Changes in the secretory acinar cells of the rat parotid gland during aging

The secretory acinar cells of parotid glands from rats of varying ages have been examined by electron microscopy to determine what age-related changes occur in these cells. The most prominent change noted in these cells is the progressive increase in the amount of lipofuscin granules with age. Lipofuscin granules are membrane-bound structures consisting of lipids, other subcomponents, and a matrix. In addition, these cells contain lipid droplets that are not associated with any other components and tend to accumulate at the base of the cells in older rats. Also, many acinar cells in the glands of old rats contain altered secretory granules which appear to be in the process of degeneration. The accumulation of lipid and degenerating secretory granules appears to be related to the reduced level of cellular secretory activity in the glands of older rats. It is possible that these two types of inclusions contribute to the formation of lipofuscin granules. Lipofuscin and degenerating secretory granules are associated with acid phosphatase, which is demonstrated cytochemically, indicating that these granules are lysosomal structures.     

Co-localization of rab4 with endocytosis-related proteins in the rat parotid glands

Small GTP-binding proteins have been implicated in the regulation of vesicular traffic. We investigated the localization of Rab4 in the rat parotid glands by Western blotting and light-microscopic immunohistochemistry. Rab4 was localized mainly on the intracellular membranes in the subapical-actin terminal web, but was not present in the basolateral region both in acinar and ductal cells. Actin, alpha-adaptin, Rab5A and aquaporin5 were detected in the Rab4-containing intracellular membrane fraction prepared using anti-Rab4 antibody covalently coupled to magnetic beads. Detection of actin indicated that the Rab4-containing intracellular membranes were attached to the actin filaments. Although alpha-adaptin was immunohistochemically distributed along the plasma membrane, this protein coexisted with Rab4 only at the apical region. Rab5A immunoreactivity was distributed all around the cytoplasm. These findings suggested that Rab4 participates in endocytosis at the apical membrane of parotid glands.     

The synthesis of amylase in parotid glands of young and old rats

The age-related changes in the rate of synthesis of total and secretory proteins were examined in parotid glands of young (2 months) and old (24 months) rats. The differences in the rate of incorporation of radioactive leucine into acid-insoluble proteins of the gland indicate that the rate of protein synthesis declines with age in this gland. To determine whether the rate of synthesis of secretory proteins changes with age in this gland, the rates of incorporation of [3H]leucine into amylase, a major secretory protein of the gland, were compared by radioactivity determinations. For this comparison, amylase was precipitated with glycogen after incubating the gland slices in the presence of the labeled amino acid. The study shows that rate of synthesis of amylase declines significantly with age in this gland. The possible relationship between the decline in protein synthesis and the reduced level of secretory activity of the gland due to aging is discussed.     

N-linked protein glycosylation in the rat parotid gland during aging

N-Linked protein glycosylation was examined in vitro in dispersed rat parotid acinar cells from young adult (3-6 months) and aged (22-24 months) rats. A small decrease in general protein production was observed with cells from aged animals (approximately 20% lower incorporation of [14C]leucine into 10% CCl3 COOH insoluble protein during continuous pulse labeling). Incorporation of [3H]mannose into N-linked glycoproteins by aged cells was further reduced (approximately 35%). Similarly microsomal membranes from parotid glands of aged animals showed approximately 50% reduction in the synthesis of mannosylphosphoryl dolichol, a key intermediate in the dolichol pathway of protein N-glycosylation. Man-P-Dol synthase, the microsomal enzyme responsible for production of this saccharide-lipid, displayed no change in apparent Km for GDP-mannose when preparations from aged animals were utilized, but did show approximately 50% reduction in Vmax. Following beta-adrenoreceptor activation, cells from both young adult and aged glands showed increased N-linked protein glycosylation almost to the same extent (approximately 2-fold). The data suggested that in aged rat parotid cells there is a basal reduction of activity in the pathway responsible for asparagine-linked protein glycosylation, but that following exocytotic stimuli this pathway responds in a manner comparable to cells from young adult glands.     

Parasympathetic non-adrenergic, non-cholinergic-induced protein synthesis and mitogenic activity in rat parotid glands

Electrical stimulation of the parasympathetic auriculo-temporal nerve (40 Hz, 30 min), in the anaesthetized rat under - and -adrenoceptor blockade, increased [3H]thymidine and [3H]leucine uptake into the parotid glands by 80 and 263 %, respectively. The increase in response to parasympathetic stimulation was almost the same ([3H]thymidine 82 % and [3H]leucine 283 %) when atropine (2 mg kg-1 I.P. or I.V.) was included in the pretreatment. Neither intravenous infusion of vasoactive intestinal peptide (0.5-20 mg kg-1 min-1, 30 min) nor of bethanechol (10 mg kg-1 min-1, 30 min), under adrenoceptor blockade, increased the uptake of [3H]thymidine into the glands. However, these drugs increased [3H]leucine uptake, and in combination they interacted positively. Whereas vasoactive intestinal peptide is likely to be involved in the parasympathetic nerve-evoked protein synthesis, the nature of the non-adrenergic, non-cholinergic component(s) involved in the mitogenic response is presently unknown.     

