|Year : 2017 | Volume
| Issue : 4 | Page : 1072-1078
Urinary 8-hydroxy-2′-deoxyguanosine as an oxidative DNA damage biomarker in chronic heart failure
Naglaa M Ghanayem1, Eman A E. Badr1, Ibrahim Elmadbouh1, Mahmoud A Soliman2, Sara K Rizk1
1 Department of Medical Biochemistry, Menoufia University, Menoufia, Egypt
2 Department of Cardiology, Menoufia University, Menoufia, Egypt
|Date of Submission||18-Dec-2016|
|Date of Acceptance||27-Feb-2017|
|Date of Web Publication||04-Apr-2018|
Sara K Rizk
Department of Medical Biochemistry, Menoufia University, Shebin Elkom, Menoufia
Source of Support: None, Conflict of Interest: None
The aim of this study was to evaluate the role of urinary 8-hydroxy-2′-deoxyguanosine (8-OHdG) as a biomarker of oxidative DNA damage in patients with chronic heart failure (CHF).
CHF is the leading cause of morbidity, mortality, and increasing healthcare costs around the world. Oxidative stress is known to play a crucial role in the pathogenesis of heart failure. In nuclear and mitochondrial DNA, 8-OHdG is one of the predominant forms of free radical-induced oxidative lesions.
Patients and methods
This study was conducted on 80 individuals: 50 CHF patients (22 men and 28 women) and 30 healthy controls (16 men and 14 women). The patients were selected from the Cardiology Department of Menoufia University Hospital during the period June 2015 to December 2015. All individuals included in this study were subjected to full history taking, clinical examination, echocardiography, and laboratory investigations that included tests for evaluation of serum lipid profiles, fasting blood sugar, 2 h postprandial glucose, serum creatinine, malondialdehyde, catalase, and urinary 8-OHdG, which was carried out by enzyme-linked immunosorbent assay technique.
Urinary 8-OHdG was significantly higher in patients than in controls. There was a significant statistical increase in malondialdehyde in patients compared with controls. There was a significant statistical decrease in catalase in patients compared with controls. There was a significant negative correlation between urinary 8-OHdG and systolic ejection fraction in the patients' group. Urinary 8-OHdG can be used as a biomarker in CHF at a cutoff of 5.5 ng/mg creatinine with an accuracy of 87.5%.
In CHF, urinary 8-OHdG can be a reliable marker of DNA damage as it reflects the level of oxidative stress and degree of CHF severity.
Keywords: 8-hydroxy-2′-deoxyguanosine, chronic heart failure, reactive oxygen species
|How to cite this article:|
Ghanayem NM, E. Badr EA, Elmadbouh I, Soliman MA, Rizk SK. Urinary 8-hydroxy-2′-deoxyguanosine as an oxidative DNA damage biomarker in chronic heart failure. Menoufia Med J 2017;30:1072-8
|How to cite this URL:|
Ghanayem NM, E. Badr EA, Elmadbouh I, Soliman MA, Rizk SK. Urinary 8-hydroxy-2′-deoxyguanosine as an oxidative DNA damage biomarker in chronic heart failure. Menoufia Med J [serial online] 2017 [cited 2018 Oct 17];30:1072-8. Available from: http://www.mmj.eg.net/text.asp?2017/30/4/1072/229237
| Introduction|| |
Chronic heart failure (CHF) represents a major public health issue with a complex pathology and disease management pattern as well as high prevalence.
It is a complex clinical syndrome that can result from a functional or structural cardiac disorder that impairs the ventricle's ability to fill with or eject blood.
Heart failure (HF)-related hospitalizations have increased substantially in the last decade, representing 1–2% of all hospital admissions and becoming the leading reason for admission in individuals aged 65 years or older.
Oxidative stress reflects an imbalance between the systemic manifestation of reactive species and a biological system's ability to readily detoxify the reactive intermediates or to repair the resulting damage.
Reactive oxygen species (ROS) are chemically reactive species containing oxygen. Examples include hydrogen peroxide, superoxide anion (O2−), hydroxyl radical (OH•), and singlet oxygen (1 O2).
