AIMS/OBJECTIVE:p16INK4A is a cyclin-dependent kinase inhibitor that plays important roles in pancreatic islet beta-cell maturation, function, and senescence. As staining pattern consistency and reproducibility have been challenging in the field, our objective was to validate a monoclonal antibody for the detection of p16INK4A in mouse and human fixed pancreas sections. METHODS:Four commercial monoclonal antibodies reported to react against mouse p16INK4A were tested by fluorescent immunohistochemistry on positive and negative control formalin-fixed paraffin-embedded pancreas sections from two different mouse strains. Lot-to-lot consistency was also evaluated. Fixed human donor pancreas sections were evaluated for reactivity with the monoclonal p16INK4A antibody that performed best on mouse tissues. RESULTS:One commercial monoclonal antibody robustly detected a nuclear antigen in both mouse and human pancreas sections, consistent with the previously reported p16INK4A staining pattern, and its role as a cyclin-dependent kinase inhibitor in beta cells. This antibody also showed lot-to-lot consistency in staining pattern. CONCLUSION:Our findings highlight the importance of antibody validation for p16INK4A detection in mouse and human pancreas sections and suggest caution in using untested commercially available antibodies to this protein for immunohistochemistry of the pancreas.
Accurate quantification of islet mass is critical for the preclinical evaluation and therapeutic application of stem cell-derived pancreatic islet organoids. Traditional methods, including Ricordi's islet equivalent (IEQ) approach and its equivalent circle diameter adaptations, often overestimate islet volume due to reliance on maximal diameters and discrete size bins. To address these limitations, we developed an automated image segmentation and three-dimensional modeling framework to quantify individual islet clusters from brightfield images. Clusters were fitted with ellipses and modeled as ellipsoids using rotation about either the minor or major axis, allowing IEQs to be calculated continuously relative to a reference 150 μm spherical islet. Major-axis (prolate) rotation provided the most conservative and physically plausible volume estimates, whereas minor-axis (oblate) rotation and diameter-based approaches systematically overestimated IEQs. Functional assessment with glucose-stimulated insulin secretion assays across multiple size categories demonstrated consistent insulin output for clusters below 250 μm, supporting the reproducibility of our 3D differentiation system. In vivo, streptozotocin-induced diabetic mice transplanted with islet doses based on major-axis modeling exhibited faster and more stable restoration of glycemia compared with groups receiving doses derived from overestimated approaches. These findings establish that major-axis ellipsoid modeling offers a mathematically consistent, conservative, and biologically relevant method for estimating IEQs, providing a practical framework to guide dosing in preclinical studies and supporting the translational development of stem cell-derived islet therapies.
Diabetes mellitus and end-stage renal disease (ESRD) are major causes of morbidity and mortality worldwide.Pancreatic islet transplantation offers potential insulin independence. Different sites and their suitability for islet transplantation have been investigated, including renal subcapsular and intraportal sites, among others. Various limitations and complications have been reported, such as the number of donor islet species, islet isolation, and site of transplantation. Combining islets and kidney transplantation in diabetic patients suffering from ESRD has several advantages.This review compares the experimental experience of the renal subcapsular and intraportal sites for islet allo- and xeno-transplantation. The subcapsular site appears preferable in rodent models. The intraportal site has been more successful in large animals. Combined islet-kidneys have been suggested as a treatment targeting both ESRD and type 1 diabetes. With our increasing ability to genetically-engineer pigs, and the availability of novel immunosuppressive agents, xenotransplantation is an emerging therapeutic option.
