AGEs/sRAGE, a novel risk factor in the pathogenesis of end-stage renal disease.

Interaction of advanced glycation end products (AGEs) with its cell-bound receptor (RAGE) results in cell dysfunction through activation of nuclear factor kappa-B, increase in expression and release of inflammatory cytokines, and generation of oxygen radicals. Circulating soluble receptors, soluble receptor (sRAGE), endogenous secretory receptor (esRAGE) and cleaved receptor (cRGAE) act as decoy for RAGE ligands and thus have cytoprotective effects. Low levels of sRAGE and esRAGE have been proposed as biomarkers for many diseases. However sRAGE and esRAGE levels are elevated in diabetes and chronic renal diseases and still tissue injury occurs. It is possible that increases in levels of AGEs are greater than increases in the levels of soluble receptors in these two diseases. Some new parameters have to be used which could be an universal biomarkers for cell dysfunction. It is hypothesized that increases in serum levels of AGEs are greater than the increases in the soluble receptors, and that the levels of AGEs is correlated with soluble receptors and that the ratios of AGEs/sRAGE, AGEs/esRAGE and AGEs/cRAGE are elevated in patients with end-stage renal disease (ESRD) and would serve as an universal risk marker for ESRD. The study subject comprised of 88 patients with ESRD and 20 healthy controls. AGEs, sRAGE and esRAGE were measured using commercially available enzyme linked immune assay kits. cRAGE was calculated by subtracting esRAGE from sRAGE. The data show that the serum levels of AGEs, sRAGE, cRAGE are elevated and that the elevation of AGEs was greater than those of soluble receptors. The ratios of AGEs/sRAGE, AGEs/esRAGE and AGEs/cRAGE were elevated and the elevation was similar in AGEs/sRAGE and AGEs/cRAGE but greater than AGEs/esRAGE. The sensitivity, specificity, accuracy, and positive and negative predictive value of AGEs/sRAGE and AGEs/cRAGE were 86.36 and 84.88 %, 86.36 and 80.95 %, 0.98 and 0.905, 96.2 and 94.8 %, and 61.29 and 56.67 % respectively. There was a positive correlation of sRAGE with esRAGE and cRAGE, and AGEs with esRAGE; and negative correlation between sRAGE and AGEs/sRAGE, esRAGE and AGES/esRAGE, and cRAGE and AGES/cRAGE. In conclusion, AGEs/sRAGE, AGEs/cRAGE and AGEs/esRAGE may serve as universal risk biomarkers for ESRD and that AGEs/sRAGE and AGEs/cRAGE are better risk biomarkers than AGEs/esRAGE.

Association between circulating thrombopoietin levels and cardiovascular risk prediction scores in renal transplant recipients.

BACKGROUND/AIMS: The 7-year Major Adverse Cardiovascular Events Calculator (CRCRTR-MACE) predicts cardiovascular events (CVE) in renal transplant recipients (RTR), and thrombopoietin (TPO) is a humoral inflammatory factor implicated in cardiovascular disease (CVD). The aim of the study was to determine if circulating TPO levels in stable RTR are positively associated with variable(s) in the CRCRTR-MACE score. METHODS: CRCRTR-MACE scores were calculated in 95 stable RTR. TPO levels were measured by multiplexed fluorescent bead-based immunoassay in all patients and 48 controls. Multivariate analysis (MVA) was performed between TPO and CV risk variables and patient demographics. Stepwise regression with backward elimination of insignificant variables estimated the impact of risk variables on TPO levels. Significance was defined at p < 0.05. Normalized data were presented as mean ± SD and non-normalized data as median (maximum to minimum). RESULTS: The risk of a CVE within 7 years as predicted by the median was 9.97% (range 1.93-84.2). The percentage of patients who were above 20% risk for a CVE was 28.4%. Control TPO level of 170.41 (4.4-995.9) pg/ml was significantly lower than that of 237.90 (32.77-1,386.79) pg/ml in RTR (p = 0.010). TPO level correlated significantly with the total CRCRTR-MACE score (R = 0.310, p = 0.004), smoking (p = 0.009) and eGFR (R = -0.275, p = 0.012) but not with age, diabetes, LDL level or history of CVE. Only the total CRCRTR-MACE score (p = 0.013) and smoking (p = 0.009) remained significant in the MVA. Stepwise regression estimated that smoking increased TPO levels by 206.28 pg/ml and each 10% increase in CRCRTR-MACE score increased TPO levels by an additional 44.4 pg/ml. CONCLUSION: TPO levels are increased in RTR with high CRCRTR-MACE, particularly in smokers with diminished eGFR. Circulating TPO may serve as a biomarker and treatment target for CVD in RTR.

