Krystyna Pawlak

Oxidative Stress – a Link between Endothelial Injury, Coagulation Activation, and Atherosclerosis in Haemodialysis Patients

Background/Aim: Recently emerging evidence suggests that oxidative stress (SOX) may participate in atherogenesis. The aim of the present study was to establish whether enhanced SOX, involving endothelial injury, activation of coagulation, and inflammatory reaction, could be implicated in atherosclerotic diseases in haemodialysis (HD) patients. Methods: Markers of SOX, endothelial injury, coagulation, and cytokines, were measured in the plasma of HD patients with and without cardiovascular disease (CVD), and of healthy controls by ELISA methods. Remodeling of the carotid arteries was assessed by measuring the intima-media thickness (IMT) as a surrogate of atherosclerotic disease in all groups. Results: Markers of SOX, endothelial injury, and extrinsic coagulation pathway activation and IMT values were significantly elevated in HD patients, especially in those with CVD when compared with the control group. The von Willebrand factor antigen (vWF:Ag) levels were more increased in the patients with CVD than in those without. Furthermore, the plasma levels of tumour necrosis factor alpha, monocyte chemo-attractant protein 1, and macrophage inflammatory protein 1 beta were significantly higher only in the HD group with CVD when compared with the controls. The IMT was strongly and directly correlated with Cu/Zn superoxide dismutase. Both IMT and Cu/Zn superoxide dismutase were positively correlated with age, thrombomodulin, vWF:Ag, tissue factor, tissue factor pathway inhibitor, prothrombin fragment F1 + 2, monocte chemo-attractant protein 1, macrophage inflammatory protein 1 beta, and tumour necrosis factor alpha levels. Multivariate analysis identified vWF:Ag as the only independent variable significantly associated with an increased IMT. Conclusions: The present study suggests that enhanced SOX, involved pro-atherogenic cytokine and chemokines levels, endothelial injury, and coagulation activation may constitute a pathway for accelerated atherosclerosis in HD patients. The significant, independent association between IMT and vWF:Ag should be assessed in future studies to determine whether vWF:Ag elevation is causative or a by-product of the increased IMT.

The coagulo-lytic system and endothelial function in cyclosporine-treated kidney allograft recipients

Since thromboembolic complications in transplanted patients are generally attributed to combined abnormalities in platelets and coagulo-lytic system, some hemostatic parameters tPA (tissue plasmogin activator):Ag and activity, its inhibitor-PAIAg and activity, tPA/PAI, thrombin-antithrombin (TAT) and plasmin-antiplasmin complexes (PAP), urokinase-uPA, euglobulin clot lysis time-ECLT, fibrinogen, plasminogen, protein C activity, D-dimer, prothrombin fragments1+2 (F1+2), fibrin monomers, fibronectin, lipoprotein-a, and von Willebrand factor(vWF), were evaluated using commercially available kits. The studies were performed on kidney transplant recipients treated with CsA, azathioprine and prednisone (n=21), and healthy volunteers (n=21). ECLT was significantly prolonged in kidney transplant recipients together with a rise in F1+2,lipoprotein-a, fibrinogen, fibronectin, and vWF when compared with controls. The TPA level was lower, whereas the PAI level was higher in kidney transplant recipients when compared with controls. In conclusion, CsA-treated kidney transplant recipients show evidence of pronounced impairment in fibrinolysis and endothelial damage in comparison with healthy volunteers.

Is Hepcidin a Link between Anemia, Inflammation and Liver Function in Hemodialyzed Patients?

Background: Hepcidin synthesis in hepatocytes is modulated in response to anemia, hypoxia or inflammation. A cross-sectional study was performed to assess hepcidin correlations with markers of iron status, erythropoietin therapy and markers of inflammation in hemodialyzed patients and in the healthy volunteers. Methods: Iron status, complete blood count, creatinine, albumin, lipids were assessed using standard laboratory methods. Hepcidin and high-sensitivity CRP were measured using commercially available kits. Results: Serum iron, TIBC, TSAT, erythrocyte count, Hb, Ht, platelet count, albumin, and cholesterol were lower, whereas ferritin and hepcidin were higher in hemodialyzed patients over controls. Hepcidin correlated positively with triglycerides, albumin, aspartate aminotransferase, lymphocyte count, ferritin and erythropoietin dose and negatively with erythrocyte count, Hb, and Ht in hemodialyzed patients. In multiple regression analysis, triglycerides (β value was 0.28, p = 0.02) and albumin (β value was –0.31, p = 0.006) were correlates of hepcidin in hemodialyzed patients. Conclusions: Elevated hepcidin levels in hemodialyzed patients may be due to functional iron deficiency and anemia. Liver plays an important role in the synthesis of hepcidin. Low-grade inflammation, frequently found in hemodialyzed patients, might also contribute to elevated hepcidin concentration. Hypothesis that hepcidin might link anemia, inflammation and liver function in kidney disease should be further evaluated.

