Structural and histochemical changes in the salivary glands of Rhipicephalus appendiculatus during feeding

The salivary glands of the brown ear tick of cattle, R. appendiculatus, from both sexes and at all stages of feeding, were examined as whole glands and as sections for ultrastructural and histochemical changes. The type 1 acinus consists of a basal labyrinth formed by the interdigitations of a central cell and four peripheral cells. These cells form a specialized border with a central constrictor cell which surrounds the acinar duct. The plasma membrane of the central cell is exposed to the duct. The type 1 acini do not appear to secrete active saliva components involved in feeding. The type 2 acini undergo a great increase in synthetic and secretory activity during feeding in both sexes and secrete a lipoprotein probably to form part of the attachment cone and also glycoproteins and esterases of unknown functions. The type 3 acini of both sexes also secrete a lipoprotein probably to form part of the attachment cone. The f cells of these acini in the females transiently secrete a glycoprotein of unknown function and then transform to become part of a water excreting unit. In the males the secretory activity of the granular cells of the type 2 and 3 acini is maintained for further attachments. The type 4 acini of the males accumulate masses of proteinaceous granules. The system of interstitial cells and intercellular spaces in types 2, 3 and 4 acini is large and increasingly active during feeding.     

Developmental regulation of granule size and numbers in larval salivary glands of drosophila by steroid hormone ecdysone

The size and number of secretory granules in late larval salivary glands of Drosophila melanogaster have been related to interecdysial and early metamorphic development represented by well-known puffs in polytene chromosomes. Interecdysial period (puff stage 1 (PS1)) is characterized by presence of numerous small granules (11,000 per cell). The transition from PSI to early metamorphic phase (PS2 and upwards), induced by rapid elevation in endogenous steroid hormone ecdysone, is accompanied by continuous growth of granule diameter with concomitant reduction in their number per cell. In the PS4, just prior to secretion, approximately 3000 mature granules occur per cell. The mature state is associated with the change from hyperbolic to Gaussian distribution of granule number over their size range. Similar changes in secretory granule parameters were observed in interecdysial salivary glands explanted from 3rd instar larvae and cultured in vitro in medium containing 5x10(-6) m ecdysone.     

Ultrastructural and histochemical study of the salivary glands of Aplysia depilans (Mollusca, Opisthobranchia)

The digestive system of the sea hare, Aplysia depilans , includes a pair of ribbon-shaped salivary glands. A central duct and a large blood vessel run close to each other along the length of these glands and both are surrounded by a layer of muscle cells. Three cell types form the glandular epithelium: granular cells, vacuolated cells and mucocytes. The granular cells possess cilia and spherical secretion granules, located primarily in the apical region. The granules of immature cells have a low electron density and are mainly formed by neutral polysaccharides with small amounts of proteins. The granules of mature cells are larger, have a high electron density and are mainly formed by proteins with lower amounts of neutral polysaccharides. Transition stages between immature and mature granular cells are observed. The vacuolated cells are large and frequently pyramidal in shape, but after the application of histochemical techniques almost all vacuoles remain uncoloured. The numerous vacuoles contain flocculent material in a clear background and the mitochondria possess large crystalline structures in the matrix. A pyramidal shape is also typical of the mucocytes, which are filled with vesicles containing granular masses surrounded by a network of secretion material. These large cells are strongly stained by Alcian blue, revealing the presence of acidic mucopolysaccharides. This is the first ultrastructural study of the salivary glands in opisthobranch gastropods.     

Morphofunctional changes of salivary glands of female ixodid ticks of subfamilies Ixodinae and Amblyomminae (Acari: Ixodidae) during feeding and their significance

Using light and electron microscopy, a dynamics of functioning of salivary glands of female ticks of the genus Ixodes was studied. Comparative analysis of morphofunctional changes and role of the salivary gland secretions of females of subfamilies Ixodinae and Amblyomminae during feeding was performed on the basis of literature data and our own studies.     

