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Background/aim: Anterior teeth are prone to traumatic dental injuries (TDIs). Although a number of techniques ranging from original tooth fragment reattachment (TFR) to direct composite restoration (DCR) can be used to restore uncomplicated crown fractures, there is no consensus on which method is best. The purpose of this study was to investigate the fracture resistance of bovine incisors restored by two different techniques (TFR and DCR) in three different fracture models., Materials and Methods: Sixty extracted bovine lower incisors were randomly divided into three groups (n = 20). Angle, oblique, or transverse sections of all the teeth in a group were prepared by using a disk. The cut surfaces were scanned, and the cross-sectional areas (CSA) of the enamel and dentin were measured. Half the teeth in each group were restored by DCR (n = 10) and the other half by TFR (n = 10). The forces required to fracture the restored teeth were then measured using a Universal testing machine, and the fracture modes were analyzed (cohesive, adhesive, or mixed)., Results: No statistically significant differences between the TFR and DCR restorations were detected for total and enamel CSAs in any of the restoration shapes (p > .067). The fracture forces required to break DCR angle and transverse restorations were significantly greater than for the corresponding shapes restored with TFR (p < .033). However, the difference in the forces needed to fracture oblique section restorations by DCR or TFR was not statistically significant (p = .239), despite a similar trend (143.4 ± 51 N and 120.9 ± 25 N, respectively)., Conclusion: This study revealed that a greater force is required to fracture teeth restored by the DCR than by the TFR technique, especially for a transverse section. This demonstrates that restoring a fractured tooth provides a superior outcome compared to reattaching the fractured fragment., (© 2023 The Authors. Dental Traumatology published by John Wiley & Sons Ltd.)

Sibeprenlimab blocks the cytokine "A Proliferation-Inducing Ligand" (APRIL), which may play a key role in immunoglobulin A nephropathy pathogenesis. A phase 1 study of subcutaneous (SC) sibeprenlimab evaluated preliminary safety, tolerability, pharmacokinetics, and pharmacodynamics in healthy participants. This was an open-label, single-ascending-dose study. Twelve participants in each of 4 sequential dosing cohorts received 1 SC dose of sibeprenlimab (200 mg [1×1 mL injection], 400 mg [2×1 mL injections], 400 mg [1×2 mL injection], or 600 mg [1 mL+2 mL injections]) and underwent 16-week follow-up for adverse events, pharmacokinetics, and pharmacodynamics (serum APRIL, immunoglobulin [Ig] levels). Sibeprenlimab in single SC doses of 200-600 mg was slowly absorbed into the systemic circulation, with a median time to maximum serum concentration of approximately 6-10.5 days, and a mean elimination half-life of approximately 8-10 days. Serum APRIL, IgA, IgM, and, to a lesser extent, IgG decreased in a dose-dependent and reversible manner. Maximal reduction in serum IgA was approximately 60% at the 400- and 600-mg doses and 40% at 200 mg. Serum APRIL rapidly decreased to near the lower limit of quantification, and duration of suppression was dose-dependent, with near complete suppression until weeks 4-6 at the 400-mg dose and week 8 at the 600-mg dose. Adverse events occurred in 30/48 (62.5%) participants; none were serious or led to study discontinuation. Sibeprenlimab rapidly and sustainably reduced target APRIL and Ig biomarkers in a dose-dependent and reversible manner, with acceptable preliminary safety and pharmacokinetics., (© 2023 The Authors. Clinical Pharmacology in Drug Development published by Wiley Periodicals LLC on behalf of American College of Clinical Pharmacology.)

Clinicians working on first-in-human clinical studies need to be able to judge whether safety signals observed on an investigational drug were more likely to have occurred by chance or to have been caused by the drug. We retrospectively reviewed 84 Novartis studies including 1,234 healthy volunteers receiving placebo to determine the expected incidence of changes in commonly measured laboratory parameters and vital signs, in the absence of any active agent. We calculated the frequency of random incidence of safety signals, focusing on the liver, cardiovascular system, kidney, and pancreas. Using the liver enzyme alanine aminotransferase (ALT) as an example, we illustrate how a predictive model can be used to determine the probability of a given subject to experience an elevation of ALT above the upper limit of the normal range under placebo, conditional on the characteristics of this subject and the study., (© 2018 The Authors. Clinical and Translational Science published by Wiley Periodicals, Inc. on behalf of American Society for Clinical Pharmacology and Therapeutics.)

