Immunohistochemistry of carbonic anhydrase isozymes (CA-I, II and III) in canine salivary glands: a distributional and comparative assessment

The immunohistolocalization of carbonic anhydrase isozymes (CA-I, II, III) in canine salivary glands was studied using antiserum against CA-I, II, III. In parotid glands, immunostaining intensely localized cytosolic CA-II antiserum throughout the cytoplasm of acinar secretory cells and ductal epithelial cells, especially in the striated duct region. CA-III reactivity in the glands was only seen selectively at the intercalated ductal cells. In contrast, no immunoreaction localized CA-I in the gland. In the submandibular and sublingual glands, CA-I, II, and III were all observed in the ductal segments of the glands, whereas serous demilune appeared devoid of all three cytosolic CA isozymes. In contrast, in zygomatic glands (i.e. dorsal buccal glands) all CA isozymes were observed in both serous demilune and ductal segments. In all of the salivary glands examined, no mucous acinar cells were found to be reactive for any CA.     

Immunohistochemistry of the bovine secretory carbonic anhydrase isozyme (CA-VI) in bovine alimentary canal and major salivary glands

In the present study, we firstly demonstrated immunohistochemical expressions of secretory carbonic anhydrase (CA-VI) isozyme in bovine forestomach, large intestine and major salivary glands. CA-VI was detected in basal layer epithelial cells of esophageal and forestomach stratified epithelium, in mucous cells of upper glandular region of large intestine, in serous acinar cells of the parotid gland, in serous demilune cells and some ductal liner cells of mandibular, monostomatic sublingual and esophageal glands. These immunohistolocalizations suggested that bovine CA-VI plays various roles in pH regulation, maintenance of ion and fluid balance, and cell proliferation.     

A pathological study of the salivary glands of rabid dogs in the Philippines

Rabies is a zoonotic disease caused by the rabies virus. While the salivary glands are important as exit and propagation sites for the rabies virus, the mechanisms of rabies excretion remain unclear. Here, we investigated the histopathology of the salivary glands of rabid dogs and analyzed the mechanism of excretion into the oral cavity. Mandibular and parotid glands of 22 rabid dogs and three control dogs were used. Mild to moderate non-suppurative sialadenitis was observed in the mandibular glands of 19 of the 22 dogs, characterized by loss of acinar epithelium and infiltration by lymphoplasmacytic cells. Viral antigens were detected in the mucous acinar epithelium, ganglion neurons and myoepithelium. Acinar epithelium and lymphocytes were positive for anti-caspase-3 antibodies and TUNEL staining. In contrast, no notable findings were observed in the ductal epithelial cells and serous demilune. In the parotid gland, the acinar cells, myoepithelium and ductal epithelium all tested negative. These findings confirmed the path through which the rabies virus descends along the facial nerve after proliferation in the brain to reach the ganglion neurons of the mandibular gland, subsequently traveling to the acinar epithelium via the salivary gland myoepithelium. Furthermore, the observation that nerve endings passing through the myoepithelium were absent from the ductal system suggested that viral proliferation and cytotoxicity could not occur there, ensuring that secretions containing the virus are efficiently excreted into the oral cavity.     

Histology and histochemistry of the major salivary glands in the southern white-breasted hedgehog (Erinaceus concolor)

This study aimed to investigate the major salivary glands in the southern white-breasted hedgehog (Erinaceus concolor) histologically and histochemically. Five adult males were included in this study. The results showed that anatomically the shape of the parotid, submandibular and sublingual glands of the Erinaceus concolor was respectively almost pear, elliptical to pyramidal and oval. Histologically, the parotid gland had serous acini and secretory cells showed negative reaction to alcian blue (AB) (pH = 2.5) and negative response to aldehyde fuchsin (AF), methylene blue (MB) and PAS stains. The submandibular gland was a mixed gland of mucous and serous acini. The mucous acini were strongly positive for PAS, AB, MB and AF. However, serous acini were week for PAS and negative against AB, MB and AF stains. The sublingual gland was purely composed of mucous acini. The mucous acini of the sublingual gland were strongly positives for PAS, AB and MB methods. While their reaction to the AF staining was negative. In conclusion, the histological and histochemical observations of the major salivary glands of the southern white-breasted hedgehog (E. concolor) indicated that these glands shown similarities and some special different histochemical features as compared to other mammalian species.     