To minimize the damage caused by free radicals, the organism utilizes enzymatic and nonenzymatic antioxidant systems. The first group comprises superoxide dismutase, glutathione peroxidase, catalase, and glutathione, whereas the second group consists of vitamin A, ascorbic acid (vitamin C), and α-tocopherol (vitamin E).
Higher production of ROS in the body may change the DNA structure, result in modification of proteins and lipids, activate several stress-induced transcription factors, and produce proinflammatory and anti-inflammatory cytokines.
DNA in the nucleus is one of the major targets of ROS, and 8-hydroxy-2′-deoxyguanosine (8-OHdG) is produced from deoxyguanosine in the DNA by ROS and is used as a marker of oxidative DNA damage.
8-OHdG was detected immunohistochemically in human cardiac tissues of patients with severe dilated cardiomyopathy. Therefore, 8-OHdG may be a suitable candidate as a cardiac biomarker to evaluate the level of oxidative stress.
The aim of this study was to assess the value of urinary 8-OHdG as a biomarker of oxidative DNA damage in patients with CHF.
| Patients and Methods|| |
This study was carried out on 80 individuals, who were classified into the following groups.
Group 1: this group included 50 patients with CHF: 22 men and 28 women with a mean age of 57.38 ± 8.70 years. The patients were attending the Outpatient Clinic and Inpatient Department of Cardiology, Menoufia University Hospital, during the period June 2015 to December 2015.
Diagnosis of HF was based on echocardiography with left ventricular ejection fraction (EF) less than 50%. Patients were classified on the basis of EF % as follows: 45–55%, mild stage; 30–44%, moderate stage; and less than 30%, severe stage.
Group 2: this group included 30 age-matched and sex-matched apparently healthy individuals. They were 16 men and 14 women with a mean age of 56.33 ± 5.12 years.
Menoufia Hospital's Review Board gave ethics approval for the study and all participants gave written informed consent before subject characterization and sample collection.
Patients with acute coronary syndrome, acute HF requiring artificial ventilation, other organ failures, cancer, or inflammatory diseases such as infection or collagen disease were excluded from this study.
All participants were subjected to the following: full assessment of history, a thorough clinical examination, pulse and blood pressure [systolic (SBP), diastolic (DBP)] measurements, determination of the presence or absence of cyanosis, and determination of the presence or absence of lower-limb edema. Laboratory investigations included measurement of lipid profile, fasting blood sugar, 2 h postprandial glucose, serum creatinine, plasma level of catalase enzyme, serum level of malondialdehyde (MDA), and urinary 8-OHdG.
After overnight fasting, 5 ml of venous blood was taken from each subject and divided as follows: 1 ml of blood was taken in a tube containing EDTA for measurement of plasma catalase enzyme level; another l ml was taken in a tube containing fluoride for measurement of fasting blood glucose level, and the remaining 3 ml was taken in a plain tube and left to clot for 30 min at room temperature. A second blood sample was taken after 2 h in a tube containing fluoride for measurement of 2 h postprandial blood glucose. All tubes were subjected to centrifugation for 10 min at 3000 g force. The serum and plasma obtained were divided into several aliquots and stored at −80°C until the time of assay.
Thirty-milliliter samples of morning urine were collected in a sterilized container and transported to the laboratory, where they were processed immediately. They were centrifuged to remove particulate matter and stored at −80°C for detection of urinary 8-OHdG.
Serum lipid profile [serum total cholesterol (TC), triglycerides (TG), and high-density lipoprotein cholesterol (HDLc)] were determined by enzymatic colorimetric method, using a BioMerieux kit (bioMérieux SA F-69280 Marcy l'Etoile France). Serum low-density lipoprotein cholesterol (LDLc) was calculated with the Friedewald's equation as LDLc=TC−(TG/5+HDLc). Fasting blood glucose and 2 h postprandial glucose were measured with a spectrophotometer using a Biodiagnostic kit (Egypt). Serum creatinine was determined by means of the enzymatic kinetic colorimetric Jaffe's method, using a Biodiagnostic kit.
