Candida Administration in 5/6 Nephrectomized Mice Enhanced Fibrosis in Internal Organs: An Impact of Lipopolysaccharide and (1→3)-ß-D-Glucan from Leaky Gut.

Uremic toxins and gut dysbiosis in advanced chronic kidney disease (CKD) can induce gut leakage, causing the translocation of gut microbial molecules into the systemic circulation. Lipopolysaccharide (LPS) and (1→3)-ß-D-glucan (BG) are the major gut microbial molecules of Gram-negative bacteria and fungi, respectively, and can induce inflammation in several organs. Here, the fibrosis in the kidney, liver, and heart was investigated in oral C. albicans-administered 5/6 nephrectomized (Candida-5/6 Nx) mice. At 20 weeks post 5/6 Nx, Candida-5/6 Nx mice demonstrated increased 24 h proteinuria, liver enzymes, and serum cytokines (TNF-α, IL-6, and IL-10), but not weight loss, systolic blood pressure, hematocrit, serum creatinine, or gut-derived uremic toxins (TMAO and indoxyl sulfate), compared to in 5/6 Nx alone. The gut leakage in Candida-5/6 Nx was more severe, as indicated by FITC-dextran assay, endotoxemia, and serum BG. The areas of fibrosis from histopathology, along with the upregulated gene expression of Toll-like receptor 4 (TLR-4) and Dectin-1, the receptors for LPS and BG, respectively, were higher in the kidney, liver, and heart. In vitro, LPS combined with BG increased the supernatant IL-6 and TNF-α, upregulated the genes of pro-inflammation and pro-fibrotic processes, Dectin-1, and TLR-4 in renal tubular (HK-2) cells and hepatocytes (HepG2), when compared with LPS or BG alone. This supported the pro-inflammation-induced fibrosis and the possible LPS-BG additive effects on kidney and liver fibrosis. In conclusion, uremia-induced leaky gut causes the translocation of gut LPS and BG into circulation, which activates the pro-inflammatory and pro-fibrotic pathways, causing internal organ fibrosis. Our results support the crosstalk among several organs in CKD through a leaky gut.

Uremia-Induced Gut Barrier Defect in 5/6 Nephrectomized Mice Is Worsened by Candida Administration through a Synergy of Uremic Toxin, Lipopolysaccharide, and (1➔3)-ß-D-Glucan, but Is Attenuated by Lacticaseibacillus rhamnosus L34.

A chronic kidney disease (CKD) causes uremic toxin accumulation and gut dysbiosis, which further induces gut leakage and worsening CKD. Lipopolysaccharide (LPS) of Gram-negative bacteria and (1➔3)-ß-D-glucan (BG) of fungi are the two most abundant gut microbial molecules. Due to limited data on the impact of intestinal fungi in CKD mouse models, the influences of gut fungi and Lacticaseibacillus rhamnosus L34 (L34) on CKD were investigated using oral C. albicans-administered 5/6 nephrectomy (5/6Nx) mice. At 16 weeks post-5/6Nx, Candida-5/6Nx mice demonstrated an increase in proteinuria, serum BG, serum cytokines (tumor necrotic factor-α; TNF-α and interleukin-6), alanine transaminase (ALT), and level of fecal dysbiosis (Proteobacteria on fecal microbiome) when compared to non-Candida-5/6Nx. However, serum creatinine, renal fibrosis, or gut barrier defect (FITC-dextran assay and endotoxemia) remained comparable between Candida- versus non-Candida-5/6Nx. The probiotics L34 attenuated several parameters in Candida-5/6Nx mice, including fecal dysbiosis (Proteobacteria and Bacteroides), gut leakage (fluorescein isothiocyanate (FITC)-dextran), gut-derived uremic toxin (trimethylamine-N-oxide; TMAO) and indoxyl sulfate; IS), cytokines, and ALT. In vitro, IS combined with LPS with or without BG enhanced the injury on Caco-2 enterocytes (transepithelial electrical resistance and FITC-dextran permeability) and bone marrow-derived macrophages (supernatant cytokines (TNF-α and interleukin-1 ß; IL-1ß) and inflammatory genes (TNF-α, IL-1ß, aryl hydrocarbon receptor, and nuclear factor-κB)), compared with non-IS activation. These injuries were attenuated by the probiotics condition media. In conclusion, Candida administration worsens kidney damage in 5/6Nx mice through systemic inflammation, partly from gut dysbiosis-induced uremic toxins, which were attenuated by the probiotics. The additive effects on cell injury from uremic toxin (IS) and microbial molecules (LPS and BG) on enterocytes and macrophages might be an important underlying mechanism.