Drugs prescribed during pregnancy affect two populations simultaneously: fetuses and their mothers. Drug-induced fetal injury (teratogenicity) has a significant impact on current and future public health. Teratogenic risk designation of many drugs relies on associating rare fetal events with rare environmental exposures. Therefore we aim to develop preclinical predictive models of clinical teratogenicity. We collated public databases for drug-target-gene relationships for 619 drugs spanning the 5 pregnancy risk classes. Genes targeted by high risk but not low risk drugs demonstrated 79% accuracy (p < 0.0001 vs. random) for predicting high vs. low fetal risk on cross validation. Functional enrichment analysis revealed that target genes of drugs known to be safe in pregnancy contained no developmentally related terms, while target genes of known teratogens contained 85 developmentally related terms. Drug target gene signatures that are enriched for known developmental genes may provide valuable preclinical predictive information regarding drug pregnancy risk., (Copyright © 2010 Elsevier Inc. All rights reserved.)

SRL has been increasingly used in renal transplantation, but limited sampling approaches for estimation of AUC remain elusive. A post-hoc analysis of 94 PK profiles in 75 patients from four previous studies was performed to generate limited sampling approaches for approximation of AUC based on two to four time points for both BID and OD SRL dosing. AUC was calculated using the trapezoid rule. Stepwise linear regression was performed to generate an abbreviated AUC from the limited sampling approaches. For BID dosing, complete AUC had a strong correlation with the trough levels (r(2) = 0.882, p < 0.0001) and with C2 level (r(2) = 0.9025, p < 0.0001). A three-point and a four-point limited sampling approach showed improved agreement with complete AUC compared with single-point sampling. A convenient and accurate (r(2) = 0.992) four-point limited sampling approach reads: AUC = 10;(1.085 + 0.117 x log C0 + 0.164 x log C1-0.131 x log C2 + 0.823 x log C4). Similarly, complete AUC had a statistically significant correlation with the trough levels (r(2) = 0.549, p < 0.0001) and with C2 level (r(2) = 0.716, p < 0.0001) for OD dosing. The estimation of AUC for OD dosing was improved over single-point sampling (r(2) = 0.951) using the formula: AUC = 10;(1.100 + 0.115 x log C0 + 0.803 x log C4). This study represented the first limited sampling approach for SRL. Further studies are required to determine the optimal SRL target AUC.

Long-term outcomes for patients with nephrotic syndrome (NS) correlate closely with the degree of steroid responsiveness. There are currently no diagnostic tests that accurately predict steroid responsiveness in NS. In children in particular, a prolonged course of daily, high-dose corticosteroid therapy is as much a diagnostic maneuver as it is a treatment. Urine proteomics has been emerging as a potentially rich source of noninvasive yet informative biomarkers of drug responsiveness in NS. In this review, we discuss some of the initial studies of the nephrotic urinary proteome as well as some ongoing and future challenges, including defining the normal urinary proteome, and extracting valuable urinary protein data from an abundance of urinary albumin.

Children with renal disease have tremendous potential for recovery, particularly when disease processes are detected early in the disease course. However, invasive diagnostic maneuvers can be challenging, especially in younger children who may require general anesthesia. Urine proteomics technologies present an opportunity to discover noninvasive yet informative diagnostic and prognostic markers of renal disease in children. In this article we review current concepts regarding the normal urine proteome, followed by an overview of urine proteomics as applied to nephrotic syndrome, and conclude with a discussion of some of the challenges of performing proteomic profiling on nephrotic urine, with its inherent abundance of proteins.