The plasma level of catalase was determined by the enzymatic colorimetric method, using a Biodiagnostic kit. MDA was determined by the enzymatic colorimetric method on the basis of the reaction of thiobarbituric acid with MDA to form thiobarbituric acid-reactive product using a Biodiagnostic kit. Urinary 8-OHdG was determined by means of the enzyme-linked immunosorbent assay, using human 8-OHdG ELISA kit (Glory science Co., Ltd, USA). Raw values of urinary 8-OHdG were normalized to urinary creatinine levels and expressed as ng/mg creatinine.
The data collected were tabulated and analyzed by statistical package for the social sciences, version 20, on an IBM compatible computer (SPSS; SPSS Inc., Chicago, Illinois, USA). Quantitative data were expressed as mean ± SD and analyzed by applying the t-test for comparison between two groups of normally distributed variables, whereas for comparison between two groups of non-normally distributed variables the Mann–Whitney test was applied. Qualitative data were expressed as number and percentage and analyzed by applying the χ2-test; for 2 × 2 tables and if any cell had an expected number less than 5 Fisher's exact test was applied. Spearman's correlation was used for non-normally distributed quantitative variables or when one of the variables was qualitative. Receiver operating characteristic curve was used to determine cutoff points, sensitivity, and specificity for quantitative variables of interest.
| Results|| |
The results of the present study are presented in [Table 1],[Table 2],[Table 3],[Table 4],[Table 5],[Table 6] and in [Figure 1].
|Figure 1: Receiver operating characteristic (ROC) curve for urinary 8-hydroxy-2'-deoxyguanosine (8-OHdG) between CHF patients and controls (A). There is a significant negative correlation between urinary 8-OHdG/urinary creatinine (ng/mg creatinine) and EF% among CHF patients (B). |
Click here to view
|Table 1: Statistical comparison of demographic and clinical data between the studied groups|
Click here to view
|Table 2: Statistical comparison of laboratory parameters between the studied groups|
Click here to view
|Table 3: Distribution of laboratory parameters according to percentage ejection fraction in the patient subgroups|
Click here to view
|Table 4: Correlation between urinary 8-hydroxy-2′-deoxyguanosine and the studied parameters in the patient group|
Click here to view
|Table 6: Distribution of urinary 8-hydroxy-2′-deoxyguanosine and diabetes mellitus in the patient group on the basis of hypertension|
Click here to view
The results showed no significant statistical difference between the two studied groups as regards age and sex distribution. However, there was a significant statistical difference between the studied groups regarding pulse, SBP and DBP, hypertension, history of ischemic heart disease, diabetes mellitus (DM), presence of edema, and EF% [Table 1].
There was significant statistical increase in blood glucose, TC, TG, LDL, serum creatinine, MDA, and urinary 8-OHdG in group I compared with group II and significant statistical decrease in HDL and catalase in group I compared with group II [Table 2].
There was a nonsignificant statistical difference between the patient subgroups according to EF% with respect to blood glucose, TC, TG, LDL, HDL, serum creatinine, catalase, MDA, serum creatinine, and urinary 8-OHdG [Table 3].
There was significant negative correlation between urinary 8-OHdG and EF%. However, there was a nonsignificant correlation between urinary 8-OHdG and other parameters in the patient group [Table 4].
The diagnostic accuracy of urinary 8-OHdG in CHF patients as a marker of DNA damage was 87.5%, with sensitivity 96%, specificity 73%, positive predictive value 86%, and negative predictive value 92% at a cutoff point of 5.5 ng/mg creatinine [Table 5] and [Figure 1]. On the basis of the cutoff point the patients were divided into those having high urinary 8-OHdG and those having low urinary 8-OHdG. High urinary 8-OHdG was present in 45 (95.7%) of 47 patients of New York Heart Association (NYHA) class III and in three (100%) of three patients of NYHA IV. High urinary 8-OHdG levels were present in 12 (85.7%) of 14 patients with mild impairment of systolic functions, in 31 (100%) of 31 patients with moderate impairment, and in five (100%) of five patients with severe impairment of systolic functions (data not shown).
