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DNASE1L3 is an extracellular nuclease that digests chromatin released from apoptotic cells. DNASE1L3 variants impair the enzyme function, enhance autoantibody production and type I interferon (IFN-I) responses, and cause different autosomal recessive phenotypes ranging from hypocomplementemic urticarial vasculitis syndrome to full-blown systemic lupus erythematosus (SLE). Kidney involvement in patients with DNASE1L3 variants is poorly characterized. Herein, we describe the clinical course of 3 children with monogenic SLE due to DNASE1L3 variants who developed refractory glomerulonephritis leading to kidney failure. They had different renal histopathological patterns (ie, membranous, endocapillary, and extracapillary glomerulonephritis and thrombotic microangiopathy), all belonging to the lupus nephritis (LN) spectrum. One patient had a mixed phenotype, showing an overlap between SLE and antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis. Using immunofluorescence, we detected glomerular expression of the IFN-I-induced human myxovirus resistance protein 1 (MXA), which was particularly evident in glomerular endothelial cells. Two of the patients had increased expression of interferon-stimulated genes in the peripheral blood, and all 3 patients had reduced serum DNAse activity. Our findings suggest that DNASE1L3-related glomerulonephritis can be included in the spectrum of IFN-I-mediated kidney disorders and provide the rationale for IFN-I-directed therapies in order to improve the poor outcome of this rare condition., (Copyright © 2024 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.)

Background and Hypothesis: Persistent distressing psychotic-like experiences (PLE) are associated with impaired functioning and future psychopathology. Prior research suggests that physical activities may be protective against psychopathology. However, it is unclear whether physical activities may interact with genetics in the development of psychosis., Study Design: This study included 4679 participants of European ancestry from the Adolescent Brain Cognitive Development Study. Persistent distressing PLE was derived from the Prodromal-Questionnaire-Brief Child Version using four years of data. Generalized linear mixed models tested the association between polygenic risk score for schizophrenia (PRS-SCZ), physical activities, and PLE. The models adjusted for age, sex, parental education, income-to-needs ratio, family history of psychosis, body mass index, puberty status, principal components for PRS-SCZ, study site, and family., Study Results: PRS-SCZ was associated with a greater risk for persistent distressing PLE (adjusted relative risk ratio (RRR) = 1.14, 95% CI [1.04, 1.24], P = .003). Physical activity was associated with less risk for persistent distressing PLE (adjusted RRR = 0.87, 95% CI [0.79, 0.96], P = .008). Moreover, physical activities moderated the association between PRS-SCZ and persistent distressing PLE (adjusted RRR = 0.89, 95% CI [0.81, 0.98], P = .015), such that the association was weaker as participants had greater participation in physical activities., Conclusions: These findings demonstrate that the interaction between genetic liability and physical activities is associated with trajectories of distressing PLE. Further research is needed to understand the mechanisms of physical activities and genetic liability for schizophrenia in the development of psychosis., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Maryland Psychiatric Research Center. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Schizophrenia may represent a trade-off in the evolution of human-specific ontogenetic mechanisms that guide neurodevelopment. Human Accelerated Regions (HARs) are evolutionary markers functioning as neurodevelopmental transcription enhancers that have been associated with brain configuration, neural information processing, and schizophrenia risk. Here, we have investigated the influence of HARs' polygenic load on neuroanatomical measures through a case-control approach (128 patients with schizophrenia and 115 controls). To this end, we have calculated the global schizophrenia Polygenic Risk Score (Global PRS SZ ) and that specific to HARs (HARs PRS SZ ). We have also estimated the polygenic burden restricted to the HARs linked to transcriptional regulatory elements active in the foetal brain (FB-HARs PRS SZ ) and the adult brain (AB-HARs PRS SZ ). We have explored the main effects of the PRSs and the PRSs x diagnosis interactions on brain regional cortical thickness (CT) and surface area (SA). The results indicate that a higher FB-HARs PRS SZ is associated with patients' lower SA in the lateral orbitofrontal cortex, the superior temporal cortex, the pars triangularis and the paracentral lobule. While noHARs-derived PRSs show an effect on the risk, our neuroanatomical findings suggest that the human-specific transcriptional regulation during the prenatal period underlies SA variability, highlighting the role of these evolutionary markers in the schizophrenia genomic architecture., (© 2023. Springer Nature Limited.)

Introduction: Immunity to Human leukocyte antigen (HLA) cannot explain all cases of ABMR, nor the differences observed in the outcome of kidney recipients with circulating DSAs endowed with similar biologic characteristics. Thus, increasing attention has recently been focused on the role of immunity to non-HLA antigenic targets., Methods: We analyzed humoral auto- and alloimmune responses to the non-HLA antigen glutathione S-transferase theta 1 (GSTT1), along with development of de novo ( dn )HLA-DSAs, in a cohort of 146 pediatric non-sensitized recipients of first kidney allograft, to analyze its role in ABMR and graft loss. A multiplex bead assay was employed to assess GSTT1 antibodies (Abs)., Results: We observed development of GSTT1 Abs in 71 recipients after transplantation, 16 with MFI > 8031 (4th quartile: Q4 group). In univariate analyses, we found an association between Q4-GSTT1Abs and ABMR and graft loss, suggesting a potential role in inducing graft damage, as GSTT1 Abs were identified within ABMR biopsies of patients with graft function deterioration in the absence of concomitant intragraft HLA-DSAs. HLA-DSAs and GSTT1 Abs were independent predictors of graft loss in our cohort. As GSTT1 Ab development preceded or coincided with the appearance of dn HLA-DSAs, we tested and found that a model with the two combined parameters proved more fit to classify patients at risk of graft loss., Discussion: Our observations on the harmful effects of GSTT1Abs, alone or in combination with HLA-DSAs, add to the evidence pointing to a negative role of allo- and auto-non-HLA Abs on kidney graft outcome., Competing Interests: Authors BR and JH were employed by Immucor Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Comoli, Cioni, Ray, Tagliamacco, Innocente, Caridi, Bruschi, Hariharan, Fontana, Trivelli, Magnasco, Nocco, Klersy, Muscianisi, Ghiggeri, Cardillo, Verrina, Nocera and Ginevri.)

