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Articles from the An insulin centennial: Past, Present, and Future Special Issue, Edited by Alexander Kokkinos and Eleuterio Ferrannini| Volume 125, 154892, December 01, 2021

Insulin resistance and insulin sensitizing agents

  • Lucia Mastrototaro
    Affiliations
    Institute for Clinical Diabetology, German Diabetes Center, Leibniz Center for Diabetes Research at Heinrich-Heine University Düsseldorf, Düsseldorf, Germany

    German Center for Diabetes Research, Partner Düsseldorf, München-Neuherberg, Germany
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  • Michael Roden
    Correspondence
    Corresponding author at: Department of Endocrinology and Diabetology, Medical Faculty and University Hospital, Heinrich-Heine University, Düsseldorf, c/o German Diabetes Center at Heinrich-Heine University, Auf dem Hennekamp 65, 40225 Düsseldorf, Germany.
    Affiliations
    Institute for Clinical Diabetology, German Diabetes Center, Leibniz Center for Diabetes Research at Heinrich-Heine University Düsseldorf, Düsseldorf, Germany

    German Center for Diabetes Research, Partner Düsseldorf, München-Neuherberg, Germany

    Department of Endocrinology and Diabetology, Medical Faculty and University Hospital, Heinrich-Heine University Düsseldorf, Düsseldorf, Germany
    Search for articles by this author
Open AccessPublished:September 22, 2021DOI:https://doi.org/10.1016/j.metabol.2021.154892

      Highlights

      • Common insulin resistance (IR) varies among people with obesity and type-2 diabetes.
      • Exosomes and miRNA contribute to altered organ crosstalk during development of IR.
      • Lifestyle interventions do not uniformly improve insulin sensitivity in IR states.
      • In addition to metformin and thiazolidinedione, new therapeutic concepts improve IR.
      • Biomarkers for early detection of IR and predicting therapy response are emerging.

      Abstract

      Insulin resistance is a common feature of obesity and type 2 diabetes, but novel approaches of diabetes subtyping (clustering) revealed variable degrees of insulin resistance in people with diabetes. Specifically, the severe insulin resistant diabetes (SIRD) subtype not only exhibits metabolic abnormalities, but also bears a higher risk for cardiovascular, renal and hepatic comorbidities. In humans, insulin resistance comprises dysfunctional adipose tissue, lipotoxic insulin signaling followed by glucotoxicity, oxidative stress and low-grade inflammation. Recent studies show that aside from metabolites (free fatty acids, amino acids) and signaling proteins (myokines, adipokines, hepatokines) also exosomes with their cargo (proteins, mRNA and microRNA) contribute to altered crosstalk between skeletal muscle, liver and adipose tissue during the development of insulin resistance. Reduction of fat mass mainly, but not exclusively, explains the success of lifestyle modification and bariatric surgery to improve insulin sensitivity. Moreover, some older antihyperglycemic drugs (metformin, thiazolidinediones), but also novel therapeutic concepts (new peroxisome proliferator-activated receptor agonists, incretin mimetics, sodium glucose cotransporter inhibitors, modulators of energy metabolism) can directly or indirectly reduce insulin resistance. This review summarizes molecular mechanisms underlying insulin resistance including the roles of exosomes and microRNAs, as well as strategies for the management of insulin resistance in humans.

      Keywords

      1. Introduction

      Insulin resistance (IR) describes an impairment of insulin sensitivity, reflected by a shift of the insulin concentration-effect curve towards higher insulin concentrations [
      • Roden M.
      Clinical diabetes research: methods and techniques.
      ]. Of note, common chronic IR, as observed in obesity and type 2 diabetes mellitus (T2DM), mostly also includes an impairment of insulin responsiveness, reflected by a reduction of the maximal insulin effect. In humans, IR is generally defined by the inability of insulin-target tissues to adequately dispose blood glucose, suppress endogenous glucose production (EGP) and lipolysis as well as to stimulate glycogen synthesis at elevated plasma insulin concentrations [
      • Roden M.
      Clinical diabetes research: methods and techniques.
      ]. In contrast to common chronic IR, conditions of increased metabolic demand, such as fasting, dehydration, stress and infection, can induce a state of reversible IR mediated by stress hormones and pro-inflammatory cytokines, which inhibit insulin action in target tissues and promote energy mobilization [
      • Tsatsoulis A.
      • Mantzaris M.D.
      • Bellou S.
      • Andrikoula M.
      Insulin resistance: an adaptive mechanism becomes maladaptive in the current environment - an evolutionary perspective.
      ]. Finally, mutations in the insulin signaling cascade can cause genetic forms of IR, such as leprechaunism, Rabson–Mendenhall syndrome and type A insulin resistance syndrome [
      • Angelidi A.M.
      • Filippaios A.
      • Mantzoros C.S.
      Severe insulin resistance syndromes.
      ]. These specific genetic causes and further epigenetic mechanisms possibly underlying IR [
      • Nilsson E.
      • Jansson P.A.
      • Perfilyev A.
      • Volkov P.
      • Pedersen M.
      • Svensson M.K.
      • et al.
      Altered DNA methylation and differential expression of genes influencing metabolism and inflammation in adipose tissue from subjects with type 2 diabetes.
      ] are beyond the scope of this review.
      Common IR mainly results from an imbalance of energy intake and expenditure [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ], although genetic predisposition is also involved [
      • Manning A.K.
      • Hivert M.F.
      • Scott R.A.
      • Grimsby J.L.
      • Bouatia-Naji N.
      • Chen H.
      • et al.
      A genome-wide approach accounting for body mass index identifies genetic variants influencing fasting glycemic traits and insulin resistance.
      ]. The still rising prevalence of obesity and T2DM [
      • Zheng Y.
      • Ley S.H.
      • Hu F.B.
      Global aetiology and epidemiology of type 2 diabetes mellitus and its complications.
      ] requires the assessment of IR not only for refined phenotyping, but even more to enable stratified prevention and treatment. The hyperinsulinemic-euglycemic clamp test (HEC) is the gold standard method for quantifying in vivo insulin sensitivity, which is calculated as glucose disposal during steady state and expressed as M-value or rate of glucose disappearance (Rd) [
      • Roden M.
      Clinical diabetes research: methods and techniques.
      ]. Under these conditions, skeletal muscle (SkM) takes up the majority of glucose so that whole-body or peripheral insulin sensitivity by M-value and Rd mainly reflect SkM insulin sensitivity [
      • Roden M.
      Clinical diabetes research: methods and techniques.
      ]. In addition, minimal model analysis of the frequently sampled intravenous or oral glucose tolerance tests allow to assess insulin sensitivity (e.g. Si, Matsuda and oral glucose insulin sensitivity indices) during dynamically changing glycemia and insulinemia. For larger studies, surrogate indices of insulin sensitivity have been developed from fasting insulin, C-peptide and glucose levels, such as the quantitative insulin sensitivity check index and the homeostatic model assessment IR (HOMA-IR) [
      • Roden M.
      Clinical diabetes research: methods and techniques.
      ], which although assessed under different metabolic conditions correlate reasonably well with clamp-derived measures [
      • Zaharia O.P.
      • Strassburger K.
      • Strom A.
      • Bonhof G.J.
      • Karusheva Y.
      • Antoniou S.
      • et al.
      Risk of diabetes-associated diseases in subgroups of patients with recent-onset diabetes: a 5-year follow-up study.
      ].
      Under typical clamp conditions, which simulate postprandial hyperinsulinemia, the IR observed in people with T2DM and their insulin-resistant offspring results mainly from reduced non-oxidative glucose disposal due to lower SkM glycogen synthesis, as a consequence of lower glucose uptake [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ]. The lower glucose transport is caused by abnormal insulin signaling via the insulin receptor kinase (IRK) and insulin receptor substrate 1 (IRS1) [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ]. Of note, a recent study showed that the pattern of IRS1 phosphorylation is not uniformly altered in all T2DM individuals, suggesting that other post-translational modifications likely contribute to the attenuation of insulin signaling [
      • Karlsson H.K.R.
      • Kasahara A.
      • Ikeda M.
      • Chibalin A.
      • Harada J.
      • Ryden M.
      • et al.
      Quantitative phosphoproteomic analysis of IRS1 in skeletal muscle from men with normal glucose tolerance or type 2 diabetes: a case-control study.
      ]. In white adipose tissue (WAT), IR may be a very early event during the development of T2DM, triggered by invasion of macrophages with subsequent local release of pro-inflammatory cytokines, which inhibit IRK activity [
      • Hotamisligil G.S.
      • Budavari A.
      • Murray D.
      • Spiegelman B.M.
      Reduced tyrosine kinase activity of the insulin receptor in obesity-diabetes. Central role of tumor necrosis factor-alpha.
      ]. Dysfunctional insulin resistant WAT exhibits excessive lipolysis with release of free fatty acids (FFA), which enter other tissues such as the liver and form lipid toxic intermediates, resulting in inhibition of IRK and hepatic IR with subsequently elevated gluconeogenesis, reduced glycogen synthesis and higher EGP [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ]. Higher cellular glucose availability also favors synthesis of fatty acids (FA) via de novo lipogenesis (DNL), which augments intrahepatic lipid (IHL) content and correlates negatively with insulin sensitivity [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ,
      • Roumans K.H.M.
      • Lindeboom L.
      • Veeraiah P.
      • Remie C.M.E.
      • Phielix E.
      • Havekes B.
      • et al.
      Hepatic saturated fatty acid fraction is associated with de novo lipogenesis and hepatic insulin resistance.
      ]. In addition to metabolites and cytokines also gut microbiota can contribute to IR. Gut dysbiosis is associated with the disruption of tight-junctions and the translocation of bacterial lipopolysaccharides (LPS) into the systemic circulation, resulting in the development of inflammation and IR [
      • Saad M.J.
      • Santos A.
      • Prada P.O.
      Linking gut microbiota and inflammation to obesity and insulin resistance.
      ]. Conversely, gut microbiota-derived short-chain FA could prevent obesity and T2DM by reducing inflammation and increasing lipid storage capacity by WAT [
      • Gancheva S.
      • Jelenik T.
      • Alvarez-Hernandez E.
      • Roden M.
      Interorgan metabolic crosstalk in human insulin resistance.
      ].

      2. Mechanisms underlying the development of insulin resistance

      Common IR associates with defective insulin signaling mediated by several mechanisms in humans, including accumulation of specific lipid mediators, abnormal features of mitochondrial function as well as increases in stress-activated protein c-Jun-N-terminal-kinase (JNK) and inflammatory pathways (Fig. 1) [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ].
      Fig. 1
      Fig. 1Altered interorgan crosstalk underlying the development of insulin resistance and possible therapeutic strategies.
      High energy intake induces both gut dysbiosis, which is associated with elevation of LPS and BCAA and decreased release of GLP-1 and SCFA into the systemic circulation, and excessive FA storage in WAT, which leads to WAT dysfunction and insulin resistance. BW loss by hypocaloric nutrition or bariatric surgery is the most effective strategy to improve WAT dysfunction and insulin resistance. Dietary modification, bariatric surgery, metformin and the non-steroidal FX-Ra, PX-104, also act via affecting gut microbiota. Metformin and GLP-1Ra further stimulate secretion of GLP-1, which mediates satiety and BW loss. Dysfunctional WAT also lowers adiponectin secretion and favors lipolysis-induced FFA release. Macrophages infiltration triggers WAT insulin resistance and subsequently release of pro-inflammatory cytokine and exosomes, which participate to the interorgan crosstalk. Several pharmacological agents including PPARδa, PPARα/δa, IL-6i, TNFαi, CCR2/5i, and synthetic ω3-FA reduce markers of systemic inflammation and insulin resistance. Anti-inflammatory drugs, such as salsalate and antagonists of IL-6 and TNFα, but also SGLT2i, PTP1Bi and FGF21a, increase plasma concentrations of adiponectin, which lowers ectopic lipid accumulation. The PPARγ ligands such as TZD also promote FFA storage and thereby lower circulating FFA levels. Once FFA enter the liver, they form lipid toxic intermediates (sn-1,2-DAG and Cer), which inhibit the insulin signaling cascade, resulting in hepatic insulin resistance with subsequently elevated EGP. As consequence, glucose is diverted to FA and favor DNL, which contributes to IHL storage. Metformin decreases EGP through inhibition of GPD2 and CI. At early stages of obesity-related insulin resistance, hepatic FAO adapts by upregulation, which is lost in NASH, leading to decreased FAO, ROS production and inflammation. Hepatic ER stress is also enhanced in obesity and inhibits insulin signaling. BW loss and panPPARa decrease IHL and improve insulin resistance. A number of further pharmacological agents including FGF21a, PPARɑa and CRMP (so far tested in nonhuman primates) aim to augment FAO, whereas KHKi target DNL. Increased intramyocellular sn-1,2-DAG activate PKCθ, which in turns causes Ser1101-phosphorylation of IRS1 and suppression of insulin-stimulated glucose uptake. Furthermore, decreased SkM mitochondrial content and FAO exacerbate lipid-induced insulin resistance. Exercise increases insulin sensitivity by enhancing GLUT4-glucose uptake and increasing mitochondrial biogenesis. Furthermore, NMN and Met stimulate GLUT4 translocation to the plasma membrane by enhancing Akt phosphorylation and AMPK phosphorylation, respectively. PPARδa also act on the SkM by enhancing FAO.
      a = agonist; Akt = protein kinase B; AMPK = AMP-activated protein kinase; BCAA = branched-chain amino acids; BW = body weight; CI = complex I; CCR2/5 = chemokine receptor; Cer = ceramides; CRMP = controlled-release mitochondrial protonophore; DAG = diacylglycerols; DNL = de novo lipogenesis; EGP = endogenous glucose production; ER = endoplasmic reticulum; FA = fatty acids; FAO = fatty acid oxidation; FFA = free fatty acids; FGF 21 = fibroblast growth factor 21; FXR = farnesoid X receptor; GLP-1R = glucagon-like peptide 1 receptor; GPD2 = glycerol-3-phosphate dehydrogenase; i = inhibitor; IHL = intrahepatic lipid; IL = interleukin; IR = insulin receptor; IRS1 = insulin receptor substrate 1; KHK = ketohexokinase; LPS = lipopolysaccharides; Met = metformin; Mt. = mitochondrial; NASH = non-alcoholic steatohepatitis; NMN = Nicotinamide mononucleotide; nPKC = novel protein kinase C; PPAR = peroxisome proliferator-activated receptor; PTP1B = protein tyrosine phosphatase-1B; ROS = reactive oxygen species; SCFA = short-chain fatty acids; (S)EV = (small) extracellular vesicles; SGLT2 = sodium glucose cotransporter 2; SkM = skeletal muscle; sn-1,2-DAG = sn-1,2-diacylglycerols; TNFα = tumor necrosis factor α; TZD = thiazolidinediones; WAT = adipose tissue.

