RESUMO
The endocrine pancreas is composed of clusters of cell groups called pancreatic islets. These cells are responsible for the synthesis and secretion of hormones crucial for glycemic homeostasis, such as insulin and glucagon. Therefore, these cells were the targets of many studies. One method to study and/or understand endocrine pancreatic physiology is the isolation of these islets and stimulation of hormone production using different concentrations of glucose, agonists, and/or antagonists of specific secretagogues and mimicking the stimulation of hormonal synthesis and secretion. Many researchers studied pancreatic physiology in murine models due to their ease of maintenance and rapid development. However, the isolation of pancreatic islets involves meticulous processes that may vary between rodent species. The present study describes a simple and effective technical protocol for isolating intact islets from mice and rats for use as a practical guide for researchers. The method involves digestion of the acinar parenchyma by intraductal collagenase. Isolated islets are suitable for in vitro endocrine secretion analyses, microscopy techniques, and biochemical analyses.
Assuntos
Ilhotas Pancreáticas , Animais , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/citologia , Camundongos , Ratos , Masculino , Camundongos Endogâmicos C57BL , Separação Celular/métodosRESUMO
Cell-to-cell interactions mediated by intercellular junctions (IJs) are crucial for beta-cell functioning and proper insulin secretion, however, their role in type-2 diabetes is still unclear. This work aimed to evaluate the cellular distribution and expression of proteins associated with adherens (AJs) and gap junctions (GJs) in pancreatic islets of C57BL6 mice fed a high-fat (HF) diet. The administration of HF diet for 30 days induced an increase in body weight, post-prandial glycemia, insulinemia, glucose intolerance, and moderate insulin resistance associated with mild perturbations in insulin secretion. The intercellular content of the AJ-associated proteins (namely, E-, N-cadherins, and α-, ß-catenins) was significantly higher in islet cells of HF-fed mice. Inversely, the gap junctional content of Cx36 was significantly decreased, as revealed by immunofluorescence, which was paralleled by a reduction in the frequency of calcium oscillations in islets of prediabetic mice. In conclusion, the endocrine pancreas displays significant changes in the content of several junctional proteins at the cell-cell contact region following short-term HF diet administration, indicating that IJs may be involved in the adaptive response of beta cells seen during this state.
Assuntos
Células Secretoras de Insulina , Ilhotas Pancreáticas , Animais , Moléculas de Adesão Celular/metabolismo , Dieta Hiperlipídica/efeitos adversos , Insulina/metabolismo , Células Secretoras de Insulina/metabolismo , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Wnt proteins act mainly as paracrine signals regulating cell proliferation and differentiation. The canonical Wnt pathway has recently been associated with pancreas development and the onset of type 2 diabetes in rodent and human but the underlying mechanisms are still unclear. The aim of this work was threefold: (a) to screen for Wnt expressed by murine pancreas/islet cells, (b) to investigate whether the Wnt gene expression profile can be changed in hyperplastic islets from type 2 prediabetic mice (fed a high-fat diet), and (c) to verify whether soluble factors (namely Wnts) released by pancreatic islets affect insulin secretion and proliferation of a beta-cell line in vitro condition. The majority of the Wnt subtypes are expressed by islet cells, such as Wnts 2, 2b, 3, 3a, 4, 5a, 5b, 6, 7a, 7b, 8a, 8b, 9a, 9b, and 11, while in the whole pancreas homogenates were found the same subtypes, except Wnts 3, 6, 7a, and 7b. Among all the Wnts, the Wnts 3a and 5b showed a significantly increased gene expression in hyperplastic islets from prediabetic mice compared with those from control mice. Furthermore, we observed that coculture with hyperplastic or nonhyperplastic islets did not change the secretory function of the mouse insulinoma clone 6 (MIN6) beta cells but induced a significant increase in cell proliferation in this lineage, which was partially blocked by the IWR-1 and IWP-2 Wnt inhibitors. In conclusion, we demonstrated that murine pancreas/islet cells can secrete Wnts, and that islet-released Wnts may participate in the regulation of beta-cell mass under normal and prediabetic conditions.
