Biomarkers in neonates receiving potential nephrotoxic drugs.

OBJECTIVE: Novel biomarkers, such as kidney injury molecule-1 (KIM-1), cystatin, and neutrophil gelatinase-associated lipocalin (NGAL) were shown to predict acute kidney injury (AKI) earlier than serum creatinine in critically ill. We carried out the present study to evaluate these biomarkers in addition to conventional in our neonates. PATIENTS AND METHODS: We recruited 70 neonates of various gestational age groups receiving one or more potential nephrotoxic drug/s. Daily urine samples were collected for estimating KIM-1, cystatin, and NGAL. Modified neonatal kidney disease improving global outcomes (mKDIGO) classification was used in defining AKI. RESULTS: A significant trend in increased urine concentrations of KIM-1, cystatin, and NGAL were observed as we proceed from term to preterm categories. Strong positive correlation was observed between urine albumin and urine albumin creatinine ratio (ACR), and strong negative correlations between urine creatinine and urine cystatin, and between urine creatinine with urine NGAL. A moderate positive correlation was observed between urine KIM-1 and urine cystatin, between urine KIM-1 and urine NGAL, and between urine cystatin and urine NGAL; and a moderate negative correlation was observed between urine creatinine and urine KIM-1. Seven neonates met the mKDIGO criteria for AKI and ROC plot revealed that baseline KIM-1 and NGAL can significantly predict possible drug-induced AKI in neonates. CONCLUSIONS: Urine KIM-1, cystatin, and NGAL are significantly correlated with several other conventional biomarkers that reflect renal function in critically ill neonates. Baseline urine KIM-1 and NGAL concentrations can predict the AKI following potential nephrotoxic drug use in this population.

Early term effect of ureterorenoscopy (URS) on the Kidney: research measuring NGAL, KIM-1, FABP and CYS C levels in urine

ABSTRACT Aim: URS is a very commonly used procedure for treatment of ureter stones. Increased hydrostatic pressure in the collecting system linked to fluids used during the procedure may cause harmful effects on the kidney. The aim of this study is to determine whether the URS procedure has a negative effect on the kidney by investigating NGAL, KIM-1, FABP and Cys C levels in urine. Material and Methods: This study included 30 patients undergoing ureterorenoscopy (URS) for ureter stones. Urine samples were collected 5 times; before the URS procedure (control) and at 1, 3, 5 and 12 hours following the procedure. NGAL, KIM-1, FBAP and Cys C levels were measured in urine and compared with the control values. Results: The NGAL levels in urine before the procedure and at 1, 3, 5 and 12 hours after the procedure were 34.59±35.34; 62.72±142.32; 47.15±104.48; 45.23±163.16 and 44.99±60.79ng/mL, respectively (p=0.001). Similarly, the urinary KIM-1, FABP and Cys C levels were found to increase compared to control values; however this increase did not reach statistical significance (p >0.05). Conclusions: After the URS procedure, there were important changes in NGAL, FABP, KIM-1 and Cys C levels. These changes reached statistical significance for NGAL, but did not reach significance for the other parameters. In conclusion, the URS procedure significantly affects the kidney; however, this effect disappears over time.

Early term effect of ureterorenoscopy (URS) on the Kidney: research measuring NGAL, KIM-1, FABP and CYS C levels in urine.

AIM: URS is a very commonly used procedure for treatment of ureter stones. Increased hydrostatic pressure in the collecting system linked to fluids used during the procedure may cause harmful effects on the kidney. The aim of this study is to determine whether the URS procedure has a negative effect on the kidney by investigating NGAL, KIM-1, FABP and Cys C levels in urine. MATERIAL AND METHODS: This study included 30 patients undergoing ureterorenoscopy (URS) for ureter stones. Urine samples were collected 5 times; before the URS procedure (control) and at 1, 3, 5 and 12 hours following the procedure. NGAL, KIM-1, FBAP and Cys C levels were measured in urine and compared with the control values. RESULTS: The NGAL levels in urine before the procedure and at 1, 3, 5 and 12 hours after the procedure were 34.59±35.34; 62.72±142.32; 47.15±104.48; 45.23±163.16 and 44.99±60.79ng/mL, respectively (p=0.001). Similarly, the urinary KIM-1, FABP and Cys C levels were found to increase compared to control values; however this increase did not reach statistical significance (p >0.05). CONCLUSIONS: After the URS procedure, there were important changes in NGAL, FABP, KIM-1 and Cys C levels. These changes reached statistical significance for NGAL, but did not reach significance for the other parameters. In conclusion, the URS procedure significantly affects the kidney; however, this effect disappears over time.