Changing myoepithelial cell distribution during regeneration of rat parotid glands

The distribution of the myoepithelial cells during regeneration of the rat parotid gland after atrophy induced by one week of parotid duct ligation was investigated by immunohistochemistry for actin and transmission electron microscopy (TEM). Immunohistochemically, residual ducts were surrounded by actin-positive cells when clips were removed from the duct. Three days later, most of the newly formed acini originating from the residual ducts were also embraced by actin-positive cells. After 10 days, actin-positivity tended to be seen as dots around acini that decreased in number day by day. On day 21 actin-positive cells mainly surrounded intercalated ducts with only a few positive reactions identified at the acinar periphery. Electron microscopically, residual ducts and newly formed acini were peripherally embraced by myoepithelial cells before day 5. After day 7, shift of myoepithelial cells from the periphery of acini to the duct-acinar junctional region was identified. Then few myoepithelial cells were identified at the periphery of acini. These observations indicate that myoepithelial cells migrate from the acinar periphery to the duct-acinar junctional region during rat parotid regeneration, and that such behaviour is closely related to that seen during rat parotid development.     

Changes in mitochondrial DNA during aging

Mitochondrial DNA (mtDNA) was isolated from liver mitochondria of rats between 2 and 24 months of age. The mtDNA was purified by cesium chloride—ethidium bromide isopycnic density gradient centrifugation. In the gradients, in addition to the two expected bands of ethidium-DNA complex, there was observed a third, more dense band ($\text{d = 1.69}\text{g}\text{cm}{}^3$). This novel band, rarely observed in preparations from younger animals, was present in most preparations from older animals. The latter was characterized using the diphenylamine assay(s) and ascertained to contain DNA and carbohydrate components. Agarose gel electrophoresis revealed the DNA of the novel band to have a migration identical to form I mtDNA. Digestion of the novel band with the restriction endonuclease Bam HI yielded products identical to those obtained upon treatment of form I mtDNA with Bam HI. The observation of mtDNA at a density of $\text{1.69 g}\text{cm}{}^3$ indicates the presence, predominantly in older animals, of a subclass of mtDNA molecules with altered ethidium binding properties. The significance of this mtDNA and its position in the gradient is unclear at this time.     

Chaperones in the parotid gland: localization of heat shock proteins in human adult salivary glands

Heat shock proteins (HSPs) are expressed or increased in response to various biological stresses. Moreover, these 'stress proteins' seem to be expressed by some cells living in physiological conditions. From then on, they could play an important physiological role in normal cell functioning. The best-known physiological role of these HSP proteins is to act as 'molecular chaperones'. In this context, we have investigated the immunohistochemical expression of HSP27, HSP70, HSP90 and HSP110 in 10 human adult salivary glands. To highlight the presence of RNAm encoding HSP70, an in situ hybridization was performed. In our material, HSP27 was strongly expressed in the cytoplasm of striated duct cells and in some myoepithelial cells. The same localization was less stained for HSP70 and HSP90. The immunocytochemical reaction was weak or negative for HSP110 in striated ducts. HSPs were not expressed in acinic cells. In situ hybridization gave a positive signal in striated ducts with a probe encoding HSP70. Epithelial cells of the striated ducts and myoepithelial cells expressed HSP27, HSP70 and HSP90. These HSPs probably act in part as molecular chaperones for protein synthesis, transport and for several interactions between HSPs and different proteins.     

Morphological changes in collagen fiber during development of human fetal parotid and submandibular glands

Parotid and submandibular glands from human fetuses (16, 20, 24, 28, 32 weeks of gestation) were examined under a scanning electron microscope. Changes were found in the arrangement of collagen fibers in the connective tissue surrounding the salivary gland. In particular, several layers around the salivary gland were formed by a collagen network structure. These structures, although in varied arrangements, were recognizable in each stage of fetal growth. They are thought to play the role of a "cushion" against pressure created by accumulation of granules because of the reflex activity of myoepithelial cells during secretion. These structural changes are related to the mechanical performance of granule formation in the salivary gland and secretion during the development of the fetus.     

Hormonal function of the adrenal glands in men and monkeys in hemoblastoses and during aging

Similar changes in the functioning of the adrenal glands (activation of cortisol synthesis due to a more complete utilization of its biochemical precursors and reduced formation of dehydroepiandrosterone and its sulfate fraction) are demonstrated in men and monkeys in hemoblastoses and during aging. It is assumed that being adaptive and nonspecific in nature, these changes lead to hormonal imbalance promoting the development of senile involution processes in patients and animals with severe chronic disease. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 124, No. 8, pp. 207–210, August, 1997     

Nerve-evoked secretion of immunoglobulin A in relation to other proteins by parotid glands in anaesthetized rat

