Membranous nephropathy is characterized by deposition of immune complexes along the glomerular basement membrane. PLA2R and THSD7A are target antigens in 70% and 1-5% of primary membranous ...nephropathy cases, respectively. In the remaining cases, the target antigen is unknown. Here, laser microdissection of glomeruli followed by mass spectrometry was used to identify novel antigen(s) in PLA2R-negative membranous nephropathy. An initial pilot mass spectrometry study in 35 cases of PLA2R-negative membranous nephropathy showed high spectral counts for neural tissue encoding protein with EGF-like repeats, NELL-1, in six cases. Mass spectrometry failed to detect NELL-1 in 23 PLA2R-associated membranous nephropathy and 88 controls. NELL-1 was localized by immunohistochemistry, which showed bright granular glomerular basement membrane staining for NELL-1 in all six cases. Next, an additional 23 NELL-1 positive cases of membranous nephropathy were identified by immunohistochemistry in a discovery cohort of 91 PLA2R-negative membranous nephropathy cases, 14 were confirmed by mass spectrometry. Thus, 29 of 126 PLA2R-negative cases were positive for NELL-1. PLA2R-associated membranous nephropathy and controls stained negative for NELL-1. We then identified five NELL-1 positive cases of membranous nephropathy out of 84 PLA2R and THSD7A-negative cases in two validation cohorts from France and Belgium. By confocal microscopy, both IgG and NELL-1 co-localized to the glomerular basement membrane. Western blot analysis showed reactivity to NELL-1 in five available sera, but no reactivity in control sera. Clinical and biopsy findings of NELL-1 positive membranous nephropathy showed features of primary membranous nephropathy. Thus, a subset of membranous nephropathy is associated with accumulation and co-localization of NELL-1 and IgG along the glomerular basement membrane, and with anti-NELL-1 antibodies in the serum. Hence, NELL-1 defines a distinct type of primary membranous nephropathy.
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Background & Aims The effects of bariatric surgery in patients with nonalcoholic fatty liver disease (NASH) are not well established. We performed a prospective study to determine the biological and ...clinical effects of bariatric surgery in patients with NASH. Methods From May 1994 through May 2013, one hundred and nine morbidly obese patients with biopsy-proven NASH underwent bariatric surgery at the University Hospital of Lille, France (the Lille Bariatric Cohort). Clinical, biological, and histologic data were collected before and 1 year after surgery. Results One year after surgery, NASH had disappeared from 85% of the patients (95% confidence interval CI: 75.8%−92.2%). Compared with before surgery, patients had significant reductions in mean ± SD body mass index (BMI, from 49.3 ± 8.2 to 37.4 ± 7) and level of alanine aminotransferase (from 52.1 ± 25.7 IU/L to 25.1 ± 20 IU/L); mean levels of γ-glutamyltransferases were reduced from 51 IU/L before surgery (interquartile range IQR, 34−87 IU/L) to 23 IU/L afterward (IQR, 14−33 IU/L) and mean insulin resistance index values were reduced from 3.6 ± 0.5 to 2.9 ± 0.5 ( P < .01 for each comparison). NASH disappeared from a higher proportion of patients with mild NASH before surgery (94%) than severe NASH (70%) ( P < .05) according to Brunt score. In histologic analysis, steatosis was detected in 60% of the tissue before surgery (IQR, 40%−80%) but only 10% 1 year after surgery (IQR, 2.5%−21.3%); the mean nonalcoholic fatty liver disease score was reduced from 5 (IQR, 4−5) to 1 (IQR, 1−2) (each P < .001). Hepatocellular ballooning was reduced in 84.2% of samples (n = 69; 95% CI: 74.4−91.3) and lobular inflammation in 67.1% (n = 55; 95% CI: 55.8−77.1). According to Metavir scores, fibrosis was reduced in 33.8% of patients (95% CI: 23.6%−45.2%). Patients whose NASH persisted 1 year after surgery (n = 12) had lost significantly less weight (change in BMI, 9.1 ± 1.5) than those without NASH (change in BMI, 12.3 ± 0.6) ( P = .005). Patients who underwent laparoscopic gastric banding lost less weight (change in BMI, 6.4 ± 0.7) than those who underwent gastric bypass (change in BMI, 14.0 ± 0.5) ( P < .0001), and a higher proportion had persistent NASH (30.4% vs 7.6% of those with gastric bypass; P = .015). Conclusions Bariatric surgery induced the disappearance of NASH from nearly 85% of patients and reduced the pathologic features of the disease after 1 year of follow-up. It could be a therapeutic option for appropriate morbidly obese patients with NASH who do not respond to lifestyle modifications. More studies are needed to determine the long-term effects of bariatric surgery in morbidly obese patients with NASH.
