Circulating beta-hydroxybutyrate levels in advanced heart failure with reduced ejection fraction: Determinants and prognostic impact.

AIMS: Patients with heart failure (HF) display metabolic alterations, including heightened ketogenesis, resulting in increased beta-hydroxybutyrate (ß-OHB) formation. We aimed to investigate the determinants and prognostic impact of circulating ß-OHB levels in patients with advanced HF and reduced ejection fraction (HFrEF). METHODS AND RESULTS: A total of 867 patients with advanced HFrEF (age 57 ± 11 years, 83% male, 45% diabetic, 60% New York Heart Association class III), underwent clinical and echocardiographic examination, circulating metabolite assessment, and right heart catheterization (n = 383). The median ß-OHB level was 64 (interquartile range [IQR] 33-161) µmol/L (normal 0-74 µmol/L). ß-OHB levels correlated with increased markers of lipolysis (free fatty acids [FFA]), higher natriuretic peptides, worse pulmonary haemodynamics, and lower humoral regulators of ketogenesis (insulin/glucagon ratio). During a median follow-up of 1126 (IQR 410-1781) days, there were 512 composite events, including 324 deaths, 81 left ventricular assist device implantations and 107 urgent cardiac transplantations. In univariable Cox regression, increased ß-OHB levels (T3 vs. T1: hazard ratio [HR] 1.39, 95% confidence interval [CI] 1.13-1.72, p = 0.002) and elevated FFA levels (T3 vs. T1: HR 1.39, 95% CI 1.09-1.79, p = 0.008) were both predictors of a worse prognosis. In multivariable Cox analysis evaluating the simultaneous associations of FFA and ß-OHB levels with outcomes, only FFA levels remained significantly associated with adverse outcomes. CONCLUSIONS: In patients with advanced HFrEF, increased plasma ß-OHB correlate with FFA levels, worse right ventricular function, greater neurohormonal activation and other markers of HF severity. The association between plasma ß-OHB and adverse outcomes is eliminated after accounting for FFA levels, suggesting that increased ß-OHB is a consequence reflecting heightened lipolytic state, rather than a cause of worsening HF.

How to verify the analytical and clinical performance of ELISA immunoanalysis in the real laboratory practice. PCSK9 as an example.

BACKGROUND: Manufacturers and diagnostic companies often recommend on-site verification of analytical performance in the clinical laboratory. The validation process used by manufacturers is rarely described in detail, and certain information on analytical performance is missing from the product sheet, especially for immunoanalytical methods. We describe an approach to the detailed validation of an ELISA method for the measurement of proprotein convertase subtilisin/kexin type 9 (PCSK9) plasma concentrations. We compared manufacturers' claims of analytical performance with data obtained in the field laboratory using several approaches. METHODS: We used the Human Proprotein Convertase 9/PCSK9 Quantikine ELISA diagnostic kit (R&D systems, Bio-Techne Ltd., Abingdon Science Park, Abingdon, UK) and three levels of quality control solution Quantikine Immunoassay Control Group 235 (R&D systems, Bio-Techne Ltd., Abingdon Science Park, Abingdon, UK) to verify precision. We measured the concentration of PCSK9 using the DS2 ELISA Reader (Dynex Technologies GmbH, Denkendorf, Germany). We used analysis of variance (ANOVA) and the R statistical package (R core team, version 1.4.5). Statistical analysis and terminology were performed according to protocol CLSI EP15-A3, and the reference interval was checked according to CLSI/IFCC C28-A3c. RESULTS: We found a significant difference between the manufacturer's claims of analytical performance and real data measured in the routine clinical laboratory. The calculated CV (%) for repeatability (calculated by simple estimation of the mean and SD, as used by the manufacturer) was between 5.5% and 7.4%, but the manufacturer's claim was between 4.1% and 6.5%. Using ANOVA, the true repeatability was between 5.0% and 6.9%. Similarly, ANOVA revealed values of CV (%) for within-laboratory imprecision between 6.5% and 9.1%, while the manufacturer's claims were between 4.1% and 5.9%. The recovery ranged from 105.5% to 121.8%. The manufacturer's recommended reference interval was checked and we didn't find any significant difference between men and women. CONCLUSIONS: We describe a comprehensive approach to verify the analytical performance of an ELISA method using the measurement of PCSK9 plasma concentration as an example. We found differences between the results of this approach based on the CLSI EP15-A3 protocol and data provided by the manufacturer. We recommend the verification of analytical performance by more complex statistical tools in laboratory practice.