There was nonsignificant statistical difference in urinary 8-OHdG between the patient subgroups on the basis of hypertension and DM [Table 6].
| Discussion|| |
HF is the leading cause of morbidity, mortality, and healthcare costs around the world. The number of HF patients is increasing worldwide and its prevalence is expected to rise more steeply as the population ages.
Oxidative stress has been implicated in the pathogenesis and development of CHF, causing cardiomyocyte death, abnormalities in transduction of myocardial adrenergic receptor signaling, as well as contractile dysfunction. There is a significant correlation between oxidative stress and severity of CHF.
8-OHdG is a product of oxidative DNA damage produced by specific enzymatic cleavage after 8-hydroxylation of the guanine base. Upon DNA repair, 8-OHdG is excreted through urine.
This study assesses the value of 8-OHdG in CHF patients.
In the present study neither age nor sex was significant. There was a predominance of female sex (56%) compared with male sex (44%) among CHF patients, with a mean age of 57.38 ± 8.70 years. This is in agreement with the studies of Rywik et al., Nichols et al., Fonarow et al., and Joubin et al..
There was a significant statistical difference between patients and controls regarding pulse, SBP and DBP, hypertension, history of ischemic heart disease, DM, presence of edema, and EF%.
These results agree with those of Joubin et al., who found a significant increase in myocardial infarction and lower-limb edema in patients with CHF (52%) compared with the control group, with significant decrease in EF% in those patients. Fonarow et al. reported that the most common comorbid conditions in HF patients were hypertension (73%), coronary artery disease (57%), and diabetes (44%). Dei Cas et al. reported that in patients with DM the prevalence of HF is greater than that of the general population, and a 1% increase in glycosylated hemoglobin in DM patients is associated with an 8% increased risk for HF and that insulin resistance in HF patients, even in the absence of overt DM, is an independent predictor of worse prognosis, suggesting a pathophysiologic involvement of insulin resistance in HF progression.
The present study showed that the patients' group had significant statistical increase in TC, MDA, and urinary 8-OHdG compared with the control group.
These results agree with those of Sakatani et al., who described hypercholesterolemia as a risk factor for coronary artery disease.
Radovanovic et al. reported that MDA level might be a useful parameter for monitoring and planning the management of CHF patients.
Tsutsui et al. reported that oxidative stress-induced DNA damage (e.g. 8-OHdG) occurred in the mitochondria of failing hearts in a mouse MI model. Rivera et al. and Suzuki et al. reported that levels of 8-OHdG and lipid peroxidation were higher in HF patients than in controls. They stated that oxidative stress plays an important part in the pathophysiology and development of HF through the production of free radicals.
This study showed that urinary 8-OHdG proportionally increases with the severity of CHF in patients. Also, there was no significant difference between patients with and those without DM with respect to urinary 8-OHdG.
These results matched those of Kobayashi et al., who found that urinary 8-OHdG was higher in HF patients than in controls and became higher as the severity increased and that there was no difference in urinary 8-OHdG levels regardless of the presence of DM.
Our results were also commensurate with those of Watanabe et al., who found that serum 8-OHdG levels increased in proportion to the severity of disease, with a mean value of 0.24 ± 0.08 ng/ml in patients of NYHA functional class I and 0.32 ± 0.11 ng/ml in NYHA class II patients versus NYHA class III (0.75 ± 0.57 ng/ml, P = 0.04 and 0.003, respectively).
Susa et al. also documented that urinary 8-OHdG may reflect clinical severity with respect to both symptoms and cardiac dysfunction in CHF and that urinary 8-OHdG is predictive of morbidity and mortality in patients with CHF.
In contrast with this study, Nagayoshi et al. showed that urinary 8-OHdG levels were not associated with the severity of HF in patients with nonischemic HF.
In the present study, the cutoff point of urinary 8-OHdG was 5.5 ng/mg creatinine, with a sensitivity of 96% and a specificity of 73%. According to the cutoff point of urinary 8-OHdG, high urinary 8-OHdG levels were present in 95.7% of patients of NYHA class III and in 100% of NYHA class IV.
