Precocious differentation of mouse parotid glands and pancreas induced by hormones

Changes of α-amylase activity (1,4-α-D-glucan glucanohydrolase, EC 3.2.1.1) in mouse parotid gland pancreas were investigated during embryonic and postnatal development. Amylase activity in the parotid gland increased from around day 12 and reached the adult level on day 30. On the other hand, the activity in the pancreas increased during the last stage of gestation, decreased after birth, and then gradually increased from around day 15, reaching the adult level on day 35. Precocious differentiation of the parotid gland was induceed by injections of hydrocortisone or thyroxine after birth, but these hormones did not have additive effects on the parotid gland. Injection of insulin had little effect when given alone, but suppressed the effects of the other two hormones on the gland. Only hydrocortisone increased the amylase activity in mouse pancreas during postnatal development, the other two hormones causing slight decrease in pancreatic amylase. Adrenalectomy and injection of hydrocortisone affected the parotid gland but not the pancreas of adult mice. These results suggest that hydrocortisone involved in cytodifferentiations of the parotid gland and pancreas, and in maintenance of the parotid gland. Thyroxine may also be important in differentiation of the parotid gland.     

Effects of hormones on differentiation of parotid glands of suckling mice in vitro

The hormonal requirements for functional differentiation of mouse parotid glands were investigated using organ cultures in chemically defined medium. The hormones tested were insulin, thyroxine and prednisolone, and the parameters examined were alpha-amylase activity and the ultrastructure of the tissue. It is found that most of the amylase in the cultures (80%) was released into the culture medium after 5 days of cultivation. Prednisolone (5 . 10(-3) mg/ml) alone resulted in a 3--4-fold increase in specific activity of amylase (total amylase activity in the medium and culture) over that in its absence, but neither insulin nor thyroxine alone induced the enzyme. Prednisolone plus thyroxine (over 1 . 10(-7) mg/ml) or insulin (over 1 . 10(-3) unit/ml) induced markedly the enzyme, amylase specific activity being as much as 4- or 6-fold that with prednisolone alone. Moreover the enzyme specific activity was dependent on the prednisolone concentration (5 . 10(-7) - 5 . 10(-3) mg/ml) in the presence of thyroxine (1 . 10(-2) mg/ml) or insulin (1 . 10(-2) unit/ml). Morphological differentiation was also observed in explants cultivated in medium containing prednisolone plus thyroxine or insulin. These results suggest that besides glucocorticoids, insulin and thyroxine are involved in increase in amylase activity in mouse parotid glands during the late suckling period.     

Ultrastructural changes of rat parotid glands induced by a diet of liquid Metrecal

Summary The ultrastructure of parotid glands was studied in rats fed a diet of liquid Metrecal for two weeks and compared with that of parotid glands of control rats which received a diet of Purina lab chow. The liquid diet induced major alterations of acinar cells, but other parenchymal components were apparently unaffected.Most acinar cells of experimental rats were atrophic and some of these were undergoing necrosis. Lipid droplets and dense bodies (believed to be lysosomes) were numerous in atrophic cells. The Golgi apparatus, quantity of secretory granules, and intercellular canaliculi were smaller than in acinar cells of control rats.Such findings suggest that the secretory process was impaired and support the conclusion that parotid glands of rats maintained on a liquid diet are physiologically less active than those of chow fed rats. The decreased activity, as previously reported, may result from reduced masticatory activity.Supported by U.S.P.H.S. grant DE 02110.     

