Metabolic and Metabo-Clinical Signatures of Type 2 Diabetes, Obesity, Retinopathy, and Dyslipidemia.
Yousri Noha A,Suhre Karsten,Yassin Esraa,Al-Shakaki Alya,Robay Amal,Elshafei Maha,Chidiac Omar,Hunt Steven C,Crystal Ronald G,Fakhro Khalid A
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.
An Expanded Genome-Wide Association Study of Fructosamine Levels Identifies RCN3 as a Replicating Locus and Implicates FCGRT as the Effector Transcript.
Riveros-Mckay Fernando,Roberts David,Di Angelantonio Emanuele,Yu Bing,Soranzo Nicole,Danesh John,Selvin Elizabeth,Butterworth Adam S,Barroso Inês
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.
MG53 E3 Ligase-Dead Mutant Protects Diabetic Hearts From Acute Ischemic/Reperfusion Injury and Ameliorates Diet-Induced Cardiometabolic Damage.
Feng Han,Shen Hao,Robeson Matthew J,Wu Yue-Han,Wu Hong-Kun,Chen Geng-Jia,Zhang Shuo,Xie Peng,Jin Li,He Yanyun,Wang Yingfan,Lv Fengxiang,Hu Xinli,Zhang Yan,Xiao Rui-Ping
Cardiometabolic diseases, including diabetes and its cardiovascular complications, are the global leading causes of death, highlighting a major unmet medical need. Over the past decade, mitsugumin 53 (MG53), also called TRIM72, has emerged as a powerful agent for myocardial membrane repair and cardioprotection, but its therapeutic value is complicated by its E3 ligase activity, which mediates metabolic disorders. Here, we show that an E3 ligase-dead mutant, MG53-C14A, retains its cardioprotective function without causing metabolic adverse effects. When administered in normal animals, both the recombinant human wild-type MG53 protein (rhMG53-WT) and its E3 ligase-dead mutant (rhMG53-C14A) protected the heart equally from myocardial infarction and ischemia/reperfusion (I/R) injury. However, in diabetic db/db mice, rhMG53-WT treatment markedly aggravated hyperglycemia, cardiac I/R injury, and mortality, whereas acute and chronic treatment with rhMG53-C14A still effectively ameliorated I/R-induced myocardial injury and mortality or diabetic cardiomyopathy, respectively, without metabolic adverse effects. Furthermore, knock-in of MG53-C14A protected the mice from high-fat diet-induced metabolic disorders and cardiac damage. Thus, the E3 ligase-dead mutant MG53-C14A not only protects the heart from acute myocardial injury but also counteracts metabolic stress, providing a potentially important therapy for the treatment of acute myocardial injury in metabolic disorders, including diabetes and obesity.
Downregulation of Erythrocyte miR-210 Induces Endothelial Dysfunction in Type 2 Diabetes.
Zhou Zhichao,Collado Aida,Sun Changyan,Tratsiakovich Yahor,Mahdi Ali,Winter Hanna,Chernogubova Ekaterina,Seime Till,Narayanan Sampath,Jiao Tong,Jin Hong,Alvarsson Michael,Zheng Xiaowei,Yang Jiangning,Hedin Ulf,Catrina Sergiu-Bogdan,Maegdefessel Lars,Pernow John
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.
Effect of General Adiposity and Central Body Fat Distribution on the Circulating Metabolome: A Multicohort Nontargeted Metabolomics Observational and Mendelian Randomization Study.
Ahmad Shafqat,Hammar Ulf,Kennedy Beatrice,Salihovic Samira,Ganna Andrea,Lind Lars,Sundström Johan,Ärnlöv Johan,Berne Christian,Risérus Ulf,Magnusson Patrik K E,Larsson Susanna C,Fall Tove
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.
Genetic Control of Splicing at SIRPG Modulates Risk of Type 1 Diabetes.
Smith Morgan J,Pastor Lucia,Newman Jeremy R B,Concannon Patrick
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.
Regulatory T Cells Control Effector T Cell Inflammation in Human Prediabetes.
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.
Human Islet MicroRNA-200c Is Elevated in Type 2 Diabetes and Targets the Transcription Factor ETV5 to Reduce Insulin Secretion.
Ofori Jones K,Karagiannopoulos Alexandros,Nagao Mototsugu,Westholm Efraim,Ramadan Shaima,Wendt Anna,Esguerra Jonathan L S,Eliasson Lena
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.
Obesity-Induced miR-455 Upregulation Promotes Adaptive Pancreatic β-Cell Proliferation Through the CPEB1/CDKN1B Pathway.
Hu Qianxing,Mu Jinming,Liu Yuhong,Yang Yue,Liu Yue,Pan Yi,Zhang Yanfeng,Li Ling,Liu Dechen,Chen Jianqiu,Zhang Fangfang,Jin Liang
Pancreatic β-cells adapt to compensate for increased metabolic demand during obesity. Although the miRNA pathway has an essential role in β-cell expansion, whether it is involved in adaptive proliferation is largely unknown. First, we report that EGR2 binding to the miR-455 promoter induced miR-455 upregulation in the pancreatic islets of obesity mouse models. Then, in vitro gain- or loss-of-function studies showed that miR-455 overexpression facilitated β-cell proliferation. Knockdown of miR-455 in ob/ob mice via pancreatic intraductal infusion prevented compensatory β-cell expansion. Mechanistically, our results revealed that increased miR-455 expression inhibits the expression of its target cytoplasmic polyadenylation element binding protein 1 (CPEB1), an mRNA binding protein that plays an important role in regulating insulin resistance and cell proliferation. Decreased CPEB1 expression inhibits elongation of the poly(A) tail and the subsequent translation of Cdkn1b mRNA, reducing the CDKN1B expression level and finally promoting β-cell proliferation. Taken together, our results show that the miR-455/CPEB1/CDKN1B pathway contributes to adaptive proliferation of β-cells to meet metabolic demand during obesity.