These results are in accordance with the findings of Takeishi, who showed that the optimal cutoff values for urinary 8-OHdG set at the point of maximum sensitivity plus specificity on each receiver operating characteristic curve was 12.4 ng/mg creatinine, with a sensitivity of 64.9% and a specificity of 70.4%.
Further, Suzuki et al. determined the normal upper limit of serum 8-OHdG as 0.40 ng/ml. HF patients were divided into two groups: those with normal serum 8-OHdG levels less than or equal to 0.40 ng/ml and those with high serum 8-OHdG levels greater than 0.40 ng/ml. High serum 8-OHdG levels were present in 21.2% of patients of NYHA class I, in 43.1% of NYHA II, in 42.6% of NYHA III, and in 69.4% of NYHA IV.
| Conclusion|| |
It can be concluded that the concentration of urinary 8-OHdG is increased in patients with HF and acts an indicator of DNA damage and oxidative stress.
Urinary 8-OHdG concentrations positively correlated with the severity of CHF. Urinary 8-OHdG may provide useful prognostic information for clinical outcomes in patients with HF, suggesting that oxidative stress plays an important role in the pathophysiology of CHF.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Heidenreich PA, Albert NM, Allen LA, Bluemke DA, Butler J, Fonarow GC,et al
. Forecasting the impact of heart failure in the United States: a policy statement from the American Heart Association. Circ Heart Fail 2013; 6
Reda A, Soliman M, Ahmed M, Abd El-Ghani R. Assessment of left atrial function in patients with systolic heart failure: strain imaging study. Menoufia Med J 2015; 28
Athanasakis K, Bilitou A, Lee D, Karampli E, Karavidas A, Parissis J,et al
. Cost-effectiveness of eplerenone in NYHA class II chronic heart failure patients with reduced LVEF: an analysis for Greece. Clinicoecon Outcomes Res 2016; 8
Chandra K, Salman A, Mohd A, Sweety R, Najam A. Protection against FCA induced oxidative stress induced DNA damage as a model of arthritis and in vitro
anti-arthritic potential of costus speciosus rhizome extract. Int J Pharmacognosy Phytochem Res 2015; 7
Hayyan M, Hashim MA, AlNashef IM. Superoxide ion: generation and chemical implications, Chem Rev 2016; 116
Lubrano V, Balzan S. Enzymatic antioxidant system in vascular inflammation and coronary artery disease. World J Exp Med 2015; 5
Birben E, Sahiner UM, Sackesen C, Erzurum S, Kalayci O. Oxidative stress and antioxidant defense. World Allergy Organ J 2012; 5
Takeishi Y. Biomarkers in heart failure. Int Heart J 2014; 55
Kobayashi S, Susa T, Tanaka T, Wada Y, Okuda S, Doi M,et al
. Urinary 8-hydroxy-2′-deoxyguanosine reflects symptomatic status and severity of systolic dysfunction in patients with chronic heart failure. Eur J Heart Fail 2011; 13
Lang R, Bierig M, Devereux R, Flachskampf F, Foster E, Pellikka P,et al
. Recommendations for chamber quantification. J Am Soc Echocardiogr 2005; 18
Yeon HK, Myung-Shik L. “Gut Microbiota and Metabolic Disorders”. Diabetes & Metabolism Journal 2015; 39
Trinder P. Determination of glucose in blood using glucose oxidase with an alternative oxygen acceptor. Ann Clin Biochem 1969; 6
Walser M. Assessing renal function from creatinine measurements in adults with chronic renal failure. Am J Kidney Dis 1998; 32
Zámocký M, Koller F. Understanding the structure and function of catalases: clues from molecular evolution and in vitro
mutagenesis. Prog Biophys Mol Biol 1999; 72
Mattson JP, Sun J, Murray DM, Poole DC. Lipid peroxidation in the skeletal muscle of hamsters with emphysema. Pathophysiology 2002; 8
