Plasma levels of soluble CD30 are increased in children with chronic renal failure and with primary growth deficiency and decrease during treatment with recombination human growth hormone.

BACKGROUND: Previous studies have suggested that in vivo Th2 lymphocyte activation is related to increased soluble CD30 (sCD30) plasma levels. As various hormones (dehydroepiandrosterone, glucocorticoids, progesterone) can regulate the Th1/Th2 balance, and because growth hormone (GH) enhances lymphocyte function, we measured sCD30 plasma levels, before and after treatment with recombinant human GH (rhGH), in children with growth failure due to chronic renal failure (CRF) or to isolated GH deficiency in order to evaluate the potential effects of rhGH treatment on Th1/Th2 balance. METHODS: sCD30 plasma levels were determined by ELISA assay in 30 children with CRF (mean age 10.7+/-3.7 years), in five children with isolated GH deficiency (mean age 11.4+/-2.6 years), and in 10 normal controls (mean age 10.1+/-3.5 years). RESULTS: sCD30 levels were higher in the 30 children with CRF than in the 10 controls (179.8+/-79.4 vs 11.3+/-10.9 U/ml, P<0.001) exhibiting an inverse correlation with glomerular filtration rate (GFR) (r=-0.7860, P<0.001). In 11 children with CRF, after 19.9+/-16.7 months of rhGH treatment, a decrease of sCD30 plasma level (170+/-50 vs 134+/-49 U/ml, P<0.01) was observed. The five children with primary GH deficiency had higher sCD30 plasma level than controls (mean 147+/-105 vs 11+/-10 U/ml, P<0.004) and sCD30 plasma levels decreased to 95.2+/-109.6 U/ml after rhGH treatment. CONCLUSIONS: The finding that rhGH treatment decreased sCD30 plasma levels in children with CRF, and that children with primary GH deficiency had higher sCD30 plasma levels than controls, suggest that GH may regulate CD30 expression and possibly the balance of Th1/Th2. Whether the uraemia-induced increase in sCD30 is due to decreased renal excretion, to overproduction or both, remains to be determined.

Inhibition of renal permeability towards albumin: a new function of apolipoproteins with possible pathogenetic relevance in focal glomerulosclerosis.

Focal segmental glomerulosclerosis (FSGS) is a degenerative renal disease characterized by the accumulation of extracellular matrix and lipids within the glomerular tuft. It has been proposed that an abnormal renal permeabilization towards proteins induced by a putative plasma factor is, in some way, involved in the pathogenesis of the disease. In this paper, we measured the plasma permeability activity (Palb) in several sera of patients with FSGS and found a mean activity of 0.82+/-0.03 which means a marked increase compared to a mean Palb of 0.16+/-0.03 in normal controls. Coincubation of FSGS and normal serum reduced the permeability activity within the normal range; normal serum added to the incubation medium after the glomeruli had already been exposed to the FSGS serum had no effect, suggesting the presence of inhibitory substances with a direct effect on a circulating substrate. Finally, the antipermeability activity was retained when heated to 60 degrees C but not to 100 degrees C. By serial fractionations of normal serum and reported activity measurements at each step, five natural occurring inhibitors of albumin permeabilization were purified and characterized by matrix assisted laser desorption/ionization-mass spectrometry (MALDI-MS), as components of apolipoproteins (apo) (apo E2 and E4, apo L, the high Mr apo J and a 28 kDa fragment of apo A-IV). Coincubation of each apolipoprotein with FSGS serum inhibited permeability, but only apo J and apo E2 and E4 were found to be crucial for the process. In conclusion, we have purified from normal serum five inhibitors of permeability induced by FSGS serum, all corresponding to apolipoproteins. An imbalance between permeability factors and apolipoproteins may play a pathogenetic role in FSGS.

Acute effects of simultaneous intraperitoneal infusion of glucose and amino acids.

