Excessive adiposity is the main cause of obesity and type two diabetes (T2D). Variants in human IMP2/IGF2BP2 gene are associated with increased risk of T2D. However, little is known about its role in adipogenesis and in insulin resistance. Here, we investigate the function of IMP2 during adipocyte development. Mice with Imp2 deletion in mesenchymal stem cells (MSC) are resistant to diet-induced obesity without glucose and insulin tolerance affected. Imp2 is essential for the early commitment of adipocyte-derived stem cells (ADSC) into preadipocytes, but the deletion of Imp2 in MSC is not required for the proliferation and terminal differentiation of committed preadipocytes. Mechanistically, Imp2 binds Wnt receptor Fzd8 mRNA and promotes its degradation by recruiting CCR4-NOT deadenylase complex in an mTOR-dependent manner. Our data demonstrate that Imp2 is required for maintaining white adipose tissue homeostasis through controlling mRNA stability in ADSC. However, the contribution of IMP2 to insulin resistance, a main risk of T2D, is not evident.

A recent discovery effort resulted in identification of novel splice variant and secretory granule antigens within the HLA class I peptidome of human islets and documentation of their recognition by CD8+ T cells from peripheral blood and human islets. In the current study, we applied a systematic discovery process to identify novel CD4+ T cell epitopes derived from these candidate antigens. We predicted 145 potential epitopes spanning unique splice junctions and within conventional secretory granule antigens and measured their in vitro binding to DRB1*04:01. We generated HLA class II tetramers for the 35 peptides with detectable binding and used these to assess immunogenicity and isolate T cell clones. Tetramers corresponding to peptides with verified immunogenicity were then used to label T cells specific for these putative epitopes in peripheral blood. T cells that recognize distinct epitopes derived from a cyclin I splice variant, neuroendocrine convertase 2, and urocortin-3 were detected at frequencies that were similar to those of an immunodominant proinsulin epitope. Cells specific for these novel epitopes predominantly exhibited a Th1-like surface phenotype. Among the three epitopes, responses to the cyclin I peptide exhibited a distinct memory profile. Responses to neuroendocrine convertase 2 were detected among pancreatic infiltrating T cells. These results further establish the contribution of unconventional antigens to the loss of tolerance in autoimmune diabetes.

The functional mass of insulin-secreting pancreatic β-cells expands to maintain glucose homeostasis in the face of nutrient excess, in part via replication of existing β-cells. Type 2 diabetes appears when these compensatory mechanisms fail. Nutrients including glucose and fatty acids are important contributors to the β-cell compensatory response, but their underlying mechanisms of action remain poorly understood. We investigated the transcriptional mechanisms of β-cell proliferation in response to fatty acids. Isolated rat islets were exposed to 16.7 mmol/L glucose with or without 0.5 mmol/L oleate (C18:1) or palmitate (C16:0) for 48 h. The islet transcriptome was assessed by single-cell RNA sequencing. β-Cell proliferation was measured by flow cytometry. Unsupervised clustering of pooled β-cells identified different subclusters, including proliferating β-cells. β-Cell proliferation increased in response to oleate but not palmitate. Both fatty acids enhanced the expression of genes involved in energy metabolism and mitochondrial activity. Comparison of proliferating versus nonproliferating β-cells and pseudotime ordering suggested the involvement of reactive oxygen species (ROS) and peroxiredoxin signaling. Accordingly, N-acetyl cysteine and the peroxiredoxin inhibitor conoidin A both blocked oleate-induced β-cell proliferation. Our study reveals a key role for ROS signaling through peroxiredoxin activation in oleate-induced β-cell proliferation.