Patel PR, Bevan RJ, Mistry N, Lunec J. Evidence of oligonucleotides containing 8-hydroxy-2'-deoxyguanosine in human urine. Free Radic Biol Med 2007; 42
Sakano N, Wang D, Takahashi N, Wang B, Sauriasari R, Kanbara S,et al
. Oxidative stress biomarkers and lifestyles in Japanese healthy people. J Clin Biochem Nutr 2009; 44
Hoon Lee J, Lim N, Cho M, Park HY. Epidemiology of heart failure in Korea: present and future. Korean Circ J 2016; 46
Sawyer DB, Colucci WS. Mitochondrial oxidative stress in heart failure. Circ Res 2000; 86
Kasai H. Chemistry-based studies on oxidative DNA damage: formation, repair and mutagenesis. Free Radic Biol Med 2002; 33
Rywik S, Wagrowska H, Broda G, Sarnecka A, Pytlak A, Polakowska M,et al
. Heart failure in patients seeking medical help at outpatients clinics. Part I. General characteristics. Eur J Heart Fail 2000; 2
Nichols A, Hillier T, Erbej, Brown J. Congestive heart failure in type 2 diabetes. Diabetes Care 2001; 24
Fonarow G, Adams K, Emerman C, LeJemtel T, Costanzo M, Abraham W,et al
. Characteristics and outcomes of patients hospitalized for heart failure in the United States: rationale, design, and preliminary observations from the first 100,000 cases in the Acute Decompensated Heart Failure National Registry (ADHERE). Am Heart J 2005; 194
Joubin L, Steg G, McCord J, Abraham W, Hollander J, Omland T,et al
. B-type natriuretic peptide and echocardiographic determination of ejection fraction in the diagnosis of congestive heart failure in patients with acute dyspnea. Chest 2005; 128
Dei Cas A, Khan S, Butler J, Mentz R, Bonow R, Avogaro A,et al
. Impact of diabetes on epidemiology, treatment, and outcomes of patients with heart failure. Heart Fail 2015; 3
Sakatani T, Shirayama T, Suzaki Y, Yamamoto T, Mani H, Kawasaki T,et al
. The association between cholesterol and mortality in heart failure comparison between patients. Comparison between patients with and without coronary artery disease. Int Heart J 2005; 46
Radovanovic A, Savic-Radojevic A, Pljesa-Ercegovac M, Djukic M, Suvakov S, Krotin M,et al
. Markers of oxidative damage and antioxidant enzyme activities as predictors of morbidity and mortality in patients with chronic heart failure. J Card Fail 2012; 18
Tsutsui H, Ide T, Shiomi T, Kang D, Hayashidani S, Suematsu N,et al
. Oxidative stress mediates tumor necrosis factor-alpha-induced mitochondrial DNA damage and dysfunction in cardiac myocytes. Circulation 2001; 104
Rivera M, Roselló-Lletí E, De Burgos F, Bertomeu V, Payá R, Cortés R,et al
. 8-Hydroxy-2'-deoxyguanosine and lipid peroxidation in patients with heart failure. Rev Esp Cardiol 2006; 59
Suzuki S, Shishido T, Ishino M, Katoh S, Sasaki T, Nishiyama A,et al
. 8-Hydroxy-2'-deoxyguanosine is a prognostic mediator for cardiac event. Eur J Clin Invest 2011; 41
Watanabe E, Matsuda N, Shiga T, Kajimoto K, Ajiro Y, Kawarai H,et al
. Significance of 8-hydroxy-2'-deoxyguanosine levels in patients with idiopathic dilated cardiomyopathy. J Card Fail 2006; 12
Susa T, Kobayashi S, Tanaka T, Murakami W, Akashi S, Kunitsugu I,et al
. Urinary 8-hydroxy-2'-deoxyguanosine as a novel biomarker for predicting cardiac events and evaluating the effectiveness of carvedilol treatment in patients with chronic systolic heart failure. Circ J 2012; 76
Nagayoshi Y, Kawano H, Hokamaki J, Uemura T, Soejima H, Kaikita K,et al
. Differences in oxidative stress markers based on the aetiology of heart failure: comparison of oxidative stress in patients with and without coronary artery disease. Free Radic Res 2009; 43
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6]