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Comprehensive molecular and cellular phenotyping of human islets can enable deep mechanistic insights for diabetes research. We established the Human Islet Data Analysis and Sharing (HI-DAS) consortium to advance goals in accessibility, usability, and integration of data from human islets isolated from donors with and without diabetes at the Alberta Diabetes Institute (ADI) IsletCore. Here we introduce HumanIslets.com, an open resource for the research community. This platform, which presently includes data on 547 human islet donors, allows users to access linked datasets describing molecular profiles, islet function and donor phenotypes, and to perform various statistical and functional analyses at the donor, islet and single-cell levels. As an example of the analytic capacity of this resource we show a dissociation between cell culture effects on transcript and protein expression, and an approach to correct for exocrine contamination found in hand-picked islets. Finally, we provide an example workflow and visualization that highlights links between type 2 diabetes status, SERCA3b Ca2+-ATPase levels at the transcript and protein level, insulin secretion and islet cell phenotypes. HumanIslets.com provides a growing and adaptable set of resources and tools to support the metabolism and diabetes research community.
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Epidemiological studies consistently link environmental toxicant exposure with increased Type 2 diabetes risk. Our study investigated the diabetogenic effects of a widely used flame retardant, Dechlorane Plus (DP), on pancreatic ß-cells using rodent and human model systems. We first examined pancreas tissues from male mice exposed daily to oral gavage of either vehicle (corn oil) or DP (10, 100, or 1000 µg/kg per day) and fed chow or high fat diet for 28-days in vivo. DP exposure did not affect islet size or endocrine cell composition in either diet group. Next, we assessed the effect of 48-hour exposure to vehicle (DMSO) or DP (1, 10, or 100 nM) in vitro using immortalized rat ß-cells (INS-1 832/3), primary mouse and human islets, and human stem-cell derived islet-like cells (SC-islets). In INS-1 832/3 cells, DP did not impact glucose-stimulated insulin secretion (GSIS) but significantly decreased intracellular insulin content. DP had no effect on GSIS in mouse islets or SC-islets but had variable effects on GSIS in human islets depending on the donor. DP alone did not affect insulin content in mouse islets, human islets, or SC-islets, but mouse islets co-exposed to DP and glucolipotoxic (GLT) stress conditions (28.7 mM glucose + 0.5 mM palmitate) had reduced insulin content compared to control conditions. Co-exposure of mouse islets to DP + GLT amplified the upregulation of Slc30a8 compared to GLT alone. Our study highlights the importance and challenges of using different in vitro models for studying chemical toxicity.
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Hidrocarbonetos Clorados , Células Secretoras de Insulina , Compostos Policíclicos , Animais , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Humanos , Camundongos , Masculino , Compostos Policíclicos/farmacologia , Hidrocarbonetos Clorados/toxicidade , Ratos , Insulina/metabolismo , Retardadores de Chama/toxicidade , Secreção de Insulina/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Células CultivadasRESUMO
OBJECTIVE: Chronic exposure to persistent organic pollutants (POPs) is associated with increased incidence of type 2 diabetes, hyperglycemia, and poor insulin secretion in humans. Dioxins and dioxin-like compounds are a broad class of POPs that exert cellular toxicity through activation of the aryl hydrocarbon receptor (AhR). We previously showed that a single high-dose injection of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, aka dioxin; 20 µg/kg) in vivo reduced fasted and glucose-stimulated plasma insulin levels for up to 6 weeks in male and female mice. TCDD-exposed male mice were also modestly hypoglycemic and had increased insulin sensitivity, whereas TCDD-exposed females were transiently glucose intolerant. Whether these effects are driven by AhR activation in ß-cells requires investigation. METHODS: We exposed female and male ß-cell specific Ahr knockout (ßAhrKO) mice and littermate Ins1-Cre genotype controls (ßAhrWT) to a single high dose of 20 µg/kg TCDD and tracked the mice for 6 weeks. RESULTS: Under baseline conditions, deleting AhR from ß-cells caused hypoglycemia in female mice, increased insulin secretion ex vivo in female mouse islets, and promoted modest weight gain in male mice. Importantly, high-dose TCDD exposure impaired glucose homeostasis and ß-cell function in ßAhrWT mice, but these phenotypes were largely abolished in TCDD-exposed ßAhrKO mice. CONCLUSION: Our study demonstrates that AhR signaling in ß-cells is important for regulating baseline ß-cell function in female mice and energy homeostasis in male mice. We also show that ß-cell AhR signaling largely mediates the effects of TCDD on glucose homeostasis in both sexes, suggesting that the effects of TCDD on ß-cell function and health are driving metabolic phenotypes in peripheral tissues.