The stress response protein regulated in development and DNA damage response 1 (REDD1) has been implicated in visual deficits in patients with diabetes. The aim here was to investigate the mechanism responsible for the increase in retinal REDD1 protein content that is observed with diabetes. We found that REDD1 protein expression was increased in the retina of streptozotocin-induced diabetic mice in the absence of a change in REDD1 mRNA abundance or ribosome association. Oral antioxidant supplementation reduced retinal oxidative stress and suppressed REDD1 protein expression in the retina of diabetic mice. In human retinal Müller cell cultures, hyperglycemic conditions increased oxidative stress, enhanced REDD1 expression, and inhibited REDD1 degradation independently of the proteasome. Hyperglycemic conditions promoted a redox-sensitive cross-strand disulfide bond in REDD1 at C150/C157 that was required for reduced REDD1 degradation. Discrete molecular dynamics simulations of REDD1 structure revealed allosteric regulation of a degron upon formation of the disulfide bond that disrupted lysosomal proteolysis of REDD1. REDD1 acetylation at K129 was required for REDD1 recognition by the cytosolic chaperone HSC70 and degradation by chaperone-mediated autophagy. Disruption of REDD1 allostery upon C150/C157 disulfide bond formation prevented the suppressive effect of hyperglycemic conditions on REDD1 degradation and reduced oxidative stress in cells exposed to hyperglycemic conditions. The results reveal redox regulation of REDD1 and demonstrate the role of a REDD1 disulfide switch in development of oxidative stress.

The phenotypic and functional plasticity of adipose tissue macrophages (ATMs) during obesity plays a crucial role in orchestration of adipose and systemic inflammation. Tonicity-responsive enhancer binding protein (TonEBP) (also called NFAT5) is a stress protein that mediates cellular responses to a range of metabolic insults. Here, we show that myeloid cell-specific TonEBP depletion reduced inflammation and insulin resistance in mice with high-fat diet-induced obesity but did not affect adiposity. This phenotype was associated with a reduced accumulation and a reduced proinflammatory phenotype of metabolically activated macrophages, decreased expression of inflammatory factors related to insulin resistance, and enhanced insulin sensitivity. TonEBP expression was elevated in the ATMs of obese mice, and Sp1 was identified as a central regulator of TonEBP induction. TonEBP depletion in macrophages decreased induction of insulin resistance-related genes and promoted induction of insulin sensitivity-related genes under obesity-mimicking conditions and thereby improved insulin signaling and glucose uptake in adipocytes. mRNA expression of TonEBP in peripheral blood mononuclear cells was positively correlated with blood glucose levels in mice and humans. These findings suggest that TonEBP in macrophages promotes obesity-associated systemic insulin resistance and inflammation, and downregulation of TonEBP may induce a healthy metabolic state during obesity.

Mitochondria play a vital role in white adipose tissue (WAT) homeostasis including adipogenesis, fatty acid synthesis, and lipolysis. We recently reported that the mitochondrial fusion protein optic atrophy 1 (OPA1) is required for induction of fatty acid oxidation and thermogenic activation in brown adipocytes. In the current study we investigated the role of OPA1 in WAT function in vivo. We generated mice with constitutive or inducible knockout of OPA1 selectively in adipocytes. Studies were conducted under baseline conditions, at thermoneutrality, following high-fat feeding or during cold exposure. OPA1 deficiency reduced mitochondrial respiratory capacity in white adipocytes, impaired lipolytic signaling, repressed expression of de novo lipogenesis and triglyceride synthesis pathways, and promoted adipose tissue senescence and inflammation. Reduced WAT mass was associated with hepatic triglycerides accumulation and glucose intolerance. Moreover, mice deficient for OPA1 in adipocytes had impaired adaptive thermogenesis and reduced cold-induced browning of subcutaneous WAT and were completely resistant to diet-induced obesity. In conclusion, OPA1 expression and function in adipocytes are essential for adipose tissue expansion, lipid biosynthesis, and fatty acid mobilization of WAT and brown adipocytes and for thermogenic activation of brown and beige adipocytes.

