ABSTRACT
Diabetes mellitus, a chronic and non-transmissible disease, triggers a wide range of micro- and macrovascular complications. The differentiation of pancreatic ß-like cells (PßLCs) from induced pluripotent stem cells (iPSCs) offers a promising avenue for regenerative medicine aimed at treating diabetes. Current differentiation protocols strive to emulate pancreatic embryonic development by utilizing cytokines and small molecules at specific doses to activate and inhibit distinct molecular signaling pathways, directing the differentiation of iPSCs into pancreatic ß cells. Despite significant progress and improved protocols, the full spectrum of molecular signaling pathways governing pancreatic development and the physiological characteristics of the differentiated cells are not yet fully understood. Here, we report a specific combination of cofactors and small molecules that successfully differentiate iPSCs into PßLCs. Our protocol has shown to be effective, with the resulting cells exhibiting key functional properties of pancreatic ß cells, including the expression of crucial molecular markers (pdx1, nkx6.1, ngn3) and the capability to secrete insulin in response to glucose. Furthermore, the addition of vitamin C and retinoic acid in the final stages of differentiation led to the overexpression of specific ß cell genes.
Subject(s)
Ascorbic Acid , Cell Differentiation , Diabetes Mellitus , Induced Pluripotent Stem Cells , Insulin-Secreting Cells , Tretinoin , Insulin-Secreting Cells/metabolism , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/cytology , Ascorbic Acid/pharmacology , Induced Pluripotent Stem Cells/metabolism , Induced Pluripotent Stem Cells/cytology , Induced Pluripotent Stem Cells/drug effects , Tretinoin/pharmacology , Cell Differentiation/drug effects , Humans , Diabetes Mellitus/metabolism , Homeodomain Proteins/metabolism , Homeodomain Proteins/genetics , Signal Transduction/drug effects , Basic Helix-Loop-Helix Transcription Factors/metabolism , Basic Helix-Loop-Helix Transcription Factors/genetics , Trans-Activators/metabolism , Trans-Activators/genetics , Insulin/metabolism , Nerve Tissue ProteinsABSTRACT
The nerve growth factor (NGF) participates in cell survival and glucose-stimulated insulin secretion (GSIS) processes in rat adult beta cells. GSIS is a complex process in which metabolic events and ionic channel activity are finely coupled. GLUT2 and glucokinase (GK) play central roles in GSIS by regulating the rate of the glycolytic pathway. The biphasic release of insulin upon glucose stimulation characterizes mature adult beta cells. On the other hand, beta cells obtained from neonatal, suckling, and weaning rats are considered immature because they secrete low levels of insulin and do not increase insulin secretion in response to high glucose. The weaning of rats (at postnatal day 20 in laboratory conditions) involves a dietary transition from maternal milk to standard chow. It is characterized by increased basal plasma glucose levels and insulin levels, which we consider physiological insulin resistance. On the other hand, we have observed that incubating rat beta cells with NGF increases GSIS by increasing calcium currents in neonatal cells. In this work, we studied the effects of NGF on the regulation of cellular distribution and activity of GLUT2 and GK to explore its potential role in the maturation of GSIS in beta cells from P20 rats. Pancreatic islet cells from both adult and P20 rats were isolated and incubated with 5.6 mM or 15.6 mM glucose with and without NGF for 4 hours. Specific immunofluorescence assays were conducted following the incubation period to detect insulin and GLUT2. Additionally, we measured glucose uptake, glucokinase activity, and insulin secretion assays at 5.6 mM or 15.6 mM glucose concentrations. We observed an age-dependent variation in the distribution of GLUT2 in pancreatic beta cells and found that glucose plays a regulatory role in GLUT2 distribution independently of age. Moreover, NGF increases GLUT2 abundance, glucose uptake, and GSIS in P20 beta cells and GK activity in adult beta cells. Our results suggest that besides increasing calcium currents, NGF regulates metabolic components of the GSIS, thereby contributing to the maturation process of pancreatic beta cells.
Subject(s)
Glucokinase , Glucose Transporter Type 2 , Glucose , Insulin-Secreting Cells , Nerve Growth Factor , Animals , Male , Rats , Cells, Cultured , Glucokinase/metabolism , Glucose/metabolism , Glucose Transporter Type 2/metabolism , Insulin/metabolism , Insulin Secretion/drug effects , Insulin-Secreting Cells/metabolism , Insulin-Secreting Cells/drug effects , Nerve Growth Factor/metabolism , Nerve Growth Factor/pharmacology , Rats, WistarABSTRACT
Lifestyle modifications, metformin, and linagliptin reduce the incidence of type 2 diabetes (T2D) in people with prediabetes. The gut microbiota (GM) may enhance such interventions' efficacy. We determined the effect of linagliptin/metformin (LM) vs metformin (M) on GM composition and its relationship to insulin sensitivity (IS) and pancreatic ß-cell function (Pßf) in patients with prediabetes. A cross-sectional study was conducted at different times: basal, six, and twelve months in 167 Mexican adults with prediabetes. These treatments increased the abundance of GM SCFA-producing bacteria M (Fusicatenibacter and Blautia) and LM (Roseburia, Bifidobacterium, and [Eubacterium] hallii group). We performed a mediation analysis with structural equation models (SEM). In conclusion, M and LM therapies improve insulin sensitivity and Pßf in prediabetics. GM is partially associated with these improvements since the SEM models suggest a weak association between specific bacterial genera and improvements in IS and Pßf.