Objective: To evaluate the economic impact of a Bayesian network model designed to predict clinical success of a new chemical entity (NCE) based on pre-phase III data., Methods: We trained our Bayesian network model on publicly accessible data on 503 NCEs, stratified by therapeutic class. We evaluated the sensitivity, specificity and accuracy of our model on an independent data set of 18 NCE-indication pairs, using prior probability data for the antineoplastic NCEs within the training set. We performed Monte Carlo simulations to evaluate the economic performance of our model relative to reported pharmaceutical industry performance, taking into account reported capitalized phase costs, cumulative revenues for a postapproval period of 7 years, and the range of possible false negative and true negative rates for terminated NCEs within the pharmaceutical industry., Results: Our model predicted outcomes on the independent validation set of oncology agents with 78% accuracy (80%sensitivity and 76% specificity). In comparison with the pharmaceutical industry's reported success rates, on average our model significantly reduced capitalized expenditures from $727 million/successful NCE to $444 million/successful NCE (P < 0.001), and significantly improved revenues from $347 million/phase III trial to $507 million/phase III trial (P < 0.001) during the first 7 years post launch. These results indicate that our model identified successful NCEs more efficiently than currently reported pharmaceutical industry performances., Conclusions: Accurate prediction of NCE outcomes is computationally feasible, significantly increasing the proportion of successful NCEs, and likely eliminating ineffective and unsafe NCEs.

Nephrotic syndrome, a malfunction of the kidney glomerular filter, leads to proteinuria, edema and, in steroid-resistant nephrotic syndrome, end-stage kidney disease. Using positional cloning, we identified mutations in the phospholipase C epsilon gene (PLCE1) as causing early-onset nephrotic syndrome with end-stage kidney disease. Kidney histology of affected individuals showed diffuse mesangial sclerosis (DMS). Using immunofluorescence, we found PLCepsilon1 expression in developing and mature glomerular podocytes and showed that DMS represents an arrest of normal glomerular development. We identified IQ motif-containing GTPase-activating protein 1 as a new interaction partner of PLCepsilon1. Two siblings with a missense mutation in an exon encoding the PLCepsilon1 catalytic domain showed histology characteristic of focal segmental glomerulosclerosis. Notably, two other affected individuals responded to therapy, making this the first report of a molecular cause of nephrotic syndrome that may resolve after therapy. These findings, together with the zebrafish model of human nephrotic syndrome generated by plce1 knockdown, open new inroads into pathophysiology and treatment mechanisms of nephrotic syndrome.

We have previously reported sirolimus (SRL) pharmacokinetics (PK) in pediatric renal transplant recipients on a calcineurin inhibitor (CNI)-free protocol. We now report pediatric SRL PK in pediatric renal transplant patients receiving SRL + CNI. SRL was dosed to achieve target trough levels between 10 and 20 ng/mL. We performed 49 SRL PK profiles in pediatric renal transplant recipients receiving SRL in combination with either cyclosporine (CsA; 25 profiles), or tacrolimus (TCL; 24 profiles). Ten of the SRL + TCL profiles were obtained from children receiving SRL on a b.i.d. dosing regimen. All other SRL profiles were q.d. regimens. We calculated, the maximum concentration (C(max)), AUC, apparent clearance (aCL; dose/AUC) for dose in mg/m(2), and mean residence time (MRT). SRL levels were measured at 6 and 7 time points for b.i.d. and q.d. dosing, respectively. Regression analysis of SRL trough values vs. AUC showed good correlation in the SRL q.d. + CsA, SRL q.d. + TCL, and SRL b.i.d. + TCL groups (r(2) = 0.95, 0.68, and 0.44, respectively). SRL aCL corrected for body surface area was higher in children aged 0-5 yr receiving SRL with either CsA or TCL. SRL dosing schedule should be tailored to each patient. Higher SRL aCL may be present in younger children when administered with CNI.