      2.1 Lipid-induced insulin resistance

      Diacylglycerols (DAG) and ceramides have been largely studied as mediators of lipid-induced IR in liver and SkM [
      • Gancheva S.
      • Jelenik T.
      • Alvarez-Hernandez E.
      • Roden M.
      Interorgan metabolic crosstalk in human insulin resistance.
      ]. During lipid oversupply from high-fat high-calorie nutrition or excessive adipose lipolysis, FFA accumulate ectopically exceeding the rate of intracellular FA oxidation (FAO) and storage [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ]. Moreover, reduced mitochondrial density and function in insulin-resistant people impede FAO and exacerbate lipid-induced IR [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ].
      Chronic elevation of certain DAG species impairs insulin signaling via activation of conventional (α, βI, βII, γ) and novel protein kinase C (nPKC) isoforms (δ, ε, ν, θ). DAG localized in the plasma membrane (PM) activate nPKCs, whereas sequestration of DAG and PKCs in lipid droplets may protect from IR [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ]. Accumulation of sn-1,2-DAG in the PM activates PKCε and inhibitory IRK Thr1160-phosphorylation in the liver [
      • Lyu K.
      • Zhang Y.
      • Zhang D.
      • Kahn M.
      • Ter Horst K.W.
      • Rodrigues M.R.S.
      • et al.
      A membrane-bound diacylglycerol species induces PKC-mediated hepatic insulin resistance.
      ] as well as in WAT at least in mice [
      • Lyu K.
      • Zhang D.
      • Song J.D.
      • Li X.
      • Perry R.J.
      • Samuel V.T.
      • et al.
      Short-term overnutrition induces white adipose tissue insulin resistance through sn-1,2-diacylglycerol - PKCepsilon - insulin receptorT1160 phosphorylation.
      ], resulting in IR. Besides PKCε, PKCδ can be also increased in livers of obese humans and may induce hepatic IR via decreased Tyr612-phosphorylation of IRS1 and Ser473-phosphorylation of protein kinase B (Akt) [
      • Bezy O.
      • Tran T.T.
      • Pihlajamaki J.
      • Suzuki R.
      • Emanuelli B.
      • Winnay J.
      • et al.
      PKCdelta regulates hepatic insulin sensitivity and hepatosteatosis in mice and humans.
      ]. In SkM of obese, T2DM or lipid-infused individuals, DAG activates PKCθ, which in turns causes IR via inhibitory Ser1101-phosphorylation of IRS1 [
      • Roden M.
      • Shulman G.I.
      The integrative biology of type 2 diabetes.
      ,
      • Szendroedi J.
      • Yoshimura T.
      • Phielix E.
      • Koliaki C.
      • Marcucci M.
      • Zhang D.
      • et al.
      Role of diacylglycerol activation of PKCtheta in lipid-induced muscle insulin resistance in humans.
      ]. Additionally, lipid infusion-induced elevation of DAG content in SkM of healthy men can activate PKCβII, PKCδ and nuclear factor (NF)-κB inflammatory pathway [
      • Itani S.I.
      • Ruderman N.B.
      • Schmieder F.
      • Boden G.
      Lipid-induced insulin resistance in human muscle is associated with changes in diacylglycerol, protein kinase C, and IkappaB-alpha.
      ].
      The role of intramyocellular ceramides in IR is controversial, indeed not all studies confirmed a relationship between SkM IR and ceramides in human obesity and T2DM [
      • Szendroedi J.
      • Yoshimura T.
      • Phielix E.
      • Koliaki C.
      • Marcucci M.
      • Zhang D.
      • et al.
      Role of diacylglycerol activation of PKCtheta in lipid-induced muscle insulin resistance in humans.
      ,
      • Adams 2nd, J.M.
      • Pratipanawatr T.
      • Berria R.
      • Wang E.
      • DeFronzo R.A.
      • Sullards M.C.
      • et al.
      Ceramide content is increased in skeletal muscle from obese insulin-resistant humans.
      ]. Nevertheless, hepatic total ceramides and certain dihydroceramides (e.g. C16:0) are increased in insulin-resistant humans with non-alcoholic steatohepatitis (NASH) and correlate positively with hepatic oxidative stress and inflammation [
      • Apostolopoulou M.
      • Gordillo R.
      • Koliaki C.
      • Gancheva S.
      • Jelenik T.
      • De Filippo E.
      • et al.
      Specific hepatic sphingolipids relate to insulin resistance, oxidative stress, and inflammation in nonalcoholic steatohepatitis.
      ]. Interestingly, the ratio of ceramide/dihydroceramide 16:0 positively associates with markers of inflammation in visceral adipose tissue (VAT), whereas it relates negatively with IHL, mitochondrial capacity and lipid peroxidation in liver [
      • Apostolopoulou M.
      • Gordillo R.
      • Gancheva S.
      • Strassburger K.
      • Herder C.
      • Esposito I.
      • et al.
      Role of ceramide-to-dihydroceramide ratios for insulin resistance and non-alcoholic fatty liver disease in humans.
      ]. These data suggest tissue-specific roles for these lipid species without supporting an involvement of ceramide C16:0 in obesity-induced IR in humans, as shown in previous studies [
      • Turpin S.M.
      • Nicholls H.T.
      • Willmes D.M.
      • Mourier A.
      • Brodesser S.
      • Wunderlich C.M.
      • et al.
      Obesity-induced CerS6-dependent C16:0 ceramide production promotes weight gain and glucose intolerance.
      ].

      2.2 Abnormal features of mitochondrial function

      Impaired muscle mitochondrial functionality, which includes mitochondrial dynamics, turnover and plasticity, and lower content are frequent features of the elderly and people with IR or overt T2DM [
      • Koliaki C.
      • Roden M.
      Alterations of mitochondrial function and insulin sensitivity in human obesity and diabetes mellitus.
      ]. Downregulation of peroxisome proliferator-activated receptor (PPAR)γ coactivator 1-α (PGC-1α) suggests altered mitochondrial biogenesis in T2DM, whereas reduced expression of lipoprotein lipase (LPL) and PPARδ is responsible for decreased mitochondrial content in SkM of nondiabetic insulin-resistant offspring [
      • Morino K.
      • Petersen K.F.
      • Sono S.
      • Choi C.S.
      • Samuel V.T.
      • Lin A.
      • et al.
      Regulation of mitochondrial biogenesis by lipoprotein lipase in muscle of insulin-resistant offspring of parents with type 2 diabetes.
      ]. Mitochondrial fusion is also decreased in SkM of obese individuals and the transcript levels of Mfn2 correlate positively with markers of insulin sensitivity [
      • Koliaki C.
      • Roden M.
      Alterations of mitochondrial function and insulin sensitivity in human obesity and diabetes mellitus.
      ]. Decreased mitochondrial content and transcript levels of oxidative phosphorylation genes have been demonstrated in WAT of obese and T2DM persons and might contribute to the development of systemic IR [
      • Koliaki C.
      • Roden M.
      Alterations of mitochondrial function and insulin sensitivity in human obesity and diabetes mellitus.
      ]. In contrast, hepatic mitochondrial respiration can be upregulated in obesity with and without steatosis, but decreases in NASH, despite cellular substrate excess and higher mitochondrial content [
      • Koliaki C.
      • Szendroedi J.
      • Kaul K.
      • Jelenik T.
      • Nowotny P.
      • Jankowiak F.
      • et al.
      Adaptation of hepatic mitochondrial function in humans with non-alcoholic fatty liver is lost in steatohepatitis.
      ], which might be a consequence of accumulation of dysfunctional mitochondria rather than increased biogenesis [
      • Wang L.
      • Liu X.
      • Nie J.
      • Zhang J.
      • Kimball S.R.
      • Zhang H.
      • et al.
      ALCAT1 controls mitochondrial etiology of fatty liver diseases, linking defective mitophagy to steatosis.
      ].

      2.3 Endoplasmic reticulum (ER) stress

      Although lipids likely play the primary role in mediating IR and inflammation [
      • Roden M.
      • Price T.B.
      • Perseghin G.
      • Petersen K.F.
      • Rothman D.L.
      • Cline G.W.
      • et al.
      Mechanism of free fatty acid-induced insulin resistance in humans.
      ], not all insulin-resistant individuals display elevated FFA concentrations. Evidence from human studies point to enhanced ER stress and JNK activation in liver and in WAT of obese individuals and to a positive correlation with BMI and percent fat [
      • Sharma N.K.
      • Das S.K.
      • Mondal A.K.
      • Hackney O.G.
      • Chu W.S.
      • Kern P.A.
      • et al.
      Endoplasmic reticulum stress markers are associated with obesity in nondiabetic subjects.
      ,
      • Gregor M.F.
      • Yang L.
      • Fabbrini E.
      • Mohammed B.S.
      • Eagon J.C.
      • Hotamisligil G.S.
      • et al.
      Endoplasmic reticulum stress is reduced in tissues of obese subjects after weight loss.
      ]. Indeed, ER stress is reduced after surgically-induced body weight (BW) loss, underlining that ER stress affects insulin action in obesity [
      • Gregor M.F.
      • Yang L.
      • Fabbrini E.
      • Mohammed B.S.
      • Eagon J.C.
      • Hotamisligil G.S.
      • et al.
      Endoplasmic reticulum stress is reduced in tissues of obese subjects after weight loss.
      ]. Conversely, the use of chemical chaperones, which enhance protein folding and protect mice against ER stress, improves hepatic and muscle insulin sensitivity, without affecting WAT insulin sensitivity and the expression of ER stress markers in obese humans [
      • Kars M.
      • Yang L.
      • Gregor M.F.
      • Mohammed B.S.
      • Pietka T.A.
      • Finck B.N.
      • et al.
      Tauroursodeoxycholic acid may improve liver and muscle but not adipose tissue insulin sensitivity in obese men and women.
      ].

      2.4 Low-grade inflammation

      Certain metabolites (e.g. cholesterol), gut-derived LPS and bacteria activate the resident hepatic macrophages which release pro-inflammatory cytokines, leading to hepatic inflammation and systemic IR [
      • Pafili K.
      • Roden M.
      Nonalcoholic fatty liver disease (NAFLD) from pathogenesis to treatment concepts in humans.
      ]. A mouse study reveals that the liver-specific transgenic expression of NF-κB in absence of steatosis and adiposity inhibits insulin-stimulated suppression of EGP and reduces glucose uptake and glycogen synthesis in SkM, suggesting a crosstalk between liver and muscles mediated by hepatic cytokines [
      • Cai D.
      • Yuan M.
      • Frantz D.F.
      • Melendez P.A.
      • Hansen L.
      • Lee J.
      • et al.
      Local and systemic insulin resistance resulting from hepatic activation of IKK-beta and NF-kappaB.
      ].
      Emerging evidences reveal that inflammation occurs also directly in SkM in obesity [
      • Wu H.
      • Ballantyne C.M.
      Skeletal muscle inflammation and insulin resistance in obesity.
      ], probably as a consequence of excessive lipid accumulation. Mechanistically, lipid oversupply activates NF-κB in myotubes, resulting in reduced mitochondrial respiratory capacity and increased reactive oxygen species (ROS) production, mitochondrial fragmentation and mitophagy [
      • Nisr R.B.
      • Shah D.S.
      • Ganley I.G.
      • Hundal H.S.
      Proinflammatory NFkB signalling promotes mitochondrial dysfunction in skeletal muscle in response to cellular fuel overloading.
      ].
      Obesity-related IR is accompanied by macrophage infiltration and inflammatory cytokines production in WAT, which activate JNK and NF-κB causing local and systemic IR [
      • Dewidar B.
      • Kahl S.
      • Pafili K.
      • Roden M.
      Metabolic liver disease in diabetes - from mechanisms to clinical trials.
      ]. However, WAT IR can exist without concomitant macrophage infiltration and inflammation, as shown in healthy humans after oral lipid ingestion [
      • Hernandez E.A.
      • Kahl S.
      • Seelig A.
      • Begovatz P.
      • Irmler M.
      • Kupriyanova Y.
      • et al.
      Acute dietary fat intake initiates alterations in energy metabolism and insulin resistance.
      ], suggesting that IR likely precedes systemic low-grade inflammation.

      2.5 Amino acids

      Several plasma metabolites, including phospholipids, ketoacids and amino acids (AA) contribute to the development of IR and are increased in T2DM [
      • Gancheva S.
      • Jelenik T.
      • Alvarez-Hernandez E.
      • Roden M.
      Interorgan metabolic crosstalk in human insulin resistance.
      ]. Elevation of plasma AA impairs insulin-stimulated glucose disposal in SkM, likely via overactivation of the mammalian target of rapamycin (mTOR)/S6 kinase pathway and phosphorylation of IRS1 [
      • Tremblay F.
      • Krebs M.
      • Dombrowski L.
      • Brehm A.
      • Bernroider E.
      • Roth E.
      • et al.
      Overactivation of S6 kinase 1 as a cause of human insulin resistance during increased amino acid availability.
      ]. Branched-chain (BC) AA are important for protein and glucose metabolism, and their plasma concentration positively correlate with IR and predict the development of T2DM in normoglycemic adults [
      • Wurtz P.
      • Soininen P.
      • Kangas A.J.
      • Ronnemaa T.
      • Lehtimaki T.
      • Kahonen M.
      • et al.
      Branched-chain and aromatic amino acids are predictors of insulin resistance in young adults.
      ]. Although BCAA are precursor of hepatic gluconeogenesis, a short-term exposure to increased plasma BCAA does not affect suppression of EGP [
      • Everman S.
      • Mandarino L.J.
      • Carroll C.C.
      • Katsanos C.S.
      Effects of acute exposure to increased plasma branched-chain amino acid concentrations on insulin-mediated plasma glucose turnover in healthy young subjects.
      ]. In SkM, BCAA impair glucose disposal during hyperinsulinemia and increase maximal ATP synthesis without affecting mitochondrial DNA abundance [
      • Tatpati L.L.
      • Irving B.A.
      • Tom A.
      • Bigelow M.L.
      • Klaus K.
      • Short K.R.
      • et al.
      The effect of branched chain amino acids on skeletal muscle mitochondrial function in young and elderly adults.
      ,
      • Smith G.I.
      • Yoshino J.
      • Stromsdorfer K.L.
      • Klein S.J.
      • Magkos F.
      • Reeds D.N.
      • et al.
      Protein ingestion induces muscle insulin resistance independent of leucine-mediated mTOR activation.
      ]. Conversely, a short-term dietary reduction of BCAA decreases postprandial insulin secretion, improves WAT metabolism and increases the Bacteroidetes in the gut microbiota of T2DM individuals [
      • Karusheva Y.
      • Koessler T.
      • Strassburger K.
      • Markgraf D.
      • Mastrototaro L.
      • Jelenik T.
      • et al.
      Short-term dietary reduction of branched-chain amino acids reduces meal-induced insulin secretion and modifies microbiome composition in type 2 diabetes: a randomized controlled crossover trial.
      ].

      3. Other mediators of interorgan crosstalk

      In addition to metabolites, cytokines (adipokines, myokines, hepatokines), microRNAs or exosomes (Exo), which carry different biologically active cargo, can contribute to the development of IR [
      • Gancheva S.
      • Jelenik T.
      • Alvarez-Hernandez E.
      • Roden M.
      Interorgan metabolic crosstalk in human insulin resistance.
      ,
      • Safdar A.
      • Saleem A.
      • Tarnopolsky M.A.
      The potential of endurance exercise-derived exosomes to treat metabolic diseases.
      ,
      • Herder C.
      • Carstensen M.
      • Ouwens D.M.
      Anti-inflammatory cytokines and risk of type 2 diabetes.
      ,
      • Molnos S.
      • Wahl S.
      • Haid M.
      • Eekhoff E.M.W.
      • Pool R.
      • Floegel A.
      • et al.
      Metabolite ratios as potential biomarkers for type 2 diabetes: a DIRECT study.
      ,
      • Raffort J.
      • Hinault C.
      • Dumortier O.
      • Van Obberghen E.
      Circulating microRNAs and diabetes: potential applications in medical practice.
      ].

      3.1 Cytokines

      WAT secretes several adipokines, which participate to the interorgan crosstalk and are associated with IR (reviewed in [
      • Herder C.
      • Carstensen M.
      • Ouwens D.M.
      Anti-inflammatory cytokines and risk of type 2 diabetes.
      ]). Adiponectin and omentin are anti-inflammatory adipokines, whose levels are inversely associated with IR [
      • Herder C.
      • Carstensen M.
      • Ouwens D.M.
      Anti-inflammatory cytokines and risk of type 2 diabetes.
      ]. The main pro-inflammatory cytokines, tumor necrosis factor (TNF)α and interleukin (IL)-6, are elevated in obesity and T2DM [
      • Shi J.
      • Fan J.
      • Su Q.
      • Yang Z.
      Cytokines and abnormal glucose and lipid metabolism.
      ]. The infusion of IL-6 increases insulin-mediated glucose uptake in healthy but not in T2DM humans [
      • Harder-Lauridsen N.M.
      • Krogh-Madsen R.
      • Holst J.J.
      • Plomgaard P.
      • Leick L.
      • Pedersen B.K.
      • et al.
      Effect of IL-6 on the insulin sensitivity in patients with type 2 diabetes.
      ], instead TNFα administration in healthy humans induces SkM IR by activating JNK [
      • Plomgaard P.
      • Bouzakri K.
      • Krogh-Madsen R.
      • Mittendorfer B.
      • Zierath J.R.
      • Pedersen B.K.
      Tumor necrosis factor-alpha induces skeletal muscle insulin resistance in healthy human subjects via inhibition of Akt substrate 160 phosphorylation.
      ]. Conversely, IL-10 negatively correlates with prevalence of T2DM, BMI and body fat, and positively correlates with insulin sensitivity in humans [
      • Bluher M.
      • Fasshauer M.
      • Tonjes A.
      • Kratzsch J.
      • Schon M.R.
      • Paschke R.
      Association of interleukin-6, C-reactive protein, interleukin-10 and adiponectin plasma concentrations with measures of obesity, insulin sensitivity and glucose metabolism.
      ].
      SkM can release myokines, which either exert their effects within SkM or in other tissues and may protect from detrimental effects of adipokines [
      • Eckardt K.
      • Gorgens S.W.
      • Raschke S.
      • Eckel J.
      Myokines in insulin resistance and type 2 diabetes.
      ]. SkM-derived IL-6 during exercise enhances EGP and SkM lipolysis, but it also triggers the production of anti-inflammatory cytokines and hinders the release of TNFα resulting in reduced systemic inflammation and risk of IR [
      • Eckardt K.
      • Gorgens S.W.
      • Raschke S.
      • Eckel J.
      Myokines in insulin resistance and type 2 diabetes.
      ]. IL-13, IL-15 and irisin, recently described as myokines, are negatively associated with IR, obesity and T2DM, and increased after exercise, as reviewed in [
      • Febbraio M.A.
      Role of interleukins in obesity: implications for metabolic disease.
      ,
      • Pedersen B.K.
      • Febbraio M.A.
      Muscles, exercise and obesity: skeletal muscle as a secretory organ.
      ].
      Proteomic studies revealed that the liver also secretes cytokines (hepatokines), which can contribute to IR and inflammation (e.g. fetuin A, hepassocin, selenoprotein P) or improve IR (e.g. adropin, sex hormone-binding globulin) [
      • Meex R.C.R.
      • Watt M.J.
      Hepatokines: linking nonalcoholic fatty liver disease and insulin resistance.
      ]. Fibroblast growth factor (FGF)21 has complex effects, as it positively associates with T2DM and obesity, but administration of its analogue reduces BW and increases adiponectin in T2DM [
      • Meex R.C.R.
      • Watt M.J.
      Hepatokines: linking nonalcoholic fatty liver disease and insulin resistance.
      ].