BACKGROUND: The feasibility of simultaneously infusing glucose and amino acid (AA)-based peritoneal dialysis solutions was tested to determine whether peritoneal dialysis patients could achieve an adequate nonprotein calorie/nitrogen ratio while preventing a marked increase in blood urea nitrogen (BUN), which is usually seen if the AAs are administered without glucose. METHODS: An automatic peritoneal dialysis cycler was used to infuse glucose and AA solutions (3:1) simultaneously during the night. Eight infusions of 1000 mL m2 of body surface area (BSA), with a 60 minute dwell time, were performed in 10 children on peritoneal dialysis. The dialytic effluent was analyzed at every exchange and totaled at eight hours to evaluate volume, glucose, and AA concentration. Blood samples for plasma, glucose, insulin, and free AA determination were drawn at the beginning of automated peritoneal dialysis (APD) session and at each instillation of peritoneal dialysate. RESULTS: The mean glucose absorption was 33.7 +/- 10.0% and the AA absorption was 55.2 +/- 13.2% of the infused amount, and the ratio of nonprotein calorie (derived from glucose) to nitrogen (derived from AA) was 115.4:1. The insulin levels returned to normal only three hours after the beginning of APD. The free AA plasma levels were already increased two hours after dinner and remained high for the entire APD treatment because of the continuous absorption of AA from the peritoneum. The BUN levels did not increase despite the supply of AA. CONCLUSIONS: This APD procedure may improve utilization of AA for protein synthesis, as suggested by the lack of increase of the BUN levels with this regimen.

Down-regulation of lysyl oxidase-induced tumorigenic transformation in NRK-49F cells characterized by constitutive activation of ras proto-oncogene.

Several investigations have suggested a putative tumor suppressor role for lysyl oxidase because it is down-regulated in many human and oncogene-induced tumors. To address this issue we down-regulated the enzyme in normal rat kidney fibroblasts by stable transfection of its cDNA in an antisense orientation. The selected clones revealed an absence of lysyl oxidase and dramatic phenotypic changes, interpretable as signs of transformation. The antisense lysyl oxidase clones showed, indeed, loose attachment to the plate and anchorage-independent growth and were highly tumorigenic in nude mice. Moreover, we found an impaired response of the PDGF and IGF-1 receptors to their ligands. In particular, the transformed cells showed a down-regulation of both PDGF receptors and expressed the 105-kDa isoform of the IGF-1 beta receptor, which was not present in the normal control cells. The lack of response to PDGF-BB has been described as a feature of many ras-transformed phenotypes. Therefore, we looked at the status of the p21(ras). Indeed, we found a significantly higher level of active p21(ras) both during steady-state growth and prolonged starvation. Our data reveal new evidence for a tumor suppressor activity of lysyl oxidase, highlighting its particular role in controlling Ras activation and growth factor dependence.

Branchio-Oto-renal syndrome: a report on nine family groups.

This study reviews nine new families with branchio-oto-renal (BOR) syndrome (Online Mendelian Inheritance in Man [OMIM] 113650). Diagnosis was made by studying 10 index cases, and then 22 other previously undetected patients were diagnosed within the nine families. The syndrome consists of conductive, sensorineural, or mixed hearing loss; preauricular pits; structural defects of the outer, middle, or inner ear; renal anomalies; lateral cervical fistulas, cysts, or sinuses; and/or nasolacrimal duct stenosis or fistulas. In our study, all patients first diagnosed in each familial group were recognized on the basis of severe renal anomalies associated with at least one of these symptoms. Our study showed that BOR syndrome is a misdiagnosed disorder, usually recognized in the presence of severe renal failure but often not diagnosed, especially in the adult in the presence of other isolated clinical signs, such as mild branchial or urological anomalies. We stress the meticulous search we performed for renal anomalies and/or hearing loss in all subjects showing minimal signs of branchial defects. BOR syndrome should be suspected in all cases of isolated urological anomalies, even if no other signs of the syndrome are present. After BOR syndrome has been diagnosed in a patient, all family members should be examined for the presence of the syndrome, even if there are only minimal stigmata of the disease.

Lysyl oxidase activates the transcription activity of human collagene III promoter. Possible involvement of Ku antigen.

Lysyl oxidase is an extracellular enzyme that controls the maturation of collagen and elastin. Lysyl oxidase and collagen III often show similar expression patterns in fibrotic tissues. Therefore, we investigated the influence of lysyl oxidase overexpression on the promoter activity of human COL3A1 gene. Our results showed that when COS-7 cells overexpressed the mature form of lysyl oxidase, the activity of the human COL3A1 promoter was increased up to an average of 12 times when tested by luciferase reporter assay. The effect was specific, because other promoters were not affected. Moreover, lysyl oxidase effect was abolished by beta-aminopropionitrile, a specific inhibitor of its catalytic activity. Electrophoretic mobility shift assay showed a binding activity in the region from -101 to -77 that was significantly increased by lysyl oxidase overexpression. The binding was specifically competed by the cold probe, and the mutagenesis of this region abolished both the binding activity in gel retardation and lysyl oxidase stimulation of COL3A1 promoter in transfection experiments. We identified the binding activity as Ku antigen in its two components: Ku80 and Ku70. This study suggests a new coordinated mechanism by which lysyl oxidase might control the development of fibrosis.