Lactobacillus rhamnosus L34 attenuates chronic kidney disease progression in a 5/6 nephrectomy mouse model through the excretion of anti-inflammatory molecules.

BACKGROUND: Although pathogenic gut microbiota causes gut leakage, increases translocation of uremic toxins into circulation and accelerates CKD progression, the local strain of Lactobacillus rhamnosus L34 might attenuate gut leakage. We explored the effects of L34 on kidney fibrosis and levels of gut-derived uremic toxins (GDUTs) in 5/6 nephrectomy (5/6Nx) mice. METHODS: At 6 weeks post-5/6Nx in mice, either L34 (1 × 106 CFU) or phosphate buffer solution (as 5/6Nx control) was fed daily for 14 weeks. In vitro, the effects of L34-conditioned media with or without indoxyl sulfate (a representative GDUT) on inflammation and cell integrity (transepithelial electrical resistance; TEER) were assessed in Caco-2 (enterocytes). In parallel, the effects on proinflammatory cytokines and collagen expression were assessed in HK2 proximal tubular cells. RESULTS: At 20 weeks post-5/6Nx, L34-treated mice showed significantly fewer renal injuries, as evaluated by (i) kidney fibrosis area (P < 0.01) with lower serum creatinine and proteinuria, (ii) GDUT including trimethylamine-N-oxide (TMAO) (P = 0.02) and indoxyl sulfate (P < 0.01) and (iii) endotoxin (P = 0.03) and serum TNF-α (P = 0.01) than 5/6Nx controls. Fecal microbiome analysis revealed an increased proportion of Bacteroidetes in 5/6Nx controls. After incubation with indoxyl sulfate, Caco-2 enterocytes had higher interleukin-8 and nuclear factor κB expression and lower TEER values, and HK2 cells demonstrated higher gene expression of TNF-α, IL-6 and collagen (types III and IV). These indoxyl sulfate-activated parameters were attenuated with L34-conditioned media, indicating the protective role of L34 in enterocyte integrity and renal fibrogenesis. CONCLUSION: L34 attenuated uremia-induced systemic inflammation by reducing GDUTs and gut leakage that provided renoprotective effects in CKD.

The failure of glomerular filtration rate estimating equations among obese population.