Biological variation of PIVKA-II in blood serum of healthy subjects measured by automated electrochemiluminescent assay.

Background: Prothrombin/Protein Induced by Vitamin K Absence-II (PIVKA-II) is a candidate biomarker of hepatocellular cancer, recommended both for diagnostics and monitoring. The aim was to evaluate biological variation (BV) of serum PIVKA-II. Methods: Within-subject (CVI) and between-subject (CVG) BV estimates were assessed in 14 healthy volunteers in a 6-week protocol. Serum concentrations of PIVKA-II were measured by a Roche Elecsys PIVKA-II diagnostic kit (cobas e8000). Precision (CVA) was assessed from duplicate measurements of all volunteers' samples. Two methods were used for the estimation of CVI: SD-ANOVA and CV-ANOVA method. We calculated the index of individuality (II) and reference change value. The experiment was fully compliant with EFLM database checklist. Results: The CVI of PIVKA-II in healthy persons, as calculated by two statistical methods, were 8.2% (SD-ANOVA with CVA of 3.2%) and 9.4% (CV-ANOVA) with CVA of 2.7%). The CVG was 19.5% (SD-ANOVA), and respective II and RCV were 0.42 and 24.4%. Conclusions: CVI and CVG of PIVKA-II were 8.2% and 19.5%, respectively, with CVA below 4%. The low II and RCV below 25% enable the use of this biomarker both for diagnostics and monitoring. More data are needed before the introduction of PIVKA-II into clinical practice.

Accuracy and Feasibility of Real-time Continuous Glucose Monitoring in Critically Ill Patients After Abdominal Surgery and Solid Organ Transplantation.

OBJECTIVE: Glycemia management in critical care is posing a challenge in frequent measuring and adequate insulin dose adjustment. In recent years, continuous glucose measurement has gained accuracy and reliability in outpatient and inpatient settings. The aim of this study was to assess the feasibility and accuracy of real-time continuous glucose monitoring (CGM) in ICU patients after major abdominal surgery. RESEARCH DESIGN AND METHODS: We included patients undergoing pancreatic surgery and solid organ transplantation (liver, pancreas, islets of Langerhans, kidney) requiring an ICU stay after surgery. We used a Dexcom G6 sensor, placed in the infraclavicular region, for real-time CGM. Arterial blood glucose measured by the amperometric principle (ABL 800; Radiometer, Copenhagen, Denmark) served as a reference value and for calibration. Blood glucose was also routinely monitored by a StatStrip bedside glucose meter. Sensor accuracy was assessed by mean absolute relative difference (MARD), bias, modified Bland-Altman plot, and surveillance error grid for paired samples of glucose values from CGM and acid-base analyzer (ABL). RESULTS: We analyzed data from 61 patients and obtained 1,546 paired glucose values from CGM and ABL. Active sensor use was 95.1%. MARD was 9.4%, relative bias was 1.4%, and 92.8% of values fell in zone A, 6.1% fell in zone B, and 1.2% fell in zone C of the surveillance error grid. Median time in range was 78%, with minimum (<1%) time spent in hypoglycemia. StatStrip glucose meter MARD compared with ABL was 5.8%. CONCLUSIONS: Our study shows clinically applicable accuracy and reliability of Dexcom G6 CGM in postoperative ICU patients and a feasible alternative sensor placement site.

Significant elevations of serum amylase caused by macroamylase: Case reports and detection possibilities.

