cytokeratins in the canine epidermis (pages 81–87)

Cytokeratins in the canine epidermis JAKOB H. WALTER Institute of Veterinary Pathology, Freie UniversitaÈt Berlin, Strasse 518 Nr. 15, 14163 Berlin, Germany (Received 6 April 1999; accepted 24 November 1999) Abstract The purpose of this study was to characterize the cytokeratins (CKs) present in the clinically normal skin of dogs. Skin samples from ®ve German shepherds, ®ve Boxers, ®ve Cocker spaniels, ®ve Yorkshire terriers and ®ve mongrels were examined biochemically (using gel electrophoresis and western blotting) and immunohistochemically (using a alkaline phosphatase anti-alkaline phosphatase technique). Results indicated that the canine epidermis expressed the cytokeratins 1, 5, 6, 10/11, 14 and 16. There were no consistent dierences in CK expression between the examined breeds with the exception of an individual polymorphism in CK1 and CK10/11. Immunohistochemical studies showed CK 14 labelling of the basal cell layer whereas CK10/11 staining was seen in the suprabasal cell layer of epidermis. Surprisingly, expression of CK6, known as `stress' cytokeratin, was demonstrated in all epidermal samples. These results indicate that there is a striking consistency of cytokeratin expression in dierent breeds which should be useful in the investigation and characterization of canine skin diseases. Keywords: cytokeratin, dogs, epidermis, keratinocytes, skin. INTRODUCTION Cytokeratins (CKs) are components of the inter- mediate ®lament network of epithelial cells. Up to 85% of the total content of the keratinocytes is composed of cytokeratins. 1 At the time of writing, 20 dierent cytokeratins (excluding hair keratin) have been identi®ed in humans, 2 which shows a remark- able epithelial tissue speci®c expression. 3 For the human epidermis it has been shown that the traditional Moll catalogue 3,4 must be supplemented with polymorphic and individual CK variations. 5±8 Although CKs have been described previously in many immunohistochemical studies of canine skin, especially in studies of neoplasia, 9±12 fundamental experiments on the composition of cytokeratins in normal dog skin have not yet been carried out. The aim of this study was to characterize the cytokeratins and possible polymorphisms in the normal canine epidermis which may be of value in the dierential diagnosis of in¯ammatory, inherited, pre-malignant and malignant diseases of skin. MATERIALS AND METHODS Unless otherwise stated, all chemicals were obtained from Sigma, Deisenhofen, Germany. Samples Skin samples (each approximately 262 cm) were taken from the lateral thorax of dogs with clinically normal skin from a selection of dierent adult breeds (®ve German shepherds, ®ve Boxers, ®ve Cocker spaniels, ®ve Yorkshire terriers, ®ve mongrels) which were presented to a veterinary hospital (Klinik u. Poliklinik f. Kleine Haustiere, FU Berlin). Nine of the dogs were killed in trac accidents and the samples were taken 4±18 h after death. Sixteen of the dogs were euthanased because of various disorders (nine cases of canine congestive cardiomyopathy, four cases of behavioural disorders, three cases of diseases of the lower respiratory system) and the samples were taken directly post mortem. The skin samples were divided: one half was used immediately for the preparation of the epidermal cytokeratins. The other half was ®xed in 10% neutral formalin for 24 h, embedded in paran wax (DDM- Tissue Embedding Medium; Medim, Giessen, Ger- many) and sectioned (4 mm). The sections were stained routinely with haematoxylin and eosin (HE) or processed for immunohistochemistry using the Sequen- za system, a disposable coverplate incubation system (Shandon; Frankfurt/Main, Germany). Histopatholo- gical examination of HE-stained skin sections used in this study showed no histopathological abnormalities. Cytokeratin preparation and separation The skin sample was placed in a water bath at 608C for 5 min and the epidermis was then peeled o the dermis using the back of a scalpel blade. Cytokeratins were extracted from the epidermis in accordance with the methods described previously. 13 The epidermal material (0.5 g) peeled from the skin was minced into smaller pieces and homogenized in 2 mL of homo- genizing buer containing 96 mM NaCl, 8 mM Correspondence: JH Walter, VetPathol., Strasse 518 Nr. 15, 14163 Berlin, Germany. E-mail: [email protected] # 2001 Blackwell Science Ltd 81 Veterinary Dermatology 2001, 12, 81±87 Ahed Bhed Ched Dhed Ref marker Fig marker Table marker Ref end Ref start 219 DISC

cytokeratins in the canine epidermis (pages 81–87)

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