BACKGROUND:Activation of islet stellate cells (ISCs) contributes to islet fibrosis and diabetes progression through excessive extracellular matrix secretion and lipid loss. Annexin A1 (ANXA1) has been reported to modulate lipid metabolism in other tissues, but its role in ISCs remains unclear. METHODS:ISCs were isolated from 9-and 28-week-old db/m and db/db mice. Lipid content analysis, qRT‒PCR, and Western blotting were used to assess lipid metabolism-related molecules. ANXA1 expression was analyzed by immunohistochemistry and Western blotting. Recombinant ANXA1 was co-cultured with db/db ISCs to evaluate lipid synthesis and lipolysis. The interaction between ANXA1 and peroxisome proliferator-activated receptor alpha (PPARα) was examined by immunoprecipitation. RESULTS:Activation of ISCs markedly reduced intracellular triglycerides, with decreased Diacylglycerol Acyltransferase 1/2 (DGAT1/2) and increased adipose triglyceride lipase (ATGL) and hormone-sensitive triglyceride lipase (HSL) expression. ANXA1 was detected in islets, MIN6 cells, and their culture supernatants. Recombinant ANXA1 treatment lowered triglyceride levels and upregulated PPARα and its downstream genes, acyl-CoA oxidase 1 (ACOX1) and cytochrome P450 4 A (CYP4A); these effects were enhanced by a PPARα agonist but reversed by inhibition. Immunofluorescence and coimmunoprecipitation confirmed that PPARα acts as a key mediator of ANXA1-regulated triglyceride metabolism in ISCs. CONCLUSION:ANXA1 promotes ISCs activation by enhancing triglyceride catabolism through the PPARα signaling pathway, suggesting a novel therapeutic target for islet fibrosis.
OBJECTIVE:Mitochondria are central to energy production and cellular homeostasis. Beyond importing diverse RNAs, they also encode hundreds of their own non-coding RNAs, contributing to a complex and dynamic RNA landscape. Early-life nutritional insults, such as fetal and postnatal protein deficiency, can impair mitochondrial function and increase the long-term diabetes risk. However, the mitochondrial non-coding transcriptome of pancreatic islets, particularly its responsiveness to nutritional cues, remains largely unexplored. METHODS:We performed RNA sequencing to profile small non-coding RNAs in mitochondrial fractions of islet cells from offspring of rats exposed to low-protein (LP) or control diets during gestation and lactation and employed mRNA-miRNA network analysis to explore the potential regulatory roles of differentially expressed mitomiRs in LP-exposed pups. RESULTS:Protein deficiency during gestation and lactation led to a profound remodeling of the small non-coding RNA landscape in whole islets, with microRNAs and piRNAs showing the most pronounced changes. In mitochondrial fractions, LP exposure resulted in a striking shift in microRNA composition, with 33 mitomiRs detected in control islets versus 23 in LP-exposed rats, and only 5 shared between groups. Notably, ten mitomiRs were selectively depleted from the cytosol and enriched in mitochondria of LP-exposed islets. Amongst these, miR-10a-5p and miR-126a-5p, are predicted to target genes involved in mitochondrial metabolism and structural organization. CONCLUSION:Early-life protein restriction triggers a highly selective reorganization of the mitomiR landscape in pancreatic islets. The identified mitomiRs may serve as regulators of mitochondrial function and intracellular signaling, potentially influencing β-cell metabolic coupling and contributing to diabetes susceptibility.
AimsType 1 diabetes is caused by immune-mediated destruction of beta cells. Interestingly, individuals with long-standing type 1 diabetes have residual beta cells, suggesting regenerative mechanisms may help beta cell survival. Islet-resident macrophages have an important role in diabetes, and can adopt a tissue-regenerating phenotype that may support beta cells. However, the roles of macrophages in beta cell survival, function, and proliferation remain poorly defined. This study aimed to elucidate how macrophage subtypes influence beta cell survival, function, and proliferation.MethodsMouse and human islets were isolated from the pancreas and co-cultured in vitro with macrophages. To investigate whether macrophages enhance beta cell survival and function, apoptosis was measured using flow cytometry, and insulin secretion was assessed via glucose-stimulated insulin secretion assays. We also examined whether macrophages increased beta cell proliferation in the presence of harmine, a DYRK1A inhibitor. Finally, we evaluated the effect of islet co-culture on macrophage phenotype by flow cytometry and cytokine secretion analysis.ResultsWe found that regenerative, but not pro-inflammatory, macrophages enhanced beta cell survival and function through mechanisms that did not require direct cell contact. Direct contact between macrophages and islets promoted a macrophage regenerative phenotype characterized by increased CD206 expression and secretion of anti-inflammatory factors. Additionally, regenerative macrophages promoted beta cell proliferation in the presence of harmine.ConclusionsOur findings demonstrate that regenerative macrophages support pancreatic beta cell survival, function, and proliferation. Harnessing the regenerative properties of macrophages could offer a novel strategy to promote beta cell survival and function.