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Diabetes Mellitus Tipo 2 , Dioxinas , Dibenzodioxinas Policloradas , Animais , Feminino , Humanos , Masculino , Camundongos , Diabetes Mellitus Tipo 2/induzido quimicamente , Glucose , Homeostase , Dibenzodioxinas Policloradas/toxicidade , Receptores de Hidrocarboneto Arílico/genética , Receptores de Hidrocarboneto Arílico/metabolismoRESUMO
BACKGROUND: Macroencapsulated pancreatic endoderm cells (PECs) can reverse diabetes in rodents and preclinical studies revealed that thyroid hormones in vitro and in vivo bias PECs to differentiate into insulin-producing cells. In an ongoing clinical trial, PECs implanted in macroencapsulation devices into patients with type 1 diabetes were safe but yielded heterogeneous outcomes. Though most patients developed meal responsive C-peptide, levels were heterogeneous and explanted grafts had variable numbers of surviving cells with variable distribution of endocrine cells. METHODS: We measured circulating triiodothyronine and thyroxine levels in all patients treated at 1 of the 7 sites of the ongoing clinical trial and determined if thyroid hormone levels were associated with the C-peptide or glucagon levels and cell fate of implanted PECs. RESULTS: Both triiodothyronine and thyroxine levels were significantly associated with the proportion of cells that adopted an insulin-producing fate with a mature phenotype. Thyroid hormone levels were inversely correlated to circulating glucagon levels after implantation, suggesting that thyroid hormones lead PECs to favor an insulin-producing fate over a glucagon-producing fate. In mice, hyperthyroidism led to more rapid maturation of PECs into insulin-producing cells similar in phenotype to PECs in euthyroid mice. CONCLUSION: These data highlight the relevance of thyroid hormones in the context of PEC therapy in patients with type 1 diabetes and suggest that a thyroid hormone adjuvant therapy may optimize cell outcomes in some PEC recipients.
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Diabetes Mellitus Tipo 1 , Humanos , Camundongos , Animais , Diabetes Mellitus Tipo 1/metabolismo , Peptídeo C/metabolismo , Tiroxina/metabolismo , Tri-Iodotironina/metabolismo , Endoderma/metabolismo , Endoderma/transplante , Glucagon/metabolismoRESUMO
Per- and polyfluoroalkyl substances (PFAS) are persistent environmental contaminants that induce immunotoxicity in experimental studies; however, epidemiological evidence-particularly during pregnancy-is scarce. We quantified associations between first trimester plasma perfluorooctanoic acid (PFOA), perfluorooctane sulfonate (PFOS), and perfluorohexane sulfonate (PFHxS) concentrations and third trimester concentrations of inflammatory biomarkers and determined if these associations were modified by fetal sex. Methods: We analyzed data from 1411 participants, recruited between 2008 and 2011, in the Maternal-Infant Research on Environmental Chemicals study. Our primary outcome was a composite inflammatory index derived by summing the z-scores of eight proinflammatory biomarkers. Using multivariable linear regression models, we quantified associations between each PFAS and the inflammatory index and individual biomarkers. We quantified the effects of the PFAS mixture using weighted quantile sum regression, and evaluated effect modification using product terms and sex-stratified models. Results: Each doubling of PFOA and PFHxS was associated with a 0.38 (95% CI, 0.09, 0.67) and 0.21 (95% CI, 0.01, 0.41) SD increase in the proinflammatory index, respectively. A one-quartile increase in the PFAS mixture was associated with a 0.40 (95% CI, 0.09, 0.71) SD increase in the proinflammatory index. In individual models, we observed positive associations between PFAS and concentrations of monocyte chemoattractant protein-1, macrophage inflammatory protein-1ß, and matrix metalloproteinases-9; however, the magnitude and precision varied according to the specific PFAS. Sex-specific findings were identified in few PFAS-biomarker associations. Conclusions: PFOA, PFOS, and PFHxS, individually and as a mixture, were positively associated with proinflammatory biomarkers during pregnancy.