Morphologic diversity of the minor salivary glands of the rat: fertile ground for studies in gene function and proteomics

In this article the locations and histologic and ultrastructural features of all of the minor salivary glands of the rat are presented; similarities and differences among them are highlighted. These glands are almost as diverse morphologically as the major salivary glands of the rat. The acini of von Ebner's glands are serous; those of the anterior and posterior buccal glands and minor sublingual glands are mucous; and those of the glossopalatal, palatal, and Weber's glands are mucous with serous demilunes. The anterior buccal, minor sublingual and von Ebner's glands have striated and stratified columnar ducts, while only the minor sublingual and von Ebner's glands have intercalated ducts. The glossopalatal, palatal, posterior buccal and Weber's glands have none of these ducts; the tubulo-acini drain abruptly into short terminal ducts composed of stratified squamous epithelium. All of the mucous acini react with an antibody to a mucin (Muc19) of the rat major sublingual gland, but in some of the glands the reaction varies in intensity among the acinar cells. Ultrastructurally, the mucous secretory granules of the anterior buccal, glossopalatal, palatal and Weber's glands are biphasic, while those of the minor sublingual and posterior buccal glands are monophasic. Although there is a considerable body of literature concerning the development, innervation, physiology and proteomics of von Ebner's glands, investigation of the other minor salivary glands of the rat ranges from modest to nearly nonexistent.     

Morphology of the salivary glands of three Squamata species: Podarcis sicula sicula, Tarentola mauritanica and Coluber viridiflavus

The histology, histochemistry and ultrastructure of the salivary glands of three species of squamata, Podarcis sicula sicula (mandibular glands), Tarentola mauritanica (sublingual gland) and Coluber viridiflavus (supra- and infralabial glands) were studied. Each gland contained acidic cells, positive for both periodic acid Schiff and Alcian blue reactions. These cells can be distinguished as seromucous and mucous types based on the different electron density of their granules. α-Amylase, until now detected only in mammalian salivary glands, was not found in any of the salivary glands examined. The ultrastructural study revealed that the salivary glandular cells of T. mauritanica lack intercellular canaliculi, which by contrast, are present between the seromucous cells in the salivary glands of C. viridiflavus and P. s. sicula . Comparable variation is also seen when the ultrastructural features of the secretory granules in salivary glands of the three Squamata species are compared. The salivary granules of T. mauritanica and C. viridiflavus are more or less dense but structureless, while the mucous granules of P. s. sicula have a distinctive and characteristic substructure. Therefore, this study, designed to obtain comparative data on the histology, histochemistry and ultrastructure of the salivary glands of three representative, but hitherto unstudied, species of Squamata, reveals great variation in the structure of these glands within the Squamata lineage, even when compared to previously documented species.     

Morphophysiological analysis of the salivary glands of Rhipicephalus sanguineus sensu lato (Acari: Ixodidae) exposed to ozonated water: A control strategy

The tick Rhipicephalus sanguineus sensu lato has great medical and veterinary importance, mainly because the ability to transmit many diseases, causing harm to pets but also risks to public health. The blood spoliation and transmission of pathogens occur because of the immunosuppressive action of these ticks' saliva, a potent mixture of bioactive substances that is secreted by the salivary glands, one of the organs responsible for their biological success, and hence the target of studies for their control. Ozone has promise for use as an alternative acaricide, due to its proven efficiency in controlling agricultural and food pests, besides posing no risk of environmental contamination or to animal and human health. Therefore, this study evaluated the acaricidal potential of exposure of females of R. sanguineus s.l. to ozonated water at many concentrations and analysed the morphophysiological alterations of the salivary glands, employing histological and light microscopic techniques. The results demonstrated that the ozonated water at the concentrations investigated caused severe alterations in the salivary glands, bringing a new perspective for control of R. sanguineus s.l., through an ecologically correct method due to the absence of harm to non‐target organisms and the environment.     

The human submandibular gland: immunohistochemical analysis of SNAREs and cytoskeletal proteins

Submandibular acinar glands secrete numerous proteins such as digestive enzymes and defense proteins on the basis of the exocrine secretion mode. Exocytosis is a complex process, including a soluble NSF attachment protein receptor (SNARE)-mediated membrane fusion of vesicles and target membrane and the additional activation of cytoskeletal proteins. Relevant data are available predominantly for animal salivary glands, especially of the rat parotid acinar cells. The authors investigated the secretory molecular machinery of acinar (serous) cells in the human submandibular gland by immunohistochemistry and immunofluorescence and found diverse proteins associated with exocytosis for the first time. SNAP-23, syntaxin-2, syntaxin-4, and VAMP-2 were localized at the luminal plasma membrane; syntaxin-2 and septin-2 were expressed in vesicles in the cytoplasm. Double staining of syntaxin-2 and septin-2 revealed a colocalization on the same vesicles. Lactoferrin and α-amylase served as a marker for secretory vesicles and were labeled positively together with syntaxin-2 and septin-2 in double-staining procedures. Cytoskeletal components such as actin, myosin II, cofilin, and profilin are concentrated at the apical plasma membrane of acinar submandibular glands. These observations complement the understanding of the complex exocytosis mechanisms.     

