The association of TMPRSS6 gene polymorphism with iron status in Egyptian children (a pilot study).

Several studies have shown association of single nucleotide polymorphisms (SNPs) of hepcidin regulatory pathways genes with impaired iron status. The most common is in the TMPRSS6 gene. In Africa, very few studies have been reported. We aimed to investigate the correlation between the common SNPs in the transmembrane protease, serine 6 (TMPRSS6) gene and iron indicators in a sample of Egyptian children for identifying the suitable candidate for iron supplementation.Patients and methods One hundred and sixty children aged 5-13 years were included & classified into iron deficient, iron deficient anemia and normal healthy controls. All were subjected to assessment of serum iron, serum ferritin, total iron binding capacity, complete blood count, reticulocyte count, serum soluble transferrin receptor and serum hepcidin. Molecular study of TMPRSS6 genotyping polymorphisms (rs4820268, rs855791 and rs11704654) were also evaluated.Results There was an association of iron deficiency with AG of rs855791 SNP, (P = 0.01). The minor allele frequency for included children were 0.43, 0.45 & 0.17 for rs4820268, rs855791 & rs11704654 respectively. Genotype GG of rs4820268 expressed the highest hepcidin gene expression fold, the lowest serum ferroportin & iron store compared to AA and AG genotypes (p = 0.05, p = 0.05, p = 0.03 respectively). GG of rs855791 had lower serum ferritin than AA (p = 0.04), lowest iron store & highest serum hepcidin compared to AA and AG genotypes (p = 0.04, p = 0.01 respectively). Children having CC of rs11704654 had lower level of hemoglobin, serum ferritin and serum hepcidin compared with CT genotype (p = 0.01, p = 0.01, p = 0.02) respectively.Conclusion Possible contribution of SNPs (rs855791, rs4820268 and rs11704654) to low iron status.

Oxidative stress and hepcidin expression in pediatric sickle cell anemia with iron overload.

BACKGROUND: Blood transfusion (BT) is essential in treating sickle cell disease (SCD); however, it leads to iron overload (IO) and oxidative stress. We studied the relationship between oxidative stress, iron status parameters, hepcidin mRNA gene expression, and IO in SCD patients. METHODS: We classified all SCD patients (n = 90) into two groups: Group I, 45 children (s.ferritin ≥ 938 ng/mL) and Group II, 45 children (s.ferritin < 938 ng/mL). A total of 55 children, age and sex matched, participated as a control group. Malondialdehyde (MDA), nitrite, s.iron, s.total iron-binding capacity (sTIBC), transferrin saturation %, s.ferritin, s.hepcidin, and hepcidin mRNA gene expression were assessed. RESULTS: Among SCD BT-dependent patients (>3 times/year), 63% were from Group I and 37% from Group II, p < .01. The two patient groups had significantly lower s.hepcidin and hepcidin gene expression than controls ( p < .001). TIBC, s.iron, s.ferritin, transferrin saturation %, ferritin/hepcidin ratio, and MDA levels were higher among SCD patients than controls ( p < .001). Group I had higher mean level of ferritin/hepcidin ratio and MDA than Group II ( p < .01). The higher level of MDA and increased frequency of BT were the significant predicting risk factors for IO ( p < .05). A receiver-operating characteristic curve indicates that MDA is the outstanding significant biomarker for high level of s.ferritin with subsequent IO progression. CONCLUSION: MDA may serve as a biomarker of oxidative stress and IO in SCD patients. This result paid attention for urgent initiation of antioxidant and chelation therapy on detecting increased MDA level.