HNF4α is important for beta cells' ability to adequately secrete insulin in response to glucose concentration and endoplasmic reticulum (ER) homeostasis. In humans, HNF4α mutations are responsible for Diabetes mellitus subtype MODY1, which has an age-determining onset. Additionally, in other forms of DM, there is evidence that sex can influence beta cell dysfunction, with possible involvement of ER stress pathways. Thus, we assessed the influence of sex and age on beta-cell dysfunction induced by HNF4α absence. We used an animal model with specific beta cells KO of HNF4α, induced after birth (Ins. CRE HNF4αloxP/loxP). Glucose intolerance is observed after 10 d of KO induction, at 50 d of age, with KO males (MKO) displaying more severe glucose intolerance than KO females (FKO). The percentage of insulin-positive cells in KO mice islets is lower compared to Control at all ages evaluated, with MKO mice showing a more pronounced decline at later ages compared to FKO. Both KO groups exhibited reduced beta cell mass and increased α-cell mass, which was more pronounced in MKO. ER stress was induced in both KO groups; however, ER stress-mediated apoptosis was observed only in MKO. FKO mice show evidence of beta cell differentiated state loss. In summary, beta cell loss in HNF4α-KO is influenced by sex and age, involves induction of ER stress, and is more severe in males, where ER stress-induced beta cell death is observed. Partial protection observed in females seems to involve dedifferentiation of beta cells.
BACKGROUND:We previously showed that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD - a persistent organic pollutant) activates the aryl hydrocarbon receptor (AHR) in pancreatic islets. The AHR is known to crosstalk with hypoxia-inducible factor 1α (HIF1α) in other cell types but AHR-HIF1α crosstalk has not been previously examined in islet cells. Islet cell function is sensitive to hypoxia; we hypothesize that AHR activation by environmental pollutant(s) will interfere with the HIF1α pathway response in islets, which may be detrimental to islet cell function and survival during periods of hypoxia. METHODS:We assessed AHR-HIF1α crosstalk by treating human donor islets and stem cell-derived islets (SC-islets) with 10 nM TCDD ± 1% O2 and measuring gene expression of downstream targets of AHR (e.g. CYP1A1) and HIF1α (e.g. HMOX1). RESULTS:In SC-islets, co-treatment with TCDD + hypoxia consistently suppressed CYP1A1 induction compared with TCDD treatment alone. In human islets, TCDD + hypoxia co-treatment suppressed CYP1A1 induction, but only in 2 of 6 donors. Both SC-islets and human donor islets displayed hypoxia-mediated suppression of glucose-6-phosphate catalytic subunit 2 (G6PC2) expression. Glucose-stimulated insulin secretion (GSIS) in human donor islets was impaired by hypoxia exposure, but unaffected by TCDD exposure. CONCLUSION:Our study shows consistent AHR-HIF1α crosstalk in SC-islets and variable crosstalk in primary human islets, depending on the donor. In both cell models, hypoxia exposure interfered with activation of the AHR pathway by TCDD but there was no evidence that AHR activation interfered with the HIF1α pathway. In summary, our data show that co-exposure to an environmental pollutant and hypoxia results in molecular crosstalk in islets.
Pancreatic and duodenal homeobox protein (PDX)1 is a major transcription factor for the regulation of insulin, glucagon and somatostatin (SST) expression. PDX1 is phosphorylated by CK2 and inhibition of this kinase results in an increased insulin and decreased glucagon secretion. Therefore, we speculated in this study that CK2 also affects SST expression. To test this, we analyzed the effects of the two CK2 inhibitors CX-4945 and SGC as well as of PDX1 overexpression on SST expression and secretion in RIN14B cells by qRT-PCR, luciferase assays, Western blot and ELISA. SST expression and secretion were additionally assessed in isolated murine and human islets exposed to the CK2 inhibitors. Moreover, we determined the expression and secretion of the pancreatic endocrine hormones in CX-4945-treated mice. We found a suppressed SST expression in RIN14B cells due to a methylated SST promoter, which could be abolished by DNA demethylation. Under these conditions, we showed that CK2 inhibition increases SST gene expression and secretion. Additional experiments with overexpression of a CK2-phosphorylation mutant of PDX1 verified that SST expression is regulated by CK2. The exposure of isolated murine and human islets to CX-4945 or SGC as well as the treatment of mice with CX-4945 revealed that CK2 also regulates SST expression under physiological conditions. Taken together, these findings not only demonstrate that CK2 controls SST expression in pancreatic δ-cells but also emphasize the crucial role of this kinase in regulating the main hormones of the endocrine pancreas.
