Lipid peroxidation is a process in which oxidants such as oxygen-free radicals attack body lipids . Phospholipids are well-known lipid targets of oxidative damage . 8-iso-prostaglandin F2α (8-iso-PGF2α) is an isoprostane that results from non-enzymatic oxidation of arachidonic acid in cell membrane phospholipids . 8-iso-PGF2α is considered the most reliable indicator of in vivo lipid peroxidation and it may contribute to the progression of diabetes and its vascular complications .
It has recently been recognized that atherosclerosis is mostly an inflammatory process and various studies have suggested a central role for the inflammatory cytokine, interleukin-6 (IL-6) in the inflammatory response responsible for atherosclerosis . IL-6 has been described as an independent predictor of type 2 diabetes mellitus (T2DM) and its cardiovascular complications. However, the existent evidence is not sufficient to create a causal relation between IL-6 levels and the progression to cardiovascular events .
Atherosclerosis is a dominant contributor to mortality and morbidity in T2DM . Carotid intima-media thickness (CIMT) is a reliable surrogate biomarker of subclinical atherosclerosis . Increased CIMT is associated with higher cardiovascular morbidity and mortality due to ischemic heart disease and stroke .
The aim of this study was to determine the relation of 8-iso-PGF2α as a marker of lipid peroxidation and IL-6 with CIMT in Egyptian T2DM patients. To the best of our knowledge, no similar studies have been previously conducted in this topic in Egypt.
Participants and methods
A total of 120 subjects were enrolled in this cross-sectional study. Ninety (90) of them were T2DM male patients with the age ≥ 40 years. The control group included 30 healthy male subjects matched with age. Patients with infectious diseases, malignancy, endocrinal or other metabolic diseases, hepatic diseases, renal impairment or cardiac dysfunction were excluded. The study protocol was approved by Alexandria University Ethics committee and all subjects gave their informed consent to be included in the study.
A complete history and a thorough physical examination including anthropometric assessment were obtained from every participant. After overnight fast for 12 hours, blood samples were withdrawn and fasting plasma glucose (FPG), glycosylated hemoglobin (HbA1c), lipid parameters, 8-iso-PGF2α and IL-6 were measured.
8-iso PGF2α was measured by ELISA technique using a kit from Cloud-Clone, Corp. (USA) and IL-6 was determined by ELISA technique using a kit from eBioscience, Inc. (Austria).
Carotid artery ultrasonography was performed for both sides of the neck using a standardized Doppler ultrasonic device (Arietta S70, Hitachi, USA) with 7.5 MHz probe. Measurements of CIMT were taken from the far wall of the common carotid artery, 2 cm proximal to the bifurcation . Average values calculated from measurements of both sides were taken for analysis . The procedure was performed by a single operator and the operator was blinded to the presence or absence of diabetes.
Statistical analyses were conducted using SPSS 18 system for Windows (SPSS Inc. Chicago, IL, USA). Data normality of distribution was tested and Mann–Whitney U test (two independent samples) was used to calculate the statistical significance between means. The strength of the association between variables was measured by using Spearman’s correlation coefficient (r). Multiple linear regression was performed to determine the predictors of carotid subclinical atherosclerosis. Differences were considered statistically significant when P ≤ 0.05.
Table 1 shows a comparison between both groups regarding to age, clinical and anthropometric characteristics with no significant difference in age, body mass index (BMI) and waist circumference (WC). Systolic blood pressure (SBP) and diastolic blood pressure (DBP) were significantly higher in diabetics (P < 0.001).
The baseline laboratory and ultrasonic characteristics of both the groups are summarized in Table 2. FPG, HbA1c and all serum lipid parameters [except high density lipoprotein (HDL)] were significantly higher in diabetic group (P < 0.05) while HDL was significantly lower in diabetic group (P < 0.001).
