756
Original Investigation
Serum cystatin C and neutrophil gelatinase-associated lipocalin in predicting the severity of coronary artery disease in diabetic patients Kaan Okyay, Aylin Yıldırır, Mutlu Çiçek, Alp Aydınalp, Haldun Müderrisoğlu Department of Cardiology, Faculty of Medicine, Başkent University Ankara Education and Research Hospital; Ankara-Turkey
ABSTRACT Objective: Cystatin C and neutrophil gelatinase-associated lipocalin (NGAL) are biomarkers of renal functions. We evaluated their roles in predicting the severity of coronary artery disease (CAD). Methods: Fifty-two consecutive type 2 diabetic patients (32 males, 65.7±8.6 years) who underwent coronary angiography (CAG) for stable CAD were included in this single-center, prospective, cross-sectional study. Patients with an estimated glomerular filtration rate 1.4 mg/ dL for men and >1.2 mg/dL for women and/or with an estimated glomerular filtration rate (eGFR) 200 mg/dL or use of hypolipidemic medications. eGFR was calculated by Modification of Diet in Renal Disease (MDRD) equation (18).
Okyay et al. Serum cystatin C and lipocalin
757
Laboratory measurements The venous blood samples for routine laboratory measurements were taken after 12-hours overnight fasting within 72 hours prior to CAG and analyzed in 1 hour. For determination of cystatin C and NGAL levels, the venous blood samples were drawn on the morning of CAG and stored at −20°C until analysis time, and were detected with sandwich enzyme immunoassay method using commercial Human Cystatin C kits (Biovendor Laboratory Medicine Inc., Brno, Czech Republic) and Human Lipocalin-2/NGAL kits (Biovendor Laboratory Medicine Inc.), respectively, by a microelisa analyser device (DSX Model; Dynex Technologies, Chantilly, VA, USA). Coronary angiography The coronary angiographies were performed using the standard Judkins technique. The angiograms were reviewed by two independent experienced invasive cardiologists for the presence and severity of CAD. Coronary vessel score was ranged between 0 and 3 according to the number of the main coronary arteries— left anterior descending (LAD), left circumflex (Cx), and right coronary artery (RCA)—with a ≥50% stenosis. Significant CAD was defined as a vessel score ≥1. Gensini score (19) was calculated for evaluation of the CAD severity in each patient. This system scores the stenosis in the epicardial coronary arteries (1 point for 1–25% stenosis, 2 for 26–50% stenosis, 4 for 51–75% stenosis, 8 for 76–90% stenosis, 16 for 91–99% stenosis, and 32 for total occlusion) and multiplies this number by a constant number in term of the anatomical position of the lesion. Multipliers are 5 for the left main coronary artery; 2.5 for the proximal segment of the LAD and Cx; 1.5 for the mid-segment of the LAD; 1 for the RCA, the distal segment of the LAD, the posterolateral artery, and the obtuse marginal artery; and 0.5 for other segments. The score in every segment is calculated, and the total score gives the Gensini score. The patients who have history of coronary revascularization (percutaneous coronary intervention or coronary artery bypass graft operation) were excluded from the study since the original Gensini’s scoring system is technically inappropriate for such cases. The median Gensini score was 23 (5–51, IQR: 46). The patients were divided into groups of severe and non-severe CAD according to this median score. Statistical analysis The normality of distribution was tested using Kolmogorov– Smirnov test. Continuous variables were presented as mean±SD if normally distributed otherwise as median with 25th and 75th percentiles. Categorical variables were presented as numbers and percentages and were compared using chi-square test (Fisher’s exact test if needed). Student’s t-test or Mann–Whitney U test was used for comparison of continuous variables where appropriate. Correlation analysis was performed using Spearman test. A multiple linear regression analysis model was generated in order to investigate the independent predictors of the Gensini score. Receiver operating characteristic (ROC) analysis was