HEMOSTASIS, PLATELET FUNCTION AND SEROTONIN IN ACUTE AND CHRONIC RENAL FAILURE

A pathogenetic role for fibrin deposition and platelet activation in the kidney is thought to play a role in the pathogenesis of acute renal failure (ARF). Thus, some fibrinolytic parameters and platelet function have been studied in 17 patients with ARF and compared to healthy volunteers and subjects with chronic renal failure (CRF). Since serotonin may participate in pathological processes resulting from platelet/vessel wall interactions, its level in the whole blood and plasma was also assayed. In ARF and CRF platelet aggregatory responses in both whole blood and in platelet rich plasma upon stimulation with various agonists (collagen, arachidonic acid, ADP, ristocetin) were lower than those obtained in healthy volunteers. Increased levels of lipoprotein (a), von Willebrand factor (vWF) and fibronectin were found in ARF relative to controls. Protein C activity was significantly lower in patients with ARF. Euglobulin clot lysis time was prolonged in ARF and CRF, reflecting a decreased overall fibrinolytic activity. Activity of tissue plasminogen activator (tPA) inhibitor (PAI) and PAI:Ag were higher in ARF, whereas tPA:Ag, urokinase, tPA/PAI complexes, thrombin-antithrombin complexes (TAT), plasmin-antiplasmin (PAP) complexes, fibrinogen, and F1+2 did not differ between ARF and controls. In CRF elevated levels of TAT, PAP, fibrinogen and prothrombin fragments F1+2 were found, whereas concentration of fibronectin was lowered when compared to controls. In both groups of renal failure patients increased levels of fibrin monomers and d-dimer were found relative to healthy volunteers. Whole blood serotonin was significantly lower, whereas plasma serotonin was significantly higher in patients with ARF and CRF relative to controls. Serotonin uptake and its release from platelets were markedly diminished in patients with ARF and CRF. Chronic renal failure exhibit a slightly different pattern of coagulopathies that acute renal failure.

The kynurenines are associated with oxidative stress, inflammation and the prevalence of cardiovascular disease in patients with end-stage renal disease

Increased oxidative stress (SOX), inflammation and prevalence of cardiovascular disease (CVD) have been reported in end-stage renal disease (ESRD), but their associations with kynurenine (KYN) pathway activation remain unknown. We determined the plasma concentrations of tryptophan (TRP), KYN, 3-hydroxykynurenine (3-HKYN); two distinct SOX markers: Cu/Zn superoxide dismutase (Cu/Zn SOD) and total peroxide; and high sensitivity C-reactive protein (hs CRP) as a indicator of inflammation in 146 ESRD patients and healthy controls. Analysis of TRP degradation through the KYN pathway demonstrated that in uremia the concentrations of this aminoacid were decreased by 40-60% in comparison with controls. In contrast, the plasma levels of KYN and 3-HKYN in ESRD patients were increased by 32-96% and 184-306%, respectively. These changes were accompanied by significant increase in the kyn/trp ratios by 140-240%, and 3-hkyn/kyn ratios by 40-154% in uremics compared to controls. ESRD patients showed a significant increase in Cu/Zn SOD, total peroxide and hs CRP levels between controls and all patients group. KYN and 3-HKYN were positively associated with inflammation and SOX markers in uremics. Logistic regression analysis showed that age, gender, presence of DM (all p<0.001), elevated hs CRP (p<0.01) and 3-HKYN levels (p<0.05) were independently associated with the presence of CVD in this population. These results suggest a relationship between KYN pathway activation and increased SOX, inflammation and CVD prevalence in ESRD patients.

