Decreased Inflammatory Markers in Diabetic Patients with Angiographically Proved Coronary Artery Disease after 18 Months of Statins Therapy DANIEL LIGHEZAN, ROXANA BUZAS, CORINA SERBAN, IOANA SUCEAVA University of Medicine and Pharmacy Victor Babes Timisoara COUNTRY: ROMANIA dr.corinaserban@yahoo.com Abstract: - The inflammation process in blood vessels can lead to complications in diabetes. The presence of diabetes in patients with coronary artery disease is a leading cause of morbidity and mortality. Objective: The aim of this study was to compare three inflammatory markers in angiographically patients with coronary artery disease with or without type 2 diabetes mellitus before and after 18 months of statins therapy. Methods: The study included 201 patients with angiographically proved coronary artery disease (CAD) that were divided considering the presence of diabetes mellitus: 69 CAD patients with DM and 132 CAD patients without DM. Three blood markers of inflammation: high-sensitivity C-reactive protein (hscrp), interleukin-6 (IL-6), and fibrinogen were compared between groups before and after 18 months of statins therapy. Serum IL- 6 values were measured by an enzyme immunoassay and serum hscrp by immunonephelometry method. Results: Initially, CAD patients with DM had higher levels of fibrinogen, hscrp and IL-6 than CAD patients without DM (360 ± 25 mg/dl vs. 353 ± 23 mg/dl, p=0.048, 4.04 ± 1.2 mg/l vs. 2.88 ± 1.4 mg/l, p<0.001 and 10.8 ± 2.54 pg/l vs 9.8 ± 1.56, p<0.001). After 18 months of statins therapy, CAD patients with DM had again significantly higher levels of fibrinogen and hscrp than CAD patients without DM (289 ± 23 mg/dl vs. 280 ± 21mg/dl, p=0.006, 1.15 ± 0.41 mg/l vs. 0.94 ± 0.42 mg/l, p<0.001), but the difference between IL-6 values was not significant. In CAD patients with DM, the levels of fibrinogen, hscrp and IL-6 significantly decreased after 18 months of statins treatment as compared with the values of inclusion (360 ± 25 mg/dl vs 289 ± 23 mg/dl, p<0.001, 4.04 ± 1.2 mg/l vs. 1.15 ± 0.4, p<0.001, 10.8 ± 2.54 pg/ml vs. 1.9 ± 0.4 pg/ml and 9.8 ± 1.56 pg/ml vs. 1.8 ± 0.38 pg/ml, p<0.001). Conclusion: Inflammatory markers (hscrp, fibrinogen, Il-6) are significantly elevated in CAD patients with DM compared with CAD patients without DM. After 18 months of statins treatment the values of studied inflammatory parameters were significantly reduced comparative with the values at inclusion. Key-Words: - fibrinogen, interleukin-6, high-sensitivity C-reactive protein, diabetes mellitus, statins 1. Introduction The number of people with diabetes grows faster than expected. In 2007, 246 million people (roughly 6%) were affected worldwide and it is estimated that this will increase to 380 million, or 7.3% by 2025 [1]. Due to systemic inflammation, oxidative stress and endothelial dysfunction with disorders of platelet function and fibrinolysis accelerated atherosclerosis can be seen in diabetic patients [2, 3]. In diabetes mellitus, multiple factors influence the prognosis in an unfavourable way, including: multi-vascular coronary affecting, diffuse coronary microangiopathy, diabetic cardiomyopathy, the atherogenic lipoprotein profile and increased inflammation markers [4]. Inflammation of the adipose tissue is considered to be a key factor in the pathogenesis of insulin resistance, but what causes this inflammation and why not all individuals with obesity develop insulin resistance remains controversial [5]. Low-grade inflammation is a common feature in patients with type 2 diabetes. Inflammatory cytokines are produced by different cell types and secreted into the circulation, where they regulate different tissues through their local, central and peripheral actions [6]. Inflammation of the vasculature might be the integrated mechanism that connects a diabetic phenotype with its attendant vascular complications [7]. Plasma fibrinogen is an independent risk factor for coronary, cerebral and peripheral artery disease and CRP may play a role in the development of atherosclerosis [8]. Recent Mendelian randomization studies failed to demonstrate a causal role between CRP levels and atherosclerosis, suggesting that CRP may more likely be a marker than an actual pathogenic component of atherosclerosis [9]. Another cytokine that regulates humoral and cellular responses and plays a central role in inflammation ISBN: 978-1-61804-122-7 294
