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Serum cystatin C as an index of renal function in kidney transplant patients
Serum Cystatin C as an Index of Renal Function in Kidney Transplant Patients S.H. Akbas, A. Yavuz, M. Tuncer, C. Ruhi, A. Gurkan, R. Cetinkaya, A. Demirbas, M. Gultekin, M. Akaydin, and F. Ersoy ABSTRACT Management of renal transplant patients requires periodic measurement of renal function, which is usually assessed by measuring the glomerular filtration rate (GFR). The most commonly used marker for GFR is serum creatinine, although muscle wasting and tubular secretion may lead to overestimation of the actual GFR. Serum concentrations of the low-molecular-weight proteins, cystatin C and 2-microglobulin (B2M), may afford useful markers to determine a reduced GFR. We investigated whether these molecules provide reliable indicators of renal function in 75 renal transplant patients. Cystatin C and B2M correlated significantly with creatinine (r ⫽ .648, P ⬍ .05 and r ⫽ .578, P ⬍ .05, respectively). Inverse serum creatinine was superior to inverse cystatin C and inverse B2M when renal function equations were used (r ⫽ .95, P ⬍ .05, according to MDRD; r ⫽ .87, P ⬍ .05, according to Cockroft–Gault). Receiver operating characteristic (ROC) analysis was performed to quantitate the accuracy of the different markers to detect reduced GFR using a cutoff value of 70 mL/min. No significant difference between the areas under the ROC curves comparing cystatin C and B2M was observed; however, serum creatinine demonstrated a significantly greater value than cystatin C (.981 vs .724, P ⫽ .001). We conclude that serum creatinine is a more efficacious marker than serum cystatin C to assess renal function.
A
CCURATE and rapid assessment of renal function is crucial in renal transplant recipients. The glomerular filtration rate (GFR) is considered the definitive indicator of renal function. Many methods to determine GFR have been described, including clearance assays using the “gold standards.” of exogenously administered substances such as inulin, 125I-iothalamate, or 51Cr EDTA. However, they are rarely used for routine measurements due to their complexity. Creatinine clearance, which entails a 24-hour urine sample, is a practical approach, but involves patient compliance and a laborious collection. Measuring serum creatinine is the most commonly used method to rapidly assess kidney function, but it also has several disadvantages such as inaccuracies attributable to gender differences, muscle mass changes, or variations in tubular secretion of creatinine.1–3 Measurement of serum concentration of the low-molecular-weight basic proteins cystatin C and 2-microglobulin (B2M) has been proposed as a suitable alternative to detect a reduced GFR. Human cystatin C is a 132-amino-acid, 13-kd cysteine protease inhibitor, which is produced by all © 2004 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710 Transplantation Proceedings, 36, 99⫺101 (2004)
nucleated cells and modulated by intracellular protein catabolism. Its endogenous production rate is unaltered during inflammatory processes.4 Its serum concentration appears to be independent of gender, age, nutrition, medications, or body mass. Furthermore, its renal clearance is similar to that of exogenous substances. It is freely filtered by the glomerulus to be reabsorbed and catabolized in the proximal tubules.1,2,5 Based on these characteristics, cystatin C has been proposed as an indicator of GFR with a higher diagnostic efficiency than serum creatinine.1–5 The purpose of our study was to evaluate the performance of serum cystatin C as a reliable screening marker of From the Central Laboratory (S.H.A., M.G.) and Departments of Nephrology (A.Y., M.T., C.R., R.C., F.E.), and General Surgery (A.G., A.D., M.A.), Akdeniz University Faculty of Medicine, Antalya, Turkey. Address reprint requests to Dr S. Halide Akbas, Central Laboratory, Akdeniz University Faculty of Medicine, Clinical Biochemistry Unit, Antalya, Turkey. E-mail: halideakbas@ akdeniz.edu.tr 0041-1345/04/$–see front matter doi:10.1016/j.transproceed.2003.11.024 99
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reduced GFR in renal transplant patients in comparison with serum creatinine, creatinine clearance, and another low-molecular-weight protein, B2M.
MATERIALS AND METHODS We studied 75 stable renal transplant patients of mean age 34 ⫾ 10.23 years and mean time posttransplantation 17.43 ⫾ 32.02 months. Immunosuppression included cyclosporine or tacrolimus, which in some patients was supplemented with prednisone and MMF. Serum concentrations of cystatin C, B2M, and creatinine were determined from individual blood samples. Creatinine clearance measurements were performed by a 24-hour urine collection. Serum and urine creatinine concentrations were estimated by the modified Jaffe method, using a Roche modular autoanalyzer. Cystatin C and B2M were analyzed by a latex-enhanced immunonephelometric assay on a BNII nephelometer (Dade Behring). Estimation of creatinine clearance was performed according to Cockroft–Gault and MDRD equations. Statistical analyses were performed by MedCalc statistical software (MedCalc software, Mariakerke, Belgium) and SPSS for Windows. Pearson correlation coefficients were calculated for creatinine clearance with reciprocal values of serum cystatin C, creatinine, and B2M. P ⬍ .05 was considered statistically significant. ROC plots were generated to show sensitivity and specificity for serum cystatin C, creatinine, and B2M.
