GRP75 (75-kDA glucose-regulated protein), defined as a major component of both the mitochondrial quality control system and mitochondria-associated membrane, plays a key role in mitochondrial homeostasis. In this study, we assessed the roles of GRP75, other than as a component, in insulin action in both in vitro and in vivo models with insulin resistance. We found that GRP75 was downregulated in mice fed a high-fat diet (HFD) and that induction of Grp75 in mice could prevent HFD-induced obesity and insulin resistance. Mechanistically, GRP75 influenced insulin sensitivity by regulating mitochondrial function through its modulation of mitochondrial-supercomplex turnover rather than mitochondria-associated membrane communication: GRP75 was negatively associated with respiratory chain complex activity and was essential for mitochondrial-supercomplex assembly and stabilization. Moreover, mitochondrial dysfunction in Grp75-knockdown cells might further increase mitochondrial fragmentation, thus triggering cytosolic mtDNA release and activating the cGAS/STING-dependent proinflammatory response. Therefore, GRP75 can serve as a potential therapeutic target of insulin resistant-related diabetes or other metabolic diseases.

Signal regulatory protein SIRPγ (CD172G) is expressed on the surface of lymphocytes, where it acts by engaging its ligand, CD47. SIRPG, which encodes SIRPγ, contains a nonsynonymous coding variant, rs6043409, which is significantly associated with risk for type 1 diabetes. SIRPG produces multiple transcript isoforms via alternative splicing, all encoding potentially functional proteins. We show that rs6043409 alters a predicted exonic splicing enhancer, resulting in significant shifts in the distribution of SIRPG transcript isoforms. All of these transcript isoforms produced protein upon transient expression in vitro. However, CRISPR/Cas9 targeting of one of the alternatively spliced exons in SIRPG eliminated all SIRPγ expression in Jurkat T cells. These targeted cells formed fewer cell-cell conjugates with each other than with wild-type Jurkat cells, expressed reduced levels of genes associated with CD47 signaling, and had significantly increased levels of cell-surface CD47. In primary CD4+ and CD8+ T cells, cell-surface SIRPγ levels in response to anti-CD3 stimulation varied quantitatively by rs6043409 genotype. Our results suggest that SIRPG is the most likely causative gene for type 1 diabetes risk in the 20p13 region and highlight the role of alternative splicing in lymphocytes in mediating the genetic risk for autoimmunity.

At present, outside of infancy, genetic testing for monogenic diabetes is typically for mutations in maturity-onset diabetes of the young (MODY) genes that predominantly result in isolated diabetes. Monogenic diabetes syndromes are usually only tested for when supported by specific syndromic clinical features. How frequently patients with suspected MODY have a mutation in a monogenic syndromic diabetes gene is unknown and thus missed by present testing regimes. We performed genetic testing of 27 monogenic diabetes genes (including 18 associated with syndromic diabetes) for 1,280 patients with a clinical suspicion of MODY who were not suspected of having monogenic syndromic diabetes. We confirmed monogenic diabetes in 297 (23%) patients. Mutations in seven different syndromic diabetes genes accounted for 19% (95% CI 15-24%) of all monogenic diabetes. The mitochondrial m.3243A>G and mutations in HNF1B were responsible for the majority of mutations in syndromic diabetes genes. They were also the 4th and 5th most common causes of monogenic diabetes overall. These patients lacked typical features, and their diabetes phenotypes overlapped with patients with nonsyndromic monogenic diabetes. Syndromic monogenic diabetes genes (particularly m.3243A>G and HNF1B) should be routinely tested in patients with suspected MODY who do not have typical features of a genetic syndrome.

Maternal genetic variants associated with offspring birth weight and adult type 2 diabetes (T2D) risk loci show some overlap. Whether T2D genetic risk influences longitudinal fetal weight and the gestational timing when these relationships begin is unknown. We investigated the associations of T2D genetic risk scores (GRS) with longitudinal fetal weight and birth weight among 1,513 pregnant women from four ancestral groups. Women had up to five ultrasonography examinations. Ancestry-matched GRS were constructed separately using 380 European- (GRSeur), 104 African- (GRSafr), and 189 East Asian- (GRSeas) related T2D loci discovered in different population groups. Among European Americans, the highest quartile GRSeur was significantly associated with 53.8 g higher fetal weight (95% CI 19.2-88.5) over the pregnancy. The associations began at gestational week 24 and continued through week 40, with a 106.8 g (95% CI 6.5-207.1) increase in birth weight. The findings were similar in analysis further adjusted for maternal glucose challenge test results. No consistent association was found using ancestry-matched or cross-ancestry GRS in non-Europeans. In conclusion, T2D genetic susceptibility may influence fetal growth starting at midsecond trimester among Europeans. Absence of similar associations in non-Europeans urges the need for further genetic T2D studies in diverse ancestries.

