The expression of keratins, vimentin, neurofilament proteins, smooth muscle actin, neuron-specific enolase, and synaptophysin in tumors of the specific glands in the canine anal region.

Eight canine tumors originating from specific glandular structures in the anal region, as well as metastatic tumor tissue of two of these cases (case Nos. 7, 8), were immunohistochemically analyzed using various monoclonal antibodies (MoAbs) directed against human keratin types, vimentin, neurofilament proteins, and alpha-smooth muscle actin. These tumors also were stained for the broad-spectrum neuroendocrine markers neuron-specific enolase (NSE) and synaptophysin. In histologically normal canine anal structures, alpha-smooth muscle actin and NSE antibodies stained basally localized (probably myoepithelial) cells in the anal glands and the anal sac glands. NSE staining also was present in a limited number of luminal cells in both anal glands and anal sac glands. Synaptophysin labeling was not observed in any of these glandular structures. Histologically, the tumors were differentiated into well- and moderately differentiated perianal gland tumors (n = 5) and carcinomas without perianal gland differentiation (n = 3), corresponding to the so-called apocrine carcinomas of the anal region. Immunohistochemically, the perianal gland tumors could be differentiated from the carcinomas by marked differences in staining pattern with the various keratin MoAbs, particularly MoAbs directed against human keratin types 7 and 18. The keratin-staining characteristics of the carcinomas suggest a glandular luminal cell origin. Metastases of the carcinomas showed loss of some keratin-staining characteristics as compared with the primary tumor. Staining for NSE was only observed in solitary cells and small cell clusters in the carcinomas and their metastases, whereas the alpha-smooth muscle actin antibody did not react with the carcinoma cells. None of the tumors stained for neurofilament proteins or synaptophysin. An unequivocal neuroendocrine nature of the carcinomas could not be substantiated by our immunohistochemical study, although the presence of a population of neuroendocrine cells within these neoplasms seems likely. Because the immunohistochemical features of the carcinomas with respect to various keratin MoAbs and NSE are similar to those of the anal glands and the anal sac glands, both these glands might be considered as site of origin of these carcinomas.

Immunohistochemistry with keratin monoclonal antibodies in canine tissues: urogenital tract, respiratory tract, (neuro-)endocrine tissues, choroid plexus and spinal cord.

Twelve oligo- or monospecific monoclonal antibodies (MoAbs) directed against human keratin types were used in an immunohistochemical study of the canine male and female urogenital tract, the respiratory tract, the adrenal gland, the (para-)thyroid gland, the choroid plexus and the spinal cord. The keratin MoAbs showed differences in staining patterns in the various epithelial tissues and the diverse epithelial cells. The kidney was characterized by a complex keratin staining pattern and the canine urothelium showed regional differences in keratin staining. Also in the female genital tract different keratin staining patterns were observed. Testicular and adrenal gland cells did not react with any of the keratin MoAbs. The keratin staining patterns in the various canine tissues showed, in addition to similarities, also distinct differences when compared to the staining patterns in corresponding tissues of other species, e.g. of man. These staining dissimilarities indicate that the reactivity patterns of the keratin MoAbs with restricted keratin immunoreactivity can not be always extrapolated from one species to another. Nevertheless, MoAbs directed against human keratin proteins can apparently be used to differentiate between various types of canine epithelia or epithelial compartments.

Keratin and vimentin distribution patterns in the epithelial structures of the canine anal region.

The intermediate filament labeling pattern of the epithelial structures of the canine anal region was studied with different polypeptide specific keratin monoclonal antibodies (MoAbs) and with a monoclonal and polyclonal vimentin antibody. The epithelial structures in this region could be discriminated and characterized by differences in their keratin staining pattern. The basal cells in the different epithelial structures showed a similar staining pattern characterized by reactivity with MoAbs staining keratins 5, 8, 14, and 17. Columnar epithelial cells showed a completely different phenotype mostly characterized by reactivity with MoAbs staining keratins 7, 5, 8, 18, and 19. A restricted number of differentiated perianal gland cells showed perinuclear vimentin staining. Myoepithelial cells did not stain for vimentin, but, as other basal cells, were positive for MoAbs staining keratins 5, 8, 14, and 17.

