Yuko Akioka

EFFECT OF PRE- AND POSTOPERATIVE PLASMAPHERESIS ON POSTTRANSPLANT RECURRENCE OF FOCAL SEGMENTAL GLOMERULOSCLEROSIS IN CHILDREN

Background. Posttransplant recurrence is frequent in patients who received renal transplantation for focal segmental glomerulosclerosis (FSGS). The recurrence has been ascribed to a circulating permeability factor or factors. We have used plasmapheresis (PP) to treat recurrent FSGS and also studied whether preoperative PP is effective in preventing recurrence of FSGS. Methods. We retrospectively analyzed 21 allografts of 20 patients with nephrotic syndrome and biopsy-proven FSGS. They were divided into two groups depending on whether they had prophylactic PP; a prophylactic (n=15) and a nonprophylactic group (n=6). PP was performed two to three times prophylactically and therapeutically until proteinuria was markedly reduced. In each session, 50–75 ml/kg of the patient’s plasma was exchanged with 5–8% albumin. Results. FSGS recurred in 9 of 21 allografts, 4 of 6 in the nonprophylactic group, and 5 of 15 in the prophylactic group. Therapeutic PP was performed in seven of nine recurrent patients without definite adverse effect, with satisfactory results except in one patient. Children lost proteinuria after 6 to >100 sessions of PP and the number correlated with the pretreatment level of proteinuria. The mean follow-up periods were 62.7 and 41.6 months for the prophylactic and nonprophylactic groups, respectively. At the last follow-up, 66.7% of relapsing and 81.8% of nonrelapsing patients had a functioning graft. Conclusion. PP appears to be effective for the prevention and treatment of posttransplant recurrence of FSGS, although further consideration of cost/benefit and risks is required before a conclusive judgement can be made.

ABO-incompatible pediatric kidney transplantation in a single-center trial

Abstract We have performed ten pediatric kidney transplantations from living-related ABO-incompatible donors. All patients underwent preoperative plasmapheresis with or without immunoadsorption to reduce isoagglutinin. Primary immunosuppression consisted of methylprednisolone, cyclosporin or tacrolimus, azathioprine, anti-lymphocyte globulin, and/or deoxyspergualin. At transplantation splenectomy was simultaneously performed in all patients. Median follow-up is 65 months (range 4–95 months). The patient and graft survival rates are 100% to date. Post-transplantation isoagglutinin titers did not increase more than 1:32, except for 1 patient, without uncontrollable vascular rejection episodes. Despite the heavy immunosuppressive regimen, cytomegalovirus infection occurred in only three patients, who were successfully treated with ganciclovir and cytomegalovirus high-titer gamma globulin. Our small series clearly shows that the preoperative reduction of isoagglutinin, splenectomy, and strict immunosuppressive therapy lead to successful long-term results in children.

Increase of Integrin‐Linked Kinase Activity in Cultured Podocytes upon Stimulation with Plasma from Patients with Recurrent FSGS

Recurrent focal segmental glomerulosclerosis (FSGS) is a major challenge in the field of transplantation. Integrin‐linked kinase (ILK) has emerged as a key mediator of podocyte–glomerular basement membrane (GBM) interactions. To clarify the involvement of plasma factors in FSGS recurrence, we examined the effects of plasma from FSGS patients with or without posttransplant recurrence on cultured podocytes, focusing particularly on ILK activity. Podocytes from a conditionally immortalized mouse podocyte cell line were treated with plasma from 11 FSGS patients, and ILK activity was determined using an immune complex kinase assay. Treatment with plasma from three patients with recurrence induced an increase in ILK activity. In contrast, no increase in ILK activity was observed in cultured podocytes treated with plasma from the remaining three patients with recurrence and five patients without recurrence. Cultured podocytes treated with plasma that induced ILK activity showed alterations of focal contact and detachment from the laminin matrix. In conclusion, this preliminary study provides experimental evidence suggesting the possible presence of circulating toxic factors in the plasma of some patients with recurrent FSGS, which induce an increase in podocyte ILK activity that may lead to the detachment of podocytes from the GBM.