Recessive Genome-Wide Meta-analysis Illuminates Genetic Architecture of Type 2 Diabetes.
Most genome-wide association studies (GWAS) of complex traits are performed using models with additive allelic effects. Hundreds of loci associated with type 2 diabetes have been identified using this approach. Additive models, however, can miss loci with recessive effects, thereby leaving potentially important genes undiscovered. We conducted the largest GWAS meta-analysis using a recessive model for type 2 diabetes. Our discovery sample included 33,139 case subjects and 279,507 control subjects from 7 European-ancestry cohorts, including the UK Biobank. We identified 51 loci associated with type 2 diabetes, including five variants undetected by prior additive analyses. Two of the five variants had minor allele frequency of <5% and were each associated with more than a doubled risk in homozygous carriers. Using two additional cohorts, FinnGen and a Danish cohort, we replicated three of the variants, including one of the low-frequency variants, rs115018790, which had an odds ratio in homozygous carriers of 2.56 (95% CI 2.05-3.19; P = 1 × 10-16) and a stronger effect in men than in women (for interaction, P = 7 × 10-7). The signal was associated with multiple diabetes-related traits, with homozygous carriers showing a 10% decrease in LDL cholesterol and a 20% increase in triglycerides; colocalization analysis linked this signal to reduced expression of the nearby PELO gene. These results demonstrate that recessive models, when compared with GWAS using the additive approach, can identify novel loci, including large-effect variants with pathophysiological consequences relevant to type 2 diabetes.
Monocyte miRNAs Are Associated With Type 2 Diabetes.
miRNAs are small noncoding RNAs that may contribute to common diseases through epigenetic regulation of gene expression. Little is known regarding the role of miRNAs in type 2 diabetes (T2D). We performed miRNA sequencing and transcriptomic profiling of peripheral monocytes from the longitudinal Multi-Ethnic Study of Atherosclerosis (MESA) (N = 1,154). We examined associations between miRNAs and prevalent impaired fasting glucose and T2D and evaluated the T2D-associated miRNA effect on incident T2D. Of 774 detected miRNAs, 6 (miR-22-3p, miR-33a-5p, miR-181c-5p, miR-92b-3p, miR-222-3p, and miR-944) were associated with prevalent T2D. For five of the six miRNAs (all but miR-222-3p), our findings suggest a dose-response relationship with impaired fasting glucose and T2D. Two of the six miRNAs were associated with incident T2D (miR-92b-3p: hazard ratio [HR] 1.64, P = 1.30E-03; miR-222-3p: HR 1.97, P = 9.10E-03) in the highest versus lowest tertile of expression. Most of the T2D-associated miRNAs were also associated with HDL cholesterol concentrations. The genes targeted by these miRNAs belong to key nodes of a cholesterol metabolism transcriptomic network. Higher levels of miRNA expression expected to increase intracellular cholesterol accumulation in monocytes are linked to an increase in T2D risk.
Methazolamide Attenuates the Development of Diabetic Cardiomyopathy by Promoting β-Catenin Degradation in Type 1 Diabetic Mice.
Methazolamide (MTZ), a carbonic anhydrase inhibitor, has been shown to inhibit cardiomyocyte hypertrophy and exert a hypoglycemic effect in patients with type 2 diabetes and diabetic db/db mice. However, whether MTZ has a cardioprotective effect in the setting of diabetic cardiomyopathy is not clear. We investigated the effects of MTZ in a mouse model of streptozotocin-induced type 1 diabetes mellitus (T1DM). Diabetic mice received MTZ by intragastric gavage (10, 25, or 50 mg/kg, daily for 16 weeks). In the diabetic group, MTZ significantly reduced both random and fasting blood glucose levels and improved glucose tolerance in a dose-dependent manner. MTZ ameliorated T1DM-induced changes in cardiac morphology and dysfunction. Mechanistic analysis revealed that MTZ blunted T1DM-induced enhanced expression of β-catenin. Similar results were observed in neonatal rat cardiomyocytes (NRCMs) and adult mouse cardiomyocytes treated with high glucose or Wnt3a (a β-catenin activator). There was no significant change in β-catenin mRNA levels in cardiac tissues or NRCMs. MTZ-mediated β-catenin downregulation was recovered by MG132, a proteasome inhibitor. Immunoprecipitation and immunofluorescence analyses showed augmentation of AXIN1-β-catenin interaction by MTZ in T1DM hearts and in NRCMs treated with Wnt3a; thus, MTZ may potentiate AXIN1-β-catenin linkage to increase β-catenin degradation. Overall, MTZ may alleviate cardiac hypertrophy by mediating AXIN1-β-catenin interaction to promote degradation and inhibition of β-catenin activity. These findings may help inform novel therapeutic strategy to prevent heart failure in patients with diabetes.
Single-Cell Transcriptomics Reveals Novel Role of Microglia in Fibrovascular Membrane of Proliferative Diabetic Retinopathy.