BACKGROUND: The presence of isolated elevated serum amylase levels can be caused by high molecular mass complexes. We describe 13 cases of hyperamylasemia detected in adult patients without clinical symptoms of a pancreatic disorder. Five of them were thoroughly examined using different tools for the detection of macrocomplexes. METHODS: We performed both screening and more advanced methods of macroamylase detection, including polyethylene glycol precipitation, sample storage at 4°C and separation by gel filtration. RESULTS: The presence of macroamylase in the suspected samples was confirmed by the methods described, except for the sample storage at 4°C. In this method, the enzyme activity did not decrease. The polyethylene glycol precipitation activity (% PPA) averaged 89.1% for amylase, whereas the control samples averaged 30.7%. Gel filtration chromatography confirmed an IgA macroamylase peak in three samples and an IgG macroamylase peak in two samples. CONCLUSION: The presence of macroamylase should be suspected whenever the clinical history and condition of the patient do not match the measured enzyme value to avoid diagnostic errors and unnecessary invasive examinations. The presence of macrocomplexes is considered a benign process that may occur in apparently healthy individuals. Cooperation between clinicians and laboratory staff is necessary.

Galectin-3 as an independent prognostic factor after heart transplantation.

BACKGROUND: Galectin-3 (GAL3) is linked to the prognosis of patients with heart failure and after heart transplantation (HTx). We assessed the prognostic role of GAL3 in a long-term follow-up after HTx. METHODS: HTx patients (N = 121) were evaluated in a single-center, noninterventional, prospective, observational study. The median follow-up was 96 months (2942 days, interquartile range (IQR) 2408-3264 days), and 40 patients died. GAL3 was measured before HTx, +10 days after HTx, and during the first posttransplant year. Survival analysis (all-cause mortality) was performed with adjustments for clinical and laboratory variables. RESULTS: The median pretransplant GAL3 level was 18.0 µg/L (IQR 14.0-25.9), and higher values were associated with older age, worse kidney function, left ventricular assist device use before HTx, a higher IMPACT score, and mortality. Increased pretransplant GAL3 predicted shorter survival time (HR 2.05, 95% CI 1.09-3.85, p < .05). Similar prognostic power had GAL3 on the 10th posttransplant day (HR 2.03, 95% CI 1.08-3.82, p < .05). GAL3 was an independent predictor of death after adjustment for clinical variables (age, infection, diabetes, smoking, IMPACT score, and troponin). CONCLUSIONS: GAL3 was significantly associated with all-cause mortality after adjusting for clinical and laboratory variables and may serve as an additional prognostic biomarker.

Biological variation of proprotein convertase subtilisin/kexin type 9 (PCSK9) in human serum.

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is involved in the regulation of LDL receptors. Inhibition of PCSK9 increase uptake of LDL-particles and pathogen-associated molecular patterns (PAMPs). The aim of our study was to evaluate biological variation of serum PCSK9. METHODS: Within-subject (CVI) and between-subject (CVG) biological variations were assessed in 14 healthy volunteers in a 6-week protocol (7 samples, equidistant time intervals). Serum concentration of PCSK9 was measured by a Quantikine ELISA assay (R&D systems, Bio-Techne Ltd., UK) on a DS2 ELISA reader (Dynex Technologies GmbH, Germany). Precision (CVA) was assessed by duplicate measurements. Two methods with different levels of robustness were used for the estimation of CVI, SD-ANOVA and CV-ANOVA method. We calculated the index of individuality and reference change values. The experiment was fully compliant with EFLM database checklist. RESULTS: The within-subject values of PCSK9 in healthy persons, as calculated by two statistical methods, were 23.2% (SD-ANOVA with CVA of 5.6%) and 26.6% (CV-ANOVA with CVA of 4.8%). The CVG was 10.9% (SD-ANOVA), index of individuality and RCV were 2.13 and 66.3%, respectively. CONCLUSIONS: The high index of individuality indicates that common reference intervals can be used to interpret serum PSCK9 values.

Very low lipoprotein(a) and increased mortality risk after myocardial infarction.