Detection of exosomal miR-18a and miR-222 levels in Egyptian patients with hepatic cirrhosis and hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is known to be the second leading cause of cancer-related mortality worldwide. For improving the prognosis as well as reducing the rate of mortality, early diagnosis of HCC is a must. AIMS: This study was conducted to assess the ability of the serum expression of exosomal miR-18a and miR-222 to differentiate and diagnose patients with HCC, patients with liver cirrhosis, and healthy controls. METHODS: This study included 51 patients with liver cirrhosis, 51 patients with HCC on top of hepatitis C virus (HCV) infection, and 50 healthy controls. RESULTS: miR-18a and miR-222 were assessed using reverse transcription-polymerase chain reaction. MiR-18a and miR-222 levels were significantly higher in the liver cirrhosis and HCC groups than the control group (p ˂ 0.001). However, no statistically significant difference was found between patients with HCC and liver cirrhosis (p = 0.4 for miR-18a and p = 0.1 for miR-222). ROC curve analyses to evaluate the diagnostic performances of the two miRNAs as important noninvasive diagnostic markers revealed a best cutoff value of 2 for miR-18a to differentiate between liver cirrhosis, HCC, and healthy controls. And for mir-222, a cutoff value of 1.7 and 1.9 showed the highest specificity for discrimination between liver cirrhosis, HCC, and healthy controls, respectively. Moreover, logistic regression model revealed that miR-18a expression was independent predictive factor in HCC patients (p = 0.004), while miR-222 expression was independent predictive factor in liver cirrhosis patients (p < 0.001). CONCLUSION: miR-18a and miR-222 were significantly discriminative markers between patients with liver cirrhosis and HCC and healthy individuals. Therefore, they have a prognostic rather than a diagnostic value. Moreover, miR-18a and miR-222 could be useful in identifying liver injuries, including fibrosis and cirrhosis.

The Relation between Serum Hepcidin, Ferritin, Hepcidin: Ferritin Ratio, Hydroxyurea and Splenectomy in Children with ß-Thalassemia.

BACKGROUND: Hepcidin, a small peptide hormone, is established as the main regulator of iron homeostasis. AIM: To estimate serum hepcidin, ferritin, and hepcidin: ferritin ratio in ß-thalassemia patients and to determine the effect of splenectomy and hydroxyurea on serum hepcidin. METHODS: A study was conducted on 30 thalassemia major (ßTM), 29 thalassemia intermedia (ßTI) and 29 healthy children's controls. Data were collected by patient interviewing where detailed history-taking and thorough clinical examinations were carried out. Serum ferritin and hepcidin were measured by ELISA assay (Bioneovan Co. Ltd Beijing, China). RESULTS: Βeta-thalassemia patients had higher serum ferritin, serum hepcidin and lower Hb and hepcidin: ferritin ratio compared to the controls (p < 0.001, 0.010, 0.001, 0.001) respectively. Β-TM patients had higher mean serum hepcidin and serum ferritin compared to ß-TI, with statistically significant difference (P = 0.042, P < 0.001, respectively). Twenty-one patients out of 29 ßTI was on hydroxyurea therapy; these patients had significantly lower levels of serum ferritin (P < 0.004) and significantly higher levels of Hb (P < 0.004). Serum ferritin was statistically significantly higher in splenectomized patients P < 0.009. Serum hepcidin level was insignificantly higher in splenectomized patients than non-splenectomized patients (21.6 ± 14.75, 17.76 ± 10.01 ng/mL). Hepcidin showed a significantly positive correlation with hepcidin: ferritin ratio in all studied groups. CONCLUSION: Serum hepcidin was elevated in ß-thalassemia children with more evident elevation in ßTM patients. Splenectomy played no major role in hepcidin regulation. Knowing that hepcidin in serum has a dynamic and multi-factorial regulation, individual evaluation of serum hepcidin and follow up, e.g. every 6 months could be valuable, and future therapeutic hepcidin agonists could be helpful in management of iron burden in such patient.

Evaluation of miRNA-21 and miRNA Let-7 as Prognostic Markers in Patients With Breast Cancer.