Diabetic patients were found to have a significantly higher 8-iso-PGF2α level when compared with non-diabetics (657 ± 476 pg/mL vs. 304 ± 243.6 pg/mL, P < 0.001) and the same was found as regards IL-6 (6.9 ± 4.5 pg/mL vs. 4.5 ± 1.5 pg/mL, P = 0.002) and CIMT (0.8 ± 0.14 mm vs. 0.58 ± 0.1 mm, P < 0.001)
The correlations between CIMT and all other parameters in diabetic subgroup are outlined in Table 3. CIMT had statistically significant positive correlations with age (P < 0.001), duration of DM (P < 0.001), SBP (P = 0.004), DBP (P < 0.001), FBG (P < 0.001), HbA1c (P < 0.001), LDL (P < 0.001), 8-iso-PGF2α (P < 0.001) (Fig. 1A) and IL-6 (P < 0.001) (Fig. 1B). In addition, CIMT was negatively correlated with HDL (P = 0.02) and had no significant correlation with BMI, WC, TC and TG.
In multiple linear regression analysis (Table 4): age, duration of DM, LDL, 8-iso-PGF2α and IL-6 were found to significantly predict the presence of subclinical atherosclerosis in T2DM with P value = (0.026, 0.02, 0.005, 0.044 and 0.001, respectively).
Oxidative stress is generally thought to share in contribution to atherosclerosis and diabetes, yet the data describing oxidized lipid in human atherosclerosis in diabetes is still insufficient .
In our study, we found a strong positive correlation between lipid peroxidation marker 8-iso-PGF2α and carotid subclinical atherosclerosis. Moreover, 8-iso-PGF2α was found to be a significant predictor of subclinical atherosclerosis in T2DM in multiple regression analysis. This is in line with what reported by Liu et al. . It is also in agreement with what revealed by Ono et al. , who studied the association between 8-iso-PGF2α and coronary subclinical atherosclerosis (assessed by coronary artery calcium score) in T2DM.
Likewise, other studies found a same correlation between both variables in different conditions like middle-age men  and hypertension [16, 17]. Moreover, 8-iso-PGF2α was reported to be directly correlated with the extent of coronary atherosclerosis .
The atherogenic role of 8-iso-PGF2α may be attributed to its mitogenic, vasoconstrictive and platelet-activating features .
Atherosclerosis is an inflammatory process involving many types of cells like macrophages, lymphocytes, endothelial cells and vascular smooth muscle cells. Activation of these cells to share in atherosclerosis is attributed to cytokines .
In spite of the presence of a large number of studies that have established a direct link between IL-6 and atherosclerosis, some studies did not reach to existence of such a relationship .
In the current study, a strong direct correlation between IL-6 and CIMT in T2DM was revealed and IL-6 was found to significantly predict carotid subclinical atherosclerosis in T2DM in multiple regression analysis. This is in good agreement with what showed by Dahlén et al. . It also goes in concordance with what found in a meta-analysis carried out by Zhang et al. .
The suggested mechanisms by which IL-6 can induce atherosclerosis include endothelial cell activation, prothrombotic effects on platelets, stimulation of vascular smooth muscle proliferation and increasing lipid accumulation in macrophages .
There were some limitations in our study. In addition to the relatively small sample size, our study population was composed of male Egyptian patients presented at a single hospital. This study was cross sectional, so we cannot explain the pathophysiological effect of lipid peroxidation and IL-6 on atherosclerosis.
In summary, this study shows an association between both lipid peroxidation and IL-6 with subclinical atherosclerosis in male subjects with T2DM. Whether targeting these risk factors with appropriate interventions can help prevent or treat atherosclerosis in patients with T2DM will require additional studies.
Conflicts of interest
There are no conflicts of interest.