The role of therapeutic drug monitoring (TDM) of valaciclovir (VA)/aciclovir (A) and valganciclovir/ganciclovir (VG/G) in critically ill patients is still a matter of debate. More data on the dose-concentration relationship might therefore be useful, especially in pediatrics where clinical practice is not adequately supported by robust PK studies. We developed and validated a new liquid chromatography-tandem mass spectrometry (LC-MS/MS) micro-method to simultaneously quantify A and G from plasma and dried plasma spots (DPS). The method was based on rapid organic extraction from DPS and separation on a reversed-phase C-18 UHPLC column after addition of deuterated internal standards. Accurate analyte quantification using SRM detection was then obtained using a Thermo Fisher Quantiva triple-quadrupole MS coupled to an Ultimate 3000 UHPLC. It was validated following international (EMA) guidelines for bioanalytical method validation and was tested on samples from pediatric patients treated with A, VG, or G for cytomegalovirus infection following solid organ or hematopoietic stem cell transplantation. Concentrations obtained from plasma and DPS were compared using Passing-Bablok and Bland-Altman statistical tests. The assay was linear over wide concentration ranges (0.01-20 mg/L) in both plasma and DPS for A and G, suitable for the expected therapeutic ranges for both Cmin and Cmax, accurate, and reproducible in the absence of matrix effects. The results obtained from plasma and DPS were comparable. Using an LC-MS/MS method allowed us to obtain a very specific, sensitive, and rapid quantification of these antiviral drugs starting from very low volumes (50 μL) of plasma samples and DPS. The stability of analytes for at least 30 days allows for cost-effective shipment and storage at room temperature. Our method is suitable for TDM and could be helpful for improving knowledge on PK/PD targets of antivirals in critically ill pediatric patients.

While de novo donor-specific HLA antibodies (dnDSAs) have a detrimental impact on kidney graft outcome, the clinical significance of de novo non donor-specific antibodies (dnNDSAs) is more controversial. We retrospectively evaluated for Ab development and characteristics of dnNDSAs serially collected post-transplant sera and, when available, graft biopsy eluates, from 144 non-sensitized, primary pediatric kidney recipients, consecutively transplanted at a single center between 2003 and 2017, using HLA class I and class II single-antigen flow-bead assays (SAB). The results were compared with clinical-pathologic data from HLA antibody negative and HLA dnDSA-positive patients. Forty-five out of 144 patients developed dnNDSAs (31%). Among the dnNDSA-positive patients, 86% displayed one or more class I/II antibodies recognizing antigens included in the CREG/shared epitope groups that also comprise the mismatched donor HLA antigens. Despite potential pathogenicity, as suggested by their occasional presence within the graft, dnNDSAs displayed significantly lower MFI, and limited complement binding and graft homing properties, when compared with dnDSAs. In parallel, the graft survival probability was significantly lower in patients with dnDSA than in those with dnNDSA or without HLA antibodies (p < 0.005). Indeed, the dnNDSA-positive patients remaining dnDSA-negative throughout the posttransplant period did not develop clinical antibody mediated rejection and graft loss, and maintained good graft function at a median follow-up of 9 years. The biological characteristics of dnNDSAs may account for the low graft damaging capability when compared to dnDSAs., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Plasma DNA fragmentomics is an emerging area in cell-free DNA diagnostics and research. In murine models, it has been shown that the extracellular DNase, DNASE1L3, plays a role in the fragmentation of plasma DNA. In humans, DNASE1L3 deficiency causes familial monogenic systemic lupus erythematosus with childhood onset and anti-dsDNA reactivity. In this study, we found that human patients with DNASE1L3 disease-associated gene variations showed aberrations in size and a reduction of a "CC" end motif of plasma DNA. Furthermore, we demonstrated that DNA from DNASE1L3-digested cell nuclei showed a median length of 153 bp with CC motif frequencies resembling plasma DNA from healthy individuals. Adeno-associated virus-based transduction of Dnase1l3 into Dnase1l3-deficient mice restored the end motif profiles to those seen in the plasma DNA of wild-type mice. Our findings demonstrate that DNASE1L3 is an important player in the fragmentation of plasma DNA, which appears to act in a cell-extrinsic manner to regulate plasma DNA size and motif frequency., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Systemic lupus erythematosus (SLE) is an autoimmune disease with variable clinical expression. It is a potentially devastating condition affecting mostly women and leading to clinically unpredictable outcomes. Remission and flares may, in fact, alternate over time and a mild involvement limited to few articular sites may be followed by severe and widespread organ damage. SLE is the prototype of any autoimmune condition and has, for this reason, attracted the interest of basic immunologists. Therapies have evolved over time and clinical prognosis has, in parallel, been improved. What clinicians still lack is the possibility to use biomarkers of the disease as predictors of outcome and, in this area, several studies are trying to find solutions. Circulating autoantibodies are clearly a milestone of clinical research and the concrete possibility is to integrate, in the future, classical markers of activation (like C3) with target organ autoantibodies. Anti-dsDNA antibodies represent a basic point in any predictive attempt in SLE and should be considered the benchmark for any innovative proposal in the wide field of target organ pathologies related to SLE. DNA is part of the nucleosome that is the basic unit of chromatin. It consists of DNA wrapped around a histone octamer made of 2 copies each of Histone 2A, 2B, 3, and 4. The nucleosome has a plastic organization that varies over time and has the potential to stimulate the formation of antibodies directed to the whole structure (anti-nucleosome) or its parts (anti-dsDNA and anti-Histones). Here, we present an updated review of the literature on antibodies directed to the nucleosome and the nucleosome constituents, i.e., DNA and Histones. Wetriedto merge the data first published more than twenty years ago with more recent results to create a balanced bridge between old dogma and more recent research that could serve as a stimulus to reconsider mechanisms for SLE. The formation of large networks would provide the chance of studying large cohorts of patients and confirm what already presented in small sample size during the last years.

Current research is focusing on identifying bioclinical parameters for risk stratification of renal allograft loss, largely due to antibody-mediated rejection (AMR). We retrospectively investigated graft outcome predictors in 24 unsensitized pediatric kidney recipients developing HLA de novo donor-specific antibodies (dnDSAs), and treated for late AMR with plasmapheresis + low-dose IVIG + Rituximab or high-dose IVIG + Rituximab. Renal function and DSA properties were assessed before and longitudinally post treatment. The estimated GFR (eGFR) decline after treatment was dependent on a negative % eGFR variation in the year preceding treatment (P = 0.021) but not on eGFR at treatment (P = 0.74). At a median follow-up of 36 months from AMR diagnosis, 10 patients lost their graft. Altered eGFR (P < 0.001) and presence of C3d-binding DSAs (P = 0.005) at treatment, and failure to remove DSAs (P = 0.01) were negatively associated with graft survival in the univariable analysis. Given the relevance of DSA removal for therapeutic success, we analyzed antibody properties dictating resistance to anti-humoral treatment. In the multivariable analysis, C3d-binding ability (P < 0.05), but not C1q-binding, and high mean fluorescence intensity (P < 0.05) were independent factors characterizing DSAs scarcely susceptible to removal. The poor prognosis of late AMR is related to deterioration of graft function prior to treatment and failure to remove C3d binding and/or high-MFI DSAs., (© 2018 Steunstichting ESOT.)

