A Pilot Study to Assess Markers of Renal Damage in the Rodent Kidney After Exposure to 7 MHz Ultrasound Pulse Sequences Designed to Cause Microbubble Translation and Disruption

Acoustic radiation force has been proposed as a mechanism to enhance microbubble concentration for therapeutic and molecular imaging applications. It is hypothesized that once microbubbles are localized, bursting them with acoustic pressure could result in local drug delivery. It is known that low-frequency, high-amplitude acoustic energy combined with cavitation nuclei can result in bioeffects. However, little is known about the bioeffects potential of acoustic parameters involved in radiation-force and microbubble destruction pulse sequences applied at higher frequencies. In this pilot study, rat kidneys are exposed to high-duty cycle, low-amplitude pulse sequences known to cause substantial bubble translation due to radiation force, as well as high-amplitude short pulse sequences known to cause microbubble destruction. Both studies are performed at 7 MHz on a clinical ultrasound system, and implemented in 3-D for entire kidney exposure. Analysis of biomarkers of renal injury and renal histopathology indicate that there was no significant renal damage due to these ultrasound parameters in conjunction with microbubbles within the study group

X-Linked Alport Dogs Demonstrate Mesangial Filopodial Invasion of the Capillary Tuft as an Early Event in Glomerular Damage

BACKGROUND: X-linked Alport syndrome (XLAS), caused by mutations in the type IV collagen COL4A5 gene, accounts for approximately 80% of human Alport syndrome. Dogs with XLAS have a similar clinical progression. Prior studies in autosomal recessive Alport mice demonstrated early mesangial cell invasion as the source of laminin 211 in the glomerular basement membrane (GBM), leading to proinflammatory signaling. The objective of this study was to verify this process in XLAS dogs. METHODS: XLAS dogs and WT littermates were monitored with serial clinicopathologic data and kidney biopsies. Biopsies were obtained at set milestones defined by the onset of microalbuminuria (MA), overt proteinuria, onset of azotemia, moderate azotemia, and euthanasia. Kidney biopsies were analyzed by histopathology, immunohistochemistry, and electron microscopy. RESULTS: XLAS dogs showed progressive decrease in renal function and progressive increase in interstitial fibrosis and glomerulosclerosis (based on light microscopy and immunostaining for fibronectin). The only identifiable structural abnormality at the time of microalbuminuria was ultrastructural evidence of mild segmental GBM multilamination, which was more extensive when overt proteinuria developed. Co-localization studies showed that mesangial laminin 211 and integrin α8β1 accumulated in the GBM at the onset of overt proteinuria and coincided with ultrastructural evidence of mild cellular interpositioning, consistent with invasion of the capillary loops by mesangial cell processes. CONCLUSION: In a large animal model, the induction of mesangial filopodial invasion of the glomerular capillary loop leading to the irregular deposition of laminin 211 is an early initiating event in Alport glomerular pathology

Concurrent renal amyloidosis and thymoma resulting in a fatal ventricular thrombus in a dog

Thymoma‐associated nephropathies have been reported in people but not in dogs. In this report, we describe a dog with thymoma and concurrent renal amyloidosis. A 7‐year‐old castrated male Weimaraner was presented for progressive anorexia, lethargy, and tachypnea. The dog was diagnosed with azotemia, marked proteinuria, and a thymoma that was surgically removed. Postoperatively, the dog developed a large left ventricular thrombus and was euthanized. Necropsy confirmed the presence of a left ventricular thrombus and histopathology revealed renal amyloidosis. We speculate that the renal amyloidosis occurred secondary to the thymoma, with amyloidosis in turn leading to nephrotic syndrome, hypercoagulability, and ventricular thrombosis. This case illustrates the potential for thymoma‐associated nephropathies to occur in dogs and that dogs suspected to have thymoma should have a urinalysis and urine protein creatinine ratio performed as part of the pre‐surgical database

Detection and Classification of Novel Renal Histologic Phenotypes Using Deep Neural Networks.

With the advent and increased accessibility of deep neural networks (DNNs), complex properties of histologic images can be rigorously and reproducibly quantified. We used DNN-based transfer learning to analyze histologic images of periodic acid-Schiff-stained renal sections from a cohort of mice with different genotypes. We demonstrate that DNN-based machine learning has strong generalization performance on multiple histologic image processing tasks. The neural network extracted quantitative image features and used them as classifiers to look for differences between mice of different genotypes. Excellent performance was observed at segmenting glomeruli from non-glomerular structure and subsequently predicting the genotype of the animal on the basis of glomerular quantitative image features. The DNN-based genotype classifications highly correlate with mesangial matrix expansion scored by a pathologist (R.E.C.), which differed in these animals. In addition, by analyzing non-glomeruli images, the neural network identified novel histologic features that differed by genotype, including the presence of vacuoles, nuclear count, and proximal tubule brush border integrity, which was validated with immunohistologic staining. These features were not identified in systematic pathologic examination. Our study demonstrates the power of DNNs to extract biologically relevant phenotypes and serve as a platform for discovering novel phenotypes. These results highlight the synergistic possibilities for pathologists and DNNs to radically scale up our ability to generate novel mechanistic hypotheses in disease

