RESUMO
BACKGROUND: Oxidative stress is an imbalance between the levels of reactive oxygen species (ROS), reactive nitrogen species (RNS) and endogenous antioxidants. The aetiology and pathogenesis of several oral diseases are attributed to this process. The antioxidant enzymes secreted in the saliva by submandibular glands maintain oral health through the scavenging of ROS. The objective of this work was to study the capacity of an aqueous extract of L. divaricata (AE), and its majority compound, nordihydroguariaretic acid (NDGA), to modulate the pro-oxidant/antioxidant status in submandibular glands in a model of oxidative stress induced by streptozotocin (STZ) in rats. METHODS: To induce oxidative stress with STZ, a group of animals was treated i.p. with 1 X PBS (control group) and other group was injected i.p. once with STZ (60 mg/kg). Ten days after the treatment, blood samples were taken from the tail vain to determine the glucose levels. Animals with glucose values ≥300 mg/ml were selected. The submandibular glands of control and STZ treated animals were incubated with either the AE (500 µg/ml) or with NDGA (1.5 µg/ml), and the content of malondialdehyde (MDA), protein carbonyl groups, ROS and RNS, and the activity and expression of peroxidase (Px), superoxide dismutase (SOD) and inducible nitric oxide synthase (iNOS) were assayed. RESULTS: AE decreased the levels of MDA (##P < 0.01) and protein carbonyl groups (#P < 0.05), and modulated the levels of ROS such as hydrogen peroxide (H2O2)(##P < 0.01), superoxide anion (O2.-) (#P < 0.05) and nitric oxide (NO) (#P < 0.05) in relation to the modulation of Px and iNOS expression. NDGA was found to be involved in these effects. CONCLUSIONS: The antioxidant activity of the AE in the submandibular glands would allow the maintenance of the antioxidant pool to prevent oral oxidative diseases.
Assuntos
Diabetes Mellitus Experimental/metabolismo , Larrea/química , Masoprocol/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Glândula Submandibular/efeitos dos fármacos , Animais , Antioxidantes/farmacologia , Feminino , Malondialdeído/análise , Oxirredutases/análise , Ratos , Ratos Wistar , Glândula Submandibular/química , Glândula Submandibular/enzimologiaRESUMO
OBJECTIVE: This study aimed to demonstrate the immunohistochemical changes associated with MMP-2 and type 1 collagen separately for the first time in the major salivary glands (the parotid, submaxillary, and sublingual glands) that occur with aging in mice. MATERIAL AND METHODS: Fourteen Balb/c white mice (50-80 g) were used in this study. The animals were divided into two equal groups. Group I consisted of young animals (2-month-old) (n=7) and Group II consisted of older animals (18-month-old) (n=7). After routine histological follow-ups, Hematoxylin-eosin (H&E), Masson's Trichrome staining and immunohistochemical staining was performed for type I collagen and MMP-2. RESULTS: We observed that there were age-related decreases in the number of acinar cells, increase in eosinophilic zymogen granules in cells, collagen accumulation in fibrotic areas and dilatation in interlobular ducts. Also, while type I collagen and MMP-2 immunoreactivity were moderate in the salivary glands of the young mice, they were high in the salivary glands of the old mice (p=0.001). In the H-score assessment, MMP-2 immunoreactivity was lower at a significant level in young mice than in old mice (p=0.001). CONCLUSIONS: This study showed that anatomical, physiological and morphological abnormalities occur in all three major salivary glands as a natural consequence of aging.
Assuntos
Colágeno Tipo I/análise , Metaloproteinase 2 da Matriz/análise , Glândula Parótida/química , Glândula Sublingual/química , Glândula Submandibular/química , Fatores Etários , Animais , Feminino , Imuno-Histoquímica , Camundongos Endogâmicos BALB C , Glândula Parótida/patologia , Glândula Parótida/fisiopatologia , Valores de Referência , Glândula Sublingual/patologia , Glândula Sublingual/fisiopatologia , Glândula Submandibular/patologia , Glândula Submandibular/fisiopatologiaRESUMO
Abstract Objective This study aimed to demonstrate the immunohistochemical changes associated with MMP-2 and type 1 collagen separately for the first time in the major salivary glands (the parotid, submaxillary, and sublingual glands) that occur with aging in mice. Material and Methods Fourteen Balb/c white mice (50-80 g) were used in this study. The animals were divided into two equal groups. Group I consisted of young animals (2-month-old) (n=7) and Group II consisted of older animals (18-month-old) (n=7). After routine histological follow-ups, Hematoxylin-eosin (H&E), Masson's Trichrome staining and immunohistochemical staining was performed for type I collagen and MMP-2. Results We observed that there were age-related decreases in the number of acinar cells, increase in eosinophilic zymogen granules in cells, collagen accumulation in fibrotic areas and dilatation in interlobular ducts. Also, while type I collagen and MMP-2 immunoreactivity were moderate in the salivary glands of the young mice, they were high in the salivary glands of the old mice (p=0.001). In the H-score assessment, MMP-2 immunoreactivity was lower at a significant level in young mice than in old mice (p=0.001). Conclusions This study showed that anatomical, physiological and morphological abnormalities occur in all three major salivary glands as a natural consequence of aging.