De novo posttransplant donor-specific HLA-antibody ( dn DSA) detection is now recognized as a tool to identify patients at risk for antibody-mediated rejection (AMR) and graft loss. It is still unclear whether the time interval from transplant to DSA occurrence influences graft damage. Utilizing sera collected longitudinally, we evaluated 114 consecutive primary pediatric kidney recipients grafted between 2002 and 2013 for dn DSA occurrence by Luminex platform. dn DSAs occurred in 39 patients at a median time of 24.6 months. In 15 patients, dn DSAs developed within 1 year ( early-onset group), while the other 24 seroconverted after the first posttransplant year ( late-onset group). The two groups were comparable when considering patient- and transplant-related factors, as well as DSA biological properties, including C1q and C3d complement-binding ability. Only recipient age at transplant significantly differed in the two cohorts, with younger patients showing earlier dn DSA development. Late AMR was diagnosed in 47% of the early group and in 58% of the late group. Graft loss occurred in 3/15 (20%) and 4/24 (17%) patients in early- and late-onset groups, respectively ( p = ns). In our pediatric kidney recipients, dn DSAs predict AMR and graft loss irrespective of the time elapsed between transplantation and antibody occurrence., Competing Interests: The authors declare that they have no competing interests.

Alloantibody-mediated graft injury is a major cause of kidney dysfunction and loss. The complement-binding ability of de novo donor-specific antibodies (dnDSAs) has been suggested as a prognostic tool to stratify patients for clinical risk. In this study, we analyzed posttransplant kinetics of complement-fixing dnDSAs and their role in antibody-mediated rejection development and graft loss. A total of 114 pediatric nonsensitized recipients of first kidney allograft were periodically monitored for dnDSAs using flow bead assays, followed by C3d and C1q assay in case of positivity. Overall, 39 patients developed dnDSAs, which were C1q(+) and C3d(+) in 25 and nine patients, respectively. At follow-up, progressive acquisition over time of dnDSA C1q and C3d binding ability, within the same antigenic specificity, was observed, paralleled by an increase in mean fluorescence intensity that correlated with clinical outcome. C3d-fixing dnDSAs were better fit to stratify graft loss risk when the different dnDSA categories were evaluated in combined models because the 10-year graft survival probability was lower in patients with C3d-binding dnDSA than in those without dnDSAs or with C1q(+) /C3d(-) or non-complement-binding dnDSAs (40% vs. 94%, 100%, and 100%, respectively). Based on the kinetics profile, we favor dnDSA removal or modulation at first confirmed positivity, with treatment intensification guided by dnDSA biological characteristics., (© Copyright 2016 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Background: Development of de novo donor-specific antibodies (dnDSA) is associated with late or chronic antibody-mediated rejection (CAMR) and poor graft outcome in low-risk kidney transplant recipients. High-level soluble B-cell activating factor (sBAFF) was observed in kidney recipients at higher risk of developing dnDSA., Methods: We longitudinally analyzed sBAFF levels in 81 consecutive primary pediatric kidney recipients monitored for de novo human leukocyte antigen (HLA) antibody (Ab) occurrence to gain insight into the events conditioning B-cell activation posttransplant and to analyze the usefulness of paired DSA-sBAFF monitoring in this clinical setting., Results: At a median follow-up of 65 months, 23 patients (28%) developed dnDSA, with 13 of 23 developing CAMR. Irrespective of HLA Ab status, sBAFF levels progressively increased in all patients in the first posttransplant year, thereafter reaching a plateau. sBAFF levels were influenced by the degree of HLA class I antigen match and donor age. Despite higher levels of sBAFF in HLA Ab-positive patients (median and 95% confidence interval sBAFF in DSA+non-DSA patients: 568, 534-608 pg/mL vs. 502, 422-548 pg/mL in Ab-negative patients; P<0.05), we found that sBAFF monitoring could not predict DSA development by a time to event longitudinal analysis. Moreover, sBAFF kinetics up to CAMR onset could not anticipate CAMR development in the DSA cohort., Conclusion: Our findings provide evidence of early posttransplant B-cell activation even in unsensitized recipients of first kidney allograft. The significance of this activation, likely induced by exposition to the allograft, is yet unclear.

Data on the different HLA-antibody (Ab) categories in pediatric kidney recipients developing de novo donor-specific Abs (DSA) after transplantation are scarce. We retrospectively evaluated 82 consecutive nonsensitized pediatric recipients of a first kidney graft for de novo HLA Ab occurrence and antigen specificity. At a median follow-up of 6 years, 29% of patients developed de novo DSA, while 45% had de novo non-DSA. DSA appeared at 25-month median time post-transplant and were mostly directed toward HLA-DQ antigens. Considering each HLA antigen, the estimated rate of DQ DSA (7.55 per 100 person-years) was much higher than the rates observed for non-DQ DSA. The HLA-DQ Ab recognized determinants of the DQβ chain in 70% of cases, α chain in 25% of cases, and both chains in one patient. Non-DSA peaked earlier than DSA, and were largely directed against HLA class I specificities that belonged to HLA-A- and HLA-B-related cross-reacting epitope groups (CREG) in 56% of cases. Our results indicate a need for evaluating HLA-DQ compatibilities in kidney allocation, in order to minimize post-transplant development of de novo DSA, known to be responsible for antibody-mediated rejection and graft loss., (© 2014 Steunstichting ESOT.)

In children with idiopathic nephrotic syndrome, rituximab can maintain short-term remission with withdrawal of prednisone and calcineurin inhibitors. Long-term effects including the number of repeated infusions to maintain remission are unknown. To test this, we treated 46 consecutive children with idiopathic nephrotic syndrome lasting for at least 1 year (mean 6.3 years), maintained in remission with oral prednisone and calcineurin inhibitors. They received 1-5 rituximab courses during a median follow-up of 3 years. Oral agents were tapered after each infusion, and completely withdrawn within 45 days. Rituximab was well tolerated. Six-month probabilities of remission were 48% after the first infusion and 37% after subsequent infusions. One- and 2-year-remission probabilities were, respectively, 20 and 10%. Median time intervals between complete oral-agent withdrawal and relapse were 5.6 and 8.5 months, respectively, following the first and subsequent courses. The time to reconstitution of CD20 cells correlated with the duration of remission, but was not associated with variation in FcyR, CD20, or SMPDL-3B polymorphisms. Podocyte Src phosphorylation was normal. Thus, rituximab can be safely and repeatedly used as a prednisone and calcineurin inhibitor-sparing therapy in a considerable proportion of children with dependent forms of idiopathic nephrotic syndrome. Further study is needed to identify patients who will benefit most from rituximab therapy.