BACKGROUND: Breast cancer remains one of the top threats to women's health. The current lack of tumor markers with desirable sensitivity and specificity is a major obstacle toward the future management of breast cancer. Many studies are directed to reveal the diagnostic and prognostic potentials of circulating miRNAs in breast cancer. In this study, we attempt to evaluate the feasibility and clinical utility of circulating miRNA-21 and let-7 as prognostic biomarkers for breast cancer. METHODS: Real-time quantitative polymerase chain reaction technique was used. Levels of miRNA-21 and let-7 expression were determined in sera from 125 participants representing 3 different groups. With fold-change analysis, the expression of miRNA-21 and let-7 in the decided groups were assessed. RESULTS: Patients with breast cancer showed significantly higher expression of miRNA-21 compared with controls and other participants with benign breast lesions (P < .001). The mean expression levels of serum miRNA-21 was 3.27 ± 2.10-fold in patients with breast cancer. The expression of miRNA let-7 was significantly decreased in patients with breast cancer (2.45 ± 2.20-fold) than the control group and the benign breast lesions group (5.27 ± 3.30-fold and 6.22 ± 4.90-fold, respectively; P < .001). Levels of miRNA let-7 expression negatively correlated with development of metastases in patients with breast cancer (P < .001). CONCLUSIONS: Our study establishes the association between altered levels of miRNA let-7 and metastases risk in patients with breast cancer, implying a role of miRNA let-7 in disease progression and prognosis.

Klotho G-395A gene polymorphism: impact on progression of end-stage renal disease and development of cardiovascular complications in children on dialysis.

BACKGROUND: Klotho G-395-A gene polymorphism may impact children with end-stage renal disease (ESRD). We investigated the relevance of Klotho G-395-A on ESRD development and progression, and its relationship with evolution of cardiovascular complications in pediatric dialysis patients. METHODS: Fifty-five children with chronic kidney disease (CKD) and seventy healthy children were genotyped for Klotho G-395A. RESULTS: Incidence of GA/AA genotypes and A allele were higher in ESRD patients compared with controls (54.5 vs. 7.1%, P < 0.001; 30.9 vs. 13.6%, P = 0.001, respectively). Also, children with GA/AA genotypes were 15.6 times more likely to develop ESRD than with GG genotype (95% CI 5.4-44.7, P < 0.001). A allele carriers have 2.8 times higher risk of developing ESRD than those with G allele (95% CI 1.5-5.35, P = 0.001). Also, the A allele could be considered a predictor of cardiovascular disease (CVD), as carriers have 161 times higher risk of cardiovascular complications than non-carriers (95% CI 21-1233, P < 0.001). All ESRD patients with CVD presented with left ventricular hypertrophy (LVH) and the frequency of A allele was significantly higher among ESRD children with LVH, whereas G allele frequency was significantly higher among ESRD children without LVH. CONCLUSIONS: The A allele of the G-395A Klotho gene polymorphism shows a significantly higher frequency among children with CKD and those with CVD and LVH. This mutant allele could be used as a risk marker for the development of ESRD as well as a predictor of CVD in these children.

Association of ficolin-2 (FCN2) functional polymorphisms and protein levels with rheumatic fever and rheumatic heart disease: relationship with cardiac function.

