Skip Navigation

European Journal of Heart Failure 2006 8(1):58-62; doi:10.1016/j.ejheart.2005.04.015
This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Search for citing articles in:
ISI Web of Science (5)
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by George, J.
Right arrow Articles by Keren, G.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by George, J.
Right arrow Articles by Keren, G.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© 2005 European Society of Cardiology

Usefulness of anti-oxidized LDL antibody determination for assessment of clinical control in patients with heart failure

Jacob George*, Dov Wexler, Arie Roth, Tomer Barak, David Sheps and Gad Keren

The Department of Cardiology Tel Aviv Sourasky Medical Center, Israel The Sackler Faculty of Medicine Tel Aviv University, Israel

* Corresponding author. E-mail address: jacobg{at}post.tau.ac.il


   Abstract
Top
 Abstract
1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 References
 
Background: It has been suggested that oxidative stress may play a role in the pathogenesis of heart failure, this may have potential implications for therapeutic strategies. However, measures of oxidative stress are subject to confounding inaccuracies. IgG antibodies to oxidized LDL reflect exposure to the lipoprotein over an extended period and may thus mirror oxidative stress over a prolonged time frame. Therefore, we tested the hypothesis that anti-oxLDL antibodies correlate with the control of heart failure (HF), as manifested by hospital admissions for cardiac dysfunction.

Methods: One hundred and two consecutive patients attending the HF clinic with either systolic or diastolic HF were enrolled and the quality of clinical control was evaluated by assessing hospital admissions over the year prior to index determination of the oxidative stress marker. Antibodies to oxLDL were determined by ELISA and pro-BNP levels were also measured.

Results: Most patients (mean age 71.5 years) had systolic HF; mean NYHA functional class was 2.7 and mean left ventricular ejection fraction was 39.7%. Anti-oxLDL antibodies, but not pro-BNP, correlated significantly with mean NYHA score (averaged from all clinic visits in the year prior to blood testing), and with hospital admissions over the year prior to blood testing. Mean IgG anti-oxLDL antibody levels in patients with hospital admissions were 3.4 times higher than those in subjects not hospitalized over the previous year.

Conclusion: IgG anti-oxLDL antibody levels correlate with the severity of HF.

Key Words: Heart failure • Oxidized-LDL • Antibodies • Pro-BNP • Oxidative stress

Received May 6, 2004; Accepted April 25, 2005


   1. Introduction
Top
 Abstract
 1. Introduction
2. Materials and methods
 3. Results
 4. Discussion
 References
 
Heart failure (HF) is responsible for a significant number of hospital admissions, and is the leading cause for hospitalization in subjects aged over 65 years [1]. Despite considerable progress in the understanding of the mechanisms involved in progression of HF, the associated morbidity and mortality are still unacceptably high. In order to achieve further improvement in the management of patients with HF, additional monitoring methods are required. Oxidative stress is a general term that denotes imbalance between factors that promote production of reactive oxygen species and the ability to oppose/scavenge and subsequently neutralize the byproducts of these reactive free radicals [2]. Several lines of evidence suggest that oxidative stress could be involved in the pathogenesis of HF [3]. Experimental studies in animal models of cardiac dysfunction such as those produced by myocardial infarction after left anterior descending artery ligation, adriamycin administration and pressure overload, all exhibit increased production of free radicals [4-6]. Accordingly, anti-oxidant therapy has been shown to attenuate myocardial damage induced by adriamycin [7]. In humans, assessment of oxidative stress is more complex, as there is no reproducible, standardized methodology. However, studies in patients with HF have used elevation of breath pentane levels [8], plasma thiobarbituric acid reactive substances and malondialdehyde-like activity [9], all of which are lipid peroxidation markers mirroring oxidative stress. Isoprostanes, which are relatively new markers of oxidative stress, have been shown to be increased in the pericardial fluid of patients with HF [10] and neutrophil superoxide-generating capacity [11] was also found to be elevated in patients with left ventricular dysfunction.

Oxidized LDL is present in the plasma and correlates with the extent of atherosclerosis [12-14]. Recently, plasma levels of oxLDL have been shown to be of prognostic value in predicting mortality in HF patients [15]. Assessment of antibodies to oxidized LDL may more reliably reflect the level of oxidative stress [16]. Indeed, antibodies to oxLDL have been shown to correlate with the extent of atherosclerosis and predict future myocardial infarction [17-19]. Therefore, we performed this study to assess whether determination of anti-oxLDL antibodies was useful for discrimination between clinically controlled and repeatedly hospitalized patients with HF.