The emerging role of humoral immunity in the pathogenesis of chronic allograft damage has prompted research aimed at assessing the role of anti-HLA antibody (Ab) monitoring as a tool to predict allograft outcome. Data on the natural history of allografts in children developing de novo Ab after transplantation are limited. Utilizing sera collected pretransplant, and serially posttransplant, we retrospectively evaluated 82 consecutive primary pediatric kidney recipients, without pretransplant donor-specific antibodies (DSA), for de novo Ab occurrence, and compared results with clinical-pathologic data. At 4.3-year follow up, 19 patients (23%) developed de novo DSA whereas 24 had de novo non-DSA (NDSA, 29%). DSA appeared at a median time of 24 months after transplantation and were mostly directed to HLA-DQ antigens. Among the 82 patients, eight developed late/chronic active C4d+ antibody-mediated rejection (AMR), and four C4d-negative AMR. Late AMR correlated with DSA (p < 0.01), whose development preceded AMR by 1-year median time. Patients with DSA had a median serum creatinine of 1.44 mg/dL at follow up, significantly higher than NDSA and Ab-negative patients (p < 0.005). In our pediatric cohort, DSA identify patients at risk of renal dysfunction, AMR and graft loss; treatment started at Ab emergence might prevent AMR occurrence and/or progression to graft failure., (© Copyright 2012 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Idiopathic nephrotic syndrome resistant to standard treatments remains a therapeutic dilemma in pediatric nephrology. To test whether the anti-CD20 monoclonal antibody rituximab may benefit these patients, we conducted an open-label, randomized, controlled trial in 31 children with idiopathic nephrotic syndrome unresponsive to the combination of calcineurin inhibitors and prednisone. All children continued prednisone and calcineurin inhibitors at the doses prescribed before enrollment, and one treatment group received two doses of rituximab (375 mg/m(2) intravenously) as add-on therapy. The mean age was 8 years (range, 2-16 years). Rituximab did not reduce proteinuria at 3 months (change, -12% [95% confidence interval, -73% to 110%]; P=0.77 in analysis of covariance model adjusted for baseline proteinuria). Additional adjustment for previous remission and interaction terms (treatment by baseline proteinuria and treatment by previous remission) did not change the results. In conclusion, these data do not support the addition of rituximab to prednisone and calcineurin inhibitors in children with resistant idiopathic nephrotic syndrome.

Background and Objectives: Prednisone and calcineurin inhibitors are the mainstay therapy of idiopathic nephrotic syndrome (INS) in children. However, drug dependence and toxicity associated with protracted use are common. Case series suggest that the anti-CD20 monoclonal antibody rituximab (RTX) may maintain disease remission., Design, Setting, Participants, & Measurements: This open-label randomized controlled trial was powered to show that a strategy based on RTX and lower doses of prednisone and calcineurin inhibitors was noninferior to standard doses of these agents in maintaining 3-month proteinuria as low as baseline or up to 1 g/d greater (noninferiority margin). Participants were stratified by the presence of toxicity to prednisone/calcineurin inhibitors and centrally assigned to add RTX (Mabthera, 375 mg/m(2) intravenously) to lower doses of standard agents or to continue with current therapy alone. The risk of relapse was a secondary outcome., Results: Fifty-four children (mean age 11 ± 4 years) with INS dependent on prednisone and calcineurin inhibitors for >12 months were randomized. Three-month proteinuria was 70% lower in the RTX arm (95% confidence interval 35% to 86%) as compared with standard therapy arm (intention-to-treat); relapse rates were 18.5% (intervention) and 48.1% (standard arm) (P = 0.029). Probabilities of being drug-free at 3 months were 62.9% and 3.7%, respectively (P < 0.001); 50% of RTX cases were in stable remission without drugs after 9 months., Conclusions: Rituximab and lower doses of prednisone and calcineurin inhibitors are noninferior to standard therapy in maintaining short-term remission in children with INS dependent on both drugs and allow their temporary withdrawal.

The mechanism responsible for proteinuria in non-genetic idiopathic nephrotic syndrome (iNS) is unknown. Animal models suggest an effect of free radicals on podocytes, and indirect evidence in humans confirm this implication. We determined the oxidative burst by blood CD15+ polymorphonucleates (PMN) utilizing the 5-(and-6)-carboxy-2',7'-dichlorofluorescin diacetate (DCF-DA) fluorescence assay in 38 children with iNS. Results were compared with PMN from normal subjects and patients with renal pathologies considered traditionally to be models of oxidative stress [six anti-neutrophil cytoplasmic autoantibody (ANCA) vasculitis, seven post-infectious glomerulonephritis]. Radicals of oxygen (ROS) production was finally determined in a patient with immunodeficiency, polyendocrinopathy, enteropathy X-linked (IPEX) and in seven iNS children after treatment with Rituximab. Results demonstrated a 10-fold increase of ROS production by resting PMN in iNS compared to normal PMN. When PMN were separated from other cells, ROS increased significantly in all conditions while a near-normal production was restored by adding autologous cells and/or supernatants in controls, vasculitis and post-infectious glomerulonephritis but not in iNS. Results indicated that the oxidative burst was regulated by soluble factors and that this regulatory circuit was altered in iNS. PMN obtained from a child with IPEX produced 100 times more ROS during exacerbation of clinical symptoms and restored to a near normal-level in remission. Rituximab decreased ROS production by 60%. In conclusion, our study shows that oxidant production is increased in iNS for an imbalance between PMN and other blood cells. Regulatory T cells (Tregs) and CD20 are probably involved in this regulation. Overall, our observations reinforce the concept that oxidants deriving from PMN are implicated in iNS.

Objectives: Little is reported in literature regarding correct management of benign lesions of the kidney. The aim of our study is to present a series of total and partial nephrectomies performed in the last 5 years and to discuss indications., Materials and Methods: Patients with benign lesions who underwent nephrectomy and partial nephrectomy at our institution in the period 2003-2008 were retrospectively included in the study. Notes were carefully reviewed and demographic data, symptoms onset, preoperative diagnosis, investigations, medical and/or surgical treatment, postoperative complications and definitive histological reports were collected., Results: Forty procedures were performed. Twelve patients were preoperatively diagnosed of having multicystic dysplastic kidney (MCDK), which was confirmed in 10, whereas the remaining 28 patients of having severe dysplasia or hypodysplasia. Thirty-four patients underwent total nephrectomy, six underwent partial nephrectomy. Histopathological analysis confirmed segmental or complete abnormalities of the involved kidney in all cases. No malignancies were detected., Discussion: Our study confirmed the extremely low malignancy rate of MCDK and hypodysplastic kidneys. The 20% mismatch of pre- and post-operative diagnosis suggests a common aetiology and shared therapeutic strategies for MCDK and hypodysplasia. At present, there is no consensus regarding correct indications for nephrectomy in paediatric age. As nephrectomy seems not to provide any advantage over preservation, but surgical and anesthesiological risks, we should be prudent in preserving every asymptomatic poorly or non-functioning kidneys maintaining a strict follow-up. Randomised controlled studies on larger multicentric series are strongly warranted to define this topic.