Assuntos
Animais , Feminino , Glândula Parótida/química , Glândula Sublingual/química , Glândula Submandibular/química , Metaloproteinase 2 da Matriz/análise , Colágeno Tipo I/análise , Glândula Parótida/fisiopatologia , Glândula Parótida/patologia , Valores de Referência , Glândula Sublingual/fisiopatologia , Glândula Sublingual/patologia , Glândula Submandibular/fisiopatologia , Glândula Submandibular/patologia , Imuno-Histoquímica , Fatores Etários , Camundongos Endogâmicos BALB CRESUMO
Saliva is the first barrier to entry of bacteria and viruses into the body. The submandibular glands (SMG) contribute to the maintenance of oral health and regulation of immune/ inflam matory responses. Previous studies suggest that transforming growth factor beta 1 (TGFB1) may contribute to salivary gland fibrosis but the expression of the TGFB1 system in the SMG has not been elucidated. Thus, the aim of this study was to analyze in rat SMG the immunolocalization of TGFB1 and its specific receptors ALK5 (profibrotic) and ALK1 (proproliferative) and the coreceptor endoglin (EDG) in a bilateral experimental periodontitis (EP) model (cotton thread ligature around the neck of the first lower molars) for 1 and 6 weeks. Fixed SMG were embedded in paraffin and serially cut for routine hematoxylin-eosin staining for histological analysis or immunohistochemical techniques by diaminobenzidine detection. SMG histology from animals with EP showed timedependent structural changes involving marked reduction in the height of the contoured ducts, cell destruction, loss of secretory granules, periductal congestion and excess connective tissue surrounding these ducts indicative of a fibrotic process, compared to control SMG. TGFB1, ALK5 and ALK1 receptors and the coreceptor EDG were mainly immunolocalized in ductal cells and in the fibrotic areas in EP groups. The expression of the profibrotic ALK5 receptor was increased in areas of fibrosis in SMG of animals with EP. In SMG of rats with EP, the localization of the TGFB1 specific receptors in the ducts and cells from fibrotic areas, due to the expression of TGFB1 in the surrounding areas, might indicate paracrine and autocrine actions exerted by TGFB1 via its specific receptors. The results of this study suggest that TGFB1 promotes fibrosis, inducing cell proliferation via ALK1 and EDG receptors and stimulates fibrosis relatedprocesses via ALK5 receptor, which could lead to abnormal secretor activity of the SMG during periodontal disease.
La saliva es la primera barrera para la entrada de bacterias y virus en el cuerpo. Las glándulas submandibulares (GSM) contribuyen al mantenimiento de la salud oral y a la regulación de las respuestas inmunoinflamatorias. Estudios previos sugieren que el factor de crecimiento transformante beta 1 (TGFB1) puede contribuir a la fibrosis de las glándulas salivales, pero la expresión y localización del sistema TGFB1 en las GSM no ha sido dilucidada. El objetivo del presente trabajo fue analizar por inmunohistoquímica en las GSM de ratas la expresión de TGFB1 y sus receptores específicos ALK5 (profibrótico) y ALK1 (proproliferativo) y el coreceptor endoglina (EDG) en un modelo de periodontitis bilateral experimental (PE) (hilo de algodón alrededor del cuello de los primeros molares inferiores) durante 1 y 6 semanas. Las GSM fueron fijadas y embebidas en parafina para realizar cortes seriados los cuales se tiñeron con hematoxilinaeosina para analizar la histología o se procesaron para realizar la técnica de inmunohistoquímica mediante detección con diaminobenzidine. La histología de las GSM de animales con PE reveló cambios estructurales tiempo dependientes, con una marcada reducción de la altura de los conductos, destrucción celular, pérdida de gránulos secretores, congestión periductal y exceso de tejido conectivo que rodea los conductos, indicando un proceso de fibrosis respecto de las GSM de animales control. TGFB1, ALK5 y ALK1 y el coreceptor EDG fueron principalmente inmunolocalizados en las células que forman los ductos y en las áreas de fibrosis en los grupos con PE. La expresión del receptor profibrótico ALK5 se incrementó en las áreas de fibrosis en GSM de animales con PE. En GSM de ratas con PE, la localización de los receptores específicos de TGFB1 en las células de los conductos y áreas de fibrosis, junto con la expresión de TGFB1 en las áreas circundantes, podría indicar acciones paracrinas y autocrinas ejercidas por TGFB1 a través de sus receptores específicos. Los resultados de este estudio sugieren que TGFB1 podría inducir un proceso de fibrosis promoviendo la proliferación celular a través de los receptores ALK1 y EDG, y favoreciendo procesos relacionados con la fibrosis a través de su receptor ALK5, lo que conduciría a una actividad secretora anormal de la GSM durante la enfermedad periodontal.