Chronic hyperglycemia impairs mitochondrial function of beta cells. Changes in mitochondrial function preceding a negative glucose effect have not been fully characterized, nor interactions with ketones. To compare effects on beta cell mitochondrial function by short and longer exposures to elevated glucose and interactions with ketones oxygen consumption rate (OCR) was measured in intact clonal beta cells by an OROBOROS and in rat islets by a Seahorse instrument. Proteins (subunits) of mitochondrial complexes (C) were measured by immunoblotting. ATP and ROS were measured in islets. In INS-1 832/13 cells, overnight exposure to 27 vs. 11 mm glucose increased OCR and uncoupled mitochondrial respiration. These effects vanished when prolonging the exposure time of elevated glucose. C1 was decreased after two days of culture with high glucose. Interactions with racemic 5 and 20 mm beta-hydroxybutyrate (BHB) were not detected. In islets, culture overnight at 27 vs.11 mm glucose enhanced basal OCR. No decrease in glucose-induced OCR was seen after prolonging 27 mm glucose for two days. Interactions with 5 mm BHB were not detected. Prolonged exposure to 27 mm glucose enhanced basal ECAR (extracellular acidification rate) and an ECAR response to acute elevation of glucose. C1 and 3 and 4 were decreased after two days of 27 vs. 11 mm glucose. ATP levels were decreased at this time-point and extracellular ROS increased. High glucose time-dependently affects mitochondrial function in clonal beta cells and islets. C1 was uniformly decreased. Interactions with BHB were not detected.
Pancreatic β-cells release insulin in response to fluctuations in plasma glucose, amino acids, and free fatty acids (FFA). Clonal cell lines and isolated islets serve as essential early models for studying the impact of nutrients and evaluating potential therapies to address β-cell dysfunction. Acute and chronic changes in FFA levels have been shown to have positive and negative effects on β-cell function both in vivo and in vitro. A key problem in comparing islet lipid studies from different laboratories is that a wide variety of methods are used to isolate, culture, and assess islet function. The current study compares bovine serum albumin (BSA) types and lipid preparation methods in clonal 832/13 cells and human islets. Changing the percentage and culture conditions when using FFA-free BSA can negatively affect β-cell function compared to regular BSA. Preparing palmitate with FFA-free BSA can rescue insulin secretion compared to treating cells alone with FFA-free BSA. Different methods of preparing palmitate can have unique effects on insulin secretion. Overall, interpreting the effects of lipids on β-cell function is complicated by a number of variables that need to be controlled for in islet experiments.
Statins are widely used to treat hyperlipidemia and atherosclerotic cardiovascular diseases (ACVD) by significantly lowering low-density lipoprotein cholesterol (LDL-C) levels. However, their use has been associated with an increased risk of type 2 diabetes (T2D), a paradox given their lipid-lowering benefits. This study investigates the role of LDL receptors (LDLR) and proprotein convertase subtilisin/kexin type 9 (PCSK9) in the diabetogenic effects of atorvastatin on pancreatic β-cells. Using the MIN6 pancreatic β-cell line, we assessed the impact of atorvastatin on LDL-C uptake, PCSK9 expression, glucose-stimulated insulin release (GSIR), and cell proliferation. Cellular cholesterol assays, EdU labeling, Dil-LDL uptake, western blot analysis, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and ELISA, were employed to measure relevant biomarkers and cellular responses. Male C57BL/6j mice were treated with atorvastatin to validate in vitro findings. Atorvastatin enhances LDL-C uptake by upregulating LDLR on the cell surface, without causing excess cholesterol accumulation. Additionally, atorvastatin suppresses PCSK9 expression, which is crucial for LDLR degradation. Interestingly, atorvastatin, combined with exogenous LDL-C, impairs glucose-stimulated insulin release (GSIR) but promotes cell proliferation, highlighting a potential mechanism for statin-associated diabetes. Oral administration of atorvastatin in mice reduced plasma PCSK9 and insulin levels, supporting the in vitro findings. These results indicate that while atorvastatin effectively lowers circulating cholesterol, it may adversely affect pancreatic β-cell function by modulating LDLR and LDL-C uptake, thereby increasing the risk of T2D. This study highlights the importance of further research to develop strategies mitigating the diabetogenic effects of statins while maintaining their cardiovascular benefits.