Accumulation of toxic products degradation of kynurenine in hemodialyzed patients

In patients that developed a chronic renal failure the augmentation intryptophan degradation is reflected in the increase in plasma metabolitesof kynurenine pathway. Hemodialaysis is one of therapeutic approachesthat significantly reduce all plasma kynurenine metabolites in uremicpatients. In spite of haemodialaysis, plasma concentration of kynurenine,kynurenic acid, 3-hydroxykynurenine, anthranilic acid, xanthurenicacid and quinolinic acid were still elevated in uremic patients in comparisonwith healthy volunteers. These data shows significant disturbances inkynurenine metabolism in uremic patients. Accumulation of thesesubstances in uremic blood is capable to account for certain uremicsymptoms.

Kynurenine, quinolinic acid--the new factors linked to carotid atherosclerosis in patients with end-stage renal disease.

Increased oxidative stress (SOX), inflammation and accelerated atherosclerosis have been reported in end-stage renal disease (ESRD), but their associations with kynurenine pathway activation remain unknown. We determined the plasma concentrations of kynurenine (KYN), kynurenic acid (KYNA) and quinolinic acid (QA); three distinct SOX markers: Cu/Zn superoxide dismutase (Cu/Zn SOD), total peroxide and malondialdehyde (MDA), high sensitivity C-reactive protein (hs CRP) as a indicator of inflammation, and intima-media thickness (IMT)--an early reflection of the systemic atherosclerosis in the population of 124 patients with ESRD. In uraemia, the concentrations of KYN, KYNA and QA were increased by 37-105%, by 84-428%, and by 394-1018% of the control values; respectively. These changes were accompanied by significant increase in kyna/kyn and qa/kyn ratios, reflecting increased activity of kynurenine pathway enzymes. KYN, QA and qa/kyn ratio were positively associated with inflammation, SOX markers, and IMT values in uraemics. Moreover, multiple stepwise regression analysis identified age, presence of diabetes mellitus, QA and qa/kyn ratio as the independent variables significantly associated with increased IMT in this population. In conclusion, the results of the present study suggest a relationship between kynurenine pathway activation and increased oxidative stress, inflammation and the progression of atherosclerosis in end-stage renal disease patients.

Kynurenine pathway – a new link between endothelial dysfunction and carotid atherosclerosis in chronic kidney disease patients

PURPOSE The endothelium dysfunction is an important component of atherosclertic cardiovascular disease. It has been also suggested that kynurenine pathway activation may be involved in the pathogenesis of this disease. MATERIAL/METHODS This is a cross-sectional study in chronic kidney disease (CKD) patients (n=106; 60 Males). The plasma markers of endothelial dysfunction and kynurenine (KYN), 3-hydroxykynurenine (3-HKYN), kynurenic acid (KYNA), anthranilic acid (AA) and quinolinic acid (QA) were measured in relation to an early indicator of the systemic atherosclerosis - intima-media thickness (IMT). RESULTS Kynurenines, von Willebrand factor (vWF), thrombomodulin (TM), soluble adhesion molecules (sICAM-1, sVCAM-1) and IMT in each uraemic group were significantly higher than in healthy people. In contrast, no significant differences in sE-selectin and sP-selectin concentrations were observed between CKD patients and controls. Kynurenines were positively associated with vWF, TM, sICAM-1 and sVCAM-1, whereas sP-selectin was inversely associated with the most of kynurenines. IMT was positively correlated both with kynurenines: KYN, 3-HKYN, QA as well as with endothelial markers: TM, vWF, sICAM-1 and sVCAM-1 (all p<0.01). Finally, multiple regression analysis identified age, vWF, sVCAM-1 and QA levels as the independent variables significantly associated with increased IMT in this population (adjusted r² = 0.51). CONCLUSIONS This study suggests a relationship between kynurenine pathway activation, endothelial dysfunction and the progression of atherosclerosis in CKD patients. It opens a new idea that the inhibition of kynurenine pathway may provide an effective strategy to slow down endothelial dysfunction and thereby the prevalence of atherosclerosis in this population.