Assuntos
Glândula Submandibular/patologia , Fator de Crescimento Transformador beta1/biossíntese , Animais , Fibrose/etiologia , Imuno-Histoquímica , Masculino , Periodontite/complicações , Ratos , Ratos Wistar , Glândula Submandibular/química , Fator de Crescimento Transformador beta1/análiseRESUMO
Saliva is the first barrier to entry of bacteria and viruses into the body. The submandibular glands (SMG) contribute to the maintenance of oral health and regulation of immune/ inflam matory responses. Previous studies suggest that transforming growth factor beta 1 (TGFB1) may contribute to salivary gland fibrosis but the expression of the TGFB1 system in the SMG has not been elucidated. Thus, the aim of this study was to analyze in rat SMG the immunolocalization of TGFB1 and its specific receptors ALK5 (profibrotic) and ALK1 (proproliferative) and the coreceptor endoglin (EDG) in a bilateral experimental periodontitis (EP) model (cotton thread ligature around the neck of the first lower molars) for 1 and 6 weeks. Fixed SMG were embedded in paraffin and serially cut for routine hematoxylin-eosin staining for histological analysis or immunohistochemical techniques by diaminobenzidine detection. SMG histology from animals with EP showed timedependent structural changes involving marked reduction in the height of the contoured ducts, cell destruction, loss of secretory granules, periductal congestion and excess connective tissue surrounding these ducts indicative of a fibrotic process, compared to control SMG. TGFB1, ALK5 and ALK1 receptors and the coreceptor EDG were mainly immunolocalized in ductal cells and in the fibrotic areas in EP groups. The expression of the profibrotic ALK5 receptor was increased in areas of fibrosis in SMG of animals with EP. In SMG of rats with EP, the localization of the TGFB1 specific receptors in the ducts and cells from fibrotic areas, due to the expression of TGFB1 in the surrounding areas, might indicate paracrine and autocrine actions exerted by TGFB1 via its specific receptors. The results of this study suggest that TGFB1 promotes fibrosis, inducing cell proliferation via ALK1 and EDG receptors and stimulates fibrosis relatedprocesses via ALK5 receptor, which could lead to abnormal secretor activity of the SMG during periodontal disease.
La saliva es la primera barrera para la entrada de bacterias y virus en el cuerpo. Las glándulas submandibulares (GSM) contribuyen al mantenimiento de la salud oral y a la regulación de las respuestas inmunoinflamatorias. Estudios previos sugieren que el factor de crecimiento transformante beta 1 (TGFB1) puede contribuir a la fibrosis de las glándulas salivales, pero la expresión y localización del sistema TGFB1 en las GSM no ha sido dilucidada. El objetivo del presente trabajo fue analizar por inmunohistoquímica en las GSM de ratas la expresión de TGFB1 y sus receptores específicos ALK5 (profibrótico) y ALK1 (proproliferativo) y el coreceptor endoglina (EDG) en un modelo de periodontitis bilateral experimental (PE) (hilo de algodón alrededor del cuello de los primeros molares inferiores) durante 1 y 6 semanas. Las GSM fueron fijadas y embebidas en parafina para realizar cortes seriados los cuales se tiñeron con hematoxilinaeosina para analizar la histología o se procesaron para realizar la técnica de inmunohistoquímica mediante detección con diaminobenzidine. La histología de las GSM de animales con PE reveló cambios estructurales tiempo dependientes, con una marcada reducción de la altura de los conductos, destrucción celular, pérdida de gránulos secretores, congestión periductal y exceso de tejido conectivo que rodea los conductos, indicando un proceso de fibrosis respecto de las GSM de animales control. TGFB1, ALK5 y ALK1 y el coreceptor EDG fueron principalmente inmunolocalizados en las células que forman los ductos y en las áreas de fibrosis en los grupos con PE. La expresión del receptor profibrótico ALK5 se incrementó en las áreas de fibrosis en GSM de animales con PE. En GSM de ratas con PE, la localización de los receptores específicos de TGFB1 en las células de los conductos y áreas de fibrosis, junto con la expresión de TGFB1 en las áreas circundantes, podría indicar acciones paracrinas y autocrinas ejercidas por TGFB1 a través de sus receptores específicos. Los resultados de este estudio sugieren que TGFB1 podría inducir un proceso de fibrosis promoviendo la proliferación celular a través de los receptores ALK1 y EDG, y favoreciendo procesos relacionados con la fibrosis a través de su receptor ALK5, lo que conduciría a una actividad secretora anormal de la GSM durante la enfermedad periodontal.
