scholarly journals Autofluorescence microscopy as a label-free tool for renal histology and glomerular segmentation

2021 ◽  
Author(s):  
Nathan Heath Patterson ◽  
Elizabeth K Neumann ◽  
Kavya Sharman ◽  
Jamie L Allen ◽  
Raymond C Harris ◽  
...  
Keyword(s):  
Recognition Performance ◽  
Periodic Acid ◽  
Kidney Tissue ◽  
Human Kidney ◽  
True Positive Rate ◽  
Label Free ◽  
Periodic Acid Schiff ◽  
Renal Histology ◽  
Fresh Frozen ◽  
Histological Staining

Functional tissue units (FTUs) composed of multiple cells like the glomerulus in the kidney nephron play important roles in health and disease. Histological staining is often used for annotation or segmentation of FTUs, but chemical stains can introduce artefacts through experimental factors that influence analysis. Secondly, many molecular -omics techniques are incompatible with common histological stains. To enable FTU segmentation and annotation in human kidney without the need for histological staining, we detail here the use of widefield autofluorescence (AF) microscopy as a simple, label-free modality that provides detailed renal morphology comparable to periodic acid-Schiff (PAS) stained tissue in both formalin-fixed paraffin-embedded (FFPE) and fresh frozen samples and with no tissue processing beyond sectioning. We demonstrate automated deep learning-based glomerular unit recognition and segmentation on PAS and AF images of the same tissue section from 9 fresh frozen samples and 9 FFPE samples. All training comparisons were carried out using registered AF microscopy and PAS stained whole slide images originating from the same section, and the recognition models were built with the exact same training and test examples. Measures of recognition performance, such as the Dice-Sorensen coefficient, the true positive rate, and the positive predictive value differed less than 2% between standard PAS and AF microscopy for both preservation methods. These results demonstrate that AF is a potentially powerful tool to study human kidney tissue, that it can serve as a label-free source for automated and manual annotation of tissue structures.

PodoSighter: A Cloud-Based Tool for Label-Free Podocyte Detection in Kidney Whole Slide Images

2021 ◽  
pp. ASN.2021050630
Author(s):  
Darshana Govind ◽  
Jan Becker ◽  
Jeffrey Miecznikowski ◽  
Avi Rosenberg ◽  
Julien Dang ◽  
...  
Keyword(s):  
Web Application ◽  
Periodic Acid ◽  
High Sensitivity ◽  
Ground Truth ◽  
Renal Diseases ◽  
Label Free ◽  
Current Standard ◽  
Periodic Acid Schiff ◽  
Glomerular Diseases ◽  
Whole Slide Images

Background: Podocyte depletion precedes progressive glomerular damage in several renal diseases. However, the current standard of visual detection and quantification of podocyte nuclei from brightfield microscopy images is laborious and imprecise. Methods: We have developed PodoSighter, an online cloud-based tool, to automatically identify and quantify podocyte nuclei from giga-pixel brightfield whole-slide images (WSIs) using deep learning. Ground-truth to train the tool used immunohistochemically or immunofluorescence-labeled images from a multi-institutional cohort of 122 histologic sections from mouse, rat, and human kidneys. To demonstrate generalizability of our tool in investigating podocyte loss in clinically relevant samples, we tested it in rodent models of glomerular diseases, including diabetic kidney disease, crescentic glomerulonephritis, and dose-dependent direct podocyte toxicity and depletion, as well as in human biopsies from steroid resistant nephrotic syndrome and from human autopsy tissues. Results: The optimal model yielded high sensitivity/specificity of 0.80/0.80, 0.81/0.86, and 0.80/0.91, in mouse, rat, and human images, respectively, from periodic-acid Schiff-stained WSIs. Furthermore, the podocyte nuclear morphometrics extracted using PodoSighter were informative in identifying diseased glomeruli. We have made PodoSighter freely available to the general public as turnkey plugins in a cloud-based web application for end-users. Conclusion: Our study demonstrates an automated computational approach to detect and quantify podocyte nuclei in standard histologically-stained WSIs, facilitating podocyte research and enabling possible future clinical applications.

