Electron microscopy study of 496 cases of lupus nephritis: A single-center experience.

INTRODUCTION: Renal involvement is seen in about 40-82% of systemic lupus erythematosus (SLE) Asian patients. The exact diagnosis and classification of lupus nephritis are important for treatment and prognosis. This study aimed to investigate the value of electron microscopy (EM) in the diagnosis and classification of lupus nephritis compared with light microscopy. METHOD: In this cross-sectional referral-center 16-year study of lupus nephritis, the final diagnosis was based on the EM study. Primary light microscopy findings were compared with EM diagnosis. Moreover, Immunofluorescence patterns distribution was assessed. RESULTS: From 496 patients diagnosed with lupus nephritis based on EM, 225(45.4%) of patients were categorized in class IV, followed by 98(19.7%), 93(18.8%), 46(9.3%), and 14(2.8%) who were categorized into classes of II, III, V, and VI respectively. Only 1(0.2%) patient belonged to class I, and 19(3.8%) cases were diagnosed with mixed two classes. Using EM was essential for diagnosing 25.6% of cases taking the correct classification by light microscopy into account; however, disregarding correct classification, this could change to a 7.4% contribution rate of EM. The most common cause of misdiagnosis, disregarding incorrect classification, was inadequate or wrong tissue. Positive associations were detected between tubular atrophy and interstitial fibrosis of both electron and light microscopy with different classes (P < 0.001). CONCLUSION: While light microscopy is highly accurate for diagnosing lupus nephritis regardless of correct classification, EM contributes substantially to the correct classification of lupus nephritis types.

A new view of macula densa cell microanatomy.

Although macula densa (MD) cells are chief regulatory cells in the nephron with unique microanatomical features, they have been difficult to study in full detail due to their inaccessibility and limitations in earlier microscopy techniques. The present study used a new mouse model with a comprehensive imaging approach to visualize so far unexplored microanatomical features of MD cells, their regulation, and functional relevance. MD-GFP mice with conditional and partial induction of green fluorescent protein (GFP) expression, which specifically and intensely illuminated only single MD cells, were used with fluorescence microscopy of fixed tissue and live MD cells in vitro and in vivo with complementary electron microscopy of the rat, rabbit, and human kidney. An elaborate network of major and minor cell processes, here named maculapodia, were found at the cell base, projecting toward other MD cells and the glomerular vascular pole. The extent of maculapodia showed upregulation by low dietary salt intake and the female sex. Time-lapse imaging of maculapodia revealed highly dynamic features including rapid outgrowth and an extensive vesicular transport system. Electron microscopy of rat, rabbit, and human kidneys and three-dimensional volume reconstruction in optically cleared whole-mount MD-GFP mouse kidneys further confirmed the presence and projections of maculapodia into the extraglomerular mesangium and afferent and efferent arterioles. The newly identified dynamic and secretory features of MD cells suggest the presence of novel functional and molecular pathways of cell-to-cell communication in the juxtaglomerular apparatus between MD cells and between MD and other target cells.NEW & NOTEWORTHY This study illuminated a physiologically regulated dense network of basal cell major and minor processes (maculapodia) in macula densa (MD) cells. The newly identified dynamic and secretory features of these microanatomical structures suggest the presence of novel functional and molecular pathways of cell-to-cell communication in the juxtaglomerular apparatus between MD and other target cells. Detailed characterization of the function and molecular details of MD cell intercellular communications and their role in physiology and disease warrant further studies.

Bevacizumab-associated glomerular microangiopathy that occurred after postoperative chemotherapy for ovarian cancer.