Glomerular targets of autoimmunity in human membranous nephropathy are poorly understood. Here, we used a combined proteomic approach to identify specific antibodies against podocyte proteins in both serum and glomeruli of patients with membranous nephropathy (MN). We detected specific anti-aldose reductase (AR) and anti-manganese superoxide dismutase (SOD2) IgG(4) in sera of patients with MN. We also eluted high titers of anti-AR and anti-SOD2 IgG(4) from microdissected glomeruli of three biopsies of MN kidneys but not from biopsies of other glomerulonephritides characterized by IgG deposition (five lupus nephritis and two membranoproliferative glomerulonephritis). We identified both antigens in MN biopsies but not in other renal pathologies or normal kidney. Confocal and immunoelectron microscopy (IEM) showed co-localization of anti-AR and anti-SOD2 with IgG(4) and C5b-9 in electron-dense podocyte immune deposits. Preliminary in vitro experiments showed an increase of SOD2 expression on podocyte plasma membrane after treatment with hydrogen peroxide. In conclusion, our data support AR and SOD2 as renal antigens of human MN and suggest that oxidative stress may drive glomerular SOD2 expression.

Cyclosporine (CsA) has considerably modified the graft survival in solid organ and bone marrow transplantations. It is also the treatment of choice in chronic diseases such as steroid resistance and/or dependence nephrotic syndrome and autoimmune-diseases, especially in those cases that require long term treatments. Renal toxicity is the major adverse effect of chronic CsA administration. Deterioration of renal function and renal histopathology are the basic elements of the diagnosis. Overall, available studies suggest a good degree of safety related to appropriate drug dosages even if they require an adequate degree of surveillance in case of rapid changes of renal functions and long term evaluation of renal pathology. CsA neurotoxicity is the second major problem that seems underestimated especially in case of subtle manifestations in children. The full blown picture of the acute form is characterized by convulsion and sudden alteration of mental function that are reversible upon drug withdrawal. The diagnosis is based on typical CT and MRI aspects of extensive bilateral white-matter abnormalities in the occipital region of the brain that mimics the posterior encephalopathy syndrome. Prospective evaluations of drug tolerance include renal histology in case of chronic renal toxicity and neuro-imaging to identify and block acute neurotoxicity.

Mesenchymal stem cells (MSCs) may be of value in regeneration of renal tissue after damage; however, lack of biological knowledge and variability of results in animal models limit their utilization. We studied the effects of MSCs on podocytes in vitro and in vivo utilizing adriamycin (ADR) as a model of renal toxicity. The in vivo experimental approach was carried out in male Sprague-Dawley rats (overall 60 animals) treated with different ADR schemes to induce acute and chronic nephrosis. MSCs were given a) concomitantly to ADR in tail vein or b) in aorta and c) in tail vein 60 days after ADR. Homing was assessed with PKH26-MSCs. MSCs rescued podocytes from apoptosis induced by ADR in vitro. The maximal effect (80% rescue) was obtained with MSCs/podocytes coculture ratio of 1:1 for 72 h. All rats treated with ADR developed nephrosis. MSCs did not modify the clinical parameters (i.e., proteinuria, serum creatinine, lipids) but protected the kidney from severe glomerulosclerosis when given concomitantly to ADR. Rats given MSCs 60 days after ADR developed the same severe renal damage. Only a few MSCs were found in renal tubule-interstitial areas 1-24 h after injection and no MSCs were detected in glomeruli. MSCs reduced apoptosis of podocytes treated with ADR in vitro. Early and repeated MSCs infusion blunted glomerular damage in chronic ADR-induced nephropathy. MSCs did not modify proteinuria and progression to renal failure, which implies lack of regenerative potential in this model.

Background: Glucose infusion test (GIT) is a new method to measure vascular access recirculation (R) based on basal glucose increase in the arterial blood line after a 20% glucose bolus (5 ml) into the venous chamber., Methods: We compared GIT with the ultrasound dilution method (HD01, Transonic Systems Inc., USA) in a circuit reproducing in vitro the phenomenon of R. We repeated the comparison in 162 chronic haemodialysis patients (133 fistulae, 17 central venous catheters, 12 prosthetic grafts)., Results: In vitro, we determined the timing for C2 sampling: QB 200 ml/min, C2 16-20 s; QB 300 ml/min, C2 13-17 s; QB 400 ml/min, C2 9-12 s. GIT showed no false positives nor false negatives (100% specificity and sensitivity) while HD01 did not recognize three cases with R=5% (91% sensitivity) and it yielded no false positive (100% specificity). The Bland-Altman analysis showed a bias of 0.2+/-1.3% and 1.3+/-2.9% for GIT and HD01, respectively. In vivo, only 16 out of 162 patients were found positive with both methods (GIT 13.5+/-13%; HD01 16.3+/-15%; P=NS) while three patients with minimal R (GIT 3.2%) were not recognized by HD01 although a low R peak was clearly evident and repeatable on the laptop plot. The Bland-Altman analysis showed an overall bias of 0.2+/-1.7% to the limits of agreement=-3.1 and 3.6% (n=162) and no correlation between the difference and the mean of positive tests. The pooled coefficient of variation of positive cases was 13.3 and 18.1% for GIT and HD01, respectively., Discussion: Our in vitro study showed a good performance of GIT and its better sensitivity compared to HD01. These results were confirmed in vivo with only 3/162 discordant results due to a low R under the HD01 limit of detection (R=5%). In conclusion, the GIT proved to be a very accurate screening test for R, with a very low threshold of detection. In addition, it is simple, user-friendly and inexpensive.