Lectin histochemistry of dog major and minor salivary glands

The distribution of different carbohydrates in dog major and minor salivary glands was investigated using a peroxidase-labelled avidin-biotin method to demonstrate binding of six lectins (Canavalia ensiformis agglutinin [Con A], Dolichos biflorus agglutinin [DBA], Arachis hypogaea (peanut) agglutinin [PNA], Glycine max agglutinin [SBA], Tetragonolobus purpurea agglutinin [TGP] and wheat germ agglutinin [WGA]). With PNA, there was only weak staining in serous acini of parotid glands. Other lectins bound, to different degrees, to different components of the salivary glands; differences could be detected between glands and between binding of different lectins to serous and mucous acinar cells and to the epithelial cell cytoplasm, luminal surface and contents of ducts. These results provide a basis for the comparison of possible changes in carbohydrates which may occur in salivary gland diseases.     

Lectin Histochemistry of Glycoconjugates in Horse Salivary Glands

The glycoconjugate content of major horse salivary glands was investigated by means of horseradish peroxidase-conjugated lectins. Qualitative differences were observed in the terminal sugar residues of secretory glycoproteins and glycoconjugates linked to the apical surface of excretory duct epithelial cells. Mucous acinar cells in mandibular and sublingual glands contained oligosaccharides with D-galactose, α- and β-N-acetylgalactosamine, N-acetylglucosamine and fucose residues, whereas mandibular, sublingual and parotid serous cells contained only oligosaccharides with terminal α- and β-N-acetylgalactosamine residues. The apical portion of striated and interlobular duct lining cells of mandibular and sublingual glands stained for α- and β-N-acetylgalactosamine and for N-acetylglucosamine. In parotid gland the cytoplasm of intercalated duct cells and the apical surface of striated duct epithelial cells stained for α-N-acetylgalactosamine.     

Immunohistochemical localization of carbonic anhydrase isoenzymes in salivary gland and intestine in adult and suckling pigs

Localizations of carbonic anhydrase isoenzymes (CA I, CA II and CA III) were investigated immunohistochemically in the salivary glands and intestine of mature and suckling pigs. Carbonic anhydrase isoenzymes were not detected in the salivary glands of sucklings, but were present in the adult. Bicarbonate ion in saliva might be important for the digestion of solid foods in mature pigs, but unnecessary for the digestion of milk in sucklings. Expressions of CA I and CA II were detected strongly in the large intestine of the adult and sucklings, and faintly only at duodenum in the small intestine. CA I and CA II isoenzymes in the large intestine may be involved, at least in part, in ion absorption and water metabolism during digestion and absorption of milk in suckling pigs. In addition, CA I and CA II expression in the duodenal villus enterocyte may support the process of bicarbonate absorption in the duodenum.     

Comparison of the distribution of carbonic anhydrase isozymes (CA-I, CA-II, CA-III) in the rat gastrointestinal tract.

The present paper described the immunohistochemical distributions of carbonic anhydrase (CA) isozymes. CA-I, CA-II and CA-III, in the epithelium lining the rat gastrointestinal tract, with rabbit antibodies to equine CA-I, CA-II and CA-III. Prior to the immunohistochemical examinations, the crossreactivities of these antibodies to the rat-antigens were confirmed in this study. In the stomach, surface epithelial cells and parietal cells of the glandular region showed an immunoreactivity only to CA-II. In the large intestine, each immunoreactivity to CA isozyme (CA-I, CA-II and CA-III) was localized in the upper portion of intestinal glands, and decreased toward the distal digestive tract, but absent in the small intestine. The present histological findings suggested that the CA isozymes might play a role in the ion-transportation during the water absorption in the rat large intestine.     