and tissue injury is interleukin-6 (IL-6). Il-6 is associated with higher all-cause mortality, unstable angina, left ventricular dysfunction, propensity to diabetes and its complications, hypertension, obesity and several types of cancer [10, 11]. A large number of randomised controlled trials demonstrated the beneficial effects of statins on major vascular events and mortality rates in diabetics [12, 13]. The risk-benefit profiles of high dose statin therapy, particularly in patients with diabetes, is not directly addressed by evidence from robust clinical investigations [14]. Several randomized clinical trials showed that intensivedose statin therapy increased the risk of new-onset diabetes by 12% when compared with moderatedose therapy although intensive-dose statin therapy reduced risk of cardiovascular events by 16% when compared with moderate-dose therapy [15]. The latest recommendations of American Diabetic Association are screening, diagnostic, and therapeutic actions that are known or believed to favourably affect health outcomes of patients with diabetes [16]. However, it is not sufficiently known what changes may be observed in the level of inflammatory markers like fibrinogen, hscrp and IL-6, which may provide a complex information about the effect of the statin treatment. Some side effects of statins treatment in a long period of time are not clear enough. It is already known that the effectiveness of treatment in diabetic patients with coronary heart disease is decreased than in patients without diabetes due to metabolic balance [17, 18]. The aim of this study was to compare three inflammatory markers (hscrp, fibrinogen, Il-6) in patients with angiographically proved coronary artery disease divided considering the presence of type 2 diabetes mellitus, before and after 18 months of statins therapy. 2. Material and methods This prospective study included 201 patients (138 men and 62 women) with angiographically proved coronary artery disease (CAD) that were divided considering the presence of diabetes mellitus: 69 CAD patients with DM and 132 CAD patients without DM, after interview information regarding age, sex, hypertension, presence of diabetes, history of smoking and medication use. Exclusion criteria from the study were: treatment with anti-inflammatory or hypolipidemic agents, estrogen therapy, antiplatelet drugs; hscrp greater than 10 mg/l; serum creatinine more than 1.4 mg/dl for women and than 1.5 mg/dl for men; proteinuria greater than 300 mg/24-hour; secondary hypertension; chronic heart failure (New York Heart Association class III and IV); positive history or clinical signs of ischemic heart disease; diabetes mellitus; neoplastic or hepatic disease. In patients aged 34 and 77 years we performed: history, completed physical examination and biochemistry then we started medication with hydroxymethylglutaryl-coenzyme a reductase inhibitors (Atorvastatin, Simvastatin or Rosuvastatin): - Simvastatin was administered at doses of 20-40 mg depending on the degree of vascular damage, the maximum doses were administered to patients with severe multiple coronary vessels affection - Atorvastatin was administered at doses of 10-80 mg/day, the maximum dose was administered to patients with severe multiple coronary vessels affection - Rosuvastatin was administered at a dose of 10 mg / day. Body mass index (BMI) was calculated as weight (kg) divided by height squared (m 2 ). A value higher of 30 Kg/m 2 was considered obesity. Blood pressure was measured using three measurements of systolic and diastolic blood pressure, using the average of the last two measurements, with a standard sphygmomanometer (Riester, Germany). The subject was placed in a seated position, with arm raised to the heart, physically and mentally after a rest of 10 to 15 min. Blood pressure values used for statistical calculations were the average of six determinations in two moments of study: 3 BP determinations every 5 min at