Is neutrophil gelatinase-associated lipocalin an optimal marker of renal function and injury in liver transplant recipients?

BACKGROUND: Recently, research has focused on the association of neutrophil gelatinase-associated lipocalin (NGAL) with acute and/or active kidney injury. However, it should be remembered that NGAL is involved in iron metabolism and antimicrobial defense mechanisms. METHODS: One hundred seven consecutive liver transplant recipients were included in this study. Plasma and urine NGAL levels were measured with the use of enzyme-linked immunosorbent assay. NGAL levels were studied as plasma concentrations (pNGAL), urine concentrations (uNGAL), urinary NGAL to creatinine ratio (uNGAL/Cr), and fractional NGAL secretion (fNGAL). RESULTS: pNGAL was found to be inversely correlated with estimated glomerular filtration rate (eGFR) and plasma cystatine C (pCysC) (r = -0.26 and P = .007, r = -0.38 and P = .00006, respectively). uNGAL was positively correlated with urinary cystatine C to creatinine ratio (uCysC/Cr) and fractional cystatine C excretion (fCysC) (r = 0.43 and P = .000004; r = 0.4 and P = .1; respectively). uNGAL/Cr was inversely correlated with hematocrit (Htc) and hemoglobin (Hb) (r = -0.35 and P = .0002; r = -0.39 and P = .00004; respectively), and positively correlated with uCysC/Cr (r = 0.5 and P < .0000001). fNGAL was directly correlated with uCysC/Cr and fCysC (r = 0.53 and P < .0000001; r = 0.39 and P = .00005; respectively) and inversely correlated with red blood cell count (RBC; r = -0.31 and P = .001). We observed significant differences of pNGAL, uNGAL/Cr, and fNGAL between sexes, with highest values of uNGAL, uNGAL/Cr, and fNGAL in women and pNGAL in men. In multivariate regression analysis, pNGAL was positively correlated with time elapsed from liver transplantation, neutrophil count, pCysC, and sex (ß = 0.36 and P = .00001; ß = 0.32 and P = .0001; ß = 0.58 and P < .0000001; ß = 0.17 and P = .025; respectively) and inversely correlated with patient's age (ß = -0.18 and P = .02) with R = 0.67 and R(2) = 0.45, independently from blood glucose, eGFR, RBC, white blood cell count, Hb, uCysC, uCysC/Cr, and fCysC. CONCLUSIONS: Plasma and urine NGAL levels are strongly correlated not only with kidney function parameters, but also with red and white blood cell parameters and patient's age and sex.

Method validation for preparing urine samples for downstream proteomic and metabolomic applications.

BACKGROUND: Formal validation of methods for biospecimen processing in the context of accreditation in laboratories and biobanks is lacking. A protocol for processing of a biospecimen (urine) was validated for fitness-for-purpose in terms of key downstream endpoints. METHODS: Urine processing was optimized for centrifugation conditions on the basis of microparticle counts at room temperature (RT) and at 4°C. The optimal protocol was validated for performance (microparticle counts), and for reproducibility and robustness for centrifugation temperature (4°C vs. RT) and brake speed (soft, medium, hard). Acceptance criteria were based on microparticle counts, cystatin C and creatinine concentrations, and the metabolomic profile. RESULTS: The optimal protocol was a 20-min, 12,000 g centrifugation at 4°C, and was validated for urine collection in terms of microparticle counts. All reproducibility acceptance criteria were met. The protocol was robust for centrifugation at 4°C versus RT for all parameters. The protocol was considered robust overall in terms of brake speeds, although a hard brake gave significantly fewer microparticles than a soft brake. CONCLUSIONS: We validated a urine processing method suitable for downstream proteomic and metabolomic applications. Temperature and brake speed can influence analytic results, with 4°C and high brake speed considered optimal. Laboratories and biobanks should ensure these conditions are systematically recorded in the scope of accreditation.

Surface Plasmon Resonance Imaging biosensor for cystatin determination based on the application of bromelain, ficin and chymopapain.