758
Okyay et al. Serum cystatin C and lipocalin
Anatol J Cardiol 2016; 16: 756-61
Table 1. Demographics and baseline clinical parameters of the patients
Age, years Gender, male/female BMI, kg/ m2
Severe CAD (n=26)
Non-severe CAD (n=26)
P
65.7±8.6
64.3±8.4
66.5±9.6
0.08
32/20
19 / 7
13/13
0.15
29.2±3.9 29.2±4.2 29.3±3.6 0.91
Smoking, %
21.2
30.8
11.5
0.17
Hypertension, %
88.5
88.5
88.5
1.00
Hyperlipidemia, %
69.2
73.1
65.4
0.76
23 (5–51)
51 (29–69)
Gensini score HbA1c, %
7.41±1.45
7.66±1.47
7.18±1.43
0.24
BUN, mg/dL
18.63±6.86
19.92±8.28
17.34±4.89
0.18
Scr, mg/dL
0.86±0.17
0.92±0.15
0.80±0.17
0.006
eGFR, mL/min /1.73 m2 88.32±18.70 82.96±19.50 93.69±18.75 0.049 13.76±1.40
13.81±1.56
13.72±1.25
0.80
Total-C, mg/dL
188.81±35.36 190.66±31.80 186.52±40.22 0.73
LDL-C, mg/dL
112.00±30.55 116.44±31.10 107.17±29.86 0.30
HDL-C, mg/dL
40.89±9.64
41.15±10.59
1500.00 1300.00 1100.00 900.00 700.00 500.00 0
40.61±8.76
0.86
Triglycerides, mg/dL 147 (119–196) 150 (132–192) 129 (99–197) 0.20 Medications, % Insulin
23.1
34.6
11.5
0.10
Oral antihypoglycemic agents
86.5
80.8
92.3 0.42
ACEI
28.8
23.1
34.6 0.54
ARB
36.8
42.3
30.8
0.56
CCB
32.7
38.5
26.9
0.55
β-blockers
53.8
61.5
46.2 0.40
Statins
57.7
53.8
61.5
0.78
Acetylsalicylic acid
59.6
65.4
53.8
0.57
Warfarin
3.8
3.8
3.8
–
Nitrates
11.8
15.4
7.7 0.67
Data are presented as mean±standard deviation, median with 25th and 75th percentiles, or number and frequencies where indicated ACEI - angiotensin converting enzyme inhibitor; ARB - angiotensin receptor blocker; BMI - body mass index; BUN - blood urea nitrogen; C - cholesterol; CAD - coronary artery disease; CCB - calcium channel blocker; eGFR - estimated glomerular filtration rate; HbA1c - hemoglobin A1c; HDL-C - high-density lipoprotein cholesterol; LDL-C low-density lipoprotein cholesterol; Scr - serum creatinine Student’s t-test or Mann–Whitney U Test and chi-square test were used for statistical analyses
used to predict the cut off points for cystatin C in determination of multi-vessel disease. Then, sensitivity and specificity were calculated. Statistical analysis was performed by SPSS software (version 17; Statistical Package for Social Sciences, Chicago, IL). A p value of
Original Investigation
Serum cystatin C and neutrophil gelatinase-associated lipocalin in predicting the severity of coronary artery disease in diabetic patients Kaan Okyay, Aylin Yıldırır, Mutlu Çiçek, Alp Aydınalp, Haldun Müderrisoğlu Department of Cardiology, Faculty of Medicine, Başkent University Ankara Education and Research Hospital; Ankara-Turkey
ABSTRACT Objective: Cystatin C and neutrophil gelatinase-associated lipocalin (NGAL) are biomarkers of renal functions. We evaluated their roles in predicting the severity of coronary artery disease (CAD). Methods: Fifty-two consecutive type 2 diabetic patients (32 males, 65.7±8.6 years) who underwent coronary angiography (CAG) for stable CAD were included in this single-center, prospective, cross-sectional study. Patients with an estimated glomerular filtration rate 1.4 mg/ dL for men and >1.2 mg/dL for women and/or with an estimated glomerular filtration rate (eGFR) 200 mg/dL or use of hypolipidemic medications. eGFR was calculated by Modification of Diet in Renal Disease (MDRD) equation (18).