inclusion in the study group, and 3 measurements of BP every 5 min before the carotid ultrasound examination. The values obtained were interpreted following the recommendations of the European Society of Hypertension in 2007 [19]. Blood samples were collected in the morning, after patient fasting 8 h. Serum total cholesterol (TC), high-density lipoprotein cholesterol (HDLcholesterol), and triglyceride (TG) concentrations were assessed using standard enzymatic methods. Low density lipoprotein cholesterol was calculated using Friedewald s formula [20]. Fibrinogen was measured by Dimension RxL Max (Dade Behring Inc., USA), using a nephelometric research assay. Serum high-sensitivity CRP (hscrp) levels were measured using a wide-range latex-enhanced immunoturbidimetric assay. ISBN: 978-1-61804-122-7 295
The study was conducted according to the Declaration of Helsinki, and the written informed consent was obtained from each subject. 2.1 Statistical methods Means were calculated for continuous variables and proportions for categorical variables. Unpaired Student t tests or analyses of variance were conducted to assess statistical significance of differences between groups using Office Excel 2007 program. 3. Results The baseline characteristics of the patients, lipid profile and inflammatory parameters (fibrinogen, IL-6 and hscrp) compared between the two groups divided considering the presence of diabetes mellitus with t-student test, were presented in table 1. Table 1. Baseline characteristics of the patients and biochemical parameters at inclusion (mean± SD) CAD+DZ CAD-DZ P (n=69) (n=132) Age (years) 57.6±9.6 59.9±9.7 NS Sex (M %) 70 69 NS Positive history of CVD 32 46 NS (%) Smokers (%) 48 59 NS Fasting glycemia (mg/dl) 78 ± 6.21 126± 37.15 <0.001 Total cholesterol (mg/dl) 239±48 224 ± 59 NS Triglycerides (mg/dl) 207±93 168±64 <0.001 LDL cholesterol (mg/dl) 166.7 ± 4.02 143.49 ± 5.9 <0.001 HDL-cholesterol (mg/dl) 39.5 ± 4.01 45 ± 4.24 0.001 Fibrinogen (mg/dl) 360 ± 25 353 ± 23 0.048 hscrp (mg/l) 4.04 ± 1.2 2.88 ± 1.4 <0.001 Interleukin-6 (pg/ml) 10.8 ± 2.54 9.8 ± 1.56 <0.001 The baseline characteristics of the patients, lipid profile and inflammatory parameters (fibrinogen, IL-6 and hscrp) after 18 months of statins therapy, compared between the two groups divided considering the presence of diabetes mellitus with t- student test, were presented in table 2. Age (years) 57.6±9.6 59.9±9.7 NS Sex M/F (%) 70 69 NS Positive history of CVD 32 46 NS (%) Smokers (%) 48 59 NS Fasting glycemia (mg/dl) 78 ± 6.21 126± 37.15 <0.001 Total cholesterol (mg/dl) 172 ±28.89 163±32.03 <0.001 Triglycerides (mg/dl) 159+71 132+47 <0.001 LDL cholesterol (mg/dl) 91.7± 28.4 81.51± 27.72 <0.001 HDL-cholesterol (mg/dl) 50.1 ± 5.9 52.2 ± 6.5 0.026 Fibrinogen (mg/dl) 360 ± 25 280 ± 21 <0.001 hscrp (mg/l) 4.04 ± 1.2 0.94 ± 0.42 <0.001 Interleukin-6 (pg/ml) 1.9 ± 0.4 1.8 ± 0.38 NS The values of lipid profile and inflammatory parameters (fibrinogen, IL-6 and hscrp) in CAD+DM group before and after 18 months of statins therapy were presented in table 3. Table 3. Biochemical parameters in CAD+DM group before and after statins therapy CAD+DZ before statins (n=69) CAD+DZ after stains therapy (n=69) Total cholesterol (mg/dl) 239±48 172 ±28.89 <0.001 Triglycerides (mg/dl) 207±93 159+71 <0.001 LDL cholesterol (mg/dl) 166.7 ± 4.02 91.7± 28.4 <0.001 HDL-cholesterol (mg/dl) 39.5 ± 4.01 50.1 ± 5.9 0.026 Fibrinogen (mg/dl) 360 ± 25 289 ± 23 0.006 hscrp (mg/l) 4.04 ± 1.2 1.15 ± 0.41 <0.001 Interleukin-6 (pg/ml) 10.8 ± 2.54 1.9 ± 0.4 <0.001 The values of lipid profile and inflammatory parameters (fibrinogen, IL-6 and hscrp) in CAD- DM group before and after 18 months of statins therapy were presented in table 4. Table 4. Biochemical parameters in CAD-DM group before and after statins therapy CAD-DZ before statins (n=132) CAD-DZ after stains therapy (n=132) Total cholesterol (mg/dl) 224 ± 59 163±32.03 <0.001 Triglycerides (mg/dl) 168 ± 64 132 ± 47 <0.001 LDL cholesterol (mg/dl) 143.49 ± 5.9 71.51±27.72 <0.001 HDL-cholesterol (mg/dl) 45 ± 4.24 52.2 ± 6.5 0.026 Fibrinogen (mg/dl) 353 ± 23 280 ± 21 <0.001 hscrp (mg/l) 2.88 ± 1.4 0.94 ± 0.42 <0.001 Interleukin-6 (pg/ml) 9.8 ± 1.56 1.8 ± 0.38 <0.001 p p Table 2. Baseline characteristics of the patients and biochemical parameters after 18 months of statins therapy (mean± SD) CAD+DZ (n=69) CAD-DZ (n=132) P ISBN: 978-1-61804-122-7 296