Lactose synthetase activity in mouse mammary glands is controlled by thyroid hormones

Epithelial cells in explants from the mammary glands of euthyroid mature virgin mice are proliferatively dormant. They must undergo DNA synthesis and traverse the cell cycle in vitro before they are able to differentiate fully in response to insulin, hydrocortisone, and prolactin, and synthesize enzymatically active alpha-lactalbumin (measured as lactose synthetase activity). In contrast, glands from hyperthyroid mature virgin mice do not require DNA synthesis in vitro to differentiate. Explants from the euthyroid virgin tissue overcome their dependence on DNA synthesis when 10(-9) M 3,5,3'-triiodo-L-thyronine is added directly to the cultures in addition to the other three hormones. Explants from involuted mammary glands from euthyroid primiparous mice do not require DNA synthesis in vitro to make the milk protein even though they, like explants from mature euthyroid virgin tissue, are proliferatively dormant and do not contain detectable lactose synthetase activity in vivo. Glands from primiparous animals made mildly hypothyroid by ingestion of 0.1% thiouracil in drinking water during 7 wk of involution remain morphologically indistinguishable from glands of their euthyroid counterparts. However, explants from the glands of these hypothyroid animals revert to a state of dependence on DNA synthesis to differentiate functionally. These observations suggest that the dependence on DNA synthesis and cell cycle traversal for hormonal induction of lactose synthetase activity in the mouse mammary gland is controlled by thyroid hormones.     

Precocious differentiation of chick embryo pancreas in vitro: Roles of prednisolone,insulin and L-thyroxine

The hormonal requirements for functional differentiation of chick embryo pancreas were investigated by using organ cultures in chemically defined medium. The hormones tested were prednisolone, insulin and thyroxine, and the parameters examined were α-amylase (EC 3.2.1.1) and chymotrypsinogen (EC 3.4.4.5) activities, and the ultrastructure of the tissues. Addition of prednisolone alone to explants from 14-day-old chicken embryo pancreas for 3 days increased the activities of amylase and chymotrypsinogen in the tissues by 3.4- and 6.6-fold, respectively, those of tissues before cultivation. Neither thyroxine or insulin alone, nor both hormones together affected pancreatic exocrine differentiation. Thyroxine enhanced the effect of prednisolone on both enzymes, but insulin did not. When the explants were cultured in the medium containing all three hormones, maximum enzyme activities were observed; amylase or chymotrypsinogen activity being 7- or 18-fold, respectively, that of tissues before cultivation. But these three hormones were not simultaneously necessary. Morphological differentiation was also observed in explants cultivated in medium containing these three hormones. These results suggest that glucocorticoids are essential for normal differentiation of chick pancreas during the late fetal period, possibly with insulin and thyroxine, and also support the idea that pancreatic enzymes are controlled separately.     

Secretory character of a group of isoproterenol-induced polypeptides in mouse parotid glands

The secretory nature of the isoproterenol-induced mouse parotid polypeptides C, D, E, F, and G (molecular weights 64,000, 61,000, 51,500, 38,000, and 37,000, respectively) is documented. Polypeptides C, D, E, F, and G, accumulated in response to successive daily stimulations with isoproterenol, were detected in a fraction enriched in hypertrophic parotid acinar cells. These cells, characterized by an increased content of cytoplasmic granules, maintain a secretory responsiveness to isoproterenol, which has been evidenced by light microscopy, enzymatic analysis, and unidimensional SDS-polyacrylamide gel electrophoresis. Thus, a parallelism in the loss and recovery of both secretory granules, alpha-amylase and polypeptides C, D, E, F, and G, was observed. Moreover, after secretion stimulation, polypeptides C, D, E, F, and G were detected in the fluid collected directly from parotid gland cannulation. Given the secretory character of polypeptides C, D, E, F, and G, mechanisms explaining both their progressive accumulation along the chronic administration of isoproterenol, as well as their progressive disappearance observed after suspending that treatment, are discussed.     

Recretion of enzymes and hormones by exocrine glands

The article reviews a poorly explored issue of secretive physiology-recretion from blood by glandulocites of various endocrinal glands of hydrolytic ferments and hormones that have been synthesized by digestive and endocrinal glands. The article discusses potential physiological role of the recretion function and the diagnostic significance of information obtained from analysis of recreted ferments and hormones in exosecretions.     

Precocious differentiation of chick embryo pancreas in vitro. Roles of prednisolone, insulin and L-thyroxine.

