conditions, and thus may contribute to the decrement in macrophage atherogenicity and foam cell formation, attenuating the development of vascular complications in diabetes mellitus.

Primed polymorphonuclear leukocytes constitute a possible link between inflammation and oxidative stress in hyperlipidemic patients.

BACKGROUND: Oxidative stress (OS) and chronic inflammation are involved and contribute to the development of atherosclerosis. Primed polymorphonuclear leukocytes (PMNLs) are a possible source for superoxide radicals and inflammatory mediators, hence can promote OS and inflammation. The involvement of primed PMNLs in clinical states associated with high risk for developing cardiovascular disease and atherosclerosis, such as hypertension, renal failure and diabetes has been described, however, little is known about PMNLs characteristics in hyperlipidemic patients. METHODS: Hyperlipidemic patients and healthy control (HC) subjects were enrolled in this cross-sectional study. PMNL priming was estimated by measuring the rate of superoxide release and by levels of membrane CD11b. PMNL priming and myeloperoxidase (MPO) levels served as OS indices. Inflammation was linked to peripheral white blood cells and PMNL counts and to apoptosis. Systemic inflammation was estimated by blood levels of fibrinogen, C-reactive protein (CRP), transferrin and albumin. PMNL priming and inflammation parameters were related to the severity of hyperlipidemia. RESULTS: PMNLs from hyperlipidemic patients are primed compared to HC. A decrease in PMNL-MPO levels with increased levels of serum MPO were found in hyperlipidemic patients. Leukocyte counts tended to be higher in hyperlipidemic patients with increased PMNL apoptosis. PMNL priming and fibrinogen levels correlated positively with the severity of hyperlipidemia (r=0.32, P=0.02 for CD11b vs. cholesterol and r=0.38, P=0.009 for CD11b vs. LDL-c; r=0.35, P=0.01 for fibrinogen vs. cholesterol and r=0.3, P=0.03 for superoxide release vs. LDL-c). CONCLUSION: PMNLs are primed in hyperlipidemic patients contributing to OS and inflammation in these patients. This study highlights primed PMNLs as an additional risk factor for promoting atherosclerosis in hyperlipidemic patients.