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.
The discovery of different podocyte autoantibodies in membranous nephropathy (MN) raises questions about their pathogenetic and clinical meaning. This study sought to define antibody isotypes and ...correlations; to compare levels in MN, other glomerulonephritides, and controls; and to determine their association with clinical outcomes.
Serum IgG(1), IgG(3), and IgG(4) against aldose reductase (AR), SOD2, and α-enolase (αENO) were measured at diagnosis in 186 consecutive MN patients, in 96 proteinuric controls (36 with FSGS, and 60 with IgA nephropathy), and in 92 healthy people recruited in four Italian nephrology units. Anti-phospholipase A2 receptor (PLA2r) and anti-neutral endopeptidase (NEP) IgG(4) were titrated in the same specimens. Association with 1-year follow-up clinical parameters was studied in 120 patients.
IgG(4) was the most common isotype for all antibodies; IgG(1) and IgG(3) were nearly negligible. IgG(4) levels were positive in a significant proportion of MN patients (AR, 34%; SOD2, 28%; αENO, 43%). Antibody titers were higher in MN than in healthy and pathologic controls (P<0.005). Anti-NEP IgG(4) did not differ from normal controls (P=0.12). Anti-PLA2r IgG(4) was detected in 60% of patients and correlated with anti-AR, anti-SOD2, and anti-αENO IgG(4) (P<0.001). In MN patients negative for the whole antibody panel (20%), 1-year proteinuria was lower compared with patients with at least one antibody positivity (P<0.05).
Our data suggest that IgG(4) is the prevalent isotype for antibodies against cytoplasmic antigens of podocytes (AR, SOD2, αENO). Their levels were higher than in other proteinuric glomerulonephritides and in normal controls and were correlated with anti-PLA2r. Only baseline negativity for all known antibodies predicted lower 1-year proteinuria.
The identification of glomerular auto-antigens in idiopathic human membranous glomerulonephritis (MGN) is a crucial step towards the definition of the mechanisms of the disease. Recent ‘in vivo’ ...studies demonstrated a heterogeneous composition of glomerular immune-deposits in MGN biopsies only a part of which have been characterized.
We studied with a proteomical approach IgGs eluted from laser capture microdissected glomeruli of 8 MGN patients and showed the existence of other three immune proteins in MGN glomeruli (α-enolase, elongation factor 2 and Glycyl Aminoacyl-tRNA Synthetase). One of these, i.e. α-enolase, fulfilled all criteria for being considered an auto-antigen. Specific IgG1 and IgG4 reacting with podocyte α-enolase were, in fact, eluted from microdissected glomeruli and Confocal- and Immuno Electron-Microscopy showed co-localization of α-enolase with IgG4 and C5b-9 in immune-deposits. Serum levels of anti a-enolase IgG4 were determined in 131 MGN patients and were found elevated in 25% of cases.
Overall, our data demonstrate that glomerular α-enolase is a target antigen of autoimmunity in human MGN. Circulating anti α-enolase auto-antibodies can be detected in sera of a significant quota of MGN patients. Like other auto-antigens, α-enolase may be implicated in the pathogenesis of human MGN.
Direct characterization of target podocyte antigens and autoantibodies in human membranous glomerulonephritis. Alpha-enolase and borderline antigens. Display omitted
► We identified α-enolase as new auto-antigens in membranous glomerulonephritis (MGN). ► We studied a-enolase using a proteomics approach. ► This new auto-antigen was validated in 131 MGN patients. > α-enolase may be implicated in the pathogenesis of human MGN.