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Epidemiological studies report associations between early-life exposure to persistent organic pollutants (POPs) and impaired metabolic homeostasis in adulthood. We investigated the impact of early-life exposure to low-dose 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or 'dioxin') on the establishment of ß-cell area during the perinatal period, as well as ß-cell health and glucose homeostasis later in life. Adult female mice were injected with either corn oil (CO; vehicle control) or TCDD (20 ng/kg/day) 2×/week throughout mating, pregnancy, and lactation; offspring were thus indirectly exposed to maternal TCDD in utero and during lactation, with pollutant exposure ending at weaning. All offspring were maintained on chow diet from weaning until 12-17 weeks of age, after which a subset of CO- and TCDD-exposed offspring were transferred to a 45% high fat diet (HFD) as a metabolic stressor for an additional 10 weeks. TCDD significantly upregulated cytochrome P450 1a1 (Cyp1a1) gene expression in offspring pancreas at birth and weaning, indicating that maternal TCDD directly reaches the developing pancreas. TCDD-exposed pups were transiently hypoglycemic at birth and females were born with reduced % ß-cell area, which persisted into adulthood. Early-life TCDD exposure had no persistent long-term effects on glucose homeostasis in chow-fed offspring, but when transferred to HFD, TCDD-exposed female offspring had a delayed onset of HFD-induced hyperglycemia, more pronounced HFD-induced hyperinsulinemia, and increase % PCNA+ ß-cells compared with CO-exposed female offspring. This study demonstrates that early-life exposure of mice to TCDD has modest effects on metabolic health in chow-fed offspring but alters metabolic adaptability to HFD feeding in females.
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Dioxinas , Dibenzodioxinas Policloradas , Efeitos Tardios da Exposição Pré-Natal , Gravidez , Masculino , Humanos , Camundongos , Animais , Feminino , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Dibenzodioxinas Policloradas/toxicidade , Reprodução , Glucose/farmacologiaRESUMO
Persistent organic pollutants (POPs) are a diverse family of contaminants that show widespread global dispersion and bioaccumulation. Humans are continuously exposed to POPs through diet, air particles, and household and commercial products; POPs are consistently detected in human tissues, including the pancreas. Epidemiological studies show a modest but consistent correlation between exposure to POPs and increased diabetes risk. The goal of this review is to provide an overview of epidemiological evidence and an in-depth evaluation of the in vivo and in vitro evidence that POPs cause ß-cell toxicity. We review evidence for six classes of POPs: dioxins, polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), organophosphate pesticides (OPPs), flame retardants, and per- and polyfluoroalkyl substances (PFAS). The available data provide convincing evidence implicating POPs as a contributing factor driving impaired glucose homeostasis, ß-cell dysfunction, and altered metabolic and oxidative stress pathways in islets. These findings support epidemiological data showing that POPs increase diabetes risk and emphasize the need to consider the endocrine pancreas in toxicity assessments. Our review also highlights significant gaps in the literature assessing islet-specific endpoints after both in vivo and in vitro POP exposure. In addition, most rodent studies do not consider the impact of biological sex or secondary metabolic stressors in mediating the effects of POPs on glucose homeostasis and ß-cell function. We discuss key gaps and limitations that should be assessed in future studies.
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Poluentes Ambientais , Hidrocarbonetos Clorados , Praguicidas , Bifenilos Policlorados , Poluentes Ambientais/toxicidade , Glucose , Hidrocarbonetos Clorados/análise , Hidrocarbonetos Clorados/toxicidade , Poluentes Orgânicos Persistentes , Praguicidas/análise , Praguicidas/toxicidade , Bifenilos Policlorados/análise , Bifenilos Policlorados/toxicidadeRESUMO
miRNAs have crucial functions in many biological processes and are candidate biomarkers of disease. Here, we show that miR-216a is a conserved, pancreas-specific miRNA with important roles in pancreatic islet and acinar cells. Deletion of miR-216a in mice leads to a reduction in islet size, ß-cell mass, and insulin levels. Single-cell RNA sequencing reveals a subpopulation of ß-cells with upregulated acinar cell markers under a high-fat diet. miR-216a is induced by TGF-ß signaling, and inhibition of miR-216a increases apoptosis and decreases cell proliferation in pancreatic cells. Deletion of miR-216a in the pancreatic cancer-prone mouse line KrasG12D;Ptf1aCreER reduces the propensity of pancreatic cancer precursor lesions. Notably, circulating miR-216a levels are elevated in both mice and humans with pancreatic cancer. Collectively, our study gives insights into how ß-cell mass and acinar cell growth are modulated by a pancreas-specific miRNA and also suggests miR-216a as a potential biomarker for diagnosis of pancreatic diseases.