Histoprotective effect of vitamin D against carbon tetrachloride nephrotoxicity in rats

2015 ◽  
Vol 35 (7) ◽  
pp. 713-723 ◽  
Author(s):  
SMEO Elmubarak ◽  
N Özsoy
Keyword(s):  
Vitamin D ◽  
Carbon Tetrachloride ◽  
Periodic Acid ◽  
Kidney Tissue ◽  
Albino Rats ◽  
Periodic Acid Schiff ◽  
Glucose Levels ◽  
Tissue Sections ◽  
Wistar Albino Rats ◽  
Hematoxylin Eosin

This study investigated the protective effect of vitamin D against carbon tetrachloride (CCl4)-induced nephrotoxicity in rats. Adult male Wistar albino rats were divided into four groups ((A) control; (B) 10-week exposure to CCl4; (C) 10-week exposure to CCl4 + vitamin D treatment; and (D) 10-week exposure to CCl4 + 12 weeks of vitamin D treatment). The CCl4 dose (1.5 ml kg−1) was injected subcutaneously twice a week, while the 0.5 mg kg−1 dose of vitamin D was administered intraperitoneally every day, as appropriate for each group. Whole animal and kidney weights as well as serum urea, creatinine, and glucose levels were measured. Kidney tissue sections were stained with hematoxylin–eosin, Masson’s trichrome, and periodic acid–Schiff. Tubular and glomerular degeneration were detected in the kidney tissues of CCl4-treated rats, together with dilatation and vacuolization within the tubules and hemorrhage in the intertubular region. In the kidney glomeruli; congestion, atrophy, and adhesion to parietal layer were observed. Tissue disorganization and aggregation of Bowman’s capsules were noted. Mononuclear cell infiltration was observed between the glomeruli and the tubules. In contrast, the kidney sections and functional parameters of vitamin D-treated rats were similar to the controls, suggesting that vitamin D treatment is able to reduce renal damage.

Symmetric Dimethylarginine Is a Sensitive Biomarker of Glomerular Injury in Rats

2021 ◽  
pp. 019262332110453
Author(s):  
Rebecca Kohnken ◽  
Lauren Himmel ◽  
Michael Logan ◽  
Richard Peterson ◽  
Sabyasachi Biswas ◽  
...  
Keyword(s):  
Renal Function ◽  
Renal Injury ◽  
Periodic Acid ◽  
Kidney Tissue ◽  
Glomerular Injury ◽  
Symmetric Dimethylarginine ◽  
Puromycin Aminonucleoside ◽  
Periodic Acid Schiff ◽  
Gold Standard Method ◽  
Toxicity Studies

Glomerular filtration rate is the gold-standard method for assessment of renal function but is rarely performed in routine toxicity studies. Standard serum biomarkers of renal function are insensitive and become elevated only with significant loss of organ function. Symmetric dimethylarginine (SDMA) is a ubiquitous analyte that is freely filtered by the glomerulus and can be detected in serum. It has shown utility for the detection of renal injury in dogs and cats in clinical veterinary practice, but the potential utility of SDMA to detect renal injury in preclinical species or toxicity studies has not been thoroughly investigated. We utilized a well-characterized glomerular toxicant, puromycin aminonucleoside, to induce podocyte injury and subsequent proteinuria in young male Sprague-Dawley rats. At the end of 1 or 2 weeks, blood, urine, and kidney tissue were collected for analysis. One week following a single 50 mg/kg dose, urea nitrogen, creatinine, and albumin mean values were within historical control ranges, while SDMA was increased. Glomerular changes in these animals included periodic acid–Schiff positive globules within podocytes, podocyte hypertrophy by light microscopy, and podocyte degeneration with effacement of foot processes by electron microscopy (EM). Taken together, our data indicate that SDMA may be a useful biomarker for early detection of glomerular toxicities in rats.

scholarly journals C4d immunohistochemistry in membranous nephropathy

2014 ◽  
Vol 6 (02) ◽  
pp. 076-079 ◽  
Author(s):  
Monalisa Hui ◽  
Megha S Uppin ◽  
Aruna K Prayaga ◽  
Sree Bhushan Raju ◽  
Liza Rajasekhar
Keyword(s):  
Basement Membrane ◽  
Membranous Nephropathy ◽  
Periodic Acid ◽  
Polyclonal Antiserum ◽  
Hepatitis C Infection ◽  
Periodic Acid Schiff ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Fresh Frozen ◽  
Hematoxylin And Eosin

ABSTRACT Background: Membranous nephropathy (MN) is the most common cause of nephropathy in adults. The diagnosis is based on characteristic light microscopic, electron microscope and immunofluorescence (IF) findings. In early MN, the light microscopic findings may be difficult to differentiate from minimal chain disease. In the absence of fresh frozen tissue for IF, immunohistochemistry with C4d aids in the diagnosis. Materials and Methods: A total 48 cases of MN diagnosed on renal biopsy were analyzed. The formalin fixed paraffin embedded tissues were stained with routine hematoxylin and eosin stains along with periodic acid-Schiff and silver methenamine stains to highlight the basement membrane. Fresh frozen tissues were available for IF in 40 cases. Immunostaining with C4d was done on paraffin-embedded sections by polymer-Horse Radish Peroxidase (HRP) technique using polyclonal antiserum to C4d (Biogenex, India). Results: There were 25 cases of idiopathic MN, 17 cases of Class V lupus nephritis and 2 cases were secondary to hepatitis C infection with cirrhosis. The glomerular basement membrane (GBM) was diffusely thickened with formation of spikes in 28 cases. In 11 cases the capillary loops were rigid but spikes were not seen and in 9 cases there was no apparent thickening of the basement membrane. All the cases showed diffuse positivity for C4d along the GBM. Conclusion: C4d is a reliable method to establish the diagnosis of MN and also a sensitive marker of complement activation reflecting the pathogenesis of MN.