Vitreous fibrovascular membranes (FVMs), the hallmark of proliferative diabetic retinopathy (PDR), cause retinal hemorrhage, detachment, and eventually blindness. However, little is known about the pathophysiology of FVM. In this study, we used single-cell RNA sequencing on surgically harvested PDR-FVMs and generated a comprehensive cell atlas of FVM. Eight cellular compositions were identified, with microglia as the major cell population. We identified a GPNMB+ subpopulation of microglia, which presented both profibrotic and fibrogenic properties. Pseudotime analysis further revealed the profibrotic microglia was uniquely differentiated from retina-resident microglia and expanded in the PDR setting. Ligand-receptor interactions between the profibrotic microglia and cytokines upregulated in PDR vitreous implicated the involvement of several pathways, including CCR5, IFNGR1, and CD44 signaling, in the microglial activation within the PDR microenvironment. Collectively, our description of the novel microglia phenotypes in PDR-FVM may offer new insight into the cellular and molecular mechanism underlying the pathogenesis of DR, as well as potential signaling pathways amenable to disease-specific intervention.
pdx1 Knockout Leads to a Diabetic Nephropathy- Like Phenotype in Zebrafish and Identifies Phosphatidylethanolamine as Metabolite Promoting Early Diabetic Kidney Damage.
The pdx1-/- zebrafish mutant was recently established as a novel animal model of diabetic retinopathy. In this study, we investigate whether knockout of pdx1 also leads to diabetic kidney disease (DKD). pdx1-/- larvae exhibit several signs of early DKD, such as glomerular hypertrophy, impairments in the filtration barrier corresponding to microalbuminuria, and glomerular basement membrane (GBM) thickening. Adult pdx1-/- mutants show progressive GBM thickening in comparison with the larval state. Heterozygous pdx1 knockout also leads to glomerular hypertrophy as initial establishment of DKD similar to the pdx1-/- larvae. RNA sequencing of adult pdx1+/- kidneys uncovered regulations in multiple expected diabetic pathways related to podocyte disruption and hinting at early vascular dysregulation without obvious morphological alterations. Metabolome analysis and pharmacological intervention experiments revealed the contribution of phosphatidylethanolamine in the early establishment of kidney damage. In conclusion, this study identified the pdx1 mutant as a novel model for the study of DKD, showing signs of the early disease progression already in the larval stage and several selective features of later DKD in adult mutants.
PACS-2 Ameliorates Tubular Injury by Facilitating Endoplasmic Reticulum-Mitochondria Contact and Mitophagy in Diabetic Nephropathy.
Mitochondria-associated endoplasmic reticulum membrane (MAM) may have a role in tubular injury in diabetic nephropathy (DN), but the precise mechanism remains unclear. Here, we demonstrate that the expression of phosphofurin acidic cluster sorting protein 2 (PACS-2), a critical regulator of MAM formation, is significantly decreased in renal tubules of patients with DN, and PACS-2 expression is positively correlated with renal function and negatively correlated with degrees of tubulointerstitial lesions. Conditional deletion of Pacs-2 in proximal tubules (PTs) aggravates albuminuria and tubular injury in a streptozotocin-induced mouse model of diabetes. Mitochondrial fragmentation, MAM disruption, and defective mitophagy accompanied by altered expression of mitochondrial dynamics and mitophagic proteins, including Drp1 and Becn1, are observed in tubules of diabetic mice; these changes are more pronounced in PT-specific Pacs-2 knockout mice. In vitro, overexpression of PACS-2 in HK-2 cells alleviates excessive mitochondrial fission induced by high glucose concentrations through blocking mitochondrial recruitment of DRP1 and subsequently restores MAM integrity and enhances mitophagy. Mechanistically, PACS-2 binds to BECN1 and mediates the relocalization of BECN1 to MAM, where it promotes the formation of mitophagosome. Together, these data highlight an important but previously unrecognized role of PACS-2 in ameliorating tubular injury in DN by facilitating MAM formation and mitophagy.
Restoration of Autophagic Flux Improves Endothelial Function in Diabetes Through Lowering Mitochondrial ROS-Mediated eNOS Monomerization.
Endothelial nitric oxide synthase (eNOS) monomerization and uncoupling play crucial roles in mediating vascular dysfunction in diabetes, although the underlying mechanisms are still incompletely understood. Increasing evidence indicates that autophagic dysregulation is involved in the pathogenesis of diabetic endothelial dysfunction; however, whether autophagy regulates eNOS activity through controlling eNOS monomerization or dimerization remains elusive. In this study, autophagic flux was impaired in the endothelium of diabetic db/db mice and in human endothelial cells exposed to advanced glycation end products or oxidized low-density lipoprotein. Inhibition of autophagic flux by chloroquine or bafilomycin A1 were sufficient to induce eNOS monomerization and lower nitric oxide bioavailability by increasing mitochondrial reactive oxygen species (mtROS). Restoration of autophagic flux by overexpressing transcription factor EB (TFEB), a master regulator of autophagy and lysosomal biogenesis, decreased endothelial cell oxidative stress, increased eNOS dimerization, and improved endothelium-dependent relaxations (EDRs) in db/db mouse aortas. Inhibition of mammalian target of rapamycin kinase (mTOR) increased TFEB nuclear localization, reduced mtROS accumulation, facilitated eNOS dimerization, and enhanced EDR in db/db mice. Moreover, calorie restriction also increased TFEB expression, improved autophagic flux, and restored EDR in the aortas of db/db mice. Taken together, the findings of this study reveal that mtROS-induced eNOS monomerization is closely associated with the impaired TFEB-autophagic flux axis leading to endothelial dysfunction in diabetic mice.
Müller Glial Expression of REDD1 Is Required for Retinal Neurodegeneration and Visual Dysfunction in Diabetic Mice.