Background: The problem of pure red cell aplasia (PRCA) prompted nephrologists to revert to a wider intravenous (i.v.) utilization of erythropoeitin (Epo). Once weekly i.v. Epo administration has been suggested to be as effective as the twice/thrice weekly i.v. dose. The aim of the present study was to test whether once weekly i.v. Epo administration is equally as cost-effective as once weekly subcutaneous (s.c.) and 2-3 times weekly i.v. administration., Methods: We prospectively studied 41 patients (23 males, aged 28-82 years), on renal replacement therapy for 18-286 months, stabilized on twice or thrice weekly s.c. Epo-alpha (basal). The patients were treated for three consecutive 6 month periods with once weekly s.c. (OWSC), once weekly i.v. (OWIV) and twice/thrice weekly i.v. (TWIV) Epo-alpha. The initial dose for each period was equal to the final dose of the previous one; when necessary, the dose was adjusted according to DOQY guidelines. Iron, folic acid and vitamin B(12) supplementations were given throughout all the study periods. At the end of each of the four study periods, the following parameters were evaluated: haemoglobin, haematocrit, hypochromic red blood cells (RBCs), iron, serum ferritin, transferrin, folate, vitamin B(12), C-reactive protein (CRP), Kt/V, parathyroid hormone (PTH) and weekly dose of Epo-alpha., Results: Thirty-three out of 41 enrolled patients completed the study (there were five deaths, two renal transplants and one transfer). No significant changes were observed as regards iron, serum ferritin, transferrin, folate, vitamin B(12), CRP, Kt/V or PTH level. Haemoglobin levels were not different at the end of the basal (11.7+/-1.21), OWSC (11.8+/-0.86) and TWIV (12.1+/-1.04) periods, while significantly lower levels were observed after the OWIV period (11.0+/-0.97, P<0.01). Weekly Epo consumption (Epo U/week/kg body weight/g haemoglobin) was: basal 11.57+/-5.96; OWSC 10.22+/-4.53; OWIV 15.99+/-7.7*(a); and TWIV 11.89+/-6.3*(a) (*P<0.01 vs basal; (a)P<0.01 vs OWSC)., Conclusions: From our results, the OWIV schedule seems to have less efficacy in the control of anaemia of chronic renal failure patients on dialysis treatment than either OWSC or TWIV schedules.

During hemodialysis access surveillance, referral for evaluation and correction of stenosis is based upon determination that a significant decrease in blood flow (Q) has occurred. However, criteria for determining when a decrease is statistically significant have not yet been established. In this study we established such criteria by analyzing Q variation with the glucose pump test (GPT). We took nine Q measurements in each of 25 patients (18 grafts, 7 fistulas) during three dialysis sessions within a 2-week period (predialysis and during hours 1 and 3). We determined thresholds that define a significant percentage decrease in Q (deltaQ) for various p values. In order to confirm the general applicability of these thresholds, we computed the average within-patient Q variation during the three sessions (computed as a coefficient of variation and referred to as short-term variation). We then determined the relative influences of biological (true) variation and analytical error on short-term variation. We found that deltaQ must be > 33% to be significant at p < 0.05, whereas the threshold is > 17% for p < 0.20. Measuring Q at uniform versus different times during the sessions did not significantly reduce these thresholds. We also found that biological variation was nearly as large as short-term Q variation, whereas analytical error contributed minimally to short-term variation. In conclusion, this study defines thresholds for a significant deltaQ that have wide application in determining access referral for evaluation and correction of stenosis. Selection of a particular threshold should consider the relative importance of avoiding thrombosis versus avoiding unnecessary procedures. If avoiding unnecessary procedures is a priority, then we recommend a threshold of > 33%. These thresholds apply to other methods of measuring Q, provided analytical error is significantly less than biological variation.

Vascular access recirculation (R) is a well-known cause of decreased dialysis dose. In this paper a new easy protocol for R detection in pre-dialysis derived from the classic Glucose Infusion Test (GIT) is introduced. The pre-dialysis GIT (GIT-pre) is based on the glucose (5%, 10 ml) bolus injection directly into the venous needle and on a simultaneous withdrawal from the arterial needle. If the glucose value increases during the glucose bolus, R is present. This new protocol was validated on 29 chronic haemodialysis patients (20 AVFs, 7 CVCs, 2 PTFE grafts), comparing the glucose increase with the classic GIT during dialysis. Only one CVC had R with the blood lines in the normal position (deltaglu = 465 mg/dl with GIT-pre and a deltaglu = 186 mg/dl, R = 9.3% with classic GIT) while in the reverse blood line position, all CVCs showed a significant glucose increase (mean GIT-pre deltaglu = 195 mg/dl; mean GIT deltaglu = 140 mg/dl corresponding to a R = 8%). There were 5 AVFs with true R (correct blood lines position) clearly identified by both methods (mean values deltaglu = 316 mg/dl with GIT-pre and a deltaglu = 390 mg/dl, R = 19.5% with classic GIT). Preliminary results show good reliability of the new protocol in identifying VA R caused either by failing VA with stenosis or by reverse blood lines position. The GIT-pre is a simpler application of the classic GIT useful for testing new VA, new needle positions or CVC performance before starting dialysis. A simpler R test could increase the frequency of the measurements and consequently the power of R in early detection of VA problems.

Vascular access recirculation (R) allows the evaluation of the adequacy of the extracorporeal blood circuit in dialysis patients. The test verifies the correct needle position in patients with arterovenous fistulae (AVF) and the effective function of central venous catheters. In clinically uncomplicated native fistulae, a normal R test could avoid more complex procedures like blood flow measure or angiography. The AVF recirculation has two components, vascular access recirculation (AR) and cardiopulmonary recirculation (CPR). While the first phenomenon is well known, the second remained undetected for many years resulting in wrong R calculations with false positives. Using the correct formula, the great majority of AVF resulted in zero recirculation. The presence of R reduces the dialysis efficiency to critical levels, mainly in unsuspected cases. Among the numerous available R tests, the urea test is the oldest and historically the most commonly used method, but unfortunately it is labor intensive, with low sensitivity and specificity and with delayed results. The "ultrasound dilution"method is considered the gold standard, easy to perform, with good repeatability, but it is expensive requiring a specific device. Finally, the glucose infusion test (GIT) is a new low-cost test with immediate results and a very low detection limit, with good repeatability and high specificity and sensitivity.

Background: Blood flow (Qa) measurements are an important step in the surveillance protocol of haemodialysis vascular access (VA). The glucose pump test (GPT) is a new test for Qa measurement based on the dilution of a constant glucose infusion. The aim of this study is to verify the clinical accuracy of GPT in a graft surveillance protocol with sequential Qa measurements., Methods: In 30 chronic haemodialysis patients with graft, we compared monthly sequential Qa measurements performed with GPT in pre-dialysis and the ultrasound dilution technique (HD01 device Transonic Systems Inc., USA) during dialysis. The colour Doppler ultrasonography study (CDU) was our reference standard for the diagnosis of stenosis. The endpoints were the graft thrombosis or PTA treatment., Results: According to the K/DOQI guidelines we could identify the thrombosis high-risk grafts when Qa was <600 ml/min or <1000 ml/min with a decrease >25% in serial Qa measurements. HD01 yielded 27 of 112 high-risk Qa measurements (21 Qa <600 ml/min; mean 406+/-145 ml/min; 6 deltaQa >25%; mean 43+/-7%). In 12 of 27 cases the CDU control did not show haemodynamically significant stenoses (false positive); 15 of 27 cases were confirmed high-risk accesses by CDU and did PTAs (HD01 specificity 86%). GPT yielded 14 of 112 high-risk Qa measurements (8 Qa <600 ml/min; mean 404+/-135 ml/min; 6 deltaQa >25%; mean 38+/-8%) and all had severe stenoses and underwent PTA treatments showing a GPT specificity of 100%. The CDU study allowed us to correctly assess the Qa negative cases. HD01 method had 10 false negative cases (treated or clotted grafts with a Qa >600 ml/min and deltaQa <25%) with a sensitivity of 60%, while GPT had 11 false negative cases with a sensitivity of 56%. The diagnostic accuracy tested with the ROC curves was similar with both tests (area under the curve was 0.762 and 0.752 with GPT and ultrasound dilution, respectively; P = 0.985). The diagnostic efficiency (percentage of grafts with agreement between test result and factual situation) was 90 and 80% (P = 0.056) for GPT and HD01, respectively., Conclusion: Compared with HD01, the GPT had a lower false positive rate and similar diagnostic accuracy and efficiency. The clinical implication is a smaller number of unnecessary, invasive procedures (angiographies or PTAs), without increasing the thrombosis risk. This study has shown that GPT is an accurate, quick and economic test for Qa monitoring.