Obesity is associated with adverse health outcomes, but the metabolic effects have not yet been fully elucidated. We aimed to investigate the association between adiposity and circulating metabolites and to address causality with Mendelian randomization (MR). Metabolomics data were generated with nontargeted ultraperformance liquid chromatography coupled to time-of-flight mass spectrometry in plasma and serum from three population-based Swedish cohorts: ULSAM (N = 1,135), PIVUS (N = 970), and TwinGene (N = 2,059). We assessed associations of general adiposity measured as BMI and central body fat distribution measured as waist-to-hip ratio adjusted for BMI (WHRadjBMI) with 210 annotated metabolites. We used MR analysis to assess causal effects. Lastly, we attempted to replicate the MR findings in the KORA and TwinsUK cohorts (N = 7,373), the CHARGE Consortium (N = 8,631), the Framingham Heart Study (N = 2,076), and the DIRECT Consortium (N = 3,029). BMI was associated with 77 metabolites, while WHRadjBMI was associated with 11 and 3 metabolites in women and men, respectively. The MR analyses in the Swedish cohorts suggested a causal association (P value <0.05) of increased general adiposity and reduced levels of arachidonic acid, dodecanedioic acid, and lysophosphatidylcholine (P-16:0) as well as with increased creatine levels. The results of the replication effort provided support for a causal association of adiposity with reduced levels of arachidonic acid (P value = 0.03). Adiposity is associated with variation of large parts of the circulating metabolome; however, further investigation of causality is required in well-powered cohorts.

One hundred years have passed since the discovery of insulin-an achievement that transformed diabetes from a fatal illness into a manageable chronic condition. The decades since that momentous achievement have brought ever more rapid innovation and advancement in diabetes research and clinical care. To celebrate the important work of the past century and help to chart a course for its continuation into the next, the Canadian Institutes of Health Research's Institute of Nutrition, Metabolism and Diabetes and the U.S. National Institutes of Health's National Institute of Diabetes and Digestive and Kidney Diseases recently held a joint international symposium, bringing together a cohort of researchers with diverse interests and backgrounds from both countries and beyond to discuss their collective quest to better understand the heterogeneity of diabetes and thus gain insights to inform new directions in diabetes treatment and prevention. This article summarizes the proceedings of that symposium, which spanned cutting-edge research into various aspects of islet biology, the heterogeneity of diabetic phenotypes, and the current state of and future prospects for precision medicine in diabetes.

One exercise session can elevate insulin-stimulated glucose uptake (ISGU) in skeletal muscle, but the mechanisms remain elusive. Circumstantial evidence suggests a role for Akt substrate of 160 kDa (AS160 or TBC1D4). We used genetic approaches to rigorously test this idea. The initial experiment evaluated the role of AS160 in postexercise increase in ISGU using muscles from male wild-type (WT) and AS160-knockout (KO) rats. The next experiment used AS160-KO rats with an adeno-associated virus (AAV) approach to determine if rescuing muscle AS160 deficiency could restore the ability of exercise to improve ISGU. The third experiment tested if eliminating the muscle GLUT4 deficit in AS160-KO rats via AAV-delivered GLUT4 would enable postexercise enhancement of ISGU. The final experiment used AS160-KO rats and AAV delivery of AS160 mutated to prevent phosphorylation of Ser588, Thr642, and Ser704 to evaluate their role in postexercise ISGU. We discovered the following: 1) AS160 expression was essential for postexercise increase in ISGU; 2) rescuing muscle AS160 expression of AS160-KO rats restored postexercise enhancement of ISGU; 3) restoring GLUT4 expression in AS160-KO muscle did not rescue the postexercise increase in ISGU; and 4) although AS160 phosphorylation on three key sites was not required for postexercise elevation in ISGU, it was essential for the full exercise effect.