The prognosis of pediatric nephrotic syndrome (NS) correlates with the responsiveness to glucocorticoid therapy. Steroid-resistant NS (SRNS) patients progress to end-stage renal disease, while steroid-sensitive NS (SSNS) and steroid-dependent (SDNS) patients do not. We have performed proteomic profiling of urine samples from a cross section of pediatric and adolescent subjects with SSNS, SRNS, and orthostatic proteinuria (OP) to identify urinary biomarkers of steroid resistance. We performed surface-enhanced laser desorption/ionization time of flight mass spectrometry (SELDI-TOF MS) on urine from 19 subjects with SSNS/SDNS in remission, 14 with SSNS/SDNS in relapse, 5 with SRNS in relapse, and 6 with OP. Genetic algorithm search of principal component space revealed a group of five peaks distinguishing SRNS subjects, with mass/charge (m/z) values of 3,917.07, 4,155.53, 6,329.68, 7,036.96, and 11,117.4. Our analyses identified the peak at m/z 11,117.4 with an accuracy of 95% for classifying SRNS. Multidimensional protein fractionation and mass spectrometric analysis of SRNS urine samples combined with immunodepletion identified the 11,117.4 protein as beta2-microglobulin (B2M). Using an unbiased protein profiling approach, we have validated previously reported findings of B2M as a biomarker associated with SRNS. Prospective studies are warranted to establish additional biomarkers that would be predictive of SRNS.

The goal of this study was to estimate the diagnostic and prognostic accuracy of renal biopsies in focal segmental glomerulosclerosis (FSGS), accounting for the focal nature of affected glomeruli. Computational simulations were performed on a total of 138,600 virtual kidneys, across a range of FSGS involvement. Simulations were designed to address the diagnostic accuracy of renal biopsies, and the biopsy characteristics required to reflect accurately the true degree of involvement of FSGS in the entire kidney or just the juxtamedullary (JM) region. The diagnostic accuracy of renal biopsies for the detection of at least one FSGS glomerulus exceeded 80% when 10-20% of the kidney was affected by FSGS. Hundreds to thousands of biopsy glomeruli were required to characterize reliably the true extent of FSGS when fewer than 75% of the kidney's glomeruli were affected. Renal biopsies with an average of 20 glomeruli did not accurately reflect the extent of FSGS in kidneys until at least 80% of all glomeruli in the kidney were affected. Targeting JM glomeruli did not result in significant improvements in the prognostic performance characteristics of renal biopsies. These findings suggest that conventional renal biopsies might be inadequate for characterizing the extent of FSGS.

Proteomic profiling with SELDI-TOF MS has facilitated the discovery of disease-specific protein profiles. However, multicenter studies are often hindered by the logistics required for prompt deep-freezing of samples in liquid nitrogen or dry ice within the clinic setting prior to shipping. We report high concordance between MS profiles within sets of quadruplicate split urine and serum samples deep-frozen at 0, 2, 6, and 24 h after sample collection. Gage R&R results confirm that deep-freezing times are not a statistically significant source of SELDI-TOF MS variability for either blood or urine.

Numerous cellular and molecular perturbations have been studied to elucidate the pathogenic mechanisms underlying nephrotic-range proteinuria, which may in turn shed light on disease-specific mechanisms. We have analyzed the publicly available data from the PhysGen partial panel of consomic rats to determine whether there are quantitative trait loci that associate with nephrotic-range proteinuria. As of this writing, consomic rat strains subjected to the renal protocol have been bred by the Program for Genomic Applications for 15 of the 22 rat chromosomes for both genders, predominantly with the Brown-Norway (BN) and Dahl salt-sensitive (SS) strains as parents. We defined chromosomes of interest as consomic SS-xBN strains whose phenotype measurements differed significantly from SS but not BN strains, stratified by gender. We filtered and clustered differentially expressed genes by function in renal tissue from relevant strains. Proteinuria was significantly higher in male SS vs. male SS-18BN, and it was significantly higher in male SS vs. female SS. Functional clustering of differentially expressed genes yielded two specific functional clusters: apoptosis (p=0.022) and angiogenesis (p=0.046). Gene expression profiles demonstrated differential expression of apoptotic and angiogenic genes. However, TUNEL stains of renal tissue showed no significant difference in the number of apoptotic nuclei. We conclude that chromosomes 18 and X are quantitative trait loci for nephrotic-range proteinuria in rats.