Subject(s)
Gastrointestinal Microbiome , Linagliptin , Metformin , Prediabetic State , Humans , Metformin/pharmacology , Metformin/therapeutic use , Gastrointestinal Microbiome/drug effects , Prediabetic State/drug therapy , Prediabetic State/microbiology , Male , Female , Middle Aged , Cross-Sectional Studies , Linagliptin/therapeutic use , Linagliptin/pharmacology , Hypoglycemic Agents/pharmacology , Hypoglycemic Agents/therapeutic use , Diabetes Mellitus, Type 2/drug therapy , Diabetes Mellitus, Type 2/microbiology , Diabetes Mellitus, Type 2/metabolism , Insulin Resistance , Adult , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/metabolism , AgedABSTRACT
SUMMARY: The therapeutic effect of a granulocyte-colony stimulating factor (G-CSF) biosimilar drug, zarzio, on non-alcoholic fatty liver disease (NAFLD) in a rat model was investigated in this study. Thirty-two rats were randomly divided into four groups. Groups I and II were fed a standard laboratory diet, whereas groups III and IV were fed a high fat diet (HFD) for 14 weeks. After 12 weeks of feeding, groups I and III were administered normal saline, and groups II and IV were intraperitoneally administered zarzio (200 mg/kg/day) for two consecutive weeks. Hematoxylin-eosin (H&E) staining was used to assess hepatic and pancreatic morphology in all groups, oil red O (ORO) staining for lipid accumulation, Masson's staining for fibrosis, and immunohistochemistry assay for hepatic protein expression of insulin receptor substrate 1 (IRS1), nuclear factor erythroid 2-related factor 2 (Nrf2), tumour necrosis factor alpha (TNF-α) and pancreatic caspase-3. The NAFLD rats (group III) developed hepatic steatosis with increased lipid accumulation, perisinusoidal fibrosis, upregulated IRS1, TNF-α (all P<0.05) without a significant increase in Nrf2 protein expression compared with normal control. In comparison, model rats treated with zarzio (group IV) showed significant rejuvenation of the hepatic architecture, reduction of fat accumulation, and fibrosis. This was accompanied by the upregulation of Nrf2, downregulation of IRS1 and TNF-α protein expression (all P<0.05). No correlation was detected between NAFLD and non-alcoholic fatty pancreas disease (NAFPD). However, the pancreatic β-cells in group III showed increased caspase-3 expression, which was decreased (P<0.05) in group IV. In conclusion, zarzio ameliorates NAFLD by improving the antioxidant capacity of liver cells, reducing hepatic IRS1, TNF-α protein expression and pancreatic β-cells apoptosis, suggesting that zarzio could be used as a potential therapy for NAFLD.
En este estudio se investigó el efecto terapéutico de un fármaco biosimilar del factor estimulante de colonias de granulocitos (G-CSF), zarzio, sobre la enfermedaddel hígado graso no alcohólico (NAFLD) en un modelo de rata. Treinta y dos ratas se dividieron aleatoriamente en cuatro grupos. Los grupos I y II fueron alimentados con una dieta estándar de laboratorio, mientras que los grupos III y IV fueron alimentados con una dieta alta en grasas (HFD) durante 14 semanas. Después de 12 semanas de alimentación, a los grupos I y III se les administró solución salina normal, y a los grupos II y IV se les administró zarzio por vía intraperitoneal (200 mg/kg/ día) durante dos semanas consecutivas. Se utilizó tinción de hematoxilina-eosina (H&E) para evaluar la morfología hepática y pancreática en todos los grupos, tinción con rojo aceite O (ORO) para la acumulación de lípidos, tinción de Masson para la fibrosis y ensayo de inmunohistoquímica para la expresión de la proteína hepática del sustrato 1 del receptor de insulina (IRS1), factor nuclear eritroide 2 relacionado con el factor 2 (Nrf2), factor de necrosis tumoral alfa (TNF-α) y caspasa-3 pancreática. Las ratas NAFLD (grupo III) desarrollaron esteatosis hepática con aumento de la acumulación de lípidos, fibrosis perisinusoidal, IRS1 y TNF-α regulados positivamente (todos P <0,05) sin un aumento significativo en la expresión de la proteína Nrf2 en comparación con el control normal. En comparación, las ratas modelo tratadas con zarzio (grupo IV) mostraron un rejuvenecimiento significativo de la arquitectura hepática, una reducción de la acumulación de grasa y fibrosis. Esto estuvo acompañado por la regulación positiva de Nrf2, la regulación negativa de la expresión de la proteína IRS1 y TNF-α (todas P <0,05). No se detectó correlación entre NAFLD y la enfermedad del páncreas graso no alcohólico (NAFPD). Sin embargo, las células β pancreáticas en el grupo III mostraron una mayor expresión de caspasa-3, que disminuyó (P <0,05) en el grupo IV. En conclusión, zarzio mejora la NAFLD al mejorar la capacidad antioxidante de las células hepáticas, reduciendo el IRS1 hepático, la expresión de la proteína TNF-α y la apoptosis de las células β pancreáticas, lo que sugiere que zarzio podría usarse como una terapia potencial para la NAFLD.