Clinical studies support a role for the protein regulated in development and DNA damage response 1 (REDD1) in ischemic retinal complications. To better understand how REDD1 contributes to retinal pathology, we examined human single-cell sequencing data sets and found specificity of REDD1 expression that was consistent with markers of retinal Müller glia. Thus, we investigated the hypothesis that REDD1 expression specifically in Müller glia contributes to diabetes-induced retinal pathology. The retina of Müller glia-specific REDD1 knockout (REDD1-mgKO) mice exhibited dramatic attenuation of REDD1 transcript and protein expression. In the retina of streptozotocin-induced diabetic control mice, REDD1 protein expression was enhanced coincident with an increase in oxidative stress. In the retina of diabetic REDD1-mgKO mice, there was no increase in REDD1 protein expression, and oxidative stress was reduced compared with diabetic control mice. In both Müller glia within the retina of diabetic mice and human Müller cell cultures exposed to hyperglycemic conditions, REDD1 was necessary for increased expression of the gliosis marker glial fibrillary acidic protein. The effect of REDD1 deletion in preventing gliosis was associated with suppression of oxidative stress and required the antioxidant transcription factor nuclear factor erythroid-2-related factor 2 (Nrf2). In contrast to diabetic control mice, diabetic REDD1-mgKO mice did not exhibit retinal thinning, increased markers of neurodegeneration within the retinal ganglion cell layer, or deficits in visual function. Overall, the findings support a key role for Müller glial REDD1 in the failed adaptive response of the retina to diabetes that includes gliosis, neurodegeneration, and impaired vision.
Nrf2 Regulates β-Cell Mass by Suppressing β-Cell Death and Promoting β-Cell Proliferation.
Finding therapies that can protect and expand functional β-cell mass is a major goal of diabetes research. Here, we generated β-cell-specific conditional knockout and gain-of-function mouse models and used human islet transplant experiments to examine how manipulating Nrf2 levels affects β-cell survival, proliferation, and mass. Depletion of Nrf2 in β-cells results in decreased glucose-stimulated β-cell proliferation ex vivo and decreased adaptive β-cell proliferation and β-cell mass expansion after a high-fat diet in vivo. Nrf2 protects β-cells from apoptosis after a high-fat diet. Nrf2 loss of function decreases Pdx1 abundance and insulin content. Activating Nrf2 in a β-cell-specific manner increases β-cell proliferation and mass and improves glucose tolerance. Human islets transplanted under the kidney capsule of immunocompromised mice and treated systemically with bardoxolone methyl, an Nrf2 activator, display increased β-cell proliferation. Thus, by managing reactive oxygen species levels, Nrf2 regulates β-cell mass and is an exciting therapeutic target for expanding and protecting β-cell mass in diabetes.
Endothelial Phospholipase Cγ2 Improves Outcomes of Diabetic Ischemic Limb Rescue Following VEGF Therapy.
Therapeutic vascular endothelial growth factor (VEGF) replenishment has met with limited success for the management of critical limb-threatening ischemia. To improve outcomes of VEGF therapy, we applied single-cell RNA sequencing (scRNA-seq) technology to study the endothelial cells of the human diabetic skin. Single-cell suspensions were generated from the human skin followed by cDNA preparation using the Chromium Next GEM Single-cell 3' Kit v3.1. Using appropriate quality control measures, 36,487 cells were chosen for downstream analysis. scRNA-seq studies identified that although VEGF signaling was not significantly altered in diabetic versus nondiabetic skin, phospholipase Cγ2 (PLCγ2) was downregulated. The significance of PLCγ2 in VEGF-mediated increase in endothelial cell metabolism and function was assessed in cultured human microvascular endothelial cells. In these cells, VEGF enhanced mitochondrial function, as indicated by elevation in oxygen consumption rate and extracellular acidification rate. The VEGF-dependent increase in cell metabolism was blunted in response to PLCγ2 inhibition. Follow-up rescue studies therefore focused on understanding the significance of VEGF therapy in presence or absence of endothelial PLCγ2 in type 1 (streptozotocin-injected) and type 2 (db/db) diabetic ischemic tissue. Nonviral topical tissue nanotransfection technology (TNT) delivery of CDH5 promoter-driven PLCγ2 open reading frame promoted the rescue of hindlimb ischemia in diabetic mice. Improvement of blood flow was also associated with higher abundance of VWF+/CD31+ and VWF+/SMA+ immunohistochemical staining. TNT-based gene delivery was not associated with tissue edema, a commonly noted complication associated with proangiogenic gene therapies. Taken together, our study demonstrates that TNT-mediated delivery of endothelial PLCγ2, as part of combination gene therapy, is effective in diabetic ischemic limb rescue.
Independent of Renox, NOX5 Promotes Renal Inflammation and Fibrosis in Diabetes by Activating ROS-Sensitive Pathways.
Excessive production of renal reactive oxygen species (ROS) plays a major role in diabetic kidney disease (DKD). Here, we provide key findings demonstrating the predominant pathological role of the pro-oxidant enzyme NADPH oxidase 5 (NOX5) in DKD, independent of the previously characterized NOX4 pathway. In patients with diabetes, we found increased expression of renal NOX5 in association with enhanced ROS formation and upregulation of ROS-sensitive factors early growth response 1 (EGR-1), protein kinase C-α (PKC-α), and a key metabolic gene involved in redox balance, thioredoxin-interacting protein (TXNIP). In preclinical models of DKD, overexpression of NOX5 in Nox4-deficient mice enhances kidney damage by increasing albuminuria and augmenting renal fibrosis and inflammation via enhanced ROS formation and the modulation of EGR1, TXNIP, ERK1/2, PKC-α, and PKC-ε. In addition, the only first-in-class NOX inhibitor, GKT137831, appears to be ineffective in the presence of NOX5 expression in diabetes. In vitro, silencing of NOX5 in human mesangial cells attenuated upregulation of EGR1, PKC-α, and TXNIP induced by high glucose levels, as well as markers of inflammation (TLR4 and MCP-1) and fibrosis (CTGF and collagens I and III) via reduction in ROS formation. Collectively, these findings identify NOX5 as a superior target in human DKD compared with other NOX isoforms such as NOX4, which may have been overinterpreted in previous rodent studies.