BACKGROUND: Inconclusive data exist on risk associated with Lp(a) in patients after myocardial infarction (MI). Aims of the present study were to evaluate the association of Lp(a) level with total mortality and recurrent cardiovascular events. DESIGN AND METHODS: Single center prospective registry of consecutive patients hospitalized for acute myocardial infarction between June 2017 and June 2020 at a large tertiary cardiac center with available blood samples drawn <24h of admission. RESULTS: Data from 851 consecutive patients hospitalized for MI were evaluated. During the median follow-up of 19 months (interquartile range 10-27), 58 (6.8%) patients died. Nonlinear modelling revealed a U-shaped association between Lp(a) and total mortality risk. Compared to patients with Lp(a) ranging between 10-30 nmol/L and after multivariate adjustment, total mortality risk was increased both in patients with Lp(a)<7 nmol/L (hazard ratio (HR) 4.08, 95% confidence interval (CI) 1.72-9.68) and Lp(a) ≥125 nmol/L (HR 2.92, 95% CI 1.16-7.37), respectively. Similarly, the risk of combined endpoint of acute coronary syndrome recurrence or cardiovascular mortality was increased both in patients with low (sub-HR 2.60, 95% CI 1.33-5.08) and high (sub-HR 2.10, 95% CI 1.00-4.39) Lp(a). Adjustment for heart failure signs at the time of hospitalization weakened the association with total mortality and recurrent cardiovascular events. CONCLUSIONS: In the present analysis, both high and low concentrations of Lp(a) were associated with an increased risk of total mortality and recurrent cardiovascular events after MI. The excess of mortality associated with Lp(a) was partially attributable to more prevalent heart failure.

Changes in Sepsis Biomarkers after Immunosuppressant Administration in Transplant Patients.

Sepsis biomarkers change continuously during the postoperative period. We aimed to demonstrate the influence of immunosuppressants after transplantation (Tx) on presepsin, procalcitonin, CRP, white blood cells, and IL-6. A group of 140 patients after major surgery (86 non-Tx, 54 Tx) without any signs of sepsis or infectious complications was followed for 7 days. The changes in biomarkers were analyzed with respect to the type of surgery, organ, and induction immunosuppressant used (antithymocyte globulin, corticosteroids, or basiliximab/rituximab). Concentrations (95th percentiles) of presepsin and procalcitonin were higher in the Tx group (presepsin: Tx < 2380 vs. non-Tx < 1368 ng/L, p < 0.05; procalcitonin: <28.0 vs. 3.49 µg/L, p < 0.05). In contrast, CRP and IL-6 were lower in the Tx group (CRP: Tx < 84.2 vs. non-Tx < 229 mg/L, p < 0.05; IL-6: <71.2 vs. 317 ng/L, p < 0.05). Decreases in CRP and IL-6 were found for all immunosuppressants, and procalcitonin was increased after antithymocyte globulin and corticosteroids. Negligible changes were found for white blood cells. Different responses of presepsin, procalcitonin, CRP, and IL-6 were therefore found in patients without any infectious complications after major surgery or transplantation. Immunosuppression decreased significantly IL-6 and CRP in comparison to non-Tx patients, while procalcitonin was increased after corticosteroids and antithymocyte globulin only. Cautious interpretation of sepsis biomarkers is needed in the early posttransplant period. This work was conducted as a noninterventional (nonregistered) study.

The effect of three major co-morbidities on quality of life and outcome of patients with heart failure with reduced ejection fraction.

