SERUM YKL-40 LEVELS IN CHRONIC HEART FAILURE

Maria Kazakova1, Petar Nikolov2, Kiril Simitchiev3, Fedya Nikolov2, Victoria Sarafian1 1) Department of Medical Biology, Medical Faculty, Medical University Plovdiv, 2) Department of Cardiology, Medical Faculty, Medical University Plovdiv, 3) Department of Analytical Chemistry and Computer Chemistry, Faculty of Chemistry, University of Plovdiv, Bulgaria. Journal of IMAB Annual Proceeding (Scientific Papers) 2014, vol. 20, issue 2


INTRODUCTION
Despite advances in the prevention and treatment of cardiovascular diseases heart failure is still a leading cause of mortality and disability.It represents a global public health problem.Chronic heart failure (CHF) is a heterogeneous disease with diverse etiologies and clinical phenotypes (1).
In CHF, activated inflammatory processes are observed.Several studies demonstrate that the increased serum levels of inflammatory cytokines and chemokines are significantly associated with altered functional cardiac performance (2).YKL-40, known as chitinase-3 like-1 protein (CHI3L1), is a glycoprotein secreted by activated macrophages and neutrophils in different tissues with inflammation, arthritic chondrocytes, vascular smooth muscle cells and cancer cells (3,4).
In patients, elevated YKL-40 levels are associated with pathological processes characterized by inflammation, extracellular remodeling and tissue fibrosis (5).The exact function of YKL-40 remains unknown, but it was suggested that the glycoprotein participated in cell proliferation and differentiation, endothelial dysfunction, atherosclerosis and neoangiogenesis (6,7,8).
The purpose of the current study was to determine serum YKL-40 levels in patients with CHF and to evaluate the potential relationship with ultrasonography and functional tests.

Subjects
The target group consisted of 24 patients with CHF with mean age 70 years (55-89; min-max), of whom 58% were males.Levels of hemoglobin, creatinine, uric acid and aminotransferases were measured with standard laboratory techniques and all of them were within the reference range.The controls were 16 age-comparable volunteers who attended a health screening.
The study was approved by the University Ethics Committee.All participants provided informed consent, according to the Helsinki Declaration.

ELISA
Blood samples were taken in the morning and serum was isolated by centrifugation at 2500 rpm for 10 minutes.The sera were stored at -80 ºC until analysis.Serum YKL-40 levels were determined by ELISA (Quidel, CA, USA).The intra-assay and inter-assay coefficients of variation were 5.8 and 6.0%, respectively.All samples were analysed in duplicates.

Six minute walk test (6MWT)
The patients had to walk the longest distance possible in interval of 6 min, through a walking course (corridor) 30 m long, according to the guidelines of American Thoracic Society.

Statistical methods, graphics and applied software
The Mann-Whitney U test was used to compare the values of a continuous variable in two independent groups.Boxplot diagrams were used for graphical visualization of a continuous variable as well as to distinguish outliers in the data series (the criteria 1.5 of the interquartile range have been accepted).The correlations between two continuous variables were estimated by the Kendall's taub correlation coefficient.All calculations have been made with MS Excel 2010.

RESULTS
Our study determined significantly higher YKL-40 levels in serum of CHF patients compared to controls (P=0.010).The median serum concentration of YKL-40 in the target group was 151 ± 64 ng/ml vs 95 ± 51 ng/ml (median ± standard deviation) in healthy people.The results are presented on Fig. 1.
Echocardiography showed that EDD and ESD levels were in upper limit values in patients.The data from EF and 6MWT decreased compared to reference range.Table 1 indicates the results from the laboratory and clinical measurements performed in the patients' group.Òhese results revealed weakness of the heart muscle, ventricular hypertrophy with impaired diastolic filling and function.Òhe damaged pumping function of the heart muscle leads to heart failure.
Next, we evaluated the correlation between serum YKL-40 levels, echocardiography examination and the 6MWT.We did not detect any relationship between the investigated parameters and YKL-40.Legend: The median level of YKL-40 in CHF patients was 151 ± 64 ng/ ml while in the control group it was 95 ± 51 ng/ml.YKL-40 was increased in patients compared to healthy subjects (P=0.010).

DISCUSSION
New therapies and biomarkers are needed to follow up the progression of CHF.Inflammatory mediators could provide important prognostic information in CHF patients (9).Although YKL-40 was supposed to be a potential biomarker in cardiovascular diseases, there is a controversy in the findings of different groups (10,11).
In our study, we demonstrated significantly higher serum YKL-40 levels in CHF patients compared to healthy subjects.These findings were in accordance with a large study of 717 patients with CHF followed up for 7 years, suggesting that YKL-40 was a new biomarker for all-cause mortality.On the other hand, some researchers claimed that the concentration of the glycoprotein could not serve as a prognostic indicator in cardiovascular pathologies (7).
The functional role of YKL-40 in CHF is not clearly defined yet.In vivo, macrophages in atherosclerotic plaques express the glycoprotein and the highest YKL-40 mRNA was determined in macrophages in early atherosclerotic lesions.The researchers concluded that the protein could be used as a marker for atherosclerosis at the onset of the disease (12).
In vitro, YKL-40 induced lymphocyte and endothelial cells adherence, migration and proliferation, leading to extracellular matrix remodeling and abnormal angiogenesis (7).Protein expression was detected by immunohistochemistry in developing rat hearts (13).
Recent investigations revealed that glycoprotein levels increased in patients after an acute myocardial infarction correlating with the number of damaged vessels (10,14).
The impact of echocardiography and the 6MWT is to provide objective evidence of structural and functional cardiac disorder (15).In our study there was no relationship between YKL-40 levels and the results from these functional tests.Weakness of the heart muscle and ventricular hypertrophy were observed.We detected impaired pumping function of the heart muscle, leading to heart failure.This fact was in agreement with a study of patients with CHF where YKL-40 was elevated compared to healthy age-matched people, but was not associated with other clinical characteristics or prognosis (7).Further investigations are needed to clarify the biological role of this glycoprotein in CHF.
The ultrasonographic and functional examinations detected evidence-based pathological changes in heart structure and function.We suppose that YKL-40 might reflect the inflammatory route in the development of CHF.

CONCLUSION
The increased serum YKL-40 levels in patients with chronic heart failure highlight the possibility this novel inflammatory biomarker to be associated with disease pathogenesis and progression.

Table 1 .
Laboratory and clinical characteristic of CHF patients.