A Surface Plasmon Resonance Imaging (SPRI) sensor based on bromelain or chymopapain or ficin has been developed for specific cystatin determination. Cystatin was captured from a solution by immobilized bromelain or chymopapain or ficin due to the formation of an enzyme-inhibitor complex on the biosensor surface. The influence of bromelain, chymopapain or ficin concentration, as well as the pH of the interaction on the SPRI signal, was investigated and optimized. Sensor dynamic response range is between 0-0.6 µg/ml and the detection limit is equal to 0.1 µg/ml. In order to demonstrate the sensor potential, cystatin was determined in blood plasma, urine and saliva, showing good agreement with the data reported in the literature.

Serum neutrophil gelatinase-associated lipocalin as a marker of renal function in non-diabetic patients with stage 2-4 chronic kidney disease.

The current Kidney Disease Outcomes Quality Initiative (K/DOQI) guidelines advocate creatinine-based equations for estimating GFR to identify patients with potential kidney disease and classify them into different stages due to the fact that serum creatinine is very insensitive to changes in the glomerular filtration rate. Very few biomarkers exist for monitoring chronic kidney disease. The aim of the study was to assess whether NGAL could represent a novel, sensitive marker of kidney function in adult patients with CKD. The study was performed on 92 non-diabetic patients with CKD stages 2-4. Serum and urinary NGAL as well as serum cystatin C were measured using commercially available kits. Serum NGAL was related, in univariate analysis, to serum creatinine, urinary NGAL, hemoglobin, hematocrit, leukocyte count, eGFR, and cystatin C. Urinary NGAL correlated with age, hemoglobin, hematocrit, serum creatinine, and eGFR. In multiple regression analysis, predictors of serum NGAL were creatinine (beta value = 0.97, p = 0.005), cystatin C (beta = 0.34, p = 0.01), and eGFR (beta value = 1.77, p = 0.001). In the healthy volunteers, serum NGAL correlated with age, serum creatinine, eGFR, leukocyte count, and cystatin C. Taking into consideration the fact that the recent DOQI (Dialysis Outcomes Quality Initiative) states that individuals with reduced GRF (glomerular filtration rate) are at greater risk for CVD and cardiac deaths, precise evaluation of renal function is important in order to select the appropriate strategy to reduce the cardiovascular risk. NGAL should be investigated as a potential early and sensitive marker of kidney impairment/injury.

Variation of serum creatinine, cystatin C, and creatinine clearance tests in persons with normal renal function.

BACKGROUND: To determine the potential sensitivity of several renal function tests for detecting early changes in renal function, we compared the within-individual (W-I) variation over 5 months of serum creatinine, serum cystatin C, and creatinine clearance. METHODS: On 31 healthy subjects, blood and timed urine specimens were collected once each month to get 6 collections. Creatinine (enzymatic) in serum and urine and cystatin C (immunonephelometric) in serum were measured and glomerular filtration rate (GFR) by creatinine clearance and the Modification of Diet in Renal Disease (MDRD) equation were calculated. To compare W-I variations between different creatinine methods, we also measured creatinine by both enzymatic and kinetic alkaline picrate methods on 15 sets of frozen samples. RESULTS: For the 31 volunteers, the mean W-I variations for serum creatinine (5.8%) and cystatin C (5.4%) were both much lower than the W-I variation of creatinine clearance (18.7%). As expected, the MDRD GFR had a similar W-I variation (6.7%) to that of serum creatinine and its values were markedly different than GFR by creatinine clearance. On the 15 sets of frozen samples, the W-I variation of creatinine measured by the enzymatic method (CV 5.2%) was slightly less than by the picrate method (CV 6.2%). CONCLUSIONS: The low W-I variation of both serum cystatin C and serum creatinine suggests that serial measurements of either would detect a changes in renal function earlier than would GFR by creatinine clearance or MDRD equation, which allows reporting only for GFRs<60 ml/min/1.7 m(2). While we measured only creatinine clearance, the large variability, difficulty, and cost of all clearance measurements make them impractical for routine monitoring of patients.

[Estimation of the glomerular filtration rate in elderly inpatients: comparison of four methods]. / Quelle méthode d'évaluation de la fonction rénale utiliser chez le sujet âgé hospitalisé en unité de court séjour gériatrique ? Comparaison de quatre méthodes.