The transition from lean to obese states involves systemic metabolic remodeling that impacts insulin sensitivity, lipid partitioning, inflammation, and glycemic control. Here, we have taken a pharmacological approach to test the role of a nutrient-regulated chromatin modifier, lysine-specific demethylase (LSD1), in obesity-associated metabolic reprogramming. We show that systemic administration of an LSD1 inhibitor (GSK-LSD1) reduces food intake and body weight, ameliorates nonalcoholic fatty liver disease (NAFLD), and improves insulin sensitivity and glycemic control in mouse models of obesity. GSK-LSD1 has little effect on systemic metabolism of lean mice, suggesting that LSD1 has a context-dependent role in promoting maladaptive changes in obesity. In analysis of insulin target tissues we identified white adipose tissue as the major site of insulin sensitization by GSK-LSD1, where it reduces adipocyte inflammation and lipolysis. We demonstrate that GSK-LSD1 reverses NAFLD in a non-hepatocyte-autonomous manner, suggesting an indirect mechanism potentially via inhibition of adipocyte lipolysis and subsequent effects on lipid partitioning. Pair-feeding experiments further revealed that effects of GSK-LSD1 on hyperglycemia and NAFLD are not a consequence of reduced food intake and weight loss. These findings suggest that targeting LSD1 could be a strategy for treatment of obesity and its associated complications including type 2 diabetes and NAFLD.

Several unrelated findings led us to hypothesize that induction of autoimmunity is a consequence of a prior major inflammatory event in individuals with susceptible HLA phenotypes and elevated sensitivity to cytokines and free fatty acids (FFA). We observed provocative enhanced responsiveness of cultured human fibroblasts from individuals with type 1 diabetes (T1D), but not control subjects, to FFA and the inflammatory cytokines TNFα and IL1-β. Major infections increase inflammatory cytokines as well as circulating FFA. Endotoxin-treated animal models of sepsis also exhibit elevated inflammatory cytokines that inhibit FFA oxidation and elevate FFA. The pancreatic β-cell possesses low reactive oxygen species (ROS) scavenging capacity and responds to both elevated FFA and cytokines with increased ROS production, a combination that increases exocytosis and trafficking of secretory vesicles to the plasma membrane. Increased trafficking is accompanied by increased cycling of secretory granule proteins and may be linked with increased surface presentation of granule proteins to the immune system. We propose that this ultimately targets β-cell granular proteins at the cell surface and is consistent with the preponderance of autoantibodies to granule proteins. Our hypothesis encourages testing of potential early therapeutic interventions to prevent progression of β-cell destruction.

Preeclampsia is a pregnancy-specific complication with long-term negative outcomes for offspring, including increased susceptibility to type 2 diabetes (T2D) in adulthood. In a rat reduced uteroplacental perfusion pressure (RUPP) model of chronic placental ischemia, maternal hypertension in conjunction with intrauterine growth restriction mimicked aspects of preeclampsia and resulted in female embryonic day 19 (e19) offspring with reduced β-cell area and increased β-cell apoptosis compared with offspring of sham pregnancies. Decreased pancreatic β-cell area persisted to postnatal day 13 (PD13) in females and could influence whether T2D developed in adulthood. Macrophage changes also occurred in islets in T2D. Therefore, we hypothesized that macrophages are crucial to reduction in pancreatic β-cell area in female offspring after chronic placental ischemia. Macrophage marker CD68 mRNA expression was significantly elevated in e19 and PD13 islets isolated from female RUPP offspring compared with sham. Postnatal injections of clodronate liposomes into female RUPP and sham offspring on PD2 and PD9 significantly depleted macrophages compared with injections of control liposomes. Depletion of macrophages rescued reduced β-cell area and increased β-cell proliferation and size in RUPP offspring. Our studies suggest that the presence of macrophages is important for reduced β-cell area in female RUPP offspring and changes in macrophages could contribute to development of T2D in adulthood.