scholarly journals Deep Learning–Based Segmentation and Quantification in Experimental Kidney Histopathology

2020 ◽  
Vol 32 (1) ◽  
pp. 52-68 ◽  
Author(s):  
Nassim Bouteldja ◽  
Barbara M. Klinkhammer ◽  
Roman D. Bülow ◽  
Patrick Droste ◽  
Simon W. Otten ◽  
...  
Keyword(s):  
Neural Network ◽  
Deep Learning ◽  
Convolutional Neural Network ◽  
High Throughput ◽  
Clinical Studies ◽  
High Performance ◽  
Periodic Acid ◽  
Kidney Tissue ◽  
Disease Models ◽  
Periodic Acid Schiff

BackgroundNephropathologic analyses provide important outcomes-related data in experiments with the animal models that are essential for understanding kidney disease pathophysiology. Precision medicine increases the demand for quantitative, unbiased, reproducible, and efficient histopathologic analyses, which will require novel high-throughput tools. A deep learning technique, the convolutional neural network, is increasingly applied in pathology because of its high performance in tasks like histology segmentation.MethodsWe investigated use of a convolutional neural network architecture for accurate segmentation of periodic acid–Schiff-stained kidney tissue from healthy mice and five murine disease models and from other species used in preclinical research. We trained the convolutional neural network to segment six major renal structures: glomerular tuft, glomerulus including Bowman’s capsule, tubules, arteries, arterial lumina, and veins. To achieve high accuracy, we performed a large number of expert-based annotations, 72,722 in total.ResultsMulticlass segmentation performance was very high in all disease models. The convolutional neural network allowed high-throughput and large-scale, quantitative and comparative analyses of various models. In disease models, computational feature extraction revealed interstitial expansion, tubular dilation and atrophy, and glomerular size variability. Validation showed a high correlation of findings with current standard morphometric analysis. The convolutional neural network also showed high performance in other species used in research—including rats, pigs, bears, and marmosets—as well as in humans, providing a translational bridge between preclinical and clinical studies.ConclusionsWe developed a deep learning algorithm for accurate multiclass segmentation of digital whole-slide images of periodic acid–Schiff-stained kidneys from various species and renal disease models. This enables reproducible quantitative histopathologic analyses in preclinical models that also might be applicable to clinical studies.

Tubular overexpression of Gremlin in transgenic mice aggravates renal damage in diabetic nephropathy

2015 ◽  
Vol 309 (6) ◽  
pp. F559-F568 ◽  
Author(s):  
Vanessa Marchant ◽  
Alejandra Droguett ◽  
Graciela Valderrama ◽  
M. Eugenia Burgos ◽  
Daniel Carpio ◽  
...  
Keyword(s):  
Diabetic Nephropathy ◽  
Renal Damage ◽  
Transforming Growth Factor ◽  
Mesangial Cells ◽  
Interstitial Fibrosis ◽  
Smooth Muscle Actin ◽  
Periodic Acid ◽  
Kidney Tissue ◽  
Renal Tubules ◽  
Periodic Acid Schiff

Diabetic nephropathy (DN) is currently a leading cause of end-stage renal failure worldwide. Gremlin was identified as a gene differentially expressed in mesangial cells exposed to high glucose and in experimental diabetic kidneys. We have described that Gremlin is highly expressed in biopsies from patients with diabetic nephropathy, predominantly in areas of tubulointerstitial fibrosis. In streptozotocin (STZ)-induced experimental diabetes, Gremlin deletion using Grem1 heterozygous knockout mice or by gene silencing, ameliorates renal damage. To study the in vivo role of Gremlin in renal damage, we developed a diabetic model induced by STZ in transgenic (TG) mice expressing human Gremlin in proximal tubular epithelial cells. The albuminuria/creatinuria ratio, determined at week 20 after treatment, was significantly increased in diabetic mice but with no significant differences between transgenic (TG/STZ) and wild-type mice (WT/STZ). To assess the level of renal damage, kidney tissue was analyzed by light microscopy (periodic acid-Schiff and Masson staining), electron microscopy, and quantitative PCR. TG/STZ mice had significantly greater thickening of the glomerular basement membrane, increased mesangial matrix, and podocytopenia vs. WT/STZ. At the tubulointerstitial level, TG/STZ showed increased cell infiltration and mild interstitial fibrosis. In addition, we observed a decreased expression of podocin and overexpression of monocyte chemoattractant protein-1 and fibrotic-related markers, including transforming growth factor-β1, Col1a1, and α-smooth muscle actin. Together, these results show that TG mice overexpressing Gremlin in renal tubules develop greater glomerular and tubulointerstitial injury in response to diabetic-mediated damage and support the involvement of Gremlin in diabetic nephropathy.