      3.2 MicroRNA

      MicroRNAs are small-non coding RNA molecules that regulate glucose homeostasis through the modulation of several components of insulin signaling pathway (Fig. 2) and might be used as biomarkers for monitoring the development and progression of metabolic disease in humans, as described in this section (Table 1) [
      • Raffort J.
      • Hinault C.
      • Dumortier O.
      • Van Obberghen E.
      Circulating microRNAs and diabetes: potential applications in medical practice.
      ,
      • Nigi L.
      • Grieco G.E.
      • Ventriglia G.
      • Brusco N.
      • Mancarella F.
      • Formichi C.
      • et al.
      MicroRNAs as regulators of insulin signaling: research updates and potential therapeutic perspectives in type 2 diabetes.
      ]. MicroRNA-34a, key component of adipocyte-secreted Exo, inhibits a shift of macrophage to M2 anti-inflammatory phenotype and its expression in VAT of overweight/obese humans correlates positively with parameters of IR and systemic inflammation [
      • Pan Y.
      • Hui X.
      • Hoo R.L.C.
      • Ye D.
      • Chan C.Y.C.
      • Feng T.
      • et al.
      Adipocyte-secreted exosomal microRNA-34a inhibits M2 macrophage polarization to promote obesity-induced adipose inflammation.
      ]. The levels of several extracellular microRNAs differ between obese and lean individuals and are correlated with metabolic parameters like circulating FFA, HbA1c and HOMA-IR [
      • Jones A.
      • Danielson K.M.
      • Benton M.C.
      • Ziegler O.
      • Shah R.
      • Stubbs R.S.
      • et al.
      miRNA signatures of insulin resistance in obesity.
      ]. MicroRNA-122 is associated with IR, inflammation and adiposity in overweight adults and regulates insulin signaling pathways, as revealed by functional analysis [
      • Shah R.
      • Murthy V.
      • Pacold M.
      • Danielson K.
      • Tanriverdi K.
      • Larson M.G.
      • et al.
      Extracellular RNAs are associated with insulin resistance and metabolic phenotypes.
      ]. Moreover, the exosomal microRNA profile correlates with BMI, transaminases and uric acid in children with non-alcoholic fatty liver disease (NAFLD) [
      • Zhou X.
      • Huang K.
      • Jia J.
      • Ni Y.
      • Yuan J.
      • Liang X.
      • et al.
      Exosomal miRNAs profile in children’s nonalcoholic fatty liver disease and the correlation with transaminase and uric acid.
      ] and differs between healthy and BMI-matched T2DM humans, who display higher levels of microRNA-20b-5p, which modulates the Akt signaling when transfected in SkM cells [
      • Katayama M.
      • Wiklander O.P.B.
      • Fritz T.
      • Caidahl K.
      • El-Andaloussi S.
      • Zierath J.R.
      • et al.
      Circulating exosomal miR-20b-5p is elevated in type 2 diabetes and could impair insulin action in human skeletal muscle.
      ]. People with T2DM display also upregulation of microRNA-144 and downregulation of its predicted target IRS1, which supports the relevance of microRNA-144 for T2DM development [
      • Karolina D.S.
      • Armugam A.
      • Tavintharan S.
      • Wong M.T.
      • Lim S.C.
      • Sum C.F.
      • et al.
      MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in type 2 diabetes mellitus.
      ]. The levels of circulating microRNAs are affected by surgically-induced BW loss, exercise and glucose lowering treatment [
      • Bae Y.U.
      • Kim Y.
      • Lee H.
      • Kim H.
      • Jeon J.S.
      • Noh H.
      • et al.
      Bariatric surgery alters microRNA content of circulating exosomes in patients with obesity.
      ,
      • Ghai V.
      • Kim T.K.
      • Etheridge A.
      • Nielsen T.
      • Hansen T.
      • Pedersen O.
      • et al.
      Extracellular vesicle encapsulated microRNAs in patients with type 2 diabetes are affected by metformin treatment.
      ,
      • Ortega F.J.
      • Mercader J.M.
      • Moreno-Navarrete J.M.
      • Rovira O.
      • Guerra E.
      • Esteve E.
      • et al.
      Profiling of circulating microRNAs reveals common microRNAs linked to type 2 diabetes that change with insulin sensitization.
      ,
      • Flowers E.
      • Aouizerat B.E.
      • Abbasi F.
      • Lamendola C.
      • Grove K.M.
      • Fukuoka Y.
      • et al.
      Circulating microRNA-320a and microRNA-486 predict thiazolidinedione response: moving towards precision health for diabetes prevention.
      ,
      • Parrizas M.
      • Brugnara L.
      • Esteban Y.
      • Gonzalez-Franquesa A.
      • Canivell S.
      • Murillo S.
      • et al.
      Circulating miR-192 and miR-193b are markers of prediabetes and are modulated by an exercise intervention.
      ], suggesting a new potential use of microRNA as predictive biomarkers for monitoring therapy response and moving towards precision medicine for IR prevention.
      Fig. 2
      Fig. 2Biogenesis and secretion of microRNAs and exosomes and their role for insulin sensitivity.
      Pri-miRNAs are transcribed in the nucleus by RNA polymerase II ① and later cleaved by Drosha to generate pre-miRNAs ②, which are exported into the cytoplasm by Exportin-5 ③ and here processed by Dicer into 21–24 nucleotide duplexes ④. The duplex unwinds ⑤ and mature miRNA either assembles into RISC and binds the mRNA target inducing translational repression ⑥ or is released to the extracellular milieu. Released miRNA can be coupled with RNA-binding proteins, such as Argonaute-2 ⑦, and lipoproteins ⑧ or selectively incorporated either into extracellular microvesicles (size 100–1000 nm), formed by the outward budding of the plasma membrane ⑨, or Exo (size 20–140 nm), derived from the endosomal route ⑩ [
      • Nigi L.
      • Grieco G.E.
      • Ventriglia G.
      • Brusco N.
      • Mancarella F.
      • Formichi C.
      • et al.
      MicroRNAs as regulators of insulin signaling: research updates and potential therapeutic perspectives in type 2 diabetes.
      ].
      After invagination of the plasma membrane ①, early endosomes mature into late endosome leading to inward budding of the endosomal membrane and formation of intraluminal vesicles ②. This leads to the formation of multivesicular bodies ③, which can either fuse with lysosomes to be degraded ④, or fuse with the plasma membrane and release Exo into the extracellular environment ⑤ [
      • Safdar A.
      • Saleem A.
      • Tarnopolsky M.A.
      The potential of endurance exercise-derived exosomes to treat metabolic diseases.
      ].
      Several miRNAs modulate insulin signaling and inflammatory pathways, and can be altered in T2DM and insulin resistance [
      • Nigi L.
      • Grieco G.E.
      • Ventriglia G.
      • Brusco N.
      • Mancarella F.
      • Formichi C.
      • et al.
      MicroRNAs as regulators of insulin signaling: research updates and potential therapeutic perspectives in type 2 diabetes.
      ,
      • Pan Y.
      • Hui X.
      • Hoo R.L.C.
      • Ye D.
      • Chan C.Y.C.
      • Feng T.
      • et al.
      Adipocyte-secreted exosomal microRNA-34a inhibits M2 macrophage polarization to promote obesity-induced adipose inflammation.
      ]. The miR-34a, carried by WAT-derived Exo, inhibits polarization of macrophages from pro-inflammatory (M1) to anti-inflammatory (M2) and its levels are increased in VAT of obese humans [
      • Pan Y.
      • Hui X.
      • Hoo R.L.C.
      • Ye D.
      • Chan C.Y.C.
      • Feng T.
      • et al.
      Adipocyte-secreted exosomal microRNA-34a inhibits M2 macrophage polarization to promote obesity-induced adipose inflammation.
      ]. Several miRNAs (miR-205-5p, miR-21 and miR-26b) control expression and activity of PTEN and SHIP2 and are downregulated in mouse models and humans with T2DM and obesity. The miR-146a and miR-122 inhibit expression of PTP1B and PTPN1, which inhibit insulin signaling. Levels of miRNAs targeting IRS1, IRS2, IR and CAV-1 are upregulated in liver and SkM of obese mice as well as in SkM (miR-29a, miR-29c and miR-135a) and serum (miR-144) of people with T2DM [
      • Nigi L.
      • Grieco G.E.
      • Ventriglia G.
      • Brusco N.
      • Mancarella F.
      • Formichi C.
      • et al.
      MicroRNAs as regulators of insulin signaling: research updates and potential therapeutic perspectives in type 2 diabetes.
      ,
      • Karolina D.S.
      • Armugam A.
      • Tavintharan S.
      • Wong M.T.
      • Lim S.C.
      • Sum C.F.
      • et al.
      MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in type 2 diabetes mellitus.
      ]. Finally, miR-20b-5p targets Akt and it is upregulated in circulating Exo of people with T2DM [
      • Katayama M.
      • Wiklander O.P.B.
      • Fritz T.
      • Caidahl K.
      • El-Andaloussi S.
      • Zierath J.R.
      • et al.
      Circulating exosomal miR-20b-5p is elevated in type 2 diabetes and could impair insulin action in human skeletal muscle.
      ]. Besides miRNAs, circulating and WAT-derived Exo from obese persons carry proteins implicated in WAT inflammation and insulin resistance (TGFB1, OGN, SDCB1) [
      • Camino T.
      • Lago-Baameiro N.
      • Bravo S.B.
      • Molares-Vila A.
      • Sueiro A.
      • Couto I.
      • et al.
      Human obese white adipose tissue sheds depot-specific extracellular vesicles and reveals candidate biomarkers for monitoring obesity and its comorbidities.
      ]; instead circulating Exo released after exercise from insulin-resistant individuals carry proteins related to insulin sensitivity (PLC, PKA, ERK/MAPK pathways) and oxidative metabolism (CAT, PRDX1/2, SOD2, G6PD2) [
      • Apostolopoulou M.
      • Mastrototaro L.
      • Hartwig S.
      • Pesta D.
      • Straßburger K.
      • de Filippo E.
      • et al.
      Metabolic responsiveness to training depends on insulin sensitivity and protein content of exosomes in insulin resistant males.
      ].
      Biogenesis and secretion pathways of miRNA are depicted by red numbers, those of Exo by green numbers. The miRNAs (and extracellular proteins) identified in serum/plasma are reported in red, those released by WAT in yellow, those found intracellularly in insulin-target tissues in black colors. The arrows indicate upregulation or downregulation in obesity and T2DM.
      Ago2 = argonaute 2; Akt = protein kinase B; C = circulating; CAT = catalase; CAV-1 = caveolin 1; E = exosome; ERK/MAPK = extracellular signal-regulated kinases/mitogen-activated protein kinases; Exo = exosome; G6PD2 = glucose-6-phosphate 1-dehydrogenase 2; HDL = high density lipoproteins; IR = insulin receptor; IRS1 = insulin receptor substrate 1; L = liver; miRNA = microRNA; MV = microvesicles; MVB = multivesicular bodies; OGN = mimecan; PKA = protein kinase A; PLC = phospholipase C; Pol = polymerase; PRDX1/2 = peroxiredoxin 1/2; PTEN = phosphatase and tensin homolog; PTP1B = Protein-tyrosine phosphatase 1B; PTPN1 = Protein Tyrosine Phosphatase, Non-receptor type 1; SDCB1 = syntenin 1; SHIP2 SH2 domain-containing inositol 5′-phosphatase 2; SkM = skeletal muscle; SOD2 = superoxide dismutase 2; T2DM = type 2 diabetes mellitus; TGFB1 = transforming growth factor β 1; VAT = visceral adipose tissue; WAT = adipose tissue.
      Table 1Human studies aimed to assess the role of microRNAs and exosomes in the development of insulin resistance.
      microRNA/ExoCohort N total (males), BMI (kg/m2)Tissue of originCorrelation with metabolic featuresPossible molecular mechanismTarget tissue/cellsRef
      miR-34aWATLean, 29(0); 21 ± 1

      Overweight/Obe, 24(0); 26 ± 2
      WATHOMA-IR↑,

      IL-6↑,

      TNFα↑
      Klf4↓,

      M1 macrophage↑
      [
      • Pan Y.
      • Hui X.
      • Hoo R.L.C.
      • Ye D.
      • Chan C.Y.C.
      • Feng T.
      • et al.
      Adipocyte-secreted exosomal microRNA-34a inhibits M2 macrophage polarization to promote obesity-induced adipose inflammation.
      ]
      miR-144c

      miR-365c

      miR-32c

      miR-451c

      miR-150c

      miR-409-3pc

      miR-151-5pc

      miR-374bc

      miR-193bc

      miR-122c

      miR-382c

      miR-136c

      miR-34ac

      miR-335c

      miR-423-5pc

      miR-19bc

      miR-152c

      miR-484c

      miR-22-5pc

      miR-22c
      Lean, 12(0); 23 ± 3

      Obe IS, 11(0); 43 ± 4

      Obe IR, 19(0); 45 ± 4

      Obe T2DM, 15(0); 44 ± 4
      PlasmaFasting lipids,

      HbA1c,

      HOMA-IR,

      Glucose,

      Insulin
      [
      • Jones A.
      • Danielson K.M.
      • Benton M.C.
      • Ziegler O.
      • Shah R.
      • Stubbs R.S.
      • et al.
      miRNA signatures of insulin resistance in obesity.
      ]
      miR-122c

      mir-192c
      Elderly, 2317(1010); 28 ± 5

      Youth, 90(36); 34 ± 10
      PlasmaInsulin ↑,

      HOMA-IR↑,

      VAT↑,

      BMI↑,

      TG:HDL↑,

      Adiponectin↓
      AMPK

      MAPK
      [
      • Shah R.
      • Murthy V.
      • Pacold M.
      • Danielson K.
      • Tanriverdi K.
      • Larson M.G.
      • et al.
      Extracellular RNAs are associated with insulin resistance and metabolic phenotypes.
      ]
      Exo

      miR122-5p e

      miR34a-5p e

      miR155-5p e ↑ miR146b-3 e
      NAFLD, 20(14); 30 ± 1

      Healthy, 20(13); 17 ± 0
      BMI↑

      ALT↑

      AST↑

      UA↑
      [
      • Zhou X.
      • Huang K.
      • Jia J.
      • Ni Y.
      • Yuan J.
      • Liang X.
      • et al.
      Exosomal miRNAs profile in children’s nonalcoholic fatty liver disease and the correlation with transaminase and uric acid.
      ]
      Exo↔

      miR-20b-5pe
      T2DM, 21(21); 29 ± 1

      NGT, 20(20); 29 ± 1
      SerumIn NGT:

      2 h Glucose↓

      Fat mass↓
      STAT3↓

      AKTIP↓

      GS↓
      hSkMc[
      • Katayama M.
      • Wiklander O.P.B.
      • Fritz T.
      • Caidahl K.
      • El-Andaloussi S.
      • Zierath J.R.
      • et al.
      Circulating exosomal miR-20b-5p is elevated in type 2 diabetes and could impair insulin action in human skeletal muscle.
      ]
      miR-144c

      miR-146ac

      miR-150c↑↓

      miR-182c↑↓

      miR-192c

      miR-29ac

      miR-30dc↑↓

      miR-320ac
      T2DM, 21(21); 24–28

      IGT, 14(14); 24–30

      Healthy, 15(15); 22–24
      SerumIRS1↓

      IRS1_Tyr612
      Serum[
      • Karolina D.S.
      • Armugam A.
      • Tavintharan S.
      • Wong M.T.
      • Lim S.C.
      • Sum C.F.
      • et al.
      MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in type 2 diabetes mellitus.
      ]
      Obe vs healthy:

      miR-122-5pe

      miR-193b-5pe

      miR-26b-3pe

      miR-4449e

      miR-1290e

      miR-193a-5pe

      miR-183-5pe

      miR-126-5pe

      miR-4461e

      miR-1273ae

      miR-6739-5pe

      miR-1273g-3pe

      miR-6739-5pe

      miR-1273g-3pe

      miR-4284e

      miR-6751-3pe

      miR-4485-5pe

      miR-8485e

      miR-1285-3pe

      miR-20a-5pe

      Pre- vs post-BS:

      miR-424-5pe
      Healthy, 18(5); 22 ± 1

      Obe, 16(7); 40 ± 5



      Obe pre-BS, 12; 40 ± 5

      Obe post-BS, 12; 31 ± 5
      SerumWNT,

      Insulin signaling
      [
      • Bae Y.U.
      • Kim Y.
      • Lee H.
      • Kim H.
      • Jeon J.S.
      • Noh H.
      • et al.
      Bariatric surgery alters microRNA content of circulating exosomes in patients with obesity.
      ]
      Healthy vs T2DM/T2DM-Met:

      miR-122-5pc

      miR-192-5pc

      miR-323b-3pc

      miR-203a-3pe

      miR-10b-5pe

      miR-142-3pe

      miR-126-3pe

      T2DM-Met vs T2DM:

      miR-15a-5pe

      miR-30c-5pe

      miR-424-5pe
      Healthy, 39(16); 28 ± 5

      T2DM, 10(6); 24 ± 5

      T2DM-Met, 31(12); 29 ± 5
      Serum[
      • Ghai V.
      • Kim T.K.
      • Etheridge A.
      • Nielsen T.
      • Hansen T.
      • Pedersen O.
      • et al.
      Extracellular vesicle encapsulated microRNAs in patients with type 2 diabetes are affected by metformin treatment.
      ]
      T2D vs NGT

      miR-140-5pc

      miR-222c

      miR-142-3pc

      miR-423-5pc

      miR-125bc

      miR192c

      miR-195c

      miR-130bc

      miR-532-5pc

      miR-126c

      Met vs placebo

      miR-140-5pc

      miR-222↓

      miR-142-3pc

      miR192c
      NGT, 35(35); 25 ± 2

      NGT-Obe, 10(10); 32 ± 2

      T2DM, 30(30); 27 ± 2

      T2DM-Obe, 18(18); 33 ± 3



      T2DM-Met, 17(8); 36 ± 6 T2DM-placebo, 18(8); 34 ± 8
      SerumFPG

      HbA1c

      TG

      BMI
      [
      • Ortega F.J.
      • Mercader J.M.
      • Moreno-Navarrete J.M.
      • Rovira O.
      • Guerra E.
      • Esteve E.
      • et al.
      Profiling of circulating microRNAs reveals common microRNAs linked to type 2 diabetes that change with insulin sensitization.
      ]
      IR vs IS:

      miR-193bc

      miR-22-3pc

      miR-320ac

      miR-375c

      miR-486-5pc

      IR-TZD-R vs IR-TZD-NR:

      miR-20b-5pc

      miR-21-5pc

      miR-214-3pc

      miR-22-3pc

      miR-320ac

      miR-486-5pc
      IS, 18(5); 26 ± 4

      IR, 75(19); 34 ± 7



      IR-TZD-R, 36(13); 32 ± 6

      IR-TZD-NR, 11(5); 31 ± 2
      Serum[
      • Flowers E.
      • Aouizerat B.E.
      • Abbasi F.
      • Lamendola C.
      • Grove K.M.
      • Fukuoka Y.
      • et al.
      Circulating microRNA-320a and microRNA-486 predict thiazolidinedione response: moving towards precision health for diabetes prevention.
      ]
      Cohort 1

      Lean vs pre:

      miR-191c↑ (IFG)

      miR-15bc↑ (IFG)

      miR-128c↓ (IFG)

      miR-125a-5pc↓ (IFG)

      miR-150c

      miR-192c

      miR-193bc

      Lean vs T2DM:

      miR-191c

      miR-139-5pc

      miR-21c

      Cohort 2

      Lean vs pre:

      miR-150c

      miR-192c

      miR-193bc

      Baseline vs post-ex:

      miR-150c↓ (Pre)

      miR-192c↓(Pre)

      miR-193bc↓(Pre)
      Cohort 1:

      Lean, 29(29); 29 ± 1

      Pre:

      - IFG, 22(22); 29 ± 1

      - IGT, 21(21); 29 ± 1

      T2DM, 20(20); 30 ± 1



      Cohort 2:

      Lean, 12(12); 25 ± 0.5

      Pre, 6(6); 27 ± 1
      SerumTG↑

      FLI↑
      [
      • Parrizas M.
      • Brugnara L.
      • Esteban Y.
      • Gonzalez-Franquesa A.
      • Canivell S.
      • Murillo S.
      • et al.
      Circulating miR-192 and miR-193b are markers of prediabetes and are modulated by an exercise intervention.
      ]
      miR-148be

      miR-4269e

      miR-23be

      miR-4429e
      Lean, 5(0); 22–25

      Obe, 7(0); 33–50
      VATACVR2B (TGF-β signaling)A549 cells[
      • Ferrante S.C.
      • Nadler E.P.
      • Pillai D.K.
      • Hubal M.J.
      • Wang Z.
      • Wang J.M.
      • et al.
      Adipocyte-derived exosomal miRNAs: a novel mechanism for obesity-related disease.
      ]
      ExoLean, 16(15); 26 ± 4SAT, VATVAT-Exo:

      ALT↑

      AST↑

      γGT↑

      SAT-Exo:

      Waist circumference↓

      Metabolic syndrome↓
      p-AKT↓↑Hepatocytes[
      • Kranendonk M.E.
      • Visseren F.L.
      • van Herwaarden J.A.
      • Nolte-’t Hoen E.N.
      • de Jager W.
      • Wauben M.H.
      • et al.
      Effect of extracellular vesicles of human adipose tissue on insulin signaling in liver and muscle cells.
      ]
      Exo↑

      miR-192e

      miR-122e
      Advanced NAFLD, 3 Early NAFLD, 3SerumTGF-β

      αSMA

      Col1a1
      HSC[
      • Lee Y.S.
      • Kim S.Y.
      • Ko E.
      • Lee J.H.
      • Yi H.S.
      • Yoo Y.J.
      • et al.
      Exosomes derived from palmitic acid-treated hepatocytes induce fibrotic activation of hepatic stellate cells.
      ]
      EV↑Obe NASH,16; >30

      Obe steatosis,16; >30

      Obe, 11; >30
      PlasmaC16↑

      S1P↑
      Liver[
      • Kakazu E.
      • Mauer A.S.
      • Yin M.
      • Malhi H.
      Hepatocytes release ceramide-enriched pro-inflammatory extracellular vesicles in an IRE1alpha-dependent manner.
      ]
      EV↑Cross-sectional cohort 1:

      NGT,16(7); 25 ± 4

      T2DM,22(8); 39 ± 9

      Cross-sectional cohort 2:

      NGT,30(10); 35 ± 7

      T2DM,30(5); 35 ± 6

      Longitudinal cohort:

      No-NoT2DM, 19(5); 33 ± 6

      No-T2DM, 19(7); 35 ± 8

      Pre-T2DM, 20(6); 36 ± 9
      SerumHOMA-IR↑

      HOMA-β↓
      LeukocytesInflammation pathway↑

      Oxidative stress pathway and gene↓
      [
      • Freeman D.W.
      • Noren Hooten N.
      • Eitan E.
      • Green J.
      • Mode N.A.
      • Bodogai M.
      • et al.
      Altered extracellular vesicle concentration, cargo, and function in diabetes.
      ]
      EV↑NGT, 21; <27

      Overweight, 7; 27–30;

      Obe, 20; >30
      PlasmaBMI↑[
      • Amosse J.
      • Durcin M.
      • Malloci M.
      • Vergori L.
      • Fleury A.
      • Gagnadoux F.
      • et al.
      Phenotyping of circulating extracellular vesicles (EVs) in obesity identifies large EVs as functional conveyors of macrophage migration inhibitory factor.
      ]
      Abbreviations: ACVR2B = Activin A Receptor Type 2B; AKTIP = Akt interacting protein; ALT = alanine aminotransferase; AMPK = 5′ adenosine monophosphate-activated protein kinaseα; SMA = α smooth muscle actin; AST = aspartate aminotransferase; BMI = body mass index; BS = bariatric surgery; c = circulating; Col1a1 = Collagen Type I Alpha 1 Chain; EV = extracellular vesicles; e = exosome; Ex = exercise; Exo = exosome; FLI = fatty liver index; FPG = fasting plasma glucose; γGT = gamma glutamyltransferase; GS = glycogen synthase; HDL = high density lipoprotein; HSC = hepatic stellate cells; hSkMc = human skeletal muscle cells; IFG = impaired fasting glucose; IGT = impaired glucose tolerant; IL-6 = interleukin 6; IR = insulin resistant; IS = insulin sensitive;; Klf4 = Krüppe-likel factor 4; MAPK = mitogen-activated protein kinase; NAFLD = non-alcoholic fatty liver disease; NGT = normal glucose tolerant; Obe = obese; Pre = pre-diabetic; S1P = sphingosine-1-phosphate; SAT = subcutaneous adipose tissue; STAT3 = Signal transducer and activator of transcription 3; T2DM = type 2 diabetes mellitus; TG = triglycerides; TGF-β = tumor growth factor-β; TNFα = tumor necrosis factor α; TZD-R = thiazolidinedione responders; TZD-NR = TZD non-responders; UA = uric acid; VAT = visceral AT.

      3.3 Exosomes

      There is growing evidence that extracellular vesicles (EV), in particular Exo derived from endosomes (Table 1, Fig. 2), are also involved in the pathophysiology of obesity and IR. WAT in vivo secrete Exo containing adipocyte-molecules, whose expression and abundance are regulated by obesity [
      • Ferrante S.C.
      • Nadler E.P.
      • Pillai D.K.
      • Hubal M.J.
      • Wang Z.
      • Wang J.M.
      • et al.
      Adipocyte-derived exosomal miRNAs: a novel mechanism for obesity-related disease.
      ]. The metabolic effects resulting from EV released by WAT and WAT-macrophages of obese mice have been investigated by injecting them in lean mice, where they cause IR, glucose intolerance and inflammation [
      • Deng Z.B.
      • Poliakov A.
      • Hardy R.W.
      • Clements R.
      • Liu C.
      • Liu Y.
      • et al.
      Adipose tissue exosome-like vesicles mediate activation of macrophage-induced insulin resistance.
      ,
      • Ying W.
      • Riopel M.
      • Bandyopadhyay G.
      • Dong Y.
      • Birmingham A.
      • Seo J.B.
      • et al.
      Adipose tissue macrophage-derived exosomal miRNAs can modulate in vivo and in vitro insulin sensitivity.
      ]. Conversely, anti-inflammatory M2 macrophages secrete Exo, which improve glucose tolerance and insulin sensitivity in obese mice [
      • Ying W.
      • Gao H.
      • Dos Reis F.C.G.
      • Bandyopadhyay G.
      • Ofrecio J.M.
      • Luo Z.
      • et al.
      MiR-690, an exosomal-derived miRNA from M2-polarized macrophages, improves insulin sensitivity in obese mice.
      ]. The deletion of AMPKα1 in WAT enhances the release of Exo, which exacerbate the lipid deposition and inflammation when injected in hepatocytes, suggesting a role of Exo in the communication between WAT and liver [
      • Yan C.
      • Tian X.
      • Li J.
      • Liu D.
      • Ye D.
      • Xie Z.
      • et al.
      A high-fat diet attenuates AMPK alpha1 in adipocytes to induce exosome shedding and nonalcoholic fatty liver development in vivo.
      ]. Exo released from WAT of lean persons could also impair insulin signaling in hepatocytes depending on their adipokine content [
      • Kranendonk M.E.
      • Visseren F.L.
      • van Herwaarden J.A.
      • Nolte-’t Hoen E.N.
      • de Jager W.
      • Wauben M.H.
      • et al.
      Effect of extracellular vesicles of human adipose tissue on insulin signaling in liver and muscle cells.
      ]. The number of circulating EV is increased in individuals with obesity, T2DM and NAFLD [
      • Lee Y.S.
      • Kim S.Y.
      • Ko E.
      • Lee J.H.
      • Yi H.S.
      • Yoo Y.J.
      • et al.
      Exosomes derived from palmitic acid-treated hepatocytes induce fibrotic activation of hepatic stellate cells.
      ,
      • Kakazu E.
      • Mauer A.S.
      • Yin M.
      • Malhi H.
      Hepatocytes release ceramide-enriched pro-inflammatory extracellular vesicles in an IRE1alpha-dependent manner.
      ,
      • Freeman D.W.
      • Noren Hooten N.
      • Eitan E.
      • Green J.
      • Mode N.A.
      • Bodogai M.
      • et al.
      Altered extracellular vesicle concentration, cargo, and function in diabetes.
      ,
      • Amosse J.
      • Durcin M.
      • Malloci M.
      • Vergori L.
      • Fleury A.
      • Gagnadoux F.
      • et al.
      Phenotyping of circulating extracellular vesicles (EVs) in obesity identifies large EVs as functional conveyors of macrophage migration inhibitory factor.
      ], allowing exploitation of EV as diagnostic tools as well as mediators of IR. A recent study analyzed the proteomic profiling of EV and revealed that plasma and WAT-derived EV shed by obese persons carry proteins implicated in WAT inflammation and IR [
      • Camino T.
      • Lago-Baameiro N.
      • Bravo S.B.
      • Molares-Vila A.
      • Sueiro A.
      • Couto I.
      • et al.
      Human obese white adipose tissue sheds depot-specific extracellular vesicles and reveals candidate biomarkers for monitoring obesity and its comorbidities.
      ]. Moreover, obesity-related circulating Exo can impair insulin signaling pathways, increase triglycerides (TG) content and decrease FGF21 secretion when they are injected in hepatocytes [
      • Afrisham R.
      • Sadegh-Nejadi S.
      • Meshkani R.
      • Emamgholipour S.
      • Paknejad M.
      Effect of circulating exosomes derived from normal-weight and obese women on gluconeogenesis, glycogenesis, lipogenesis and secretion of FGF21 and fetuin A in HepG2 cells.
      ]. Finally, 12-w exercise increases the number of circulating Exo, along with insulin sensitivity, and differently affects Exo proteome in nondiabetic and diabetic humans suggesting a role for Exo as mediators of the individual metabolic exercise responsiveness [
      • Apostolopoulou M.
      • Mastrototaro L.
      • Hartwig S.
      • Pesta D.
      • Straßburger K.
      • de Filippo E.
      • et al.
      Metabolic responsiveness to training depends on insulin sensitivity and protein content of exosomes in insulin resistant males.
      ].

      4. Strategies to improve insulin resistance

      Current guidelines primarily advise lifestyle modification (healthy nutrition and exercise) and BW loss to ameliorate insulin sensitivity and thereby to prevent or treat T2DM. Nevertheless, also several antihyperglycemic drugs are required to treat T2DM and some can directly or indirectly improve IR.