Subject(s)
Animals , Male , Rats , Granulocyte Colony-Stimulating Factor/administration & dosage , Biosimilar Pharmaceuticals/administration & dosage , Non-alcoholic Fatty Liver Disease/drug therapy , Immunohistochemistry , Tumor Necrosis Factor-alpha/drug effects , Disease Models, Animal , Insulin-Secreting Cells/drug effects , NF-E2-Related Factor 2 , Caspase 3 , Diet, High-Fat/adverse effectsABSTRACT
Vitamin D3 is associated with improvements in insulin resistance and glycemia. In this study, we investigated the short-term effect of 1α,25(OH)2 Vitamin D3 (1,25-D3) and cholecalciferol (vitamin D3) on the glycemia and insulin sensitivity of control and dexamethasone-induced insulin-resistance rats. 45Ca2+ influx responses to 1,25-D3 and its role in insulin secretion were investigated in isolated pancreatic islets from control rats. In vivo, 5 d treatment with 1,25-D3 (i.p.) prevented insulin resistance in dexamethasone-treated rats. Treatment with 1,25-D3 improved the activities of hepatic enzymes, serum lipids and calcium concentrations in insulin-resistant rats. 25-D3 (o.g.) does not affect insulin resistance. In pancreatic islets, 1,25-D3 increased insulin secretion and stimulated rapid response 45Ca2+ influx. The stimulatory effect of 1,25-D3 on 45Ca2+ influx was decreased by diazoxide, apamine, thapsigargin, dantrolene, 2-APB, nifedipine, TEA, PKA, PKC, and cytoskeleton inhibitor, while it was increased by glibenclamide and N-ethylmaleimide. The stimulatory effect of 1,25-D3 on 45Ca2+ influx involves the activation of L-type VDCC, K+-ATP, K+-Ca2+, and Kv channels, which augment cytosolic calcium. These ionic changes mobilize calcium from stores and downstream activation of PKC, PKA tethering vesicle traffic and fusion at the plasma membrane for insulin secretion. This is the first study highlighting the unprecedented role of 1,25-D3 (short-term effect) in the regulation of glucose homeostasis and on prevention of insulin resistance. Furthermore, this study shows the intracellular ß-cell signal transduction of 1,25-D3 through the modulation of pivotal ionic channels and proteins exhibiting a coordinated exocytosis of vesicles for insulin secretion.
Subject(s)
Cholecalciferol/analogs & derivatives , Exocytosis/drug effects , Insulin Resistance , Insulin Secretion/drug effects , Insulin/metabolism , Animals , Calcium/metabolism , Calcium Channels, L-Type/genetics , Calcium Channels, L-Type/metabolism , Cholecalciferol/metabolism , Cyclic AMP-Dependent Protein Kinases/genetics , Cyclic AMP-Dependent Protein Kinases/metabolism , Humans , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/metabolism , Islets of Langerhans/drug effects , Islets of Langerhans/metabolism , Male , Rats , Rats, WistarABSTRACT
A high caloric intake, rich in saturated fats, greatly contributes to the development of obesity, which is the leading risk factor for type 2 diabetes (T2D). A persistent caloric surplus increases plasma levels of fatty acids (FAs), especially saturated ones, which were shown to negatively impact pancreatic ß-cell function and survival in a process called lipotoxicity. Lipotoxicity in ß-cells activates different stress pathways, culminating in ß-cells dysfunction and death. Among all stresses, endoplasmic reticulum (ER) stress and oxidative stress have been shown to be strongly correlated. One main source of oxidative stress in pancreatic ß-cells appears to be the reactive oxygen species producer NADPH oxidase (NOX) enzyme, which has a role in the glucose-stimulated insulin secretion and in the ß-cell demise during both T1 and T2D. In this review, we focus on the acute and chronic effects of FAs and the lipotoxicity-induced ß-cell failure during T2D development, with special emphasis on the oxidative stress induced by NOX, the ER stress, and the crosstalk between NOX and ER stress.
Subject(s)
Diabetes Mellitus, Type 2/pathology , Endoplasmic Reticulum Stress , Insulin-Secreting Cells/pathology , Lipids/toxicity , NADPH Oxidases/metabolism , Oxidative Stress , Animals , Endoplasmic Reticulum Stress/drug effects , Humans , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/metabolism , Lipids/chemistry , Oxidative Stress/drug effectsABSTRACT
The endoplasmic reticulum (ER) stress is one of the mechanisms related to decreased insulin secretion and beta cell death, contributing to the progress of type 2 diabetes mellitus (T2D). Thus, investigating agents that can influence this process would help prevent the development of T2D. Recently, the growth-hormone-releasing hormone (GHRH) action has been demonstrated in INS-1E cells, in which it increases cell proliferation and insulin secretion. As the effects of GHRH and its agonists have not been fully elucidated in the beta cell, we proposed to investigate them by evaluating the role of the GHRH agonist, MR-409, in cells under ER stress. Our results show that the agonist was unable to ameliorate or prevent ER stress. However, cells exposed to the agonist showed less oxidative stress and greater survival even under ER stress. The mechanisms by which GHRH agonist, MR-409, leads to these outcomes require further investigation.