V2 Vasopressin Receptor (V2R) Mutations in Partial Nephrogenic Diabetes Insipidus Highlight Protean Agonism of V2R Antagonists

Background: Inactivating mutations of the V2 receptor (V2R) cause nephrogenic diabetes insipidus (NDI). Results: Two V2R mutants discovered in partial NDI show partial defects, and V2R antagonists rescued them. Conclusion: V2R antagonists operate as pharmacochaperones for defective mutants, whereas they operate as inverse agonists for normal receptors. Significance: V2R antagonists can act as protean agonists, potentially underlying their dual effects. Inactivating mutations of the V2 vasopressin receptor (V2R) cause cross-linked congenital nephrogenic diabetes insipidus (NDI), resulting in renal resistance to the antidiuretic hormone AVP. In two families showing partial NDI, characterized by an apparently normal response to diagnostic tests and an increase in the basal ADH levels suggesting AVP resistance, we have identified two V2R mutations, Ser-333del and Y128S. Both mutant V2Rs, when expressed in COS-7 cells, show partial defects in vasopressin-stimulated cAMP accumulation and intracellular localization. The inhibition of internalization does not rescue their localization. In contrast, the non-peptide V2R antagonists OPC41061 and OPC31260 partially rescue the membrane localization and basal function of these V2R mutants, whereas they inhibit the basal activity of the wild-type V2R. These results indicate that a partial loss of function of Ser-333del and Y128S mutant V2Rs results from defective membrane trafficking. These findings further indicate that V2R antagonists can act as protean agonists, serving as pharmacological chaperones for inactivating V2R mutants and also as inverse agonists of wild-type receptors. We speculate that this protean agonism could underlie the possible dual beneficial effects of the V2R antagonist: improvement of hyponatremia with heart failure or polycystic kidney disease and potential rescue of NDI.

Decreased glomerular filtration as the primary factor of elevated circulating suPAR levels in focal segmental glomerulosclerosis

BackgroundCirculating factor(s) has been thought to be the underlying cause of focal segmental glomerulosclerosis (FSGS), and recent studies foster this idea by demonstrating increased soluble urokinase receptor (suPAR) levels in the serum of FSGS patients.MethodsTo explore the possible contribution of suPAR in FSGS pathogenesis, we analyzed serum suPAR levels in 17 patients with FSGS and compared them with those in patients with steroid-sensitive nephrotic syndrome, chronic glomerulonephritis, or non-glomerular kidney diseases.ResultsSerum suPAR levels in patients with FSGS were higher than those in patients with steroid-sensitive nephrotic syndrome or chronic glomerulonephritis, but not higher than those in patients with non-glomerular kidney diseases. suPAR levels negatively correlate with estimated glomerular filtration rate and were decreased after renal transplantation in patients with FSGS as well as in those with non-glomerular kidney diseases. Furthermore, 6 FSGS patients with post-transplant recurrence demonstrated that suPAR levels were not high during the recurrence.ConclusionsBased on our results, elevated suPAR levels in FSGS patients were attributed mainly to decreased glomerular filtration. These data warrant further analysis for involvement of possible circulating factor(s) in FSGS pathogenesis.

Asymptomatic high Epstein–Barr viral load carriage in pediatric renal transplant recipients

Tanaka E, Sato T, Ishihara M, Tsutsumi Y, Hisano M, Chikamoto H, Akioka Y, Dohno S, Maeda A, Hattori M, Wakiguchi H, Fujieda M. Asymptomatic high Epstein‐Barr viral load carriage in pediatric renal transplant recipients.
Pediatr Transplantation 2011: 15: 306–313.

Antineutrophil cytoplasmic antibody-positive crescentic glomerulonephritis associated with propylthiouracil therapy.

Abstract A teenage girl with crescentic glomerulonephritis had antineutrophil cytoplasmic antibody (ANCA) detected after she had received propylthiouracil (PTU) for hyperthyroidism without cutaneous vasculitis. ANCA was detected on admission; renal biopsy showed crescentic glomerulonephritis with focal necrotizing glomerulonephritis but no immune deposits. Administration of steroid and decreasing the dose of PTU produced a good clinical response and the ANCA disappeared. It was concluded that ANCA is closely related to the pathogenesis of crescentic glomerulonephritis and that treatment with PTU appeared to induce ANCA.

Screening of vesicoureteral reflux in pediatric patients with kidney transplantation showing non-specific interstitial fibrosis and tubular atrophy with interstitial Tamm–Horsfall protein deposits in protocol allograft biopsy

