Maternal obesity may lead to epigenetic alterations in the offspring and might thereby contribute to disease later in life. We investigated whether a lifestyle intervention in pregnant women with obesity is associated with epigenetic variation in cord blood and body composition in the offspring. Genome-wide DNA methylation was analyzed in cord blood from 208 offspring from the Treatment of Obese Pregnant women (TOP)-study, which includes pregnant women with obesity randomized to lifestyle interventions comprised of physical activity with or without dietary advice versus control subjects (standard of care). DNA methylation was altered at 379 sites, annotated to 370 genes, in cord blood from offspring of mothers following a lifestyle intervention versus control subjects (false discovery rate [FDR] <5%) when using the Houseman reference-free method to correct for cell composition, and three of these sites were significant based on Bonferroni correction. These 370 genes are overrepresented in gene ontology terms, including response to fatty acids and adipose tissue development. Offspring of mothers included in a lifestyle intervention were born with more lean mass compared with control subjects. Methylation at 17 sites, annotated to, for example, DISC1, GBX2, HERC2, and HUWE1, partially mediates the effect of the lifestyle intervention on lean mass in the offspring (FDR <5%). Moreover, 22 methylation sites were associated with offspring BMI z scores during the first 3 years of life (P < 0.05). Overall, lifestyle interventions in pregnant women with obesity are associated with epigenetic changes in offspring, potentially influencing the offspring's lean mass and early growth.

We and others previously demonstrated that a type 1 diabetes genetic risk score (GRS) improves the ability to predict disease progression and onset in at-risk subjects with islet autoantibodies. Here, we hypothesized that GRS and islet autoantibodies, combined with age at onset and disease duration, could serve as markers of residual β-cell function following type 1 diabetes diagnosis. Generalized estimating equations were used to investigate whether GRS along with insulinoma-associated protein-2 autoantibody (IA-2A), zinc transporter 8 autoantibody (ZnT8A), and GAD autoantibody (GADA) titers were predictive of C-peptide detection in a largely cross-sectional cohort of 401 subjects with type 1 diabetes (median duration 4.5 years [range 0-60]). Indeed, a combined model with incorporation of disease duration, age at onset, GRS, and titers of IA-2A, ZnT8A, and GADA provided superior capacity to predict C-peptide detection (quasi-likelihood information criterion [QIC] = 334.6) compared with the capacity of disease duration, age at onset, and GRS as the sole parameters (QIC = 359.2). These findings support the need for longitudinal validation of our combinatorial model. The ability to project the rate and extent of decline in residual C-peptide production for individuals with type 1 diabetes could critically inform enrollment and benchmarking for clinical trials where investigators are seeking to preserve or restore endogenous β-cell function.

MicroRNAs (miRNAs) are noncoding small RNAs that regulate various pathophysiological cellular processes. Here, we report that expression of the miR-378 family was significantly induced by metabolic inflammatory inducers, a high-fructose diet, and inflammatory cytokine tumor necrosis factor-α. Hepatic miRNA profiling revealed that expression of miR-378a was highly upregulated, which, in turn, targeted the 3'-untranslated region of PPARα mRNA, impaired mitochondrial fatty acid β-oxidation, and induced mitochondrial and endoplasmic reticulum stress. More importantly, the upregulated miR-378a can directly bind to and activate the double-strand RNA (dsRNA)-dependent protein kinase R (PKR) to sustain the metabolic stress. In vivo, genetic depletion of miR-378a prevented PKR activation and ameliorated inflammatory stress and insulin resistance. Counterbalancing the upregulated miR-378a using nanoparticles encapsulated with an anti-miR-378a oligonucleotide restored PPARα activity, inhibited PKR activation and ER stress, and improved insulin sensitivity in fructose-fed mice. Our study delineated a novel mechanism of miR-378a in the pathogenesis of metabolic inflammation and insulin resistance through targeting metabolic signaling at both mRNA (e.g., PPARα) and protein (e.g., PKR) molecules. This novel finding of functional interaction between miRNAs (e.g., miR-378a) and cellular RNA binding proteins (e.g., PKR) is biologically significant because it greatly broadens the potential targets of miRNAs in cellular pathophysiological processes.