   2. Materials and methods
Top
 Abstract
 1. Introduction
 2. Materials and methods
3. Results
 4. Discussion
 References
 
2.1. Patients
One hundred and two consecutive patients with mild to severe HF, with either diastolic HF, dilated cardiomyopathy or ischemic cardiomyopathy, who attended the HF clinic at the Tel Aviv Sourasky Medical Center between June 2001 and January 2002, entered this study. Patients with infection, inflammatory diseases, malignancy, congenital malformations of the heart or vessels, severe angina pectoris or a history of acute myocardial infarction (MI) within the past three months, were excluded. Clinical characteristics of the patients are shown in Table 1. The data were obtained from the medical records of the patients in the year prior to the index clinical visit. Details of HF hospitalizations were obtained both from medical records and from specific questioning of the patient.


View this table:
[in this window]
[in a new window]

  		Table 1 Clinical characteristics of HF patients (n=102)

 
Informed consent was obtained from all patients before participation in the study, and the protocol was approved by Ethics Committee of the local institution.

2.2. Determination of antibodies to oxLDL
Native and oxLDL were prepared as previously described [20]. Ninety-six well polystyrene plates (Nunc Maxisorp, Denmark) were coated with either copper oxLDL, native LDL (at concentration of 10 µg/ml in PBS) or PBS overnight at 4 °C. After washing four times with phosphate-buffered saline containing 0.05% Tween and 0.001% aprotinin (Sigma, USA) the plates were blocked with 2% BSA for 2 h at room temperature. Serum samples were diluted to 1:50 in PBS 0.05% Tween 0.2% BSA. After additional overnight incubation the sera were washed and alkaline phosphatase-conjugated goat anti-human IgG (Jackson ImmunoResearch laboratory Inc., USA) was added diluted 1:10,000 in PBS 0.05% Tween-0.2% BSA for 1 h at room temperature. After extensive washing, 1 mg/ml p-nitrophenyl-phosphate (Sigma) in 50 mM carbonate buffer containing 1 mM MgCl2 pH 9.8 was added as a substrate. The reaction was stopped after 30 min by adding 1 M of NaOH. The colour was read at a 405 nm wavelength in a Titertek ELISA reader (S.L.T Laboratory Instruments, Vienna, Austria) and results expressed as optical density at 405 nm. The values of oxLDL antibodies were calculated by subtracting the value obtained from binding to native LDL from the binding to oxLDL. Optical density levels were multiplied by a factor of 1000.

2.3. Determination of proBNP levels
Serum pro-BNP levels were measured by automated immuno-assay (Elecsys pro-BNP, Roche Diagnostics, Germany). The test principle includes using two polyclonal antibodies directed against N-terminal pro-BNP; epitope 1: amino acid 1-21 and epitope 2: amino acid 39-50. The results are calibrated against a synthetic N-terminal proBNP (amino acid 1-76). The range of results is between 5-35,000 pg/ml.

2.4. Statistical analysis
Comparisons between scalar parameters were performed using the Mann-Whitney non-parametric test whereas comparisons between continuous variables, including proBNP and anti-oxLDL antibodies was performed using the Student's t-test.

Correlation between oxLDL IgG levels and continuous parameters was performed using Spearman's correlation coefficient due to the skewed distribution of values. The software used for analysis was the SAS system for Windows version 8.02.


   3. Results
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
4. Discussion
 References
 
Of the 102 patients attending the HF clinic, 72% were males, 55% had systolic dysfunction, and 73% had ischemic cardiomyopathy. Hospital admissions were defined as only those documented to be due to deterioration in HF control, and requiring intravenous use of diuretics. The mean number of admissions was 0.44 (from 0 to 5 over the year prior to blood testing). The mean number of clinical visits over the one-year period was 9 per patient, which reflects the relatively severe HF in this group of patients (mean NYHA score 2.7). The clinical data for patients enrolled in the study are presented in Tables 1 and 2. The median level of pro-BNP was 3155 pg/ml, the median optical density value of IgG anti-oxLDL levels was 104 units.