AIMS: Diabetes mellitus, chronic obstructive pulmonary disease, and chronic kidney disease are prevalent in patients with heart failure with reduced ejection fraction (HFrEF). We have analysed the impact of co-morbidities on quality of life (QoL) and outcome. METHODS AND RESULTS: A total of 397 patients (58.8 ± 11.0 years, 73.6% with New York Heart Association functional class ≥3) with stable advanced HFrEF were followed for a median of 1106 (inter-quartile range 379-2606) days, and 68% of patients (270 patients) experienced an adverse outcome (death, urgent heart transplantation, and implantation of mechanical circulatory support). Chronic obstructive pulmonary disease was present in 16.4%, diabetes mellitus in 44.3%, and chronic kidney disease in 34.5% of patients; 33.5% of patients had none, 40.0% had one, 21.9% had two, and 3.8% of patient had three co-morbidities. Patients with more co-morbidities reported similar QoL (assessed by Minnesota Living with Heart Failure Questionnaire, 45.46 ± 22.21/49.07 ± 21.69/47.52 ± 23.54/46.77 ± 23.60 in patients with zero to three co-morbidities, P for trend = 0.51). Multivariable regression analysis revealed that furosemide daily dose, systolic blood pressure, New York Heart Association functional class, and body mass index, but not the number of co-morbidities, were significantly (P < 0.05) associated with QoL. Increasing co-morbidity burden was associated with worse survival (P < 0.0001), lower degree of angiotensin-converting enzyme inhibitor/angiotensin receptor blocker treatment (P = 0.001), and increasing levels of BNP (mean of 685, 912, 1053, and 985 ng/L for patients with zero to three co-morbidities, P for trend = 0.008) and cardiac troponin (sm-cTnI, P for trend = 0.0496), which remained significant (P < 0.05) after the adjustment for left ventricular ejection fraction, left ventricular end-diastolic diameter, right ventricular dysfunction grade, body mass index, and estimated glomerular filtration rate. CONCLUSIONS: In stable advanced HFrEF patients, co-morbidities are not associated with impaired QoL, but negatively affect the prognosis both directly and indirectly through lower level of HF pharmacotherapy and increased myocardial stress and injury.

Myocardial ketone body utilization in patients with heart failure: The impact of oral ketone ester.

AIMS: Upregulation of ketone body (ß-hydroxybutyrate, ßHB) utilization has been documented in human end-stage heart failure (HF), but is unclear if this is due to intrinsic cardiac metabolic remodeling or a HF-related catabolic state. This study sought to evaluate the maximal ketone body utilization capacity and its determinants in controls and in patients with moderate HF and reduced ejection fraction (HFrEF). METHODS AND RESULTS: 19 HFrEF patients and 9 controls underwent sampling from the arterial circulation (A) and coronary sinus (CS) to measure transmyocardial extraction of energy-providing substrates and oxygen. In a separate experiment, measurements were performed 80-min after oral administration of 25 g of ketone ester (KE, (R)-3-hydroxybutyl(R)-3-hydroxybutyrate) drink in 11 HFrEF and 6 control subjects. There were no statistically significant differences in fasting substrate levels and fractional extractions between HF and controls. Administration of KE increased ßHB by 12.9-fold, revealing an increased ability to utilize ketones in HFrEF as compared to controls (fractional extraction, FE%: 52 vs 39%, p = 0.035). ßHB FE% correlated directly with ßHB myocardial delivery (r = 0.90), LV mass (r = 0.56), LV diameter (r = 0.65) and inversely with LV EF (-0.59) (all p < 0.05). ßHB FE% positively correlated with lactate FE% (p < 0.01), but not with FFA or glucose FE%, arguing against substrate competition. CONCLUSIONS: Acute nutritional ketosis enhances ßHB extraction in patients with HFrEF compared to controls, and this enhancement correlates with degree of cardiac dysfunction and remodeling. Data suggest that subclinical metabolic remodeling occurs early in HF progression. Further studies are needed to determine whether exogenous ketones may have a potential therapeutic role.

Spironolactone-furosemide combination therapy and acid-base disorders in liver cirrhosis patients
.

OBJECTIVE: Respiratory alkalosis (RA) and dilutional hyperchloremic acidosis (DHA) are the most common acid-base balance (ABB) disorders in patients with liver cirrhosis. The aims of this study were to clarify whether RA develops in relation to DHA via respiratory compensation of metabolic acidosis and whether spironolactone in combination with low-dose furosemide - diuretics known to ameliorate DHA - positively affects RA in liver cirrhosis patients. MATERIALS AND METHODS: 59 patients with advanced cirrhosis were divided into two groups. Group D consisted of individuals (urine sodium concentration (UNa+) > 20 mmol/L) who responded to combination therapy consisting of spironolactone and low-dose furosemide. The non-D group consisted of individuals (UNa+ ≤ 20 mmol/L) who either did not respond to the treatment or who were not administered it. In both groups, we examined serum and urine concentrations of electrolytes and ABB parameters, including SNa+-SCl- and SNa+/SCl- values. RESULTS: In group D, we found a statistically significant relationship between pCO2 and SHCO3-: r = 0.756 (p < 0.001) and between pCO2 and SNa+-SCl-: r = 0.522 (p = 0.001). Neither Salb nor the corrected anion gap were associated with changes in SHCO3- or pCO2 values. Although SHCO3- values were normal, abnormal pCO2 values were observed in one third of group D patients. Based on multivariable analysis, SHCO3- proved to be a statistically significant influencing factor on pCO2 values. CONCLUSION: DHA contributes to the development of RA in individuals with liver cirrhosis. Reducing DHA by means of effective diuretic therapy comprising spironolactone and furosemide has a beneficial effect on RA in such patients.