INTRODUCTION: A role for ficolin (FCN) 2 gene polymorphisms in the pathogenesis of recurrent severe streptococcal infections and rheumatic carditis has been suggested. The aim of the study was to evaluate a possible relationship between single nucleotide polymorphisms located at positions -602 and -4 of the FCN2 gene and FCN2 serum levels and risk of development of rheumatic fever (RF) and rheumatic heart disease (RHD). MATERIAL AND METHODS: Seventy-seven Caucasian Egyptian patients with RF were recruited with a control group of 43 healthy subjects. DNA was extracted for analysis of the FCN2 gene at positions -602 and -4 and serum protein level was measured by ELISA. RESULTS: FCN2 AA genotype at the -4 position was more frequently observed in RF and RHD patients, as compared to healthy subjects (p = 0.005 and p = 0.013, respectively); furthermore, the A allele was identified as a possible risk factor for the development of RF (p = 0.023, OR = 1.852, 95% CI: 1.085-3.159). The haplotype -602/-4 G/A, which was associated with low median levels of L-ficolin, was observed more frequently in the RF group when compared to the healthy subjects (74/162, 48.1% vs. 29/420, 33.7%, OR = 1.834, 95% CI: 1.034-3.252, p = 0.038). Low serum ficolin-2 level was associated with ESV and EDV increases. FCN 2 level was significantly lower with AA genotypes than GG+AG genotypes of the -4 position (56.68 ±17.90 vs. 66.05 ±18.79, p = 0.008). CONCLUSIONS: Polymorphisms linked to low levels of L-ficolin may render an individual at risk of recurrent and/or severe streptococcal infection. The -4 AA genotype and -602/-4 G/A haplotype are possible risk factors for the development of carditis.

FcγRIIa defunctioning polymorphism in paediatric patients with renal allograft.

INTRODUCTION: Fc gamma receptor (FcγR) IIa is considered the most widely distributed of the three classes of Fc receptors, and it expresses an allelic polymorphism. This type of polymorphism may modify the immune response and may be an important factor for some diseases. The aim of the study reported herein was to evaluate the association between the FcγRIIa polymorphism and susceptibility to both end-stage renal disease (ESRD) and acute kidney graft rejection (AR) in children who have undergone renal transplantation. MATERIAL AND METHODS: The study evaluated 70 children who had undergone transplantation and 60 healthy subjects. AR was observed in 25 children. RESULTS: FcγRIIa genotypes and alleles were significantly different between transplantation patients and the control group. The assessment for FcγR of the groups in which AR was present showed that there was only a risk of having an acute rejection in homozygous genotype RR. CONCLUSIONS: FcγRIIa RR genotype and allele frequency was increased in paediatric renal transplant recipients. The present findings showed that FcγRIIa genotype may be related to ESRD disease susceptibility, and FcγRIIa polymorphisms seemed to affect AR.

Matrix Metalloproteinase (MMP)-9 Levels in Children on Hemodialysis: Association with MMP-9 C-1562T Gene Polymorphism and Vitamin D Levels.

BACKGROUND AND OBJECTIVES: Data concerning the concentration of matrix metalloproteinase-9 (MMP-9) and its functional polymorphisms in chronic kidney diseases (CKD) are conflicting. The present study aimed to evaluate the levels of MMP-9in children with end stage renal diseases (ESRD) on hemodialysis (HD) and to explore its association with MMP-9 polymorphism and vitamin D levels as an important risk factors for cardiovascular diseases (CVD). METHODS: We studied 55 children with ESRD on hemodialysis and 18 healthy children served as controls. MMP-9 and vitamin D levels were measured by ELISA in serum of all patients and controls. Genotypes for MMP-9 polymorphism(C-1562T) were determined by RFLP for only 28 of the patients and all the controls. RESULTS: There were insignificantly reduced MMP-9levels of patients vs. controls, however, there was significant increase in MMP-9 levels associated with CC genotypes for(C-1562T) polymorphism compared with CT genotype (p=0.01). We found that at MMP-9 base position-1562, the frequencies of the genotypes CC and CT in Children on HD were 71.4% and 28.6% respectively while all our controls were of the CC genotype. The alleles frequencies of C and T in patients were 85.7% and 14.29% as compared to 100% and 0%, respectively in the controls. Significant decrease in vitamin D was observed in children on HD versus that in controls (p=0.008). Serum MMP9 levels and age were variables that were independently associated with CVD. CONCLUSIONS: MMP9 genetic polymorphism (C-1562T) affects MMP9alterations in ESRD children on HD and vitamin D deficiency is common in our HD pediatric patients who require attention in accordance with current practice guidelines. They probably require supplementation with higher doses of cholecalciferol.

Usefulness of serum procalcitonin as a diagnostic biomarker of infection in children with chronic kidney disease.