Bevacizumab is a monoclonal antibody against vascular endothelial growth factor (VEGF) that is used to treat patients with various cancers. However, it is known to be associated with adverse events, such as hypertension and proteinuria. The histology of bevacizumab-induced nephropathy is known as thrombotic microangiopathy or minimal change nephrotic syndrome. Recently, however, the terms "bevacizumab-associated glomerular microangiopathy" and "anti-VEGF therapy-induced glomerular microangiopathy" have been proposed. We present a case of a 68-year-old woman who was administered postoperative chemotherapy (carboplatin, paclitaxel, and bevacizumab) for stage IV ovarian cancer. Proteinuria and hypertension appeared after three courses; however, six courses were completed. Then, gemcitabine and carboplatin were administered for recurrence of her cancer. She was diagnosed with nephrotic syndrome after eight courses. Renal biopsy showed accumulation of periodic acid-Schiff (PAS)-positive substances in the capillary walls and para-mesangial areas. Double contouring of basement membranes was also observed. Immunofluorescence microscopy revealed positive staining for IgG, IgA, IgM, C3, C4, and C1q. Immunosuppressive therapy was administered, but was ineffective. Further examination by electron microscopy and immunostaining led to a diagnosis of bevacizumab-associated glomerular microangiopathy.

Immunoglobulin-Negative DNAJB9-Associated Fibrillary Glomerulonephritis: A Report of 9 Cases.

Fibrillary glomerulonephritis (FGN) was previously defined by glomerular deposition of haphazardly oriented fibrils that stain with antisera to immunoglobulins but do not stain with Congo red. We report what is to our knowledge the first series of immunoglobulin-negative FGN, consisting of 9 adults (7 women and 2 men) with a mean age at diagnosis of 66 years. Patients presented with proteinuria (100%; mean protein excretion, 3g/d), hematuria (100%), and elevated serum creatinine level (100%). Comorbid conditions included carcinoma in 3 and hepatitis C virus infection in 2; no patient had hypocomplementemia or monoclonal gammopathy. Histologically, glomeruli were positive for DNAJB9, showed mostly mild mesangial hypercellularity and/or sclerosis, and were negative for immunoglobulins by immunofluorescence on frozen and paraffin tissue. Ultrastructurally, randomly oriented fibrils measuring 13 to 20nm in diameter were seen intermingling with mesangial matrix in all and infiltrating glomerular basement membranes in 5. On follow-up (mean duration, 21 months), 2 had disease remission, 4 had persistently elevated serum creatinine levels and proteinuria, and 3 required kidney replacement therapy. Thus, rare cases of FGN are not associated with glomerular immunoglobulin deposition, and the diagnosis of FGN in these cases can be confirmed by DNAJB9 immunostaining. Pathogenesis remains to be elucidated.

IgM nephropathy complicated by cerebral venous sinus thrombosis: a case study.

BACKGROUND: IgM nephropathy is a rare disease with variable clinical presentations and is an unusual cause of nephrotic syndrome. Histopathological findings typically include mesangial hypercellularity with IgM and complement deposition, though the spectrum may range from normal glomeruli through to focal and segmental glomerulosclerosis. Thromboembolism is a well recognised complication of nephrotic syndrome, but cerebral venous sinus thrombosis is rarely described. CASE PRESENTATION: This is the case of a 23-year-old male presenting with the nephrotic syndrome, whose initial renal biopsy was consistent with minimal change disease. Complete remission was achieved with prednisone, however multiple relapses and steroid dependence prompted re-biopsy, the results of which were more consistent with IgM nephropathy. His last relapse was complicated by cerebral venous sinus thrombosis. He then received rituximab and a weaning course of prednisone to again enter remission. CONCLUSIONS: This case highlights the need to consider IgM nephropathy in the differential diagnosis of nephrotic syndrome. Additionally, it emphasises the risk of thrombosis in patients with severe nephrosis.

Renal injury following long-term exposure to carbon disulfide: analysis of a case series.