The field of proteomics is developing at a rapid pace in the post-genome era. Translational proteomics investigations aim to apply a combination of established methods and new technologies to learn about protein expression profiles predictive of clinical events, therapeutic response, and underlying mechanisms. However, in contrast to genetic studies and in parallel with gene expression studies, the dynamic nature of the proteome in conjunction with the challenges of accounting for post-translational modifications requires the translational proteomics investigator to understand the strengths and limitations of proteomics approaches. In this review, we provide an overview of proteomics approaches and techniques, and proteomics informatics for clinical transplantation investigators. We also review recent publications pertaining to transplantation proteomics, and discuss the implications and utility of urine proteomics for non-invasive investigation of transplant outcomes.

Many hypothesize that subtle inflammation and immune activity detected in the intraoperative period are linked to adverse postkidney transplant clinical outcomes. To this end, renal allografts were analyzed for expression of pro-inflammatory, inflammation-induced adhesion molecules, immune activation as well as anti-apoptotic genes expressed 15 min after vascular reperfusion (zero-hour) to determine whether this analysis can aid in predicting the occurrence of delayed graft function (DGF), acute rejection (AR), and the quality of graft function at 6 mo. Intraoperative biopsies were obtained from 75 consecutively performed renal allografts in which consent was obtained 15 min after vascular reperfusion. These biopsies were analyzed by quantitative real-time PCR for transcription of 15 select genes and by standard histopathology. Posttransplant clinical outcomes were also analyzed in respect to intraoperative transcriptional profiles and clinical parameters available at the time of transplantation. This study demonstrates that a limited and hypothesis-driven PCR-based transcriptional profile of the zero-hour kidney biopsy predicts posttransplant clinical outcomes including DGF, early AR, and the quality of renal function 6 mo posttransplantation. For some clinical endpoints, the combined use of molecular analysis and established clinical indicators available at the time of transplantation further enhances the quality of prognosis. The transcriptional profiling data provide absolutely essential data to the predictive models, particularly with respect to AR and renal function 6 mo posttransplantation.

Idiopathic nephrotic syndrome is the most common glomerular disorder of childhood. Recurrence of nephrotic syndrome immediately following renal transplantation is rapid, results in a high rate of graft loss, and represents the most severe form of nephrotic syndrome. This review discusses the molecular heterogeneity of pediatric nephrotic syndrome across the spectrum of disease activity. A schema is offered for a molecular approach to pediatric nephrotic syndrome, including immune-mediated and structural/genetic factors., (Copyright 2004 Blackwell Munksgaard)

Immunosuppression with SRL may provide an opportunity to avoid long-term exposure to the nephrotoxicity of CNI. Thus, we have initiated an experimental protocol of IL-2r antibody induction, prednisone, MMF and SRL in pediatric renal transplant recipients (median age 15.5 yr, IQR 8.5, range 1.3-21.7). The recipients were treated with daclizumab every 2 wk for the first 2 months, prednisone on a tapering schedule, MMF at 1200 mg/m(2)/day and SRL given b.i.d. The SRL was dosed to achieve defined target whole blood 12-h trough levels. We performed 24 SRL PK profiles in 13 stable pediatric renal transplant recipients at 1 and 3 months post-transplant. Half-life (T(1/2)) and terminal T(1/2) were 9.7 (7.1-24.6) and 10.8 (4.4-95.2) hours (median, range) respectively at month 1, and were 9.6 (5-17.8) and 12.1 (4.7-71.0) hours respectively at month 3. SRL trough levels correlated with AUC (r(2) = 0.84, p < 0.001). There was no relationship between SRL and mycophenolic acid (MPA) AUC values (r(2) = 0.04). During the first 3 months post-transplant only one patient experienced severe neutropenia and another patient had subclinical (histologic) evidence of a mild acute rejection episode with no change in renal function. We conclude that the T(1/2) of SRL in pediatric renal transplant recipients not treated with CNI is much shorter than what has been reported for adults, due to rapid metabolism. We conclude that children require SRL dosing every 12 h, higher doses and frequent drug monitoring to achieve target SRL concentrations.