      4.1 Lifestyle modification (LSM)

      While hypocaloric nutrition and exercise training remain key to the prevention and treatment of obesity and T2DM, only a limited number of people adheres to long-term LSM (Table 2). A recent meta-analysis of 19 LSM studies reported that diet with physical activity achieve a greater diabetes risk reduction than either strategy alone. However, their effects decline over time, therefore maintenance strategies are needed for prolonged effects [
      • Haw J.S.
      • Galaviz K.I.
      • Straus A.N.
      • Kowalski A.J.
      • Magee M.J.
      • Weber M.B.
      • et al.
      Long-term sustainability of diabetes prevention approaches: a systematic review and meta-analysis of randomized clinical trials.
      ]. Caloric restriction (CR) for 16-wk in obese individuals enhances whole-body insulin sensitivity, measured by HEC, but it does not improve SkM mitochondrial oxidative capacity and intramyocellular lipid (IMCL) content. However, CR reduces protein content of thioredoxin-interacting protein, which partly explains the increase in insulin-stimulated glucose disposal [
      • Johnson M.L.
      • Distelmaier K.
      • Lanza I.R.
      • Irving B.A.
      • Robinson M.M.
      • Konopka A.R.
      • et al.
      Mechanism by which caloric restriction improves insulin sensitivity in sedentary obese adults.
      ]. In line, a 12-wk moderate hypocaloric diet reduces BW, fasting EGP, hyperglycemia and IHL in obesity and T2DM, despite no decrease in IMCL [
      • Petersen K.F.
      • Dufour S.
      • Befroy D.
      • Lehrke M.
      • Hendler R.E.
      • Shulman G.I.
      Reversal of nonalcoholic hepatic steatosis, hepatic insulin resistance, and hyperglycemia by moderate weight reduction in patients with type 2 diabetes.
      ]. In contrast, a short-term very-low calorie diet (VLCD, 700 kcal/day) reduces IMCL and enhances muscle glucose uptake [
      • Lara-Castro C.
      • Newcomer B.R.
      • Rowell J.
      • Wallace P.
      • Shaughnessy S.M.
      • Munoz A.J.
      • et al.
      Effects of short-term very low-calorie diet on intramyocellular lipid and insulin sensitivity in nondiabetic and type 2 diabetic subjects.
      ]. Studies in larger cohorts demonstrated that a VLCD normalizes IHL without improving metabolic control and insulin sensitivity [
      • Taylor R.
      • Al-Mrabeh A.
      • Zhyzhneuskaya S.
      • Peters C.
      • Barnes A.C.
      • Aribisala B.S.
      • et al.
      Remission of human type 2 diabetes requires decrease in liver and pancreas fat content but is dependent upon capacity for beta cell recovery.
      ], while a VLCD followed by LCD reduces IR, inflammation, ER stress and ROS production in obese people [
      • Lopez-Domenech S.
      • Abad-Jimenez Z.
      • Iannantuoni F.
      • de Maranon A.M.
      • Rovira-Llopis S.
      • Morillas C.
      • et al.
      Moderate weight loss attenuates chronic endoplasmic reticulum stress and mitochondrial dysfunction in human obesity.
      ]. Other studies investigating the different macronutrients showed improvements of insulin sensitivity in healthy individuals after a 4-week daily intake of sodium butyrate [
      • Bouter K.
      • Bakker G.J.
      • Levin E.
      • Hartstra A.V.
      • Kootte R.S.
      • Udayappan S.D.
      • et al.
      Differential metabolic effects of oral butyrate treatment in lean versus metabolic syndrome subjects.
      ] or resistant starch [
      • Robertson M.D.
      • Bickerton A.S.
      • Dennis A.L.
      • Vidal H.
      • Frayn K.N.
      Insulin-sensitizing effects of dietary resistant starch and effects on skeletal muscle and adipose tissue metabolism.
      ], as measured via HEC. Isocaloric protein-rich diets reduces IHL, markers of IR and inflammation [
      • Markova M.
      • Pivovarova O.
      • Hornemann S.
      • Sucher S.
      • Frahnow T.
      • Wegner K.
      • et al.
      Isocaloric diets high in animal or plant protein reduce liver fat and inflammation in individuals with type 2 diabetes.
      ]. Supplementation with long chain ω3-FA decreases WAT and systemic inflammation, resulting in enhanced glucose disposal [
      • Hernandez J.D.
      • Li T.
      • Rau C.M.
      • LeSuer W.E.
      • Wang P.
      • Coletta D.K.
      • et al.
      omega-3PUFA supplementation ameliorates adipose tissue inflammation and insulin-stimulated glucose disposal in subjects with obesity: a potential role for apolipoprotein E.
      ].
      Table 2Effects of lifestyle interventions for >4-wk in crossover and before-after studies demonstrating improvements of insulin resistance.
      StudyDesign; intervention, duration; methodCohort

      N total (males), BMI (kg/m2)
      Changes in insulin sensitivityMetabolic changesMolecular mechanism
      Karusheva Y et al. [
      • Karusheva Y.
      • Koessler T.
      • Strassburger K.
      • Markgraf D.
      • Mastrototaro L.
      • Jelenik T.
      • et al.
      Short-term dietary reduction of branched-chain amino acids reduces meal-induced insulin secretion and modifies microbiome composition in type 2 diabetes: a randomized controlled crossover trial.
      ]
      R/Co;

      Diet; BCAA- vs BCAA+;

      4-wk;

      HEC, MTT
      T2DM, 12(8); 31 ± 3OGIS↑
      Significant effect between groups after intervention.


      PREDIM↑
      Significant effect between groups after intervention.


      M/I↔

      iEGP↔

      iFFA↔
      BW↓
      Significant effect of intervention vs baseline.
      FGF21s↑
      Significant effect between groups after intervention.


      pAKTAT
      Significant effect between groups after intervention.


      pmTORAT
      Significant effect between groups after intervention.


      RCRAT
      Significant effect between groups after intervention.


      Firmicutes↓
      Significant effect between groups after intervention.


      Baceroidetes↑
      Significant effect between groups after intervention.
      Johnson ML et al. [
      • Johnson M.L.
      • Distelmaier K.
      • Lanza I.R.
      • Irving B.A.
      • Robinson M.M.
      • Konopka A.R.
      • et al.
      Mechanism by which caloric restriction improves insulin sensitivity in sedentary obese adults.
      ]
      B/A;

      Diet;

      CR;

      16-wk;

      EPC
      Obe, 11; 35 ± 1Rd↑
      Significant effect of intervention vs baseline.


      iEGP↔
      BW↓
      Significant effect of intervention vs baseline.


      FPI↓
      Significant effect of intervention vs baseline.
      mt function↔

      IMCL↔

      TXNIP↓
      Significant effect of intervention vs baseline.
      Petersen K et al. [
      • Petersen K.F.
      • Dufour S.
      • Befroy D.
      • Lehrke M.
      • Hendler R.E.
      • Shulman G.I.
      Reversal of nonalcoholic hepatic steatosis, hepatic insulin resistance, and hyperglycemia by moderate weight reduction in patients with type 2 diabetes.
      ]
      B/A;

      Diet;

      LCD;

      12-wk;

      HEC
      T2DM, 8(5); 30 ± 1iEGP↑
      Significant effect of intervention vs baseline.
      BW↓
      Significant effect of intervention vs baseline.


      IHL↓
      Significant effect of intervention vs baseline.
      Taylor R et al. [
      • Taylor R.
      • Al-Mrabeh A.
      • Zhyzhneuskaya S.
      • Peters C.
      • Barnes A.C.
      • Aribisala B.S.
      • et al.
      Remission of human type 2 diabetes requires decrease in liver and pancreas fat content but is dependent upon capacity for beta cell recovery.
      ]
      B/A;

      Diet; LCD;

      4-mo;

      SISTA
      T2DM-R, 40(23); 35 ± 1

      T2DM-NR, 18(9); 36 ± 1
      T2DM-R:

      β-cells function↑
      Significant effect of intervention vs baseline.
      Both:

      BW↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      Liver fat↓
      Significant effect of intervention vs baseline.


      TGP
      Significant effect of intervention vs baseline.


      Pancreas fat↓
      Significant effect of intervention vs baseline.


      FPI↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      T2DM-R:

      FPG↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      VLDL1-TG↓
      Significant effect of intervention vs baseline.


      HbA1c↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.
      López-Domènech S et al. [
      • Lopez-Domenech S.
      • Abad-Jimenez Z.
      • Iannantuoni F.
      • de Maranon A.M.
      • Rovira-Llopis S.
      • Morillas C.
      • et al.
      Moderate weight loss attenuates chronic endoplasmic reticulum stress and mitochondrial dysfunction in human obesity.
      ]
      B/A;

      Diet; VLCD, 6-wk → LCD, 18-wk;

      HOMA-IR
      Obe, 64(18); >35HOMA-IR↓
      Significant effect of intervention vs baseline.
      BW↓
      Significant effect of intervention vs baseline.


      Systemic inflammation↓
      Significant effect of intervention vs baseline.


      HbA1c↓
      Significant effect of intervention vs baseline.
      ER stress↓
      Significant effect of intervention vs baseline.


      ROS production↓
      Significant effect of intervention vs baseline.


      GPX1↑
      Significant effect of intervention vs baseline.
      Bouter KEC et al., 2018 [
      • Bouter K.
      • Bakker G.J.
      • Levin E.
      • Hartstra A.V.
      • Kootte R.S.
      • Udayappan S.D.
      • et al.
      Differential metabolic effects of oral butyrate treatment in lean versus metabolic syndrome subjects.
      ]
      B/A;

      Diet; Na butyrate 4 g/d;

      4-wk;

      HEC
      Healthy, 9(9); 22 ± 2

      MS, 10(10); 33 ± 4
      Healthy:

      Rd↑
      Significant effect of intervention vs baseline.


      iEGP↑
      Significant effect of intervention vs baseline.
      MS:

      PropionateP
      Significant effect of intervention vs baseline.


      SCFA F
      Significant effect of intervention vs baseline.


      TCP
      Significant effect of intervention vs baseline.


      LDLP
      Significant effect of intervention vs baseline.
      Healthy:

      Lachnospiraceae
      Significant effect of intervention vs baseline.


      Bacteroides
      Significant effect of intervention vs baseline.


      MS:

      Coriobacteriaceae
      Significant effect of intervention vs baseline.


      Clostridiales
      Significant effect of intervention vs baseline.
      Robertson MD et al. [
      • Robertson M.D.
      • Bickerton A.S.
      • Dennis A.L.
      • Vidal H.
      • Frayn K.N.
      Insulin-sensitizing effects of dietary resistant starch and effects on skeletal muscle and adipose tissue metabolism.
      ]
      R/Co;

      Diet; Resistant starch, 30 g/d vs placebo;

      4-wk;

      HEC; MTT
      Healthy, 10(4); 23 ± 1HOMA-IR↔

      HOMA-β↔

      M/I↑
      Significant effect between groups after intervention.
      Lean mass↓
      Significant effect between groups after intervention.


      Glucose uptakeAT,SkM
      Significant effect between groups after intervention.


      SCFAP
      Significant effect between groups after intervention.


      NEFAAT
      Significant effect between groups after intervention.


      FPGh↑
      Significant effect between groups after intervention.


      PPI↓
      Significant effect between groups after intervention.
      IRS1AT
      Significant effect between groups after intervention.


      HSLAT
      Significant effect between groups after intervention.
      Hernandez JD et al. [
      • Hernandez J.D.
      • Li T.
      • Rau C.M.
      • LeSuer W.E.
      • Wang P.
      • Coletta D.K.
      • et al.
      omega-3PUFA supplementation ameliorates adipose tissue inflammation and insulin-stimulated glucose disposal in subjects with obesity: a potential role for apolipoprotein E.
      ]
      B/A;

      Diet;

      ω3-PUFA;

      3-mo;

      HEC
      IR, 13(3); 39 ± 2iEGP↔

      Rd↑
      Significant effect of intervention vs baseline.
      FFA↓
      Significant effect of intervention vs baseline.


      Pro-inflammatory cytokines↓
      Significant effect of intervention vs baseline.


      Adiponectin↑
      Significant effect of intervention vs baseline.
      Pro-inflammatory macrophage markersWAT
      Significant effect of intervention vs baseline.


      ApoEWAT
      Significant effect of intervention vs baseline.


      ApoC1WAT
      Significant effect of intervention vs baseline.
      Lee S et al. [
      • Lee S.
      • Gulseth H.L.
      • Langleite T.M.
      • Norheim F.
      • Olsen T.
      • Refsum H.
      • et al.
      Branched-chain amino acid metabolism, insulin sensitivity and liver fat response to exercise training in sedentary dysglycaemic and normoglycaemic men.
      ]
      B/A;

      Endurance exercise;

      12-wk;

      HEC
      IR, 13(13); 29 ± 2

      Healthy, 13(13); 24 ± 2
      GIR↑
      Significant effect of intervention vs baseline.
      BW↓
      Significant effect of intervention vs baseline.
      (IR)

      Total fat↓
      Significant effect of intervention vs baseline.


      VO2max
      Significant effect of intervention vs baseline.
      BCAA catabolic pathway↑
      Significant effect of intervention vs baseline.


      mRNA BCAA catabolic genes↑
      Significant effect of intervention vs baseline.
      Camastra S et al. [
      • Camastra S.
      • Gastaldelli A.
      • Mari A.
      • Bonuccelli S.
      • Scartabelli G.
      • Frascerra S.
      • et al.
      Early and longer term effects of gastric bypass surgery on tissue-specific insulin sensitivity and beta cell function in morbidly obese patients with and without type 2 diabetes.
      ]
      B/A;

      RYGB;

      2-mo, 1-y;

      HEC
      RYGB:

      T2DM, 13(4); 50 ± 2

      Obe, 12(1); 54 ± 2



      CTRL:

      Lean, 8(0); 23 ± 0

      Obe, 14(2); 36 ± 1;
      2-mo:

      RaGly↑
      Significant effect between groups after intervention.


      EGP↓
      Significant effect between groups after intervention.


      M value↑
      Significant effect between groups after intervention.


      1-y:

      M value↑
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      RaGly↓
      Significant effect of intervention vs baseline.


      EGP↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      β-cell function (T2DM)↓
      Significant effect of intervention vs baseline.
      2-mo:

      BW↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      BMI↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      FPG↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      FPI↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      NEFAP
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      Glucose ox↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      Lipid ox↑
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      1-y:

      BW↓
      Significant effect of intervention vs baseline.


      BMI↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      FPG↓
      Significant effect of intervention vs baseline.


      FPI↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      HbA1c↓
      Significant effect of intervention vs baseline.


      AIRg↑
      Significant effect of intervention vs baseline.


      Glucose ox↑
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.


      Lipid ox↓
      Significant effect of intervention vs baseline.
      Significant effect between groups after intervention.
      Gancheva S et al. [
      • Gancheva S.
      • Ouni M.
      • Jelenik T.
      • Koliaki C.
      • Szendroedi J.
      • Toledo F.G.S.
      • et al.
      Dynamic changes of muscle insulin sensitivity after metabolic surgery.
      ]
      B/A;

      BS;

      2-wk, 12-wk, 24-wk, 52-wk;

      HEC
      Obe, 49(14); 51 ± 72-wk:

      AT-IR↑
      Significant effect of intervention vs baseline.


      12/24/52-wk:

      AT-IR↓
      Significant effect of intervention vs baseline.


      M value↑
      Significant effect of intervention vs baseline.
      2-wk:

      PKCθ↑
      Significant effect of intervention vs baseline.


      DAG↑
      Significant effect of intervention vs baseline.


      FFA↑
      Significant effect of intervention vs baseline.


      BMI↓
      Significant effect of intervention vs baseline.


      BW↓
      Significant effect of intervention vs baseline.


      HbA1c↓
      Significant effect of intervention vs baseline.


      12/24/52-wk:

      FPG↓
      Significant effect of intervention vs baseline.


      FPI↓
      Significant effect of intervention vs baseline.


      FFA↓
      Significant effect of intervention vs baseline.
      52-wk:

      DNA methylation
      Data are presented as mean ± SD.
      Abbreviations: AIRg = acute insulin response; AKT = protein kinase B; AT = adipose tissue; AT-IR = adipose tissue insulin resistance; β-HAD = β-hydroxyacid dehydrogenase; B/A = controlled trial before/after; BCAA = branched chain amino acid; BS = bariatric surgery; BW = body weight; COX = cytochrome c oxidase; CR = caloric restriction; CSA = citrate synthase activity; CTRL = control; DAG = diacylglycerol; EPC = euglycemic pancreatic camp; ER = endoplasmic reticulum; ERK = extracellular signal-regulated kinases; F = fecal; FFA = free fatty acids; FGF21 = fibroblast growth factor 21; FPG = fasting plasma glucose; FPGh = fasting plasma ghrelin; FPI = fasting plasma insulin; GIR = glucose infusion rate; GPX1 = glutathione peroxidase 1; HEC = hyperinsulinemic euglycemic clamp; HIIT = high intensity interval training; H-ND = non-diabetic control; H-off = healthy offspring of mother with T2D; HSL = hormone sensitive lipase; iEGP = insulin-simulated suppression of endogenous glucose production; iFFA = insulin-stimulated free fatty acids suppression; IR = insulin resistant; IRS = insulin receptor substrate; LCD = low-calorie diet; M/I = HEC-derived M value adjusted for insulin concentrations; LDL = low density lipoprotein; MS = metabolic syndrome; mt = mitochondrial; mTOR = mechanistic target of rapamycin; MTT = meal tolerance test; NEFA = non-esterified fatty acids; NR = non-responder; Obe = obese; OGIS = oral glucose sensitivity index; ω3-PUFA = omega3 polyunsaturated fatty acids; P = plasma; PKC = protein kinase C; PPI = postprandial insulin secretion; PREDIM = predicted M index; R = responders; RaGly = rate of appearance of glycerol; R/Co = randomized crossover; RCR = mitochondrial respiratory control ratio; Rd = glucose rate of disappearance; ROS = reactive oxygen species; RYGB = Roux-en-Y gastric bypass surgery; SCFA = short chain fatty acid; SISTA = Stepped Insulin Secretion Test with Arginine stimulation; SkM = skeletal muscle; T2DM = type 2 diabetes mellitus; TC = total cholesterol; TG = triglycerides; TXNIP = thioredoxin-interacting protein; VAT = visceral AT; VLCD = very-low-calorie diet; VLDL = very low density lipoprotein.
      a Significant effect of intervention vs baseline.
      b Significant effect between groups after intervention.
      In addition, physical activity increases insulin sensitivity by (i) enhancing muscle mitochondrial biogenesis, GLUT4 protein content and glucose uptake, (ii) repartitioning IMCL and (iii) reducing fat mass [
      • Balkau B.
      • Mhamdi L.
      • Oppert J.M.
      • Nolan J.
      • Golay A.
      • Porcellati F.
      • et al.
      Physical activity and insulin sensitivity: the RISC study.
      ]. A short-term exercise training increases glucose disposal rate and EGP suppression during HEC in the absence of BW loss [
      • Kirwan J.P.
      • Solomon T.P.
      • Wojta D.M.
      • Staten M.A.
      • Holloszy J.O.
      Effects of 7 days of exercise training on insulin sensitivity and responsiveness in type 2 diabetes mellitus.
      ], but exercise with BW loss promotes a greater reduction in VAT and improves EGP suppression as well as glucose disposal [
      • Coker R.H.
      • Williams R.H.
      • Yeo S.E.
      • Kortebein P.M.
      • Bodenner D.L.
      • Kern P.A.
      • et al.
      The impact of exercise training compared to caloric restriction on hepatic and peripheral insulin resistance in obesity.
      ]. Of note, 12-wk exercise intervention enhances BCAA catabolism in SkM and subcutaneous adipose tissue (SAT), which might explain the improvement of insulin sensitivity [
      • Lee S.
      • Gulseth H.L.
      • Langleite T.M.
      • Norheim F.
      • Olsen T.
      • Refsum H.
      • et al.
      Branched-chain amino acid metabolism, insulin sensitivity and liver fat response to exercise training in sedentary dysglycaemic and normoglycaemic men.
      ]. However, up to 20% of T2DM individuals fail to respond to physical training with improved glucose metabolism [
      • Stephens N.A.
      • Sparks L.M.
      Resistance to the beneficial effects of exercise in type 2 diabetes: are some individuals programmed to fail?.
      ]. Single nucleotide polymorphisms in mitochondrial components correlate with the effectiveness of the training to ameliorate aerobics fitness and insulin sensitivity in glucose tolerant relatives of T2DM individuals [
      • Kacerovsky-Bielesz G.
      • Kacerovsky M.
      • Chmelik M.
      • Farukuoye M.
      • Ling C.
      • Pokan R.
      • et al.
      A single nucleotide polymorphism associates with the response of muscle ATP synthesis to long-term exercise training in relatives of type 2 diabetic humans.
      ]. Furthermore, variable training responses can reflect a different SkM epigenomic profile and gene expression at baseline [
      • Stephens N.A.
      • Brouwers B.
      • Eroshkin A.M.
      • Yi F.
      • Cornnell H.H.
      • Meyer C.
      • et al.
      Exercise response variations in skeletal muscle PCr recovery rate and insulin sensitivity relate to muscle epigenomic profiles in individuals with type 2 diabetes.
      ]. Exercise intensity and volume can differently affect IR. A meta-analysis including 2033 participants suggests that high intensity interval training (HIIT) reduces BW by 1.3 kg compared with the non-exercising group and it improves IR, assessed with less accurate methods than HEC, more effectively than continuous exercise, with the largest effects among insulin-resistant individuals [
      • Jelleyman C.
      • Yates T.
      • O’Donovan G.
      • Gray L.J.
      • King J.A.
      • Khunti K.
      • et al.
      The effects of high-intensity interval training on glucose regulation and insulin resistance: a meta-analysis.
      ]. However, one study implying a two-step HEC confirmed that HIIT enhances mitochondrial function and peripheral insulin sensitivity in young and elderly people [
      • Robinson M.M.
      • Dasari S.
      • Konopka A.R.
      • Johnson M.L.
      • Manjunatha S.
      • Esponda R.R.
      • et al.
      Enhanced protein translation underlies improved metabolic and physical adaptations to different exercise training modes in young and old humans.
      ], instead another study showed that HIIT does not uniformly improve insulin sensitivity among healthy and T2DM individuals, but it allows to identify responders to training, who are mainly insulin-resistant at baseline [
      • Apostolopoulou M.
      • Mastrototaro L.
      • Hartwig S.
      • Pesta D.
      • Straßburger K.
      • de Filippo E.
      • et al.
      Metabolic responsiveness to training depends on insulin sensitivity and protein content of exosomes in insulin resistant males.
      ].