In type 1 diabetes (T1D), a lifelong autoimmune disease, T cells infiltrate the islets and the exocrine pancreas in high numbers. CD8+ T cells are the main cell type found in the insulitic lesion, and CD8+ T cells reactive against β-cell antigens have been detected in peripheral blood and in the pancreas of patients with short- or long-term disease. In the Diabetes Virus Detection (DiViD) study, researchers collected pancreatic tissue, by pancreatic tail resection, from living patients with recent-onset T1D. These tissues have been extensively studied by the scientific community, but the autoreactive nature of the T-cell infiltrate has remained unexplored. Our objective was to determine the number and localization of these cells in pancreas samples obtained through the DiViD study. Here, we demonstrate the presence of high frequencies of CD8+ T cells reactive against a highly relevant epitope derived from the preproinsulin signal peptide in pancreatic tissue samples from these donors. We also show the heterogeneity of islet distribution and CD8+ T-cell infiltration. Our findings contribute to the current limited existing knowledge of T-cell reactivity in the pancreas of donors with recent-onset T1D and indicate that antigen-specific therapies directed toward preproinsulin could have high clinical impact.

Genome-wide association studies (GWAS) and linkage studies have had limited success in identifying genome-wide significantly linked regions or risk loci for diabetic nephropathy (DN) in individuals with type 1 diabetes (T1D). As GWAS cohorts have grown, they have also included more documented and undocumented familial relationships. Here we computationally inferred and manually curated pedigrees in a study cohort of >6,000 individuals with T1D and their relatives without diabetes. We performed a linkage study for 177 pedigrees consisting of 452 individuals with T1D and their relatives using a genome-wide genotyping array with >300,000 single nucleotide polymorphisms and PSEUDOMARKER software. Analysis resulted in genome-wide significant linkage peaks on eight chromosomal regions from five chromosomes (logarithm of odds score >3.3). The highest peak was localized at the HLA region on chromosome 6p, but whether the peak originated from T1D or DN remained ambiguous. Of other significant peaks, the chromosome 4p22 region was localized on top of ARHGAP24, a gene associated with focal segmental glomerulosclerosis, suggesting this gene may play a role in DN as well. Furthermore, rare variants have been associated with DN and chronic kidney disease near the 4q25 peak, localized on top of CCSER1.

An adverse intrauterine environment is associated with the future risk of obesity and type 2 diabetes. Changes in placental function may underpin the intrauterine origins of adult disease, but longitudinal studies linking placental function with childhood outcomes are rare. Here, we determined the abundance and phosphorylation of protein intermediates involved in insulin signaling, inflammation, cortisol metabolism, protein glycosylation, and mitochondrial biogenesis in placental villus samples from healthy mothers from the Healthy Start cohort. Using MANOVA, we tested the association between placental proteins and offspring adiposity (fat mass percentage) at birth (n = 109) and infancy (4-6 months, n = 104), and adiposity, skinfold thickness, triglycerides, and insulin in children (4-6 years, n = 66). Placental IGF-1 receptor protein was positively associated with serum triglycerides in children. GSK3β phosphorylation at serine 9, a readout of insulin and growth factor signaling, and the ratio of phosphorylated to total JNK2 were both positively associated with midthigh skinfold thickness in children. Moreover, peroxisome proliferator-activated receptor γ coactivator (PGC)-1α abundance was positively associated with insulin in children. In conclusion, placental insulin/IGF-1 signaling, PGC-1α, and inflammation pathways were positively associated with metabolic outcomes in 4- to 6-year-old children, identifying a novel link between placental function and long-term metabolic outcomes.