Histochemistry of sialoglycoconjugates in goat submandibular glands

The functional properties of sialic acids appear to be manifold. Therefore, they are considered as essential components of saliva. In this study, the localization of sialoglycoconjugates in the submandibular glands of Japanese miniature (Shiba) goat was examined by light and electron microscopic histochemical methods. The submandibular glands exhibited a large amount of sialic acids. Additionally, sialic acids with O-acetyl substitutions were detectable in the mucous acinar cells and serous demilunar cells. According to lectin histochemical methods, the mucous and serous cells mainly contained the Siaα2-6Gal/GalNAc sequence. These sialoglycoconjugates generated by the submandibular glands may specifically participate in the maintenance of the viscoelastic properties of saliva, protection of oral tissues and prevention of pathogenic microbial attacks. Therefore, our results suggest that they are essential components of saliva to maintain oral health.     

Idiopathic salivary gland enlargement (sialadenosis) in dogs: a microscopic study

A histological, histochemical and morphometric study was performed on submandibular salivary glands from 13 dogs which had presented with a submandibular mass or swelling that proved to be a portion of non-inflammatory and non-neoplastic submandibular salivary gland. There were no consistent changes in lectin-binding histochemistry or immunohistochemical expression of various cell markers, and, in most cases, there was no measurable difference in acinar size in the affected gland. The possible explanation for the clinical salivary gland enlargement is therefore unclear.     

Immunohistochemical localization of CB1 receptor in canine salivary glands

CB1 is a member of the G-protein-linked receptor superfamily that is present in the central nervous system as well as in certain peripheral neuronal and non-neuronal tissues. Recently, the presence of CB1 was found in the ductal system of the major salivary glands of laboratory animals, but no data are available for domestic mammals. Thus, in the present study, we examined the presence and distribution of CB1 in the major salivary glands of dogs using immunohistochemical techniques. CB1 was found in the parotid and mandibular glands of adult dogs; positive immunoreaction was localized to the cells of the striated ducts, with a peculiar localization on or near the apical membrane. This particular localization may be explained by the characteristics of this receptor as membrane-associated. The acinar structures were completely negative for CB1. We conclude that CB1 is involved in the control of dog salivary secretion via endogenous substances, likely endocannabinoids. The localization of CB1 highlights that endocannabinoids promote qualitative and/or quantitative changes of the primary saliva in the ductal system.     

Cytoarchitectural differences of myoepithelial cells among goat major salivary glands

Previously, the distribution of myoepithelial cells (mecs) in the salivary glands was studied by both immunohistochemistry, and transmission electron microscopy; however, little was elucidated concerning their morphological features, especially in goats. This study was performed to investigate the correlation between the cytoarchitecture of the mecs in goat major salivary glands (parotid, mandibular, and sublingual glands) and the nature of the saliva secretion. The cytoarchitectural features of the mecs were examined by scanning and transmission electron microscopy as well as immunohistochemically. The secretory endpieces in the parotid gland are of the pure serous type, but in both the mandibular and sublingual glands they are of the mixed type. In all studied glands, the intercalated ducts were covered by mecs which, unlike the large stellate cells that surrounded the secretory endpieces, were spindle-shaped with few cytoplasmic processes. Interestingly, the mecs were found to bulge on the basal surfaces of the serous acini and intercalated ducts in all glands and to be in close contact to the seromucous tubules surface in the mandibular and sublingual glands forming a continuous network around it. In conclusion, the differences in the degree of development of the mecs as well as the number of their cytoplasmic processes may be correlated with the nature of the secretion and the number of the secretory granules. Thus these observations may have some relevance in the diagnosis of atrophy and pathogenic conditions of these glands.     