A DNA element in the alpha1 type III collagen promoter mediates a stimulatory response by angiotensin II.

BACKGROUND: Angiotensin II (Ang II) plays an important role in extracellular matrix deposition and tissue scarring in the kidney and the heart. The mechanism for extracellular matrix stimulation by Ang II is currently hypothetical, with one possibility pointing to a direct effect on cell synthesis of specific collagens. METHODS: We studied the molecular mechanism for activation of type III collagen synthesis by Ang II in an in vitro cell model of myofibroblasts by evaluating (1) alpha1(III) collagen mRNA expression; (2) alpha1(III) collagen promoter activity; (3) DNA/protein binding with characterization of binding sites; (4) expression of transcription factors; and (5) the role of a short DNA segment as Ang II responsive element. RESULTS: We found a specific dose-dependent stimulation of alpha1(III) collagen mRNA expression and a parallel effect on alpha1(III) collagen promoter activity. Transfection of constructs containing alpha1(III) collagen promoter fragments of different lengths localized the site of activation within the shortest 178 bp construct. By gel-retardation experiments, we observed the formation of a DNA-protein complex with crude extracts from Ang II-stimulated cells and an oligonucleotide spanning the 3 to 20 sequence. This complex was due to a sequence-specific interaction and was abolished by a 3 bp substitution mutation. The introduction of this mutation into the 178 bp construct abolished the stimulatory effect of Ang II. CONCLUSIONS: These results demonstrate that Ang II stimulates the expression of alpha1(III) collagen mRNA in myofibroblasts in vitro by activating the alpha1(III) collagen promoter at the level of a factor recognition site localized immediately downstream of the transcription start site. This mechanism could be involved in Ang II-induced renal and heart fibrosis.

Clinical and molecular heterogeneity of juvenile nephronophthisis in Italy: insights from molecular screening.

Autosomal recessive nephronophthisis (NPH) is a renal disorder histologically characterized by tubulointerstitial lesions that are, in some cases, associated with extrarenal manifestations such as tapeto-retinal degeneration or liver fibrosis. The disease is usually pauci-symptomatic in an early phase but invariably evolves to end-stage renal failure in childhood or early adulthood. The recent discovery of the NPHP1 gene (nephrocystin) has prompted research into putative genotype-phenotype correlations. We screened a population of 68 Italian children (10 multiplex families, 47 sporadic cases) with a clinical and histopathologic picture of NPH and found a large homozygous deletion at 2q13 involving nephrocystin in 30 cases, and heterozygous deletion associated with new point mutations at exons 15 (Tyr518Ter) and 17 (Arg585Ter) of the gene in two other cases. The remaining 36 children had no apparent molecular defects of nephrocystin. In spite of this genetic heterogeneity, the two groups, with and without detectable molecular defects of nephrocystin, showed similar renal defects and comparable cumulative survival considering the start of dialysis as an end-point. The unique difference observed was a less frequent requirement of dialysis in NPH1 patients with pure renal form. Finally, tapeto-retinal degeneration was associated with renal lesions in seven cases presenting deletion of the nephrocystin gene and in five sporadic cases without molecular defects. These data show that a molecular defect of nephrocystin is involved in approximately 50% of patients with NPH, and another 50% require further molecular characterization. Research therefore should now be aimed at characterizing a new locus. In spite of the molecular heterogeneity, NPH in children presents similar renal and extrarenal manifestations, thus suggesting the involvement of common pathological routes.

Vesico-ureteral reflux in pediatric kidney transplants: clinical relevance to graft and patient outcome.

