Rat Submandibular Gland Research Articles

Post-weaning xenohormone intake affects adult rat submandibular gland in a sex-dependent manner.

We previously reported that maternal exposure to genistein and vinclozolin, ingested alone or in combination, affects submandibular salivary glands of rat offspring. Here, we investigated the responsiveness of submandibular gland when such xenohormone exposure occurs later in life. Chemicals were given orally to male and female Wistar rats (1mg/kg body weight per day), from weaning to adulthood. Submandibular glands and plasma were collected at postnatal day 100 for histologic and molecular analysis. Whereas no effect was observed in females, increases in granular convoluted tubules area coupled with a modification of salivary secretions were found in male submandibular glands. Genistein and vinclozolin similarly increased the mRNA expression of Cystatin C, Mucin 10, Growth factors, and plasmatic EGF. Negative correlations were found between the expressions of androgen receptor and EGF (-0.34; p<0.05), TGFα (-0.52; p<0.01), Mucin 10 (-0.43; p<0.05), and Cystatin C (-0.42; p<0.05) as well as between progesterone receptor and EGF (-0.56; p<0.01). The Spearman correlation test revealed also a positive correlation between salivary EGF-mRNA expression and EGF in plasma (+0.32; p<0.05). Our findings confirm the sex-dependent sensitivity of submandibular salivary glands to dietary xenohormones and underline the influence of the exposure period.

Effect of low radiation dose on the expression and location of aquaporins in rat submandibular gland.

Head and neck cancers are common in several regions of the world and the treatment usually includes radiotherapy. This treatment can generate adverse effects to the salivary flow, with a relationship between the dose and the damage caused. Salivary gland cells are highly permeable to water and therefore, they express aquaporins (AQPs). This study analyzed changes in the expression and location of these proteins and identified morphological changes induced by low radiation in rat submandibular gland. Female rats were divided into control and irradiated groups. Immunohistochemistry analysis allowed confirming the presence of AQP1 in the blood vessel endothelium. Intense and steady labelling granules were also observed in the cytoplasm of submandibular gland ductal cells. In addition, there was AQP5 positive labelling in ductal cells delimiting the lumen of intercalated duct, in the cytoplasm and membrane of acinar cells. Finally, the decrease of AQP labelling in irradiated animal glands validated their radiosensitivity. Thus, the decrease in AQP1 protein levels in the endothelium and AQP5 in gland ductal cells of irradiated animals may have hindered the removal of water from the lumen of ductal cells, inducing a delay in water absorption and triggering a slight lumen increase.

Differences in mucin expression in the submandibular glands of rats during peridontitis induction

Background: Porphyromonas gingivalis (Pg) produces lipopolysacharide (LPS) which acts as a stimulator of inflammation in periodontal tissues. Periodontitis-induced apoptosis and vacuolation of the salivary gland, therefore, causes hyposalivation. Mucin secretion is produced by the submandibular gland under stimulation by the cholinergic and adrenergic receptors. Both forms of stimulation influence the volume of mucin secretion. Mucin saliva plays an important role in the early stages of Pg colonization in the oral cavity. On the other hand, it serves to protect against bacterial invasion. Purpose: The aim of this research was to identify differences in mucin expression in the submandibular gland during periodontitis induction. Methods: 32 male Wistar rats were assigned to either a sham periodontitis or a periodontitis group. The former group received a daily injection of a vehicle solution (n = 16), while members of the periodontitis induction group (n=16) were injected each day with 500 µL of Pg 108 into the mesial area of the upper molar. Mucin in the submandibular gland was analyzed at the 7th, 14th, 21th and 28th days after injection by means of periodic acid schiff (PAS) staining. Results: 28 days after injection mild gingivitis was developed in the periodontitis experiment group. Junctional epithelium (JE) thickness decreased gradually following the increase of PG injection periods (p&lt;0.05). However, mucin expression increased prominently at 7th, 14th, and 21th days after injection and decreased on day 28th after PG injection. Mucin was expressed in the duct cells of the submandibular gland. Conclusion: The result of this study suggests that there are different levels of mucin expression in the submandibular gland during periodontitis induction.

Preparation of Murine Submandibular Salivary Gland for Upright Intravital Microscopy.