Genome-Wide Association Study Identifies Genetic Loci Associated With Fat Cell Number and Overlap With Genetic Risk Loci for Type 2 Diabetes.
Interindividual differences in generation of new fat cells determine body fat and type 2 diabetes risk. In the GENetics of Adipocyte Lipolysis (GENiAL) cohort, which consists of participants who have undergone abdominal adipose biopsy, we performed a genome-wide association study (GWAS) of fat cell number (n = 896). Candidate genes from the genetic study were knocked down by siRNA in human adipose-derived stem cells. We report 318 single nucleotide polymorphisms (SNPs) and 17 genetic loci displaying suggestive (P < 1 × 10-5) association with fat cell number. Two loci pass threshold for GWAS significance, on chromosomes 2 (lead SNP rs149660479-G) and 7 (rs147389390-deletion). We filtered for fat cell number-associated SNPs (P < 1.00 × 10-5) using evidence of genotype-specific expression. Where this was observed we selected genes for follow-up investigation and hereby identified SPATS2L and KCTD18 as regulators of cell proliferation consistent with the genetic data. Furthermore, 30 reported type 2 diabetes-associated SNPs displayed nominal and consistent associations with fat cell number. In functional follow-up of candidate genes, RPL8, HSD17B12, and PEPD were identified as displaying effects on cell proliferation consistent with genetic association and gene expression findings. In conclusion, findings presented herein identify SPATS2L, KCTD18, RPL8, HSD17B12, and PEPD of potential importance in controlling fat cell numbers (plasticity), the size of body fat, and diabetes risk.
The Role of Long Noncoding RNA MALAT1 in Diabetic Polyneuropathy and the Impact of Its Silencing in the Dorsal Root Ganglion by a DNA/RNA Heteroduplex Oligonucleotide.
Diabetic polyneuropathy (DPN) is the most common complication of diabetes, yet its pathophysiology has not been established. Accumulating evidence suggests that long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) plays pivotal roles in the regulation of cell growth and survival during diabetic complications. This study aimed to investigate the impact of MALAT1 silencing in dorsal root ganglion (DRG) sensory neurons, using an α-tocopherol-conjugated DNA/RNA heteroduplex oligonucleotide (Toc-HDO), on the peripheral nervous system of diabetic mice. We identified MALAT1 upregulation in the DRG of chronic diabetic mice that suggested either a pathological change or one that might be protective, and systemic intravenous injection of Toc-HDO effectively inhibited its gene expression. However, we unexpectedly noted that this intervention paradoxically exacerbated disease with increased thermal and mechanical nociceptive thresholds, indicating further sensory loss, greater sciatic-tibial nerve conduction slowing, and additional declines of intraepidermal nerve fiber density in the hind paw footpads. Serine/arginine-rich splicing factors, which are involved in pre-mRNA splicing by interacting with MALAT1, reside in nuclear speckles in wild-type and diabetic DRG neurons; MALAT1 silencing was associated with their disruption. The findings provide evidence for an important role that MALAT1 plays in DPN, suggesting neuroprotection and regulation of pre-mRNA splicing in nuclear speckles. This is also the first example in which a systemically delivered nucleotide therapy had a direct impact on DRG diabetic neurons and their axons.
lncRNA-H19 in Fibroblasts Promotes Wound Healing in Diabetes.
Cutaneous wound healing in diabetes is impaired and would develop into nonhealing ulcerations. However, the molecular mechanism underlying the wound-healing process remains largely obscure. Here, we found that cutaneous PDGFRα+ fibroblast-expressing lncRNA-H19 (lncH19) accelerates the wound-healing process via promoting dermal fibroblast proliferation and macrophage infiltration in injured skin. PDGFRα+ cell-derived lncH19, which is lower in contents in the wound-healing cutaneous tissue of patients and mice with type 2 diabetes, is required for wound healing through promoting proliferative capacity of dermis fibroblasts as well as macrophage recruitments. Mechanistically, lncH19 relieves the cell cycle arrest of fibroblasts and increases macrophage infiltration in injured tissues via inhibiting p53 activity and GDF15 releasement. Furthermore, exosomes derived from adipocyte progenitor cells efficiently restore the impaired diabetic wound healing via delivering lncH19 to injured tissue. Therefore, our study reveals a new role for lncRNA in regulating cutaneous tissue repair and provides a novel promising insight for developing clinical treatment of diabetes.
Glucose-Dependent miR-125b Is a Negative Regulator of β-Cell Function.
Impaired pancreatic β-cell function and insulin secretion are hallmarks of type 2 diabetes. miRNAs are short, noncoding RNAs that silence gene expression vital for the development and function of β cells. We have previously shown that β cell-specific deletion of the important energy sensor AMP-activated protein kinase (AMPK) results in increased miR-125b-5p levels. Nevertheless, the function of this miRNA in β cells is unclear. We hypothesized that miR-125b-5p expression is regulated by glucose and that this miRNA mediates some of the deleterious effects of hyperglycemia in β cells. Here, we show that islet miR-125b-5p expression is upregulated by glucose in an AMPK-dependent manner and that short-term miR-125b-5p overexpression impairs glucose-stimulated insulin secretion (GSIS) in the mouse insulinoma MIN6 cells and in human islets. An unbiased, high-throughput screen in MIN6 cells identified multiple miR-125b-5p targets, including the transporter of lysosomal hydrolases M6pr and the mitochondrial fission regulator Mtfp1. Inactivation of miR-125b-5p in the human β-cell line EndoCβ-H1 shortened mitochondria and enhanced GSIS, whereas mice overexpressing miR-125b-5p selectively in β cells (MIR125B-Tg) were hyperglycemic and glucose intolerant. MIR125B-Tg β cells contained enlarged lysosomal structures and had reduced insulin content and secretion. Collectively, we identify miR-125b as a glucose-controlled regulator of organelle dynamics that modulates insulin secretion.