Membranous nephropathy results from subepithelial antigen-antibody complex deposition along the glomerular basement membrane. Although PLA2R, THSD7A, and NELL-1 account for a majority (about 80%) of ...the target antigens, the target antigen in the remaining cases is not known. Using laser microdissection of PLA2R-negative glomeruli of patients with membranous nephropathy followed by mass spectrometry we identified a unique protein, Semaphorin 3B, in three cases. Mass spectrometry failed to detect Semaphorin-3B in 23 PLA2R-associated cases of membranous nephropathy and 88 controls. Semaphorin 3B in all three cases was localized to granular deposits along the glomerular basement membrane by immunohistochemistry. Next, an additional eight cases of Semaphorin 3B-associated membranous nephropathy were identified in three validation cohorts by immunofluorescence microscopy. In four of 11 cases, kidney biopsy also showed tubular basement membrane deposits of IgG on frozen sections. Confocal microscopy showed that both IgG and Semaphorin 3B co-localized to the glomerular basement membrane. Western blot analysis of five available sera showed reactivity to reduced Semaphorin 3B in four of four patients with active disease and no reactivity in one patient in clinical remission; there was also no reactivity in control sera. Eight of the 11 cases of Semaphorin 3B-associated membranous nephropathy were pediatric cases. Furthermore, in five cases, the disease started at or below the age of two. Thus, Semaphorin 3B-associated membranous nephropathy appears to be a distinct type of disease; more likely to be present in pediatric patients.
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In alcoholic hepatitis (AH), development of targeted therapies is crucial and requires improved knowledge of cellular and molecular drivers in liver dysfunction. The unique opportunity of using ...explanted livers from patients with AH having undergone salvage liver transplantation allowed to perform more in-depth molecular translational studies.
We studied liver explants from patients with AH submitted to salvage transplantation (n=16), from patients with alcoholic cirrhosis without AH (n=12) and fragments of normal livers (n=16). Hepatic cytokine content was quantified. Hepatocyte function and proliferation and the presence of hepatic progenitor cells (HPCs) were evaluated by immunohistochemistry, western blot or quantitative PCR. Mitochondrial morphology was evaluated by electron microscopy.
Livers from patients with AH showed decreased cytokine levels involved in liver regeneration (tumour necrosis factor α and interleukin-6), as well as a virtual absence of markers of hepatocyte proliferation compared with alcoholic cirrhosis and normal livers. Electron microscopy revealed obvious mitochondrial abnormalities in AH hepatocytes. Importantly, livers from patients with AH showed substantial accumulation of HPCs that, unexpectedly, differentiate only into biliary cells. AH livers predominantly express laminin (extracellular matrix protein favouring cholangiocyte differentiation); consequently, HPC expansion is inefficient at yielding mature hepatocytes.
AH not responding to medical therapy is associated with lack of expression of cytokines involved in liver regeneration and profound mitochondrial damage along with lack of proliferative hepatocytes. Expansion of HPCs is inefficient to yield mature hepatocytes. Manoeuvres aimed at promoting differentiation of HPCs into mature hepatocytes should be tested in AH.