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Progressão da Doença , Deleção de Genes , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , MicroRNAs/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Animais , Apoptose , Sequência de Bases , Linhagem Celular Tumoral , Movimento Celular , Dieta Hiperlipídica , Humanos , Secreção de Insulina , Camundongos Endogâmicos C57BL , Camundongos Knockout , MicroRNAs/metabolismo , Especificidade de Órgãos , RatosRESUMO
CONTEXT: Human studies consistently show an association between exposure to persistent organic pollutants, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, aka "dioxin"), and increased diabetes risk. We previously showed that a single high-dose TCDD exposure (20 µg/kg) decreased plasma insulin levels in male and female mice in vivo, but effects on glucose homeostasis were sex-dependent. OBJECTIVE: The current study assessed whether prolonged exposure to a physiologically relevant low-dose of TCDD impacts glucose homeostasis and/or the islet phenotype in a sex-dependent manner in chow-fed or high-fat diet (HFD)-fed mice. METHODS: Male and female mice were exposed to 20 ng/kg/d TCDD 2×/week for 12 weeks and simultaneously fed standard chow or a 45% HFD. Glucose homeostasis was assessed by glucose and insulin tolerance tests, and glucose-induced plasma insulin levels were measured in vivo. Histological analysis was performed on pancreas from male and female mice, and islets were isolated from females for TempO-Seq transcriptomic analysis. RESULTS: Low-dose TCDD exposure did not lead to adverse metabolic consequences in chow-fed male or female mice, or in HFD-fed males. However, TCDD accelerated the onset of HFD-induced hyperglycemia and impaired glucose-induced plasma insulin levels in females. TCDD caused a modest increase in islet area in males but reduced the percent beta cell area within islets in females. TempO-Seq analysis suggested abnormal changes to endocrine and metabolic pathways in female TCDDHFD islets. CONCLUSION: Our data suggest that prolonged low-dose TCDD exposure has minimal effects on glucose homeostasis and islet morphology in chow-fed male and female mice but promotes maladaptive metabolic responses in HFD-fed females.
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Adaptação Fisiológica/efeitos dos fármacos , Dieta Hiperlipídica/efeitos adversos , Dioxinas/farmacologia , Animais , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Doença Crônica , Relação Dose-Resposta a Droga , Metabolismo Energético/efeitos dos fármacos , Exposição Ambiental/efeitos adversos , Poluentes Ambientais/farmacologia , Feminino , Insulina/metabolismo , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Dibenzodioxinas Policloradas/farmacologia , Caracteres Sexuais , Fatores de TempoRESUMO
The potential for persistent organic pollutants (POPs), including dioxins and dioxin-like polychlorinated biphenyls (DL-PCBs), to increase the risk of incident diabetes in adults has been extensively studied. However, there is substantial variability in the reported associations both between and within studies. Emerging data from rodent studies suggest that dioxin disrupts glucose homeostasis in a sex-specific manner. Thus, we performed a review and meta-analysis of relevant epidemiological studies to investigate sex differences in associations between dioxin or DL-PCB exposure and type 2 diabetes incidence. Articles that met our selection criteria (n = 81) were organized into the following subcategories: data stratified by sex (n = 13), unstratified data (n = 45), and data from only 1 sex (n = 13 male, n = 10 female). We also considered whether exposure occurred either abruptly at high concentrations through a contamination event ("disaster exposure") or chronically at low concentrations ("non-disaster exposure"). There were 8 studies that compared associations between dioxin/DL-PCB exposure and diabetes risk in males versus females within the same population. When all sex-stratified or single-sex studies were considered in the meta-analysis (n = 18), the summary relative risk (RR) for incident diabetes among those exposed relative to reference populations was 1.78 (95% CI = 1.37-2.31) and 1.95 (95% CI = 1.56-2.43) for female and males, respectively. However, when we restricted the meta-analysis to disaster-exposed populations, the RR was higher in females than males (2.86 versus 1.59, respectively). In contrast, in non-disaster exposed populations the RR for females was lower than males (1.40 and 2.02, respectively). Our meta-analysis suggests that there are sex differences in the associations between dioxin/DL-PCBs exposure and incident diabetes, and that the mode of exposure modifies these differences.