PURPOSE: High variability has been observed in the estimation of the glomerular filtration rate (GFR) in older patients, according to the formula used and no single formula has been recommended to date. The aim of this study was to quantify the precision and accuracy of the GFR estimated by means of three formulas and the measurement of cystatin C. METHODS: This prospective study was conducted in an acute care geriatric unit. Participants were patients, aged 70 years and over, having a possible 24-hour urine collection. The GFR was estimated using the Cockroft-Gault (CG), the Modification of Diet in Renal Disease (MDRD), and the Creatinine Clearance (Cl-Cr) formulas. The serum level of cystatin C was also measured. RESULTS: Eighty-one patients were included in the study. CG formula underestimated the GFR by a mean difference of 8.65 ml/min, compared with MDRD formula. Cl-Cr underestimated the GFR by a mean difference of 7.56 ml/min, compared with CG formula, and by a mean difference of 16.79 ml/min, compared with the MDRD formula. The degree of discrepancy between CG and Cl-Cr estimates, and between Cl-Cr and MDRD estimates decreased as the estimated GFR approached normal values. MDRD best matched the measurement of cystatin C, followed by CG and Cl-Cr (Kappa coefficient=0.43, 0.22 and 0.16, respectively). CONCLUSION: Our study confirms the high variability of GFR in older patients and particularly in those with abnormal renal function, depending on the formula used. Serum cystatine C level and MDRD formula appear to be the most concordant estimates of GFR in this population.

Early detection of acute kidney injury: emerging new biomarkers.

Acute kidney injury (AKI) has recently become the preferred term to describe the syndrome of acute renal failure (ARF) with 'failure' or 'ARF' restricted to patients who have AKI and need renal replacement therapy.(1) This allows capture of the broader clinical spectrum of modest reductions in creatinine, which are themselves known to be associated with major increases in both short- and long-term mortality risk.(2-5) It is hoped that this change in nomenclature will facilitate an expansion of our understanding of the underlying pathophysiology and also facilitate definitions of AKI, which allow comparisons among clinical trials of patients with similar duration and severity of illness. This review will cover the need for early detection of AKI and the role of urinary and plasma biomarkers, including enzymuria. The primary message is that use of existing criteria to diagnose AKI, namely elevation of the serum creatinine with or without oliguria, results in identification that is too late to allow successful intervention. New biomarkers are essential to change the dire prognosis of this common condition.

NGAL (neutrophil gelatinase-associated lipocalin) and cystatin C: are they good predictors of contrast nephropathy after percutaneous coronary interventions in patients with stable angina and normal serum creatinine?

The aim of the study was to assess whether NGAL and cystatin C could predict contrast-induced nephropathy in non-diabetic patients (n=60, mean age 60+/-11 years) with normal serum creatinine undergoing elective PCI. We found a significant rise in serum NGAL after 2, 4 and 8 h, and in urinary NGAL after 4, 8 and 24 h after PCI. Cystatin C rose significantly 8 and 24 h after the procedure. Prevalence of CIN was 10%. We found 90% sensitivity and 74% specificity of serum and 76% sensitivity and 80% specificity of urinary NGAL increase. NGAL may represent a sensitive early biomarkers of renal impairment after PCI.

[Assessment of the accuracy and precision of cystatin C-based GFR estimates and Cr-based GFR estimates in comparison with Cr51-EDTA GFR].