The submandibular salivary gland (SMG) is one of the three major salivary glands, and is of interest for many different fields of biological research, including cell biology, oncology, dentistry, and immunology. The SMG is an exocrine gland comprised of secretory epithelial cells, myofibroblasts, endothelial cells, nerves, and extracellular matrix. Dynamic cellular processes in the rat and mouse SMG have previously been imaged, mostly using inverted multi-photon microscope systems. Here, we describe a straightforward protocol for the surgical preparation and stabilization of the murine SMG in anesthetized mice for in vivo imaging with upright multi-photon microscope systems. We present representative intravital image sets of endogenous and adoptively transferred fluorescent cells, including the labeling of blood vessels or salivary ducts and second harmonic generation to visualize fibrillar collagen. In sum, our protocol allows for surgical preparation of mouse salivary glands in upright microscopy systems, which are commonly used for intravital imaging in the field of immunology.

Morphological and functional state of the epithelial cells in submandibular salivary glands after repeated amputations of the incisor teeth in adult rats

Purpose. To evaluate a morphofunctional state of the epithelial cells of the acini and ducts of submandibular glands of mature rats after repeated incisors amputation. Materials and methods. The experiment was performed on mature (2 months) white male rats, divided into groups: intact, control group and a group of rats subjected to repeated incisors amputation. Applying histological, histochemical and morphometric methods, we evaluated the morphological and functional state of the epithelial cells in submandibular glands of adult rats at 2, 3, 4, 6, 8, 10 and 12 weeks after repeated amputation of the incisor teeth. Results. There is an increase of submandibular gland acini area at the 3–10 week after repeated incisors amputation. The functional activity of the submandibular glands ducts and acini cells decreases at the 2–4 week of the experiment. Conclusion. As a result of repeated incisors amputation in rat submandibular glands acini and ducts epithelial cells develop reversible structural and functional changes, which are leveled by the 12th and 6th week of the experiment, respectively.

Effect of Thyroid Hormone on the Histology of Rat Submandibular Salivary Gland During Postnatal Development

Effect of Thyroid Hormone on the Histology of Rat Submandibular Salivary Gland During Postnatal Development

Antimicrobial protein secretion in the rat submandibular gland induced by aroma inhalation

Antimicrobial protein secretion in the rat submandibular gland induced by aroma inhalation

Distribution of aquaporin-5, transforming growth factor-β1 and laminin during regeneration of atrophic rat submandibular glands after duct ligation.

The aim of this study was to determine the localization of aquaporin-5 (AQP5), transforming growth factor-β1 (TGF-β1) and laminin during regeneration of the rat submandibular gland. After duct ligation for 7 days, the regenerating glands were collected on days 0, 1, 3, 7, and 14 after ligation release to study the process of regeneration. Immunohistochemical staining revealed apical expression of AQP5 in many acinar cells, strong expression in intercalated ducts (ICDs) of the normal submandibular gland at Day 14, and strong expression in duct-like structures (DLSs) during regeneration from Day 0 to 7. However, a few AQP5-negative acinar cells were detected during regeneration. At Day 0, immunopositivity for TGF-β1 was detected in connective tissue. At Days 3 and 7 during regeneration, TGF-β1 immunostaining was observed in DLSs, which were surrounded by α-smooth muscle actin-positive thickened myoepithelial cells. Laminin staining was strong in the thickened basement membrane of DLSs at Day 3 during regeneration, but weak around acinar cells at Day 14. These findings suggest that TGF-β1 is involved in the environment around DLSs, myoepithelial cells and laminin, that DLSs have the same functional properties as ICDs, and that AQP5-negative acinar cells may be mucous cells.

Simultaneous monitoring of Ca&lt;sup&gt;2+&lt;/sup&gt; response and salivary secretion in rat submandibular gland in live animals

Simultaneous monitoring of Ca&lt;sup&gt;2+&lt;/sup&gt; response and salivary secretion in rat submandibular gland in live animals

Ca2+-CaMKKβ pathway is required for adiponectin-induced secretion in rat submandibular gland.