The hormonal requirement for functional differentiation of chick embryo pancreas were investigated by using organ cultures in chemically defined medium. The hormones tested were prednisolone, insulin and thyroxine, and the parameters examined were alpha-amylase (EC 3.2.1.1) and chymotrypsinogen (EC 3.4.4.5) activities, and the ultrastructure of the tissues. Addition of prednisolone alone to explants from 14-day-old chicken embryo pancreas for 3 days increased the activities of amylase and chymotrypsinogen in the tissues by 3.4- and 6.6-fold, respectively, those of tissues before cultivation. Neither thyroxine or insulin alone, nor both hormones together affected pancreatic exocrine differentiation. Thyroxine enhanced the effect of prednisolone on both enzymes, but insulin did not. When the explants were cultured in the medium containing all three hormones, maximum enzyme activities were observed; amylase or chymotrypsinogen activity being 7- or 18-fold, respectively, that of tissues before cultivation. But these three hormones were not simultaneously necessary. Morphological differentiation was also observed in explants cultuvated in medium containing these three hormones. These results suggest that glucocorticoids are essential for normal differentiation of chick pancreas during the late fetal period, possibly with insulin and thyroxine, and also support the idea that pancreatic enzymes are controlled separately.     

Ultrastructure of bovine parotid glands

Bovine parotid glands exhibit outstanding structural differences when compared with those of non-ruminant mammals. The acini are tortuous, branched and lined with cells of different heights, imparting a scalloped appearance to acinar lumina. Numerous microvilli, ca. 1.5 μ in length, extend into the lumina and intercellular canaliculi. Intercellular canaliculi measure ca. 3 μ in diameter and interweave in close association with intercellular tissue spaces. Intercellular tissue spaces are separated from the extraacinar spaces across a basal lamina only, whereas junctional complexes guard canaliculi from direct continuity with tissue spaces and/or extraacinar spaces. Flattened cytoplasmic lamellae extend from adjacent acinar cells and loosely interdigitate with one another across the tissue spaces. Acinar cells contain more mitochondria and less granular endoplasmic reticulum than parotid glands of non-ruminant mammals. Two types of secretory material, in the form of inclusions which vary in size and electron density, are present in the acinar cells. Intercalated ducts connect acini with striated ducts which in turn, empty into collecting ducts located between gland lobules. In terms of frequency of “basal infoldings” and numbers of mitochondria, striated ducts of calf parotid glands are not as well developed as those of certain other salivary glands. Myoepithelial cells are most often present at junctions of acini and intercalated ducts where they may attach to both acinar and ductal epithelium. Nerve “terminals” were not observed on the epithelial side of basement membranes in relation to the secretory cells.     

Changes in the polypeptide composition of mouse parotid glands after stimulation of secretion and DNA synthesis by isoproterenol

The polypeptide composition of mouse parotid glands has been analysed by unidimensional SDS-polyacrylamide slab gel electrophoresis and Coomassie blue staining after isoproterenol stimulation of secretion and DNA synthesis. Two polypeptides (polypeptides A and B) are lost within 2 h and their restoration in the glands occurs according to a chronology which is identical to that of the alpha-amylase activity. On the other hand, five clearly defined new bands appear consistently during the late prereplicative period of isoproterenol-stimulated mouse parotid acinar cells (polypeptides C, D, E, F and G). These new polypeptides are induced by doses of isoproterenol which provoke secretion and DNA synthesis, but not by doses which provoke only secretion. Although no function has been assigned to any of the above-described polypeptides, a relation between polypeptides A and B and secretion and between polypeptides C, D, E, F and G and the proliferative response is suggested.     