BACKGROUND: Obesity is a major public health with increasing numbers of obese individuals are at risk for kidney disease. However, the validity of serum creatinine-based glomerular filtration rate (GFR) estimating equations in obese population is yet to be determined. METHODS: We evaluated the performance of the reexpressed Modification of Diet in Renal Disease (MDRD), reexpressed MDRD with Thai racial factor, Thai estimated GFR (eGFR) as well as Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equations among obese patients, defined as body mass index (BMI) ≥25 kg/m2 with the reference measured GFR (mGFR) determined by 99mTc-diethylene triamine penta-acetic acid (99mTc-DTPA) plasma clearance method. Serum creatinine levels were measured using standardized enzymatic method simultaneously with GFR measurement. The statistical methods in assessing agreement for continuous data including total deviation index (TDI), concordance correlation coefficient (CCC), and coverage probability (CP) for each estimating equation were compared with the reference mGFR. Accuracy within 10% representing the percentage of estimations falling within the range of ±10% of mGFR values for all equations were also tested. RESULTS: A total of 240 Thai obese patients were finally recruited with mean BMI of 31.5 ± 5.8 kg/m2. In the total population, all eGFR equations underestimated the reference mGFR. The average TDI values were 55% indicating that 90% of the estimates falling within the range of -55 to +55% of the reference mGFR. The CP values averaged 0.23 and CCC scores ranged from 0.75 to 0.81, reflecting the low to moderate levels of agreement between each eGFR equation and the reference mGFR. The proportions of patients achieving accuracy 10% ranged from 23% for the reexpressed MDRD equation to 33% for the Thai eGFR formula. Among participants with BMI more than 35 kg/m2 (n = 48), the mean error of all equations was extremely wide and significantly higher for all equations compared with the lower BMI category. Also, the strength of agreement evaluated by TDI, CCC, and CP were low in the subset of patients with BMI ≥35 kg/m2. CONCLUSION: Estimating equations generally underestimated the reference mGFR in subjects with obesity. The overall performance of GFR estimating equations demonstrated poor concordance with the reference mGFR among individuals with high BMI levels. In certain clinical settings such as decision for dialysis initiation, the direct measurements of GFR are required to establish real renal function among obese population.

Effect of Maintenance Intravenous Iron Treatment on Erythropoietin Dose in Chronic Hemodialysis Patients: A Multicenter Randomized Controlled Trial.

BACKGROUND: There is no consensus on intravenous (IV) iron supplement dose, schedule, and serum ferritin target in functional iron deficiency anemia to maintain optimum target levels of iron stores by several guidelines. OBJECTIVE: To examine the effect of IV iron supplementation to different targets of serum ferritin on erythropoietin dose and inflammatory markers in chronic hemodialysis (HD) patients with functional iron deficiency anemia. DESIGN: A multicenter, randomized, open-label study. SETTING: In a developing country, Thailand. PATIENTS: Chronic HD patients with functional iron deficiency anemia. MEASUREMENTS: Erythropoietin resistance index, high-sensitivity C-reactive protein, and fibroblast growth factor 23. METHODS: Two hundred adult chronic HD patients with transferrin saturation less than 30% and serum ferritin of 200 to 400 ng/mL were randomized 1:1 to maintain serum ferritin 200 to 400 ng/mL (low-serum ferritin group, N = 100) or 600 to 700 ng/mL (high-serum ferritin group, N = 100). During a 6-week titration period, participants randomized to the high-serum ferritin group initially received 600 mg IV iron (100 mg every week), while the participants in the low-serum ferritin group did not receive IV iron. During the 6-month follow-up period, the dose of IV iron was adjusted by protocol. RESULTS: The mean dose of IV iron was 108.3 ± 28.2 mg/month in the low-serum ferritin group and 192.3 ± 36.2 mg/month in the high-serum ferritin group. The mean serum ferritin was 367.0 ± 224.9 ng/mL in the low ferritin group and 619.6 ± 265.2 ng/mL in the high ferritin group. The erythropoietin resistance index was significantly decreased in the high-serum ferritin group compared to the low-serum ferritin group after receiving IV iron in the 6-week titration period (mean difference: -113.43 ± 189.14 vs 41.08 ± 207.38 unit/week/g/dL; P < .001) and 3-month follow-up period (mean differences: -88.88 ± 234.43 vs -10.48 ± 217.75 unit/week/g/dL; P = .02). LIMITATIONS: Short follow-up period. CONCLUSION: Maintaining a serum ferritin level of 600 to 700 ng/mL by IV iron administration of approximately 200 mg per month as a maintenance protocol can decrease erythropoietin dose requirements in chronic HD patients with functional iron deficiency anemia. TRIALS REGISTRATION: The study was registered with the Thai Clinical Trials Registry TCTR20180903003.