RESULTS
Among 75 transplant patients, the mean serum creatinine concentration was 1.35 ⫾ 0.45 mg/dL and the mean creatinine clearance calculated by the Cockroft–Gault equation was 78.78 ⫾ 26.98 mL/min per 1.73 m2. Using the MDRD, the mean creatinine clearance was 74.28 ⫾ 32.20 mL/min. The mean serum cystatin C concentration was 1.55 ⫾ 0.74 mg/L and the mean serum B2M concentration as 3.42 ⫾ 1.48 mg/L. Cystatin C and B2M both correlated significantly with creatinine (r ⫽ .648, P ⬍ .05 and r ⫽ .578, P ⬍ .05, respectively). Inverse serum creatinine was superior to inverse cystatin C or inverse B2M to estimate renal function by clearance equations (r ⫽ .95, P ⬍ .05 according to MDRD [Fig 1a] and (r ⫽ .87, P ⬍ .05 according to Cockroft–Gault). Inverse cystatin C correlated significantly better with MDRD (r ⫽ .63, P ⬍ .05; Fig 1b) than did estimation of the creatinine clearance according to the Cockroft–Gault formula (r ⫽ .51), whereas inverse B2M showed lower correlation coefficients (not shown). ROC analysis was performed to quantitate the accuracy of different markers to detect a reduced GFR using a cutoff value of 70 mL/min. No significant difference was observed between the areas under the ROC curves for cystatin C and B2M (0.724 and 0.799, respectively; P ⫽ .17 [not shown]). The areas under the ROC curves for serum creatinine and cystatin C were 0.981 and 0.724, respectively (P ⫽ .001). As can be seen in Fig 2, serum creatinine demonstrated a significantly greater area under the curve compared with cystatin C.
Fig 1. Correlations of the reciprocals of serum creatinine (r ⫽ .95, P ⬍ .05) and cystatin C (r ⫽ .63, P ⬍ .05) with creatinine clearance according to MDRD.
DISCUSSION
Sensitive and reliable detection of a reduced GFR is important for management of transplant patients. Because recent studies suggest that low molecular-weight proteins, such as cystatin C and B2M, might provide better markers of GFR than serum creatinine,6,7 we investigated the efficacy of these markers in renal transplant patients. We observed a significant correlation between the levels of serum cystatin C and creatinine. Keevil et al suggested that cystatin C displays a larger intraindividual variation than serum creatinine.3 Risch et al showed that serum creatinine is a better monitoring parameter than cystatin C because it displays lower variability.6 Consistent with these findings, our results reveal that serum creatinine was more efficacious than serum cystatin C and/or B2M to detect a reduced GFR as estimated by creatinine clearance. Hermida et al reported that, in renal transplant patients, impaired glomerular filtration of cystatin C may be due to the formation of cystatin C–immunoglobulin complexes.4 Additional factors that might explain the lower utility of cystatin C include assay interference, backleak of cystatin C into the circulation, or impaired filtration. The other measured low-molecular-weight protein in our study, B2M, did not exhibit
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suppressive medications in accord with reports of Risch et al.6 Finally, we measured serum cystatin C concentrations with an automated latex particle– enhanced immunonephelometric assay, an assay that is more costly than creatinine measurement. Thus, further studies are needed to evaluate the cost-effectiveness of this assay in renal transplant patients. REFERENCES
Fig 2. Nonparametric ROC plots for the diagnostic accuracy of serum concentrations of creatinine (solid line) and cystatin C (dotted line) in reduced GFR (⬍70 mL/min) in 75 renal transplant patients. The areas under the curve for serum creatinine and cystatin C were .981 and .724, respectively (P⫽.001).