Adiponectin functions as a promoter of saliva secretion in rat submandibular gland via activation of adenosine monophosphate-activated protein kinase (AMPK) and increased paracellular permeability. Ca2+ mobilization is the primary signal for fluid secretion in salivary acinar cells. However, whether intracellular Ca2+ mobilization is involved in adiponectin-induced salivary secretion is unknown. Here, we found that full-length adiponectin (fAd) increased intracellular Ca2+ and saliva secretion in submandibular glands. Pre-perfusion with ethylene glycol-bis (2-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA) combined with thapsigargin (TG), an endoplasmic reticulum Ca2+-ATPase inhibitor, abolished fAd-induced salivary secretion, AMPK phosphorylation, and enlarged tight junction (TJ) width. Furthermore, in cultured SMG-C6 cells, co-pretreatment with EGTA and TG suppressed fAd-decreased transepithelial electrical resistance and increased 4-kDa FITC-dextran flux responses. Moreover, fAd increased phosphorylation of calcium/calmodulin-dependent protein kinase (CaMKKβ), a major kinase that is activated by elevated levels of intracellular Ca2+, but not liver kinase B1 phosphorylation. Pre-perfusion of the isolated gland with STO-609, an inhibitor of CaMKKβ, abolished fAd-induced salivary secretion, AMPK activation, and enlarged TJ width. CaMKKβ shRNA suppressed, whereas CaMKKβ re-expression rescued fAd-increased paracellular permeability. Taken together, these results indicate that adiponectin induced Ca2+ modulation in rat submandibular gland acinar cells. Ca2+-CaMKKβ pathway is required for adiponectin-induced secretion through mediating AMPK activation and increase in paracellular permeability in rat submandibular glands.

Daily voluntary exercise enhances pilocarpine-induced saliva secretion and aquaporin 1 expression in rat submandibular glands.

Saliva—a water‐based fluid containing electrolytes, immunoglobulins, and enzymes—has many functions, including the protection and hydration of mucosal structures within the oral cavity and the initiation of digestion. Aquaporins (AQPs) are proteins that act as water channels through membranes. We have previously reported upregulation of the expression levels of AQP 1 and 5 in the submandibular glands (SMGs) in heat‐acclimated rats. In this study, we investigated pilocarpine‐induced saliva secretion and AQP expression in rats after voluntary exercise. Male, 10‐week‐old Wistar rats were initially maintained at an ambient temperature of 24 °C for 10 days and were then kept for 40 days in cages either with a running wheel (exercise rats, n = 6) or with a locked wheel [control rats (CN), n = 6]. After the training period, the rats were anesthetized and pilocarpine, an M3 muscarinic receptor agonist, was intraperitoneally injected (0.5 mg·kg−1) to stimulate saliva secretion. Saliva was collected, and the SMGs were sampled and subjected to western blot, RT‐PCR, and immunohistochemical analyses. Pilocarpine induced a greater amount of saliva in the exercised rats than in the CN. Expression levels of AQP1 mRNA and protein were significantly higher in SMGs of exercised rats than in those of the CN, but the expression of AQP5 was not affected by voluntary exercise. Voluntary exercise increased the expression of vascular endothelial growth factor (VEGF) and cluster of differentiation 31 (CD31), a marker for endothelial cells, in the SMGs. Voluntary exercise promoted pilocarpine‐induced saliva secretion, probably via an increase in the expression level of AQP1 due to VEGF‐induced CD31‐positive angiogenesis in the SMG.

Histamine stimulates secretion of extracellular vesicles with nucleotidase activity in rat submandibular gland

BackgroundExtracellular vesicles released by different cells have been isolated from diverse fluids including saliva. We previously reported that rat submandibular glands secrete nanovesicles that catalyze hydrolysis of ATP, ADP and AMP, which are actors of the purinergic signaling system along with adenosine. Extracellular nucleotides like ATP and adenosine are involved in the regulation of inflammatory processes and apoptosis. Histamine, a widely distributed biogenic amine, is involved in inflammatory response. ObjectiveTo test if activation of histamine receptors in rat submandibular gland promotes changes in the release of vesicles with nucleotidase activity that could modulate purinergic signaling. MethodsRat submandibular glands were incubated in the absence or presence of histamine and JNJ7777120, an antagonist for H4 receptors. Extracellular vesicles were isolated from incubation media by differential centrifugation. Vesicular nucleotidase activity was measured following Pi release by 3mM MgATP, MgADP or MgAMP. ResultsHistamine increased the release of vesicles with nucleotidase activity in a concentration dependent manner. JNJ7777120 significantly reduced this effect. Vesicular nucleotidases obtained in the absence or presence of histamine promoted Pi production from ATP, ADP and AMP. ConclusionThe results show a relationship between histamine and the regulation of purinergic signaling, which could be important in the modulation of inflammatory processes.