Background: Knockout (KO) ferrets with the cystic fibrosis transmembrane conductance regulator (CFTR) exhibit distinct phases of dysglycemia and pancreatic remodeling prior to cystic fibrosis-related diabetes (CFRD) development. Following normoglycemia during the first month of life (Phase l), hyperglycemia occurs during the subsequent 2 months (Phase Il) with decreased islet mass, followed by a period of near normoglycemia (Phase Ill) in which the islets regenerate. We aimed to characterize islet hormone expression patterns across these Phases. Methods: Immunofluorescence staining per islet area was performed to characterize islet hormone expression patterns in age matched CFTR KO and wild type (WT) ferrets, focusing on the first three phases. Results: In Phase I, insulin staining intensity was higher in CF (p < 0.01) than WT but decreased in Phase III (p < 0.0001). Glucagon was lower in CF during Phases I and increased in Phase III, while proinsulin decreased (p < 0.0001) Phases II and III. CF sections showed lower proinsulin-to-insulin ratio in Phase I (p < 0.01) and in Phase III (p < 0.05) compared to WT. Conversely, glucagon-to-insulin ratio was lower in CF in Phase I (p < 0.0001) but increased in Phase III (p < 0.0001). Mender's coefficient overlap showed higher overlap of insulin over proinsulin in CF sections in Phase II (p < 0.001) and Phase III (p < 0.0001) compared to WT. Mender's coefficient rate was higher in CF sections during Phase II (p < 0.001). Conclusion: CF ferret islets revealed significant immunofluorescent staining changes compared to WT during various phases of disease, providing insights into CRFD pathophysiology.
Neuregulin 4 (Nrg4) is a brown fat-enriched endocrine factor that ameliorates lipid metabolism disorders. Autophagy is critical for pancreatic beta-cell to counteract lipotoxicity-induced apoptosis. This study aimed at exploring whether Nrg4 attenuates lipotoxicity-induced beta-cell apoptosis by regulating autophagy. The mouse pancreatic beta-cell line MIN6 was cultured in palmitic acid (PA) with or without Nrg4 administration. Apoptosis rate, together with anti-apoptotic and pro-apoptotic protein levels, was investigated. Autophagic flux and autophagy-related protein levels along with related signaling pathways that regulate autophagy were also evaluated. Results showed that Nrg4 decreased PA-induced MIN6 apoptosis, enhanced anti-apoptotic protein B-cell lymphoma 2 (Bcl-2) expression and reduced pro-apoptotic proteins Bcl-2-associated X protein (Bax) and cleaved-caspase 3 expressions. Autophagy levels in MIN6 also decreased with PA treatment and Nrg4 administration reactivated autophagy. Further, Nrg4 administration activated autophagy via the mammalian target of rapamycin (mTOR) signaling pathway. In addition, when the mTOR pathway was stimulated or autophagy was suppressed, the beneficial effects of Nrg4 administration on MIN6 apoptosis were diminished. These results imply that Nrg4 administration attenuates MIN6 apoptosis by promoting mTOR-dependent autophagy and thus may lead to a new therapeutic method for type 2 diabetes mellitus (T2DM).
Background: Islet or beta-cell transplantation is a therapeutical approach to substitute the insulin-producing cells which are abolished in type 1 diabetes mellitus. The shortage of human islets as well as the complicated and costly isolation process limit the application of these techniques in daily clinical practice. EndoC-beta H is a human beta-cell line that readily forms aggregates termed pseudoislets, providing an alternative to primary human islets or beta-cells. Methods: EndoC-beta H3 cells were seeded and incubated to form pseudoislets. Their insulin secretion was analyzed by ELISA and compared with cell monolayers. Pseudoislets were transplanted into streptozotocin-treated NMRi nu/nu mice. Blood glucose was monitored before and after transplantation and compared with wild types. Grafts were analyzed by immunohistology. Results: This study shows that EndoC-beta H cells are able to form pseudoislets by aggregation, leading to an enhanced glucose stimulated insulin secretion in vitro. These pseudoislets were then successfully transplanted into the livers of diabetic mice and produced insulin in vitro. Blood glucose levels of the streptozocin-treated recipient mice were significantly decreased when compared to pre-transplantation and matched the levels found in control mice. Conclusion: We suggest pseudoislets aggregated from EndoC-beta H cells as a valuable and promising model for islet transplantation research. [GRAPHICS]
Islet amyloid polypeptide (IAPP) is a factor that regulates food intake and is secreted from both pancreatic islets and insulinoma cells. Here, we aimed to evaluate IAPP immunohistochemically in islets or insulinoma cells in association with clinical characteristics. We recruited six insulinoma patients and six body mass index-matched control patients with pancreatic diseases other than insulinoma whose glucose tolerance was confirmed to be normal preoperatively. IAPP and IAPP-insulin double staining were performed on pancreatic surgical specimens. We observed that the IAPP staining level and percentage of IAPP-positive beta cells tended to be lower (p = 0.1699) in the islets of insulinoma patients than in those of control patients, which might represent a novel IAPP expression pattern under persistent hyperinsulinemia and hypoglycemia.