Assuntos
Animais , Masculino , Ratos , Glândula Submandibular/patologia , Fator de Crescimento Transformador beta1/biossíntese , Periodontite/complicações , Glândula Submandibular/química , Fibrose/etiologia , Imuno-Histoquímica , Ratos Wistar , Fator de Crescimento Transformador beta1/análiseRESUMO
The aim of the present work was to study the effect of a restricted diet on carbohydrate metabolism in submandibular glands of female prepuber rats and the influence of arachidonic acid derivatives. Rats of 21 days of age were divided into three groups. Group I: normally fed rats. Group II: restricted diet (50% of the normal food intake). Group III: normally restricted diet with re-feeding. The baseline concentration of tissue glycogen was significantly higher in Group II than in I or III and after 60 min of incubation in a glucose free medium all groups showed a glycogen drop. In addition, the glucose metabolism was increased in Group II. Cycloxigenase inhibitors failed to alter (14)CO(2) levels in Groups I and III. In Group II, indomethacin and acetylsalicylic acid inhibited glucose metabolism, which was reverted by PGE(2) addition. The exogenous arachidonic acid metabolism and different eicosanoids showed that restricted diet significantly increased the production of PGE(2) but diminished PGF(2)(alpha) production. Our results suggest that a restricted diet would lead to a new dynamic equilibrium of glucose homeostasis. Prostaglandins E(2) and F(2)(alpha) would participate by adapting the source of energy to tissue demands while maintaining the metabolic features that characterize puberty.
Assuntos
Dieta , Dinoprostona/fisiologia , Glucose/metabolismo , Glândula Submandibular/metabolismo , Animais , Ácido Araquidônico/administração & dosagem , Ácido Araquidônico/metabolismo , Metabolismo dos Carboidratos/efeitos dos fármacos , Dióxido de Carbono/análise , Inibidores de Ciclo-Oxigenase/farmacologia , Feminino , Glicogênio/análise , Indometacina/farmacologia , Ratos , Ratos Wistar , Glândula Submandibular/químicaRESUMO
The mandibular gland secretions of worker castes from wild colonies of the leaf-cutting ants Atta sexdens sexdens and Atta opaciceps were analyzed quantitatively by gas chromatography-mass spectrometry. The secretions comprised a complex mixture of volatile, mainly oxygenated compounds, and their profiles exhibited considerable qualitative and quantitative variations among species and castes. The known alarm pheromone 4-methyl-3-heptanone was common to both species. The elevated relative proportions of this ketone found in the secretions of gardeners and generalists suggest that such castes are primarily responsible for the production and release of the alarm pheromone. Quantitative variations (but no qualitative differences) in the profiles of secretions of soldiers from different colonies of A. sexdens sexdens were detected, supporting the view that intraspecific colony recognition is mediated through mandibular gland secretions. Subsequent laboratory assays showed that, among the compounds identified by GC-MS, 4-methyl-3-heptanone elicited a strong alarm response in workers of A. sexdens sexdens and A. opaciceps.
Assuntos
Formigas/química , Folhas de Planta/fisiologia , Glândula Submandibular/química , Glândula Submandibular/metabolismo , Animais , Comportamento Animal , Espectrometria de Massas , Óleos Voláteis/análise , Óleos Voláteis/químicaRESUMO
It was analyzed the forms of renin produced by a mouse immortalized mesangial cell line (MIC) and their ability to generate angiotensin II (AII). The synthesis, localization and secretion of renin and AII by MIC were evaluated under conditions of normal (10 mM) or high (30 mM) glucose concentration. Two major bands of 35 kDa and 70 kDa were observed in SDS-PAGE. The amino-terminal sequencing revealed the presence of prorenin and renin in these bands with higher homology with the submaxillary gland form of renin. Renin and AII were detected in cell lysate and in culture medium, indicating that MIC synthesize and secrete these peptides. Renin was localized in the cytoplasm while AII was seen predominantly inside the nucleus. High glucose induced an increase in the synthesis and secretion of renin and AII. Results suggest that MIC produce AII and a renin form similar to the submandibular. Intracellular AII may be directed at the nucleus and/or be secreted, indicating that AII may directly influences gene expression in these cells. The mechanisms of synthesis and secretion of renin and AII are potentially modified by high glucose concentration, suggesting a possible role of AII produced by mesangial cells in diabetic nephropathy.