ROLE OF GHRELIN ON CISPLATIN INDUCED MORPHOLOGICAL CHANGES ON SUBMANDIBULAR SALIVARY GLANDS

Background: Cisplatin is a potent antineoplastic agent widely used for a variety of malignancies. However, it has many complications such as neuropathy and cachexia. Ghrelin is a recently discovered hormone with a potent orexigenic and neuroprotective properties that may prevent or reduce these complications. Objective: This present study designed to determine the effectiveness of ghrelin administration as a protective agent against cisplatin-induced cytotoxicity in the submandibular glands of rats. Design: Twenty –four adult male Sprague Dawley rats, were divided into three groups (n=8). Group (I) rats were received ghrelin 0.8 mg/kg twice daily for 14 day. Group (II) rats were received three doses cisplatin 0.5 mg /kg given daily. Group (III) rats were received cisplatin and ghrelin in dose as in group I, II. Daily measurement of body weight and blood glucose level. After 14 days, animals were scarified and the submandibular glands were removed and the prepared sections were examined histologically by H&E and immunohistochemical stain for inducible nitric oxide synthase (iNOS). Results: Ghrelin administration in parallel with cisplatin-based chemotherapy were markedly stimulate appetite, with improvement of body weight and blood glucose level. Histological assessment revealed that ghrelin hormone reduce necrotic changes induced by chemotherapy.Conclusions: Exogenous ghrelin markedly increase food intake and general nutritional health and also improve the induced cytotoxic changes during chemotherapy.

In vivo Endocrine Secretion of Prostacyclin Following Expression of a Cyclooxygenase-1/Prostacyclin Fusion Protein in the Salivary Glands of Rats Via Nonviral Gene Therapy

Pulmonary arterial hypertension (PAH) is a progressive disease that culminates in right heart failure and death. Prostacyclin (PGI2) and its derivatives are effective treatments for PAH when administered as continuous parenteral infusions. This treatment paradigm requires medical sophistication, and patients are at risk for complications from an indewelling catheter; drug interruptions may result in rebound pulmonary hypertension and death. We hypothesized that the salivary gland can be repurposed into an endogenous production site for circulating PGI2 through the expression of a fusion protein embodying cyclooxygenase-1 (Cox1) and prostacyclin synthase (PGIS) domains. We utilized ultrasound-assisted gene transfer, a nonviral gene transfer strategy that achieves robust gene transfer to the salivary gland. We initially found that Cox1-PGIS expression in livers of mice using an adenoviral vector dramatically increased circulating PGI2 relative to untreated rats or rats treated with PGIS alone. We then utilized ultrasound-assisted gene transfer to express Cox1-PGIS in the submandibular glands of rats and showed a significant elevation of circulating PGI2 that corresponded to approximately 30% of that seen in humans undergoing intravenous infusion therapy for PAH. These results suggest the feasibility of gene therapy to drive endogenous biosynthesis of PGI2 as a therapeutic strategy for the treatment of PAH.

Histopathological and Ultrastructural evaluation of the Effect of Electronic Cigarette Smoke on the Submandibular Gland of Albino Rats