CONTEXTE: Il n'existe aucun consensus sur la dose et la posologie du supplément de fer administré par voie intraveineuse (IV) dans le traitement de l'anémie ferriprive fonctionnelle, ni sur la cible de ferritine sérique permettant de maintenir les réserves ferriques optimales définies par les différentes recommandations. OBJECTIF: Examiner l'effet d'un supplément de fer IV, à différentes cibles de ferritine sérique, sur la dose d'érythropoïétine et les marqueurs inflammatoires de patients sous hémodialyse chronique atteints d'une anémie ferriprive fonctionnelle. TYPE D'ÉTUDE: Essai multicentrique ouvert à répartition aléatoire. CADRE: L'étude s'est tenue en Thaïlande, un pays en développement. SUJETS: Des patients sous hémodialyse chronique atteints d'anémie ferriprive fonctionnelle. MESURES: L'indice de résistance à l'érythropoïétine, la protéine C réactive très sensible et le facteur de croissance des fibroblastes 23. MÉTHODOLOGIE: Deux cents adultes sous HD chronique présentant une saturation en transferrine inférieure à 30 % et un taux de ferritine sérique entre 200 et 400 ng/mL ont été répartis en deux groupes (ratio 1:1). On visait le maintien d'un taux de ferritine sérique entre 200 et 400 ng/mL dans le premier groupe (faible taux de ferritine sérique; n=100) et entre 600 et 700 ng/mL dans le deuxième groupe (taux élevé de ferritine sérique; n=100). Au cours d'une période d'ajustement de six semaines, les sujets du groupe à taux élevé de ferritine sérique ont initialement reçu une dose de 600 mg de fer par IV (100 mg par semaine) alors que les sujets de l'autre groupe n'en ont pas reçu. La dose de fer IV a été ajustée selon le protocole au cours des six mois de suivi. RÉSULTATS: La dose moyenne de fer administrée par IV était de 108,3 ±28,2 mg/mois pour les sujets du groupe à faible taux de ferritine sérique et de 192,3 ±36,2 mg/mois pour les sujets du groupe à taux élevé de ferritine sérique. En ce qui concerne les taux de ferritine sérique, ceux-ci s'établissaient respectivement à 367,0 ±224,9 ng/mL et à 619,6 ±265,2 ng/mL. Les sujets du groupe à taux élevé de ferritine sérique présentaient un indice de résistance à l'érythropoïétine significativement réduit par rapport à ceux du groupe à faible taux après avoir reçu un supplément de fer IV durant la période d'ajustement de six semaines (différence moyenne: -113,43 ±189,14 contre 41,08 ±207,38 unités/semaine/g/dL; p<0,001) et après trois mois de suivi (différence moyenne: -88,88 ±234,43 contre -10,48 ±217,75 unités/semaine/g/dL; p=0,02). LIMITES: Courte période de suivi. CONCLUSION: Le maintien d'un taux de ferritine sérique entre 600 et 700 ng/mL par l'administration IV d'une supplémentation en fer, à raison d'environ 200 mg par mois (protocole de maintien), peut contribuer à réduire la posologie d'érythropoïétine chez les patients hémodialysés et atteints d'anémie ferriprive fonctionnelle. ENREGISTREMENT DE L'ESSAI: L'essai a été enregistré selon le registre des essais cliniques thaïlandais TCTR20180903003.

Bone turnover markers predict type of bone histomorphometry and bone mineral density in Asian chronic haemodialysis patients.