Natural killer cells constitute 50-90% of lymphocytes in human uterine decidua in early pregnancy. Here, CD56(bright) uterine decidual NK (dNK) cells were compared with the CD56(bright) and CD56(dim) peripheral NK cell subsets by microarray analysis, with verification of results by flow cytometry and RT-PCR. Among the approximately 10,000 genes studied, 278 genes showed at least a threefold change with P < or = 0.001 when comparing the dNK and peripheral NK cell subsets, most displaying increased expression in dNK cells. The largest number of these encoded surface proteins, including the unusual lectinlike receptors NKG2E and Ly-49L, several killer cell Ig-like receptors, the integrin subunits alpha(D), alpha(X), beta1, and beta5, and multiple tetraspanins (CD9, CD151, CD53, CD63, and TSPAN-5). Additionally, two secreted proteins, galectin-1 and progestagen-associated protein 14, known to have immunomodulatory functions, were selectively expressed in dNK cells.

Mutations in NPHS2, encoding podocin, have been identified in childhood onset focal and segmental glomerulosclerosis (FSGS). The role of NPHS2 in adult disease is less well defined. We studied 30 families with FSGS and apparent autosomal recessive inheritance and 91 individuals with primary FSGS. We screened family members for NPHS2 mutations. NPHS2 mutations appeared to be responsible for disease in nine of these families. In six families, the affected individuals were compound heterozygotes for a nonconservative R229Q amino acid substitution. This R229Q variant has an allele frequency of 3.6% in a control population. In these families, R229Q was the only mutation identified on one of the two disease-associated NPHS2 alleles. We used in vitro-translated podocin and purified nephrin to investigate the effect of R229Q on their interaction and found decreased nephrin binding to the R229Q podocin. These data suggest that this common polymorphism contributes to the development of FSGS. Chromosomes bearing the R229Q mutation share a common haplotype defining an approximately 0.2-Mb region. R229Q appears to enhance susceptibility to FSGS in association with a second mutant NPHS2 allele. Identification of R229Q mutations may be of clinical importance, as NPHS2-associated disease appears to define a subgroup of FSGS patients unresponsive to corticosteroids.

Background: The pathogenesis of childhood nephrotic syndrome (NS), whether the lesion is minimal change disease (MCD) or focal segmental glomerulosclerosis (FSGS), remains elusive. Based on the presence of elevated cytokine levels in peripheral blood, a T cell-induced injury could be postulated., Methods: To test the hypothesis that infiltrating T cells actively contribute to the glomerular injury in children with NS, we studied the intrarenal transcription of various T cell-related chemokines, cytokines and cytotoxic T-lymphocyte (CTL) effector molecules in the renal biopsy tissue of 52 nephrotic children with a variety of histologic lesions. Intrarenal gene expression was studied using reverse transcription (RT)-assisted-polymerase chain reaction (PCR)., Results: Interleukin-2 (IL-2) and IL-4 transcripts were not observed in any of the specimens. IL-2 receptor alpha mRNA was detected in 24 of 40 proteinuric patients, but also in 6 of 10 patients in remission and showed no significant differences with regard to steroid response. Intrarenal gene expression of CTL mediators and transforming growth factor-beta1 (TGF-beta1) was noted particularly in patients with progressive disease leading to chronic renal failure. TGF-beta1 gene expression was noted in 23 of 29 steroid resistant (SR) children with NS not caused by lupus nephritis and in 18 of 20 FSGS patients. In contrast TGF-beta1 gene expression was detected in only 3 of 14 steroid-sensitive patients (P < 0.001). Two of these patients later developed FSGS. In patients with steroid-resistant NS, intrarenal TGF-beta1 gene expression showed a positive predictive value of 90% and a negative predictive value of 88% to identify FSGS (P < 0.0001)., Conclusion: These results support the notion that immunologically mediated events contribute to the progressive renal damage seen in children with FSGS.