Contribution of impaired renal function to cardiovascular risk prediction models in renal transplant recipients.

BACKGROUND: The Framingham risk score (FRS) and cardiovascular risk calculator for renal transplant recipients (CRCRTR-MACE) quantify cardiovascular risk in renal transplant recipients (RTR). In contrast to the FRS, the CRCRTR-MACE includes serum creatinine as a variable in the risk prediction equation. OBJECTIVE: To determine the influence of impaired renal function on performances of the two equations. METHODS: A chart review of 270 RTR transplanted from 1979 to 2012. High risk was defined at scores ≥20%. Standard statistical analyses included multivariate analysis (MVA), stepwise analysis, and odds ratio to estimate contributions of risk factors. RESULTS: Mean transplant duration was 9.51 ± 6.65 yr. Mean eGFR was 59.19 ± 28.26 mL/min/1.73 m(2) . FRS and CRCRTR-MACE scores of least 20% were present in 9.3% and 24.8%, respectively, while 7.2% and 11.2% of RTR with eGFR ≥60 mL/min/1.73 m(2) were high risk, respectively. Mean age, blood pressure, TC:HDL ratio, smoking, and diabetes were evenly distributed in patients with varying eGFR. FRS scores remained similar at wide eGFR range (≤30 mL/min/1.73 m(2) -≥90 mL/min/1.73 m(2) ), while CRCRTR-MACE scores significantly increased as eGFR decreased. CONCLUSIONS: CRCRTR-MACE identified more patients at high cardiovascular risk, even in those with more favorable renal function, suggesting a fundamental difference between the two calculators beyond renal function.

The Sask formula to estimate glomerular filtration rate in renal transplant patients.

The aim of this study was to develop a glomerular filtration rate (GFR) equation for renal transplant and compare its performance with Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) and isotope dilution mass spectrometry (IDMS) equations. Using genetic symbolic regression analysis, the Sask equation was developed from a training sample of 772 isotope GFR (iGFR) scans performed in 99 transplanted patients. It was then validated in two other samples of 269 scans with the same number of patients. Standard methods including accuracy at 30% range from reference values were compared. In two validation samples, the Sask equation maintained the lowest bias of -0.7 ± 19.0 and 0.4 ± 18.4 ml/min/1.73 m(2) (p < 0.05) versus -3.1 ± 19.6 and -7.2 ± 18.8 ml/min/1.73 m(2) for CKD-EPI and -2.2 ± 19.2 and -6.5 ± 18.3 ml/min/1.73 m(2) for IDMS, respectively. In those with iGFR between 90 and 30 ml/min/1.73 m(2), the Sask equation demonstrated: (1) the lowest bias of -1.0 ± 15.7 and -0.4 ± 15.7 ml/min/1.73 m(2) (p < 0.05 vs. other tests); (2) an accuracy of 75.5 and 76.1% (p < 0.05 vs. other tests), and (3) a mean percentage error of 1.9 ± 30.5 and -4.1 ± 31.4 ml/min/1.73 m(2) (p < 0.05 vs. other tests). Analysis based on gender demonstrated improved performance in the total and subtotal female populations with GFR between 90 and 30 ml/min/1.73 m(2). The CKD-EPI and Sask equations performed better than IDMS. The Sask equation demonstrated improved bias over CKD-EPI, with iGFR between 90 and 30 ml/min/1.73 m(2), particularly in females.

Interaction of Serum Phosphate with Age as Predictors of Cardiovascular Risk Scores in Stable Renal Transplant Recipients.