Abstract Interstitial fibrosis and tubular atrophy (IF/TA) in kidney allografts are induced by multiple factors, and although much effort has been devoted on the classification of IF/TA, clarification of the causes of non‐specific IF/TA is equally important for appropriate therapy. Tamm–Horsfall protein (THP) in renal tissue can be a useful marker for the histological expression of urine backflow and suspected vesicoureteral reflux (VUR). Here, we examined the presence of VUR in pediatric recipients with interstitial THP deposits in kidney allografts to clarify the cause of non‐specific IF/TA. Ten pediatric patients showing interstitial THP deposits with non‐specific IF/TA were enrolled and voiding cystourethrography was performed. Major histological findings of these patients were interstitial mononuclear cell infiltration and fibrosis associated with interstitial THP deposits. The semiquantitative grading scores of IF/TA were as follows: (i) mild IF/TA (n = 3); (ii) moderate IF/TA (n = 3); and (iii) severe IF/TA (n = 4). In the severe grade, diffuse interstitial mononuclear cell infiltrates were prominent with the appearance of thyroidization classically observed in chronic pyelonephritis. Eight of ten patients (80%) had VUR into the graft. Although symptomatic pyelonephritis was not observed in any of the patients, asymptomatic bacteriuria was detected in 40.0% of the patients. There was no significant correlation between VUR grade and IF/TA histological score. The patients without VUR also showed mild or severe IF/TA. Therefore, VUR and urinary flow stasis accompanied by asymptomatic urinary tract infection appear to be the causes of interstitial mononuclear cell infiltration and fibrosis associated with interstitial THP deposits in kidney allografts.

Epstein-Barr virus load for early detection of lymphoproliferative disorder in pediatric renal transplant recipients.

AIM The aims of this study were to establish a protocol for monitoring Epstein-Barr virus (EBV) infection for identification of pediatric renal transplant recipients with a high risk of developing posttransplant lymphoproliferative disorder (PTLD) and to predict the development of PTLD. SUBJECTS AND METHODS Peripheral blood mononuclear cells (PBMCs) and plasma EBV loads were measured by nested PCR (n-PCR) and real-time PCR (r-PCR) every 1 - 3 months after grafting in 17 pediatric recipients who were seronegative for EBV before grafting (4 with EBV-associated symptoms, including 2 with PTLD (Group A); 6 with asymptomatic persistent high EBV loads in PBMCs of > 1,000 copies/µgDNA for over 6 months (Group B); and 7 with neither EBV-associated symptoms nor persistent high EBV loads in PBMCs (Group C)). RESULTS n-PCR revealed EBV-DNA in PBMCs from all patients. The EBV genome was present in plasma in 3 (75%), 1 (17%), and 0 (0%) in Groups A, B and C (p < 0.01 for A vs. B and A vs. C). EBV loads detected by r-PCR in PBMCs were significantly higher in Groups A (p < 0.05) and B (p < 0.01) compared to Group C. EBV genomes in plasma were detected by n- and r-PCR in only the 2 cases with PTLD. One patient with lymphadenitis in Group A and 1 patient in Group B had EBV-DNA in plasma based on n-PCR, but the viral loads using r-PCR were < 250 copies/ml. CONCLUSION EBV loads in PBMCs alone are insufficient for predicting EBV-associated symptoms including PTLD. Plasma EBV loads (over 250 copies/ml) estimated by r-PCR may be useful to distinguish PTLD from other EBV-associated diseases or asymptomatic viremia.

Monitoring of Epstein-Barr virus load and killer T cells in pediatric renal transplant recipients.

AIM The aim of this study is to establish a monitoring method to prevent Epstein-Barr virus (EBV)-associated symptoms including post-transplant lymphoproliferative disorder (PTLD) that occur after pediatric renal transplantation. SUBJECTS AND METHODS Circulating EBV loads were quantified by real-time PCR every 1 - 3 months after grafting in 22 pediatric recipients (13 EBV-seronegative [R(-)] and 9 EBV-seropositive [R(+)] recipients before grafting). The peripheral blood cell populations of non-specific activated killer cells (CD8+HLA-DR+ phenotype) in 13 R(-) recipients and EBV-specific cytotoxic T cells (CTLs) reactive with a tetramer expressing HLA-A24-restricted EBV-specific antigens in 8 of 13 R(-) recipients were determined by flow cytometry. RESULTS EBV-associated symptoms including PTLD (2 cases) were found in 4 R(-) and none of the R(+) recipients. The maximum of EBV load in the R(-) group was significantly higher that in the R(+) group. In R(-) recipients, 4 symptomatic cases had significantly more EBV genome than asymptomatic cases. EBV-specific CTLs were detected in 6 of the 8 R(-) recipients, but these CTLs could not be detected in 1 of the 2 cases at onset of PTLD. The percentage of CD8+HLA-DR+ cells was significantly higher in asymptomatic recipients than in recipients with EBV-associated symptoms whose EBV loads were over 400 copies/microg DNA. CONCLUSION Monitoring of killer T cells and EBV loads may allow assessment of the risk of EBV-associated symptoms, and high EBV loads and low EBV-specific and/or non-specific CTL responses may be predictive for development of EBV-associated symptoms such as PTLD.