Kyohei Yasuno

Collagenofibrotic Glomerulonephropathy with Fibronectin Deposition in a Dog

We report herein a case of collagenofibrotic glomerulonephropathy in a 3-year-old Shiba Inu with severe proteinuria. Histologically, renal glomeruli were enlarged with massive deposition of a homogeneous eosinophilic substance within the mesangium and capillary walls. The deposits reacted weakly with periodic acid-Schiff, stained deep blue with Massons trichrome, and were positive by immunofluorescence for type III collagen and fibronectin. Ultrastructurally, the deposits consisted of fibrils and amorphous material in the mesangial matrix and beneath the glomerular capillary endothelium. The fibrils had transverse bands analogous to those of collagen fibrils. Electron microscopy also revealed focal detachment of podocytes and foot process effacement in glomerular tufts, which suggested that podocyte injury had contributed to the development of proteinuria in this dog. The current case resembles collagenofibrotic glomerulonephropathy (CFGN) in humans in histopathologic, immunofluorescence, and electron microscopic findings. This is the first report of CFGN in a nonhuman species with glomerular deposition of fibronectin and type III collagen.

Expression of Nephrin, Podocin, α-Actinin-4 and α3-Integrin in Canine Renal Glomeruli

The biological features of podocytes that contribute to the pathogenesis of proteinuria have not been investigated in dogs. The aim of this study was to investigate the expression and localization of nephrin, podocin, α-actinin-4 and α3-integrin in canine renal glomeruli. Renal cortical tissue was collected from the kidneys of five normal adult beagles. Western blotting and immunofluorescence microscopy revealed specific expression and localization of the four proteins in canine glomeruli. Expression of genes encoding the four molecules in isolated glomeruli was detected by reverse transcriptase polymerase chain reaction. The results of this study will permit future exploration of podocyte injury and its involvement in protein leakage from the capillary wall in canine glomerular diseases.

Expression of Periostin in Normal, Atopic, and Nonatopic Chronically Inflamed Canine Skin

In humans, periostin plays a critical role in the enhancement and chronicity of allergic skin inflammation; however, whether it is involved in the pathogenesis of canine dermatitis remains unknown. The aim of this study was to examine the expression patterns of periostin in healthy, atopic, and nonatopic chronically inflamed canine skin. Biopsy specimens from 47 dogs with skin disease and normal skin tissue from 5 adult beagles were examined by light microscopy, immunohistochemistry, and in situ hybridization. In normal skin, periostin was localized just beneath the epidermis and around the hair follicles. In chronically inflamed skin, periostin expression was most intense in the dermis with inflammatory cell infiltrates. In contrast, low levels of periostin were detected in acutely inflamed and noninflamed skin. Conversely, all canine atopic dermatitis tissues characteristically showed the most intense expression of periostin in the superficial dermis, particularly at the epidermal–dermal junction. In situ hybridization showed that periostin mRNA was broadly expressed in the basal epidermal keratinocytes, outer root sheath cells, and dermal fibroblasts in normal dog skin. High expression of periostin mRNA was observed in fibroblasts in dog skin with chronically inflamed dermatitis. Moreover, in some chronically inflamed skin specimens, periostin mRNA expression was increased in basal keratinocytes. The severity score of chronic pathologic changes and CD3+ cell number in the dermis were correlated with distribution pattern of periostin in the atopic skin. These data suggest that periostin could play a role in the pathophysiology of chronic dermatitis, including atopic dermatitis, in dogs.

Dynamics of absolute amount of nephrin in a single podocyte in puromycin aminonucleoside nephrosis rats calculated by quantitative glomerular proteomics approach with selected reaction monitoring mode

BACKGROUND The slit diaphragm (SD) is a complex of podocyte-specific proteins and plays a significant role in glomerular filtration. To understand podocyte biology, it is important to determine the expression amount of the SD complex proteins. This study aimed to quantify the absolute amount of nephrin, which is believed to be a major component of SD, in podocytes and to apply that method to normal and puromycin aminonucleoside (PAN) nephrosis rats. METHODS The counting method for podocyte number in a glomerulus was developed by three-dimensional reconstruction imaging of Wilms tumor (WT-1) immunofluorescence on isolated glomeruli. Absolute amount of nephrin was quantified by mass spectrometry using the selected reaction monitoring (SRM) mode with a stable isotope-labeled peptide. RESULTS The number of podocytes per glomerulus was 95.5±17.6 in the control rats, 90.7±19.2 on Day 4 and 90.7±26.2 on Day 7 in PAN nephrosis rats. The amount of nephrin per glomerulus in control rats was 1.02±0.11 fmol and those in PAN nephrosis rats were reduced to 0.46±0.06 fmol and 0.35±0.04 fmol on Day 4 and Day 7. The nephrin amount per podocyte was significantly decreased association with the development of proteinuria in PAN nephrosis rats. CONCLUSIONS This study established the absolute quantification of nephrin and determined the amount of nephrin in a podocyte of normal and PAN nephrosis rat kidneys. This highly sensitive and selective quantification method for protein is a useful tool for the analysis of SD protein in a podocyte.

Trichoblastoma with Abundant Plump Stromal Cells in a Dog

ABSTRACT Histopathological and immunohistochemical examinations were made on a cutaneous tumor on the head of an 11-year-old female mixed-breed dog. The tumor was well demarcated and comprised multilobular structures of neoplastic epithelial cells with abundant plump peritumoral stromal cells. The neoplastic cells formed irregular cell cords or trabeculae and were arranged in characteristic palisades at the periphery. Immunohistochemically, neoplastic cells were positive for p63 and the several cytokeratins examined. In contrast, the plump peritumoral stromal cells were positive for vimentin and unevenly for nestin, a neuroepithelial stem cell protein. The stromal cells prominently proliferated in proximity to epithelial neoplastic cells, suggesting a close interaction between these two cell types.