Background: The Glucose Pump Test (GPT) is a new method for measuring the access blood flow (Qa) based on a constant glucose infusion (Qi), with known glucose concentration (Ci), in the arterial needle and on glucose determination in two blood samples from the venous needle, the first (C1) in basal conditions, the second (C2) during the infusion. Qa depends on the difference in glucose between the two samples and is computed from the formula: QaGPT = Qi x (Ci-C2)/(C2-C1)., Methods: The new method, previously evaluated by ultrasound dilution and color-Doppler techniques, was compared with the Urea Test (UT) in 20 patients measuring recirculation (R) during reversal of the arterial and venous needles (QaUT = Qb x (1/R -1)). All Qa determinations were done twice by both methods. Glucose and urea were determined respectively two and three times., Results: Mean QaGPT = 841, SD 347 ml/min, mean QaUT = 872, SD 417 ml/min (p = n.s.); mean percent difference QaGPT-QaUT= 16%, SD 14; mean coefficients of variation of paired determinations: 8.1% and 12.1% respectively; Pearson coefficient between the two methods: r= 0.91., Conclusions: The comparison showed a good correlation between the two methods and similar mean values. The coefficient of variation of the new method was acceptable and lower than with the UT. The GPT is a reliable technique for measuring blood flow in vascular accesses.

Background: The glucose pump test (GPT) is a recently introduced method of measuring hemodialysis access blood flow (Qa). A validation of GPT during dialysis has not yet been done, and performance characteristics of the method have not yet been fully analyzed., Methods: The authors studied 33 patients (25 synthetic grafts, 8 autogenous arteriovenous fistulae). Qa measurements by ultrasound dilution (UD) and GPT were done in triplicate during dialysis. In GPT, a baseline blood sample (C(1)) was obtained, followed by infusion of a 10% glucose solution (C(i)) through the arterial needle into the access at 16 mL/min (Q(i)). After 11 seconds, a downstream blood sample (C(2)) was aspirated from the venous needle. C(1) and C(2) glucose were measured by glucometer. Qa was computed by the equation: Qa = Q(i)(C(i) - C(2))/(C(2) - C(1)). A model of the access vascular circuit was used to determine the influence of C(2) aspiration on the Qa measurement., Results: Mean Qa was 1413 mL/min by UD versus 1,496 mL/min by GPT (P = 0.11). There was a strong linear correlation between the 2 methods (r = 0.905; P <0.001). The pooled coefficient of variation was 6.4% for UD and 9.6% for GPT. The circuit model showed that aspiration of C(2) causes an increase in Qa (DeltaQa) that depends on the aspiration rate (Q(ASP)) and fraction of resistance in the circuit that is downstream to the venous needle: DeltaQa = Q(ASP)(Downstream resistance)/(Total resistance). The model predicts the overestimate is approximately 62 mL/min for grafts and 120 mL/min for fistulae but may vary depending on the balance of resistances upstream and downstream to the venous needle., Conclusion: This study shows that GPT closely correlates with UD, and the method has adequate precision. GPT is an inexpensive method that may help make Qa measurements more widely available than previously possible.

Background: A good test for monitoring blood flow (Q(a)) must be accurate, rapid and economical in order to allow frequent easy measurements. The glucose pump test (GPT) is based on a constant glucose infusion as a dilutional indicator of Q(a)., Methods: GPT protocol requires a constant glucose infusion, by a syringe pump, into the arterial needle and two blood withdrawals from the venous needle, one basal before the infusion (C(a1)), the other (C(a2)) 11 s after the start of the infusion. At the bedside we measure glucose on C(a1) and C(a2). Knowing the infused glucose concentration (C(i)) and the pump infusion rate (Q(i)) we can easily calculate Q(a)=Q(i)x(C(i)-C(a2))/(C(a2)-C(a1)). We verified the accuracy of this new method by comparing it with the in vitro results from a circuit reproducing vascular access circulation, and in vivo comparing GPT-Q(a) with Doppler ultrasound in pre-dialysis to the Transonic HD01-Q(a) during dialysis in 23 chronic haemodialysis patients., Results: GPT-Q(a) values were highly correlated with the in vitro Q(a)=1.01 x GPT-Q(a)-16.6; r=0.94. There was agreement between the mean flow values of GPT and Doppler (927.5 and 927.1 ml/min, respectively; P=NS) while the mean value of HD01 was significantly lower (HD01-Q(a)=690 ml/min; P<0.001 vs GPT-Q(a) and Doppler-Q(a)). The regression analysis showed a good correlation between GPT and Transonic results (r=0.95; HD01-Q(a)=0.86 x GPT-Q(a)-111.9), while there was a significant difference between the two measurements (mean Delta 235+/-117 ml/min; range from 15 to 451 ml/min). This difference could be caused by the large haemodynamic variations (different blood pressure, cardiac output, circulating effective volume, haematocrit) between pre-dialysis and intra-dialysis and in addition by the counter current flow during the reversal blood lines Transonic measurements., Conclusions: GPT offers the advantage of a simple bedside procedure easily performed before dialysis: it does not interfere with the dialysis treatment and it is less intrusive for the patient as it does not involve reversal of the blood lines. The preliminary data indicate that our method could be a useful, simple and cheap test for monitoring access flow in every dialysis unit.