BACKGROUND: Cystatin C (cysC) is said to be an ideal marker for glomerular filtration rate (GFR), independent of external factors such as age, nutrition and inflammation. The authors compared the accuracy and precision of cysC-based and creatinine (Cr)-based GFR estimates using Cr51-EDTA GFR method as a reference. METHODS: Serum concentrations of cysC and Cr were measured in adults over 17 yr (n=170) and children below 17 yr (n=79) who had had GFR estimated by Cr51-EDTA method. CysC-based GFR was estimated by the formula of Thierry [CysC-based GFR estimates (mL/min/1.73 m2)=78 x (1/cysC, in mg/L)+4] and Cr-based GFR by the formula of modified Modification of Diet in Renal Disease [MDRD II, Cr-based GFR estimates (mL/min/1.73 m2)=186 x (Scr)(-1.154) x (Age)(-0.203) x 0.742 (for a female patient) x 1.212 (for a black patient). RESULTS: In comparison with Cr51-EDTA GFR, in children below 17 yr, the bias +/- standard deviation (SD) of cysC-based and Cr-based GFR estimates were 7.5 +/- 6.1 and 106.5 +/- 98.2, respectively, in the range of below 90 of Cr51-EDTA GFR (mL/min/1.73 m2), and 33.7 +/- 33.0 and 174.4 +/- 18.8 in the range of over 90. In adults over 17 yr, the respective figures were 13.1 +/- 11.0 and 17.4 +/- 29.8 in below 90, and 21.2 +/- 20.1 and 83.6 +/- 108.8 in over 90 of Cr51-EDTA GFR. CONCLUSIONS: CysC-based GFR estimates show acceptable ranges of biases over the whole age and GFR ranges. CysC-based GFR estimates is considered to be the marker for GFR, which could be used without limitation of age and GFR ranges.

Serum cystatin C-A useful endogenous marker of renal function in intensive care unit patients at risk for or with acute renal failure?

Critically ill patients are at high risk for developing acute renal failure (ARF). The prevention of ARF is of outmost importance in order to improve the increased morbidity and mortality associated with ARF. Unfortunately, there is lack of adequate endogenous markers that can identify renal dysfunction early - this hampers timely application of measures to prevent further renal damage. The use of exogenous markers of renal function is not only time-consuming but also expensive, and therefore can not be used on a regular basis in the intensive care unit. Both the presently used endogenous and exogenous markers are not reliable during continuous renal replacement therapy (CRRT) because these markers are removed by the therapy itself impeding early detection of recovering of renal function. Cystatin C has been proposed as an alternative endogenous marker of renal function for more than 15 years. In this manuscript we review the literature on the role of cystatin C as marker for renal function, focusing on the critically ill patient. Serum cystatin C concentrations have been found to relate to renal impairment and suggest that cystatin C is more sensitive to detect mild decreases in GFR. Cystatin C could be an important tool both to recognize early renal dysfunction and to identify renal recovery while on CRRT in the critically ill patient, however, we are in need of more studies.

Serum and urine cystatin C levels in children with post-pyelonephritic renal scarring: a pilot study.

AIM: We aimed to investigate in children with a history of acute pyelonephritis the influence of unilateral post-pyelonephritic renal scarring detected by DMSA scan on serum (S(CysC)) and urine cystatin C (U(CysC)) as well as upon other traditional markers of renal damage. METHODS: Children with DMSA proven pyelonephritis (n = 28) were grouped as either scar [+] (n = 19, unilateral renal scarring) or scar [-] (no scarring, n = 9). The scar [+] group was further divided into scar-1 (differential DMSA uptake, Delta(DMSA) 10%, n = 11) subgroups. S(CysC), serum creatinine, urine NAG, microalbumin, protein, fractional sodium excretion (FE(Na)), tubular phosphate reabsorption (TPR), and U(CysC/Cr) were evaluated in all patients. RESULTS: Neither S(CysC) nor U(CysC) were affected by age, height, and weight. scar [+] versus scar [-] groups and scar-1 versus scar-2 subgroups were not different with regard to all studied parameters. S(CysC) did not increase in children with post-pyelonephritic unilateral renal scarring. However, 11 children with slightly increased (>0.95 mg/l) S(CysC) levels in scar [+] group tended to have higher Delta(DMSA), albeit not significantly. Furthermore, U(CysC/Cr) correlated well with urine microalbumin, NAG, and FE(Na) in all children and the scar [+] group (P < 0.05). CONCLUSION: S(CysC) and U(CysC) did not differ among pediatric patients with and without unilateral post-pyelonephritic renal scarring. However, Delta(DMSA) uptake between the two kidneys tended to be raised in children with S(CysC) levels higher than the reference ranges. Additionally, U(CysC/Cr) exhibits parallelism with tubular functions.

Increased urinary cystatin C reflects structural and functional renal tubular impairment independent of glomerular filtration rate.