Mammary adenoma with sebaceous differentiation in a dog

The current report describes a complex canine mammary adenoma with a rare histological feature characterized by sebaceous differentiation of tumor cells. A 13-year-old, mixed-breed, intact female dog had mammary tumors on the right mammary chain. Histologically, one of the masses was composed of bilayered ductal structures with luminal epithelial cells together with basaloid or myoepithelial cell components. Within the tumor, there were a number of lobules and nests of large foamy cells associated with basaloid reserve–like cells similar to sebaceous gland. Squamous metaplasia was also seen within the tumor. Immunohistochemical staining indicated that the tumor cells with sebaceous differentiation were positive for cytokeratin (CK)14 and that the associated basaloid reserve–like cells were positive for p63. In contrast, other luminal epithelial tumor cells were positive for CK18 and CK19, but not for CK14 and p63. The myoepithelial cells were positive for α-smooth muscle actin and p63. The expression of p63 in both sebaceous basaloid reserve–like cells and myoepithelial cells, and their structural continuity within the tumor tissue, suggested a common origin of these 2 components.

Caudal vascular hamartoma accompanied by aberrant arteriovenous structures in a dog

A 9-year-old, female German Shepherd Dog presented with a firm bulging lesion at the tip of the tail. Histologically, the lesion was characterized by numerous poorly circumscribed clusters of vascular structures resembling capillaries that were separated by normal or mucinous connective tissues. The capillary-like vascular structures were composed of several layers: a single von Willebrand factor–positive endothelial cell layer with round, oval, or flattened hyperchromatic nuclei; and 1 or 2 surrounding spindle-shaped smooth muscle actin–positive pericyte layers. In the deep portion of the lesion, there were large vessels that showed morphological aberrations. These thin-to-thick–walled blood vessels corresponded to arteries and veins of varying diameters and were surrounded by fibrosis. The present case report describes a rare cutaneous vascular hamartoma, accompanied by aberrant arteriovenous structures.

Analysis of ultrastructural glomerular basement membrane lesions and podocytes associated with proteinuria and sclerosis in Osborne-Mendel rats with progressive glomerulonephropathy.

The renal glomeruli of 12 male Osborne-Mendel (OM) rats 3 to 24 weeks old were examined by electron microscopy. Effacement of podocyte foot processes (FPs) developed at 3 weeks of age and became progressively worse over time. Loss or dislocation of the slit membrane was also found. Vacuoles and osmiophilic lysosomes appeared in the podocytes starting at 6 weeks of age. Podocyte detachment from the glomerular basement membrane (GBM) was apparent at 18 weeks of age. Laminated GBM was occasionally observed in all animals. These features might lead to the development of spontaneous proteinuria and glomerulosclerosis in OM rats.

Development of Podocyte Injuries in Osborne–Mendel Rats is Accompanied by Reduced Expression of Podocyte Proteins

Osborne-Mendel (OM) rats spontaneously develop glomerulopathy with progressive podocyte injury. Changes in protein expression levels in the foot processes of podocytes have been suggested to play an important role in the development of renal disease. The aim of this study was to investigate the temporal relationship between the expression of five podocyte proteins (nephrin, podocin, synaptopodin, α-actinin-4 and α3-integrin) and the development of podocyte injuries, proteinuria and glomerulosclerosis in OM rats. Male OM rats 5-20 weeks of age and age-matched Fischer 344 rats were used. Semiquantitative analysis of expression of the five podocyte proteins was performed by immunofluorescence labelling. Nephrin mRNA expression was determined by quantitative real-time reverse transcriptase polymerase chain reaction and nephrin protein expression was determined by mass spectrometry. Progressive reduction in expression of the podocyte proteins correlated with the progression of podocyte injuries, the development of proteinuria and the subsequent development of glomerulosclerosis. Nephrin mRNA expression and nephrin concentration also showed temporal decreases in OM rats. Altered expression of podocyte proteins preceded the development of proteinuria and glomerulosclerosis, suggesting that this event contributes to podocyte dysfunction and progression to glomerulosclerosis.

A malignant and metastasizing feline cardiac ganglioneuroma

In the current study, a case of a cardiac ganglioneuroma with systemic metastases in a cat is described. A 12-year-old male neutered Japanese domestic cat was brought to a veterinary hospital for dysorexia, coughing, vomiting, and diarrhea. Ultrasonography revealed a mass adjacent to the right atrium. The animal died of respiratory failure 1 month after the first visit to the hospital. At necropsy, an oval-shaped white mass 1.5 cm in diameter was observed within the right auricle. Diffusely, the right ventricle was infiltrated and thickened by the neoplastic lesion. Histologically, the mass was composed of 3 types of neoplastic cells: spindle cells, large polygonal cells, and small undifferentiated cells. Immunohistochemically, the neoplasia was positive for neuronal markers such as βIII tubulin, S-100a, and protein gene product 9.5. Ultrastructurally, the large polygonal cells were characterized by abundant cytoplasm that included compressed Golgi cisternae and rough endoplasmic reticula and abundant intermediate filaments. A discontinuous basement membrane surrounded the spindle cells. Metastatic foci were found in the lungs, kidney, pancreas, urinary bladder, and adrenal glands. The morphological, immunohistochemical, and ultrastructural characteristics of the tumor cells were consistent with those of ganglioneuroma. The tumor was presumed to originate from the intramural parasympathetic ganglia in the right atrium.