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.

Proliferative lesions of intra-epidermal cytokeratin CAM5.2-positive cells in canine nipples.

Non-keratinocyte cells with clear or vacuolated cytoplasm are frequently observed in the epidermis of canine nipples. Most of these cells express cytokeratin (CK) CAM5.2, a marker of luminal epithelial cells. The morphological and immunohistochemical characteristics of these clear cells were investigated. Nipple tissue from 36 dogs of both sexes was collected and labelled immunohistochemically for CAM5.2, CK7, CK14, CK18, CK20, α-smooth muscle actin, p63, melan-A, E-cadherin, epidermal growth factor receptor and oestrogen receptor (OR). The intra-epidermal CAM5.2(+) clear cells were present singly or as small clusters, mostly within the basal layer, in 22 dogs (61%). These cells also expressed CK7, CK18, E-cadherin and OR. Electron microscopy revealed that some of these cells had surface microvilli. Multifocal proliferative lesions consisting of these cells were observed in the nipples of four dogs. In these lesions, proliferating cells formed bilayered tubules with CAM5.2(+) inner and CK14/p63(+) outer cells. This is the first report describing intra-epidermal CAM5.2(+) clear cells, distinct from melanocytes and Merkel cells in dog nipples. These cells might arise from the luminal epithelium of the papillary duct.

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.

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.

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 Masson's 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.

Interaction of the C-terminal domain of the E. coli RNA polymerase alpha subunit with the UP element: recognizing the backbone structure in the minor groove surface.

The C-terminal domain of the alpha-subunit of Escherichia coli RNA polymerase (alphaCTD) is responsible for transcriptional activation through interaction with both activator proteins and UP element DNA. Previously, we determined the solution structure of alphaCTD. Here, we investigated the interaction between alphaCTD and UP element DNA by NMR. DNA titration curves and intermolecular NOE measurements indicate that alphaCTD can bind to multiple sites on the UP element DNA. Unlike many transcription factors, alphaCTD does not have a strict base sequence requirement for binding. There is a good correlation between the strength of the interaction and the extent of intrinsic bending of the DNA oligomer estimated from the gel retardation assay. We propose that alphaCTD recognizes the backbone structure of DNA oligomers responsible for the intrinsic bending. Moreover, NMR studies and drug competition experiments indicated that alphaCTD interacts with the UP element on the minor groove side of the DNA. The C-terminal end of helix-1, the N-terminal end of helix-4, and the loop between helices 3 and 4 are used for the interaction. Based on these observations, we propose a model for the UP element-alphaCTD complex.

Solution structure of an RNA duplex including a C-U base pair.

The formation of the C-U base pair in a duplex was observed in solution by means of the temperature profile of (15)N chemical shifts, and the precise geometry of the C-U base pair was also determined by NOE-based structure calculation. From the solution structure of the RNA oligomer, r[CGACUCAGG].r[CCUGCGUCG], it was found that a single C-U mismatch preferred being stacked in the duplex rather than being flipped-out even in solution. Moreover, it adopts an irregular geometry, where the amino nitrogen (N4) of the cytidine and keto-oxygen (O4) of the uridine are within hydrogen-bonding distance, as seen in crystals. To further prove the presence of a hydrogen bond in the C-U pair, we employed a point-labeled cytidine at the exocyclic amino nitrogen of the cytidine in the C-U pair. The temperature profile of its (15)N chemical shift showed a sigmoidal transition curve, indicating the presence of a hydrogen bond in the C-U pair in the duplex.

False aneurysm of the popliteal artery treated successfully by surgery: report of two cases.

Blunt trauma to the popliteal artery may result in pseudoaneurysm formation. We herein report two patients who had a pseudoaneurysm of the popliteal arteries without any orthopedic injuries. One patient, who had been struck by an iron plate in the midthigh portion, demonstrated a pulsatile mass in this area 2 months later. The other patient had a pseudoaneurysm that was diagnosed by a histological examination; however, he could not remember having suffered any noticeable trauma.

[A combination chemotherapy of carboplatin and etoposide for malignant fibrous histiocytoma with marked effect on the metastatic lung region].

For metastatic lung region, a 82-year-old woman with malignant fibrous histiocytoma was successfully treated with combination chemotherapy of Carboplatin and Etoposide. First, the tumor appeared on her right thigh, and we operated on August 1991. However, at the same time, a metastatic lung region was also noticed. Over a year, it worsened. We gave her 350 mg Carboplatin once a week and 100 mg Etoposide 3 times a week. After 3 weeks' treatment, we stopped treatment because of sudden granulocytopenia and thrombocytopenia. But to our surprise, the metastatic lung region almost disappeared 2 months after therapy. Considering the difficulty of medical treatment for malignant fibrous histiocytoma, this case suggests that combination chemotherapy of Carboplatin and Etoposide is valuable. However, due care must be taken for side effects such as granulocytopenia and thrombocytopenia.

A freeze-fracture study on the basolateral plasma membrane of the gastric parietal cells in fasting and refed rats.

The basolateral plasma membrane of the gastric parietal cells in rats after food-deprivation and food-restitution was studied by thin-section and freeze-fracture electron microscopy. A decrease in the number of tubulovesicles and dilatation of the canalicular lumen were generally observed in the parietal cells of the refed rats. Basal folds, demonstrated both in fasting and food-resupplied rat groups parietal cells by thin-section electron microscopy, appears as branched furrows on the P face and branched wrinkles on the E face plasma membrane by the freeze-fracture replica method. Parietal cells providing numerous basal folds appeared to be more numerous in refed rat gastric gland. The basal folds may represent, at least to some extent, a reservoir of a surplus plasma membrane and play an important role in secreting substances into and also absorbing substances from the blood capillary via the underlying connective tissue.