Assuntos
Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/metabolismo , Proteínas Wnt/metabolismo , Tecido Adiposo/metabolismo , Animais , Linhagem Celular , Proliferação de Células , Dieta Hiperlipídica , Embrião de Mamíferos/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Secreção de Insulina , Masculino , Camundongos Endogâmicos C57BL , Estado Pré-Diabético/genética , Estado Pré-Diabético/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Wnt/genética , Via de Sinalização WntRESUMO
Background: The increase in the life expectancy of dogs and cats in recent decades has heightened the incidence of aging-associated diseases. Among such diseases, endocrinopathies, such as obesity, stand out. In addition, there exists a lack of professionals with expertise in this area. Therefore, proper knowledge of the characteristics of patients affected by endocrinopathies as well as the frequency of these diseases aid in the recognition of endocrine syndromes in the general population. Thus, the aim of this study was to conduct a retrospective analysis of medical charts of dogs and cats compiled by a Division of Endocrinology in southern Brazil during a 10-year period. Materials, Methods & Results: An epidemiological survey of the medical charts of 1,400 dogs and cats compiled by a Division of Endocrinology between 2004 and 2014 was performed. The diagnostic data based on laboratory and hormone tests and on ultrasound scans were used as gold standard. The data were tabulated using Excel for Windows for the stratified analysis of the diagnoses. The major diseases (93.4% of the cases) detected in dogs were hyperadrenocorticism (37%), diabetes mellitus (22%), hypothyroidism (11%), and overweight/obesity (8%), whereas the main diseases (6.6% of the cases) detected in cats were diabetes mellitus (42%) and hyperthyroidism (23%). Some rare and uncommon diseases were diagnosed [...](AU)
Assuntos
Animais , Gatos , Cães , Doenças do Sistema Endócrino/epidemiologia , Glândulas Suprarrenais/fisiologia , Obesidade/epidemiologia , Obesidade/veterinária , Hiperfunção Adrenocortical/veterinária , Diabetes Mellitus/veterinária , Hipotireoidismo/veterinária , Sobrepeso/veterináriaRESUMO
Background: The increase in the life expectancy of dogs and cats in recent decades has heightened the incidence of aging-associated diseases. Among such diseases, endocrinopathies, such as obesity, stand out. In addition, there exists a lack of professionals with expertise in this area. Therefore, proper knowledge of the characteristics of patients affected by endocrinopathies as well as the frequency of these diseases aid in the recognition of endocrine syndromes in the general population. Thus, the aim of this study was to conduct a retrospective analysis of medical charts of dogs and cats compiled by a Division of Endocrinology in southern Brazil during a 10-year period. Materials, Methods & Results: An epidemiological survey of the medical charts of 1,400 dogs and cats compiled by a Division of Endocrinology between 2004 and 2014 was performed. The diagnostic data based on laboratory and hormone tests and on ultrasound scans were used as gold standard. The data were tabulated using Excel for Windows for the stratified analysis of the diagnoses. The major diseases (93.4% of the cases) detected in dogs were hyperadrenocorticism (37%), diabetes mellitus (22%), hypothyroidism (11%), and overweight/obesity (8%), whereas the main diseases (6.6% of the cases) detected in cats were diabetes mellitus (42%) and hyperthyroidism (23%). Some rare and uncommon diseases were diagnosed [...]
Assuntos
Animais , Gatos , Cães , Doenças do Sistema Endócrino/epidemiologia , Glândulas Suprarrenais/fisiologia , Diabetes Mellitus/veterinária , Hiperfunção Adrenocortical/veterinária , Hipotireoidismo/veterinária , Obesidade/epidemiologia , Obesidade/veterinária , Sobrepeso/veterináriaRESUMO
Diabetes mellitus represents a serious public health problem owing to its global prevalence in the last decade. The causes of this metabolic disease include dysfunction and/or insufficient number of β cells. Existing diabetes mellitus treatments do not reverse or control the disease. Therefore, β-cell mass restoration might be a promising treatment. Several restoration approaches have been developed: inducing the proliferation of remaining insulin-producing cells, de novo islet formation from pancreatic progenitor cells (neogenesis), and converting non-β cells within the pancreas to β cells (transdifferentiation) are the most direct, simple, and least invasive ways to increase β-cell mass. However, their clinical significance is yet to be determined. Hypothetically, β cells or islet transplantation methods might be curative strategies for diabetes mellitus; however, the scarcity of donors limits the clinical application of these approaches. Thus, alternative cell sources for β-cell replacement could include embryonic stem cells, induced pluripotent stem cells, and mesenchymal stem cells. However, most differentiated cells obtained using these techniques are functionally immature and show poor glucose-stimulated insulin secretion compared with native β cells. Currently, their clinical use is still hampered by ethical issues and the risk of tumor development post transplantation. In this review, we briefly summarize the current knowledge of mouse pancreas organogenesis, morphogenesis, and maturation, including the molecular mechanisms involved. We then discuss two possible approaches of β-cell mass restoration for diabetes mellitus therapy: β-cell regeneration and β-cell replacement. We critically analyze each strategy with respect to the accessibility of the cells, potential risk to patients, and possible clinical outcomes.
Assuntos
Humanos , Animais , Camundongos , Diabetes Mellitus/terapia , Células Secretoras de Insulina/transplante , Técnicas de Cultura de Células/métodos , Proliferação de Células , Reprogramação Celular , Células Secretoras de Insulina/citologia , Transplante das Ilhotas Pancreáticas , RegeneraçãoRESUMO
In this study, we investigated a possible sexual dimorphism regarding metabolic response and structural and functional adaptations of the endocrine pancreas after exposure to a high-fat diet (HFd). On chow diet, male and female C57BL/6/JUnib mice showed similar metabolic and morphometric parameters, except that female islets displayed a relatively lower ß-cell:non-ß-cell ratio. After 30 days on HFd, both male and female mice showed increased weight gain, however only the males displayed glucose intolerance associated with high postprandial glycemia when compared to their controls. After 60 days on HFd, both genders became obese, hyperglycemic, hyperinsulinemic, insulin resistant and glucose intolerant, although the metabolic changes were more pronounced in males, while females displayed greater weight gain. In both genders, insulin resistance induced by HFd feeding was compensated by expansion of ß-cell mass without changes in islet cytoarchitecture. Interestingly, we found a strong correlation between the degree of ß-cell expansion and the levels of hyperglycemia in the fed state: male mice fed a 60d-HFd, showing higher glycemic levels also displayed a greater ß-cell mass increase in comparison with female mice. Additionally, sexual dimorphism was also observed regarding the source of ß-cell mass expansion following 60d-HFd: while in males, both hypertrophy and hyperplasia (revealed by morphometry and Ki67 immunoreaction) of ß-cells were observed, female islets displayed only a significant increase in ß-cell size. In conclusion, this study describes gender differences in metabolic response to high fat diet, paralleled by distinct compensatory morphometric changes in pancreatic islets.