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OBJECTIVE: Exposure to persistent organic pollutants is consistently associated with increased diabetes risk in humans. We investigated the short- and long-term impact of transient low-dose dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD) exposure during pregnancy and lactation on glucose homeostasis and beta cell function in female mice, including their response to a metabolic stressor later in life. METHODS: Female mice were injected with either corn oil (CO; vehicle control) or 20 ng/kg/d TCDD 2x/week throughout mating, pregnancy and lactation, and then tracked for 6-10 weeks after chemical exposure stopped. A subset of CO- and TCDD-exposed dams was then transferred to a 45% high-fat diet (HFD) or remained on a standard chow diet for an additional 11 weeks to assess the long-term effects of TCDD on adaptability to a metabolic stressor. To summarize, female mice were transiently exposed to TCDD and then subsequently tracked beyond when TCDD had been excreted to identify lasting metabolic effects of TCDD exposure. RESULTS: TCDD-exposed dams were hypoglycemic at birth but otherwise had normal glucose homeostasis during and post-TCDD exposure. However, TCDD-exposed dams on a chow diet were modestly heavier than controls starting 5 weeks after the last TCDD injection, and their weight gain accelerated after transitioning to a HFD. TCDD-exposed dams also had an accelerated onset of hyperglycemia, impaired glucose-induced plasma insulin levels, reduced islet size, increased MAFA-ve beta cells, and increased proinsulin accumulation following HFD feeding compared to controls. Overall, our study demonstrates that low-dose TCDD exposure during pregnancy has minimal effects on metabolism during the period of active exposure, but has detrimental long-term effects on metabolic adaptability to HFD feeding. CONCLUSIONS: Our study suggests that transient low-dose TCDD exposure in female mice impairs metabolic adaptability to HFD feeding, demonstrating that dioxin exposure may be a contributing factor to obesity and diabetes pathogenesis in females.
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Dioxinas/efeitos adversos , Obesidade/metabolismo , Animais , Diabetes Mellitus , Dieta Hiperlipídica , Dioxinas/metabolismo , Dioxinas/farmacologia , Suscetibilidade a Doenças/induzido quimicamente , Feminino , Glucose/metabolismo , Insulina/metabolismo , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/fisiologia , Lactação/efeitos dos fármacos , Lactação/metabolismo , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , GravidezRESUMO
Epidemiological studies have consistently shown an association between exposure to environmental pollutants and diabetes risk in humans. We have previously shown that direct exposure of mouse and human islets (endocrine pancreas) to the highly persistent pollutant TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) causes reduced insulin secretion ex vivo. Furthermore, a single high-dose of TCDD (200 µg/kg) suppressed both fasting and glucose-induced plasma insulin levels and promoted beta-cell apoptosis after 7 days in male mice. The current study investigated the longer-term effects of a single high-dose TCDD injection (20 µg/kg) on glucose metabolism and beta cell function in male and female C57Bl/6 mice. TCDD-exposed males displayed modest fasting hypoglycemia for ~4 weeks post-injection, reduced fasting insulin levels for up to 6 weeks, increased insulin sensitivity, decreased beta cell area, and increased delta cell area. TCDD-exposed females also had long-term suppressed basal plasma insulin levels, and abnormal insulin secretion for up to 6 weeks. Unlike males, TCDD did not impact insulin sensitivity or islet composition in females, but did cause transient glucose intolerance 4 weeks post-exposure. Our results show that a single exposure to dioxin can suppress basal insulin levels long-term in both sexes, but effects on glucose homeostasis are sex-dependent.