We calculated rate of changes in the cardiovascular risk calculator for renal transplant recipients (CRCRTR) major adverse cardiac events (MACE) in clinically stable renal transplant recipients (RTRs) to identify covariables that associate with fast cardiovascular (CV) risk progression. CRCRTR-MACE scores were calculated on 139 patients in 2011 and 2014. Score changes above and below median changes in scores were labeled fast or slow CV risk progression. Multivariate analysis (MVA) was performed to identify variables significant to percentage changes in scores. Receiver-operating characteristic (ROC) analysis was performed to define sensitivity and specificity of factors significant to fast score progression. Follow-up was 2.61 (2.02-4.47) years. Slow and fast progressions were present in 50.4 and 49.6% of patients, with a median change of 25.8% (- 92.1 to 1,444.7%). MVA showed percentage changes in age and serum phosphate were the only significant variables impacting fast progression in scores. ROC showed 2011 serum phosphate of 1.15 mmol/L to predict fast progression (area under the curve [AUC] of 0.628, p > 0.0126). Age older than 45 years combined with 2011 serum phosphate above 1.15 mmol/L had a significant AUC of 0.781, p < 0.0010 interleukin (IL)-1A and IL-28A were significant associates with serum phosphate above 1.1 mmol/L in the MVA. Changes in CV risk in RTR over time are highly variable. Serum phosphate, even within upper normal levels, predicts worsening of CV risk scores in stable RTR.

Circulating chemokine ligand levels before and after successful kidney transplantation.

BACKGROUND: Chemokine ligands (CCLs) play a pivotal role in tissue injury before and after kidney transplantation. Meanwhile, transplantation improves patient's survival and diminishes morbidity. It is hypothesized, then, that kidney transplantation diminishes pre-transplant (pre-TX) levels of circulating inflammatory CCLs. This retrospective study compared circulating levels and profiles of CCLs before transplantation (pre-TX) and after transplantation (post-TX). METHODS: Nineteen CCLs (1, 2, 3, 4, 5, 8, 11, 13, 15, 17, 21, 24, 26, 27, CXCL 5, 8, 10, 12 and 13) were measured in 47 stable post-TX recipients, and their stored pre-TX plasma was analyzed by multiplexed fluorescent bead-based immunoassay. Twenty normal controls were included for comparisons. Normalized data was presented as mean ± SD and non-normalized data as median (5-95 % CI). Significance was measured at p < 0.01. Arbitrary upper and lower margins for each CCL at the 95 % CI or 2SD levels in each group were chosen to calculate the percentile of patients in the other group who exceeded these limits. Significant CCL levels present in more than 75 % of patients in a group that exceeded the arbitrary upper or lower set margins in the other two groups were labeled as preferentially characteristic for the respective group. RESULTS: More than 75 % of pre- and post-TX patients had levels that exceeded the upper control for CCL1, 11, 15 and CCL15, CCL26 and CXCL13 levels, respectively. More than 75 % of pre- and post-TX patients exceeded the lower control for CCL3, 21, and CCL5 limits, respectively. More than 75 % of post-TX patients demonstrated elevated levels of CCL2, 3, 21, 26 and CXCL13 above the upper pre-TX cut offs. Meanwhile, more than 75 % of post-TX patients exceeded the lower pre-TX levels for CCL1, 4, 5, 8, 13, 15, 17, 24 and CXCL8 and10. Pre-TX was preferentially characterized by elevated CCL1 and 15 and diminished CCL3 and 21. Post-TX was preferentially characterized by elevated CCL26 and CXCL13 and diminished CCL4 and 5. CONCLUSION: End stage kidney disease is associated with enhanced circulating inflammatory chemokine levels. Stable kidney transplantation is associated with 1) lowered burden of circulating inflammatory chemokine levels and, 2) elevation in the pro T-helper2 chemokine, CCL26 and the homeostatic CXCL13.

Elevated Circulating Interleukin 33 Levels in Stable Renal Transplant Recipients at High Risk for Cardiovascular Events.