Introduction. Vascular access recirculation (AR), which is often unacknowledged, remains an important cause of inadequate dialytic dose. The glucose infusion test (GIT) is a new method for detecting and quantifying AR. This paper reports on a polycentric evaluation of the new test and a comparison with the classical Urea-test (UT). Methods. GIT protocol comprises withdrawal from the arterial port (sample A), injection into the venous drip chamber of 1 g glucose in 4 seconds, withdrawal from the arterial port (sample B) continuously from 13 to 17 seconds. Glucose is determined on A and B by a reflectance photometer. If B = A then there is no recirculation. If B exceeds A by at least 20 mg/dl there is recirculation. AR quantification: AR% = (B-A) / 20. GIT was performed on 623 patients from eleven dialysis centers to screen the patients for AR. Subsequently, GIT and Urea-test (UT) were compared in 189 paired tests. The reproducibility of GIT and UT was studied in 28 paired tests performed in sequence. Results. The screening test by GIT was positive in 68 cases (11 %). The majority of positivities was found in central venous catheters (CVC, 27/50 cases, 54 %), whereas only 7 % of artero-venous fistulas (AVF) were positive. In the CVC group, Tesio catheters were more frequently positive compared to Dual Lumen Catheters (64 % vs. 29 %). The comparison GIT - UT showed that results matched in 162 tests (79 negative and 83 positive both by GIT and UT), showing that on the grounds of UT, GIT has high sensitivity and specificity. In 27 tests GIT was positive, but UT negative. This disagreement is due to the different minimal limit of detection, 1 % for GIT and 5% for UT. The reproducibility was greater with GIT than with UT with a lower D% (respectively -0.6 +/- 2.5 and -0.4 +/- 6.1 %, p<0.001) and a lower coefficient of variation (17 vs 33 %). Conclusions. The screening of 623 patients by GIT confirmed that AR in AVF is normally absent, whereas an un-expectedly high frequency of moderate AR in CVC was found. The GIT-UT comparison showed that the new test is simple and immediate, and gives results with higher accuracy, sensitivity and reproducibility than UT.

Background: Vascular access recirculation is an important cause of diminished dialysis efficiency. We propose a new screening test based on glucose infusion as a tracer for recirculation., Methods: The glucose infusion test (GIT) protocol comprises a basal blood sample (A) from the arterial port, a 5 mL bolus of 20% glucose into the venous chamber (time 0), followed by a second sample (B) in four seconds (from 13 to 17 s with QB 300 mL/min) from the same port. The blood glucose level is determined at the bedside on A and B with a reflectance photometer (CV 1.8%). Interpretation of the test is straightforward: If B = A, there is no recirculation, whereas if B > A, recirculation can be calculated from the regression equation: 0.046 x (B - A) + 0.07, obtained from in vitro tests reproducing artificial recirculation at 0, 5, and 10%. To validate this new method in vivo, we compared GIT and the urea test on 39 hemodialysis patients, obtaining a good correlation (r = 0.93). The two tests were considered positive (recirculation present) when the lower 95% confidence intervals were more than zero., Results: Our patients were divided into two groups: those with (22 out of 39, mean recirculation 11.8%) or without recirculation (17 out of 39, mean 0.06%). The urea test did not recognize 7 out of 22 patients because they had a small recirculation below the urea test limit of detection., Conclusions: GIT was more sensitive (detection limit 0.3%), simpler, and immediate in showing the results than the urea test. It is an accurate and low-cost technique for screening and follow-up of vascular access in a dialysis unit.

Background: Intraperitoneal spillage of dermoid cyst content, if not followed immediately by abundant peritoneal lavage, can cause a chemical peritonitis with subsequent adhesion formation., Study Design: We performed an open clinical study in a university hospital. Forty-four consecutive ovarian dermoid cysts were removed intact from 40 premenopausal women operated on between October 1993 and December 1997. The laparoscopic technique included: 1) creation of a cleavage plane between the cyst and the ovary; 2) dissection of the cyst by a combination of water, scissors, and gravity without direct traction on the cyst; and 3) extraction of the cyst after its placement inside a laparoscopic bag., Results: The mean cyst diameter was 6.5 cm (range 3 to 12 cm). Mean operating time was 125 minutes (range 50 to 180 minutes). All patients were discharged within 48 hours. The cysts were dissected completely intact and were extracted without spillage in the abdominal cavity in all cases. Operative followup was available in 15 of the 40 patients; mild adhesions were found on the treated ovary in 3 (20%)., Conclusions: It is always possible to prevent rupture and spillage of dermoid cysts during laparoscopic operations, but this approach is time consuming and needs expert surgical technique.

This study aimed to standardize the clomiphene citrate test (CC-t) in our laboratory while comparing two different protocols of controlled ovarian stimulation in poor responders. One hundred and forty-four patients scheduled for assisted reproductive techniques were submitted to the CC-t within 3 months before starting stimulation; 133 underwent controlled ovarian stimulation with a blocking protocol. Poor responders in the first cycle (n = 30) were subsequently treated with a flare-up protocol. Although it was not statistically significant, more patients reached oocyte retrieval with the flare-up protocol. In the completed cycles, more gonadotropin ampules (55 +/- 15 vs. 34 +/- 13; p < 0.001) and more stimulation days (12.6 +/- 1 vs. 11.6 +/- 1.2; p < 0.005) were needed in the blocking than in the flare-up protocol. No difference was observed in peak 17 beta-estradiol levels, preovulatory follicles, oocytes retrieved or pregnancy rate between the two protocols. According to the threshold values, established on CC-t of patients who obtained a clinical pregnancy (n = 44), the incidence of abnormal results was 10%. All but one patient with abnormal CC-t were poor responders during the first stimulation cycle. The flare-up protocol did not improve the ovarian response in these patients.

A previous report suggests that treatment with recombinant human erythropoietin (rH-EPO) significantly improves many abnormalities in circulating amino acids (AA) in hemodialysis patients. We evaluated the effects of a 12-month treatment with rH-EPO (150-250 U/kg/week) on blood AA levels in 10 patients with chronic renal failure under regular dialytic treatment. During treatment, hemoglobin levels increased from 7.0 +/- 0.3 to 10.1 +/- 0.3 g/dl at 3 months remaining steady thereafter. Before the treatment, patients showed reduced levels of essential AA (EAA), mainly valine, leucine and threonine (p < 0.05-0.01); among non-EAA (NEAA), aspartate and serine were reduced, whereas glycine, alanine, proline, citrulline and cyst(e)ine were increased (p < 0.05-0.001). Val/Gly, Ser/Gly and Tyr/Phe ratios were low (p < 0.05-0.01). Total EAA and total NEAA (619 +/- 21 and 1,382 +/- 75 mumol/l, respectively, before the study) were unchanged (639 +/- 22 and 1,410 +/- 89 mumol/l, respectively) at 12 months. Abnormalities in AA levels observed before the treatment persisted throughout the study. Only serine increased at the end of the study (p < 0.05). In conclusion, contrary to what has been reported, treatment with rH-EPO is not associated with an amelioration of AA metabolism in hemodialysis patients.