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Diabetes Mellitus/epidemiologia , Poluentes Ambientais/efeitos adversos , Células Secretoras de Insulina/fisiologia , Dibenzodioxinas Policloradas/efeitos adversos , Fatores Sexuais , Animais , Diabetes Mellitus/etiologia , Modelos Animais de Doenças , Feminino , Homeostase , Humanos , Hipoglicemia , Insulina/metabolismo , Secreção de Insulina , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RiscoRESUMO
Diabetes prevalence is increasing worldwide, and epidemiological studies report an association between diabetes incidence and environmental pollutant exposure. There are >84,000 chemicals in commerce, many of which are released into the environment without a clear understanding of potential adverse health consequences. While in vivo rodent studies remain an important tool for testing chemical toxicity systemically, we urgently need high-throughput screening platforms in biologically relevant models to efficiently prioritize chemicals for in depth toxicity analysis. Given the increasing global burden of obesity and diabetes, identifying chemicals that disrupt metabolism should be a high priority. Pancreatic endocrine cells are key regulators of systemic metabolism, yet often overlooked as a target tissue in toxicology studies. Immortalized ß-cell lines and primary human, porcine, and rodent islets are widely used for studying the endocrine pancreas in vitro, but each have important limitations in terms of scalability, lifespan, and/or biological relevance. Human pluripotent stem cell (hPSC) culture is a powerful tool for in vitro toxicity testing that addresses many of the limitations with other ß-cell models. Current in vitro differentiation protocols can efficiently generate glucose-responsive insulin-secreting ß-like cells that are not fully mature, but still valuable for high-throughput toxicity screening in vitro. Furthermore, hPSCs can be applied as a model of developing pancreatic endocrine cells to screen for chemicals that influence endocrine cell formation during critical windows of differentiation. Given their versatility, we recommend using hPSCs to identify potential ß-cell toxins, which can then be prioritized as chemicals of concern for metabolic disruption.
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Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/patologia , Células Endócrinas/patologia , Células Secretoras de Insulina/patologia , Pâncreas/patologia , Preparações Farmacêuticas/administração & dosagem , Células-Tronco Pluripotentes/patologia , Testes de Toxicidade/métodos , Animais , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/etiologia , Células Endócrinas/efeitos dos fármacos , Humanos , Células Secretoras de Insulina/efeitos dos fármacos , Pâncreas/efeitos dos fármacos , Células-Tronco Pluripotentes/efeitos dos fármacosRESUMO
AIMS/HYPOTHESIS: Exposure to environmental pollution has been consistently linked to diabetes incidence in humans, but the potential causative mechanisms remain unclear. Given the critical role of regulated insulin secretion in maintaining glucose homeostasis, environmental chemicals that reach the endocrine pancreas and cause beta cell injury are of particular concern. We propose that cytochrome P450 (CYP) enzymes, which are involved in metabolising xenobiotics, could serve as a useful biomarker for direct exposure of islets to pollutants. Moreover, functional CYP enzymes in islets could also impact beta cell physiology. The aim of this study was to determine whether CYP1A enzymes are activated in islets following direct or systemic exposure to environmental pollutants. METHODS: Immortalised liver (HepG2) and rodent pancreatic endocrine cell lines (MIN6, ßTC-6, INS1, α-TC1, α-TC3), as well as human islets, were treated in vitro with known CYP1A inducers 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and 3-methylcholanthrene (3-MC). In addition, mice were injected with either a single high dose of TCDD or multiple low doses of TCDD in vivo, and islets were isolated 1, 7 or 14 days later. RESULTS: CYP1A enzymes were not activated in any of the immortalised beta or alpha cell lines tested. However, both 3-MC and TCDD potently induced CYP1A1 gene expression and modestly increased CYP1A1 enzyme activity in human islets after 48 h. The induction of CYP1A1 in human islets by TCDD was prevented by cotreatment with a cytokine mixture. After a systemic single high-dose TCDD injection, CYP1A1 enzyme activity was induced in mouse islets ~2-fold, ~40-fold and ~80-fold compared with controls after 1, 7 and 14 days, respectively, in vivo. Multiple low-dose TCDD exposure in vivo also caused significant upregulation of Cyp1a1 in mouse islets. Direct TCDD exposure to human and mouse islets in vitro resulted in suppressed glucose-induced insulin secretion. A single high-dose TCDD injection resulted in lower plasma insulin levels, as well as a pronounced increase in beta cell death. CONCLUSIONS/INTERPRETATION: Transient exposure to TCDD results in long-term upregulation of CYP1A1 enzyme activity in islets. This provides evidence for direct exposure of islets to lipophilic pollutants in vivo and may have implications for islet physiology.