South Asian women have a higher risk of type 2 diabetes after gestational diabetes mellitus (GDM) than Nordic women; however, the mechanisms behind this difference remain unclear. We investigated insulin sensitivity, β-cell function, and hepatic insulin clearance in 179 South Asian and 108 Nordic women ∼17 months after GDM (mean age 35.3 years, BMI 29.1 kg/m2) by oral glucose tolerance test using deconvolution of C-peptide kinetics. Thirty-one percent of South Asian and 53% of Nordic participants were normoglycemic at the time of measurement. South Asian women had higher areas under the curve (AUCs) for glucose, prehepatic insulin, and peripheral insulin and lower insulin sensitivity, disposition index, and fasting hepatic insulin clearance than Nordic women. In the group with prediabetes or diabetes, South Asian women had similar AUCs for glucose and prehepatic insulin but a higher AUC for peripheral insulin, lower disposition index, and lower fasting hepatic insulin clearance than Nordic women. The waist-to-height ratio mediated ∼25-40% of the ethnic differences in insulin sensitivity in participants with normoglycemia. Overall, our novel data revealed that South Asian women with normoglycemia after GDM showed lower insulin secretion for a given insulin resistance and lower hepatic insulin clearance than Nordic women. South Asian women are at high risk of developing type 2 diabetes after GDM, and preventive efforts should be prioritized.

In our previous data-driven analysis of evolving patterns of islet autoantibodies (IAb) against insulin (IAA), GAD (GADA), and islet antigen 2 (IA-2A), we discovered three trajectories, characterized according to multiple IAb (TR1), IAA (TR2), or GADA (TR3) as the first appearing autoantibodies. Here we examined the evolution of IAb levels within these trajectories in 2,145 IAb-positive participants followed from early life and compared those who progressed to type 1 diabetes (n = 643) with those remaining undiagnosed (n = 1,502). With use of thresholds determined by 5-year diabetes risk, four levels were defined for each IAb and overlaid onto each visit. In diagnosed participants, high IAA levels were seen in TR1 and TR2 at ages <3 years, whereas IAA remained at lower levels in the undiagnosed. Proportions of dwell times (total duration of follow-up at a given level) at the four IAb levels differed between the diagnosed and undiagnosed for GADA and IA-2A in all three trajectories (P < 0.001), but for IAA dwell times differed only within TR2 (P < 0.05). Overall, undiagnosed participants more frequently had low IAb levels and later appearance of IAb than diagnosed participants. In conclusion, while it has long been appreciated that the number of autoantibodies is an important predictor of type 1 diabetes, consideration of autoantibody levels within the three autoimmune trajectories improved differentiation of IAb-positive children who progressed to type 1 diabetes from those who did not.

The aim of this cross-sectional study was to assess retinal oxygen metabolism in patients with type 2 diabetes and different stages of nonproliferative diabetic retinopathy (DR) (n = 67) compared with healthy control subjects (n = 20). Thirty-four patients had no DR, 15 had mild DR, and 18 had moderate to severe DR. Retinal oxygen saturation in arteries and veins was measured using the oxygen module of a retinal vessel analyzer. Total retinal blood flow (TRBF) was measured using a custom-built Doppler optical coherence tomography system. Retinal oxygen extraction was calculated from retinal oxygen saturation and TRBF. Arteriovenous difference in oxygen saturation was highest in healthy subjects (34.9 ± 7.5%), followed by patients with no DR (32.5 ± 6.3%) and moderate to severe DR (30.3 ± 6.5%). The lowest values were found in patients with mild DR (27.3 ± 8.0%, P = 0.010 vs. healthy subjects). TRBF tended to be higher in patients with no DR (40.1 ± 9.2 μL/min) and mild DR (41.8 ± 15.0 μL/min) than in healthy subjects (37.2 ± 5.7 μL/min) and patients with moderate to severe DR (34.6 ± 10.4 μL/min). Retinal oxygen extraction was the highest in healthy subjects (2.24 ± 0.57 μL O2/min), followed by patients with no DR (2.14 ± 0.6 μL O2/min), mild DR (1.90 ± 0.77 μL O2/min), and moderate to severe DR (1.78 ± 0.57 μL O2/min, P = 0.040 vs. healthy subjects). These results indicate that retinal oxygen metabolism is altered in patients with type 2 diabetes. Furthermore, retinal oxygen extraction decreases with increasing severity of DR.