Subject(s)
Endoplasmic Reticulum Stress/drug effects , Indoles/adverse effects , Insulin-Secreting Cells/cytology , Sermorelin/analogs & derivatives , Animals , Cell Line , Cell Proliferation/drug effects , Cell Survival , Gene Expression Regulation/drug effects , Growth Hormone-Releasing Hormone/agonists , Growth Hormone-Releasing Hormone/metabolism , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/metabolism , Oxidative Stress/drug effects , Proto-Oncogene Proteins c-bcl-2/metabolism , Rats , Sermorelin/pharmacologyABSTRACT
Since prenatal glucocorticoids (GC) excess increases the risk of metabolic dysfunctions in the offspring and its effect on ß-cell recovery capacity remains unknown we investigated these aspects in offspring from mice treated with dexamethasone (DEX) in the late pregnancy. Half of the pups were treated with streptozotocin (STZ) on the sixth postnatal day (PN). Functional and molecular analyses were performed in male offspring on PN25 and PN225. Prenatal DEX treatment resulted in low birth weight. At PN25, both the STZ-treated offspring developed hyperglycemia and had lower ß-cell mass, in parallel with higher α-cell mass and glucose intolerance, with no impact of prenatal DEX on such parameters. At PN225, the ß-cell mass was partially recovered in the STZ-treated mice, but they remained glucose-intolerant, irrespective of being insulin sensitive. Prenatal exposition to DEX predisposed adult offspring to sustained hyperglycemia and perturbed islet function (lower insulin and higher glucagon response to glucose) in parallel with exacerbated glucose intolerance. ß-cell-specific knockdown of the Hnf4α in mice from the DS group resulted in exacerbated glucose intolerance. We conclude that high GC exposure during the prenatal period exacerbates the metabolic dysfunctions in adult life of mice exposed to STZ early in life, resulting in a lesser ability to recover the islets' function over time. This study alerts to the importance of proper management of exogenous GCs during pregnancy and a healthy postnatal lifestyle since the combination of adverse factors during the prenatal and postnatal period accentuates the predisposition to metabolic disorders in adult life.
Subject(s)
Dexamethasone/toxicity , Glucocorticoids/toxicity , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/physiology , Animals , Animals, Genetically Modified , Animals, Newborn , Dexamethasone/administration & dosage , Female , Gene Expression Regulation/drug effects , Glucocorticoids/administration & dosage , Glucose Tolerance Test , Insulin/pharmacology , Mice , Neoplasms, Experimental , Pregnancy , Prenatal Exposure Delayed Effects , RNA, Messenger/genetics , RNA, Messenger/metabolismABSTRACT
Metformin is an antidiabetic drug used for the treatment of diabetes and metabolic diseases. Imbalance in the autonomic nervous system (ANS) is associated with metabolic diseases. This study aimed to test whether metformin could improve ANS function in obese rats. Obesity was induced by neonatal treatment with monosodium L-glutamate (MSG). During 21-100 days of age, MSG-rats were treated with metformin 250 mg/kg body weight/day or saline solution. Rats were euthanized to evaluate biometric and biochemical parameters. ANS electrical activity was recorded and analyzed. Metformin normalized the hypervagal response in MSG-rats. Glucose-stimulated insulin secretion in isolated pancreatic islets increased in MSG-rats, while the cholinergic response decreased. Metformin treatment normalized the cholinergic response, which involved mostly the M3 muscarinic acetylcholine receptor (M3 mAChR) in pancreatic beta-cells. Protein expression of M3 mAChRs increased in MSG-obesity rats, while metformin treatment decreased the protein expression by 25%. In conclusion, chronic metformin treatment was effective in normalizing ANS activity and alleviating obesity in MSG-rats.
Subject(s)
Autonomic Nervous System/drug effects , Hypoglycemic Agents/therapeutic use , Metformin/therapeutic use , Obesity/drug therapy , Acetylcholine/pharmacology , Animals , Glucose/pharmacology , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/metabolism , Male , Neostigmine/pharmacology , Obesity/chemically induced , Obesity/metabolism , Obesity/physiopathology , Rats, Wistar , Receptor, Muscarinic M3/metabolism , Sodium Glutamate , Vagus Nerve/drug effects , Vagus Nerve/physiologyABSTRACT
PURPOSE: The overexposure to synthetic glucocorticoids (GC) during pregnancy can predispose to metabolic diseases during adulthood. Vitamin D is not only crucial for fetal development, but also exerts direct effects on the GC sensitivity and down-regulates GC receptors. Given the vitamin D effects on glucocorticoid-related parameters, we aimed to investigate a possible protective role of maternal vitamin D administration on the glucose homeostasis of rats exposed to dexamethasone in utero. METHODS: Pregnant rats received dexamethasone (0.1 mg/kg, Dex) daily between the 14th and 19th days of pregnancy. A subgroup of dexamethasone-treated dams received oral administration of vitamin D (500UI, DexVD) during the whole gestation. The corresponding control groups of dams were included (CTL and VD groups, respectively). Male and female offspring were evaluated at 3, 6 and 12 months of age. RESULTS: Prenatal exposure to dexamethasone caused metabolic disruption in an age and sex-dependent manner being the older male offspring more susceptible to insulin resistance, fatty liver and beta-cell mass expansion than females. Furthermore, we demonstrated that prenatal GC led to glucose intolerance in male and female offspring in an age-dependent manner. Maternal vitamin D administration did not influence glucose intolerance but attenuated the insulin resistance, liver lipid accumulation and prevented the beta-cell mass expansion caused by prenatal dexamethasone in the male offspring. CONCLUSION: Maternal vitamin D administration mitigates metabolic disturbances that occur later in life in male rats exposed to GC in utero. Moreover, our data suggest vitamin D as an important nutritional supplement for pregnant overexposed to GC during gestation.