Background & Aims Severe obesity is implicated in development of nonalcoholic fatty liver disease (NAFLD). Bariatric surgery induces weight loss and increases survival time of obese patients, but ...little is known about its effects on liver damage. We performed a 5-year prospective study to evaluate fibrosis and nonalcoholic steatosis (NASH) in severely obese patients after bariatric surgery. Methods Bariatric surgery was performed on 381 patients. Clinical and biological data, along with liver biopsies, were collected before and at 1 and 5 years after surgery. Results Five years after surgery, levels of fibrosis increased significantly, but 95.7% of patients maintained a fibrosis score ≤ F1. The percentage of patients with steatosis decreased from 37.4% before surgery to 16%, the NAFLD score from 1.97 to 1, ballooning from 0.2 to 0.1. Inflammation remained unchanged. The percentage of patients with probable or definite NASH decreased significantly over 5 years, from 27.4% to 14.2%. The kinetics of insulin resistance (IR) paralleled that of steatosis and ballooning; the greatest improvements occurred within the first year and were sustained 5 years later. Steatosis and ballooning occurred more frequently in patients with a refractory IR profile. In multivariate analysis, the refractory IR profile independently predicted the persistence of steatosis and ballooning 5 years later. Conclusions Five years after bariatric surgery for severe obesity, almost all patients had low levels of NAFLD, whereas fibrosis slightly increased. Steatosis and ballooning were closely linked to IR; long-term effects could be predicted by early improvement in IR.
MicroRNAs (miRNAs) are a class of noncoding RNA acting at a post-transcriptional level to control the expression of large sets of target mRNAs. While there is evidence that miRNAs deregulation plays ...a causative role in various complex disorders, their role in fibrotic kidney diseases is largely unexplored. Here, we found a strong up-regulation of miR-21 in the kidneys of mice with unilateral ureteral obstruction and also in the kidneys of patients with severe kidney fibrosis. In addition, mouse primary fibroblasts derived from fibrotic kidneys exhibited higher miR-21 expression level compared to those derived from normal kidneys. Expression of miR-21 in normal primary kidney fibroblasts was induced upon TGFβ exposure, a key growth factor involved in fibrogenesis. Finally, ectopic expression of miR-21 in primary kidney fibroblasts was sufficient to promote myofibroblast differentiation. As circulating miRNAs have been suggested as promising non-invasive biomarkers, we further assess whether circulating miR-21 levels are associated with renal fibrosis using sera from 42 renal transplant recipients, categorized according to their renal fibrosis severity, evaluated on allograft biopsies (Interstitial Fibrosis/Tubular Atrophy (IF/TA). Circulating miR-21 levels are significantly increased in patients with severe IF/TA grade (IF/TA grade 3: 3.0±1.0 vs lower grade of fibrosis: 1.5±1.2; p = 0.001). By contrast, circulating miR-21 levels were not correlated with other renal histological lesions. In a multivariate linear regression model including IF/TA grade and estimated GFR, independent associations were found between circulating miR-21 levels and IF/TA score (ß = 0.307, p = 0.03), and between miR-21 levels and aMDRD (ß = -0.398, p = 0.006). Altogether, these data suggest miR-21 has a key pathogenic role in kidney fibrosis and may represent a novel, predictive and reliable blood marker of kidney fibrosis.
Primary hyperparathyroidism (PHPT) is commonly diagnosed in the setting of hypercalcemia, whereas normocalcemic primary hyperparathyroidism (NHPT) may be misdiagnosed.
Our objective was to compare ...hypercalcemic hyperparathyroidism (HPHPT) versus NHPT hypercalciuric renal stone patients.
We took advantage of a routine calcium load test performed in hypercalciuric renal stone patients to assess retrospectively among PHPT patients, prevalence and characteristics of NHPT and HPHPT under a calcium restricted diet.