Visfatin and apelin, new adipocytokines, and their relation to endothelial function in patients with chronic renal failure

PURPOSE Visfatin and apelin are novel adipocytokines that have recently generated much interest. The aim of the study was to assess visfatin and apelin in correlation with markers of endothelial cell injury and inflammation in 22 patients with chronic kidney disease-CKD and 22 age- and sex-matched healthy volunteers. METHODS We assessed visfatin, apelin, markers of coagulation: TAT (thrombin-antithrombin complexes), prothrombin fragments 1+2; fibrinolysis: tPA (tissue plasminogen activator), PAI-1 (plasminogen activator inhibitor), PAP (plasmin-antiplasmin complexes); endothelial function/injury: vWF (von Willebrand factor), thrombomodulin, ICAM (intracellular adhesion molecule), VCAM (vascular cell adhesion molecule), CD146, CD40L, CD44, E-selectin, inflammation: hsCRP. RESULTS Triglycerides, hsCRP, creatinine, vWF, prothrombin fragments 1+2, TAT, thrombomodulin, ICAM, VCAM, CD146, CD44, CD40L, PAI-1, PAP, visfatin and E-selectin were elevated in chronic kidney disease patients when compared with the control group. Visfatin correlated significantly in patients with chronic kidney disease, in univariate analysis, with CD40L (r=-0.27, p<0.05), apelin (r=0.27, p<0.05), ICAM (r=0.26, p<0.05), VCAM (r=0.31, p<0.05) and tended to correlate with CD146 (r=0.21, p=0.10). Apelin correlated significantly with E-selectin (r=0.31, p<0.05) and VCAM (r=0.31, p<0.05). In the healthy volunteers visfatin correlated significantly with ICAM (r=-0.37, p<0.05) and serum creatinine (0.38, p<0.05). CONCLUSIONS Elevated visfatin in CKD patients may be due to renal failure and/or inflammation. Adipocytokines related to adhesion molecules might support the importance of inflammation/endothelial cell injury in the pathogenesis of atherosclerosis and its consequences in CKD.

Oxidative stress influences CC-chemokine levels in hemodialyzed patients.

Background: Increased oxidative stress (SOX) has been reported in hemodialyzed (HD) patients, but its influence on CC-chemokine levels remains unknown. Methods: The levels of 3 distinct SOX markers (Cu/Zn superoxide dismutase (Cu/Zn SOD), total lipid peroxide, and autoantibodies against oxidized LDL (OxLDL-Ab)), as well as those of 4 CC-chemokines (monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory proteins (MIP-1α, MIP-1β), regulated upon activation, normal T cell expressed and secreted (RANTES), and tumor necrosis factor-α (TNF-α)) were measured pre- and post-HD session in 15 HD patients with cardiovascular disease (CVD) and 12 HD patients without CVD (pre- and post-HD session) and 17 controls. Results: Cu/Zn SOD levels were elevated in HD patients (both before and after HD) compared to controls (p < 0.001). Total lipid peroxide levels were similar in the controls in pre-dialysis samples, but were increased after a HD session (p < 0.001). Pre-dialysis OxLDL-Ab levels were increased only in the HD group with CVD compared to controls (p < 0.05). The pre-dialysis plasma levels of TNF-α (p < 0.001), MCP-1 (p < 0.001) and MIP-1β (p < 0.01) were increased both in all HD patients and subgroups with CVD when compared to the controls, and remained significantly elevated when measured after HD. Dialysis increased MCP-1 (p < 0.05) and MIP-1β (p < 0.001) levels as compared to the controls as well as in patients without CVD. Plasma RANTES was significantly lower before HD (p < 0.05) and after HD (p < 0.001) in patients with CVD. Patients without CVD and all HD patients also had lower RANTES before and after HD when compared to controls (all p < 0.001). A positive correlation was observed between plasma pre-dialysis Cu/Zn SOD levels and the other SOX markers (all p < 0.05), age (p < 0.05) and duration of hemodialysis (p < 0.001). A positive relationship existed between plasma Cu/Zn SOD levels and those of MCP-1, MIP-1β and TNF-α (all p < 0.001). RANTES levels negatively correlated with Cu/Zn SOD (p < 0.0001).TNF-α positively correlated with age (p < 0.05), total lipid peroxide (p = 0.010), MCP-1 (p < 0.01) and MIP-1β levels (p < 0.01). An identifiable association exists between MCP-1 and MIP-1β levels (p < 0.01), whereas both MCP-1 and MIP-1β were inversely correlated with RANTES (both p < 0.05). Conclusion: Our data indicate for the first time a probable functional relationship between oxidative stress and CC-chemokine levels in hemodialyzed patients, particularly in those with cardiovascular disease. This relationship may represent one of the mechanisms involved in the progression of atherosclerosis in these patients.