OBJECTIVES: The ratio of the urinary mass concentrations of cystatin C and creatinine (UcysC/Ucrea)>/=11.3 mg/mmol has recently been proposed as an accurate marker for the detection of GFR/=11.3 mg/mmol and factors associated with increased UcysC/Ucrea in 72 children and adults with a wide variety of renal disorders. UcysC/Ucrea was calculated, and GFR wad estimated from serum creatinine and cystatin C by equations. RESULTS: UcysC/Ucrea>/=11.3 mg/mmol had a low diagnostic value to detect GFR values/=11.3 mg/mmol is not an accurate marker to detect GFR

Elevated cystatin C concentration and progression to pre-diabetes: the Western New York study.

OBJECTIVE: We conducted a nested case-control investigation to examine whether elevated baseline concentrations of cystatin C predicted progression from normoglycemia to pre-diabetes over 6 years of follow-up from the Western New York Health Study. RESEARCH DESIGN AND METHODS: In 2002-2004, 1,455 participants from the Western New York Health Study, who were free of type 2 diabetes and known cardiovascular disease at baseline (1996-2001), were reexamined. An incident case of pre-diabetes was defined as an individual with fasting glucose < 100 mg/dl at the baseline examination and > or = 100 and < or = 125 mg/dl at the follow-up examination, thereby eliminating individuals with prevalent pre-diabetics. All case patients (n = 91) were matched 1:3 to control participants based on sex, race/ethnicity, and year of study enrollment. All control subjects had fasting glucose levels < 100 mg/dl at both baseline and follow-up examinations. Cystatin C concentrations and the urinary albumin-to-creatinine ratio were measured from frozen (-196 degrees C) baseline blood and urine samples. Serum creatinine concentrations were available from the baseline examination only. RESULTS: Multivariate conditional logistic regression analyses adjusted for age, baseline glucose level, homeostasis model assessment of insulin resistance, BMI, hypertension, estimated glomerular filtration rate, cigarette smoking, and alcohol use revealed a significantly increased risk of progression to pre-diabetes among those with elevated baseline concentrations of cystatin C (odds ratio 3.28 [95% CI 1.43-7.54]) (upper quintile versus the remainder). Results of secondary analyses that considered high-sensitivity C-reactive protein, interleukin-6, E-selectin, or soluble intercellular adhesion molecule-1 did not alter these results. CONCLUSIONS: These results suggest that cystatin C was associated with a threefold excess risk of progression to pre-diabetes in this population.

Diagnosis of acute kidney injury: from classic parameters to new biomarkers.

A change in serum creatinine is the standard metric used to define and monitor the progression of acute kidney injury (AKI). This marker is inadequate for a number of reasons including the fact that changes in serum creatinine are delayed in time after kidney injury and hence creatinine is not a good indicator to use in order to target therapy in a timely fashion. There is an urgent need for early biomarkers for the diagnosis of AKI. There is also a need for biomarkers that will be predictive of outcome and which can be used to monitor therapy. There are a limited number of biomarkers that are being validated by a number of groups and from this list clinically useful reagents are likely to be derived over the next few years. In this article the status of 5 potential urinary biomarkers for AKI are discussed: kidney injury molecule-1, N-acetyl-Beta-D-glucosaminidase, neutrophil gelatinase-associated lipocalin, cystatin C, and interleukin-18. Considerable progress has been made although much continues to be needed to validate these markers for routine clinical use. Armed with these new tools the future will look much brighter for the patient with AKI as it is likely that early diagnosis and better predictors of outcome will lead to new therapies which can be introduced earlier in the course of disease.

Bias and precision of estimated glomerular filtration rate in children.

Determining true glomerular filtration rate (GFR) using an exogenous marker is time-consuming and cumbersome. Therefore, creatinine-based estimates of GFR are used. Recent papers using new population-specific/local parameters in their prediction equations, standardizing creatinine determination or adding other endogenous surrogate markers of GFR, like cystatin C, could demonstrate an improvement of bias inherent in the results of the prediction equations. Precision, however, is still poor. Currently, we have to accept a precision (as defined in the so-called Bland-Altman plot) of +/-20% in adults and +/-30-40% in children. This problem of poor precision/uncertainty is especially bothering in the higher, near normal GFR range. Caution should be exercised when applying prediction equations in individuals in need of an accurate GFR determination. In that case, a real clearance procedure has to be performed. In the long run, the true clearance procedure should be simplified using new exogenous GFR markers and developing new devices, allowing GFR measurements to be performed, for example, transcutaneously. Such a procedure would be more acceptable for both patients and physicians.