INTRODUCTION: Serum procalcitonin (PCT) levels are known to be low in healthy individuals in healthy subjects but are increased in patients with a severe bacterial infection. It has not been extensively studied in children with chronic kidney disease (CKD), treated either with hemodialysis (HD) or with renal transplantation. MATERIAL AND METHODS: During a 6-month period, blood samples were taken from 102 (55 HD children and 47 renal transplant recipients) children with a strong clinical suspicion of infection. Procalcitonin levels were measured by ELISA. RESULTS: Thirty-four/102 cases had proven infections as defined previously. Children with proven infections had a significantly higher PCT (0.920 ±0.24 ng/ml) than those without (0.456 ±0.53 ng/ml), p = 0.04. The ideal cutoff value derived for serum PCT was 0.5 ng/ml. This threshold value established a sensitivity of 94.1% and a specificity of 87.9%. CONCLUSIONS: This study indicates that significantly increased PCT concentration is a promising predictor of systemic bacterial infection in children with CKD, with good sensitivity and specificity. This study proposes that serum PCT is a convenient index of infection in CKD children at a cutoff value of 0.5 ng/ml.

Lymphocyte Activation Markers in Pediatric Kidney Transplant Recipients.

BACKGROUND AND OBJECTIVES: The role of CD4+CD25+ T regulatory cells (Tregs) in immune tolerance in experimental transplantation is very important but the clinical significance of circulating Tregs in the peripheral blood is undetermined. We evaluated the association between the frequency of T cell activation markers CD25 and CD71 and clinical parameters that may affect the level of these T cell markers. METHODS: In 47peditric kidney transplant (KT) recipients and 20 healthy controls, the frequency of T cell activation markers, CD25 and CD71 was measured with flow cytometry after transplantation. Two clinical protocols of induction immunosuppression were used: (1) anti-thymocyte globulin (THYMO) group (n =29) and Basiliximab (BSX) group (n=10). RESULTS: The percentage of circulating CD25 after KT was significantly lower than that in the controls. There is no significant difference between KT and the controls s regard to circulating CD71. The percentage of CD25 was significantly increased in children with acute rejection compared with those without acute rejection. Calcineurin inhibitors (CNIs) decreased the frequency of CD25 but mammalian target rapamycin (mTOR) inhibitor did not. The proportion of CD25 significantly decreased in THYMO group during the first year after transplantation. CONCLUSION: The frequency of circulating T cell activation marker CD25 in pediatric KT recipients is strongly affected by CNIs, and a high frequency of CD25 is associated with acute rejection during the early posttransplant period. The measurement of T cell activation markers, may become a useful immune monitoring tool after kidney transplantation.

Advanced glycation end products and soluble receptor as markers of oxidative stress in children on hemodialysis.

BACKGROUND: Advanced glycation end products (AGEs) have biological properties that may contribute to the mortality of children on hemodialysis (HD). This study examines the relationship of LMW fluorescence AGEs, oxidized LDL (ox-LDL), soluble receptor AGE (sRAGE) as markers of oxidative stress in children with end stage renal disease (ESRD) undergoing HD. METHOD: Thirty children with ESRD undergoing HD, and 30 healthy, age- and sex-matched children were included. Serum levels of LMW fluorescence AGEs, sRAGE, oxidized LDL (ox-LDL), pre- and post-dialysis urea, high-sensitivity C-reactive protein (hs-CRP), hemoglobin (Hb) and serum albumin (ALB), were measured. RESULTS: Abnormal serum inflammatory changes: elevated levels of LMW AGEs, sRAGE, oxLDL, CRP and urea were exhibited in HD children compared with healthy controls; more so in anemic when compared to non-anemic patients. Significant positive correlation was found between serum levels of AGEs and sRAGE. CONCLUSION: The low molecular weight form of AGEs is associated with oxidative stress in children receiving chronic HD, and may be important in the mechanisms leading to atherosclerosis and inflammation in such patients. LMW AGEs levels showed a negative correlation with sRAGE and both exhibit a significant negative relation to seum urea.