Measurement of carbonic anhydrase isozyme VI (CA‐VI) in swine sera,colostrums, saliva,bile, seminal plasma and tissues

Swine secretory carbonic anhydrase VI (CA‐VI) was purified from swine saliva and an antibody to CA‐VI was generated. A specific and sensitive enzyme‐linked immunosorbent assay (ELISA) has been developed for the measurement of swine CA‐VI. The assay can detect as little as 5 ng/mL of swine CA‐VI. Typical standard curves were determined for a range of CA‐VI solutions (7.8 to 500 ng/mL). The coefficients of variation for these solutions were less than 5%. When 500, 250 or 100 ng/mL of swine CA‐VI was added to swine sera, the recoveries were 102.0%, 109.7% and 100.2%, respectively. The concentrations of CA‐VI in the saliva (26.2 ± 30.4 µg/mL), sera (3.3 ± 4.9 ng/mL), bile (153.0 ± 114.0 ng/mL), seminal plasma (124.0 ± 39.0 ng/mL) and parotid gland (441.3 ± 90.0 µg/g wet tissue), submaxillary gland (88.1 ± 124.4 µg/g wet tissue), sublingual gland (58.6 ± 24.6 µg/g wet tissue) and gallbladder (2.4 ± 1.3 µg/1g wet tissue) were determined by ELISA. The concentration of CA‐VI in colostrum was 163.3 ± 101.4 ng/mL and did not decrease within 10 days following parturition. An immunohistochemical reaction to anti‐CA‐VI antiserum was observed in the columnar epithelial cells lining the gallbladder. These data suggest that secretory CA‐VI plays various roles in pH regulation and the maintenance of ion and fluid balance.     

Postnatal development of the mouse volatile papilla taste bud cells

In the present study, we examined specific markers for taste bud cells in the mouse and the postnatal development of volatile papilla taste bud cells in ddY mice. We examined the immunoreactivity of 4 types of carbonic anhydrase isoenzymes, CA I, CA II, CA III and CA VI, as specific markers for taste bud cells, and K8.13 cytokeratin antibody as a specific marker for the lingual epithelial cells. Of the carbonic anhydrase isoenzymes, only CA III immunoreactivity was clearly detected in the spindle shaped gustatory cells. CA VI immunoreactivity was detectable in suspentacular cells. CA I and CA II antibodies did not recognize any taste bud cell specifically. K8.13 cytokeratin immunoreactivity was detected in the lingual epithelial cells, but not in taste bud cells. At 7 days after birth, the suckling phase, very small taste buds developed from the anaplastic gustatory cells. At 14 days after birth, the taste buds showed larger size than those at 7 days after birth. At 21 days birth, after the weaning phase, taste bud structure approximated the mature structure. These results demonstrate the specificity of anti-CA III and anti-CA VI for gustatory cells and suspentacular cells, respectively. These markers should be useful for an analysis of taste bud development in mice.     

Preliminary studies of the development of Anaplasma marginale in salivary glands of adult, feeding Dermacentor andersoni ticks

On each day of feeding on susceptible calves, salivary glands obtained from groups of adult ticks that transmitted Anaplasma marginale were examined for A marginale colonies by use of light microscopy and transmission electron microscopy. On day 8 of feeding, salivary glands were examined, using fluorescein-labeled antibody and methyl green-pyronine stain. Use of fluorescein-labeled antibody consistently revealed small numbers of fluorescent foci in salivary gland acinar cells obtained from ticks that had fed for 8 days. Colonies of A marginale were seen by transmission electron microscopy only in salivary gland acini of male ticks; these colonies could not be identified, using light microscopy, in companion 1-micron plastic sections stained with Mallory stain. Methyl green-pyronine stain, used commonly to detect theilerial parasites in tick salivary glands, did not differentiate A marginale from cytoplasmic inclusions normally found in salivary gland acinar cells.     

Glycoconjugates in the mandibular salivary gland of adult dogs revealed by lectin histochemistry

The glycosidic residues in the mandibular glands of five adult dogs were studied by using seven different lectin-horseradish peroxidase conjugates. In some cases a treatment with sialidase preceded the lectin staining. The mucous acinar cells contained oligosaccharides with α- and β-N-acetylgalactosamine, N-acetylglucosamine and fucose residues, whereas the demilunar cells contained glycoconjugates rich in sialic acid linked to the penultimate disaccharide galactosyl((β1→3) N-acetylgalactosamine.