BACKGROUND: Although the levels of intact parathyroid hormone (iPTH) are well-controlled following the Kidney Disease Outcomes Quality Initiative guideline, the incidence of osteoporosis and fracture are still high in haemodialysis (HD) patients. This study was conducted to investigate the correlation between bone turnover markers, bone mineral density (BMD), and bone histomorphometry in HD patients. METHODS: Twenty-two chronic HD patients were enrolled. Serum levels of bone turnover markers were measured. Double tetracycline-labelled iliac crest bone specimens were evaluated using specialized a computer program (Osteomeasure). The types of bone histomorphometry were classified based on turnover, mineralization and volume. BMD and coronary artery calcification were also determined. RESULTS: Bone histomorphometry revealed osteitis fibrosa (50%), adynamic bone disease (45%) and mixed uremic osteodystrophy (5%). Serum iPTH level predicted high bone turnover with area under the receiver operating characteristic (ROC) of 0.833 (95% CI = 0.665-1.000, P = 0.008). Serum TRAP-5b also had ROC of 0.733 (95% CI = 0.517-0.950, P = 0.065). In addition, when using serum iPTH (cut-off 484.50 ng/mL) or serum TRAP-5b (cut-off 1.91 pg./mL) to predict high turnover, the sensitivity was 0.917. On the other hand, when both iPTH and TRAP-5B were above these cut-off, the specificity was 1.000. Low BMD and severe vascular calcification were commonly identified. However, there were no significant correlations between bone biomarkers and BMD or severe vascular calcification. CONCLUSION: Although iPTH levels were close to the target of Kidney Disease Outcomes Quality Initiative guideline, abnormal bone histomorphometry, BMD, and severe vascular calcification are still common. Bone biopsy is still crucially required as an accurate diagnostic tool in providing optimal guide for the treatment. © 2019 Asian Pacific Society of Nephrology.

Furosemide Stress Test as a Predicting Biomarker for Delayed Graft Function in Kidney Transplantation.

BACKGROUND: Delayed graft function (DGF) could worsen early and long-term outcomes of kidney transplantation (KT). DGF is caused by several pre-transplantation and perioperative factors in both donors and recipients. At present, there are no biomarkers or tests during the immediate post-KT period that can accurately predict the development of DGF. MATERIALS AND METHODS: This prospective study was conducted in deceased donor KT (DDKT) at King Chulalongkorn Memorial Hospital, Thailand. All recipients underwent furosemide stress test (FST) by receiving a single dose of intravenous furosemide, 1.5 mg/kg at 3 h after allograft reperfusion. We determined the correlations between DGF (requiring dialysis within the first week after transplantation) and the values of urine volume recorded hourly after FST until 6 h, the parameters of postoperative dynamic tests, including resistive index (RI) of renal arteries and effective renal plasma flow (ERPF), and urine neutrophil gelatinase-associated lipocalin (NGAL). RESULTS: Of the 59 total DDKT recipients enrolled, 24 developed DGF. The FST is a more accurate biomarker than urine NGAL, RI of renal arteries, and ERPF in the prediction of DGF. The 4-h urine volume less than 350 mL (FST non-responsive) was the best cut-off value in predicting DGF with 87.5% sensitivity, 82.9% specificity, and 82.5% accuracy. Multiple logistic regression analyses showed an odds ratio of 0.993 (0.986-0.999, p = 0.035) for the 4-h urine volume to predict DGF. CONCLUSIONS: The FST is a simple and accurate biomarker for predicting DGF in early post-KT period. Close monitoring and well prepared dialysis are suggested in patients with urine volume < 350 mL after 4 h of FST. The FST non-responsive patients could be the target for further DGF preventive intervention. ClinicalTrials.gov identifier: NCT03071536.

Immunogenicity and safety of quadrivalent human papillomavirus types 6/11/16/18 recombinant vaccine in chronic kidney disease stage IV, V and VD.