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Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP1A2/metabolismo , Poluentes Ambientais/toxicidade , Animais , Glicemia/efeitos dos fármacos , Linhagem Celular , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A2/genética , Células Hep G2 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dibenzodioxinas Policloradas/toxicidade , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Pancreatic progenitors derived from human embryonic stem cells (hESCs) are now in clinical trials for insulin replacement in patients with type 1 diabetes. Animal studies indicate that pancreatic progenitor cells can mature into a mixed population of endocrine cells, including glucose-responsive ß cells several months after implantion. However, it remains unclear how conditions in the recipient may influence the maturation and ultimately the function of these hESC-derived cells. Here, we investigated the effects of (1) pregnancy on the maturation of human stage 4 (S4) pancreatic progenitor cells and (2) the impact of host sex on both S4 cells and more mature stage 7 (S7) pancreatic endocrine cells implanted under the kidney capsule of immunodeficient SCID-beige mice. Pregnancy led to increased proliferation of endogenous pancreatic ß cells, but did not appear to affect proliferation or maturation of S4 cells at midgestation. Interestingly, S4 and S7 cells both acquired glucose-stimulated C-peptide secretion in females before males. Moreover, S4 cells lowered fasting blood glucose levels in females sooner than in males, whereas the responses with S7 cells were similar. These data indicate that the host sex may impact the maturation of hESC-derived cells in vivo and that this effect can be minimized by more advanced differentiation of the cells before implantation.
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Peptídeo C/metabolismo , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Glucose/farmacologia , Células-Tronco Embrionárias Humanas/citologia , Células Secretoras de Insulina/citologia , Caracteres Sexuais , Animais , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Feminino , Células-Tronco Embrionárias Humanas/efeitos dos fármacos , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Masculino , CamundongosRESUMO
Pancreatic progenitors derived from human embryonic stem cells (hESCs) are a potential source of transplantable cells for treating diabetes and are currently being tested in clinical trials. Yet, how the milieu of pancreatic progenitor cells, including exposure to different factors after transplant, may influence their maturation remains unclear. Here, we examined the effect of thyroid dysregulation on the development of hESC-derived progenitor cells in vivo. Hypothyroidism was generated in SCID-beige mice using an iodine-deficient diet containing 0.15% propyl-2-thiouracil, and hyperthyroidism was generated by addition of L-thyroxine (T4) to drinking water. All mice received macroencapsulated hESC-derived progenitor cells, and thyroid dysfunction was maintained for the duration of the study ("chronic") or for 4 weeks posttransplant ("acute"). Acute hyperthyroidism did not affect graft function, but acute hypothyroidism transiently impaired human C-peptide secretion at 16 weeks posttransplant. Chronic hypothyroidism resulted in severely blunted basal human C-peptide secretion, impaired glucose-stimulated insulin secretion, and elevated plasma glucagon levels. Grafts from chronic hypothyroid mice contained fewer ß-cells, heterogenous MAFA expression, and increased glucagon(+) and ghrelin(+) cells compared to grafts from euthyroid mice. Taken together, these data suggest that long-term thyroid hormone deficiency may drive the differentiation of human pancreatic progenitor cells toward α- and ε-cell lineages at the expense of ß-cell formation.