Immunohistochemistry with keratin and smooth muscle actin monoclonal antibodies in canine digestive tract and extramural glands.

The canine digestive system and its extramural glands (parotid gland, liver, pancreas) were immunohistochemically studied using a panel of twelve monoclonal antibodies (MoAbs) specific for human keratin proteins and for alpha-smooth muscle actin. Various epithelial tissues and cells were characterized by different keratin staining patterns. So, the epithelial lining of the upper alimentary tract was characterized by staining with the MoAb 6B10, specific for keratin-type (K) 4, and the absence of staining with the MoAbs directed against K 8 and 18 (CAM 5.2 and RGE 53, DE-K18 respectively), whereas the lower alimentary tract epithelium was not labeled by 6B10, but stained by the latter MoAbs. In the salivary glands the luminal and basal cells of the adenomeres as well as the different ductal structures could be immunohistochemically differentiated. The duct epithelium in liver and pancreas showed next to keratin staining characteristics in common with hepatocytes and exocrine pancreatic cells, additional staining by several keratin MoAbs. The keratin staining patterns in the canine tissues showed, in addition to similarities also distinct discrepancies when compared to the staining patterns in corresponding human tissues. Myoepithelial cells in salivary and oesophageal glands could be differentiated from other basally located epithelial cells by their exclusive immunoreactivity for alpha-smooth muscle actin. Canine pancreatic endocrine cells were not labeled by any of the keratin MoAbs. It is concluded that immunohistochemistry with polypeptide specific MoAbs specific for human keratin-types can be used to differentiate between different types of canine epithelial tissues and epithelial cells in the digestive tract. As a result such reagents may find their application in developmental biology and pathology of this species.

An immunohistochemical study of canine tissues with vimentin, desmin, glial fibrillary acidic protein, and neurofilament antisera.

In a wide range of canine tissues the immunoreactivity with commercially available antisera against intermediate filament antigens viz. vimentin, desmin, glial fibrillary acidic protein (GFAP) and neurofilament proteins, was studied. In addition, the results of formalin and Carnoy fixation were compared. Carnoy fixation appeared to result in optimal reactivity for all antisera. Epithelial cells did not react with any of the antisera, with exception of ovarian surface epithelium, which showed staining with the vimentin and desmin antisera. The vimentin antiserum induced staining of several cell types viz. fibroblasts, endothelial cells, chondrocytes, Schwann cells, ependymal cells, astrocytes, Leydig cells, synovial cells, podocytes and some parietal cells of Bowman's capsule. Sertoli cells showed a faint staining reaction. Muscle cells in various tissues reacted with the desmin antiserum. In the kidney a varying number of parietal cells appeared to react as did a restricted number of epithelial cells of proximal tubules and loops of Henle. GFAP reactivity was confined to glial cells, predominantly fibrous astrocytes, Schwann cells and axons. Additionally, some neuronal cell bodies in peripheral ganglia showed staining of varying intensity. Neurofilament staining was restricted to axons and some neurons. The immunoreactivity of canine tissues with these antisera is compared to findings in other species. The results confirm a broad interspecies cross-reactivity of these antisera. They can be used in studying the nature of canine tissues.

Keratin staining of canine epithelial tissues by a polyclonal antiserum.

The keratin distribution pattern in various canine epithelial tissues has been studied using a commercially available rabbit antiserum, raised against human skin keratins, in an immunoperoxidase staining method with the peroxidase-antiperoxidase complex (PAP). The staining results of two fixation methods were compared. Paraffin sections after fixation in Carnoy's solution showed optimal results, whereas paraffin sections after fixation in 10% buffered formalin resulted in a strongly reduced keratin staining reaction. Keratinizing and non-keratinizing stratified epithelial showed a strong staining reaction. Glandular epithelium and the non-stratified epithelia of internal organs e.g. liver, kidney and intestine did not react with the antiserum. However, glandular ductal epithelium, myoepithelial cells and basal cells in various epithelial tissues showed a positive staining reaction. The results indicate the presence of different keratin types in these canine epithelial tissues. The keratin distribution pattern is compared with the distribution pattern observed in tissues of other species.