higher diagnostic accuracy than cystatin C or creatinine. Serum B2M concentrations may be influenced by immuno-
1. Li FK, Ho SK, Yip TP, et al: Cystatin C assay for the detection of renal dysfunction in Chinese renal transplant patients. Clin Chim Acta 322:133, 2002 2. Risch L, Blumberg A, Huber A: Rapid and accurate assessment of glomerular filtration rate in patients with renal transplants using serum cystatin C. Nephrol Dial Transplant 14:1991, 1999 3. Keevil BG, Kilpatrick ES, Nichols SP, et al: Biological variation of cystatin C implications for the assessment og glomerular filtration rate. Clin Chem 44:1535, 1998 4. Hermida J, Romero R, Tutor JC: Relationship between serum cystatin C and creatinine in kidney and liver transplant patients. Clin Chim Acta 316:165, 2002 5. Le Bricon T, Thervet E, Benlakehal M, et al: Changes in plasma cystatin C after renal transplantation and acute rejection in adults. Clin Chem 45:2243, 1999 6. Risch L, Blumberg A, Huber AR: Assessment of renal function in renal transplant patients using cystatin C. A comparison to other renal function markers and estimates. Renal Failure 23:439, 2001 7. Woitas RP, Stoffel-Wagner B, Poege U, et al: Low molecular weight proteins as marker of glomerular filtration rate. Clin Chem 47:2179, 2001
A
CCURATE and rapid assessment of renal function is crucial in renal transplant recipients. The glomerular filtration rate (GFR) is considered the definitive indicator of renal function. Many methods to determine GFR have been described, including clearance assays using the “gold standards.” of exogenously administered substances such as inulin, 125I-iothalamate, or 51Cr EDTA. However, they are rarely used for routine measurements due to their complexity. Creatinine clearance, which entails a 24-hour urine sample, is a practical approach, but involves patient compliance and a laborious collection. Measuring serum creatinine is the most commonly used method to rapidly assess kidney function, but it also has several disadvantages such as inaccuracies attributable to gender differences, muscle mass changes, or variations in tubular secretion of creatinine.1–3 Measurement of serum concentration of the low-molecular-weight basic proteins cystatin C and 2-microglobulin (B2M) has been proposed as a suitable alternative to detect a reduced GFR. Human cystatin C is a 132-amino-acid, 13-kd cysteine protease inhibitor, which is produced by all © 2004 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710 Transplantation Proceedings, 36, 99⫺101 (2004)
nucleated cells and modulated by intracellular protein catabolism. Its endogenous production rate is unaltered during inflammatory processes.4 Its serum concentration appears to be independent of gender, age, nutrition, medications, or body mass. Furthermore, its renal clearance is similar to that of exogenous substances. It is freely filtered by the glomerulus to be reabsorbed and catabolized in the proximal tubules.1,2,5 Based on these characteristics, cystatin C has been proposed as an indicator of GFR with a higher diagnostic efficiency than serum creatinine.1–5 The purpose of our study was to evaluate the performance of serum cystatin C as a reliable screening marker of From the Central Laboratory (S.H.A., M.G.) and Departments of Nephrology (A.Y., M.T., C.R., R.C., F.E.), and General Surgery (A.G., A.D., M.A.), Akdeniz University Faculty of Medicine, Antalya, Turkey. Address reprint requests to Dr S. Halide Akbas, Central Laboratory, Akdeniz University Faculty of Medicine, Clinical Biochemistry Unit, Antalya, Turkey. E-mail: halideakbas@ akdeniz.edu.tr 0041-1345/04/$–see front matter doi:10.1016/j.transproceed.2003.11.024 99
100
AKBAS¸ , YAVUZ, TUNCER ET AL
reduced GFR in renal transplant patients in comparison with serum creatinine, creatinine clearance, and another low-molecular-weight protein, B2M.
MATERIALS AND METHODS We studied 75 stable renal transplant patients of mean age 34 ⫾ 10.23 years and mean time posttransplantation 17.43 ⫾ 32.02 months. Immunosuppression included cyclosporine or tacrolimus, which in some patients was supplemented with prednisone and MMF. Serum concentrations of cystatin C, B2M, and creatinine were determined from individual blood samples. Creatinine clearance measurements were performed by a 24-hour urine collection. Serum and urine creatinine concentrations were estimated by the modified Jaffe method, using a Roche modular autoanalyzer. Cystatin C and B2M were analyzed by a latex-enhanced immunonephelometric assay on a BNII nephelometer (Dade Behring). Estimation of creatinine clearance was performed according to Cockroft–Gault and MDRD equations. Statistical analyses were performed by MedCalc statistical software (MedCalc software, Mariakerke, Belgium) and SPSS for Windows. Pearson correlation coefficients were calculated for creatinine clearance with reciprocal values of serum cystatin C, creatinine, and B2M. P ⬍ .05 was considered statistically significant. ROC plots were generated to show sensitivity and specificity for serum cystatin C, creatinine, and B2M.