PAS Staining of Fresh Frozen or Paraffin Embedded Human Kidney Tissue v2 (protocols.io.buaknscw)

protocols.io ◽  
2021 ◽  
Author(s):  
Elizabeth Neumann ◽  
Jamie Allen ◽  
Jennifer Harvey ◽  
Maya Brewer ◽  
Carrie not provided ◽  
...  
Keyword(s):  
Kidney Tissue ◽  
Human Kidney ◽  
Fresh Frozen ◽  
Pas Staining

scholarly journals Conversion to Sirolimus Ameliorates Cyclosporine-Induced Nephropathy in the Rat: Focus on Serum, Urine, Gene, and Protein Renal Expression Biomarkers

2014 ◽  
Vol 2014 ◽  
pp. 1-17 ◽  
Author(s):  
José Sereno ◽  
Sara Nunes ◽  
Paulo Rodrigues-Santos ◽  
Helena Vala ◽  
Petronila Rocha-Pereira ◽  
...  
Keyword(s):  
Renal Damage ◽  
Molecular Mechanisms ◽  
Periodic Acid ◽  
Kidney Tissue ◽  
Periodic Acid Schiff ◽  
Gene And Protein Expression ◽  
Hematoxylin And Eosin ◽  
Kidney Lesions ◽  
Renal Expression ◽  
Relevant Gene

Protocols of conversion from cyclosporin A (CsA) to sirolimus (SRL) have been widely used in immunotherapy after transplantation to prevent CsA-induced nephropathy, but the molecular mechanisms underlying these protocols remain nuclear. This study aimed to identify the molecular pathways and putative biomarkers of CsA-to-SRL conversion in a rat model. Four animal groups (n=6) were tested during 9 weeks: control, CsA, SRL, and conversion (CsA for 3 weeks followed by SRL for 6 weeks). Classical and emergent serum, urinary, and kidney tissue (gene and protein expression) markers were assessed. Renal lesions were analyzed in hematoxylin and eosin, periodic acid-Schiff, and Masson’s trichrome stains. SRL-treated rats presented proteinuria and NGAL (serum and urinary) as the best markers of renal impairment. Short CsA treatment presented slight or even absent kidney lesions and TGF-β, NF-κβ, mTOR, PCNA, TP53, KIM-1, and CTGF as relevant gene and protein changes. Prolonged CsA exposure aggravated renal damage, without clear changes on the traditional markers, but with changes in serums TGF-βand IL-7, TBARs clearance, and kidney TGF-βand mTOR. Conversion to SRL prevented CsA-induced renal damage evolution (absent/mild grade lesions), while NGAL (serum versus urine) seems to be a feasible biomarker of CsA replacement to SRL.

scholarly journals PAS Staining of Fresh Frozen or Paraffin Embedded Human Kidney Tissue v1 (protocols.io.4qngvve)

protocols.io ◽  
2019 ◽  
Author(s):  
Jamie Allen ◽  
Jennifer Harvey ◽  
Maya Brewer ◽  
Mark de ◽  
Jeff Spraggins
Keyword(s):  
Kidney Tissue ◽  
Human Kidney ◽  
Fresh Frozen ◽  
Pas Staining

Ultrastructure of the Parotid Gland of the Nine-Banded Armadillo

1977 ◽  
Vol 35 ◽  
pp. 640-641
Author(s):  
J. R. Ruby
Keyword(s):  
Endoplasmic Reticulum ◽  
Parotid Gland ◽  
Periodic Acid ◽  
Acinar Cells ◽  
Duct System ◽  
Parotid Glands ◽  
Dasypus Novemcinctus ◽  
Anterior Portion ◽  
Periodic Acid Schiff ◽  
Thin Sections

Parotid glands were obtained from five adult (four male and one female) armadillos (Dasypus novemcinctus) which were perfusion-fixed. The glands were located in a position similar to that of most mammals. They extended interiorly to the anterior portion of the submandibular gland.In the light microscope, it was noted that the acini were relatively small and stained strongly positive with the periodic acid-Schiff (PAS) and alcian blue techniques, confirming the earlier results of Shackleford (1). Based on these qualities and other structural criteria, these cells have been classified as seromucous (2). The duct system was well developed. There were numerous intercalated ducts and intralobular striated ducts. The striated duct cells contained large amounts of PAS-positive substance.Thin sections revealed that the acinar cells were pyramidal in shape and contained a basally placed, slightly flattened nucleus (Fig. 1). The rough endoplasmic reticulum was also at the base of the cell.

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