Red blood cells (RBC) act as mediators of vascular injury in type 2 diabetes mellitus (T2DM). miR-210 plays a protective role in cardiovascular homeostasis and is decreased in whole blood of T2DM mice. We hypothesized that downregulation of RBC miR-210 induces endothelial dysfunction in T2DM. RBC were coincubated with arteries and endothelial cells ex vivo and transfused in vivo to identify the role of miR-210 and its target protein tyrosine phosphatase 1B (PTP1B) in endothelial dysfunction. RBC from patients with T2DM and diabetic rodents induced endothelial dysfunction ex vivo and in vivo. miR-210 levels were lower in human RBC from patients with T2DM (T2DM RBC) than in RBC from healthy subjects. Transfection of miR-210 in human T2DM RBC rescued endothelial function, whereas miR-210 inhibition in healthy subjects RBC or RBC from miR-210 knockout mice impaired endothelial function. Human T2DM RBC decreased miR-210 expression in endothelial cells. miR-210 expression in carotid artery plaques was lower in T2DM patients than in patients without diabetes. Endothelial dysfunction induced by downregulated RBC miR-210 involved PTP1B and reactive oxygen species. miR-210 mimic attenuated endothelial dysfunction induced by RBC via downregulating vascular PTP1B and oxidative stress in diabetic mice in vivo. These data reveal that the downregulation of RBC miR-210 is a novel mechanism driving the development of endothelial dysfunction in T2DM.

MicroRNAs (miRNAs) are part of deregulated insulin secretion in type 2 diabetes (T2D) development. Rodent models have suggested miR-200c to be involved, but the role and potential as therapeutic target of this miRNA in human islets are not clear. Here we report increased expression of miR-200c in islets from T2D as compared with nondiabetic (ND) donors and display results showing reduced glucose-stimulated insulin secretion in EndoC-βH1 cells overexpressing miR-200c. We identify transcription factor ETV5 as the top rank target of miR-200c in human islets using TargetScan in combination with Pearson correlation analysis of miR-200c and mRNA expression data from the same human donors. Among other targets were JAZF1, as earlier shown in miR-200 knockout mice. Accordingly, linear model analysis of ETV5 and JAZF1 gene expression showed reduced expression of both genes in islets from human T2D donors. Western blot analysis confirmed the reduced expression of ETV5 on the protein level in EndoC-βH1 cells overexpressing miR-200c, and luciferase assay validated ETV5 as a direct target of miR-200c. Finally, LNA knockdown of miR-200c increased glucose-stimulated insulin secretion in islets from T2D donors approximately threefold. Our data reveal a vital role of the miR-200c-ETV5 axis in β-cell dysfunction and pathophysiology of T2D.

Adipose-derived stem cells (ADSCs) can differentiate into vascular lineages and participate in vascular remodeling. Perivascular ADSCs (PV-ADSCs) draw attention because of their unique location. The heterogeneity of subcutaneous (SUB) and abdominal ADSCs were well addressed, but PV-ADSCs' heterogeneity has not been investigated. In this study, we applied single-cell analysis to compare SUB-ADSCs and PV-ADSCs regarding their subpopulations, functions, and cell fates. We uncovered four subpopulations of PV-ADSCs (Dpp4+, Col4a2+/Icam1+, Clec11a+/Cpe+, and Sult1e1+ cells), among which the Clec11a+ subpopulation potentially participated in and regulated PV-ADSC differentiation toward a smooth muscle cell (SMC) phenotype. Distinct characteristics between PV-ADSCs and SUB-ADSCs were revealed.

Fructosamine is a measure of short-term glycemic control, which has been suggested as a useful complement to glycated hemoglobin (HbA1c) for the diagnosis and monitoring of diabetes. To date, a single genome-wide association study (GWAS) including 8,951 U.S. White and 2,712 U.S. Black individuals without a diabetes diagnosis has been published. Results in Whites and Blacks yielded different association loci, near RCN3 and CNTN5, respectively. In this study, we performed a GWAS on 20,731 European-ancestry blood donors and meta-analyzed our results with previous data from U.S. White participants from the Atherosclerosis Risk in Communities (ARIC) study (Nmeta = 29,685). We identified a novel association near GCK (rs3757840, βmeta = 0.0062; minor allele frequency [MAF] = 0.49; Pmeta = 3.66 × 10-8) and confirmed the association near RCN3 (rs113886122, βmeta = 0.0134; MAF = 0.17; Pmeta = 5.71 × 10-18). Colocalization analysis with whole-blood expression quantitative trait loci data suggested FCGRT as the effector transcript at the RCN3 locus. We further showed that fructosamine has low heritability (h2 = 7.7%), has no significant genetic correlation with HbA1c and other glycemic traits in individuals without a diabetes diagnosis (P > 0.05), but has evidence of shared genetic etiology with some anthropometric traits (Bonferroni-corrected P < 0.0012). Our results broaden knowledge of the genetic architecture of fructosamine and prioritize FCGRT for downstream functional studies at the established RCN3 locus.