Identification of Novel Disease-Relevant Genes and Pathways in the Pathogenesis of Type 1 Diabetes: A Potential Defect in Pancreatic Iron Homeostasis.
Multiple pathways contribute to the pathophysiological development of type 1 diabetes (T1D); however, the exact mechanisms involved are unclear. We performed differential gene expression analysis in pancreatic islets of NOD mice versus age-matched congenic NOD.B10 controls to identify genes that may contribute to disease pathogenesis. Novel genes related to extracellular matrix development and glucagon and insulin signaling/secretion were changed in NOD mice during early inflammation. During "respective" insulitis, the expression of genes encoding multiple chemosensory olfactory receptors were upregulated, and during "destructive" insulitis, the expression of genes involved in antimicrobial defense and iron homeostasis were downregulated. Islet inflammation reduced the expression of Hamp that encodes hepcidin. Hepcidin is expressed in β-cells and serves as the key regulator of iron homeostasis. We showed that Hamp and hepcidin levels were lower, while iron levels were higher in the pancreas of 12-week-old NOD versus NOD.B10 mice, suggesting that a loss of iron homeostasis may occur in the islets during the onset of "destructive" insulitis. Interestingly, we showed that the severity of NOD disease correlates with dietary iron intake. NOD mice maintained on low-iron diets had a lower incidence of hyperglycemia, while those maintained on high-iron diets had an earlier onset and higher incidence of disease, suggesting that high iron exposure combined with a loss of pancreatic iron homeostasis may exacerbate NOD disease. This mechanism may explain the link seen between high iron exposure and the increased risk for T1D in humans.
Topical Aspirin Administration Improves Cutaneous Wound Healing in Diabetic Mice Through a Phenotypic Switch of Wound Macrophages Toward an Anti-inflammatory and Proresolutive Profile Characterized by LXA4 Release.
Patients with diabetes present a persistent inflammatory process, leading to impaired wound healing. Since nonhealing diabetic wound management shows limited results, the introduction of advanced therapies targeting and correcting the inflammatory status of macrophages in chronic wounds could be an effective therapeutic strategy to stop the sustained inflammation and to return to a healing state. In an excisional skin injury in a diet-induced diabetic murine model, we demonstrate that topical administration of low-dose aspirin (36 μg/wound/day) improves cutaneous wound healing by increasing wound closure through the promotion of the inflammation resolution program of macrophages. This treatment increased efferocytosis of wound macrophages from aspirin-treated diabetic mice compared with untreated diabetic mice. We also show that aspirin treatment of high-fat-fed mice oriented the phenotype of wound macrophages toward an anti-inflammatory and proresolutive profile characterized by a decrease of LTB4 production. The use of diabetic mice deficient for 5-LOX or 12/15-LOX demonstrated that these two enzymes of acid arachidonic metabolism are essential for the beneficial effect of aspirin on wound healing. Thus, aspirin treatment modified the balance between pro- and anti-inflammatory eicosanoids by promoting the synthesis of proresolving LXA4 through 5-LOX, LTA4, 12/15-LOX signaling. In conclusion, the restoration of an anti-inflammatory and proresolutive phenotype of wound macrophages by the topical administration of low-dose aspirin represents a promising therapeutic approach in chronic wounds.
Exercise-Linked Skeletal Irisin Ameliorates Diabetes-Associated Osteoporosis by Inhibiting the Oxidative Damage-Dependent miR-150-FNDC5/Pyroptosis Axis.
Recent evidence suggests that physical exercise (EX) promotes skeletal development. However, the impact of EX on the progression of bone loss and deterioration of mechanical strength in type 2 diabetic (T2DM) mice remains unexplored. In the present study, we investigated the effect of EX on bone mass and mechanical quality using a diabetic mouse model. The T2DM mouse model was established by a high-fat diet (HFD) with two streptozotocin (STZ) injections (50 mg/kg/body weight) in C57BL/6 female mice. The diabetic mice underwent treadmill exercises (5 days/week at 7-11 m/min for 60 minutes per day) for 8 weeks. The data showed that diabetes upregulated miR-150 expression through an oxidative stress and suppressed FNDC5/irisin by binding to its 3'-UTR. The decreased level of irisin further triggers the pyroptosis response in diabetic bone tissue. EX or N-acetyl cysteine (NAC) or anti-miRNA-150 transfection in T2DM mice restored FNDC5/irisin expression and bone formation. Furthermore, EX or r-irisin administration prevented T2DM-induced hyperglycemia and improved glucose intolerance in diabetic mice. Furthermore, osteoblastic knockdown of Nlrp3 silencing (siNlrp3) or pyroptosis inhibitor (AYC) treatment restores bone mineralization in diabetic mice. Micro-CT scans and mechanical testing revealed that trabecular bone microarchitecture and bone mechanical properties were improved after EX in diabetic mice. Irisin, either induced by skeleton or daily EX or directly administered, prevents bone loss by mitigating inflammasome-associated pyroptosis signaling in diabetic mice. This study demonstrates that EX-induced skeletal irisin ameliorates diabetes-associated glucose intolerance and bone loss and possibly provides a mechanism of its effects on metabolic osteoporosis.