Presepsin: what is important for correct interpretation.

INTRODUCTION: Presepsin (soluble CD14 subtype) is a new biomarker of infection and sepsis. Correct interpretation is based on the knowledge of analytical reliability, biological variation, decision limits, and diagnostic effectivity. AIM: The aim of the study was to verify analytical precision of presepsin measurements, to assess biological variation in healthy subjects, to verify reference and decision limits, to assess diagnostic effectivity, and to compare data with commonly used septic biomarkers - procalcitonin (PCT), CRP and interleukin 6 (IL6). MATERIAL AND METHODS: Analyti-cal precision (repeatability and intermediate precision) was estimated by repeated measurements of commercial control materials. Biological variation was evaluated in a group of 20 healthy volunteers in a 7-week experi-ment. Reference ranges were extracted from the literature and compared with data from healthy subjects. RESULTS: Precisions of presepsin measurements were 2.0-4.0 % (“within-run”) and 6.1-9.5 % (“between-run”). Intraindividual biological variation of presepsin was 22.3 %, interindividual variation 20.8 %. Index of individuality was 1.07, reference change value (RCV - critical difference) was 72 %. Upper reference limit was around 180 ng/l. CONCLUSION: Ana-lytical quality of presepsin measurement is suitable for clinical use. Biological variation parameters enable the use of reference limits, upper reference limit of presepsin is around 180 ng/l. None of the tested biomarkers has universal cut-off value, multiple biomarkers are needed and repeated measurements are advisable.

Immunoglobulin abnormalities in 1677 solid organ transplant recipients. Implications for posttransplantation follow-up.

BACKGROUND: Posttransplant lymphoproliferative disorder (PTLD) is a severe complication of solid organ transplantation (SOT). However, there is no consensus on PTLD screening methods. Gammopathies (GP), which occur in 10-25% of SOT recipients, have been linked to subsequent development of PTLD. Therefore, GP detection methods, such as serum protein electrophoresis (SPE), serum protein immunofixation (SIFE), urine protein immunofixation (UIFE) and the quantitative measurement of serum free light chains (SFLC) are candidate methods for PTLD screening. OBJECTIVE: We aimed to assess the frequency of PTLD and GP, association of GP with subsequent PTLD, allograft loss or death and the diagnostic performance of SPE/SIFE in PTLD screening. The main objective was to explore, whether GP detection methods can be used to enhance the efficiency of PTLD screening and to formulate a concise algorithm for posttransplantation (post-Tx) follow-up. METHODS: We performed a cohort study on 1677 SOT recipients with SPE/SIFE data who underwent kidney, liver, heart, pancreas, Langerhans islets or multiple organ transplantation at the Institute of Clinical and Experimental Medicine between 1966 and 2015. The median (IQR) of follow-up time was 8.0 (4.0-12.0) years. RESULTS: The frequencies of PTLD and GP in SOT recipients were 2.8% and 6.4%, respectively. The frequencies of transient GP, GP of undetermined significance and malignant GP were 33%, 63% and 4% respectively. The median time between SOT and GP detection was 2.0 (interquartile range 1.0-7.0) years. GP was associated with a significantly higher risk of PTLD, allograft loss and death, with hazard ratios (95% confidence intervals) of a 6.06 (2.51-14.64), 2.61 (1.49-4.6) and 1.99 (1.2-3.3), respectively. Additionally, GP was associated with 2.98-fold increased risk of allograft loss in kidney transplant patients. SPE diagnostic sensitivity and specificity for PTLD were 14.8% and 93.9%, respectively. PTLD was diagnosed more often and earlier if SPE/SIFE was included in the post-Tx follow-up. CONCLUSIONS: GP after SOT is associated with a high risk of PTLD, allograft loss and poor survival. The combination of SPE, SIFE, SFLC and UIFE is optimal for GP detection. These methods aid in identifying patients who are at risk for PTLD or allograft damage and should be included in regular post-Tx follow-up.