Morphology of the foveal glands and foveae dorsales in male Hyalomma truncatum and Rhipicephalus evertsi mimeticus ticks (Acari; Ixodidae) before and during feeding

The ultrastructure of the foveae dorsales and foveal glands in unfed and attached male Hyalomma truncatum and Rhipicephalus evertsi mimeticus ticks was studied. Both species are provided with a paired foveal gland system, which is similar in unfed as well as in attached ticks. This gland system consists of the fovea dorsalis with pores and pore tubes as the external part, the foveal neck zone as a link between the fovea dorsalis and the lobes of the gland and the bulbous lobes as the innermost part. The fovea dorsalis is located on either side of the dorsal midline in the midsection of the body and appears as a roundish plate containing 15±6.5 and 21±7 slit-like pores in R. evertsi mimeticus (n=210) and H. truncatum (n=210), respectively. Each pore leads into a cuticular lined channel containing a pore tube. Below each fovea, the foveal neck zone is located within a groove of the cuticle and consists of the termini of the pore tubes which enlarge basally to form a cup-shaped ampulla each. Furthermore, secretory lobes are located below the foveal neck zone. Each lobe consists of secretory cells and a central excretory duct which leads into the ampulla. The ducts are lined with microvilli. The secretory cells contain numerous vesicles of varying size with one or more granules. In male ticks of both species the secretory lobe cells remained unchanged in size, structure and granule content irrespective of whether they were unfed or attached for up to 30 days. Axons occur in the fascicles between the secretory lobe cells containing numerous neurosecretory vesicles. A possible role of the foveal glands in the production of pheromones is hypothesized.     

Theileria annulata and Babesia ovis: ultracytochemical lactic dehydrogenase activity of sporozoites in salivary glands of female ticks, Hyalomma anatolicum excavatum and Rhipicephalus bursa

The occurrence of a marker enzyme of glycolysis, lactic dehydrogenase (LDH) (EC 1.1.1.27.), was studied ultracytochemically in sporozoites of Babesia ovis in the tick Rhipicephalus bursa and in sporozoites of Theileria annulata in the tick Hyalomma anatolicum excavatum. Female ticks infected transstadially were fed on rabbits and dissected 3–5 days post infestationem. The salivary glands were removed and incubated in the cytochemical medium unfixed or after fixation in buffered paraformaldehyde solution. A modified ferricyanide medium adjusted to pH 6.5 was used for incubation. Controls were performed by preincubating specimens in 10^?3 M iodine or by omitting NAD⁺ or lactate in the incubation medium. Following incubation, the specimens were fixed in buffered solutions of paraformaldehyde or glutaraldehyde, postfixed in osmium tetroxide, and embedded in Durcupan ACM. Mature “schizonts” consisting of an abundant number of sporozoites were examined in both piroplasmean species. In sporozoites of B. ovis the final enzymic product was deposited within the nucleus. No cytoplasmic reaction was observed. However, the membrane delimiting the schizont from the adjacent glandular cells was distinctly reactive. In T. annulata reactivity was usually confined to the cytoplasm. Sometimes, a reaction within mitochondria could be observed. The reaction product had formed aggregations which often appeared to be attached to micronemes. There was no nuclear reactivity in this species.The results suggest the existence of a glycolytic metabolic pathway with different subcellular localizations in sporozoites of the two piroplasmean species.     