BAF60a Deficiency in Macrophage Promotes Diet-Induced Obesity and Metabolic Inflammation.
Adipose tissue macrophage (ATM) has been shown to play a key role in the pathogenesis of obesity-associated adipose tissue inflammation and metabolic diseases. However, the upstream factors that integrate the environmental signals to control ATM activation and adipose inflammation in obesity remain elusive. Here, we identify BAF60a, a subunit of the switch/sucrose-nonfermentable (SWI/SNF) chromatin remodeling complexes, as the central checkpoint regulator of obesity-induced ATM activation, adipose tissue inflammation, and systemic metabolic impairment. BAF60a expression was robustly downregulated in the adipose tissue stromal vascular fractions in type 2 diabetic mice. Myeloid-specific BAF60a knockout (BaMKO) promotes ATM proinflammatory activation, exacerbating diet-induced obesity, insulin resistance, and metabolic dysfunction. Conversely, myeloid-specific overexpression of BAF60a in mice attenuates macrophage proinflammatory activation. Mechanistically, transcriptome and chromatin landscape analyses demonstrate that BAF60a inactivation triggers the expression of proinflammatory gene program through chromatin remodeling. Moreover, motif analysis of ATAC-Seq and CUT&Tag-Seq data identifies the transcription factor Atf3 that physically interacts with BAF60a to suppress the proinflammatory gene expression, thereby controlling ATM activation and metabolic inflammation in obesity. Consistently, myeloid-specific Atf3 deficiency also promotes the proinflammatory activation of macrophage. This work uncovers BAF60a/Atf3 axis as the key regulator in obesity-associated ATM activation, adipose tissue inflammation, and metabolic diseases.
Inhibition of CCL28/CCR10-Mediated eNOS Downregulation Improves Skin Wound Healing in the Obesity-Induced Mouse Model of Type 2 Diabetes.
Chronic, nonhealing skin wounds, such as diabetic foot ulcers (DFUs), are common in patients with type 2 diabetes. Here, we investigated the role of chemokine (C-C motif) ligand 28 (CCL28) and its receptor C-C chemokine receptor type 10 (CCR10) in downregulation of endothelial nitric (NO) oxide synthase (eNOS) in association with delayed skin wound healing in the db/db mouse model of type 2 diabetes. We observed reduced eNOS expression and elevated CCL28/CCR10 levels in dorsal skin of db/db mice and subdermal leg biopsy specimens from human subjects with type 2 diabetes. Further interrogation revealed that overexpression of CCR10 reduced eNOS expression, NO bioavailability, and tube formation of human dermal microvascular endothelial cells (HDMVECs) in vitro, which was recapitulated in mouse dorsal skin. In addition, incubation of HDMVECs with CCL28 led to internalization of the CCR10/eNOS complex and colocalization with lysosome-associated membrane protein 1. Finally, topical application of myristoylated CCR10 binding domain 7 amino acid (Myr-CBD7) peptide prevented CCR10-eNOS interaction and subsequent eNOS downregulation, enhanced eNOS/NO levels, eNOS/VEGF-R2+ microvessel density, and blood perfusion, reduced inflammatory cytokine levels, and importantly, decreased wound healing time in db/db mice. Thus, endothelial cell CCR10 activation in genetically obese mice with type 2 diabetes promotes eNOS depletion and endothelial dysfunction, and targeted disruption of CCR10/eNOS interaction improves wound healing.
MicroRNA-29 Ameliorates Fibro-Inflammation and Insulin Resistance in HIF1α-Deficient Obese Adipose Tissue by Inhibiting Endotrophin Generation.
Dysregulation of extracellular matrix proteins in obese adipose tissue (AT) induces systemic insulin resistance. The metabolic roles of type VI collagen and its cleavage peptide endotrophin in obese AT are well established. However, the mechanisms regulating endotrophin generation remain elusive. Herein, we identified that several endotrophin-containing peptides (pre-endotrophins) were generated from the COL6A3 chain in a stepwise manner for the efficient production of mature endotrophin, partly through the action of hypoxia-induced matrix metalloproteinases (MMPs), including MMP2, MMP9, and MMP16. Hypoxia is an upstream regulator of COL6A3 expression and the proteolytic processing that regulates endotrophin generation. Hypoxia-inducible factor 1α (HIF1α) and the hypoxia-associated suppression of microRNA-29 (miR-29) cooperatively control the levels of COL6A3 and MMPs, which are responsible for endotrophin generation in hypoxic ATs. Adipocyte-specific Hif1α knock-out (APN-HIF1αKO) mice fed a chronic high-fat diet exhibited the significant amelioration of both local fibro-inflammation in AT and systemic insulin resistance compared with their control littermates, partly through the inhibition of endotrophin generation. Strikingly, adenovirus-mediated miR-29 overexpression in the ATs of APN-HIF1αKO mice in obesity significantly decreased endotrophin levels, suggesting that miR-29, combined with HIF1α inhibition in AT, could be a promising therapeutic strategy for treating obesity and related metabolic diseases.
Predictive Metabolomic Markers in Early to Mid-pregnancy for Gestational Diabetes Mellitus: A Prospective Test and Validation Study.