Among 1671 hypercalciuric patients included, 91 patients have a final diagnosis of PHPT(post load ionized calcium (iCa)>1.31 mmol/L and PTH>30 pg/ml). Prevalence of NHPT is 40% of all PHPT, however according to total serum calcium 4/35 NHPT and 7/56 HPHPT would have been misclassified in the other group. 18/35 NHPT and 40/56 HPHPT underwent parathyroidectomy. No significant characteristics related to parathyroid weight, stone composition or bone remodeling biomarkers is detected between groups. Whereas iCa is higher in HPHPT in fasting state and after calcium load, we found no difference for calcium diet, 24-hour calciuria, or calcitriol. Of notice, renal calcium excretion (FECa) post load increases by 303% in NHPT but only 176% in HPHPT (p=0.01) likely explained by a lesser PTH decrease (p=0.02). However, a strong negative association (p<0.0001) detected between pooled pre and post load iCa and PTH only in NHPT group suggests a persistent efficient PTH-CaSR control within parathyroid glands in this group.
Our data show the relevance of dynamic tests to unmask NHPT in hypercalciuric renal stone patients.
Thrombotic microangiopathy (TMA) is a poorly recognized cause of collapsing glomerulopathy. The frequency and significance of collapsing glomerulopathy associated with renal TMA have not been ...specifically studied in native kidney biopsy specimens. Here we retrospectively documented clinicopathologic features of 53 patients with histologically proven TMA in the native kidney, with special emphasis on changes due to focal segmental glomerulosclerosis (FSGS). Histological TMA was related to hypertensive nephropathy in 21 patients, genetic complement abnormalities in 9, drugs in 7, and to other causes in 16 patients. Almost half (26 patients) presented with arteriolar, 6 with glomerular, and 21 with mixed TMA. Using the Columbia classification system for the 53 patients with histological TMA, 33 had concurrent FSGS lesions with collapsing glomerulopathy the dominant variant in 19 patients (58% of the FSGS cases), not otherwise specified in 9 patients, cellular in 3, and perihilar or tip lesions in 1 patient each. The presence of FSGS was associated with a poor renal prognosis, with no prognostic difference between collapsing glomerulopathy and other FSGS variants. Thus, collapsing glomerulopathy is frequently found in native kidney biopsies with TMA, suggesting that endothelial injury may play an important role in the pathophysiology of FSGS.
Cardiorenal syndromes type 1 and 2 are complex disorders in which cardiac dysfunction leads to kidney dysfunction. However, the mechanisms remain incompletely explained, during pulmonary hypertension ...in particular. The objective of this study is to develop an original preclinical model of cardiorenal syndrome secondary to a pulmonary hypertension in piglets. Twelve 2-month-old Large White piglets were randomized in two groups: (1) induction of pulmonary hypertension by ligation of the left pulmonary artery and iterative embolizations of the right lower pulmonary artery, or (2) Sham interventions. We evaluated the cardiac function using right heart catheterization, echocardiography and measurement of biochemistry markers). Kidney was characterized using laboratory blood and urine tests, histological evaluation, immunostainings for renal damage and repair, and a longitudinal weekly assessment of the glomerular filtration rate using creatinine-based estimation and intravenous injection of an exogenous tracer on one piglet. At the end of the protocol (6 weeks), the mean pulmonary artery pressure (32 ± 10 vs. 13 ± 2 mmHg; p = 0.001), pulmonary vascular resistance (9.3 ± 4.7 vs. 2.5 ± 0.4 WU; p = 0.004) and central venous pressure were significantly higher in the pulmonary hypertension group while the cardiac index was not different. Piglets with pulmonary hypertension had higher troponin I. We found significant tubular damage and an increase in albuminuria in the pulmonary hypertension group and negative correlation between pulmonary hypertension and renal function. We report here the first porcine model of cardiorenal syndrome secondary to pulmonary hypertension.
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