      4.2 Bariatric surgery

      Bariatric surgery is the most effective anti-obesity intervention, providing an impressive and sustained BW loss, along with improvements in whole-body insulin sensitivity and even remission of T2DM [
      • Camastra S.
      • Gastaldelli A.
      • Mari A.
      • Bonuccelli S.
      • Scartabelli G.
      • Frascerra S.
      • et al.
      Early and longer term effects of gastric bypass surgery on tissue-specific insulin sensitivity and beta cell function in morbidly obese patients with and without type 2 diabetes.
      ]. Of note, the initially BW reduction in the range of 10 kg is not necessarily accompanied by improved IR, likely due to transient excessive WAT lipolysis, IMCL accumulation and SkM mitochondrial abnormalities, whereas the subsequent persistent BW loss markedly improves glucose metabolism, partly due to epigenetic changes in SkM [
      • Gancheva S.
      • Ouni M.
      • Jelenik T.
      • Koliaki C.
      • Szendroedi J.
      • Toledo F.G.S.
      • et al.
      Dynamic changes of muscle insulin sensitivity after metabolic surgery.
      ]. Since peripheral insulin sensitivity after surgery remains lower than in healthy lean individuals, Coen et al. conducted a randomized exercise trial in severe obese individuals after bariatric surgery and found that exercise further enhances insulin sensitivity and mitochondrial respiration and reduces IMCL [
      • Coen P.M.
      • Menshikova E.V.
      • Distefano G.
      • Zheng D.
      • Tanner C.J.
      • Standley R.A.
      • et al.
      Exercise and weight loss improve muscle mitochondrial respiration, lipid partitioning, and insulin sensitivity after gastric bypass surgery.
      ].

      4.3 Antihyperglycemic drugs

      Different drug concepts used to lower blood glucose in T2DM may also exert beneficial effects on IR as described in the next section and listed in Table 3.
      Table 3Randomized double-blinded controlled and paralleled group trials investigating the effects of pharmacological intervention for >3 months on insulin resistance in overweight/obese individuals with or without T2DM.
      StudyDesign; cohort, durationIntervention, N total (males); BMI (kg/m2)Primary outcomeMetabolic parameter
      Miyazaki Y et al. [
      • Miyazaki Y.
      • Mahankali A.
      • Matsuda M.
      • Mahankali S.
      • Hardies J.
      • Cusi K.
      • et al.
      Effect of pioglitazone on abdominal fat distribution and insulin sensitivity in type 2 diabetic patients.
      ]
      R/PC;

      T2DM;

      16-wk
      PIO 45 mg/d, 12(11); 29 ± 1

      Plb, 11(6); 29 ± 1
      iEGP↑
      Significant effect of intervention.


      Rd↑
      Significant effect of intervention.
      BW↑
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FM↑
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FPG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      HbA1c↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      TGP
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FFAP
      Significant effect of intervention.


      Sc fat↑
      Significant effect of intervention.


      Vs fat↓
      Significant effect of intervention.
      Krishnappa M et al. [
      • Krishnappa M.
      • Patil K.
      • Parmar K.
      • Trivedi P.
      • Mody N.
      • Shah C.
      • et al.
      Effect of saroglitazar 2 mg and 4 mg on glycemic control, lipid profile and cardiovascular disease risk in patients with type 2 diabetes mellitus: a 56-week, randomized, double blind, phase 3 study (PRESS XII study).
      ]
      R/P;

      T2DM;

      24-wk → 32-wk
      Saro 2 mg/d, 380(216); 26 ± 4

      Saro 4 mg/d, 386(243); 26 ± 4

      PIO 30 mg/d, 389(222); 26 ± 4
      24-wk:

      HbA1C↓
      Significant effect of intervention.


      FPG↓ (PIO)
      Significant effect of intervention.


      56-wk:

      HbA1c↓
      Significant effect of intervention.


      FPG↓
      Significant effect of intervention.
      24-wk:

      Saro 4 mg, PIO: TG↓
      Significant effect of intervention.


      VLDL-C↓
      Significant effect of intervention.


      Saro 2/4 mg: LDL-C↓
      Significant effect of intervention.


      Saro 4 mg: TC↓
      Significant effect of intervention.


      56-wk:

      LDL-C↓
      Significant effect of intervention.


      Saro 4 mg:TG↓
      Significant effect of intervention.


      HDL-C↓
      Significant effect of intervention.
      Jain N et al. [
      • Jain N.
      • Bhansali S.
      • Kurpad A.V.
      • Hawkins M.
      • Sharma A.
      • Kaur S.
      • et al.
      Effect of a dual PPAR alpha/gamma agonist on insulin sensitivity in patients of type 2 diabetes with hypertriglyceridemia - randomized double-blind placebo-controlled trial.
      ]
      R/PC;

      T2DM with TG > 150 mg/d;

      4-mo
      Saro 4 mg/d, 15(15); 27 ± 2

      Plb, 15(12); 28 ± 2
      M/I↑
      Significant effect of intervention.
      Significant effect vs control after intervention.


      M↑
      Significant effect of intervention.


      HOMA-β↑
      Significant effect of intervention.
      Significant effect vs control after intervention.
      TG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      HbA1c↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FPG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      HDL-C↑
      Significant effect of intervention.
      Significant effect vs control after intervention.
      Miyagawa J et al. [
      • Miyagawa J.
      • Odawara M.
      • Takamura T.
      • Iwamoto N.
      • Takita Y.
      • Imaoka T.
      Once-weekly glucagon-like peptide-1 receptor agonist dulaglutide is non-inferior to once-daily liraglutide and superior to placebo in Japanese patients with type 2 diabetes: a 26-week randomized phase III study.
      ]
      R/PC;

      T2DM;

      52-wk
      Dul 0.75 mg/d, 280(228); 36 ± 4

      Lgt 0.9 mg/d, 137(113); 26 ± 4

      Plb,70(55); 25 ± 3
      HbA1c↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      (Dul vs Plb)
      HbA1c↓
      Significant effect of intervention.
      (Dul, Lgt)

      FPG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      (vs Plb)

      BW↔

      HOMA2-%β↑
      Significant effect of intervention.
      (Dul, Lgt)

      HOMA2-%S↓
      Significant effect of intervention.
      (Dul)
      Dungan KM et al. [
      • Dungan K.M.
      • Povedano S.T.
      • Forst T.
      • Gonzalez J.G.
      • Atisso C.
      • Sealls W.
      • et al.
      Once-weekly dulaglutide versus once-daily liraglutide in metformin-treated patients with type 2 diabetes (AWARD-6): a randomised, open-label, phase 3, non-inferiority trial.
      ]
      R/P;

      T2DM;

      26-wk
      Dul 1.5 mg/d, 299(138); 34 ± 5

      Lgt 1.8 mg/d, 300(149); 34 ± 5
      HbA1c↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      HbA1c <7%↔

      FPG↓
      Significant effect of intervention.


      PPG↓
      Significant effect of intervention.


      BW↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      Β-cells function↑
      Significant effect of intervention.
      Ahren B et al. [
      • Ahren B.
      • Leguizamo Dimas A.
      • Miossec P.
      • Saubadu S.
      • Aronson R.
      Efficacy and safety of lixisenatide once-daily morning or evening injections in type 2 diabetes inadequately controlled on metformin (GetGoal-M).
      ]
      R/PC;

      T2DM;

      24-wk
      Lix 20 μg/d morning, 255(97); 33 ± 7

      Lix 20 μg/d evening, 255(114); 33 ± 6

      Plb, 170(81); 33 ± 6
      HbA1c↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      FPG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      BW↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      HOMA-β↑
      Significant effect of intervention.
      Significant effect vs control after intervention.


      2 h-PPG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      Dutour A et al. [
      • Dutour A.
      • Abdesselam I.
      • Ancel P.
      • Kober F.
      • Mrad G.
      • Darmon P.
      • et al.
      Exenatide decreases liver fat content and epicardial adipose tissue in patients with obesity and type 2 diabetes: a prospective randomized clinical trial using magnetic resonance imaging and spectroscopy.
      ]
      R/PC;

      T2DM;

      26-wk
      Exe 2x10μg/d, 22(13); 37 ± 2

      Plb, 22(8); 35 ± 1
      HbA1c↓
      Significant effect of intervention.


      EAT↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      HTGC↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      PTGC↔

      MTGC↔
      BW↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      Leptin↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      Rodbard HW et al. [
      • Rodbard H.W.
      • Rosenstock J.
      • Canani L.H.
      • Deerochanawong C.
      • Gumprecht J.
      • Lindberg S.O.
      • et al.
      Oral semaglutide versus empagliflozin in patients with type 2 diabetes uncontrolled on metformin: the PIONEER 2 trial.
      ]
      R/AC;

      T2DM;

      52-wk
      Sem 14 mg/d, 411(206); 33 ± 6

      Empa 25 mg/d, 410(209); 33 ± 6
      HbA1c↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      BW↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FPG↓
      Significant effect of intervention.


      FPI↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FCP↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      CRP↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      Schiavon M et al. [
      • Schiavon M.
      • Visentin R.
      • Gobel B.
      • Riz M.
      • Cobelli C.
      • Klabunde T.
      • et al.
      Improved postprandial glucose metabolism in type 2 diabetes by the dual glucagon-like peptide-1/glucagon receptor agonist SAR425899 in comparison with liraglutide.
      ]
      R/P;

      T2DM;

      26-wk
      SAR425899 0.12 mg/d, 21; 32[30, 36]
      Median [25th, 75th] percentile.


      SAR425899 0.16 mg/d, 15; 34[32, 38]
      Median [25th, 75th] percentile.


      SAR425899 0.20 mg/d, 10; 32[30, 35]
      Median [25th, 75th] percentile.


      Lgt 1.80 mg/d, 17; 35[29, 40]
      Median [25th, 75th] percentile.


      Plb, 7; 34[29, 35]
      Median [25th, 75th] percentile.
      HOMA2-%S↑
      Significant effect of intervention.
      (Lira)

      HOMA2-%β↑
      Significant effect of intervention.
      Significant effect vs control after intervention.
      (SAR vs Plb)

      Si
      Significant effect of intervention.


      HbA1c↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      (SAR and Lira vs Plb)
      Cusi et al. [
      • Cusi K.
      • Bril F.
      • Barb D.
      • Polidori D.
      • Sha S.
      • Ghosh A.
      • et al.
      Effect of canagliflozin treatment on hepatic triglyceride content and glucose metabolism in patients with type 2 diabetes.
      ]
      R/PC;

      T2DM;

      24-wk
      Cana 300 mg/d, 26(16); 32 ± 4

      Plb, 30(21); 31 ± 5
      HTG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      iEGP↑
      Significant effect of intervention.
      Significant effect vs control after intervention.


      β-cell function↑
      Significant effect of intervention.
      Significant effect vs control after intervention.
      HbA1c↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      BW↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FPG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FPI↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      Insulin clearance↑
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FFA↑
      Significant effect of intervention.
      Significant effect vs control after intervention.
      Kahl S et al. [
      • Kahl S.
      • Gancheva S.
      • Strassburger K.
      • Herder C.
      • Machann J.
      • Katsuyama H.
      • et al.
      Empagliflozin effectively lowers liver fat content in well-controlled type 2 diabetes: a randomized, double-blind, phase 4.
      ]
      R/PC;

      T2DM;

      24-wk
      Empa 25 mg/d, 42(29); 32 ± 4

      Plb, 42(29); 32 ± 4
      IHLC↓
      Significant effect of intervention.
      Significant effect vs control after intervention.
      M↔

      iEGP↔

      iFFA↔

      BW↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      FBG↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      UA↓
      Significant effect of intervention.
      Significant effect vs control after intervention.


      Adiponectin↑
      Significant effect of intervention.
      Significant effect vs control after intervention.
      Data are presented as mean ± SD.
      Abbreviations: ALT = alanine aminotransferase; AST = aspartate aminotransferase; BW = body weight; Cana = canagliflozin; Dul = dulaglutide; EAT = epicardial adipose tissue; Empa = empagliflozin; Exe = exenatide; FCP = fasting C peptide; FFA = free fatty acids; FM = fat mass; FPG = fasting plasma glucose; FPGluc = fasting plasma glucagone; FPI = fasting plasma insulin; Gla = glargine; HDL-C = high density lipoproteins-cholesterol; HLC = hepatic lipid content; HTGC = hepatic triglyceride content; iEGP = insulin-stimulated suppression of endogenous glucose production; LDL-C = low density lipoprotein-cholesterol; IHLC = intrahepatic lipid content; LGT = liraglutide; Lix = lixisenatide; M = glucose metabolism; M/I = insulin sensitivity; MRI = magnetic resonance imaging; MTGC = myocardial triglyceride content; PDFF = proton density fat fraction; Peg = Pegbelfermin; PIO = pioglitazone; plb = placebo; PPG = postprandial glucose; PTGC = pancreatic triglyceride content; R/AC = randomized with active control; R/P = randomized paralleled; R/PC = randomized placebo-controlled; Rd = rate of glucose disposal; Saro = saroglitazar; Sc = subcutaneous; Sem = semaglutide; Si = oral minimal model indices of insulin sensitivity; T2DM = type 2 diabetes mellitus; TC = total cholesterol; TGP = plasma triglycerides; UA = uric acid; VLDL = very-low density lipoprotein; Vs = visceral; WB-IS = whole body insulin sensitivity.
      a Significant effect of intervention.
      b Significant effect vs control after intervention.
      c Median [25th, 75th] percentile.