Replacement of beta cells through transplantation is a potential therapeutic approach for individuals with pancreas removal or poorly controllable type 1 diabetes. However, stress and death of beta cells pose significant challenges. Circulating miRNA has emerged as potential biomarkers reflecting early beta cell stress and death, allowing for timely intervention. The aim of this study was to identify miRNAs as potential biomarkers for beta cell health. Literature review combined with small RNA sequencing was employed to select islet-enriched miRNA. The release of those miRNA was assessed by RT-qPCR in vivo, using a streptozotocin induced diabetes mouse model and in vitro, through mouse and human islets exposed to varying degrees of hypoxic and cytokine stressors. Utilizing the streptozotocin induced model, we identified 18 miRNAs out of 39 candidate islet-enriched miRNA to be released upon islet stress in vivo. In vitro analysis of culture supernatants from cytokine and/or hypoxia stressed islets identified the release of 45 miRNAs from mouse and 8 miRNAs from human islets. Investigation into the biological pathways targeted by the cytokine- and/or hypoxia-induced miRNA suggested the involvement of MAPK and PI3K-Akt signaling pathways in both mouse and human islets. We have identified miRNAs associated with beta cell health and stress. The findings allowed us to propose a panel of 47 islet-related human miRNA that is potentially valuable for application in clinical contexts of beta cell transplantation and presymptomatic early-stage type 1 diabetes.
Human islets from deceased organ donors have made important contributions to our understanding of pancreatic endocrine function and continue to be an important resource for research studies aimed at understanding, treating, and preventing diabetes. Understanding the impacts of isolation and culture upon the yield of human islets for research is important for planning research studies and islet distribution to distant laboratories. Here, we examine islet isolation and cell culture outcomes at the Alberta Diabetes Institute (ADI) IsletCore (n = 197). Research-focused isolations typically have a lower yield of islet equivalents (IEQ), with a median of 252,876 IEQ, but a higher purity (median 85%) than clinically focused isolations before culture. The median recovery of IEQs after culture was 75%, suggesting some loss. This was associated with a shift toward smaller islet particles, indicating possible islet fragmentation, and occurred within 24 h with no further loss after longer periods of culture (up to 136 h). No overall change in stimulation index as a measure of islet function was seen with culture time. These findings were replicated in a representative cohort of clinical islet preparations from the Clinical Islet Transplant Program at the University of Alberta. Thus, loss of islets occurs within 24 h of isolation, and there is no further impact of extended culture prior to islet distribution for research.
Pancreatic beta cells are among the slowest replicating cells in the human body. Human beta cells usually do not increase in number with exceptions being during the neonatal period, in cases of obesity, and during pregnancy. This project explored maternal serum for stimulatory potential on human beta cell proliferation and insulin output. Gravid, full-term women who were scheduled to undergo cesarean delivery were recruited for this study. A human beta cell line was cultured in media supplemented with serum from pregnant and non-pregnant donors and assessed for differences in proliferation and insulin secretion. A subset of pregnant donor sera induced significant increases in beta cell proliferation and insulin secretion. Pooled serum from pregnant donors also increased proliferation in primary human beta cells but not primary human hepatocytes indicating a cell-type specific effect. This study suggests stimulatory factors in human serum during pregnancy could provide a novel approach for human beta cell expansion.
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 beta-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 mu g/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 beta-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.