Leucine deprivation improves insulin sensitivity; however, whether and how this effect can be extended are unknown. We hypothesized that intermittent leucine deprivation (ILD) might produce a long-term effect on improved insulin sensitivity via the formation of metabolic memory. Consistently, seven ILD cycles of treatment (1-day leucine-deficient diet, 3-day control diet) in mice produced a long-lasting (after a control diet was resumed for 49 days) effect on improved whole-body and hepatic insulin sensitivity in mice, indicating the potential formation of metabolic memory. Furthermore, the effects of ILD depended on hepatic general control nondepressible 2 (GCN2) expression, as verified by gain- and loss-of-function experiments. Moreover, ILD increased Gcn2 expression by reducing its DNA methylation at two CpG promoter sites controlled by demethylase growth arrest and DNA damage inducible b. Finally, ILD also improved insulin sensitivity in insulin-resistant mice. Thus, ILD induces long-lasting improvements in insulin sensitivity by increasing hepatic Gcn2 expression via a reduction in its DNA methylation. These results provide novel insights into understanding of the link between leucine deprivation and insulin sensitivity, as well as potential nutritional intervention strategies for treating insulin resistance and related diseases. We also provide evidence for liver-specific metabolic memory after ILD and novel epigenetic mechanisms for Gcn2 regulation.

To evaluate the effect of liraglutide, a glucagon-like peptide 1 receptor agonist, on pulmonary function and serum levels of surfactant protein D (SP-D) in type 2 diabetes. A double-blind, randomized, crossover, placebo-controlled clinical trial comprising 76 patients with a baseline forced expiratory volume in 1 s <90% of that predicted. Liraglutide was administered for 7 weeks (2 weeks of titration plus 5 weeks at 1.8 mg daily). This short duration was intentional to minimize weight loss as a potential confounding factor. Serum level of SP-D was used as a biomarker of alveolar-capillary barrier integrity. Liraglutide exerted a positive impact on forced vital capacity (FVC) in comparison with placebo (ΔFVC 5.2% of predicted [from 0.8 to 9.6]; P = 0.009). No differences in the other pulmonary variables were observed. Participants under liraglutide treatment also experienced a decrease in serum SP-D (P = 0.038). The absolute change in FVC correlated with final serum SP-D in participants receiving liraglutide (r = -0.313, P = 0.036). Stepwise multivariate regression analysis showed that final serum SP-D independently predicted changes in FVC. In conclusion, liraglutide increased FVC in patients with type 2 diabetes. This effect was associated with a significant decrease of circulating SP-D, thus pointing to a beneficial effect in the alveolar-capillary function.

A disparate array of plasma/serum markers provides evidence for chronic inflammation in human prediabetes, a condition that is most closely replicated by standard mouse models of obesity and metaflammation. These remain largely nonactionable and contrast with our rich understanding of inflammation in human type 2 diabetes. New data show that inflammatory profiles produced by CD4+ T cells define human prediabetes as a unique inflammatory state. Regulatory T cells (Treg) control mitochondrial function and cytokine production by CD4+ effector T cells (Teff) in prediabetes and type 2 diabetes by supporting T helper (Th)17 or Th1 cytokine production, respectively. These data suggest that Treg control of Teff metabolism regulates inflammation differentially in prediabetes compared with type 2 diabetes. Queries of genes that impact mitochondrial function or pathways leading to transcription of lipid metabolism genes identified the fatty acid importer CD36 as highly expressed in Treg but not Teff from subjects with prediabetes. Pharmacological blockade of CD36 in Treg from subjects with prediabetes decreased Teff production of the Th17 cytokines that differentiate overall prediabetes inflammation. We conclude that Treg control CD4+ T cell cytokine profiles through mechanisms determined, at least in part, by host metabolic status. Furthermore, Treg CD36 uniquely promotes Th17 cytokine production by Teff in prediabetes.