between CAD patients with DM group and CAD patients without DM, at inclusion (p <0.001). It was not observed a statistically significant difference between mean values of IL-6 in the two groups, determined after 18 months of treatment (p = 0.083). For both groups of patients it was observed a statistically significant difference between mean values of IL-6 at inclusion and 18 months of treatment with statins (p<0.001). Fig. 1. Lipid profile parameters at inclusion Fig.3. Fibrinogen values at inclusion and after 18 months of statins therapy Fig. 2. Lipid profile parameters after 18 months of statins therapy The parameters of lipid profile were significantly increased in CAD patients with DM as compared with CAD patients without DM. It was noticed that lipid profile values significantly decreased after 18 months of statin therapy (figure 1 and figure 2). Statistically significant differences were obtained when we compared the mean baseline values of fibrinogen between CAD patients with DM and CAD patients without DM (p=0.048). After 18 months of treatment with statins it was also observed a statistically significant difference between mean values of fibrinogen between CAD group with DM and the group without DM (p=0.006). In both groups of patients, it was noticed a statistically significant difference between mean values of fibrinogen at inclusion and mean values of fibrinogen after 18 months of statin treatment for both groups (p<0.001) (figure 3). It was obtained a statistically significant difference between mean values of hscrp in CAD patients with DM compared with the CAD without DM (p<0.001). After 18 months of statin therapy remained statistically significant differences between mean values of hscrp between CAD group with DM and CAD group without DM (p<0.001) (figure 4). It were obtained statistically significant differences when we compared the mean values of IL-6 Fig.4. hscrp values at inclusion and after 18 months of statins therapy Fig.5. Il-6 values at inclusion and after 18 months of statins therapy In this study we evaluated diabetic patients according to degree of vascular damage and the presence or absence of diabetes, diabetic as: 31.57% of patients with severe monovascular affection, 33.33% of patients with severe bivascular ISBN: 978-1-61804-122-7 297
affection, and 45.45% of patients with severe trivascular affection as shown in figure 6. Figure 6. The distribution of the patients considering vascular affection 4. Discussions Our study showed that diabetes mellitus determines significant modifications of the lipid parameters, and inflammatory parameters (fibrinogen, hs-crp, IL-6) in patients with angiographically proven coronary artery disease. After 18 months therapy with statins, lipid and inflammation parameters significantly decreased. In our study, IL-6 values were significantly increased in patients with diabetes. IL-6 represents such an important factor in the development of atherosclerotic lesions and endothelial dysfunction in patients with diabetes, consistent with the literature. A significant reducing of inflammation parameters after statins therapy was observed in both groups of coronary patients. A possible cause of differences in the groups may be due to the coexistence of diabetes in association with obesity and hypertension. Our results are consistent with those from others studies. A subgroup analysis by the CTT collaborators investigated the statin effects in nearly 19,000 patients with diabetes mellitus, mostly type 2, which comprised ~ 20% of the total number of the CTT collaborators study subjects. The study revealed that the reductions in all-cause mortality and major vascular events observed in diabetics were comparable to those in non-diabetics and were demonstrated irrespective of the patients history of vascular disease [12, 21]. The management of patients with angiographically proven coronary artery disease and diabetes should focus mainly on the evaluation and overall control of all cardiovascular risk factors, rather than individual risk factors, and the final judgment of statin therapy should be individualized according to the profile and characteristics of each patient in part. Our findings have implications of the mechanism of benefit of statin therapy. Lipid lowering and reduction of inflammation have important implications for current and future treatment of atherosclerosis. 