Intrauterine exposure to metabolic dysfunction leads to offspring metabolic dysfunction in human and rodent models, but underlying mechanisms are unclear. The mediobasal hypothalamus (MBH) is involved in energy homeostasis and weight regulation, and MBH gliosis is associated with obesity and insulin resistance. We tested the hypothesis that offspring exposed to gestational diabetes mellitus (GDM) in utero versus those unexposed would show evidence of MBH gliosis. Participants in the BrainChild Study (age 7-11 years with confirmed GDM exposure or no GDM exposure) underwent brain MRI to acquire T2-weighted images. By using the amygdala (AMY) and white matter (WM) as reference regions, MBH:AMY and MBH:WM T2 signal ratios were calculated as a radiologic measure of MBH gliosis. Linear regressions were used to examine associations between GDM exposure (GDM overall) and by timing of GDM exposure (≤26 weeks or >26 weeks) and MBH gliosis. Associations between prepregnancy BMI and child MBH gliosis were examined in secondary analyses. There were no differences in T2 signal ratios in children exposed versus not exposed to GDM overall, but children exposed to early GDM (≤26 weeks of gestation) had higher MBH:WM signal ratios than those not exposed (β = 0.147; SE 0.06; P = 0.03), adjusting for child's age, sex, and BMI z score and maternal prepregnancy BMI, whereas no associations were seen for the control ratio (AMY:WM). Prepregnancy BMI was not associated with evidence of MBH gliosis. Early exposure to GDM was associated with radiologic evidence of MBH gliosis in children. These data provide mechanistic insight into brain pathways by which exposure to GDM may increase risk for metabolic dysfunction.

Fatty acids (FAs) have been implicated in the development of gestational diabetes mellitus (GDM), but the role of monounsaturated FAs (MUFAs) remains understudied. We investigated the associations of plasma phospholipid MUFAs in early to mid-pregnancy with cardiometabolic biomarkers and GDM risk. From the Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD) Fetal Growth Studies-Singletons cohort (2009-2013), we identified 107 women with GDM according to Carpenter and Coustan criteria and 214 control participants without GDM matched (2:1) on age, race/ethnicity, and gestational week (GW) of blood collection. MUFAs were measured at 10-14, 15-26, 23-31, and 33-39 GWs by gas chromatography mass spectrometry. We found that the concentration of total 18:1 MUFAs was significantly lower among women with GDM than those without GDM at 15-26 GWs. Each SD increment in the level of total 18:1 MUFAs was associated with a 40% lower risk of GDM at 15-26 GWs. Moreover, each SD increment in vaccenic acid (18:1n-7) levels at 10-14 and 15-26 GWs were associated with a 36% and 45% lower risk of GDM, respectively. Our extensive assessments of MUFAs advance our understanding of the unique associations of FA composition with GDM risk, suggesting the potentially beneficial role of MUFAs in GDM pathophysiology.

This study examined the incidence trends of new-onset type 1 and type 2 diabetes in children and adolescents in Florida before and during the coronavirus disease 2019 (COVID-19) pandemic. In this observational descriptive cohort study, we used a validated computable phenotype to identify incident diabetes cases among individuals <18 years of age in the OneFlorida+ network of the national Patient-Centered Clinical Research Network between January 2017 and June 2021. We conducted an interrupted time series analysis based on the autoregressive integrated moving average model to compare changes in age-adjusted incidence rates of type 1 and type 2 diabetes before and after March 2020, when COVID-19 was declared a national health emergency in the U.S. The age-adjusted incidence rates of both type 1 and type 2 diabetes increased post-COVID-19 for children and adolescents. These results highlight the need for longitudinal cohort studies to examine how the pandemic might influence subsequent diabetes onset in young individuals.