Gestational diabetes mellitus (GDM) predisposes pregnant individuals to perinatal complications and long-term diabetes and cardiovascular diseases. We developed and validated metabolomic markers for GDM in a prospective test-validation study. In a case-control sample within the PETALS cohort (GDM n = 91 and non-GDM n = 180; discovery set), a random PETALS subsample (GDM n = 42 and non-GDM n = 372; validation set 1), and a case-control sample within the GLOW trial (GDM n = 35 and non-GDM n = 70; validation set 2), fasting serum untargeted metabolomics were measured by gas chromatography/time-of-flight mass spectrometry. Multivariate enrichment analysis examined associations between metabolites and GDM. Ten-fold cross-validated LASSO regression identified predictive metabolomic markers at gestational weeks (GW) 10-13 and 16-19 for GDM. Purinone metabolites at GW 10-13 and 16-19 and amino acids, amino alcohols, hexoses, indoles, and pyrimidine metabolites at GW 16-19 were positively associated with GDM risk (false discovery rate <0.05). A 17-metabolite panel at GW 10-13 outperformed the model using conventional risk factors, including fasting glycemia (area under the curve: discovery 0.871 vs. 0.742, validation 1 0.869 vs. 0.731, and validation 2 0.972 vs. 0.742; P < 0.01). Similar results were observed with a 13-metabolite panel at GW 17-19. Dysmetabolism is present early in pregnancy among individuals progressing to GDM. Multimetabolite panels in early pregnancy can predict GDM risk beyond conventional risk factors.
Plasma exosomal microRNAs Associated with Metabolism as Early Predictor of Gestational Diabetes Mellitus.
To date, the microRNA (miRNA) expression profile of plasma exosomes in women whose pregnancy is complicated by gestational diabetes mellitus (GDM) has not been fully clarified. In this study, differentially expressed miRNAs in plasma exosomes were identified by high-throughput small RNA sequencing in 12 GDM and 12 normal glucose tolerance (NGT) pregnant women and validated in 102 GDM and 101 NGT pregnant women. A total of 22 exosomal miRNAs were found and five of them were verified by qRT-PCR. Exosmal miR-423-5p was upregulated, while miR-122-5p, miR-148a-3p, miR-192-5p, and miR-99a-5p were downregulated in women whose pregnancy is complicated by GDM. IGF1R and GYS1 as target genes of miR-423-5p, and G6PC3 and FDFT1 as target genes of miR-122-5p, were associated with insulin and AMPK signaling pathways and may participate in the regulation of metabolism in GDM. The five exosomal miRNAs showed an AUC of 0.82 (95%CI, 0.73∼0.91) in early prediction of GDM. Our study demonstrated that dysregulated exosomal miRNAs in plasma from GDM pregnant women might influence the insulin and AMPK signaling pathways and could contribute to the early prediction of GDM.
miR-130b/301b is a Negative Regulator of Beige Adipogenesis and Energy Metabolism in vitro and in vivo.
Thermogenic brown or beige adipocytes dissipate energy in the form of heat and thereby counteract obesity and related metabolic complications. The microRNA cluster, miR-130b/301b, is highly expressed in adipose tissues and has been implicated in metabolic diseases as a post-transcriptional regulator of mitochondrial biogenesis and lipid metabolism. We investigated the roles of miR-130b/301b in regulating beige adipogenesis in vivo and in vitro. miR-130b/301b declined in adipose progenitor cells during beige adipogenesis, while forced overexpression of miR-130b-3p or miR-301b-3p suppressed uncoupling protein 1 (UCP1) and mitochondrial respiration, suggesting a decline in miR-130b-3p or miR-301b-3p is required for adipocyte precursors to develop the beige phenotype. Mechanistically, miR-130b/301b directly targeted AMP-activated protein kinase (AMPKα1) and suppressed peroxisome proliferator-activated receptor gamma coactivator 1-alpha (Pgc-1α), key regulators of brown adipogenesis and mitochondrial biogenesis. Mice lacking the miR-130b/301b microRNA cluster showed reduced visceral adiposity and less weight gain. miR-130b/301b null mice exhibited improved glucose tolerance, increased UCP1 and AMPK activation in subcutaneous fat (iWAT), and increased response to cold-induced energy expenditure. Together, these data identify the miR-130b/301b cluster as a new regulator that suppresses beige adipogenesis involving PGC-1α and AMPK signaling in iWAT and is therefore a potential therapeutic target against obesity and related metabolic disorders.
Long Noncoding RNA lncRHL Regulates Hepatic VLDL Secretion by Modulating hnRNPU/BMAL1/MTTP Axis.
Dysregulation of hepatic VLDL secretion contributes to the pathogenesis of metabolic diseases, such as nonalcoholic fatty liver disease (NAFLD) and hyperlipidemia. Accumulating evidence has suggested that long noncoding RNAs (lncRNAs) had malfunctioning roles in the pathogenesis of NAFLD. However, the function of lncRNAs in controlling hepatic VLDL secretion remains largely unillustrated. Here, we identified a novel lncRNA, lncRNA regulator of hyperlipidemia (lncRHL), which was liver-enriched, downregulated on high-fat diet feeding, and inhibited by oleic acid treatment in primary hepatocytes. With genetic manipulation in mice and primary hepatocytes, depletion of lncRHL induces hepatic VLDL secretion accompanied by decreased hepatic lipid contents. Conversely, lncRHL restoration reduces VLDL secretion with increased lipid deposition in hepatocytes. Mechanistic analyses indicate that lncRHL binds directly to heterogeneous nuclear ribonuclear protein U (hnRNPU), and thereby enhances its stability, and that hnRNPU can transcriptional activate Bmal1, leading to inhibition of VLDL secretion in hepatocytes. lncRHL deficiency accelerates the protein degradation of hnRNPU and suppresses the transcription of Bmal1, which in turn activates VLDL secretion in hepatocytes. With results taken together, we conclude that lncRHL is a novel suppressor of hepatic VLDL secretion. Activating the lncRHL/hnRNPU/BMAL1/MTTP axis represents a potential strategy for the maintenance of intrahepatic and plasma lipid homeostasis.