In patients with IgA nephropathy (IgAN), circulatory IgA1 and IgA1 in mesangial deposits contain elevated amounts of galactose-deficient IgA1 (Gd-IgA1). We hypothesized that a fraction of Gd-IgA1 ...from the glomerular deposits and/or circulation may be excreted into the urine and thus represent a disease-specific biomarker. Levels of urinary IgA and Gd-IgA1 were determined in 207 patients with IgAN, 205 patients with other renal diseases, and 57 healthy controls, recruited in USA, Japan, and Italy. Urinary IgA was similarly elevated in patients with IgAN and renal-disease controls compared with healthy controls. However, urinary Gd-IgA1 levels were higher in patients with IgAN (IgAN, 28.0 ± 17.9 ; disease controls, 20.6 ± 17.4 units/mg urinary creatinine; P < 0.0001 ). Lectin western blotting data confirmed these results. In IgAN patients, levels of urinary Gd-IgA1 correlated with proteinuria ( P < 0.001 ). When we purified IgA from serum and urine of an IgAN patient, the relative proportion of Gd-IgA1 to total IgA1 was higher in the urine compared with serum, suggesting selective excretion of Gd-IgA1 in IgAN. In summary, urinary excretion of Gd-IgA1 was elevated in patients with IgAN and the urinary Gd-IgA1 levels correlated with proteinuria. Urinary Gd-IgA1 may thus represent a disease-specific biomarker of IgAN.
Nephrosis and a rapid decline in kidney function characterize HIV-associated nephropathy (HIVAN). Histologically, HIVAN is a collapsing focal segmental glomerulosclerosis with prominent tubular ...damage. We explored the expression of neutrophil gelatinase-associated lipocalin (NGAL), a marker of tubular injury, to determine whether this protein has the potential to aid in the noninvasive diagnosis of HIVAN. We found that expression of urinary NGAL was much higher in patients with biopsy-proven HIVAN than in HIV-positive and HIV-negative patients with other forms of chronic kidney disease. In the HIV-transgenic mouse model of HIVAN, NGAL mRNA was abundant in dilated, microcystic segments of the nephron. In contrast, urinary NGAL did not correlate with proteinuria in human or in mouse models. These data show that marked upregulation of NGAL accompanies HIVAN and support further study of uNGAL levels in large cohorts to aid in the noninvasive diagnosis of HIVAN and screen for HIVAN-related tubular damage.
Mutations in nephrin (NPHS1) and podocin (NPHS2) genes represent a major cause of idiopathic nephrotic syndrome (NS) in children. It is not yet clear whether the presence of a single mutation acts as ...a modifier of the clinical course of NS.
We reviewed the clinical features of 40 patients with NS associated with heterozygous mutations or variants in NPHS1 (n = 7) or NPHS2 (n = 33). Long-term renal survival probabilities were compared with those of a concurrent cohort with idiopathic NS.
Patients with a single mutation in NPHS1 received a diagnosis before those with potentially nongenetic NS and had a good response to therapies. Renal function was normal in all cases. For NPHS2, six patients had single heterozygous mutations, six had a p.P20L variant, and 21 had a p.R229Q variant. Age at diagnosis and the response to drugs were comparable in all NS subgroups. Overall, they had similar renal survival probabilities as non-NPHS1/NPHS2 cases (log-rank chi(2) 0.84, P = 0.656) that decreased in presence of resistance to therapy (P < 0.001) and in cases with renal lesions of glomerulosclerosis and IgM deposition (P < 0.001). Cox regression confirmed that the only significant predictor of dialysis was resistance to therapy.
Our data indicate that single mutation or variant in NPHS1 and NPHS2 does not modify the outcome of primary NS. These patients should be treated following consolidated schemes and have good chances for a good long-term outcome.
Neutrophil extracellular traps (NET) expose modified antigens for autoantibodies in vasculitis. Little is known about levels and removal pathways of NET in systemic lupus erythematosus (SLE), ...especially in lupus nephritis (LN). We determined circulating levels and defined NET removal in large subsets of patients with incident SLE (iSLE), some of whom had new-onset nephritis.
Serum levels of NET (ELISA), DNase1/DNase1L3 (ELISA), and DNase activity (functional assay) were determined in 216 patients with iSLE 103 had incident LN (iLN), in 50 patients with other primary glomerulonephritis, and in healthy controls.
NET production by neutrophils purified from a random selection of patients was quantified as elastase/DNA release and by immunofluorescence techniques.