Introduction: E-cigarettes (E-cigs) have received a considerable attention within the last years and are among the newest products for reduction of tobacco injury. Recently, it has been postulated that they are not considered a safe substitute for smoking and may have a limited degree of effectiveness in arresting smoking. Aim: The aim of this study was to detect the histopathological and ultrastructural changes of tissue architecture of the submandibular glands in rats after passive E-cig exposure.Methods: 12 adult male albino rats were divided into three groups (four rats each). Group I (Control group): Rats were exposed to fresh air. Group II (E-cigarette 4 weeks): Each rat was exposed to passive E-cig smoke for duration of one hour daily. The exposure time was divided into 6 equal intervals of 10 mins /hr, 7 days a week for four successive weeks. Group III (E-cigarette 12 weeks): Rats of this group were exposed to the same dose and manner of treatment of E-cig smoke as in group II for 12 weeks duration. Animals were sacrificed following each treatment period and specimens were prepared for histological examination and transmission electron microscopic study.Results: H & E results revealed signs of atrophy and degeneration within the parenchymal elements together with dilatation and congestion of blood vessels with interstitial hemorrhage in 4 weeks E-cig exposed rats. More deleterious changes with marked distortion of the gland’s architecture were detected in 12 weeks E-cig exposed group. These data were supported by the electron microscopic results which reflected degenerative changes in cytoplasmic organelles. Conclusion: E-cig is not safe and its use to replace cigarette smoking cannot be recommended.

Immunohistochemical localization and expression of TRPs in rat submandibular gland development

Immunohistochemical localization and expression of TRPs in rat submandibular gland development

Temporal Expression Patterns of Clock Genes and Aquaporin 5/Anoctamin 1 in Rat Submandibular Gland Cells.

Circadian rhythms are essential for health and regulate various physiological functions. These rhythms are regulated by a negative-feedback loop involving clock genes in the suprachiasmatic nucleus (SCN) and peripheral tissues. The rate of secretion of salivary substances, ions, and water follows a circadian rhythm, however, the relationship between the molecular mechanism of salivary secretion and peripheral circadian rhythm is not yet clear. Anoctamin 1 (ANO1, also known as TMEM16A) and Aquaporin 5 (AQP5) play an important role in the transport of ions and water in the submandibular glands (SGs). We examined the interaction between the rhythmic expression pattern of the clock genes, Ano1 and Aqp5, in rat whole SGs as well as isolated acinar and ductal cells. Circadian rhythmic expression for Bmal1, Per1, Per2, Clock, Cry1, Cry2, Rorα, and Rev-erbα mRNAs, also called the clock genes, was observed in rat SGs by semi-quantitative RT-PCR analysis. We also observed rhythmic patterns in Ano1 and Aqp5 mRNA expression. The expression of ANO1 protein also showed circadian rhythm, as confirmed by western blot analysis. We could not observe any time delay between the peak expression of ANO1 protein and its mRNA. Expression levels of the clock gene mRNAs in the ductal cells was higher than that in acinar cells, however, rhythmic oscillations were observed in both. Our results suggest that SGs have peripheral clocks, and rhythmic expressions of Ano1 and Aqp5 along with the clock genes, may play an important role in the circadian regulation of salivary secretion.

Hyperlipidemia is involved in apoptosis in rat submandibular glands

ObjectiveThe aim of the present study was to investigate the effects of hyperlipidemia on histological changes and apoptosis in submandibular glands using apolipoprotein E (apoE)-deficient rats. DesignHistopathological findings related to induced apoptosis in the submandibular glands were compared between apoE-deficient rats (n=6; male; age, 16 weeks) and the corresponding wild-type rats (n=6). ResultsApoE-deficient rats showed significantly higher plasma levels of oxidized low-density lipoprotein (LDL), total cholesterol, very LDL and LDL, and lower plasma levels of high-density lipoprotein when compared to control rats (P<0.05). Lipid deposition in the submandibular gland was observed in apoE-deficient rat group and in none of the control group. Significant increases in vacuolization and apoptosis in acinar cells were observed in apoE-deficient rats, as compared to control rats (P<0.05). The number of active caspase-3-positive cells was also higher in the apoE-deficient rat group when compared with the control group (P<0.01). ConclusionsAccording to our results, hyperlipidemia induced apoptosis in apoE-deficient rat submandibular glands. Oxidized LDL generation in case of hyperlipidemia may trigger off a reaction of apoptotic acinar cells with vacuolization in the submandibular glands.

ラット顎下腺腺房部の細胞のtight junctionにおける物質透過性に対するE-cadherinの役割

ラット顎下腺腺房部の細胞のtight junctionにおける物質透過性に対するE-cadherinの役割

2 ラット顎下腺におけるIII型コラーゲンとテネイシンの加齢変動 (第445回 大阪歯科学会例会)

2 ラット顎下腺におけるIII型コラーゲンとテネイシンの加齢変動 (第445回 大阪歯科学会例会)