Subject(s)
Dexamethasone/adverse effects , Glucocorticoids/adverse effects , Metabolic Diseases/drug therapy , Prenatal Exposure Delayed Effects/drug therapy , Vitamin D/therapeutic use , Vitamins/therapeutic use , Animals , Female , Insulin-Secreting Cells/drug effects , Lipid Metabolism/drug effects , Male , Maternal-Fetal Exchange , Metabolic Diseases/blood , Metabolic Diseases/chemically induced , Metabolic Diseases/metabolism , Pregnancy , Prenatal Exposure Delayed Effects/blood , Prenatal Exposure Delayed Effects/chemically induced , Prenatal Exposure Delayed Effects/metabolism , Rats, Wistar , Sex Characteristics , Triglycerides/blood , Triglycerides/metabolism , Vitamin D/pharmacology , Vitamins/pharmacologyABSTRACT
Obesity and type 2 diabetes (T2D) are growing health problems associated with a loss of insulin sensitivity. Both conditions arise from a long-term energy imbalance, and frequently, lifestyle measures can be useful in its prevention, including physical activity and a healthy diet. Pancreatic ß-cells are determinant nutrient sensors that participate in energetic homeostasis needs. However, when pancreatic ß-cells are incapable of secreting enough insulin to counteract the reduced sensitivity, the pathology evolves to an insulin resistance condition. The primary nutrient that stimulates insulin secretion is glucose, but also, there are multiple dietary and hormonal factors influencing that response. Many studies of the physiology of ß-cells have highlighted the importance of glucose, fructose, amino acids, and free fatty acids on insulin secretion. The present review summarizes recent research on how ß-cells respond to the most abundant nutrients that influence insulin secretion. Taken together, understand the subjacent mechanisms of each nutrient on ß-cells can help to unravel the effects of mixed variables and complexity in the context of ß-cell pathology.
Subject(s)
Insulin Secretion/drug effects , Insulin-Secreting Cells/physiology , Insulin/metabolism , Nutrients/pharmacology , Animals , Humans , Insulin-Secreting Cells/cytology , Insulin-Secreting Cells/drug effectsABSTRACT
We evaluated whether early-life protein restriction alters structural parameters that affect ß-cell mass on the 15th day and 20th day of gestation in control pregnant (CP), control non-pregnant (CNP), low-protein pregnant (LPP) and low-protein non-pregnant (LPNP) rats from the fetal to the adult life stage as well as in protein-restricted rats that recovered after weaning (recovered pregnant (RP) and recovered non-pregnant). On the 15th day of gestation, the CNP group had a higher proportion of smaller islets, whereas the CP group exhibited a higher proportion of islets larger than the median. The ß-cell mass was lower in the low-protein group than that in the recovered and control groups. Gestation increased the ß-cell mass, ß-cell proliferation frequency and neogenesis frequency independently of the nutritional status. The apoptosis frequency was increased in the recovered groups compared with that in the other groups. On the 20th day of gestation, a higher proportion of islets smaller than the median was observed in the non-pregnant groups, whereas a higher proportion of islets larger than the median was observed in the RP, LPP and CP groups. ß-Cell mass was lower in the low-protein group than that in the recovered and control groups, regardless of the physiological status. The ß-cell proliferation frequency was lower, whereas the apoptosis rate was higher in recovered rats compared with those in the low-protein and control rats. Thus, protein malnutrition early in life did not alter the mass of ß-cells, especially in the first two-thirds of gestation, despite the increase in apoptosis.