The Role of GDF-15 in Heart Failure Patients With Chronic Kidney Disease.

BACKGROUND: Growth differentiation factor-15 (GDF-15) is a stress-inducible cytokine and member of the transforming growth factor-ß cytokine superfamily that refines prognostic assessment in subgroups of patients with heart failure (HF). We evaluated its role in HF patients with chronic kidney disease (CKD, estimated glomerular filtration rate <60 mL/min/1.73 m2). METHODS: A total of 358 patients with stable systolic HF were followed for a median of 1121 (interquartile range, 379-2600) days. Comprehensive evaluation including B-type natriuretic peptide (BNP) and GDF-15 testing was performed at study entry; the analysis was stratified according to kidney function. RESULTS: Patients with CKD (33.8%) were older, had more often diabetes, and were less often treated with angiotensin converting enzyme inhibitors (ACEi)/angiotensin receptor blockers (ARB). GDF-15 was associated with estimated glomerular filtration rate, whereas BNP was associated with left ventricular-end diastolic diameter and ejection fraction (P < 0.01). During follow-up, 244 patients (68.2%) experienced an adverse outcome (death, urgent transplantation, implantation of mechanical circulatory support). In patients with HF and CKD, the Cox proportional hazard model identified BNP, GDF-15, sex, systolic blood pressure, sodium, total cholesterol, and ACEi/ARB treatment as significant variables associated with an adverse outcome (P < 0.05). In multivariable analysis, BNP was replaced by GDF-15. Net reclassification improvement confirmed prognostic superiority of the model encompassing GDF-15 (GDF-15, sodium, total cholesterol, ACEi/ARB treatment) compared with the model without GDF-15 (BNP, sex, sodium, ACEi/ARB treatment), net reclassification improvement 0.62, P = 0.005. In contrast, in patients with HF and normal kidney function, BNP remained superior to GDF-15 in a multivariable model. CONCLUSIONS: In patients with systolic HF and CKD, GDF-15 is more strongly associated with adverse outcomes than the conventionally used BNP.

Biological variation of intact fibroblast growth factor 23 measured on a fully automated chemiluminescent platform.

BACKGROUND: Fibroblast growth factor 23 (FGF23), a potent regulator of phosphate and vitamin D metabolism, is a new biomarker of kidney, bone and cardiovascular disorders. The aim of this study was to assess the biological variation of intact fibroblast growth factor 23 (iFGF23). METHODS: The within-subject (CVI) and between-subject (CVG) biological variations were assessed in 14 healthy volunteers in a six-week protocol (seven samples). Imprecision (CVA) was assessed by duplicate measurements and the EP15-A2 protocol. Intact FGF23 was measured using a fully automated chemiluminescent assay (Liaison XL, DiaSorin S.p.A., Saluggia, Italy). Two methods with different sensitivities to non-Gaussian distribution were used to estimate the CVI, SD ANOVA and CV ANOVA methods. We calculated the index of individuality (II) and reference change values. RESULTS: Depending on the statistical method used, the CVI and CVA were 14.2 and 3.7% (SD ANOVA) or 12.5 and 3.9% (CV ANOVA), respectively. The corresponding reference change values were 40.5 and 36.4%, respectively. The CVG was 13.4% (SD ANOVA was the only option), and the total imprecision (EP15-A2) was less than 7%. CONCLUSIONS: The measurement of iFGF23 demonstrated a CVA less than 4% during the experimental estimation of biological variation. The total imprecision was less than 7% in the EP15-A2 experiment. The CVI values of iFGF23 in healthy persons were 14.2 (SD ANOVA) and 12.5% (CV ANOVA), respectively. The CVG was 13.4%, and the resulting index of individuality was 1.06. The reference change value was less than 41%. The availability of this automated assay for iFGF23 with well-characterized biological variation data delivers opportunities for improved availability and application of this assay clinically.