Assuntos
Diferenciação Celular , Diabetes Mellitus Tipo 1/cirurgia , Modelos Animais de Doenças , Xenoenxertos/patologia , Células-Tronco Embrionárias Humanas/transplante , Hipotireoidismo/complicações , Células Secretoras de Insulina/transplante , Animais , Antitireóideos/intoxicação , Biomarcadores/sangue , Biomarcadores/metabolismo , Linhagem Celular , Células Imobilizadas/citologia , Células Imobilizadas/patologia , Células Imobilizadas/transplante , Diabetes Mellitus Tipo 1/complicações , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patologia , Xenoenxertos/citologia , Xenoenxertos/metabolismo , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/metabolismo , Células-Tronco Embrionárias Humanas/patologia , Humanos , Hipertireoidismo/induzido quimicamente , Hipertireoidismo/complicações , Hipotireoidismo/etiologia , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Iodo/deficiência , Masculino , Camundongos SCID , Propiltiouracila/intoxicação , Distribuição Aleatória , Tiroxina/intoxicação , Transplante Heterólogo , Transplante HeterotópicoRESUMO
Pancreatic ß cells are mostly post-mitotic, but it is unclear what locks them in this state. Perturbations including uncontrolled hyperglycemia can drive ß cells into more pliable states with reduced cellular insulin levels, increased ß cell proliferation, and hormone mis-expression, but it is unknown whether reduced insulin production itself plays a role. Here, we define the effects of â¼50% reduced insulin production in Ins1(-/-):Ins2(f/f):Pdx1Cre(ERT):mTmG mice prior to robust hyperglycemia. Transcriptome, proteome, and network analysis revealed alleviation of chronic endoplasmic reticulum (ER) stress, indicated by reduced Ddit3, Trib3, and Atf4 expression; reduced Xbp1 splicing; and reduced phospho-eIF2α. This state was associated with hyper-phosphorylation of Akt, which is negatively regulated by Trib3, and with cyclinD1 upregulation. Remarkably, ß cell proliferation was increased 2-fold after reduced insulin production independently of hyperglycemia. Eventually, recombined cells mis-expressed glucagon in the hyperglycemic state. We conclude that the normally high rate of insulin production suppresses ß cell proliferation in a cell-autonomous manner.
Assuntos
Proliferação de Células , Estresse do Retículo Endoplasmático , Células Secretoras de Insulina/fisiologia , Insulina/biossíntese , Animais , Células Cultivadas , Metaboloma , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Mapas de Interação de Proteínas , Proteoma/metabolismo , Transdução de Sinais , TranscriptomaRESUMO
Pluripotent human embryonic stem cells (hESCs) are a potential source of transplantable cells for treating patients with diabetes. To investigate the impact of the host recipient on hESC-derived pancreatic progenitor cell maturation, cells were transplanted into immunodeficient SCID-beige mice or nude rats. Following the transplant, basal human C-peptide levels were consistently higher in mice compared with rats, but only rats showed robust meal- and glucose-responsive human C-peptide secretion by 19-21 weeks. Grafts from rats contained a higher proportion of insulin:glucagon immunoreactivity, fewer exocrine cells, and improved expression of mature ß cell markers compared with mice. Moreover, ECM-related genes were enriched, the collagen network was denser, and blood vessels were more intricately integrated into the engrafted endocrine tissue in rats relative to mice. Overall, hESC-derived pancreatic progenitor cells matured faster in nude rats compared with SCID-beige mice, indicating that the host recipient can greatly influence the fate of immature pancreatic progenitor cells post-transplantation.
Assuntos
Células-Tronco Embrionárias/citologia , Células Secretoras de Insulina/citologia , Pâncreas/citologia , Transplante de Células-Tronco , Animais , Peptídeo C/análise , Diferenciação Celular , Linhagem Celular , Células Cultivadas , Feminino , Humanos , Masculino , Camundongos , Camundongos SCID , Pessoa de Meia-Idade , Ratos , Ratos Nus , Especificidade da EspécieRESUMO
Pluripotent stem cells are a scalable source of pancreatic cells for transplantation into patients with diabetes. Here, we describe how the field is gaining momentum toward a ß cell replacement therapy.
Assuntos
Diabetes Mellitus/terapia , Células Secretoras de Insulina/citologia , Células-Tronco Pluripotentes/citologia , Animais , Glicemia/metabolismo , Transplante de Células , Diabetes Mellitus/imunologia , Humanos , Sistema Imunitário , Transplante das Ilhotas Pancreáticas , CamundongosRESUMO
Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) hold great potential as the basis for cell-based therapies of degenerative diseases, including diabetes. Current insulin-based therapies for diabetes do not prevent hyperglycaemia or the associated long-term organ damage. While transplantation of pancreatic islets can achieve insulin independence and improved glycemic control, it is limited by donor tissue scarcity, challenges of purifying islets from the pancreas, and the need for immunosuppression to prevent rejection of transplants. Large-scale production of ß-cells from stem cells is a promising alternative. Recent years have seen considerable progress in the optimization of in vitro differentiation protocols to direct hESCs/iPSCs into mature insulin-secreting ß-cells and clinical trials are now under way to test the safety and efficiency of hESC-derived pancreatic progenitor cells in patients with type 1 diabetes. Here, we discuss key milestones leading up to these trials in addition to recent developments and challenges for hESC/iPSC-based diabetes therapies and disease modeling.