Serum NET levels were very high in iSLE/iLN compared to all groups of controls and correlated with anti-dsDNA, C3-C4, and proteinuria; iLN had the highest levels. DNase activity was decreased in iLN compared to SLE (20% had one-half DNase activity) despite similar serum levels of DNase1/DNase1L3. In these cases, pretreatment of serum with protein A restored DNase efficiency; 1 patient was homozygous for a c.289_290delAC variant of
.
NET production by neutrophils purified from LN, SLE, and normal controls was similar in all cases.
Patients with iLN have increased circulating NET and reduced DNase activity, the latter being explained by the presence of inhibitory substances in circulation and/or by rare
mutations. Accumulation of NET derives from a multifactorial mechanism, and is associated and may contribute to disease severity in SLE, in particular to renal lesions. (Clinical trial registration: The Zeus study was registered at ClinicalTrials.gov, study number NCT02403115).
Genetic Drivers of Kidney Defects in the DiGeorge Syndrome Lopez-Rivera, Esther; Liu, Yangfan P; Verbitsky, Miguel ...
New England journal of medicine/The New England journal of medicine,
02/2017, Letnik:
376, Številka:
8
Journal Article
Recenzirano
Odprti dostop
The DiGeorge syndrome, the most common of the microdeletion syndromes, affects multiple organs, including the heart, the nervous system, and the kidney. It is caused by deletions on chromosome ...22q11.2; the genetic driver of the kidney defects is unknown.
We conducted a genomewide search for structural variants in two cohorts: 2080 patients with congenital kidney and urinary tract anomalies and 22,094 controls. We performed exome and targeted resequencing in samples obtained from 586 additional patients with congenital kidney anomalies. We also carried out functional studies using zebrafish and mice.
We identified heterozygous deletions of 22q11.2 in 1.1% of the patients with congenital kidney anomalies and in 0.01% of population controls (odds ratio, 81.5; P=4.5×10
). We localized the main drivers of renal disease in the DiGeorge syndrome to a 370-kb region containing nine genes. In zebrafish embryos, an induced loss of function in snap29, aifm3, and crkl resulted in renal defects; the loss of crkl alone was sufficient to induce defects. Five of 586 patients with congenital urinary anomalies had newly identified, heterozygous protein-altering variants, including a premature termination codon, in CRKL. The inactivation of Crkl in the mouse model induced developmental defects similar to those observed in patients with congenital urinary anomalies.
We identified a recurrent 370-kb deletion at the 22q11.2 locus as a driver of kidney defects in the DiGeorge syndrome and in sporadic congenital kidney and urinary tract anomalies. Of the nine genes at this locus, SNAP29, AIFM3, and CRKL appear to be critical to the phenotype, with haploinsufficiency of CRKL emerging as the main genetic driver. (Funded by the National Institutes of Health and others.).
Congenital Anomalies of the Kidney and Urinary Tract (CAKUT) are a major cause of morbidity in children. We measured the risk of progression to end-stage renal disease in 312 patients with CAKUT ...preselected for the presence of anomalies in kidney number or size. A model of dialysis-free survival from birth was established as a function of the renal CAKUT categories of solitary kidney; unilateral and bilateral hypodysplasia; renal hypodysplasia associated with posterior urethral valves; and multicystic and horseshoe kidney. Cox regression analysis took into account the concomitant presence of vesicoureteral reflux, year of diagnosis, and time-varying values of serum creatinine, proteinuria, and hypertension. By 30 years of age, 58 patients had started dialysis, giving a yearly incidence of 0.023 over a combined 2474 patient risk years. The risk for dialysis was significantly higher for patients with a solitary kidney or with renal hypodysplasia associated with posterior urethral valves (hazard ratios of 2.43 and 5.1, respectively) compared to patients with unilateral or bilateral renal hypodysplasia, or multicystic or horseshoe kidney, and was independent of other prognostic factors. Our study shows that sub-clinical defects of the solitary kidney may be responsible for a poorer prognosis compared to more benign forms of CAKUT. Prospective studies are needed to validate these results.
PDF