Subject(s)
Apoptosis , Dietary Proteins/administration & dosage , Insulin-Secreting Cells/physiology , Malnutrition , Prenatal Nutritional Physiological Phenomena , Animal Feed/analysis , Animal Nutritional Physiological Phenomena , Animals , Animals, Newborn , Diet/veterinary , Female , Gene Expression Regulation/drug effects , Glucose Tolerance Test , Insulin/metabolism , Insulin-Secreting Cells/drug effects , Pregnancy , RNA/genetics , RNA/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Rats, Wistar , Weight GainABSTRACT
Alzheimer's disease (AD) is a neurodegenerative disorder and the major cause of dementia. According to predictions of the World Health Organization, more than 150 million people worldwide will suffer from dementia by 2050. An increasing number of studies have associated AD with type 2 diabetes mellitus (T2DM), since most of the features found in T2DM are also observed in AD, such as insulin resistance and glucose intolerance. In this sense, some bile acids have emerged as new therapeutic targets to treat AD and metabolic disorders. The taurine conjugated bile acid, tauroursodeoxycholic (TUDCA), reduces amyloid oligomer accumulation and improves cognition in APP/PS1 mice model of AD, and also improves glucose-insulin homeostasis in obese and type 2 diabetic mice. Herein, we investigated the effect of TUDCA upon glucose metabolism in streptozotocin-induced AD mice model (Stz). The Stz mice that received 300 mg/kg TUDCA during 10 days (Stz + TUDCA), showed improvement in glucose tolerance and insulin sensitivity, reduced fasted and fed glycemia, increased islet mass and ß-cell area, as well as increased glucose-stimulated insulin secretion, compared with Stz mice that received only PBS. Stz + TUDCA mice also displayed lower neuroinflammation, reduced protein content of amyloid oligomer in the hippocampus, improved memory test and increased protein content of insulin receptor ß-subunit in the hippocampus. In conclusion, TUDCA treatment enhanced glucose homeostasis in the streptozotocin-induced Alzheimer's disease mice model, pointing this bile acid as a good strategy to counteract glucose homeostasis disturbance in AD pathology.
Subject(s)
Alzheimer Disease/drug therapy , Amyloid beta-Peptides/metabolism , Bile Acids and Salts/metabolism , Blood Glucose/drug effects , Hippocampus/drug effects , Insulin-Secreting Cells/drug effects , Taurochenodeoxycholic Acid/pharmacology , Alzheimer Disease/chemically induced , Alzheimer Disease/metabolism , Alzheimer Disease/pathology , Animals , Cytokines/genetics , Cytokines/metabolism , Disease Models, Animal , Glucose/metabolism , Glucose/pharmacology , Hippocampus/metabolism , Hippocampus/pathology , Inflammation/drug therapy , Insulin/blood , Insulin/metabolism , Insulin-Secreting Cells/metabolism , Insulin-Secreting Cells/pathology , Male , Memory and Learning Tests , Mice , Mice, Inbred C57BL , Real-Time Polymerase Chain Reaction , Streptozocin/toxicity , Taurochenodeoxycholic Acid/administration & dosageABSTRACT
ARHGAP21 is a RhoGAP protein implicated in the modulation of insulin secretion and energy metabolism. ARHGAP21 transient-inhibition increase glucose-stimulated insulin secretion (GSIS) in neonatal islets; however, ARHGAP21 heterozygote mice have a reduced insulin secretion. These discrepancies are not totally understood, and it might be related to functional maturation of beta cells and peripheral sensitivity. Here, we investigated the real ARHGAP21 role in the insulin secretion process using an adult mouse model of acute ARHGAP21 inhibition, induced by antisense. After ARHGAP21 knockdown induction by antisense injection in 60-day old male mice, we investigated glucose and insulin tolerance test, glucose-induced insulin secretion, glucose-induced intracellular calcium dynamics, and gene expression. Our results showed that ARHGAP21 acts negatively in the GSIS of adult islet. This effect seems to be due to the modulation of important points of insulin secretion process, such as the energy metabolism (PGC1α), Ca2+ signalization (SYTVII), granule-extrusion (SNAP25), and cell-cell interaction (CX36). Therefore, based on these finds, ARHGAP21 may be an important target in Diabetes Mellitus (DM) treatment.
Subject(s)
GTPase-Activating Proteins/physiology , Gene Expression Regulation/drug effects , Glucose/pharmacology , Hyperinsulinism/prevention & control , Insulin Secretion , Insulin-Secreting Cells/drug effects , Animals , Homeostasis , Hyperinsulinism/metabolism , Hyperinsulinism/pathology , Insulin-Secreting Cells/metabolism , Insulin-Secreting Cells/pathology , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Sweetening Agents/pharmacologyABSTRACT
Cholesterol is one of the triggers of oxidative stress in the pancreatic-ß cell, generating high levels of reactive oxygen species, which leads to impairment of insulin synthesis and secretion. Bioactive compounds, such as citrus flavanones, which possess anti-inflammatory and antioxidant activities, could reduce oxidative stress in ß-cells and improve their function. We describe for the first time the protective effects of the phase-II flavanone metabolites [naringenin 7-O-glucuronide, hesperetin 3'-O-glucuronide, and hesperetin 7-O-glucuronide], and two flavanones-catabolites derived from gut microbiota metabolism [hippuric acid and 3-(4-hydroxyphenyl)propionic acid], on pancreatic ß-cell line MIN6 under oxidative stress, at physiologically relevant concentration. Cholesterol reduced cell viability in a dose and time-dependent manner, with an improvement in the presence of the metabolites. Moreover, flavanone metabolites attenuated oxidative stress by reducing levels of lipid peroxides, superoxide anions, and hydrogen peroxide. In response to the reduction of reactive oxygen species, a decrease in superoxide dismutase and glutathione peroxidase activities was observed; these activities were elevated by cholesterol. Moreover, all the flavanone metabolites improved mitochondrial function and insulin secretion, and reduced apoptosis. Flavanone metabolites were found uptake by ß-cells, and therefore could be responsible for the observed protective effects. These results demonstrated that circulating phase-II hesperetin and naringenin metabolites, and also phenolics derived from gut microbiota, protect pancreatic-ß cells against oxidative stress, leading to an improvement in ß-cell function and could be the bioactive molecules derived from the citrus consumption.