Some amino acids levels: glutamine,glutamate, and homocysteine, in plasma of children with chronic kidney disease.

BACKGROUND: The high prevalence of protein-energy malnutrition is a critical issue for patients with chronic kidney disease (CKD). Serum albumin is the most commonly used nutritional marker. Another index is plasma amino acid (AA) profile. Of these, the plasma levels of glutamine, glutamate and homocysteine, correlate well with nutritional status. We measured some plasma AAs in children with different stages CKD to provide information in monitoring the therapeutic strategy, particularly in AA supplementary therapy or protein restriction. METHODS: Three amino acids were evaluated along with albumin and high sensitivity C-reactive protein (hs-CRP) in 30 patients with advanced CKD stages 4 and 5. They were divided into two groups undergoing conservative treatment (CT) (n=15) or hemodialysis (HD) (n=15). An additional group of patients with nephrotic syndrome [CKD stage 2] was also studied to assess the alterations of plasma free amino acids with the early stage of CKD. Another 30 age- and sex-matched healthy children served as controls. RESULTS: A significant increase in plasma concentration of amino acid glutamine was observed in children with advanced CKD stages 4 and 5 when compared with controls (P=0.02).Plasma glutamine level was significantly higher in ESRD children on HD than in children with nephrotic syndrome (P=0.02). We did not find a significant difference between HD children and CT children as regard to glutamine level. Notable differences were in the plasma homocysteine level detected in the CKD groups patients, which was greater than that in controls (P=0.0001). Plasma homocysteine level was significantly higher in children on HD than in children with nephrotic syndrome (P=0.01). A significant differences was observed in hs-CRP levels between the CKD groups and the controls (P=0.04). Albumin levels were lower in CKD groups than in controls (p=0.01). Glutamine showed significant positive correlations with blood urea level (r=0.84, P=0.002) and blood ammonia level (r=0.72, P=0.0001). On multiple linear regression, urea was the only variable independently associated with an elevated plasma glutamine level (Beta=0.77, P=0.02). CONCLUSION: This study indicates that the advanced stages of CKD are associated with increased plasma concentrations of glutamine and homocysteine. Glutamine retained in the plasma of children with CRF, possibly producing higher levels of the waste products (urea and NH3). Dialysis alone is insufficient to redress completely the abnormalities in AA metabolism in ESRD children. Careful consideration of dialysis and dietary measures are necessary.

Adiponectin: an adipocyte-derived hormone, and its gene encoding in children with chronic kidney disease.

BACKGROUND: The prevalence of cardiovascular disease (CVD) and inflammation is high in patients with chronic kidney disease (CKD). Adiponectin (ADPN) is an adipocytokine that may have significant anti-inflammatory and anti-atherosclerotic effects. Low adiponectin levels have previously been found in patients with high risk for CVD. METHODS: On seventy eight advanced CKD (stages 4 and 5) pediatric patients undergoing maintenance hemodialysis( MHD) or conservative treatment (CT) the following parameters were studied: body mass index, left ventricular mass index(LVMI), serum adiponectin , cholesterol, HDL-cholesterol, high sensitivity C-reactive protein (hs CRP),interleukin 6(IL6) and single-nucleotide polymorphisms (SNPs) in the ADIPOQ gene at positions 45, and 276. Seventy age-and gender-matched healthy subjects served as control subjects. RESULTS: Markedly (P = 0.01) elevated plasma adiponectin levels were observed in CKD patients, especially CT patients, compared to control subjects. The wild type of ADIPOQ 45T > G (T) allele is the main gene for patients and controls. MHD and CT patients had significantly higher frequency of the TT genotypes of +276G > T gene (P = 0.04) compared with control subjects. A significant positive correlation was observed between plasma adiponectin and IL6 level, whereas negative correlations were found between adiponectin level, cholesterol, HDL cholesterol and hs CRP. In a stepwise backward multiple regression model only IL6 (P = 0.001) was independently associated with plasma adiponectin levels. The adiponectin gene the 276 GT+TT genotypes were associated with a higher level of adiponectin . CONCLUSIONS: The present study demonstrated that ADPN is related to several metabolic and inflammatory CV risk factors in a manner consistent with the hypothesis that this protein might have a protective role against these factors. We observed an association between the +276G>T SNP in the adiponectin gene and CKD in children. Genetic variation of +276 gene seemed to have a positive impact on circulating adiponectin levels in CKD patients.

