Immunohistochemical and ultrastructural correlates of muscle-actin expression in pleomorphic adenomas and myoepitheliomas based on comparison of formalin and methanol fixation.

The degree and range of differentiation of the cells referred to as myoepithelial-like in pleomorphic adenomas and the tumour cells of myoepitheliomas are not definitely established. This type of information is critical for establishing reliable diagnostic criteria, such as expression of muscle-specific actin and ultrastructural identification of myofilaments, in these and other salivary gland tumours. Pleomorphic adenomas (18) and myoepitheliomas (5), of which 10 cases were fixed only in formalin and 13 cases where tissues were fixed in both formalin and methanol/acetic acid, were studied. Each tumour and normal accompanying parotid was immunostained with two monoclonal antibodies for smooth muscle actin, HHF35 and MSA. Staining of myoepithelial cells was absent in certain samples of normal gland with both HHF35 (15%) and MSA (69%) when formalin-fixed tissue was used. Using formalin-fixed tissue from 15 pleomorphic adenomas/myoepitheliomas, 2 (14%) had focal positivity with HHF35, while 8 cases (57%) were positive with MSA. However, a certain degree of false positivity was suspected since in samples of normal parotid, both acinar and duct cells were frequently stained, particularly with MSA. With methanol/acetic acid-fixed tissue only 4 of 13 cases (31%) were positive with either MSA or HHF35 and 2 of these only had a minor proportion of the tumour cells expressing muscle-specific actin. Using alcohol-fixed tissue, myoepithelial cells were strongly stained in all examples of normal parotid gland with both anti-actin antibodies. In 5 cases examined by electron microscopy, there was no apparent correlation between immunohistochemical results and the presence or absence of cytoplasmic filament accumulation. The results indicate considerable tumour cell heterogeneity in muscle-specific actin expression and suggest that non-luminal cells in pleomorphic adenomas and the tumour cells in myoepitheliomas may differentiate as classical myoepithelial cells, as partially differentiated (i.e. modified myoepithelial cells) or as the counterpart of basal cells present in the intra- and interlobular ducts of normal salivary gland.

The mechanism of ceruminolysis.

In a previous study comparing the efficacy of a selection of commonly used ceruminolytics, the authors noted that aqueous-based preparations, and in particular solutions of sodium bicarbonate, were more effective in disintegrating cerumen than most organic-based preparations. In that study, the authors also observed that not only had the wax truly disintegrated following exposure to the aqueous-based preparations, but also that a marked degree of swelling of the wax spheres had occurred with these preparations. In this paper the mechanism of ceruminolysis was investigated by means of a number of commonly available histological techniques. Our findings show that desquamated sheets of corneocytes are the major constituent of cerumen plugs and provide the structural framework of the wax bolus. Ceruminolytics work by hydrating the keratin cells of these sheets of desquamated stratum corneum and subsequently inducing keratolysis, with disintegration of the wax.

Corneocyte architecture in the human external auditory canal.

Using skin dissected from the external auditory canals of fresh cadaveric temporal bones, the structure of the stratum corneum was investigated by the techniques of alkaline expansion of cryostat sections, silver staining of epidermal sheets, and scanning electron microscopy. Ordered epidermal column formation was observed in the stratum corneum from skin obtained from both the bony and cartilaginous portions of the canal. Occasionally, in areas of skin where the stratum corneum was considerably thickened, this vertical organization was lost, and the corneocytes randomly arranged. This orderly cellular architectural arrangement is a feature of the non-migratory skin of lower mammals, and has also been reported to occur in some areas of human skin. The functional relevance of this finding in the skin of the external canal is that the whole of the stratum corneum must migrate as one in order to preserve a regular vertical structure, suggesting that epidermal migration occurs in the deeper layers of the meatal epidermis.

Bone resorption in chronic otitis media. The role of mast cells.

Twenty-two surgical specimens of eroded middle ear ossicles were removed from patients with chronic otitis media, with and without cholesteatoma. By using specific mast cell stains, increased numbers of mast cells were found in connective or granulation tissue adjacent to eroded surface of the bone. Mast cells possess the biological machinery necessary for enhancing bone resorption, and the population density of mast cells is increased in a variety of disorders that are associated with bone resorption. It is hypothesized that mast cells contribute to bone resorption in chronic otitis media, and the possible mechanisms by which mast cells exert their action are discussed.

Mast cells in human middle ear mucosa in health and in disease.

The distribution of mast cells was studied in normal human middle ear mucoperiosteal lining and in middle ear biopsies of patients with acute and chronic otitis media. The mast cells were identified on the basis of the metachromatic staining for their cytoplasmic granules with Giemsa and toluidine blue. Only a few mast cells located in proximity to blood vessels in the lamina propria underneath the epithelial layer were observed in normal middle ear mucoperiosteum. The number of mast cells in acute inflammatory reactions and in the normal middle ear lining was similar. By contrast, the mast cell count was significantly increased in chronic inflammatory reactions. The population density of the mast cells was the highest in the subepithelial layer of cholesteatoma, in regions where the lamina propria showed fibrosis and infiltration with chronic inflammatory cells, and around mucous glands. The presence of increased numbers of mast cells in chronic otitis media is consistent with our previous finding of high levels of histamine in middle ear effusions. It is postulated that mast cells play an important role in the pathogenesis of chronic otitis media through the release of their active biochemical mediators.

Studies on the proliferation and fate of oval cells in the liver of rats treated with 2-acetylaminofluorene and partial hepatectomy.

The kinetics of oval cell proliferation in the liver and their fate were studied by combined autoradiography and immunohistochemical staining for epidermal prekeratin and epoxide hydrolase (EH). The oval cell proliferation was induced in rats by exposure to dietary 2-acetylaminofluorene (2-AAF) for 2 weeks with the midway performance of partial hepatectomy (PH). The labeling with 3H-thymidine [3H-TdR] was done in different groups of rats by two procedures: continuous exposure for 1 week with the aid of a minipump and brief exposure by the administration of a single dose. The livers of groups of animals were examined from 1 to 10 weeks after PH. Oval cells and duct epithelium showed positive staining for prekeratin and negative for EH, whereas hepatocytes showed the reverse pattern of staining. A critical finding was the observation that the exposure to the 2-AAF inhibited virtually completely the labeling of hepatocytes with [3H]-TdR in the caudate lobe and incompletely in the right lobe without interfering with the labeling of the oval cells in either lobe. This made it possible to study the fate of the oval cells vis-à-vis hepatocytes. This qualitative-quantitative study of oval cells and hepatocytes clearly indicates that oval cells under these experimental conditions do not become hepatocytes within 10 weeks. Over 80% of oval cells disappear within this period, and the remainder persist as such. These results indicate that under one set of experimental conditions related to hepatocarcino-genesis in the rat, no evidence for the conversion of oval cells to hepatocytes was obtained.

Comparison of formalin- and acetone-fixation for immunohistochemical detection of carcinoembryonic antigen (CEA) and keratin.

The purpose of the present study was to compare the relative merits of cold acetone and buffered formalin as fixatives for the detection of carcinoembryonic antigen and keratin in permanently embedded tissues using a peroxidase-antiperoxidase (PAP) immunohistochemical procedure. The effect of treatment with the proteolytic enzyme pronase also was examined in the formalin-fixed tissues. Cold acetone was found to be superior to formalin for the preservation of CEA and keratin antigenic activities in a variety of benign and malignant tissues. Pronase treatment markedly increased the staining intensities of both antigens in formalin-fixed tissues. For many tissues, however, superior results were obtained using the cold acetone method, and this technic is recommended for the optimum retention of antigenic activity in permanently embedded tissues.