From June 1985 to December 1998, 173 pediatric renal transplants were carried out in 170 patients at our center. From this pool, 73 patients (34 males and 39 females) with a follow-up of 48 months were examined. In all patients, ureteroneocystostomy was performed according to the Lich-Grégoire procedure. All patients were treated with cyclosporin A (CsA)-based immunosuppression, including prednisone and sometimes azathioprine (AZA). Six months after transplantation, voiding cystography (VCU) was performed in all patients and reflux was classified from Grade I to Grade IV. The patients were divided into two groups: those with reflux (Group A: 25 patients) and those without (Group B: 48 patients). Grade I reflux was found in four patients, Grade II in seven patients, Grade III in seven patients, and Grade IV in seven patients. All the patients with severe reflux (Grade IV) underwent a corrective surgical procedure. Both groups were examined for immunologic and non-immunologic risk factors and no significant differences were found. Analysis of patient and graft survival rates revealed no statistical differences (NS) between Groups A and B. Mean serum creatinine (mg/dL) was 1.06 +/- 0.28 and 1.12 +/- 0.41 at 4 yr in Groups A and B, respectively (NS). Mean calculated creatinine clearance (cCrC; ml/min) was 76.74 +/- 15.92 and 77.96 +/- 15.66 in Groups A and B, respectively (NS). The analysis was further extended by considering the grade of reflux (I to IV). Again, no significant differences in the above parameters emerged between the reflux sub-groups; only in the Grade IV sub-group was a slight decrease in cCrC detected, although this difference was not statistically significant when compared with the other sub-groups. In conclusion, vesico-ureteral reflux (VUR) does not seem to negatively affect graft function. However, as all severe reflux patients (Grade IV) were surgically corrected, no conclusions can be drawn with regard to the influence of Grade IV reflux on long-term graft function.

Disseminated autoimmune disease during levamisole treatment of nephrotic syndrome.

Side effects such as cutaneous vasculitis, which occur during prolonged levamisole treatment, may discourage the utilization of the drug in relapsing nephrotic syndrome. We describe a child who developed disseminated vasculitis during prolonged treatment with levamisole. The acute phase was characterized by hepatosplenomegaly, hemolytic anemia, IgM anticardiolipin and p-antineutrophil cytoplasmic antibodies. One month after withdrawal of therapy all symptoms had disappeared and tests normalized. This case report, together with other reports on cutaneous vasculitis, suggest caution and close monitoring during prolonged levamisole therapy.

Correlation between angiotensin-converting enzyme gene insertion/deletion polymorphism and kidney graft long-term outcome in pediatric recipients: a single-center analysis.

BACKGROUND: Despite numerous advances in the areas of organ preservation, histocompatibility, and immunosuppression, chronic deterioration of organ allograft function, referred to as "chronic rejection," still remains the main obstacle to long-term graft survival. The common feature of chronic rejection is a concentric generalized graft arteriosclerosis associated with interstitial fibrosis that reflects an allogeneic injury to graft arteries, possibly worsened by other alloantigen-independent risk factors. The presence of the angiotensin I-converting enzyme (ACE) gene-deleted (D) allele has been associated, when in homozygosity, with increased risk of cardiovascular diseases and with an accelerated progression of organ damage in a variety of kidney diseases. In this study, we analyzed whether the insertion/deletion polymorphism of the ACE gene, because of its negative prognostic impact on cardiovascular and renal pathology, could have any influence on kidney graft survival in pediatric recipients. METHODS: DNA was isolated from peripheral blood mononuclear cells from 146 pediatric dialysis patients (mean age: 12.9 years) who received a first kidney graft at our center between December 1985 and July 1997. To rule out any bias due to acute graft losses, only 119 patients who reached a minimum of 12 months of graft survival were considered for statistical analysis. The insertion/deletion polymorphism of the ACE gene was detected using a polymerase chain reaction technique with two flanking primers. RESULTS: The results demonstrated that (i) the distribution of DD and non-DD (ID + II) genotypes was 36.1% (43 patients) and 63.8% (76 patients), respectively; (ii) actuarial graft survival at 7, 8, 9, and 10 years in patients with non-DD genotype was significantly higher than that in patients with DD genotype (7 years: 94.6% vs. 72.4%, P<0.05; 8 years: 94.6% vs. 62%, P<0.025; 9 years: 87.3% vs. 51.4%, P<0.025; 10 years: 76.3% vs. 25.7%, P<0.01). CONCLUSIONS: In conclusion, the above data indicate that DD genotype is associated in pediatric kidney graft recipients with a shorter long-term kidney graft survival and suggest a possible role of this genotype as a cofactor in the progression of nonimmunological injuries leading to chronic kidney graft failure.

Peritoneal mesothelial cells produce complement factors and express CD59 that inhibits C5b-9-mediated cell lysis.