MicroRNAs (miRNAs) are noncoding small RNAs that regulate various pathophysiological cellular processes. Here we reported that expression of the miR-378 family was significantly induced by metabolic inflammatory inducers, a high-fructose diet, and inflammatory cytokine TNFα. Hepatic miRNA profiling revealed that expression of miR-378a was highly upregulated which, in turn, targeted the 3'-UTR of PPARα mRNA, impaired mitochondrial fatty acid β-oxidation and induced mitochondrial and ER stress. More importantly, the upregulated miR-378a can directly bind to and activate the dsRNA-dependent protein kinase R (PKR) to sustain the metabolic stress. In vivo, genetic depletion of miR-378a prevented PKR activation, ameliorated inflammatory stress and insulin resistance. Counterbalancing the upregulated miR-378a using nanoparticles encapsulated with an anti-miR-378a oligonucleotide restored PPARα activity, inhibited PKR activation and ER stress, and improved insulin sensitivity in the fructose-fed mice. Conclusion: Our study delineated a novel mechanism of miRNA-378a in the pathogenesis of metabolic inflammation and insulin resistance through targeting metabolic signaling at both mRNA (e.g., PPARα) and protein (e.g., PKR) molecules. This novel finding of functional interaction between miRNAs (e.g., miR-378a) and cellular RNA binding protein(s) (e.g., PKR) is biologically significant as it greatly broadens the potential targets of miRNAs in cellular pathophysiological processes.

The liver is a key player for maintaining glucose homeostasis. Excessive hepatic glucose production is considered to be a key for the onset of type 2 diabetes. The primary function of heme oxygenase-1 (HO1) is to catalyze the degradation of heme into biliverdin, ferrous iron, and carbon monoxide. Previous studies have demonstrated that the degradation of heme by HO1 in the liver results in mitochondrial dysfunction and drives insulin resistance. In this study, by overexpressing HO1 in hepatocytes and mice, we showed that HO1 promotes gluconeogenesis in a Foxo1-dependent manner. Importantly, HO1 overexpression increased the generation of ferrous iron in the liver, which further activates nuclear factor-κB and phosphorylates Foxo1 at Ser273 to enhance gluconeogenesis. We further assessed the role of HO1 in insulin-resistant liver-specific knockout of IRS1 and IRS2 genes (L-DKO) mice, which exhibit upregulation of HO1 in the liver and hepatic ferrous iron overload. HO1 knockdown by shRNA or treatment of iron chelator rescued the aberrant gluconeogenesis in L-DKO mice. In addition, we found that systemic iron overload promotes gluconeogenesis by activating the hepatic protein kinase A→Foxo1 axis. Thus, our results demonstrate the role of HO1 in regulating hepatic iron status and Foxo1 to control gluconeogenesis and blood glucose.

Elucidation of mechanisms that govern lipid storage, oxidative stress, and insulin resistance may lead to improved therapeutic options for type 2 diabetes and other obesity-related diseases. Here, we find that adipose expression of the small neutral amino acid transporter SLC7A10, also known as alanine-serine-cysteine transporter-1 (ASC-1), shows strong inverse correlates with visceral adiposity, insulin resistance, and adipocyte hypertrophy across multiple cohorts. Concordantly, loss of Slc7a10 function in zebrafish in vivo accelerates diet-induced body weight gain and adipocyte enlargement. Mechanistically, SLC7A10 inhibition in human and murine adipocytes decreases adipocyte serine uptake and total glutathione levels and promotes reactive oxygen species (ROS) generation. Conversely, SLC7A10 overexpression decreases ROS generation and increases mitochondrial respiratory capacity. RNA sequencing revealed consistent changes in gene expression between human adipocytes and zebrafish visceral adipose tissue following loss of SLC7A10, e.g., upregulation of SCD (lipid storage) and downregulation of CPT1A (lipid oxidation). Interestingly, ROS scavenger reduced lipid accumulation and attenuated the lipid-storing effect of SLC7A10 inhibition. These data uncover adipocyte SLC7A10 as a novel important regulator of adipocyte resilience to nutrient and oxidative stress, in part by enhancing glutathione levels and mitochondrial respiration, conducive to decreased ROS generation, lipid accumulation, adipocyte hypertrophy, insulin resistance, and type 2 diabetes.