BACKGROUND: To investigate the clinicopathological characteristics of renal damage caused by long-term exposure to carbon disulfide (CS2) in nine patients. METHODS: All the patients underwent ultrasound-guided renal biopsy. All specimens were examined by light microscopy and immunohistochemistry (IHC). Samples form one patient were further analyzed using transmission electron microscopy. RESULTS: Similar pathological changes were observed in all patients, but the degrees of lesions were different. All cases had moderate to severe nodular mesangial hyperplasia; among these, type "Kimme1stie1-Wi1son" (K-W nodule for short) was observed in four cases, type "K - W nodule" refer to nodular hyperplasia of mesangial membrane like letter K or W. four cases had proliferative extracapillary glomerulonephritis (GN), while there were no concomitant changes in one patient. Besides, six cases had diffuse basement membrane thickening, focal segmental sclerosis or bulbar sclerosis; two cases had diffuse glomerular sclerosis, and one case had focal segmental capillary hyperplasia. Moreover, all patients had renal tubular atrophy/interstitial fibrosis with less to moderate chronic inflammatory cell infiltration, as well as renal arteriosclerosis. IHC showed that the depositions of IgA, IgM, C3d, C4d, C1q and Fib were not specific; while IgG, type III collagen, Fibronectin, Amyloid A, Igκ, Igλ, HBsAg and HBcAg were all negative. CONCLUSION: Diffuse nodular mesangial hyperplasia/sclerosing glomerular nephropathy is characterized by nodular mesangial hyperplasia with type "K-W nodules" formation, which we speculate is a special pathological manifestation of renal damage caused by carbon disulfide (CS2).

Renal crystal-storing histiocytosis involving glomeruli - A comprehensive clinicopathologic analysis.

Crystal-storing histiocytosis (CSH) is a rare manifestation of monoclonal gammopathy in which histiocytes containing monoclonal proteins in their cytoplasm are found in various organs of the body including the kidney. Within the kidney, these monoclonal crystal-laden histiocytes have been described to occur in the interstitium (most commonly) or in the glomerular mesangium. CSH within glomerular capillary loops has rarely been reported. We describe three cases of CSH primarily affecting the glomerular capillaries and review the literature of CSH in general. Twenty cases of CSH involving the kidney are present in the literature; three describe CSH in glomeruli, only one of which showed histiocytes predominantly in glomerular capillary loops, while 15 had predominantly or solely interstitial CSH. Most cases involve IgG kappa crystals with only one case involving lambda light chain. Patients with CSH predominantly involving the glomerular capillaries showed a trend toward lower serum creatinine and proteinuria at presentation, and several patients with CSH lacked a definitive diagnosis of a monoclonal gammopathy at the time of diagnosis, emphasizing the role that kidney biopsy and particularly electron microscopy play in diagnosis of this entity.

Three Dimensional Glomerular Reconstruction: A Novel Approach to Evaluate Renal Microanatomy in Diabetic Kidney Disease.

Mesangial metrics reflect glomerular filtration surface area in diabetes. The point-sampled intercept (PSI) method is the conventional method to calculate these parameters. However, this is time consuming and subject to underestimation. We introduce a novel three-dimensional (3D) reconstruction method applicable to light microscopy to measure mesangial metrics. Transmission electron microscopy (TEM), PSI and our new 3D imaging methods were used to quantify mesangial metrics from 22 patients with type 2 diabetes, normo-, micro- and macroalbuminuria and an estimated glomerular filtration rate of <60 mL/min/1.73 m2. Repeated-measures ANOVA test was used to test the equality of the measurement means from the three methods and the degree of inter method variability. Repeated-measures and post-estimation ANOVA tests together with correlation coefficient measurements were used to compare the methods with TEM as reference. There was a statistically significant difference in mesangial volume measurements (F(2, 16) = 15.53, p = 0.0002). The PSI method underestimated measurements compared to TEM and 3D methods by 30% (p = 0.001) and 15%, respectively (p < 0.001). 3D and TEM measurements did not differ significantly. 3D reconstruction is a reliable and time efficient method for calculating mesangial metrics. It may prove to be a useful tool in clinical and experimental diabetic kidney disease.

Nephron-specific knockin of the PIKfyve-binding-deficient Vac14L156R mutant results in albuminuria and mesangial expansion.