The CD59 membrane protein confers protection from C5b-9-mediated cell lysis. Because evidence exists for complement (C) activation and generation of C5b-9 in the peritoneal cavity during chronic peritoneal dialysis (CPD), we investigated, on mesothelial cell (MC) lines, the expression of CD59 and the production of C components. Four MC lines were obtained from children on CPD, and two from non uremic children. CD59 expression on MCs was investigated with anti-CD59 monoclonal antibody (mAb) and polyclonal goat immunoglobulin G (IgG). MC lines were positive for staining with anti-CD59 mAb. Western blotting analysis of MC membrane demonstrated a band with the same molecular weight as CD59. Incubation of MC with anti-CD59 mAb abrogated the protective effect of CD59 (100% cytotoxicity). C3, C4, and C6 were detected in the supernatants of MC; in non uremic MC supernatants, C5, C7, C8, and C9 were also detectable, and C4 concentration was tenfold higher. CD59 expression confers to MCs protection from C5b-9-mediated lysis. MCs produce C factors. These findings suggest that production of complement components and expression of CD59 on MCs could play a role both in peritoneal cavity infection (decreased complement production) and in peritoneal membrane damage (decreased CD59 expression and reduced remesothelialization owing to MC lysis).

[Critical evaluation of growth hormone treatment in children with chronic renal insufficiency]. / Evaluation critique du traitement par l'hormone de croissance chez l'enfant atteint d'insuffisance rénale chronique.

Growth failure remains a major complication of chronic renal insufficiency in children, which greatly affects their quality of life. Based upon the data published in the literature it appears that growth hormone therapy improves growth in these children, with little secondary effects. However, some uncertainties persist concerning the safety and efficacy of this therapy, so that careful evaluations must continue.

Renal-retinal syndromes: association of retinal anomalies and recessive nephronophthisis in patients with homozygous deletion of the NPH1 locus.

Tapeto-retinal degeneration is frequent in patients with nephronophthisis. Association of the most severe forms of tapeto-retinal dystrophy with NPH identifies a syndrome described first by Senior et al and Loken et al. This syndrome is distinct on molecular grounds from pure renal nephronophthisis (NPH1), which has its gene locus mapped on chromosome 2q13. We describe three families with large homozygous deletion of the NPH1 locus in which mild to moderate ocular lesions due to tapeto-retinal degeneration coexisted and were correlated to renal defects. This new association of NPH1 with retinal dystrophy is characterized by focal lesions of retina and is pauci-symptomatic in clinical presentation. For this reason it may remain unrecognized in most NPH1 patients.

Chronic renal replacement therapy in children: which index is best for adequacy?

BACKGROUND: The dialysis dose, Kt/V, and Solute Removal Index (SRI) have been proposed as tools to measure and compare adequacy of different renal replacement therapies in adults. The aim of our study was to elucidate whether the Kt/V and SRI could be appropriate parameters to compare different treatments and define adequacy targets in children. METHODS: Twenty-two pediatric chronic dialysis patients (2 to 17 years) were prospectively studied. Six patients were on continuous ambulatory peritoneal dialysis (CAPD), 7 patients were on automatic nightly peritoneal dialysis (ANPD), and 9 were on hemodialysis (HD). Patients had no peritonitis and were not hospitalized during the previous two months and, as proved by growth and subjective well being, were in steady state condition at the initiation of the protocol. As a consequence, the treatment delivered was assumed to be adequate and the prospective analysis was carried out within one month. Urea levels in dialysate, plasma and urine were measured to determine urea kinetics and measure adequacy parameters. RESULTS: Instantaneous urea clearance was much higher when hemodialysis was used (124.67 +/- 32.04 ml/min) compared to CAPD (2.79 +/- 0.29 ml/min) and ANPD (6.60 +/- 1.42 ml/min), as expected. The Urea dialytic clearance per week was greater in HD (67320 +/- 17299 ml) than in CAPD(28144 +/- 2895 ml) and ANPD (29910 +/- 4234 ml). Residual renal function contributed to the overall weekly clearance by 47% in CAPD, while it was only by 19% in HD and 26% in ANPD. The overall weekly clearance was therefore 79,842 ml/week in HD, 53,340 ml/week in CAPD and 41,012 ml/week in ANPD. Weekly dialytic Kt/V results were much higher in HD (3.75) than in CAPD (1.78) and ANPD (2.37). To these values, the renal Kt/V was added, reaching the values of overall (dialytic + renal) weekly Kt/V of 4.53 in HD, 3.41 in CAPD and 3.41 in ANPD. Although higher Kt/V values were observed in HD, when the SRI % was considered, HD appeared to be less efficient compared with the other two techniques. Since postdialytic rebound in HD patients averaged 22.5%, we may speculate that hemodialysis in children is less efficient than continuous or daily peritoneal dialysis because of a remarkable cardipulmonary recirculation and solute sequestration. CONCLUSION: In the global evaluation, dialysis SRI% appears to be more reliable as an index of adequacy compared to Kt/V in children. At least an integration between the two indices is strongly recommended.