View this table:
[in this window]
[in a new window]

  		Table 2 Concomitant medications in the HF patients

 
The ELISA for IgG anti-oxLDL antibodies was performed on a single batch of oxLDL, which was produced from the plasma of 20 healthy donors. The plate-to-plate variability, which was always less that 10%, was corrected by putting several control samples on all plates to serve as references. Moreover, repeated assays of all sera showed an inter-assay variability of less than 5%.

IgG anti-oxLDL antibodies increased as NYHA score increased When levels of oxidized LDL antibodies were correlated with binary factors, there was a significant association with chronic atrial fibrillation ({chi}2=5.96; p=0.015) and with chronic renal failure ({chi}2=6.82; p=0.009)(Table 3).


View this table:
[in this window]
[in a new window]

  		Table 3 Association of IgG anti-oxLDL antibody levels with baseline clinical characteristics in patients with HF-multivariate analysis

 
When IgG anti-oxidized LDL antibody levels were correlated with continuous variables by multivariate analysis, we found a significant association with NYHA score (p=.021), evidence of ischemia on thallium scan (0.02) and most significantly with hospital admissions for HF deterioration over the previous year (R=0.32; p=0.002) (Table 3). No significant differences in anti-oxLDL levels were evident between patients with systolic or diastolic HF (107±28 vs. 104±34 units, respectively, p=n.s.). By subgroup analysis, correlation between IgG anti-oxLDL levels and the number of hospital admissions did not differ between patients with systolic and diastolic HF (R=0.33, p=0.007 vs. R=0.31, p<0.009, respectively).

When IgG anti-oxLDL antibody levels were compared between patients with or without hospitalization in the year prior to enrollment, we found the levels were 3.4 times higher in the former group (221±101 OD units) as compared to the latter (65±16 OD units; p=0.006, Fig. 1A). A similar analysis of pro-BNP showed a 2.14 fold increase in patients admitted to hospital (5377±1384 pg/ml) as compared to non-admitted subjects (2516±419 pg/ml; p=0.009, Fig. 1B).


Figure 1
View larger version (8K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 1 Measurement of oxLDL antibody (A) and pro-BNP (B) levels in patients with HF with and without prior hospital admissions in the previous year. Levels of anti-oxLDL antibodies and pro-BNP were assayed as described in the text. Comparisons between admitted and non-admitted HF patients were performed using Student's t-test. *p=0.006; **p=0.009. (C) Lack of correlation between anti-oxLDL antibody levels and pro-BNP levels.

 
No correlation was evident between IgG anti-oxLDL antibody levels and serum values of pro-BNP (Fig. 1C).


   4. Discussion
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
References
 
In the current study, we sought to determine whether assessment of IgG antibody levels to oxidized LDL, were able to discriminate between patients with clinically controlled HF and patients requiring hospital admissions. Furthermore, we compared the discriminative power of anti-oxLDL antibodies with pro-BNP, which is a well-validated measure of HF severity.

The huge healthcare burden associated with HF morbidity and mortality necessitates continuous research for newer and more effective treatment modalities. However, this growing need also underscores the requirement for a simple and cost effective means of following up and predicting clinical control in patients. In this respect, recent years have witnessed the growing interest in measurement of BNP and pro-BNP as surrogate markers for the follow up of patients with HF [21,22]. However, as the pathogenesis of HF is multifactorial and involves, other than activating the neurohumoral axis, also enhanced oxidative stress, we reasoned that measurement of a marker of oxidative stress could prove useful. Unlike other indirect methods of assessment of oxidative stress, which require meticulous and extremely rapid removal and preservation of the serum, assessment of anti-oxLDL antibodies uses the more stable measurement of immunoglobulin G. Measurement of anti-oxLDL antibodies also reflects the status of lipoprotein oxidation over a prolonged period.

We found that anti-oxLDL antibody levels correlated significantly with the clinical status of HF patients, as defined by their NYHA score. Moreover, anti-oxLDL antibody levels correlated with the quality of HF control, as reflected by the number of hospital admissions recorded in the year prior to enrollment. We found that the discriminative power of anti-oxLDL antibodies was better than that obtained for serum pro-BNP, in patients admitted due to HF deterioration. These results, although from a case controlled study, nevertheless provide data to support the observation of elevated oxidative stress in patients with HF. Importantly, no association was found between pro-BNP and anti-oxLDL antibody levels suggesting that determination of the latter may have an incremental value over that provided by the former.