RESULTS
Among 75 transplant patients, the mean serum creatinine concentration was 1.35 ⫾ 0.45 mg/dL and the mean creatinine clearance calculated by the Cockroft–Gault equation was 78.78 ⫾ 26.98 mL/min per 1.73 m2. Using the MDRD, the mean creatinine clearance was 74.28 ⫾ 32.20 mL/min. The mean serum cystatin C concentration was 1.55 ⫾ 0.74 mg/L and the mean serum B2M concentration as 3.42 ⫾ 1.48 mg/L. Cystatin C and B2M both correlated significantly with creatinine (r ⫽ .648, P ⬍ .05 and r ⫽ .578, P ⬍ .05, respectively). Inverse serum creatinine was superior to inverse cystatin C or inverse B2M to estimate renal function by clearance equations (r ⫽ .95, P ⬍ .05 according to MDRD [Fig 1a] and (r ⫽ .87, P ⬍ .05 according to Cockroft–Gault). Inverse cystatin C correlated significantly better with MDRD (r ⫽ .63, P ⬍ .05; Fig 1b) than did estimation of the creatinine clearance according to the Cockroft–Gault formula (r ⫽ .51), whereas inverse B2M showed lower correlation coefficients (not shown). ROC analysis was performed to quantitate the accuracy of different markers to detect a reduced GFR using a cutoff value of 70 mL/min. No significant difference was observed between the areas under the ROC curves for cystatin C and B2M (0.724 and 0.799, respectively; P ⫽ .17 [not shown]). The areas under the ROC curves for serum creatinine and cystatin C were 0.981 and 0.724, respectively (P ⫽ .001). As can be seen in Fig 2, serum creatinine demonstrated a significantly greater area under the curve compared with cystatin C.
Fig 1. Correlations of the reciprocals of serum creatinine (r ⫽ .95, P ⬍ .05) and cystatin C (r ⫽ .63, P ⬍ .05) with creatinine clearance according to MDRD.
DISCUSSION
Sensitive and reliable detection of a reduced GFR is important for management of transplant patients. Because recent studies suggest that low molecular-weight proteins, such as cystatin C and B2M, might provide better markers of GFR than serum creatinine,6,7 we investigated the efficacy of these markers in renal transplant patients. We observed a significant correlation between the levels of serum cystatin C and creatinine. Keevil et al suggested that cystatin C displays a larger intraindividual variation than serum creatinine.3 Risch et al showed that serum creatinine is a better monitoring parameter than cystatin C because it displays lower variability.6 Consistent with these findings, our results reveal that serum creatinine was more efficacious than serum cystatin C and/or B2M to detect a reduced GFR as estimated by creatinine clearance. Hermida et al reported that, in renal transplant patients, impaired glomerular filtration of cystatin C may be due to the formation of cystatin C–immunoglobulin complexes.4 Additional factors that might explain the lower utility of cystatin C include assay interference, backleak of cystatin C into the circulation, or impaired filtration. The other measured low-molecular-weight protein in our study, B2M, did not exhibit
CYSTATIN C AS INDEX OF FUNCTION
101
suppressive medications in accord with reports of Risch et al.6 Finally, we measured serum cystatin C concentrations with an automated latex particle– enhanced immunonephelometric assay, an assay that is more costly than creatinine measurement. Thus, further studies are needed to evaluate the cost-effectiveness of this assay in renal transplant patients. REFERENCES
Fig 2. Nonparametric ROC plots for the diagnostic accuracy of serum concentrations of creatinine (solid line) and cystatin C (dotted line) in reduced GFR (⬍70 mL/min) in 75 renal transplant patients. The areas under the curve for serum creatinine and cystatin C were .981 and .724, respectively (P⫽.001).
higher diagnostic accuracy than cystatin C or creatinine. Serum B2M concentrations may be influenced by immuno-
1. Li FK, Ho SK, Yip TP, et al: Cystatin C assay for the detection of renal dysfunction in Chinese renal transplant patients. Clin Chim Acta 322:133, 2002 2. Risch L, Blumberg A, Huber A: Rapid and accurate assessment of glomerular filtration rate in patients with renal transplants using serum cystatin C. Nephrol Dial Transplant 14:1991, 1999 3. Keevil BG, Kilpatrick ES, Nichols SP, et al: Biological variation of cystatin C implications for the assessment og glomerular filtration rate. Clin Chem 44:1535, 1998 4. Hermida J, Romero R, Tutor JC: Relationship between serum cystatin C and creatinine in kidney and liver transplant patients. Clin Chim Acta 316:165, 2002 5. Le Bricon T, Thervet E, Benlakehal M, et al: Changes in plasma cystatin C after renal transplantation and acute rejection in adults. Clin Chem 45:2243, 1999 6. Risch L, Blumberg A, Huber AR: Assessment of renal function in renal transplant patients using cystatin C. A comparison to other renal function markers and estimates. Renal Failure 23:439, 2001 7. Woitas RP, Stoffel-Wagner B, Poege U, et al: Low molecular weight proteins as marker of glomerular filtration rate. Clin Chem 47:2179, 2001