家蚕蛹-成虫变态期中肠和涎腺超微结构变化与功能

为进一步明确家蚕Bombyx mori变态期间消化系统的生理功能以及溶茧酶的来源器官,通过透射电镜观察和酶活性检测,对家蚕蛹 成虫变态期中肠和涎腺的超微结构、水解酶活力以及中肠内容物的变化进行了观察和分析。结果表明: 蛹第7日到羽化前1日家蚕中肠细胞和刚羽化成虫的涎腺细胞中,均可观察到大量的分泌泡、分泌颗粒、微绒毛等分泌细胞的结构特征以及活跃的分泌现象。潜成虫的中肠和涎腺中都存在活性较高的水解酶活力,其中每毫克中肠组织中蛋白酶活力、酯酶活力和纤溶酶活力分别为726 U、751 U和263 U,每毫克涎腺组织中上述3种酶的活力分别为603 U、523 U和147 U,说明中肠和涎腺可能都具有分泌溶茧酶的功能。家蚕蛹期中肠内容物的主要成分是蛋白质、脂质和糖,三者占内含物总量的95%以上,其中蛋白质含量占78.8%～80.2%。中肠内容物的重量在刚化蛹时为20.1～21.9 mg/头,化蛹后7日内无明显变化,化蛹第9日内容物重量减少63.01%～66.17%,到成虫羽化时已所剩无几,可能是因内容物被消化吸收所致。据此推测,在家蚕变态期中肠还具有贮存和释放营养物质的功能,而溶茧酶的另一个功能可能是分解消化中肠内容物。     

Ultrastructural changes of Drosophila larval and prepupal salivary glands cultured in vitro with ecdysone

Summary Alterations in the ultrastructure of in vitro cultured larval salivary glands of Drosophila melanogaster in response to the steroid hormone ecdysone were studied in relation to complex changes in puffing patterns. We found that the changes in the fine structure of cultured glands reflected progression of the puffing pattern, and they paralleled those seen in vivo. We observed that glue secretion by exocytosis, the main function of salivary glands, took place between puff stage 5 (PS5) and PS7. Glue could not be expectorated under culture conditions but was slowly released from the lumen through a duct into the medium. After the cultured glands reached PS13/PS14, further progress of puffing and fine structural alterations required that the ecdysteroid titer be transiently extremely low or absent. Under in vitro conditions we did not observe the putative new secretory program(s) described for glands in vivo after PS12. However, ultrastructural changes which unambiguously indicated that an autohistolytic process had begun in vitro started to appear after PS17. Many salivary gland cells developed numerous features of progressive self-degradation between PS18 and PS21. Actual degradation of salivary glands in vivo seemed to be rapid, but in vitro degradation was never completed, probably due to a lack of exogenous factors from the hemolymph. Manipulations of ecdysone titer in vitro in the culture medium, known during the larval puffing cycle to cause premature induction of developmentally specific puffing patterns, did not affect the normal development of ultrastructural features of the cytoplasm and nucleus.     

Serial cultivation of epithelial cells from human and macaque salivary glands

Summary To study the regulation of human salivary-type gene expression we developed cell culture systems to support the growth and serial cultivation of salivary gland epithelial and fibroblastic cell types. We have established 22 independent salivary gland epithelial cell strains from parotid or submandibular glands of human or macaque origin. Nineteen strains were derived from normal tissues and three from human parotid gland tumors. Both the normal and the tumor-derived salivary gland epithelial cells could be serially cultivated with the aid of a 3T3 fibroblast feeder layer in a mixture of Ham’s F12 and Dulbecco’s modified Eagle’s media supplemented with fetal bovine serum, calcium, cholera toxin, hydrocortisone, insulin, and epidermal growth factor. Salivary gland epithelial cells cultured under these conditions continued to express the genes for at least two acinar-cell-specific markers at early passages. Amylase enzyme activity was detected in conditioned media from cultured rhesus parotid epithelial cells as late as Passage 5. Proline-rich-protein-specific RNAs were detected in primary cultures of both rhesus and human parotid epithelial cells. Neither amylase enzyme activity nor PRP-specific RNAs were detected in fibroblasts isolated from the same tissues. In addition, salivary gland epithelial cells cultured under our conditions retain the capacity to undergo dramatic morphologic changes in response to different substrata. The cultured salivary gland epithelial cells we have established will be important tools for the study of salivary gland differentiation and the tissue-specific regulation of salivary-type gene expression.     

Sequential changes in salivary gland structure during attachment and feeding of the cattle tick, Boophilus microplus

A study of the morphology and histochemistry of the salivary glands of the parasitic stages of Boophilus microplus has been made, glands of feeding females being studied in greatest detail. Of 9 granular cell types present in the female and 10 in the male, 3 probably secrete attachment cement and 4 others glycoproteins and enzymes, possible functions of which are discussed. Two cell types, c₄ and g (the latter being present only in males) are of unknown function. The most likely functions of non-granular epithelial cells and those forming acinus I are in osmoregulation.     