Background: Up to >80% of sexually active adults will become infected with human papillomavirus (HPV) during their lifetime. Persistent HPV infection can result in cervical, vulvovaginal, penile and anogenital cancer. Clinical studies have shown the efficacy of three doses of quadrivalent HPV-6/11/16/18 L1 virus-like particle (VLP) vaccination, at Day 0, Month 2 and Month 6, to lower the occurrence of HPV infection and its complications. However, immunogenicity and safety of the HPV vaccine have not been proven in the chronic kidney disease (CKD) population. Methods: Sixty CKD stage IV, V and VD patients were enrolled for quadrivalent HPV-6/11/16/18 vaccination. A dose of vaccine was given at Day 0, Month 2 and Month 6. Each dose contained 20 µg HPV-6 L1 VLP, 40 µg HPV-11 L1 VLP, 40 µg HPV-16 L1 VLP and 20 µg HPV-18 L1 VLP, along with 225 µg of amorphous aluminum hydroxyphosphate sulfate adjuvant. HPV type-specific antibody response to neutralizing epitopes on HPV-6/11/16/18 was performed by multiplexed, competitive Luminex® immunoassays (cLIA) at Day 0 and Month 7. Results: At Day 0, anti-HPV seropositivity was 0-6.6% depending on HPV genotype. Patients who received three doses of vaccine had 98.2, 100, 100 and 98.2% seropositivity for genotypes 6/11/16/18, respectively. The average cLIA at Month 7 for genotypes 6/11/16/18 were 928.4 ± 231.1, 1136.1 ± 264.6, 6951.0 ± 1872.3 and 2196.3 ± 761.2 milliMerck units (mMu)/mL, respectively. No serious vaccine-related adverse events were observed. Conclusions: Quadrivalent HPV vaccine has been well tolerated, is safe and provides excellent immunogenicity in late-stage CKD.

Exfoliated mesothelial cell and CA-125 in automated peritoneal dialysis (APD) and continuous ambulatory peritoneal dialysis (CAPD) patients.

OBJECTIVE: Automated peritoneal dialysis (APD) becomes the first option for peritoneal dialysis, nowadays overtaking continuous ambulatory peritoneal dialysis (CAPD) in many countries. The comparison of peritoneal membrane alteration in CAPD and APD is inconclusive. The authors therefore compared the peritoneal membrane changes in patients undergoing CAPD and APD. MATERIAL AND METHOD: In naive end stage renal disease patients, the choice of PD modes (CAPD or APD) was dependent on the patient's decision. Thirty-six CAPD and 25APD patients with a total of 287 patient-months were compared. The peritoneal mass parameter, exfoliated mesothelial cell (MTC) and dialysate CA-125, as well as modified peritoneal equilibrium test (mPET) with 4.25% dextrose solution was simultaneously evaluated at 1 and 6 month follow-up. RESULTS: Although the peritoneal function (as measured by D/P creatinine, D/D0 glucose, sodium dipping, and dialysate protein loss), adequacy, serum albumin, nutritional status, and residual renal function showed no significant differences between groups at 1 and 6 months, CA-125 but not MTC was higher in APD compared with CAPD at the first month of PD beginning. Due to the single time-point measurement limitation, the authors compared the peritoneal mass parameter differences between 1 and 6 month. During 6-month follow-up, CA-125 decreased 30 +/- 5% vs. 7 +/- 5% and MTC decreased 5 +/- 12% vs. 40 +/- 11% in APD and CAPD, respectively. The higher CA-125 reduction in APD and greater changes of MTC in CAPD suggested that there was less viable mesothelial cell in APD compared with CAPD. CONCLUSION: The authors observed that both APD and CAPD damaged peritoneum. However, there might be higher peritoneal injury in APD patients. The proper randomization study in longer follow-up period is mandatory to confirm this observation.

Effects of different levels of endotoxin contamination on inflammatory cytokine production by peripheral blood mononuclear cells after high-flux hemodialysis.