Genetic polymorphism of ACE and the angiotensin II type1 receptor genes in children with chronic kidney disease.

AIM AND METHODS: We investigated the association between polymorphisms of the angiotensin converting enzyme-1 (ACE-1) and angiotensin II type one receptor (AT1RA1166C) genes and the causation of renal disease in 76 advanced chronic kidney disease (CKD) pediatric patients undergoing maintenance hemodialysis (MHD) or conservative treatment (CT). Serum ACE activity and creatine kinase-MB fraction (CK-MB) were measured in all groups. Left ventricular mass index (LVMI) was calculated according to echocardiographic measurements. Seventy healthy controls were also genotyped. RESULTS: The differences of D allele and DI genotype of ACE were found significant between MHD group and the controls (p = 0.0001). ACE-activity and LVMI were higher in MHD, while CK-MB was higher in CT patients than in all other groups. The combined genotype DD v/s ID+II comparison validated that DD genotype was a high risk genotype for hypertension .~89% of the DD CKD patients were found hypertensive in comparison to ~ 61% of patients of non DD genotype(p = 0.02). The MHD group showed an increased frequency of the C allele and CC genotype of the AT1RA1166C polymorphism (P = 0.0001). On multiple linear regression analysis, C-allele was independently associated with hypertension (P = 0.04). CONCLUSION: ACE DD and AT1R A/C genotypes implicated possible roles in the hypertensive state and in renal damage among children with ESRD. This result might be useful in planning therapeutic strategies for individual patients.

Endothelial nitric oxide synthase gene intron4 VNTR polymorphism in patients with chronic kidney disease.

Nitric oxide production is reduced in renal disease, partially due to decreased endothelial nitric oxide production. Evidence indicates that nitric oxide deficiency contributes to cardiovascular events and progression of kidney damage. A polymorphism in intron 4 of the endothelial constitutive nitric oxide synthase (ecNOS) gene is a candidate gene in cardiovascular and renal diseases. We investigated a potential involvement of this polymorphism in chronic renal failure. A case-control study involved 78 children with chronic kidney disease (CKD) and 30 healthy controls. All participants were genotyped for the ecNOS4 polymorphism by the polymerase chain reaction (PCR). Dialyzed (maintenance hemodialysis) and conservative treatment children had significantly higher frequency of the aa genotype and ecNOS4a allele (P<0.05) compared with controls. The combined genotype aa+ab vs. bb comparison validated that a allele is a high-risk allele for end-stage renal disease (ESRD) (P<0.05). Serum nitric oxide level was found to be lower in carriers of the ecNOS 4a allele than in noncarriers (100.29±27.32 vs. 152.73±60.39 µmol/l, P=0.04). Interestingly, 85.95% of the ecNOS 4a allele ESRD patients were found hypertensive in comparison to the 60.67% patients of non noncarriers (bb genotype) (P=0.04). Also, 35.90% of the ecNOS 4a allele ESRD patients were found to have cardiovascular disease in comparison to the 5.13% patients of noncarriers (bb genotype) (P=0.01). On multiple linear regression analysis, a allele was independently associated with hypertension (P=0.03). There was a significantly higher frequency of the ecNOS4a allele carriers among CKD children, both on MHD and conservative treatment than in controls. This suggests that the ecNOS gene polymorphism may be associated with an increased risk of chronic renal failure.