We have devised a gene-clustering algorithm that is completely unsupervised in that no parameters need be set by the user, and the clustering of genes is self-optimizing to yield the set of clusters that minimizes within-cluster distance and maximizes between-cluster distance. This algorithm was implemented in Java, and tested on a randomly selected 200-gene subset of 3000 genes from cell-cycle data in S. cerevisiae. AlignACE was used to evaluate the resulting optimized cluster set for upstream cis-regulons. The optimized cluster set was found to be of comparable quality to cluster sets obtained by two established methods (complete linkage and k-means), even when provided with only a small, randomly selected subset of the data (200 vs 3000 genes), and with absolutely no supervision. MAP and specificity scores of the highest ranking motifs identified in the largest clusters were comparable.

Recurrence of nephrotic syndrome (NS) after transplantation (Tx) occurs in 20-50% of renal transplant recipients, with a median time to recurrence of 14 days and a 50% rate of graft loss. We performed a retrospective analysis of 22 pediatric patients with NS who received renal transplants at the Children's Hospital, Boston, between 1982 and 1999. During the first 14 days following Tx, 13 (59%) patients developed clinical recurrent nephrotic syndrome (RNS). RNS developed in 50% of living donor recipients and in 70% of cadaveric donor recipients (p= non-significant). Seven of the 13 patients with RNS were treated with plasmapheresis, while six received standard immunosuppressive induction therapy only. Two of the seven treated patients and one of the six untreated patients lost their grafts to RNS, yielding a total RNS graft loss rate of 23%. However, patients with RNS who achieved remission had significantly higher cumulative graft survival at 5 yr than did RNS patients who did not achieve remission (p< 0.001). Overall cumulative graft survival at 5 yr was not significantly different between the two groups: 67% in those with non-recurrent nephrotic syndrome (NRNS) vs. 64% in those with RNS, p= non-significant. We conclude that successful reversal of early RNS improves long-term graft survival in pediatric RNS. Multi-center studies are sorely needed to develop novel, less toxic therapies for native and recurrent NS.

In an attempt to identify potential markers of steroid-resistance in focal segmental glomerulosclerosis (FSGS) we evaluated intra-graft gene expression of IkappaBalpha, nuclear factor-kappaB (NF-kappaB), and angiotensinogen in 60 biopsies from 27 pediatric renal transplant recipients. Intra-graft NF-kappaB expression was significantly elevated in recurrent FSGS (R-FSGS) (218.3 + 55.6 ag/fg versus NON-FSGS 121.1 + 19.9, P=0.04) but not in acute rejection. NF-kappaB:IkappaBalpha ratios were higher in cadaveric donor versus living related donor recipients (15.7 + 2.8 vs. 8.8 + 1.3, respectively, P=0.015), and in African-American versus Caucasian recipients (15.6 + 2.9 vs. 9.1 + 1.3, respectively, P=0.03). Intra-graft angiotensinogen gene expression was significantly elevated in R-FSGS (30.5 + 8.8 ag/fg R-FSGS vs. 16.0 + 4.7 NON-FSGS, P=0.009). We conclude that increased NF-kappaB and angiotensinogen gene expression are associated with R-FSGS. Increased NF-kappaB:IkappaBalpha ratios are associated with cadaveric donor recipients and African-American race.

Advances in the understanding of genetic engineering, protein expression, and Ig function have come together to allow for the rapid synthesis and production of a novel generation of potent immunomodulating reagents. Selective approaches that allow the isolation of desired specificities will be elucidated through meticulous engineering techniques. This, in turn, may eventually result in the fulfillment of the tremendous potential of engineered proteins for therapeutic and diagnostic applications.