Intracellular trafficking processes play a key role for the establishment and maintenance of membrane surfaces in renal epithelia. Therefore, dysfunctions of these trafficking processes could be key events and important determinants in the onset and progression of diseases. The presence of cellular vacuoles-observed in many histologic analyses of renal diseases-is a macroscopic hint for disturbed intracellular trafficking processes. However, how vacuoles develop and which intracellular pathways are directly affected remain largely unknown. Previous studies showed that in some cases, vacuolization is linked to malfunction of the Vac14 complex. This complex, including the scaffold protein Vac14, the lipid kinase PIKfyve, and its counteracting lipid phosphatase Fig4, regulates intracellular phosphatidylinositol phosphate levels, which in turn, control the maturation of early-into-late endosomes, as well as the processing of autophagosomes into autophagolysosomes. Here, we analyzed the role of Vac14 in mice and observed that the nephron-specific knockin of the PIKfyve-binding-deficient Vac14L156R mutant led to albuminuria, accompanied by mesangial expansion, increased glomerular size, and an elevated expression of several kidney injury markers. Overexpression of this Vac14 variant in podocytes did not reveal a strong in vivo phenotype, indicating that Vac14-dependent trafficking processes are more important for tubular than for glomerular processes in the kidney. In vitro overexpression of Vac14L156R in Madin-Darby canine kidney cells had no impact on apico-basal polarity defects but resulted in a faster reassembly of junctional structures after Ca2+ depletion and delayed endo- and transcytosis rates. Taken together, our data suggest that increased albuminuria of Vac14L156R-overexpressing mice is a consequence of a lowered endo- and transcytosis of albumin in renal tubules.

CCR7 Is Important for Mesangial Cell Physiology and Repair.

The homeostatic chemokine receptor CCR7 serves as key molecule in lymphocyte homing into secondary lymphoid tissues. Previous experiments from our group identified CCR7 also to be expressed by human mesangial cells. Exposing cultured human mesangial cells to the receptor ligand CCL21 revealed a positive effect on these cells regarding proliferation, migration, and survival. In the present study, we localized CCR7 and CCL21 during murine nephrogenesis. Analyzing wild-type and CCR7 deficient (CCR7-/-) mice, we observed a retarded glomerulogenesis during renal development and a significantly decreased mesangial cellularity in adult CCR7-/- mice, as a consequence of less mesangial cell proliferation between embryonic day E17.5 and week 5 postpartum. Cell proliferation assays and cell-wounding experiments confirmed reduced proliferative and migratory properties of mesangial cells cultured from CCR7-/- kidneys. To further emphasize the role of CCR7 as important factor for mesangial biology, we examined the chemokine receptor expression in rats after induction of a mesangioproliferative glomerulonephritis. Here, we demonstrated for the first time that extra- and intraglomerular mesangial cells that were CCR7-negative in control rats exhibited a strong CCR7 expression during the phase of mesangial repopulation and proliferation.

Accumulation of worn-out GBM material substantially contributes to mesangial matrix expansion in diabetic nephropathy.

Thickening of the glomerular basement membrane (GBM) and expansion of the mesangial matrix are hallmarks of diabetic nephropathy (DN), generally considered to emerge from different sites of overproduction: GBM components from podocytes and mesangial matrix from mesangial cells. Reevaluation of 918 biopsies with DN revealed strong evidence that these mechanisms are connected to each other, wherein excess GBM components fail to undergo degradation and are deposited in the mesangium. These data do not exclude that mesangial cells also synthesize components that contribute to the accumulation of matrix in the mesangium. Light, electron microscopic, immunofluorescence, and in situ hybridization studies clearly show that the thickening of the GBM is due not only to overproduction of components of the mature GBM (α3 and α5 chains of collagen IV and agrin) by podocytes but also to resumed increased synthesis of the α1 chain of collagen IV and of perlecan by endothelial cells usually seen during embryonic development. We hypothesize that these abnormal production mechanisms are caused by different processes: overproduction of mature GBM-components by the diabetic milieu and regression of endothelial cells to an embryonic production mode by decreased availability of mediators from podocytes.