Subject(s)
Cholesterol/pharmacology , Citrus/chemistry , Flavanones/pharmacology , Insulin-Secreting Cells/drug effects , Insulin-Secreting Cells/metabolism , Oxidative Stress/drug effects , Animals , Antioxidants/metabolism , Apoptosis/drug effects , Cell Line , Cell Survival/drug effects , Flavanones/metabolism , Insulin/metabolism , Mice , Mitochondria/drug effects , Mitochondria/metabolism , Protective Agents/pharmacologyABSTRACT
We evaluated whether protein restriction during pregnancy alters the morphometry of pancreatic islets, the intra-islet glucagon-like peptide-1 (GLP-1) production, and the anti-apoptotic signalling pathway modulated by GLP-1. Control non-pregnant (CNP) and control pregnant (CP) rats were fed a 17% protein diet, and low-protein non-pregnant (LPNP) and low-protein pregnant (LPP) groups were fed a 6% protein diet. The masses of islets and ß-cells were similar in the LPNP group and the CNP group but were higher in the CP group than in the CNP group and were equal in the LPP group and the LPNP group. Both variables were lower in the LPP group than in the CP group. Prohormone convertase 2 and GLP-1 fluorescence in α-cells was lower in the low-protein groups than in the control groups. The least PC2/glucagon colocalization was observed in the LPP group, and the most was observed in the CP group. There was less prohormone convertase 1/3/glucagon colocalization in the LPP group than in the CP group. GLP-1/glucagon colocalization was similar in the LPP, CP and CNP groups, which showed less GLP-1/glucagon colocalization than the LPNP group. The mRNA Pka, Creb and Pdx-1 contents were higher in islets from pregnant rats than in islets from non-pregnant rats. Protein restriction during pregnancy impaired the mass of ß-cells and the intra-islet GLP-1 production but did not interfere with the transcription of genes of the anti-apoptotic signalling pathway modulated by GLP-1.
Subject(s)
Diet, Protein-Restricted/adverse effects , Glucagon-Like Peptide 1/metabolism , Insulin-Secreting Cells/metabolism , Islets of Langerhans/metabolism , Animals , Down-Regulation , Female , Gene Expression Regulation/drug effects , Gene Regulatory Networks/drug effects , Glucagon/metabolism , Insulin-Secreting Cells/drug effects , Islets of Langerhans/drug effects , Pregnancy , Proprotein Convertase 2/metabolism , RatsABSTRACT
AIMS: The aim of the present study was to clarify if in utero exposure to DEX would affect the development of different types of pancreatic endocrine cells during postnatal life. MAIN METHODS: We investigated morphological and transcriptional features of both pancreatic ß- and α-cell populations within the pancreatic islets during the early postnatal life of rats born to mothers treated with DEX (0.1 mg/kg) from day 14 to 19 of pregnancy. Untreated pregnant Wistar rats of the same age (12-week-old) were used as control (CTL). Pups were euthanized on the 1st, 3rd and 21st (PND1, PND3 and PND21, respectively) days of life, regardless of sex. Serum insulin and glucagon levels were also evaluated. KEY FINDINGS: Rats born to DEX-treated mothers exhibited increased pancreatic α-cell mass, circulating glucagon levels and Gcg, Pax6, MafB and Nkx2.2 expression. Rats born to DEX-treated mothers also presented a rise in serum insulin levels on the PND3 that was paralleled by reduced ß-cell mass. Such increase in serum insulin levels, instead, was associated with increased expression of genes associated to insulin secretion such as Gck and Slc2a2. SIGNIFICANCE: Altogether, the present data reveals yet unknown changes in endocrine pancreas during early postnatal life of rats exposed to DEX in utero. Such data may contribute to the understanding of the metabolic features of rats born to DEX-treated mothers.