Assuntos
Mesângio Glomerular/efeitos dos fármacos , Mesângio Glomerular/metabolismo , Glucose/farmacologia , Renina/metabolismo , Glândula Submandibular/química , Sequência de Aminoácidos , Angiotensina II/metabolismo , Animais , Western Blotting , Mesângio Glomerular/citologia , Imuno-Histoquímica , Manitol/metabolismo , Camundongos , Dados de Sequência Molecular , Processamento de Proteína Pós-Traducional , Transporte Proteico , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Renina/química , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alinhamento de Sequência , Tripsina/metabolismoRESUMO
Cell proliferation and apoptosis as well as cell-cell adhesion and communication are essential processes that assure cell survival, renewal and coordination. Since junctional proteins have a tumor suppressor activity, their immunohistochemical characterization has diagnostic and prognostic value. The purpose of this report is to review the role played by junctional and proliferation-related proteins in the salivary glands and to illustrate their immunohistochemical localisation in normal murine submandibular gland. Normal salivary gland tissue was obtained from normal adult male BALB/c mice. After immediate fixation in formalin and ethanol, the samples were immunohistochemically stained for E-cadherin (HECD-1), Bcl-2, Ki67 (MIB-1), connexin26 and connexin 32, beta-catenin and gamma-catenin. Their topological distribution and reactivity were evaluated by light microscopy. The nuclei of submandibular acinar cells exhibited low to moderate staining for Ki67, but no reaction was observed in ductal cells. Murine Bcl-2 was light to moderately expressed in the latero-basal domain of cells of submandibular acini but was only lightly expressed in striated and eosinophilic ducts. The lateral domain of acinar cells were heavily stained with anti-E-cadherin, while only low levels were expressed at the cellular surface of ducts. beta-Catenin was consistently and evenly distributed along the latero-apical boundaries of eosinophilic secretory duct cells as well as on the lateral domain of acinar cells. On the contrary, gamma-catenin was generally expressed at lower levels than beta-catenin, was not expressed in ductal cells and was only lightly stained on the lateral membranes of acinar cells. No expression of connexin 32 was observed in ducts but it was significantly expressed in a spotted pattern along the plasma membrane of acinic cells. Connexin 26 showed similar localization to that of connexin 32 but the staining was much more intense. Since these proteins have been reported to play key roles in maintaining homeostasis via control of cell growth, differentiation and death, their analysis in normal salivary tissue will hopefully contribute to the study of salivary tumorigenesis.
Assuntos
Caderinas/análise , Proteínas do Citoesqueleto/análise , Antígeno Ki-67/análise , Proteínas Proto-Oncogênicas c-bcl-2/análise , Glândula Submandibular/química , Transativadores/análise , Animais , Apoptose , Biomarcadores/análise , Adesão Celular , Técnicas Imunoenzimáticas , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Glândula Submandibular/citologia , beta CateninaRESUMO
Ablation of host submaxillary glands modifies Ehrlich tumor growth and tumor-infiltrating leukocytes, possibly by modifications in the serum level of growth factors produced by this gland. To extend this research, 7-month-old male EPM-1 mice (N = 30) were divided into two groups: 1) inoculated with tumor cells previously incubated with submaxillary salivary gland extract (SGE) in PBS for 30 min at 37%; 2) inoculated with tumor cells previously incubated with PBS, under the same conditions. Animals were inoculated into the footpad with 40 microl of a suspension containing 4.5 x 10(7) tumor cells/ml, and footpad thickness was measured daily for 10 days. Sections and smears of tumor cells were prepared from the tumor mass to determine mitosis frequency, percent of tumor cells immunopositive to nerve (NGF) and epidermal (EGF) growth factors and percent of tumor-infiltrating leukocytes. The incubation of tumor cells with SGE produced a tumor reduction of about 30% in size (P<0.01). This effect was not related to loss of cell viability during incubation, but a 33% increase (P<0.05) in the percentage of dead or dying tumor cells and a 15% increase in the percent of NGF/EGF-positive tumor cells (P<0.01) were observed in vivo at the end of experiment. Tumor-infiltrating lymphocytes and mitosis frequency did not differ between groups. These data suggest a direct effect of factors present in SGE on tumor cells, which induce degeneration of tumor cells.