Clinical and pathological characteristics of Fabry disease combined with IgA nephropathy in Chinese patients
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AIMS: To improve diagnosis and treatment, we characterized Fabry disease combined with IgA nephropathy and its response to treatment clinically and pathologically in Chinese patients. MATERIALS AND METHODS: Clinical and pathological characteristics of 6 Chinese patients with renal biopsy-proven Fabry disease combined with IgA nephropathy were retrospectively analyzed. RESULTS: There were 4 males and 2 females in this study. All of the 6 patients presented with proteinuria. Microscopic hematuria was observed in case 4. Extrarenal symptoms included: acroparesthesia in case 1, 2, and 6, hypohidrosis in case 6 and angiokeratomas with hearing loss in case 3. By light microscopy, podocyte distension, with vacuolization, mesangial expansion, and interstitial lesions were found in all 6 cases; and focal segmental glomerulosclerosis was observed in 3 cases. Immunofluorescence microscopy showed deposition of IgA or predominant IgA with C3 in the mesangium. By electron microscopy, myelin figures and/or zebra bodies as well as electron-dense materials, were observed in the mesangium in the 6 cases. After admission and treatment with angiotensin converting enzyme inhibitors (ACEI) or angiotensin receptor blocker (ARB) together with immunosuppressant, glomerular filtration rate (GFR) decreased in 3 cases while it increased in the other 3 without statistical differences. CONCLUSIONS: The clinical and pathological features of Fabry disease combined with IgA nephropathy are diverse. Therefore, it is important to combine family history, clinical manifestations, α-galactosidase A activity and pathological features, especially ultrastructural changes, to improve the diagnosis and treatment of the disease.
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Prevalence of Glomerulopathies in Canine Mammary Carcinoma.

The incidence and prevalence of paraneoplastic glomerulopathy, especially associated with carcinoma, are a matter of debate and the causal link between cancer and glomerular diseases remains unclear. The aim of this study was to evaluate renal biopsies of selected bitches with spontaneous mammary gland carcinoma. We hypothesized that dogs with mammary carcinomas would show histologic evidence of glomerular pathology. A prospective study was performed in dogs with naturally occurring mammary carcinoma that were undergoing tumor resection and ovariohysterectomy. We evaluated renal biopsies of 32 bitches with spontaneous mammary gland carcinoma and 11 control dogs without mammary gland neoplasia. Samples were obtained from the left kidney and the biopsy material was divided for light microscopy (LM), immunofluorescence (IF) and transmission electron microscopy (TEM). Light microscopy abnormalities were identified in 78.1% of dogs with mammary carcinoma (n = 25) and in none of the dogs in the control group. Focal glomerular mesangial matrix expansion was the most common alteration (n = 15, 60.0%), but mesangial cell proliferation (n = 9, 36.0%) and focal segmental glomerulosclerosis (n = 9, 36.0%), synechiae (n = 7, 28.0%), and globally sclerotic glomeruli (n = 6, 24.0%) were also frequent in dogs with malignancy. Immunofluorescence microscopy revealed strong IgM staining was demonstrated in 64.3% (n = 18) of carcinoma dogs. Transmission electron microscopy from dogs with carcinoma revealed slight changes, the most frequent of which was faint sub-endothelial and mesangial deposits of electron-dense material (78%). Mesangial cell interpositioning and segmental effacement of podocyte foot processes were identified in some specimens (45%). Changes in the glomerulus and proteinuria are common in dogs with naturally occurring mammary carcinoma and this condition appears to provide an excellent large animal model for cancer-associated glomerulopathy in humans.

Comparative Proteomics Analysis of Mouse Habu Nephritis Models with and without Unilateral Nephrectomy.