Additional findings in the current study relate to the significant association between anti-oxLDL antibodies and chronic renal failure and serum creatinine levels. This finding is in line with previous studies showing increased levels of anti-oxLDL antibodies in patients with chronic renal failure [23,24]. Antibody levels also correlated with the presence of chronic atrial fibrillation, a finding that could be related to a more severe HF or to the possible involvement of oxidative stress in the pathogenesis of atrial fibrillation.

We also compared levels of anti-oxLDL antibodies in the subgroups of patients with systolic and diastolic heart failure and found no significant difference. This may imply that the state of oxidative stress is not specific for the pathophysiological mechanism of HF but rather, to the patient's functional status. In this respect, the natriuretic peptides may exhibit a similar pattern as it has been shown that they are of value in predicting diastolic as well as systolic HF [25]. We also found that correlation between IgG oxLDL antibodies and the type of HF (systolic versus diastolic) was similar, again supporting a more functional value for antibody measurements.

In conclusion, we describe for the first time, the usefulness of determination of anti-oxidized LDL antibodies as a means of assessing the quality of clinical control in patients with HF. Thus, oxLDL antibodies correlate with the severity of HF and further studies are required to test their feasibility as predictive markers in HF patients.


   References
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 References
 

  1. Lenfant C. Report of the task force on research in heart failure. Circulation (1994) 90:1118–1123.[Free Full Text]
  2. Griendling K.K., FitzGerald G.A. Oxidative stress and cardiovascular injury: Part II. Animal and human studies. Circulation (2003) 108:2034–2040.[Free Full Text]
  3. MacCarthy P.A., Shah A.M. Oxidative stress and heart failure. Coron Artery Dis (2003) 14:109–113.[CrossRef][Web of Science][Medline]
  4. Myers C.E., McGuire W.P., Liss R.H., Ifrim I., Grotzinger K., Young R.C. Adriamycin: the role of lipid peroxidation in cardiac toxicity and tumour response. Science (1977) 197:165–167.[Abstract/Free Full Text]
  5. Singal P.K., Kapur N., Dhillon K.S., Beamish R.E., Dhalla A.K. Role of free radicals in catecholamine-induced cardiomyopathy. Can J Physiol Pharmacol (1982) 60:1390–1397.[Web of Science][Medline]
  6. Dhalla A.K., Hill M.F., Singal P.K. Role of oxidative stress in transition of hypertrophy to heart failure. J Am Coll Cardiol (1996) 28:506–514.[Abstract]
  7. Siveski-Iliskovic N., Hill M., Chow D.A., Singal P.K. Probucol protects against adriamycin cardiomyopathy without interfering with its antitumor effect. Circulation (1995) 91:10–15.[Abstract/Free Full Text]
  8. Sobotka P.A., Brottman M.D., Weitz Z., Birnbaum A.J., Skosey J.L., Zarling E.J. Elevated breath pentane in heart failure reduced by free radical scavenger. Free Radic Biol Med (1993) 14:643–647.[CrossRef][Web of Science][Medline]
  9. McMurray J., Chopra M., Abdullah I., Smith W.E., Dargie H.J. Evidence of oxidative stress in chronic heart failure in humans. Eur Heart J (1993) 14:1493–1498.[Abstract/Free Full Text]
  10. Mallat Z., Philip I., Lebret M., Chatel D., Maclouf J., Tedgui A. Elevated levels of 8-iso-prostaglandin F2[alpha] in pericardial fluid of patients with heart failure: a potential role for in vivo oxidant stress in ventricular dilatation and progression to heart failure. Circulation (1998) 97:1536–1539.[Abstract/Free Full Text]
  11. Ellis G.R., Anderson R.A., Lang D., Blackman D.J., Morris R.H.K., Morris-Thurgood J., et al. Neutrophil superoxide anion-generating capacity, endothelial function and oxidative stress in chronic heart failure: effects of short and long-term vitamin C therapy. J Am Coll Cardiol (2000) 36:1474–1482.[Abstract/Free Full Text]