Pancreatic islets are highly interconnected structures that produce pulses of insulin and other hormones, maintaining normal homeostasis of glucose and other nutrients. Normal stimulus-secretion and intercellular coupling are essential to regulated secretory responses, and these hallmarks are known to be altered in diabetes. In the current study, we used calcium imaging of isolated human islets to assess their collective behavior. The activity occurred in the form of calcium oscillations, was synchronized across different regions of islets through calcium waves, and was glucose dependent: higher glucose enhanced the activity, elicited a greater proportion of global calcium waves, and led to denser and less fragmented functional networks. Hub regions were identified in stimulatory conditions, and they were characterized by long active times. Moreover, calcium waves were found to be initiated in different subregions and the roles of initiators and hubs did not overlap. In type 2 diabetes, glucose dependence was retained, but reduced activity, locally restricted waves, and more segregated networks were detected compared with control islets. Interestingly, hub regions seemed to suffer the most by losing a disproportionately large fraction of connections. These changes affected islets from donors with diabetes in a heterogeneous manner.

Islet/β-cell transplantation offers great hope for patients with type 1 diabetes. We assessed the mechanisms of how intrahepatic coinfusion of human α-1 antitrypsin (hAAT)-engineered mesenchymal stromal cells (hAAT-MSCs) improves survival of human islet grafts posttransplantation (PT). Longitudinal in vivo bioluminescence imaging studies identified significantly more islets in the livers bearing islets cotransplanted with hAAT-MSCs compared with islets transplanted alone. In vitro mechanistic studies revealed that hAAT-MSCs inhibit macrophage migration and suppress IFN-γ-induced M1-like macrophages while promoting IL-4-induced M2-like macrophages. In vivo this translated to significantly reduced CD11c+ and F4/80+ cells and increased CD206+ cells around islets cotransplanted with hAAT-MSCs as identified by multiplex immunofluorescence staining. Recipient-derived F4/80+and CD11b+ macrophages were mainly present in the periphery of an islet, while CD11c+ and CD206+ cells appeared inside an islet. hAAT-MSCs inhibited macrophage migration and skewed the M1-like phenotype toward an M2 phenotype both in vitro and in vivo, which may have favored islet survival. These data provide evidence that hAAT-MSCs cotransplanted with islets remain in the liver and shift macrophages to a protective state that favors islet survival. This novel strategy may be used to enhance β-cell survival during islet/β-cell transplantation for the treatment of type 1 diabetes or other diseases.

Hybrid insulin peptides (HIPs) form in pancreatic β-cells through the formation of peptide bonds between proinsulin fragments and other peptides. HIPs have been identified in pancreatic islets by mass spectrometry and are targeted by CD4 T cells in patients with type 1 diabetes (T1D) as well as by pathogenic CD4 T-cell clones in nonobese diabetic (NOD) mice. The mechanism of HIP formation is currently poorly understood; however, it is well established that proteases can drive the formation of new peptide bonds in a side reaction during peptide bond hydrolysis. Here, we used a proteomic strategy on enriched insulin granules and identified cathepsin D (CatD) as the primary protease driving the specific formation of HIPs targeted by disease-relevant CD4 T cells in T1D. We also established that NOD islets deficient in cathepsin L (CatL), another protease implicated in the formation of disease-relevant HIPs, contain elevated levels of HIPs, indicating a role for CatL in the proteolytic degradation of HIPs. In summary, our data suggest that CatD may be a therapeutic target in efforts to prevent or slow the autoimmune destruction of β-cells mediated by HIP-reactive CD4 T cells in T1D.