BACKGROUND/AIMS: Individuals possessing a single kidney are at greater risk of renal injury upon exposure to harmful stimuli. This study aimed to explore the pathogenesis of renal injury in glomerulonephritis with versus without unilateral nephrectomy (UNX). METHODS: Histological analysis and label-free quantitative proteomics were performed on two models-the Habu snake venom-induced glomerulonephritis model with versus without UNX (HabuU and Habu models, respectively). The role of villin 1, a differentially expressed protein (DEP) in mouse mesangial cells, was investigated. RESULTS: Persistent mesangiolysis and focal hypercellularity together with reduced activation of cell proliferation in the HabuU model induced more serious renal injury compared with that in the Habu model. The DEPs between the two models were identified by label-free liquid chromatography-mass spectrometry. The KEGG pathway results indicated that regulation of actin cytoskeleton and focal adhesion were specifically enriched in the HabuU model. The cytoskeleton regulation protein villin 1 was downregulated in the HabuU model, but unchanged in the Habu model. Knockdown of villin 1 promoted apoptosis and inhibited the proliferation of mouse mesangial cells, suggesting villin 1 to be involved in qlomerular lesion self-repair insufficiency. CONCLUSION: By assessing the proteomic profiles of the two models, this study identified several important differences, particularly villin 1 expression, in regulatory mechanisms between the two models. Our findings provide novel insight into the mechanism of serious renal injury in glomerulonephritis with UNX.

Vascular mesangial channels in human nodular diabetic glomerulopathy.

The presence of vascular mesangial channels has been reported in idiopathic nodular glomerulosclerosis and diabetic glomerulopathy. However, only limited information on the morphology and immunohistochemical phenotype of these channels is available. This study aims to describe the light and electron microscopic features of these channels and delineate their immunohistochemical phenotype. Thirty-eight cases of human nodular diabetic glomerulopathy with mesangial channels identified by light microscopy were prospectively selected (2010-2012). The cases were stained with CD31/periodic acid-Schiff combined stain. Selected cases were immunostained for CD34, podoplanin, ERG, and Ki-67. Frequent, small and peripheral vascular mesangial channels were seen in all cases, whereas larger and more centrally located vascular channels were also observed. Communication between peripheral capillary loops and peripheral vascular mesangial channels was seen as was communication between peripheral and central vascular mesangial channels. The vascular mesangial channel lining cells showed a typical endothelial phenotype with strong expression of CD31, CD34, and ERG by immunohistochemistry. The lymphatic channel marker podoplanin was negative in all channels, and the proliferation marker Ki-67 showed no evidence of increased proliferation. By electron microscopy, mesangial channels show angulated, irregular borders with lining cells compatible with endothelium and surrounded by mesangial matrix. No basement membranes were identified surrounding the mesangial channels. These findings support the existence of vascular mesangial channels in nodular diabetic glomerulopathy and suggest neovascularization and altered blood flow within these glomeruli.

Three-dimensional microanatomy of the pericapillary mesangial tissues in the renal glomerulus: Comparative observations in four vertebrate classes.

The renal glomeruli in lower vertebrates display mesangium-like cells and matrices interposed between the capillary endothelium and the basement membrane, while those in mammals reportedly lack such interpositions except in pathological conditions. By combined scanning and transmission electron microscopic observations, the pericapillary mesangial tissues were comparatively analyzed in four vertebrate classes: mammals (rats and rabbits), reptiles (green iguanas), amphibians (bullfrogs), and teleosts (carps). The observations discriminated three types of pericapillary interposition. The first, acellular interpositions, occurred universally, with mammalians displaying rudimental ones. This tissue type corresponded with extracellular matrices held in subendothelial grooves which were supported by fine endothelial projections anchored to the basement membrane. In lower vertebrates these grooves constituted an anastomosed system of subendothelial channels that communicated with the mesangial region, to favor cleaning of the glomerular filter. The second, compound type was specific to reptiles and amphibians, affecting the entire capillary circumference in the latter. In this tissue type, fine mesangial processes--which accompanied considerable amounts of fibrillar matrices--were loosely associated with the endothelial bases, indicating their possible nature as a kind of myofibroblast. Occurrence of the third, cellular interpositions was confined to small incidental loci in mammalian and teleost glomeruli. This tissue type was mostly occupied by thick processes or main bodies of the mesangial cells that tightly interlocked their short marginal microvilli with corresponding indentations on the endothelial bases.