In diabetes, increasing albuminuria and decreasing glomerular filtration rate are hallmarks of chronic kidney disease in diabetes and increase the risk of atherosclerotic cardiovascular events and mortality as well as the risk for end-stage kidney disease. For two decades, standard of care has been controlling risk factors, such as glucose, blood pressure, lipids, and lifestyle factors, and specifically use of agents blocking the renin-angiotensin system. This has improved outcome, but a large unmet need has been obvious. After many failed attempts to advance the therapeutic options, the past few years have provided several new promising treatment options such as sodium-glucose cotransporter 2 inhibitors, endothelin receptor antagonists, glucagon-like peptide 1 agonists, and nonsteroidal mineralocorticoid receptor antagonists. The benefits and side effects of these agents demonstrate the link between kidney and heart; some have beneficial effects on both, whereas for other potentially renoprotective agents, development of heart failure has been a limiting factor. They work on different pathways such as hemodynamic, metabolic, inflammatory, and fibrotic targets. We propose that treatment may be personalized if biomarkers or physiological investigations assessing activity in these pathways are applied. This could potentially pave the way for precision medicine, where treatment is optimized for maximal benefit and minimal adverse outcomes. At least it may help prioritizing agents for an individual subject.

The study of humans with genetic mutations which lead to a substantial disturbance of physiological processes has made a contribution to biomedical science that is disproportionate to the rarity of affected individuals. In this lecture, I discuss examples of where such studies have helped to illuminate two areas of human metabolism. First, the control of insulin sensitivity and its disruption in states of insulin resistance and second, the regulation of energy balance and its disturbances in obesity.

Pancreatic islets are clusters of hormone-secreting endocrine cells that rely on intricate cell-cell communication mechanisms for proper function. The importance of multicellular cooperation in islet cell physiology was first noted nearly 30 years ago in seminal studies showing that hormone secretion from endocrine cell types is diminished when these cells are dispersed. These studies showed that reestablishing cellular contacts in so-called pseudoislets caused endocrine cells to regain hormone secretory function. This not only demonstrated that cooperation between islet cells is highly synergistic but also gave birth to the field of pancreatic islet organoids. Here we review recent advances related to the mechanisms of islet cell cross talk. We first describe new developments that revise current notions about purinergic and GABA signaling in islets. Then we comment on novel multicellular imaging studies that are revealing emergent properties of islet communication networks. We finish by highlighting and discussing recent synthetic approaches that use islet organoids of varied cellular composition to interrogate intraislet signaling mechanisms. This reverse engineering of islets not only will shed light on the mechanisms of intraislet signaling and define communication networks but also may guide efforts aimed at restoring islet function and β-cell mass in diabetes.

Sodium-glucose cotransporter 2 (SGLT2) inhibition reduces cardiovascular morbidity and mortality in individuals with type 2 diabetes. Beneficial effects have been attributed to increased ketogenesis, reduced cardiac fatty acid oxidation, and diminished cardiac oxygen consumption. We therefore studied whether SGLT2 inhibition altered cardiac oxidative substrate consumption, efficiency, and perfusion. Thirteen individuals with type 2 diabetes were studied after 4 weeks' treatment with empagliflozin and placebo in a randomized, double-blind, placebo-controlled crossover study. Myocardial palmitate and glucose uptake were measured with 11C-palmitate and 18F-fluorodeoxyglucose positron emission tomography (PET)/computed tomography (CT). Oxygen consumption and myocardial external efficiency (MEE) were measured with 11C-acetate PET/CT. Resting and adenosine stress myocardial blood flow (MBF) and myocardial flow reserve (MFR) were measured using 15O-H2O PET/CT. Empagliflozin did not affect myocardial free fatty acids (FFAs) uptake but reduced myocardial glucose uptake by 57% (P < 0.001). Empagliflozin did not change myocardial oxygen consumption or MEE. Empagliflozin reduced resting MBF by 13% (P < 0.01), but did not significantly affect stress MBF or MFR. In conclusion, SGLT2 inhibition did not affect myocardial FFA uptake, but channeled myocardial substrate utilization from glucose toward other sources and reduced resting MBF. However, the observed metabolic and hemodynamic changes were modest and most likely contribute only partially to the cardioprotective effect of SGLT2 inhibition.