Okyay et al. Serum cystatin C and lipocalin
757
Laboratory measurements The venous blood samples for routine laboratory measurements were taken after 12-hours overnight fasting within 72 hours prior to CAG and analyzed in 1 hour. For determination of cystatin C and NGAL levels, the venous blood samples were drawn on the morning of CAG and stored at −20°C until analysis time, and were detected with sandwich enzyme immunoassay method using commercial Human Cystatin C kits (Biovendor Laboratory Medicine Inc., Brno, Czech Republic) and Human Lipocalin-2/NGAL kits (Biovendor Laboratory Medicine Inc.), respectively, by a microelisa analyser device (DSX Model; Dynex Technologies, Chantilly, VA, USA). Coronary angiography The coronary angiographies were performed using the standard Judkins technique. The angiograms were reviewed by two independent experienced invasive cardiologists for the presence and severity of CAD. Coronary vessel score was ranged between 0 and 3 according to the number of the main coronary arteries— left anterior descending (LAD), left circumflex (Cx), and right coronary artery (RCA)—with a ≥50% stenosis. Significant CAD was defined as a vessel score ≥1. Gensini score (19) was calculated for evaluation of the CAD severity in each patient. This system scores the stenosis in the epicardial coronary arteries (1 point for 1–25% stenosis, 2 for 26–50% stenosis, 4 for 51–75% stenosis, 8 for 76–90% stenosis, 16 for 91–99% stenosis, and 32 for total occlusion) and multiplies this number by a constant number in term of the anatomical position of the lesion. Multipliers are 5 for the left main coronary artery; 2.5 for the proximal segment of the LAD and Cx; 1.5 for the mid-segment of the LAD; 1 for the RCA, the distal segment of the LAD, the posterolateral artery, and the obtuse marginal artery; and 0.5 for other segments. The score in every segment is calculated, and the total score gives the Gensini score. The patients who have history of coronary revascularization (percutaneous coronary intervention or coronary artery bypass graft operation) were excluded from the study since the original Gensini’s scoring system is technically inappropriate for such cases. The median Gensini score was 23 (5–51, IQR: 46). The patients were divided into groups of severe and non-severe CAD according to this median score. Statistical analysis The normality of distribution was tested using Kolmogorov– Smirnov test. Continuous variables were presented as mean±SD if normally distributed otherwise as median with 25th and 75th percentiles. Categorical variables were presented as numbers and percentages and were compared using chi-square test (Fisher’s exact test if needed). Student’s t-test or Mann–Whitney U test was used for comparison of continuous variables where appropriate. Correlation analysis was performed using Spearman test. A multiple linear regression analysis model was generated in order to investigate the independent predictors of the Gensini score. Receiver operating characteristic (ROC) analysis was
758
Okyay et al. Serum cystatin C and lipocalin
Anatol J Cardiol 2016; 16: 756-61
Table 1. Demographics and baseline clinical parameters of the patients
Age, years Gender, male/female BMI, kg/ m2
Severe CAD (n=26)
Non-severe CAD (n=26)
P
65.7±8.6
64.3±8.4
66.5±9.6
0.08
32/20
19 / 7
13/13
0.15
29.2±3.9 29.2±4.2 29.3±3.6 0.91
Smoking, %
21.2
30.8
11.5
0.17
Hypertension, %
88.5
88.5
88.5
1.00
Hyperlipidemia, %
69.2
73.1
65.4
0.76
23 (5–51)
51 (29–69)
Gensini score HbA1c, %
7.41±1.45
7.66±1.47
7.18±1.43
0.24
BUN, mg/dL
18.63±6.86
19.92±8.28
17.34±4.89
0.18
Scr, mg/dL
0.86±0.17
0.92±0.15
0.80±0.17
0.006
eGFR, mL/min /1.73 m2 88.32±18.70 82.96±19.50 93.69±18.75 0.049 13.76±1.40
13.81±1.56
13.72±1.25
0.80
Total-C, mg/dL
188.81±35.36 190.66±31.80 186.52±40.22 0.73
LDL-C, mg/dL
112.00±30.55 116.44±31.10 107.17±29.86 0.30
HDL-C, mg/dL
40.89±9.64
41.15±10.59
1500.00 1300.00 1100.00 900.00 700.00 500.00 0
40.61±8.76
0.86
Triglycerides, mg/dL 147 (119–196) 150 (132–192) 129 (99–197) 0.20 Medications, % Insulin
23.1
34.6
11.5
0.10
Oral antihypoglycemic agents
86.5
80.8
92.3 0.42
ACEI
28.8
23.1
34.6 0.54
ARB
36.8
42.3
30.8
0.56
CCB
32.7
38.5
26.9
0.55
β-blockers
53.8
61.5
46.2 0.40
Statins
57.7
53.8
61.5
0.78
Acetylsalicylic acid
59.6
65.4
53.8
0.57
Warfarin
3.8
3.8
3.8
–
Nitrates
11.8
15.4
7.7 0.67
Data are presented as mean±standard deviation, median with 25th and 75th percentiles, or number and frequencies where indicated ACEI - angiotensin converting enzyme inhibitor; ARB - angiotensin receptor blocker; BMI - body mass index; BUN - blood urea nitrogen; C - cholesterol; CAD - coronary artery disease; CCB - calcium channel blocker; eGFR - estimated glomerular filtration rate; HbA1c - hemoglobin A1c; HDL-C - high-density lipoprotein cholesterol; LDL-C low-density lipoprotein cholesterol; Scr - serum creatinine Student’s t-test or Mann–Whitney U Test and chi-square test were used for statistical analyses
used to predict the cut off points for cystatin C in determination of multi-vessel disease. Then, sensitivity and specificity were calculated. Statistical analysis was performed by SPSS software (version 17; Statistical Package for Social Sciences, Chicago, IL). A p value of