      4.3.1 Sulfonylurea drugs

      Sulfonylurea drugs decrease hyperglycemia by increasing meal-independent insulin secretion, but therefore also increase the risk of hypoglycemia and BW gain. While lowering hyperglycemia would decrease glucotoxicity, their effects on IR are controversial. For example, glimepiride increases adiponectin levels and decreases IR when measured by HOMA-IR [
      • Koshiba K.
      • Nomura M.
      • Nakaya Y.
      • Ito S.
      Efficacy of glimepiride on insulin resistance, adipocytokines, and atherosclerosis.
      ], but not when assessed by HEC in another study [
      • Korytkowski M.
      • Thomas A.
      • Reid L.
      • Tedesco M.B.
      • Gooding W.E.
      • Gerich J.
      Glimepiride improves both first and second phases of insulin secretion in type 2 diabetes.
      ]. Because of their side effects, their use is progressively decreasing in countries where newer drug classes are available.

      4.3.2 Metformin

      The biguanide metformin has been the guideline-recommended first-line treatment of T2DM for decades. Its clinical glucose lowering effect mainly results from decreasing EGP. Although the underlying mechanism in humans is still unclear, preclinical studies indicate that metformin likely inhibits the activity of hepatic glycerol-3-phosphate dehydrogenase, which impairs glycerol-induced gluconeogenesis and increases the cytosolic redox state, resulting in decreased lactate dehydrogenase and lactate-induced EGP [
      • LaMoia T.E.
      • Shulman G.I.
      Cellular and molecular mechanisms of metformin action.
      ]. This mechanism also helps to explain lactate acidosis, a frequent side effect of biguanides other than metformin. Alternative mechanisms comprise the inhibition of complex I followed by increased AMP, which activates AMPK, in turn promoting hepatic FAO and downregulating gluconeogenic genes. AMP can also antagonize glucagon signaling by preventing the accumulation of cyclic AMP and can directly inhibit gluconeogenesis through fructose 1,6-biphopshatase, although these effects are not observed at clinically relevant concentration of metformin [
      • LaMoia T.E.
      • Shulman G.I.
      Cellular and molecular mechanisms of metformin action.
      ]. Metformin has also extrahepatic effects. It increases whole-body insulin-stimulated glucose uptake [
      • LaMoia T.E.
      • Shulman G.I.
      Cellular and molecular mechanisms of metformin action.
      ] and AMPK activity in SkM of T2DM individuals, which is associated with higher glucose disposal and muscle glycogen concentration [
      • Musi N.
      • Hirshman M.F.
      • Nygren J.
      • Svanfeldt M.
      • Bavenholm P.
      • Rooyackers O.
      • et al.
      Metformin increases AMP-activated protein kinase activity in skeletal muscle of subjects with type 2 diabetes.
      ]. The intestinal mechanisms of metformin include alterations in gut microbiome, increased secretion of GLP-1 and growth differentiation factor 15, which reduces BW and appetite [
      • LaMoia T.E.
      • Shulman G.I.
      Cellular and molecular mechanisms of metformin action.
      ]. Despite this plethora of effects, the effect of metformin on IR is still under debate. A systematic review and meta-analysis of 11 randomized controlled trials (RCT) in obese and overweight adolescents, reveals that metformin reduces the fasting plasma glucose (FPG) in interventions <6 months, but has no effects on IR [
      • Sun J.
      • Wang Y.
      • Zhang X.
      • He H.
      The effects of metformin on insulin resistance in overweight or obese children and adolescents: a PRISMA-compliant systematic review and meta-analysis of randomized controlled trials.
      ]. Conversely a larger meta-analysis of 31 RCT of ≥8 weeks in persons at risk for diabetes shows that metformin improves BMI, lipid profiles and IR, and reduces the incidence of new-onset diabetes [
      • Salpeter S.R.
      • Buckley N.S.
      • Kahn J.A.
      • Salpeter E.E.
      Meta-analysis: metformin treatment in persons at risk for diabetes mellitus.
      ]. Finally a meta-analysis including 417 individuals with NAFLD confirms the benefit of metformin on BMI and HOMA-IR to some extent, without any effect on liver histology [
      • Li Y.
      • Liu L.
      • Wang B.
      • Wang J.
      • Chen D.
      Metformin in non-alcoholic fatty liver disease: a systematic review and meta-analysis.
      ].

      4.3.3 Peroxisome proliferator-activated receptor (PPAR) agonists

      PPAR agonists modulate the transcription of nuclear transcription factors such as PPARα/γ/δ, which primarily stimulate WAT remodeling and modulate lipid fluxes to improve insulin signaling and glucose homeostasis. Thiazolidinediones (TZD), PPARγ ligands, promote FFA storage in SAT, reducing lipid ectopic accumulation. TZD inhibit also the production of pro-inflammatory cytokines and trigger the release of adiponectin [
      • Phielix E.
      • Szendroedi J.
      • Roden M.
      The role of metformin and thiazolidinediones in the regulation of hepatic glucose metabolism and its clinical impact.
      ]. A systematic review reports that TZD reduce FPG and hepatic IR similar to metformin, but only the TZD enhance insulin-mediated glucose uptake [
      • Natali A.
      • Ferrannini E.
      Effects of metformin and thiazolidinediones on suppression of hepatic glucose production and stimulation of glucose uptake in type 2 diabetes: a systematic review.
      ]. The TZD, pioglitazone, a potent insulin sensitizer approved for treating people with T2DM and with beneficial effects on NAFLD [
      • Dewidar B.
      • Kahl S.
      • Pafili K.
      • Roden M.
      Metabolic liver disease in diabetes - from mechanisms to clinical trials.
      ], induces a shift of fat from VAT to SAT, which is associated with improved hepatic and peripheral IR [
      • Miyazaki Y.
      • Mahankali A.
      • Matsuda M.
      • Mahankali S.
      • Hardies J.
      • Cusi K.
      • et al.
      Effect of pioglitazone on abdominal fat distribution and insulin sensitivity in type 2 diabetic patients.
      ]; however, its side effects such as BW gain, edema, cardiovascular events, atypical bone fractures need to be considered [
      • Pafili K.
      • Roden M.
      Nonalcoholic fatty liver disease (NAFLD) from pathogenesis to treatment concepts in humans.
      ]. Conversely, lobeglitazone, a recently developed TZD licensed for T2DM, shows a satisfactory safety profile and ameliorates IR and hepatic steatosis in T2DM people with NAFLD [
      • Lee Y.H.
      • Kim J.H.
      • Kim S.R.
      • Jin H.Y.
      • Rhee E.J.
      • Cho Y.M.
      • et al.
      Lobeglitazone, a novel thiazolidinedione, improves non-alcoholic fatty liver disease in type 2 diabetes: its efficacy and predictive factors related to responsiveness.
      ]. MSDC-0602 K is a PPARγ-sparing TZD with insulin sensitizing effects without safety limitation and excellent tolerability [
      • Gastaldelli A.
      • Stefan N.
      • Haring H.U.
      Liver-targeting drugs and their effect on blood glucose and hepatic lipids.
      ]. The PPARα agonists fibrates, which primarily lower plasma TG and cholesterol content by stimulating hepatic FAO [
      • Staels B.
      • Fruchart J.C.
      Therapeutic roles of peroxisome proliferator-activated receptor agonists.
      ], decrease FPG, insulin levels and HOMA-IR, as shown in a recent meta-analysis [
      • Simental-Mendia L.E.
      • Simental-Mendia M.
      • Sanchez-Garcia A.
      • Banach M.
      • Atkin S.L.
      • Gotto Jr., A.M.
      • et al.
      Effect of fibrates on glycemic parameters: a systematic review and meta-analysis of randomized placebo-controlled trials.
      ]. However, fibrates fail to ameliorate IR when assessed with HEC [
      • Riserus U.
      • Sprecher D.
      • Johnson T.
      • Olson E.
      • Hirschberg S.
      • Liu A.
      • et al.
      Activation of peroxisome proliferator-activated receptor (PPAR)delta promotes reversal of multiple metabolic abnormalities, reduces oxidative stress, and increases fatty acid oxidation in moderately obese men.
      ]. Activation of PPARδ increases FAO in SkM and suppresses inflammation in macrophages [
      • Staels B.
      • Fruchart J.C.
      Therapeutic roles of peroxisome proliferator-activated receptor agonists.
      ]. The synthetic PPARδ agonists, seladelpar and GW501516, reduce HOMA-IR and TG in overweight individuals [
      • Riserus U.
      • Sprecher D.
      • Johnson T.
      • Olson E.
      • Hirschberg S.
      • Liu A.
      • et al.
      Activation of peroxisome proliferator-activated receptor (PPAR)delta promotes reversal of multiple metabolic abnormalities, reduces oxidative stress, and increases fatty acid oxidation in moderately obese men.
      ,
      • Bays H.E.
      • Schwartz S.
      • Littlejohn 3rd, T.
      • Kerzner B.
      • Krauss R.M.
      • Karpf D.B.
      • et al.
      MBX-8025, a novel peroxisome proliferator receptor-delta agonist: lipid and other metabolic effects in dyslipidemic overweight patients treated with and without atorvastatin.
      ]. Agents that activate multiple PPAR isoforms have been developed. Elafibranor, a PPARα/δ agonist, reduces inflammation and enhances both peripheral and hepatic IR assessed by HEC in obese humans [
      • Cariou B.
      • Hanf R.
      • Lambert-Porcheron S.
      • Zair Y.
      • Sauvinet V.
      • Noel B.
      • et al.
      Dual peroxisome proliferator-activated receptor alpha/delta agonist GFT505 improves hepatic and peripheral insulin sensitivity in abdominally obese subjects.
      ], but the effects on HOMA-IR are contradictory [
      • Westerouen Van Meeteren M.J.
      • Drenth J.P.H.
      • Tjwa E.
      Elafibranor: a potential drug for the treatment of nonalcoholic steatohepatitis (NASH).
      ]. The dual PPARα/γ agonist saroglitazar improves whole-body insulin sensitivity along with β-cells function, hyperglycemia and lipid profile in metformin-treated T2DM, without the safety concerns reported for other PPARα/γ agonists [
      • Krishnappa M.
      • Patil K.
      • Parmar K.
      • Trivedi P.
      • Mody N.
      • Shah C.
      • et al.
      Effect of saroglitazar 2 mg and 4 mg on glycemic control, lipid profile and cardiovascular disease risk in patients with type 2 diabetes mellitus: a 56-week, randomized, double blind, phase 3 study (PRESS XII study).
      ,
      • Jain N.
      • Bhansali S.
      • Kurpad A.V.
      • Hawkins M.
      • Sharma A.
      • Kaur S.
      • et al.
      Effect of a dual PPAR alpha/gamma agonist on insulin sensitivity in patients of type 2 diabetes with hypertriglyceridemia - randomized double-blind placebo-controlled trial.
      ]. Currently an ongoing phase-2 trial evaluates whether lanifibranor, a pan-PPAR agonist, decreases IHL and IR in people with T2DM and NAFLD (NCT03459079).

      4.3.4 Incretin mimetics

      GLP-1 receptor agonists (GLP-1Ra) increase meal-dependent insulin secretion, but also mediate satiety in the hypothalamus resulting in reduced BW and improved insulin sensitivity [
      • Baggio L.L.
      • Drucker D.J.
      Glucagon-like peptide-1 receptor co-agonists for treating metabolic disease.
      ]. A prospective study shows that 4-wk liraglutide improves BMI and insulin sensitivity measured by HEC [
      • Jinnouchi H.
      • Sugiyama S.
      • Yoshida A.
      • Hieshima K.
      • Kurinami N.
      • Suzuki T.
      • et al.
      Liraglutide, a glucagon-like peptide-1 analog, increased insulin sensitivity assessed by hyperinsulinemic-euglycemic clamp examination in patients with uncontrolled type 2 diabetes mellitus.
      ]. Conversely, administration of dulaglutide (0.75 mg) and liraglutide (0.9 mg) monotherapies in Japanese individuals with T2DM reduces HbA1c and increases HOMA-β without affecting BW [
      • Miyagawa J.
      • Odawara M.
      • Takamura T.
      • Iwamoto N.
      • Takita Y.
      • Imaoka T.
      Once-weekly glucagon-like peptide-1 receptor agonist dulaglutide is non-inferior to once-daily liraglutide and superior to placebo in Japanese patients with type 2 diabetes: a 26-week randomized phase III study.
      ]. This could be explained with the low BW of Japanese population and with the lower doses employed in this trial. Indeed, another phase-3 RCT in T2DM adults demonstrated BW reduction with dulaglutide and even more with liraglutide [
      • Dungan K.M.
      • Povedano S.T.
      • Forst T.
      • Gonzalez J.G.
      • Atisso C.
      • Sealls W.
      • et al.
      Once-weekly dulaglutide versus once-daily liraglutide in metformin-treated patients with type 2 diabetes (AWARD-6): a randomised, open-label, phase 3, non-inferiority trial.
      ]. The greater effectiveness of liraglutide for achieving BW loss might reflect enhanced uptake into the brain, which probably cannot be achieved with high-molecular-weight GLP-1Ra as dulaglutide. Phase-3 RCTs demonstrated that lixisenatide, exenatide and semaglutide reduce BW in T2DM persons and ameliorate hyperglycemia, which is accompanied by improved β-cell function and IR based on HOMA [
      • Ahren B.
      • Leguizamo Dimas A.
      • Miossec P.
      • Saubadu S.
      • Aronson R.
      Efficacy and safety of lixisenatide once-daily morning or evening injections in type 2 diabetes inadequately controlled on metformin (GetGoal-M).
      ,
      • Dutour A.
      • Abdesselam I.
      • Ancel P.
      • Kober F.
      • Mrad G.
      • Darmon P.
      • et al.
      Exenatide decreases liver fat content and epicardial adipose tissue in patients with obesity and type 2 diabetes: a prospective randomized clinical trial using magnetic resonance imaging and spectroscopy.
      ,
      • Rodbard H.W.
      • Rosenstock J.
      • Canani L.H.
      • Deerochanawong C.
      • Gumprecht J.
      • Lindberg S.O.
      • et al.
      Oral semaglutide versus empagliflozin in patients with type 2 diabetes uncontrolled on metformin: the PIONEER 2 trial.
      ]. Recent systematic reviews and meta-analyses confirmed that GLP-1Ra reduce all-cause (oral semaglutide, liraglutide, extended-release exenatide) and cardiovascular mortality (oral semaglutide, liraglutide), non-fatal myocardial infarction and kidney failure [
      • Tsapas A.
      • Avgerinos I.
      • Karagiannis T.
      • Malandris K.
      • Manolopoulos A.
      • Andreadis P.
      • et al.
      Comparative effectiveness of glucose-lowering drugs for type 2 diabetes: a systematic review and network meta-analysis.
      ,
      • Palmer S.C.
      • Tendal B.
      • Mustafa R.A.
      • Vandvik P.O.
      • Li S.
      • Hao Q.
      • et al.
      Sodium-glucose cotransporter protein-2 (SGLT-2) inhibitors and glucagon-like peptide-1 (GLP-1) receptor agonists for type 2 diabetes: systematic review and network meta-analysis of randomised controlled trials.
      ]. Also, odds ratios of stroke are lower with subcutaneous semaglutide and dulaglutide [
      • Tsapas A.
      • Avgerinos I.
      • Karagiannis T.
      • Malandris K.
      • Manolopoulos A.
      • Andreadis P.
      • et al.
      Comparative effectiveness of glucose-lowering drugs for type 2 diabetes: a systematic review and network meta-analysis.
      ], whereas GLP-1Ra have no effect on heart failure [
      • Palmer S.C.
      • Tendal B.
      • Mustafa R.A.
      • Vandvik P.O.
      • Li S.
      • Hao Q.
      • et al.
      Sodium-glucose cotransporter protein-2 (SGLT-2) inhibitors and glucagon-like peptide-1 (GLP-1) receptor agonists for type 2 diabetes: systematic review and network meta-analysis of randomised controlled trials.
      ]. Because of their clinical efficacy, GLP-1Ra are emerging as a preferred treatment option for T2DM with or at risk of cardiorenal complications.
      Also, glucagon inhibits food intake and increases energy expenditure, leading to BW loss. Several unimolecular glucagon/GLP-1Ra have been recently assessed in short-term trials in obesity and T2DM. They demonstrate meaningful reductions in glycemia and BW [
      • Ambery P.
      • Parker V.E.
      • Stumvoll M.
      • Posch M.G.
      • Heise T.
      • Plum-Moerschel L.
      • et al.
      MEDI0382, a GLP-1 and glucagon receptor dual agonist, in obese or overweight patients with type 2 diabetes: a randomised, controlled, double-blind, ascending dose and phase 2a study.
      ], and a greater improvement of β-cell function compared to liraglutide [
      • Schiavon M.
      • Visentin R.
      • Gobel B.
      • Riz M.
      • Cobelli C.
      • Klabunde T.
      • et al.
      Improved postprandial glucose metabolism in type 2 diabetes by the dual glucagon-like peptide-1/glucagon receptor agonist SAR425899 in comparison with liraglutide.
      ]. The glucose-dependent insulinotropic polypeptide (GIP) has been also utilized in dual agonists with GLP-1. Phase-2 RCT demonstrated that tirzepatide ameliorates HbA1c, BW, waist circumference and HOMA-IR in T2DM participants, suggesting a possible insulin-sensitizing effect secondary to VAT reduction [
      • Baggio L.L.
      • Drucker D.J.
      Glucagon-like peptide-1 receptor co-agonists for treating metabolic disease.
      ]. Preclinical studies have shown that the tri-agonist (glucagon/GIP/GLP-1Ra), HM15211, reduces inflammation, BW and IHL in obese mice more effectively than any dual co-agonist [
      • Baggio L.L.
      • Drucker D.J.
      Glucagon-like peptide-1 receptor co-agonists for treating metabolic disease.
      ] and a phase-1b/2a study confirmed these data in obese individuals with NAFLD [
      • Gastaldelli A.
      • Stefan N.
      • Haring H.U.
      Liver-targeting drugs and their effect on blood glucose and hepatic lipids.
      ].