1. Yin H, Xu L, Porter NA. Free radical lipid peroxidation
: mechanisms and analysis. Chem Rev. 2011; 111:5944–5972
2. Ayala A, Muñoz MF, Argüelles S. Lipid peroxidation
: production, metabolism, and signaling mechanisms of malondialdehyde and 4-hydroxy-2-nonenal. Oxid Med Cell Longev. 2014; 2014:360438
3. Kim JY, Lee JW, Youn YJ, Ahn MS, Ahn SG, Yoo BS, et al. Urinary levels of 8-iso-prostaglandin f2α and 8-hydroxydeoxyguanine as markers of oxidative stress in patients with coronary artery disease. Korean Circ J. 2012; 42:614–617
4. Zhang Y, Du Y, He JF, Li KJ. 8-iso-prostaglandin-f2α: a possible trigger or accelerator of diabetic retinopathy. Int J Ophthalmol. 2016; 9:163–165
5. Hartman J, Frishman WH. Inflammation and atherosclerosis
: a review of the role of interleukin-6
in the development of atherosclerosis
and the potential for targeted drug therapy. Cardiol Rev. 2014; 22:147–151
6. Qu D, Liu J, Lau CW, Huang Y. IL-6 in diabetes and cardiovascular complications. Br J Pharmacol. 2014; 171:3595–3603
7. Leon BM, Maddox TM. Diabetes and cardiovascular disease: epidemiology, biological mechanisms, treatment recommendations and future research. World J Diabetes. 2015; 6:1246–1258
8. Bauer M, Caviezel S, Teynor A, Erbel R, Mahabadi AA, Schmidt-Trucksäss A. Carotid intima-media thickness as a biomarker of subclinical atherosclerosis
. Swiss Med Wkly. 2012; 142:w13705
9. Provost EB, Madhloum N, Int Panis L, De Boever P, Nawrot TS. Carotid intima-media thickness, a marker of subclinical atherosclerosis
, and particulate air pollution exposure: the meta-analytical evidence. Plos One. 2015; 10:e0127014
10. Amer MS, Khater MS, Omar OH, Mabrouk RA, Mostafa SA. Association between Framingham risk score and subclinical atherosclerosis
among elderly with both type 2 diabetes mellitus and healthy subjects. Am J Cardiovasc Dis. 2014; 4:14–19
11. Kadoglou NP, Sailer N, Kapelouzou A, Lampropoulos S, Vitta I, Kostakis A. Effects of atorvastatin on apelin, visfatin (nampt), ghrelin and early carotid atherosclerosis
in patients with type 2 diabetes. Acta Diabetol. 2012; 49:269–276
12. Kathir K, Dennis JM, Croft KD, Mori TA, Lau AK, Adams MR. Equivalent lipid oxidation profiles in advanced atherosclerotic lesions of carotid endarterectomy plaques obtained from symptomatic type 2 diabetic and nondiabetic subjects. Free Radic Biol Med. 2010; 49:481–486
13. Liu JB, Li WJ, Fu FM, Zhang XL, Jiao L, Cao LJ. Inverse correlation between serum adiponectin and 8-iso-prostaglandin F2α in newly diagnosed type 2 diabetes patients. Int J Clin Exp Med. 2015; 8:6085–6090
14. Ono M, Takebe N, Oda T, Nakagawa R, Matsui M, Sasai T. Association of coronary artery calcification with MDA-LDL-C/LDL-C and urinary 8-isoprostane in Japanese patients with type 2 diabetes. Inter Med. 2014; 53:391–396
15. Yoon JH, Kim JY, Park JK, Ko SB. Oxidative damage markers are significantly associated with the carotid artery intima-media thickness after controlling for conventional risk factors of atherosclerosis
in men. Plos One. 2015; 10:e0119731
16. Guarneri M, Geraci C, Incalcaterra F, Arsena R, Mulè G, Vaccaro F, et al. Subclinical atherosclerosis
and fetuin-A plasma levels in essential hypertensive patients. Hypertens Res. 2013; 36:129–133
17. Yavuzer H, Yavuzer S, Cengiz M, Erman H, Doventas A, Balci H, et al. Biomarkers of lipid peroxidation
related to hypertension in aging. Hypertens Res. 2016; 39:342–348
18. Kuchta A, Strzelecki A, Ćwiklińska A, Totoń M, Gruchała M, Zdrojewski Z, et al. PON-1 activity and plasma 8-isoprostane concentration in patients with angiographically proven coronary artery disease. Oxid Med Cell Longev. 2016; 2016:9
19. Autieri MV. Pro- and anti-inflammatory cytokine networks in atherosclerosis
. ISRN Vas Med. 2012; 2012:17
20. Zhang B, Wang J, Xu Y, Zhou X, Liu J, Xu J, et al. Correlative association of interleukin-6
with intima media thickness: a meta-analysis. Int J Clin Exp Med. 2015; 8:4731–4743
21. Dahlén EM, Tengblad A, Länne T, Clinchy B, Ernerudh J, Nystrom FH. Abdominal obesity and low-grade systemic inflammation as markers of subclinical organ damage in type 2 diabetes. Diabetes Metab. 2014; 40:76–81
22. Reiss AB, Siegart NM, De Leon J. Interleukin-6
: atherogenic or atheroprotective? Clin Lipidol. 2017; 12:14–23