Patients with diabetes often experience visual defects before any retinal pathologies are detected. The molecular mechanism for the visual defects in early diabetes has not been elucidated. Our previous study reported that in early diabetic retinopathy (DR), rhodopsin levels were reduced due to impaired 11-cis-retinal regeneration. Interphotoreceptor retinol-binding protein (IRBP) is a visual cycle protein and important for 11-cis-retinal generation. IRBP levels are decreased in the vitreous and retina of DR patients and animal models. To determine the role of IRBP downregulation in the visual defects in early DR, we induced diabetes in transgenic mice overexpressing IRBP in the retina. IRBP overexpression prevented diabetes-induced decline of retinal function. Furthermore, IRBP overexpression also prevented decreases of rhodopsin levels and 11-cis-retinal generation in diabetic mice. Diabetic IRBP transgenic mice also showed ameliorated retinal oxidative stress, inflammation, apoptosis, and retinal degeneration compared with diabetic wild-type mice. These findings suggest that diabetes-induced IRBP downregulation impairs the regeneration of 11-cis-retinal and rhodopsin, leading to retinal dysfunction in early DR. Furthermore, increased 11-cis-retinal-free opsin constitutively activates the phototransduction pathway, leading to increased oxidative stress and retinal neurodegeneration. Therefore, restored IRBP expression in the diabetic retina may confer a protective effect against retinal degeneration in DR.

Aberrantly elevated expression in obesity of microRNAs (miRNAs), including the miRNA miR-802, contributes to obesity-associated metabolic complications, but the mechanisms underlying the elevated expression are unclear. Farnesoid X receptor (FXR), a key regulator of hepatic energy metabolism, has potential for treatment of obesity-related diseases. We examined whether a nuclear receptor cascade involving FXR and FXR-induced small heterodimer partner (SHP) regulates expression of miR-802 to maintain glucose and lipid homeostasis. Hepatic miR-802 levels are increased in FXR-knockout (KO) or SHP-KO mice and are decreased by activation of FXR in a SHP-dependent manner. Mechanistically, transactivation of miR-802 by aromatic hydrocarbon receptor (AHR) is inhibited by SHP. In obese mice, activation of FXR by obeticholic acid treatment reduced miR-802 levels and improved insulin resistance and hepatosteatosis, but these beneficial effects were largely abolished by overexpression of miR-802. In patients with nonalcoholic fatty liver disease (NAFLD) and in obese mice, occupancy of SHP is reduced and that of AHR is modestly increased at the miR-802 promoter, consistent with elevated hepatic miR-802 expression. These results demonstrate that normal inhibition of miR-802 by FXR-SHP is defective in obesity, resulting in increased miR-802 levels, insulin resistance, and fatty liver. This FXR-SHP-miR-802 pathway may present novel targets for treating type 2 diabetes and NAFLD.