BACKGROUND: In Thailand, dialysate endotoxin contamination levels vary from less than 0.001 to 2.0 EU/ml. This difference has prompted an investigation on the production of proinflammatory cytokines and counter-inflammatory mediators of peripheral blood mononuclear cells (PBMCs) after high-flux dialysis. METHODS: Patients from four hemodialysis (HD) centers who met the inclusion/exclusion criteria were enrolled into the study. PBMCs were isolated by Ficoll density gradient centrifugation and cultured. Supernatants were tested for interleukin 6 (IL-6), IL-1ß and IL-1 receptor antagonist (IL-1Ra) concentration by ELISA. RESULTS: HD centers 1, 2, 3 and 4 had mean dialysate endotoxin contamination levels of 0.001, 0.026, 0.558 and 1.960 EU/ml, respectively. HD center 4 had the highest levels of IL-6 (1,052.3 ± 240.7 pg/10(6) PBMCs), IL-1ß (1,297.1 ± 334.6 pg/10(6) PBMCs) and IL-1Ra (2,713.4 ± 1,255.3 pg/10(6) PBMCs). There were no significant differences in cytokine production between HD centers 1 and 2. CONCLUSION: Our study showed that ultrapure dialysate can minimize the risk of stimulating inflammatory cells. Ultrapure dialysate may prevent or delay endotoxin exposure-related complications.

Genome-wide association scan for diabetic nephropathy susceptibility genes in type 1 diabetes.

OBJECTIVE: Despite extensive evidence for genetic susceptibility to diabetic nephropathy, the identification of susceptibility genes and their variants has had limited success. To search for genes that contribute to diabetic nephropathy, a genome-wide association scan was implemented on the Genetics of Kidneys in Diabetes collection. RESEARCH DESIGN AND METHODS: We genotyped approximately 360,000 single nucleotide polymorphisms (SNPs) in 820 case subjects (284 with proteinuria and 536 with end-stage renal disease) and 885 control subjects with type 1 diabetes. Confirmation of implicated SNPs was sought in 1,304 participants of the Diabetes Control and Complications Trial (DCCT)/Epidemiology of Diabetes Interventions and Complications (EDIC) study, a long-term, prospective investigation of the development of diabetes-associated complications. RESULTS: A total of 13 SNPs located in four genomic loci were associated with diabetic nephropathy with P < 1 x 10(-5). The strongest association was at the FRMD3 (4.1 protein ezrin, radixin, moesin [FERM] domain containing 3) locus (odds ratio [OR] = 1.45, P = 5.0 x 10(-7)). A strong association was also identified at the CARS (cysteinyl-tRNA synthetase) locus (OR = 1.36, P = 3.1 x 10(-6)). Associations between both loci and time to onset of diabetic nephropathy were supported in the DCCT/EDIC study (hazard ratio [HR] = 1.33, P = 0.02, and HR = 1.32, P = 0.01, respectively). We demonstratedexpression of both FRMD3 and CARS in human kidney. CONCLUSIONS: We identified genetic associations for susceptibility to diabetic nephropathy at two novel candidate loci near the FRMD3 and CARS genes. Their identification implicates previously unsuspected pathways in the pathogenesis of this important late complication of type 1 diabetes.

Albumin suppresses vascular endothelial growth factor via alteration of hypoxia-inducible factor/hypoxia-responsive element pathway.

Reduction of vascular endothelial growth factor (VEGF) expression plays a crucial role in chronic kidney disease (CKD). In order to clarify a cause of VEGF suppression in CKD, we examined an interaction between proteinuria and VEGF. Rat proximal tubular cells were subjected to hypoxia with or without albumin to mimic proteinuric conditions, and VEGF expression was assessed by real-time quantitative PCR and enzyme-linked immunosorbent assays. Albumin significantly reduced VEGF expression under hypoxia. Luciferase activity controlled by hypoxia-responsive element (HRE) was suppressed by albumin, demonstrating suppression of the hypoxia-inducible factor (HIF)/HRE pathway. Studies utilizing a proteasome inhibitor and a prolyl hydroxylase inhibitor showed that mechanisms of HIF/HRE pathway suppression by albumin load did not involve degradation of HIF protein levels. Further, albumin did not change HIF mRNA levels. Our data, for the first time, suggest a clear 'link' between proteinuria and hypoxia, the two principal pathogenic factors for CKD progression.