Secretion of fluid and proteins by salivary cells is under the control of parasympathetic and sympathetic autonomic nerves. In a recent study we have shown that, in the rat submandibular gland, autonomic nerves can also increase the secretion of IgA, a product of plasma cells secreted into saliva as SIgA (IgA bound to Secretory Component, the cleaved poly-immunoglobulin receptor). The present study aimed to determine if parotid secretion of SIgA is increased by autonomic nerves and to compare SIgA secretion with other parotid proteins stored and secreted by acinar and ductal cells. Assay of IgA in saliva evoked by parasympathetic nerve stimulation immediately following an extended rest period under anaesthesia indicated that it had been secreted into intraductal saliva in the absence of stimulation during the rest period. The mean rate of unstimulated IgA secretion (2.77+/-0.28 microg min(-1) g(-1)) and the 2.5-fold increase in IgA secretion evoked by parasympathetic stimulation were similar to results found previously in the rat submandibular gland. Sympathetic nerve stimulation increased SIgA secretion 2.7-fold, much less than in the submandibular gland. SDS-PAGE and Western blot analysis with anti-IgA and anti-Secretory Component antibodies confirmed that SIgA was the predominant form of IgA in saliva. Acinar-derived amylase and ductal-derived tissue kallikrein were more profoundly increased by parasympathetic and particularly sympathetic stimulation than SIgA. Overall, the results of the present study indicate that SIgA forms a prominent component of unstimulated parotid salivary protein secretion and that its secretion is similarly increased by stimulation of either autonomic nerve supply. The secretion of other parotid salivary proteins that are synthesized and stored by acinar or ductal cells is upregulated to a much greater extent by parasympathetic and particularly sympathetic stimulation.     

Changes in the functioning of the parotid gland of guinea pigs during regeneration

Summary In guinea pigs, resection of the left parotid gland accompanied by simultaneous extirpation of the right parotid gland caused cessation of secretion during 2–7 days (animals with fistula of the duct) and prolonged the latent period before the appearance of saliva in response to pilocarpine injection. Secretion was restored in all animals towards the 7th day. The initial level of secretion was recovered at different dates between the fourth and twenty-first days.Submitted by Active Member of the Academy of Medical Sciences of the USSR, V. N. Zhukov-Verezhnikov     

Beta-adrenergic regulation of rat parotid gland exocrine protein secretion during aging

beta-Adrenergic regulation of exocrine protein secretion from the parotid gland was studied over the adult rat life span. Enzymatically dispersed cell aggregates were prepared from 3-, 6-, 12-, and 24-month-old rats and exocrine protein secretion (amylase release) measured. No age differences were seen in the time course of amylase release following (-)-isoproterenol stimulation or in the (-)-isoproterenol dose-response curve. the beta-adrenergic antagonist (+/-)-propanolol inhibited (-)-isoproterenol-stimulated protein secretion from parotid cell aggregates of young and old animals equally. Similarly dibutyryl cyclic AMP induced comparable rates of protein secretion from cells of different aged rats. Direct examination of beta-adrenergic receptor characteristics in parotid gland membranes from 3- to 24-month-old rats revealed no differences in the equilibrium dissociation constant (Kd) or maximum specific ligand binding capacity (receptor number). These results suggest that the rat parotid beta-adrenergic system remains functionally intact throughout the animals' lifetime.     

Analysis of the immunoactivator sites of parotid protein isolated from bovine parotid glands

Parotid protein (parotin) was isolated from bovine parotid glands. To analyze the active site of parotid protein, the parotid subunit (PS) was digested by trypsin and then fractionated on a Sephadex G-25 column. Fraction A induced mainly polyclonal antibody responses and interleukin 1 (IL-1)-like activities. FrAA-1, consisting of 58 amino acids (LYILYFFQSDNEDKEKVVRQEEGEE-RITALLMNGSALKQEEWWEKEDTDDTAIVLLK) was isolated from fraction A by chromatography on QAE-Sephadex A-25 columns. FrAA-1 was found to possess IL-1-like activity. There was only 29% homology between FrAA-1 and four IL-1 molecules. When 10- and 20-residue peptides based on the amino acid sequence of FrAA-1 were synthesized, P-10.2 (TDDTAIVLLK) alone exerted an IL-1-like effect on C3H/HeJ thymocytes, whereas P-20.1 (SDNEDKEKVVRQEEGEERIT) alone elicted polyclonal IgM and IgG antibody production in human lymphocytes. These results suggest that the active sites for polyclonal B-cell activator (PBA) and IL-1-like activity have different locations in FrAA-1.     

Changes in the sensitivity of the carotid sinus receptors during aging

Summary In experiments on animals (rabbits and cats) of various age groups, a study was made of the reflexes from the mechano- and chemoreceptors of the carotid sinus. In old animals the changes occurring in the sensitivity of mechano- and chemoreceptors are not uniform. The pressor and depressor reflexes from the mechanoreceptors are weakened, and the sensitivity of chemoreceptors of the carotid area increases. The reflexes in old animals are not stable: with repeated action of the stimulus they decline rapidly and an adaptation phenomenon occurs.(Presented by Active Member AMN SSSR N. N. Gorev) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 54, No. 8, pp. 37–40, August, 1962