BACKGROUND: The Major Adverse Cardiovascular Events calculator (CRCRTR-MACE) estimates the burden of cardiovascular risk in renal transplant recipients (RTR). Our recent study of 95 RTR reported the 7-year median risk of cardiovascular events (CVE) to be 9.97%, ranging from 1.93 to 84.27%. Nearly a third (28.4%) of the cohort was above 20% risk for a CVE. Since interleukins (ILs) as part of the inflammatory response may play a role in the pathogenesis of cardiovascular disease (CVD), we extended this study to identify which ILs are associated with high cardiovascular risk in this population. METHODS: Twenty-two ILs were measured by multiplexed fluorescent bead-based immunoassay in 95 RTR and 56 normal controls. Stepwise analysis after multivariate determination of significant demographic and inflammatory variables was performed between the high and low-CVD risk groups (which were arbitrarily set at scores <10% and ≥20%, respectively). Normalized data was presented as mean ± SD and non-normalized data as median (minimum-maximum). Significance was measured at <0.05. RESULTS: 27.5% of the low-risk and 31.3% of the high-risk groups had mean IL levels above the 95 percentile of the normal control levels. In the non-parametric analysis IL-6, 9, 16, 17 and 33 were significantly higher in the high-risk group compared to the control. Univariate analysis (UVA) of the high-risk group identified IL-33 as the only IL that remained significantly higher than the control and low-risk groups (p = 0.000). The percentage of patients with IL-33 levels above the 90 percentile of control value in the low and high-risk groups were 15.6% and 52.0%, respectively (p<0.002). UVA of factors significant to high IL-33 levels included estimated glomerular filtration rate (eGFR), while diabetes mellitus, serum phosphorus, microalbuminuria and age also remained significant in the multivariate analysis. CONCLUSION: Circulating IL-33 level is positively associated with high CRCRTR-MACE score. Diminished eGFR, age, diabetes, serum phosphorus and microalbuminurea demonstrate significant relationship with elevated IL-33 levels, supporting the possible pathognomonic role of IL-33 in the cardiovascular burden in RTR.

Study of uremic toxin fluxes across nanofabricated hemodialysis membranes using irreversible thermodynamics.

INTRODUCTION: The flux of uremic toxin middle molecules through currently used hemodialysis membranes is suboptimal, mainly because of the membranes' pore architecture. AIM: Identifying the modifiable sieving parameters that can be improved by nanotechnology to enhance fluxes of uremic toxins across the walls of dialyzers' capillaries. METHODS: We determined the maximal dimensions of endothelin, cystatin C, and interleukin - 6 using the macromolecular modeling software, COOT. We also applied the expanded Nernst-Plank equation to calculate the changes in the overall flux as a function of increased electro-migration and pH of the respective molecules. RESULTS: In a high flux hemodialyzer, the effective diffusivities of endothelin, cystatin C, and interleukin - 6 are 15.00 x 10(-10) cm(2)/s, 7.7 x 10(-10) cm(2)/s, and 5.4 x 10(-10) cm(2)/s, respectively, through the capillaries' walls. In a nanofabricated membrane, the effective diffusivities of endothelin, cystatin C, and interleukin - 6 are 13.87 x 10(-7) cm(2)/s, 5.73 x 10(-7) cm(2)/s, and 3.45 x 10(-7) cm(2)/s, respectively, through a nanofabricated membrane. Theoretical modeling showed that a 96% reduction in the membrane's thickness and the application of an electric potential of 10 mV across the membrane could enhance the flux of endothelin, cystatin C, and interleukin - 6 by a factor of 25. A ΔpH of 0.07 altered the fluxes minimally. CONCLUSIONS: Nanofabricated hemodialysis membranes with a reduced thickness and an applied electric potential can enhance the effective diffusivity and electro-migration flux of the respective uremic toxins by 3 orders of magnitude as compared to those passing through the high flux hemodialyzer.

Theoretical Application of Irreversible (Nonequilibrium) Thermodynamic Principles to Enhance Solute Fluxes across Nanofabricated Hemodialysis Membranes.

Objective. Nanotechnology has the potential to improve hemodialysis membrane technology. Thus, a major objective is to understand how to enhance toxic solute fluxes across these membranes. The aim of this concept building study is to review the application of irreversible thermodynamic (IT) to solute fluxes. Methods. We expanded the application of the Nernst-Planck equation to include the Kedem-Katchalsky equation, pH, membrane thickness, pore size, and electric potential as variables. Results. (1) Reducing the membrane's thickness from 25 µm to 25 nm increased the flux of creatinine, ß(2)-microglobulin, and tumor necrosis factor-α (TNF-α) by a thousand times but prevented completely albumin flux, (2) applying an electric potential of 50-400 mV across the membrane enhanced the flux of the respective molecules by 71.167 × 10(-3), 38.7905 × 10(-8), and 0.595 × 10(-13) mol/s, and (3) changing the pH from 7.35 to 7.42 altered the fluxes minimally. Conclusions. The results supported an argument to investigate the application of IT to study forces of fluxes across membranes. Reducing the membrane's thickness-together with the application of an electrical potential-qualities achievable by nanotechnology, can enhance the removal of uremic toxins by many folds. However, changing the pH at a specific membrane thickness does not affect the flux significantly.