Involvement of apoptosis and proliferation of acinar cells in atrophy of rat parotid glands induced by liquid diet

Parotid glands of experimental animals fed a liquid diet are reported to show atrophy (Hall and Schneyer 1964; Wilborn and Schneyer 1970; Hand and Ho 1981; Scott et al. 1990; Scott and Gunn 1991). To clarify whether apoptosis and proliferation of acinar cells participate in atrophy of rat parotid glands induced by liquid diet, rats were fed a liquid diet and compared to pellet-fed controls. Parotid glands were removed at 3, 7, 14 or 21?days, weighed, and examined using transmission electron microscopy (TEM), and studied immunohistochemically for cleaved-caspase-3 (Casp-3), a marker of apoptotic cells, and 5-bromo-2′-deoxyuridine (BrdU), a marker for proliferating cells. Body weights of experimental rats fed liquid diets were not significantly different from controls fed pellet diets; however weights of experimental parotid glands were smaller than those of controls. In the experimental parotid glands, structures like apoptotic bodies were histologically observed in acini at each time point; more Casp-3-positive acinar cells were identified in experimental parotid glands than in the controls on days 3, 7, and 14. Experimental glands showed fewer BrdU-positive acinar cells at each time point. TEM confirmed typical apoptotic acinar cells in the atrophic glands. These findings suggest that increased acinar cell apoptosis and reduced acinar cell proliferation occur in atrophic parotid glands of rats fed a liquid diet.     

Electron microscopic immunocytochemical localization of proline-rich proteins in normal mouse parotid salivary glands

Summary Rabbit polyclonal antibodies against isoproterenol-induced mouse proline-rich proteins (PRPs) were used to localize PRPs in the parotid salivary glands of normal adult BALB/c mice. The antibodies recognized both acidic-type and basic-type PRPs. Immunoblotting experiments revealed that the glands contained an acidic-type and a basic-type PRP. Parotid gland tissue was fixed with Karnosky's fixative and embedded in Lowicryl resin at low temperature. PRPs were localized at the electron microscope level using an indirect post-embedding staining technique with protein A-gold. The secretion granules of the acinar cells were strongly labelled. Pre-absorption of the antibody with purified acidic-type and basic-type PRPs indicated that the basic-type PRP is mainly located at the periphery of the granules but that the acidic-type PRP is more evenly distributed within the granules. Pre-absorption of the antibody with -amylase did not affect the staining pattern, suggesting minimal cross-reactivity. PRPs were also detected within the rough endoplasmic reticulum and the Golgi apparatus of acinar cells, within the granules of the proacinar cells and in the lumena of the ducts, but not within the intercalated or striated duct cell granules.     

Precocious induction of pepsinogen in the stomach of suckling mice by hormones

Effects of hormones on pepsinogen activity in mouse stomach were investigated by enzyme assay and electron microscopy. Administration of hydrocortisone alone to mice on days 5–10 increased the enzyme activity in the stomach to as much as 4.5-fold that of untreated mice and the increase was dose dependent. Thyroxine also evoked precocious differentiation of the stomach. The effects of thyroxine and hydrocortisone were additive. Injections of insulin had little effect when given alone, or in combination with other hormones. Injection of hydrocortisone alone or plus thyroxine also caused morphological differentiation of the chief cells in the stomach mucosa. Administration of thyroxine to mice on days 15–20 induced as much enzyme activity as that induced by hydrocortisone, but neither of these hormones had any effect when injected after day 23.These results suggest that besides hydrocortisone, thyroxine is also involved in differentiation of the stomach in mice for the first 20 days after birth and that the normal increase of pepsinogen activity in the stomach of mice during the late suckling period is brought about by serum glucocorticoids, possibly with thyroxine.     

Precocious decrease of chymotrypsinogen by thyroxine in the pancreas of suckling mice

Chymotrypsinogen activity in mouse pancreas gradually decreased from birth, reaching the adult level on day 20 after birth. In suckling mice the enzyme activity was decreased to 1/9 of the control value by injection of thyroxine. The activity was not affected by insulin, and was slightly increased, rather than decreased, by daily injection of hydrocortisone. The effect of thyroxine seemed to be direct, not due to modification of adrenal function.