High glucose blunts vascular endothelial growth factor response to hypoxia via the oxidative stress-regulated hypoxia-inducible factor/hypoxia-responsible element pathway.

Vascular endothelial growth factor (VEGF) is an important survival factor for endothelial cells in hypoxic environments. High glucose regulates certain aspects of VEGF expression in various cell types, including proximal tubular cells. Thus, ambient glucose levels may modulate the progression of chronic kidney disease, especially diabetic nephropathy. Immortalized rat proximal tubular cells (IRPTC) were cultured for 24 h under hypoxic conditions (1% O(2)), with or without high d-glucose (25 mM), or with or without high l-glucose (25 mM). Controls included culture in normoxic conditions and normal d-glucose (5.5 mM). VEGF mRNA expression was assessed by real-time quantitative PCR, and VEGF protein in the supernatant was assessed by ELISA. Hypoxia increased VEGF expression. This response was significantly blunted by high d-glucose (1.98 +/- 0.11- versus 2.65 +/- 0.27-fold increase for VEGF mRNA expression, 252.8 +/- 14.7 versus 324.0 +/- 11.5 pg/10(5) cells for VEGF protein; P < 0.05 both) but not by high l-glucose. It is interesting that hydrogen peroxide also blunted this response, whereas alpha-tocopherol restored the VEGF response to hypoxia in the presence of high d-glucose. For determination of involvement of the hypoxia-inducible factor (HIF)/hypoxia-responsible element (HRE) pathway, IRPTC that were stably transfected with HRE-luciferase were cultured under the previous conditions. High d-glucose also reduced luciferase activity under hypoxia, whereas alpha-tocopherol restored activity. In vivo experiments using streptozotocin-induced diabetic rats confirmed that hyperglycemia blunted HIF-HRE pathway activation. Insulin treatment restored activation of the HIF-HRE pathway in streptozotocin-induced diabetic rats. In conclusion, high glucose blunts VEGF response to hypoxia in IRPTC. This effect is mediated by the oxidative stress-regulated HIF-HRE pathway.

Water and sodium disorders in children undergoing surgical treatment of brain tumors.

BACKGROUND: Many children with brain tumors have abnormalities in water and sodium homeostasis, such as diabetes insipidus (DI), the syndrome of inappropriate antidiuretic hormone secretion (SIADH) and cerebral salt wasting syndrome (CSW), either pre-operatively or post-operatively. But little data have been published on the incidence, time of onset and clinical course of water and sodium disorders in children with brain tumors. OBJECTIVES: To characterize the water and sodium disorders in children undergoing surgical treatment for brain tumors. PATIENTS AND METHOD: The medical records of children with brain tumors admitted for surgery at King Chulalongkorn Memorial Hospital from January 1997 to December 2002 were retrospectively reviewed. RESULTS: There were 79 patients, 46 males and 33 females included in this study. Water and sodium disorders occurred in 36 patients (45.57%), 23 patients had DI, 12 patients had SIADH and 1 patient had cerebral salt wasting syndrome (CSW). Nine of the 36 patients had pre-operative onset of water and sodium disorders (8 DI and 1 SIADH). The development of DI and SIADH was associated with the location of the brain tumors. Patients with sellar and suprasellar tumors had the highest incidence of DI (75.86%). Patients with tumors of the cerebral hemispheres had the highest incidence of SIADH (44.44%). Onset of post-operative DI ranged from 2 to 19 hours (mean 9.68 hours) and most patients developed SIADH within 24 hours post-operatively. CONCLUSIONS: DI and SIADH are common problems in children with brain tumors, especially after surgical treatment and the onset usually occurs within 24 hours after surgery. Patients with sellar and suprasellar tumors are more likely to have DI, patients with tumors of cerebral hemispheres and posterior fossa tumors are more likely to have SIADH.