Proliferative, necrotizing and crescentic immune complex-mediated glomerulonephritis in a cat

Case Summary A 5-year-old cat was examined for vomiting and anorexia of 2 days’ duration. Azotemia, hyperphosphatemia and hypoalbuminemia were the main biochemical findings. Serial analyses of the urine revealed isosthenuria, proteinuria and eventual glucosuria. Hyperechoic perirenal fat was detected surrounding the right kidney by ultrasonography. Histopathologic evaluation of ante-mortem ultrasound-guided needle biopsies of the right kidney was consistent with proliferative, necrotizing and crescentic glomerulonephritis with fibrin thrombi, proteinaceous and red blood cell casts, and moderate multifocal chronic-active interstitial nephritis. Owing to a lack of clinical improvement, the cat was eventually euthanized. Post-mortem renal biopsies were processed for light microscopy, transmission electron microscopy and immunofluorescence. This revealed severe focal proliferative and necrotizing glomerulonephritis with cellular crescent formation, podocyte injury and secondary segmental sclerosis. Ultrastructural analysis revealed scattered electron-dense deposits in the mesangium, and immunofluorescence demonstrated positive granular staining for λ light chains, consistent with immune complex-mediated glomerulonephritis. Severe diffuse acute tubular epithelial injury and numerous red blood cell casts were also seen. Relevance and novel information To our knowledge, this is the first report of naturally occurring proliferative, necrotizing and crescentic immune complex glomerulonephritis in a cat

Assessment of peritubular capillary rarefaction in kidneys of cats with chronic kidney disease

Abstract Background Hypoxia is a key driver of fibrosis and is associated with capillary rarefaction in humans. Objectives Characterize capillary rarefaction in cats with chronic kidney disease (CKD). Animals Archival kidney tissue from 58 cats with CKD, 20 unaffected cats. Methods Cross‐sectional study of paraffin‐embedded kidney tissue utilizing CD31 immunohistochemistry to highlight vascular structures. Consecutive high‐power fields from the cortex (10) and corticomedullary junction (5) were digitally photographed. An observer counted and colored the capillary area. Image analysis was used to determine the capillary number, average capillary size, and average percent capillary area in the cortex and corticomedullary junction. Histologic scoring was performed by a pathologist masked to clinical data. Results Percent capillary area (cortex) was significantly lower in CKD (median 3.2, range, 0.8‐5.6) compared to unaffected cats (4.4, 1.8‐7.0; P = <.001) and was negatively correlated with serum creatinine concentrations (r = −.36, P = .0013), glomerulosclerosis (r = −0.39, P = <.001), inflammation (r = −.30, P = .009), and fibrosis (r = −.30, P = .007). Capillary size (cortex) was significantly lower in CKD cats (2591 pixels, 1184‐7289) compared to unaffected cats (4523 pixels, 1801‐7618; P = <.001) and was negatively correlated with serum creatinine concentrations (r = −.40, P = <.001), glomerulosclerosis (r = −.44, P < .001), inflammation (r = −.42, P = <.001), and fibrosis (r = −.38, P = <.001). Conclusions and Clinical Importance Capillary rarefaction (decrease in capillary size and percent capillary area) is present in kidneys of cats with CKD and is positively correlated with renal dysfunction and histopathologic lesions

Concurrent renal amyloidosis and thymoma resulting in a fatal ventricular thrombus in a dog

Thymoma‐associated nephropathies have been reported in people but not in dogs. In this report, we describe a dog with thymoma and concurrent renal amyloidosis. A 7‐year‐old castrated male Weimaraner was presented for progressive anorexia, lethargy, and tachypnea. The dog was diagnosed with azotemia, marked proteinuria, and a thymoma that was surgically removed. Postoperatively, the dog developed a large left ventricular thrombus and was euthanized. Necropsy confirmed the presence of a left ventricular thrombus and histopathology revealed renal amyloidosis. We speculate that the renal amyloidosis occurred secondary to the thymoma, with amyloidosis in turn leading to nephrotic syndrome, hypercoagulability, and ventricular thrombosis. This case illustrates the potential for thymoma‐associated nephropathies to occur in dogs and that dogs suspected to have thymoma should have a urinalysis and urine protein creatinine ratio performed as part of the pre‐surgical database.This article is published as Loewen, Jennifer M., Rachel E. Cianciolo, Liwen Zhang, Michael Yaeger, Jessica L. Ward, Jodi D. Smith, and Dana N. LeVine. "Concurrent renal amyloidosis and thymoma resulting in a fatal ventricular thrombus in a dog." Journal of veterinary internal medicine (2018). doi: 10.1111/jvim.15062.</p