Low-grade persistent inflammation is a feature of diabetes-driven vascular complications, in particular activation of the Nod-like receptor family pyrin domain containing 3 (NLRP3) inflammasome to trigger the maturation and release of the inflammatory cytokine interleukin-1β (IL-1β). We investigated whether inhibiting the NLRP3 inflammasome, through the use of the specific small-molecule NLRP3 inhibitor MCC950, could reduce inflammation, improve vascular function, and protect against diabetes-associated atherosclerosis in the streptozotocin-induced diabetic apolipoprotein E-knockout mouse. Diabetes led to an approximately fourfold increase in atherosclerotic lesions throughout the aorta, which were significantly attenuated with MCC950 (P < 0.001). This reduction in lesions was associated with decreased monocyte-macrophage content, reduced necrotic core, attenuated inflammatory gene expression (IL-1β, tumor necrosis factor-α, intracellular adhesion molecule 1, and MCP-1; P < 0.05), and reduced oxidative stress, while maintaining fibrous cap thickness. Additionally, vascular function was improved in diabetic vessels of mice treated with MCC950 (P < 0.05). In a range of cell lines (murine bone marrow-derived macrophages, human monocytic THP-1 cells, phorbol 12-myristate 13-acetate-differentiated human macrophages, and aortic smooth muscle cells from humans with diabetes), MCC950 significantly reduced IL-1β and/or caspase-1 secretion and attenuated leukocyte-smooth muscle cell interactions under high glucose or lipopolysaccharide conditions. In summary, MCC950 reduces plaque development, promotes plaque stability, and improves vascular function, suggesting that targeting NLRP3-mediated inflammation is a novel therapeutic strategy to improve diabetes-associated vascular disease.

CD11c+ macrophages/dendritic cells (MDCs) are increased and display the classically activated M1-like phenotype in obese adipose tissue (AT) and may contribute to AT inflammation and insulin resistance. Stat1 is a key transcription factor for MDC polarization into the M1-like phenotype. Here, we examined the role of Stat1 in obesity-induced AT MDC polarization and inflammation and insulin resistance using mice with specific knockout of Stat1 in MDCs (cKO). Stat1 was upregulated and phosphorylated, indicating activation, early and persistently in AT and AT MDCs of wild-type mice fed a high-fat diet (HFD). Compared with littermate controls, cKO mice fed an HFD (16 weeks) had reductions in MDC (mainly CD11c+ macrophage) M1-like polarization and interferon-γ-expressing T-helper type 1 (Th1) cells but increases in interleukin 5-expressing Th2 cells and eosinophils in perigonadal and inguinal AT, and enhanced inguinal AT browning, with increased energy expenditure. cKO mice compared with controls also had significant reductions in triglyceride content in the liver and skeletal muscle and exhibited improved insulin sensitivity and glucose tolerance. Taken together, our results demonstrate that Stat1 in MDCs plays an important role in obesity-induced MDC M1-like polarization and AT inflammation and contributes to insulin resistance and metabolic dysfunctions in obese mice.

The causes of the increased risk of severe coronavirus disease 2019 (COVID-19) in people with diabetes are unclear. It has been speculated that renin-angiotensin system (RAS) blockers may promote COVID-19 by increasing ACE2, which severe acute respiratory syndrome coronavirus 2 uses to enter host cells, along with the host protease TMPRSS2. Taking a reverse translational approach and by combining in situ hybridization, primary cell isolation, immunoblotting, quantitative RT-PCR, and liquid chromatography-tandem mass spectrometry, we studied lung and kidney ACE2 and TMPRSS2 in diabetic mice mimicking host factors linked to severe COVID-19. In healthy young mice, neither the ACE inhibitor ramipril nor the AT1 receptor blocker telmisartan affected lung or kidney ACE2 or TMPRSS2, except for a small increase in kidney ACE2 protein with ramipril. In contrast, mice with comorbid diabetes (aging, high-fat diet, and streptozotocin-induced diabetes) had heightened lung ACE2 and TMPRSS2 protein levels and increased lung ACE2 activity. None of these parameters were affected by RAS blockade. ACE2 was similarly upregulated in the kidneys of mice with comorbid diabetes compared with aged controls, whereas TMPRSS2 (primarily distal nephron) was highest in telmisartan-treated animals. Upregulation of lung ACE2 activity in comorbid diabetes may contribute to an increased risk of severe COVID-19. This upregulation is driven by comorbidity and not by RAS blockade.