  12. Boullier A., Bird D.A., Chang M.K., Dennis E.A., Friedman P., Gillotre-Taylor K., et al. Scavenger receptors, oxidized LDL, and atherosclerosis. Ann N Y Acad Sci (2001) 947:214–222.[Web of Science][Medline]
  13. Holvoet P., Vanhaecke J., Janssens S., Van de Werf F., Collen D. Oxidized LDL and malondialdehyde-modified LDL in patients with acute coronary syndromes and stable coronary artery disease. Circulation (1998) 98:1487–1494.[Abstract/Free Full Text]
  14. Holvoet P., Mertens A., Verhamme P., Bogaerts K., Beyens G., Verhaeghe R., et al. Circulating oxidized LDL is a useful marker for identifying patients with coronary artery disease. Arterioscler Thromb Vasc Biol (2001) 21:844–848.[Abstract/Free Full Text]
  15. Takashi T., Takayoshi T., Atsuyuki W., et al. Plasma oxidized low-density lipoprotein as a prognostic predictor in patients with chronic congestive heart failure. J Am Coll Cardiol (2002) 39:957–962.[Abstract/Free Full Text]
  16. Steirova A., Racek J., Stozicky F., et al. Antibodies against oxidized LDL- theory and clinical use. Physiol Res (2001) 50:131–141.[Web of Science][Medline]
  17. Salonen J.T., Yla-Herttuala S., Yamamoto R., et al. Autoantibodies against oxidized LDL and progression of carotid atherosclerosis. Lancet (1992) 339:883–887.[CrossRef][Web of Science][Medline]
  18. Lehtimaki T., Lehtinen S., Solakivi T., et al. Autoantibodies against low density lipoprotein in patients with angiographicly verified coronary artery disease. Arterioscler Thromb Vasc Biol (1999) 19:23-2.
  19. Puurunen M., Manttari M., Manninen V., et al. Antibody against oxidized low density lipoprotein predicting myocardial infarction. Arch Intern Med (1994) 154:2605–2609.[Abstract/Free Full Text]
  20. George J., Harats D., Bakshi E., Adler Y., Levy Y., Gilburd B., et al. Anti-oxidized low density lipoprotein antibody determination as a predictor of restenosis following percutaneous transluminal coronary angioplasty. Immunol Lett (1999) 68:263–266.[CrossRef][Web of Science][Medline]
  21. Wright S.P., Doughty R.N., Pearl A., Gamble G.D., Whalley G.A., Walsh H.J., et al. Plasma amino-terminal pro-brain natriuretic peptide and accuracy of heart-failure diagnosis in primary care: a randomized, controlled trial. J Am Coll Cardiol (2003) 42:1793–1800.[Abstract/Free Full Text]
  22. Anand I.S., Fisher L.D., Chiang Y.T., Latini R., Masson S., Maggioni A.P., et al. Val-HeFT Investigators. Changes in brain natriuretic peptide and norepinephrine over time and mortality and morbidity in the Valsartan Heart Failure Trial (Val-HeFT). Circulation (2003) 107:1278–1283.[Abstract/Free Full Text]
  23. Maggi E., Bellazzi R., Gazo A., et al. Autoantibodies against oxidatively-modified LDL in uremic patients undergoing dialysis. Kidney Int (1994) 46:869–876.[Web of Science][Medline]
  24. Bergesio F., Monzani G., Ciuti R., et al. Autoantibodies against oxidized LDL in chronic renal failure: role of renal function, diet, and lipids. Nephron (2001) 87:127–133.[CrossRef][Web of Science][Medline]
  25. Doust J.A., Glasziou P.P., Pietrzak E., Dobson A.J. A systematic review of the diagnostic accuracy of natriuretic peptides for heart failure. Arch Intern Med (2004) 164:1978–1984.[Abstract/Free Full Text]

Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 Eur J Heart FailHome page
O. Forster, D. Hilfiker-Kleiner, A. A. Ansari, J. B. Sundstrom, E. Libhaber, W. Tshani, A. Becker, A. Yip, G. Klein, and K. Sliwa
Reversal of IFN-{gamma}, oxLDL and prolactin serum levels correlate with clinical improvement in patients with peripartum cardiomyopathy
Eur J Heart Fail, September 1, 2008; 10(9): 861 - 868.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Search for citing articles in:
ISI Web of Science (5)
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by George, J.
Right arrow Articles by Keren, G.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by George, J.
Right arrow Articles by Keren, G.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?