The increasing prevalence of obesity has resulted in demands for the development of new effective strategies for obesity treatment. Withaferin A (WA) shows a great potential for prevention of obesity by sensitizing leptin signaling in the hypothalamus. However, the mechanism underlying the weight- and adiposity-reducing effects of WA remains to be elucidated. In this study, we report that WA treatment induced white adipose tissue (WAT) browning, elevated energy expenditure, decreased respiratory exchange ratio, and prevented high-fat diet-induced obesity. The sympathetic chemical denervation dampened the WAT browning and also impeded the reduction of adiposity in WA-treated mice. WA markedly upregulated the levels of Prdm16 and FATP1 (Slc27a1) in the inguinal WAT (iWAT), and this was blocked by sympathetic denervation. Prdm16 or FATP1 knockdown in iWAT abrogated the WAT browning-inducing effects of WA and restored the weight gain and adiposity in WA-treated mice. Together, these findings suggest that WA induces WAT browning through the sympathetic nerve-adipose axis, and the adipocytic Prdm16-FATP1 pathway mediates the promotive effects of WA on white adipose browning.

Macro- and microvascular complications of type 2 diabetes (T2D), obesity, and dyslipidemia share common metabolic pathways. In this study, using a total of 1,300 metabolites from 996 Qatari adults (57% with T2D) and 1,159 metabolites from an independent cohort of 2,618 individuals from the Qatar BioBank (11% with T2D), we identified 373 metabolites associated with T2D, obesity, retinopathy, dyslipidemia, and lipoprotein levels, 161 of which were novel. Novel metabolites included phospholipids, sphingolipids, lysolipids, fatty acids, dipeptides, and metabolites of the urea cycle and xanthine, steroid, and glutathione metabolism. The identified metabolites enrich pathways of oxidative stress, lipotoxicity, glucotoxicity, and proteolysis. Second, we identified 15 patterns we defined as "metabo-clinical signatures." These are clusters of patients with T2D who group together based on metabolite levels and reveal the same clustering in two or more clinical variables (obesity, LDL, HDL, triglycerides, and retinopathy). These signatures revealed metabolic pathways associated with different clinical patterns and identified patients with extreme (very high/low) clinical variables associated with extreme metabolite levels in specific pathways. Among our novel findings are the role of N-acetylmethionine in retinopathy in conjunction with dyslipidemia and the possible roles of N-acetylvaline and pyroglutamine in association with high cholesterol levels and kidney function.

In my lecture given on the occasion of the 2021 Banting Medal for Scientific Achievement, I briefly described the history of the incretin effect and summarized some of the developments leading to current therapies of obesity and diabetes based on the incretin hormones, glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP). In the text below, I discuss in further detail the role of these two hormones for postprandial insulin secretion in humans on the basis of recent studies with antagonists. Their direct and indirect actions on the β-cells are discussed next as well as their contrasting actions on glucagon secretion. After a brief discussion of their effect on insulin sensitivity, I describe their immediate actions in patients with type 2 diabetes and emphasize the actions of GLP-1 on β-cell glucose sensitivity, followed by a discussion of their extrapancreatic actions, including effects on appetite and food intake in humans. Finally, possible mechanisms of action of GIP-GLP-1 coagonists are discussed, and it is concluded that therapies based on incretin actions are likely to change the current hesitant therapy of both obesity and diabetes.

Adipose tissues are not homogeneous and show site-specific properties. An elusive and understudied adipose tissue depot, most likely due to its limited accessibility, is the intermuscular adipose tissue (IMAT) depot. Adipose tissue is a pliable organ with the ability to adapt to its physiological context, yet whether that adaptation is harmful or beneficial in the IMAT depot remains to be explored in humans. Potential reasons for IMAT accumulation in humans being deleterious or beneficial include 1) sex and related circulating hormone levels, 2) race and ethnicity, and 3) lifestyle factors (e.g., diet and physical activity level). IMAT quantity per se may not be the driving factor in the etiology of insulin resistance and type 2 diabetes, but rather the quality of the IMAT itself is the true puppeteer. Adipose tissue quality likely influences its secreted factors, which are also likely to influence metabolism of surrounding tissues. The advent of molecular assessments such as transcriptome sequencing (RNAseq), assay for transposase-accessible chromatin using sequencing (ATACseq), and DNA methylation at the single-cell and single-nucleus levels, as well as the potential for ultrasound-guided biopsies specifically for IMAT, will permit more sophisticated investigations of human IMAT and dramatically advance our understanding of this enigmatic adipose tissue.