Oxidative stress markers and C-reactive protein in pediatric patients on hemodialysis.

BACKGROUND: Pediatric patients with end-stage renal disease undergoing hemodialysis (HD) are exposed to oxidative stress associated with an impairment of antioxidant defense and an overproduction of oxidative stress markers. Oxidative stress plays a significant role in the development of inflammation in these patients. OBJECTIVES: The high incidence of cardiovascular disease in HD pediatric patients is now well established and the involvement of oxidative stress has been hypothesized. This study focuses on a comparison of plasma total antioxidant capacity (TAC) and lipid peroxidation product and evaluates the relationship between these parameters and high-sensitivity C-reactive protein (hsCRP) in pediatric patients on HD. SUBJECTS AND METHODS: Plasma TAC, lipid peroxidation products, malondialdehyde (MDA) as well as hsCRP were determined in 30 pediatric patients on HD and in 20 healthy controls (HC). RESULTS: TAC and MDA levels were significantly higher in children on HD than in the HC (p < 0.001). The hsCRP values were also significantly higher in HD patients than in HC (p < 0.001). The percentage of HD pediatric patients with CRP >10 mg/l was 30%. The concentrations of TAC and MDA correlated positively with hsCRP in HD patients (TAC: r = 0.52, p < 0.08; MDA: r = 0.75, p < 0.04), but not in HC. CONCLUSION: Our study demonstrates an increase in oxidative stress in children on HD and that the susceptibility to oxidative stress is strongly related to the levels of MDA produced in plasma. hsCRP levels are higher in children on HD than in HC and this is indicative of a higher degree of inflammatory activity in the former patients. These profound disturbances in oxidative stress markers may provide an explanation for the cardiovascular complications in HD patients.

Intradialytic and postdialytic platelet activation, increased platelet phosphatidylserine exposure and ultrastructural changes in platelets in children with chronic uremia.

The present research evaluated the intradialytic and postdialytic changes in platelet factor-4 and beta-thromboglobulin plasma levels by enzyme-linked immunoadsorbent assay method and platelet aggregation by ADP as well as flow cytometric percentage of annexin-V-positive platelets as a measure of phosphatidylserine externalization and ultrastructural examination of platelets in 37 uremic patients on regular hemodialysis and 25 age-matched and sex-matched controls. Platelet factor-4 plasma levels increased, remain consistently high during hemodialysis session (20.24 +/- 3.05 IU/ml after 30 min, P < 0.001 and 23.67 +/- 3.68 IU/ml after 240 min, P < 0.001) and returned to control values (6.10 +/- 1.54 IU/ml) only after 24 h following the end of the session. beta-Thromboglobulin showed a trend similar to that of platelet factor-4. Platelet aggregation by ADP showed reduced function in comparison with controls (69.32 +/- 12.37 versus 91.95 +/- 1.59%, P < 0.001). Flow cytometric percentage of annexin-V-positive platelet was significantly elevated (P < 0.001) in uremic patients when compared with normal controls. Ultrastructural studies of platelets 30 min after starting of dialysis showed degranulation of its granules and at 240 min showed complete degranulation, whereas in the postdialytic phase (12 h after the end of dialysis) refilled alpha-granules started to appear. Positive correlations were found between platelet concentration and platelet factor-4 and beta-thromboglobulin plasma levels during and after dialysis (P < 0.001) and with annexin-V-positive platelets percentage (P < 0.001). In conclusion, activated platelets were found in chronic hemodialysis patients, a finding that may explain why uremic patients often suffer from thrombotic accidents. The platelet activation is associated with exposure of phosphatidylserine on the platelet exterior. Platelet factor-4 and beta-thromboglobulin are released from platelets as a result of a defect in their granules membrane as shown by the electron microscopy, mainly as a consequence of the blood-membrane contact during dialysis, and they return only slowly to control values.