Subject(s)
Dexamethasone/toxicity , Glucagon-Secreting Cells/drug effects , Glucocorticoids/toxicity , Insulin-Secreting Cells/drug effects , Animals , Dexamethasone/administration & dosage , Female , Gene Expression Regulation , Glucagon/blood , Glucagon-Secreting Cells/cytology , Glucocorticoids/administration & dosage , Homeobox Protein Nkx-2.2 , Insulin/blood , Insulin Secretion/physiology , Insulin-Secreting Cells/cytology , Male , Pregnancy , Prenatal Exposure Delayed Effects/physiopathology , Rats , Rats, WistarABSTRACT
BACKGROUND: Free fatty acid receptor 1 (FFAR1) is G-protein coupled receptor predominantly expressed in pancreatic ß-cells that is activated by a variety of free fatty acids (FFAs). Once activated, it promotes glucose-stimulated insulin secretion (GSIS). However, increased levels of FFAs lead to lipotoxicity, inducing loss of ß-cell function. FFAR1 plays a key role in the development of type 2 diabetes (T2D), and previous studies have indicated the importance of developing anti-diabetic therapies against FFAR1, although its role in the regulation of ß-cell function remains unclear. The present study investigated the role of FFAR1 under lipotoxic conditions using palmitic acid (PA). The rat insulinoma 1 clone 832/13 (INS-1 832/13) cell line was used as a model as it physiologically resembles native pancreatic ß-cells. Key players of the insulin signaling pathway, such as mTOR, Akt, IRS-1, and the insulin receptor (INSR1ß), were selected as candidates to be analyzed under lipotoxic conditions. RESULTS: We revealed that PA-induced lipotoxicity affected GSIS in INS-1 cells and negatively modulated the activity of both IRS-1 and Akt. Reduced phosphorylation of both IRS-1 S636/639 and Akt S473 was observed, in addition to decreased expression of both INSR1ß and FFAR1. Moreover, transient knockdown of FFAR1 led to a reduction in IRS-1 mRNA expression and an increase in INSR1ß mRNA. Finally, PA affected localization of FFAR1 from the cytoplasm to the perinucleus. CONCLUSIONS: In conclusion, our study suggests a novel regulatory involvement of FFAR1 in crosstalk with mTOR-Akt and IRS-1 signaling in ß-cells under lipotoxic conditions.
Subject(s)
Insulin-Secreting Cells/drug effects , Lipid Metabolism/drug effects , Palmitic Acid/toxicity , Proto-Oncogene Proteins c-akt/metabolism , Receptors, G-Protein-Coupled/metabolism , TOR Serine-Threonine Kinases/metabolism , Animals , Apoptosis , Cell Line , Insulin-Secreting Cells/metabolism , Rats , Signal TransductionABSTRACT
Metabolic overload by saturated fatty acids (SFA), which comprises ß-cell function, and impaired glucose-stimulated insulin secretion are frequently observed in patients suffering from obesity and type 2 diabetes mellitus. The increase of intracellular Ca2+ triggers insulin granule release, therefore several mechanisms regulate Ca2+ efflux within the ß-cells, among others, the plasma membrane Ca2+-ATPase (PMCA). In this work, we describe that lipotoxicity mediated mainly by the saturated palmitic acid (PA) (16C) is associated with loss of protein homeostasis (proteostasis) and potentially cell viability, a phenomenon that was induced to a lesser extent by stearic (18C), myristic (14C) and lauric (12C) acids. PA was localized on endoplasmic reticulum, activating arms of the unfolded protein response (UPR), as also promoted by lipopolysaccharides (LPS)-endotoxins. In particular, our findings demonstrate an alteration in PMCA1/4 expression caused by PA and LPS which trigger the UPR, affecting not only insulin release and contributing to ß-cell mass reduction, but also increasing reactive nitrogen species. Nonetheless, stearic acid (SA) did not show these effects. Remarkably, the proteolytic degradation of PMCA1/4 prompted by PA and LPS was avoided by the action of monounsaturated fatty acids such as oleic and palmitoleic acid. Oleic acid recovered cell viability after treatment with PA/LPS and, more interestingly, relieved endoplasmic reticulum (ER) stress. While palmitoleic acid improved the insulin release, this fatty acid seems to have more relevant effects upon the expression of regulatory pumps of intracellular Ca2+. Therefore, chain length and unsaturation of fatty acids are determinant cues in proteostasis of ß-cells and, consequently, on the regulation of calcium and insulin secretion.
Subject(s)
Diabetes Mellitus, Type 2/metabolism , Fatty Acids, Monounsaturated/pharmacology , Insulin-Secreting Cells/drug effects , Oleic Acid/pharmacology , Palmitic Acid/toxicity , Proteostasis/drug effects , Animals , Calcium/metabolism , Cell Line , Insulin Secretion/drug effects , Insulin-Secreting Cells/metabolism , Lipopolysaccharides/toxicity , Plasma Membrane Calcium-Transporting ATPases/metabolism , Rats , Unfolded Protein Response/drug effectsABSTRACT
Venom from mammals, amphibians, snakes, arachnids, sea anemones and insects provides diverse sources of peptides with different potential medical applications. Several of these peptides have already been converted into drugs and some are still in the clinical phase. Diabetes type 2 is one of the diseases with the highest mortality rate worldwide, requiring specific attention. Diverse drugs are available (e.g., Sulfonylureas) for effective treatment, but with several adverse secondary effects, most of them related to the low specificity of these compounds to the target. In this context, the search for specific and high-affinity compounds for the management of this metabolic disease is growing. Toxins isolated from animal venom have high specificity and affinity for different molecular targets, of which the most important are ion channels. This review will present an overview about the electrical activity of the ion channels present in pancreatic ß cells that are involved in the insulin secretion process, in addition to the diversity of peptides that can interact and modulate the electrical activity of pancreatic ß cells. The importance of prospecting bioactive peptides for therapeutic use is also reinforced.