5. Conclusion Inflammatory markers (hscrp, fibrinogen, Il-6) are significantly elevated in diabetic patients with coronary artery disease compared with non-diabetic patients. After 18 months of statins treatment the values of studied inflammatory parameters were significantly reduced comparative with the values at inclusion. References: 1. Van den Oever, I.A., et al., Endothelial dysfunction, inflammation, and apoptosis in diabetes mellitus. Mediators Inflamm, 2010. 2010: p. 792393. 2. Hayden, J.M. and P.D. Reaven, Cardiovascular disease in diabetes mellitus type 2: a potential role for novel cardiovascular risk factors. Curr Opin Lipidol, 2000. 11(5): p. 519-28. 3. Sobel, B.E., R. Frye, and K.M. Detre, Burgeoning dilemmas in the management of diabetes and cardiovascular disease: rationale for the Bypass Angioplasty Revascularization Investigation 2 Diabetes (BARI 2D) Trial. Circulation, 2003. 107(4): p. 636-42. 4. Anavekar, N.S., et al., Predictors of cardiovascular events in patients with type 2 diabetic nephropathy and hypertension: a case for albuminuria. Kidney Int Suppl, 2004(92): p. S50-5. 5. Vaarala, O. and H. Yki-Jarvinen, Diabetes: Should we treat infection or inflammation to prevent T2DM? Nat Rev Endocrinol, 2012. 6. Calle, M.C. and M.L. Fernandez, Inflammation and type 2 diabetes. Diabetes Metab, 2012. 7. Shirwany, N.A. and M.H. Zou, Vascular inflammation is a missing link for diabetesenhanced atherosclerotic cardiovascular diseases. Front Biosci, 2012. 17: p. 1140-64. 8. Lefebvre, P., T. Ledent, and J. Ducobu, [Recent progress in the "fibrinogen hypothesis"]. Rev Med Brux, 2003. 24(2): p. 82-7. 9. Blankenberg, S. and S. Yusuf, The inflammatory hypothesis: any progress in risk stratification and therapeutic targets? Circulation, 2006. 114(15): p. 1557-60. 10. Fisman, E.Z. and A. Tenenbaum, The ubiquitous interleukin-6: a time for reappraisal. Cardiovasc Diabetol, 2010. 9: p. 62. 11. Abeywardena, M.Y., et al., Cardiovascular biology of interleukin-6. Curr Pharm Des, 2009. 15(15): p. 1809-21. ISBN: 978-1-61804-122-7 298
12. Rutishauser, J., Statins in clinical medicine. Swiss Med Wkly, 2011. 141: p. w13310. 13. Lardizabal, J.A. and P. Deedwania, Lipidlowering therapy with statins for the primary and secondary prevention of cardiovascular disease. Cardiol Clin, 2011. 29(1): p. 87-103. 14. Koh, K.K., I. Sakuma, and M.J. Quon, Differential metabolic effects of distinct statins. Atherosclerosis, 2011. 215(1): p. 1-8. 15. Preiss, D., et al., Risk of incident diabetes with intensive-dose compared with moderate-dose statin therapy: a meta-analysis. JAMA, 2011. 305(24): p. 2556-64. 16. Executive summary: Standards of medical care in diabetes--2012. Diabetes Care, 2012. 35 Suppl 1: p. S4-S10. 17. Berry, C., J.C. Tardif, and M.G. Bourassa, Coronary heart disease in patients with diabetes: part II: recent advances in coronary revascularization. J Am Coll Cardiol, 2007. 49(6): p. 643-56. 18. Berry, C., J.C. Tardif, and M.G. Bourassa, Coronary heart disease in patients with diabetes: part I: recent advances in prevention and noninvasive management. J Am Coll Cardiol, 2007. 49(6): p. 631-42. 19. Mancia, G., et al., 2007 ESH-ESC Practice Guidelines for the Management of Arterial Hypertension: ESH-ESC Task Force on the Management of Arterial Hypertension. J Hypertens, 2007. 25(9): p. 1751-62. 20. Friedewald, W.T., R.I. Levy, and D.S. Fredrickson, Estimation of the concentration of lowdensity lipoprotein cholesterol in plasma, without use of the preparative ultracentrifuge. Clin Chem, 1972. 18(6): p. 499-502. 21. Kearney, P.M., et al., Efficacy of cholesterollowering therapy in 18,686 people with diabetes in 14 randomised trials of statins: a meta-analysis. Lancet, 2008. 371(9607): p. 117-25. ISBN: 978-1-61804-122-7 299