Tacrolimus has immunosuppressive effects on heavy/light chain pairs and free light chains in patients after heart transplantation: A relationship with infection.

The aim of the study was to investigate the relationship between tacrolimus (TAC) immunosuppressive treatment and serum concentrations of immunoglobulin heavy/light chain pairs (sHLC) and free light chains (sFLC) in patients after heart transplantation (HTX) and to use these biomarkers to predict the risk of infection in these patients. A total of 88 patients with an immunosuppressive regimen involving tacrolimus who underwent HTX were analyzed over 24 months of follow-up. sFLC and sHLC levels were determined before and at three time points after HTX. TAC concentrations were determined at several time points after HTX, and mean TAC concentrations and areas under the curve (AUCs) of TAC concentration were calculated. Relevant clinical data were obtained from patients' medical records. A larger AUC of TAC was associated with decreases in the concentrations of IgG total (p < 0.05); similarly, cumulative AUC of TAC during 18 post-transplant months correlated inversely with sHLC IgG kappa (r = -0.228, p < 0.05) and IgG total (r = -0.352, p < 0.05). Concentrations of sFLC kappa, sFLC lambda, sHLC IgG kappa, and sHLC IgG total were significantly lower in infected patients (in the 9th month after HTX, all p < 0.05). Combined criteria for increased AUC (greater than the median of 12.9 mg·d/l) and decreased sFLC kappa (less than the median of 12.5 mg/l) correlated with the presence of infection (p < 0.03) in the 9th month after HTX. Ratio of concentration of TAC to sFLC kappa or lambda was significantly higher in infected patients (both p < 0.05). Intensive treatment with tacrolimus after HTX is possibly reflected by decreases in sFLC and sHLC (mainly sHLC IgG). Patients with decreased concentrations of these biomarkers are at increased risk for infection, primarily in the 9th month after HTX, when the concentrations of tacrolimus were the highest.

Enhanced liver fibrosis (ELF) score: Reference ranges, biological variation in healthy subjects, and analytical considerations.

BACKGROUND: The Enhanced Liver Fibrosis score has been recognized as a non-invasive test for liver fibrosis. However, reference intervals, biological variation and analytical performance have not been studied in detail so far. The aim was to acquire data that are essential for correct interpretation. METHODS: A total of 40 apparently healthy volunteers were evaluated for reference ranges of serum concentration of hyaluronic acid, aminoterminal propeptide of type III collagen, and tissue inhibitor of metalloproteases-1, and calculated ELF score. A subgroup of 20 subjects was evaluated repeatedly for 7 weeks. For all variables, reference intervals, within-subject and between-subject biological variations, reference change values, and the indexes of individuality were assessed. Analytical performance (intermediate precision) and interlaboratory comparison were also evaluated. RESULTS: The reference ranges were 5.1-62.7 µg/L for HA, 3.56-12.6 µg/L for PIIINP, 143.6-265.3 µg/L for TIMP-1, and 7.14-9.55 for the ELF score. The within-subject variations were 32.7, 10.6, 4.2, and 3.2% for HA, PIIINP, TIMP-1, and ELF score, respectively. Similarly, the between-subject variations were 59.0, 13.3, 12.8, and 5.2%. For the ELF score, RCV was 10.1% and II was 0.62. The intermediate precisions were <5%, <6%, and <10% for HA, PIIINP, and TIMP-1, respectively. CONCLUSION: The reference range of the ELF score overlap with the area defined as moderate fibrosis by the manufacturer. High biological variation of HA was diminished by the natural logarithm in the calculation of the ELF score. The use of the ELF score has suitable analytical and acceptable biological performance characteristics for clinical practice. However, the transfer of results evaluated in healthy persons to the populations with chronic liver diseases deserves caution.