Female tick Hyalomma marginatum marginatum salivary glands: preliminary study on protein changes during feeding process and antigens recognized by repeatedly infested cattle

Proteins extracted from salivary glands of unfed, three days and five days fed adult Hyalomma marginatum marginatum were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). We have noticed changes during the three feeding steps. Some proteins disappeared during feeding process (23, 38, 39, 40 to 50, 95 and 112 kDa), they might be proteins which were converted in other substances and are secreted. Other antigens (13 to 14, 20, 25, 29, 165 and 210 kDa) were synthesized as a result of tick attachment and feeding. They may be related to growth and development or are the ciment which fixed the adult. Also, three Holstein calves were infested five times with 100 pairs of adult ticks of the same species. The five infestations were performed two weeks from the previous infestation. The sera before infestations and after each infestation were used in western-blot analyses to identify antigens from five days salivary gland extracts of the primary infestation of ticks. Three antigens (18.7, 50 and 80 kDa) were revealed weakly after the first and the second infestations by sera samples but not at infestation onward. Others (13.5, 17 to 18.5, 25, 30, 70, 133, 176 and 193 kDa) were revealed only by sera taken after manifestation of resistance (third infestation). A 13.5 kDa antigen was particularly revealed when resistance had appeared and became more evident after the fourth and fifth infestations. The late antigens recognized might be associated with establishment of calves resistance against ticks.     

Mechanisms of programmed cell death in the midgut and salivary glands from Bradysia hygida (Diptera: Sciaridae) during pupal–adult metamorphosis

Programmed cell death is involved with the degeneration/remodeling of larval tissues and organs during holometabolous development. The midgut is a model to study the types of programmed cell death associated with metamorphosis because its structure while degenerating is a substrate for the formation of the adult organ. Another model is the salivary glands from dipteran because their elimination involves different cell death modes. This study aimed to investigate the models of programmed cell death operating during midgut replacement and salivary gland histolysis in Bradysia hygida. We carried out experiments of real‐time observations, morphological analysis, glycogen detection, filamentous‐actin localization, and nuclear acridine orange staining. Our findings allow us to establish that an intact actin cytoskeleton is required for midgut replacement in B. hygida and nuclear condensation and acridine orange staining precede the death of the larval cells. Salivary glands in histolysis present cytoplasmic blebbing, nuclear retraction, and acridine orange staining. This process can be partially reproduced in vitro. We propose that the larval midgut death involves autophagic and apoptotic features and apoptosis is a mechanism involved with salivary gland histolysis.     

On approaches to the functional restoration of salivary glands damaged by radiation therapy for head and neck cancer,with a review of related aspects of salivary gland morphology and development

Radiation therapy for cancer of the head and neck can devastate the salivary glands and partially devitalize the mandible and maxilla. As a result, saliva production is drastically reduced and its quality adversely altered. Without diligent home and professional care, the teeth are subject to rapid destruction by caries, necessitating extractions with attendant high risk of necrosis of the supporting bone. Innovative techniques in delivery of radiation therapy and administration of drugs that selectively protect normal tissues can reduce significantly the radiation effects on salivary glands. Nonetheless, many patients still suffer severe oral dryness. I review here the functional morphology and development of salivary glands as these relate to approaches to preventing and restoring radiation-induced loss of salivary function. The acinar cells are responsible for most of the fluid and organic material in saliva, while the larger ducts influence the inorganic content. A central theme of this review is the extent to which the several types of epithelial cells in salivary glands may be pluripotential and the circumstances that may influence their ability to replace cells that have been lost or functionally inactivated due to the effects of radiation. The evidence suggests that the highly differentiated cells of the acini and large ducts of mature glands can replace themselves except when the respective pools of available cells are greatly diminished via apoptosis or necrosis owing to severely stressful events. Under the latter circumstances, relatively undifferentiated cells in the intercalated ducts proliferate and redifferentiate as may be required to replenish the depleted pools. It is likely that some, if not many, acinar cells may de-differentiate into intercalated duct-like cells and thus add to the pool of progenitor cells in such situations. If the stress is heavy doses of radiation, however, the result is not only the death of acinar cells, but also a marked decline in functional differentiation and proliferative capacity of all of the surviving cells, including those with progenitor capability. Restoration of gland function, therefore, seems to require increasing the secretory capacity of the surviving cells, or replacing the acinar cells and their progenitors either in the existing gland remnants or with artificial glands.