We have followed four patients with Bartter syndrome for a mean of 25.4 years (range 21.5-28.8 years) after diagnosis. All patients received non-steroidal anti-inflammatory drugs (NSAID). In all patients, various degrees of renal dysfunction were noted to be temporally associated with NSAID therapy. In two patients, renal dysfunction resolved after discontinuing NSAID therapy, while maintaining other chronic medications such as potassium-sparing diuretics. Renal dysfunction persisted after NSAID withdrawal in two patients. We report these cases as a warning that NSAID should be considered an important cause of either reversible or irreversible renal dysfunction in Bartter syndrome.

Background: We have shown previously that heightened expression of the cytotoxic lymphocyte (CL) effector genes perforin (P), granzyme B (GB), and Fas ligand (FasL), is closely correlated with acute allograft rejection, particularly when two or more target genes are up-regulated., Methods: We used quantitative reverse transcription-polymerase chain reaction to analyze CL gene expression from peripheral blood leukocytes (PBLs) and renal allograft biopsies in 31 paired samples of PBLs and renal tissue from 25 renal allograft recipients. Our aims were (1) to determine whether the expression of CL gene expression in PBLs correlates with expression of these genes in renal allograft biopsy tissue and (2) to determine whether CL gene expression in PBLs correlates with the histological diagnosis., Results: Coordinate gene expression in PBLs and acutely rejecting allografts was found in 9/11 (82%) for P, 07/11 (64%) for GB, and 10/11 (91%) for FasL. Coordinate absence was found in 15/20 (75%) for P, 17/20 (85%) for GB, and 16/20 (80%) for FasL in nonrejecting allografts. Furthermore, up-regulation of any two genes in PBLs correlated with pathological diagnosis of rejection with excellent positive (100%) and negative (95%) predictive values., Conclusion: Coordinate CL gene expression in PBLs and the allograft is usually detected. CL gene expression in PBLs is closely associated with a pathologic diagnosis of rejection. CL gene expression in PBLs may serve as a noninvasive method of monitoring for renal allograft rejection.

Background: Interleukin (IL)-2, IL-4, IL-7, IL-9, and IL-15, all T-cell growth factors (TCGFs), utilize the common IL-2 receptor gammac chain as a critical signaling component in their receptor complexes. We have bred IL-2-/- and IL-4-/- double knockout (DKO) mice and showed vigorous islet allograft rejection by DKO hosts. The identity of TCGFs that support the IL-2- and IL-4-independent allograft rejection is unclear., Methods: We analyzed IL-9 gene expression in rejecting islet allografts in wild-type and in DKO mice, as well as in human renal transplant biopsy specimens, by reverse transcriptase polymerase chain reaction and compared the expression of IL-9 with that of other TCGFs., Results: IL-9 gene expression was not detected in rejecting murine islet allografts in either wild-type or DKO recipient mice despite robust expression of other TCGFs, including IL-7 and IL-15. IL-9 transcripts were also not expressed in any of the human renal transplant biopsies obtained 4 to 251 days after transplantation, regardless of the presence or absence of histological evidence of rejection. Despite expression of IL-9 by DKO splenic cells upon in vitro mitogenic stimulation, IL-9 alone was unable to stimulate the proliferation of concanavalin A-activated splenic leukocytes harvested from DKO mice., Conclusion: IL-9 is conspicuously absent despite vigorous expression of IL-2, IL-4, IL-7, and IL-15 genes during acute allograft rejection.

Background: The interaction between CD40 and its ligand CD40L is essential for the development and maintenance of vigorous immunity. We have sought to determine: (i) whether a heightened level of CD40L transcripts is evident during acute allograft rejection and (ii) the kinetics of CD40L gene expression during allograft rejection., Methods: By using quantitative reverse transcriptase-assisted polymerase chain reaction techniques, we found that heightened CD40L gene expression is a correlate of acute human renal allograft rejection (P<0.01)., Results: In a murine model of MHC-mismatched islet allografts, our results showed that CD40L transcripts were rarely detected at day 2 after transplantation, but were remarkably heightened at day 5 after transplantation. The transcript levels then steadily increased and peaked at the time of rejection., Conclusion: These data suggest that therapy aimed at blocking the CD40 to CD40L interaction should be applied during the immediate posttransplant period.