      4.3.5 α-Glucosidase inhibitors (AGi)

      AGi lower postprandial hyperglycemia by delaying intestinal carbohydrate absorption [
      • Pafili K.
      • Roden M.
      Nonalcoholic fatty liver disease (NAFLD) from pathogenesis to treatment concepts in humans.
      ]. In T2DM, acarbose does not affect IR after 3-m treatment (75 mg daily) [
      • Jenney A.
      • Proietto J.
      • O'Dea K.
      • Nankervis A.
      • Traianedes K.
      • D'Embden H.
      Low-dose acarbose improves glycemic control in NIDDM patients without changes in insulin sensitivity.
      ], but improves IR and insulin secretion after 4-m treatment (100 mg daily) [
      • Delgado H.
      • Lehmann T.
      • Bobbioni-Harsch E.
      • Ybarra J.
      • Golay A.
      Acarbose improves indirectly both insulin resistance and secretion in obese type 2 diabetic patients.
      ]. Also, 3-m treatment with miglitol has favorable effects on BMI, HOMA-IR and adiponectin [
      • Yokoyama H.
      • Kannno S.
      • Ishimura I.
      • Node K.
      Miglitol increases the adiponectin level and decreases urinary albumin excretion in patients with type 2 diabetes mellitus.
      ]. These effects most likely result from the improvement of glucotoxicity.

      4.3.6 Sodium glucose cotransporter (SGLT)-2 inhibitors (SGLT2i)

      SGLT2i not only inhibit reabsorption of glucose in the proximal renal tubule resulting in glucosuria and lower blood glucose, but also improve insulin sensitivity in individuals with T2DM by reducing BW or glucotoxicity. However, glucosuria elicits an adaptive increase in energy intake that causes less BW loss than expected [
      • Ferrannini G.
      • Hach T.
      • Crowe S.
      • Sanghvi A.
      • Hall K.D.
      • Ferrannini E.
      Energy balance after sodium-glucose cotransporter 2 inhibition.
      ]. Canagliflozin induces changes in BW that are positively associated with decreased IHL and hepatic IR [
      • Cusi K.
      • Bril F.
      • Barb D.
      • Polidori D.
      • Sha S.
      • Ghosh A.
      • et al.
      Effect of canagliflozin treatment on hepatic triglyceride content and glucose metabolism in patients with type 2 diabetes.
      ], instead empagliflozin reduces BW and hepatic fat without effect on tissue-specific insulin sensitivity. Furthermore, empagliflozin decreases the levels of circulating uric acid and raises plasma adiponectin concentrations, which likely ameliorate inflammation [
      • Kahl S.
      • Gancheva S.
      • Strassburger K.
      • Herder C.
      • Machann J.
      • Katsuyama H.
      • et al.
      Empagliflozin effectively lowers liver fat content in well-controlled type 2 diabetes: a randomized, double-blind, phase 4.
      ]. Treatment with dapagliflozin reduces glucotoxicity and thereby improves SkM insulin sensitivity with a modest BW reduction, but it increases plasma glucagon and EGP [
      • Merovci A.
      • Solis-Herrera C.
      • Daniele G.
      • Eldor R.
      • Fiorentino T.V.
      • Tripathy D.
      • et al.
      Dapagliflozin improves muscle insulin sensitivity but enhances endogenous glucose production.
      ]. Recent systematic reviews and meta-analyses confirmed that SGLT2i reduce all-cause (empagliflozin, dapagliflozin) and cardiovascular mortality (empagliflozin), non-fatal myocardial infarction and kidney failure [
      • Tsapas A.
      • Avgerinos I.
      • Karagiannis T.
      • Malandris K.
      • Manolopoulos A.
      • Andreadis P.
      • et al.
      Comparative effectiveness of glucose-lowering drugs for type 2 diabetes: a systematic review and network meta-analysis.
      ,
      • Palmer S.C.
      • Tendal B.
      • Mustafa R.A.
      • Vandvik P.O.
      • Li S.
      • Hao Q.
      • et al.
      Sodium-glucose cotransporter protein-2 (SGLT-2) inhibitors and glucagon-like peptide-1 (GLP-1) receptor agonists for type 2 diabetes: systematic review and network meta-analysis of randomised controlled trials.
      ]. These effects were observed in metformin-treated people with T2DM at increased cardiovascular risk receiving background treatment, whereas in individuals at low cardiovascular risk the effects of SGLT2i similarly to other antidiabetic drugs did not differ from placebo for vascular outcomes [
      • Tsapas A.
      • Avgerinos I.
      • Karagiannis T.
      • Malandris K.
      • Manolopoulos A.
      • Andreadis P.
      • et al.
      Comparative effectiveness of glucose-lowering drugs for type 2 diabetes: a systematic review and network meta-analysis.
      ]. Of note, SGLT2i also decrease mortality and hospital admission for heart failure more than GLP-1Ra, but have no effect on stroke [
      • Palmer S.C.
      • Tendal B.
      • Mustafa R.A.
      • Vandvik P.O.
      • Li S.
      • Hao Q.
      • et al.
      Sodium-glucose cotransporter protein-2 (SGLT-2) inhibitors and glucagon-like peptide-1 (GLP-1) receptor agonists for type 2 diabetes: systematic review and network meta-analysis of randomised controlled trials.
      ]. Because of their clinical efficacy, SGLT2i are emerging as a treatment option for people with cardiorenal complications regardless of the presence of T2DM.

      4.4 Drugs in clinical trials affecting IR

      4.4.1 Anti-inflammatory treatment concepts

      Given that inflammation induces IR and that low-grade inflammation associates with IR, obesity and T2DM, it is suggestive to improve IR by inhibiting inflammation, i. e. by anti-inflammatory drugs. Of note, glucocorticoids rather cause IR and hyperglycemia due to their metabolic effects and statins diminish circulating inflammatory markers, but increase the risk of IR and of T2DM. Among the non-steroidal anti-inflammatory drugs, acetylsalicylic acid decreases EGP, FA and TG and enhances peripheral glucose disposal [
      • Hundal R.S.
      • Petersen K.F.
      • Mayerson A.B.
      • Randhawa P.S.
      • Inzucchi S.
      • Shoelson S.E.
      • et al.
      Mechanism by which high-dose aspirin improves glucose metabolism in type 2 diabetes.
      ]. Likewise, the salicylate, salsalate, increases adiponectin, glucose utilization during HEC and glycemic control in obesity and T2DM [
      • Goldfine A.B.
      • Shoelson S.E.
      Therapeutic approaches targeting inflammation for diabetes and associated cardiovascular risk.
      ].
      The non-steroidal farnesoid X (FX)-Ra, PX-104, also improves insulin sensitivity and alters gut microbiota in NAFLD [
      • Traussnigg S.
      • Halilbasic E.
      • Hofer H.
      • Munda P.
      • Stojakovic T.
      • Fauler G.
      • et al.
      Open-label phase II study evaluating safety and efficacy of the non-steroidal farnesoid X receptor agonist PX-104 in non-alcoholic fatty liver disease.
      ]. Of note, the steroidal FX-Ra, obeticholic acid, reduces markers of hepatic inflammation and ameliorates insulin sensitivity in humans with T2DM and NAFLD [
      • Mudaliar S.
      • Henry R.R.
      • Sanyal A.J.
      • Morrow L.
      • Marschall H.U.
      • Kipnes M.
      • et al.
      Efficacy and safety of the farnesoid X receptor agonist obeticholic acid in patients with type 2 diabetes and nonalcoholic fatty liver disease.
      ], but increases HOMA-IR in NASH [
      • Neuschwander-Tetri B.A.
      • Loomba R.
      • Sanyal A.J.
      • Lavine J.E.
      • Van Natta M.L.
      • Abdelmalek M.F.
      • et al.
      Farnesoid X nuclear receptor ligand obeticholic acid for non-cirrhotic, non-alcoholic steatohepatitis (FLINT): a multicentre, randomised, placebo-controlled trial.
      ]. Also, cenicriviroc, a dual chemokine receptor (CCR) 2/5 antagonist, reduces hepatic fibrosis, and biomarkers of systemic inflammation, but fails to affect insulin sensitivity in a phase-2 RCT in adults with NASH [
      • Friedman S.L.
      • Ratziu V.
      • Harrison S.A.
      • Abdelmalek M.F.
      • Aithal G.P.
      • Caballeria J.
      • et al.
      A randomized, placebo-controlled trial of cenicriviroc for treatment of nonalcoholic steatohepatitis with fibrosis.
      ].
      Another concept relies on the use of blocking antibodies directed against pro-inflammatory cytokines. Inhibition of TNFα by using various monoclonal antibodies (infliximab, adalimumab, etanercept), reduces IR in individuals with inflammatory diseases, although these studies were not randomized [
      • Goldfine A.B.
      • Shoelson S.E.
      Therapeutic approaches targeting inflammation for diabetes and associated cardiovascular risk.
      ], but not in a phase-4 RCT in obese humans [
      • Wascher T.C.
      • Lindeman J.H.
      • Sourij H.
      • Kooistra T.
      • Pacini G.
      • Roden M.
      Chronic TNF-alpha neutralization does not improve insulin resistance or endothelial function in “healthy” men with metabolic syndrome.
      ]. Inhibition of IL-1β by monoclonal antibodies (gevokizumab, canakinumab, LY2189102) or an IL-1R antagonist in phase-2/3 RCTs reduces HbA1c and increases insulin secretion without effects on insulin sensitivity in T2DM [
      • Goldfine A.B.
      • Shoelson S.E.
      Therapeutic approaches targeting inflammation for diabetes and associated cardiovascular risk.
      ]. The anti-IL-6 antibody tocilizumab increases adiponectin levels and reduces HOMA-IR in obese individuals in a placebo-controlled RCT [
      • Wueest S.
      • Seelig E.
      • Timper K.
      • Lyngbaek M.P.
      • Karstoft K.
      • Donath M.Y.
      • et al.
      IL-6 receptor blockade increases circulating adiponectin levels in people with obesity: an explanatory analysis.
      ].

      4.4.2 Modulation of lipid and energy metabolism

      Several drugs that target hepatic lipid metabolism have beneficial effects on IR, such as a ketohexokinase inhibitor, which decreases DNL in humans with NAFLD, or a protein tyrosine phosphatase-1B inhibitor, which reduces BW and increases adiponectin and insulin sensitivity in obese individuals with T2DM, or a synthetic ω3-FA, which decreases TG, markers of inflammation and HOMA-IR in obese people with NAFLD. These drug concepts have been recently reviewed elsewhere [
      • Gastaldelli A.
      • Stefan N.
      • Haring H.U.
      Liver-targeting drugs and their effect on blood glucose and hepatic lipids.
      ].
      Modulators of energy metabolism, which enhance FAO (mitochondrial uncouplers, thyromimetics, FGF analogs) and regulate mitochondrial biogenesis and homeostasis in SkM (nicotinamide adenine dinucleotide precursors), are briefly summarized in the next sections.

      4.4.2.1 Mitochondria uncouplers

      The mitochondrial uncoupler 2,4-dinitrophenol (DNP) was used for the treatment of obesity, before being discontinued due to its severe adverse effects [
      • Dewidar B.
      • Kahl S.
      • Pafili K.
      • Roden M.
      Metabolic liver disease in diabetes - from mechanisms to clinical trials.
      ]. To overcome its adverse effects, the liver-targeted agents DNP-methyl ether (DNPME) and controlled-release mitochondrial protonophore (CRMP) were developed. Both agents enhance hepatic FAO and reduce ectopic lipid deposition without changes in BW, resulting in improved hepatic steatosis and whole-body IR in rodent models and nonhuman primates [
      • Goedeke L.
      • Shulman G.I.
      Therapeutic potential of mitochondrial uncouplers for the treatment of metabolic associated fatty liver disease and NASH.
      ]. Conversely, BAM15 decreases BW without affecting food intake, and improves whole-body insulin sensitivity, hepatic FAO, steatosis and inflammation in obese mice [
      • Goedeke L.
      • Shulman G.I.
      Therapeutic potential of mitochondrial uncouplers for the treatment of metabolic associated fatty liver disease and NASH.
      ].

      4.4.2.2 Thyromimetics

      The thyroid hormone receptor (THR)-β agonists, sobetirome and KB-2115, stimulate hepatic FAO and reduce steatosis in rodents but they worsen SkM IR [
      • Vatner D.F.
      • Weismann D.
      • Beddow S.A.
      • Kumashiro N.
      • Erion D.M.
      • Liao X.H.
      • et al.
      Thyroid hormone receptor-beta agonists prevent hepatic steatosis in fat-fed rats but impair insulin sensitivity via discrete pathways.
      ]. Currently, none of these agents reached phase-2 trials. Resmetirom and VK2809 improve hepatic lipid metabolism and NAFLD in phase-2 studies without effects on IR [
      • Gastaldelli A.
      • Stefan N.
      • Haring H.U.
      Liver-targeting drugs and their effect on blood glucose and hepatic lipids.
      ].

      4.4.2.3 Fibroblast growth factor (FGF) analogs

      FGF21 stimulates hepatic FAO and modulates lipid and glucose metabolism. In a phase-2 RCT, administration of a PEGylated analogue of FGF21, enhances insulin sensitivity and adiponectin concentrations in obese humans with T2DM [
      • Charles E.D.
      • Neuschwander-Tetri B.A.
      • Pablo Frias J.
      • Kundu S.
      • Luo Y.
      • Tirucherai G.S.
      • et al.
      Pegbelfermin (BMS-986036), PEGylated FGF21, in patients with obesity and type 2 diabetes: results from a randomized phase 2 study.
      ].

      4.4.2.4 Nicotinamide adenine dinucleotide (NAD+) precursors

      Nicotinamide riboside (NR) and mononucleotide (NMN) are rate limiting factors in the biosynthesis of NAD+. Animal studies showed beneficial effects of NR on oxidative metabolism, IR, hyperglycemia, BW and hepatic steatosis [
      • Dollerup O.L.
      • Christensen B.
      • Svart M.
      • Schmidt M.S.
      • Sulek K.
      • Ringgaard S.
      • et al.
      A randomized placebo-controlled clinical trial of nicotinamide riboside in obese men: safety, insulin-sensitivity, and lipid-mobilizing effects.
      ]. However, 12-wk of NR supplementation does not improve IR and whole-body glucose metabolism in obese men [
      • Dollerup O.L.
      • Christensen B.
      • Svart M.
      • Schmidt M.S.
      • Sulek K.
      • Ringgaard S.
      • et al.
      A randomized placebo-controlled clinical trial of nicotinamide riboside in obese men: safety, insulin-sensitivity, and lipid-mobilizing effects.
      ]. Conversely, a recent 10-wk RCT demonstrated that NMN supplementation increases insulin-stimulated glucose disposal, assessed by HEC, and SkM insulin signaling in overweight/obese women [
      • Yoshino M.
      • Yoshino J.
      • Kayser B.D.
      • Patti G.
      • Franczyk M.P.
      • Mills K.F.
      • et al.
      Nicotinamide mononucleotide increases muscle insulin sensitivity in prediabetic women.
      ].

      5. Conclusions

      During the last decade, the discovery of new molecular mechanisms underlying the development of IR has led to the identification of new therapeutic concepts in addition to metformin and TZD, such as incretin mimetics and SGLT2i, which together can elicit greater benefits than monotherapies. Also, monoclonal antibodies targeting cytokines should be considered for treating IR in obesity and T2DM. Moreover, modulators of energy metabolism represent an alternative strategy to increase cellular energy expenditure and improve IR with or without BW loss. However, the adverse effects of some drugs and the presence of non-responders to lifestyle interventions, contribute to the progressive pandemic of obesity and T2DM. Therefore, there is an urgent need to develop effective biomarkers for early detection of IR and for the prediction of individual responses to treatment, such as Exo and microRNA, which still need further validation before their clinical application.

      CRediT authorship contribution statement

      Lucia Mastrototaro: Writing – original draft, Writing – review & editing. Michael Roden: Writing – original draft, Writing – review & editing, Supervision.

      Declaration of competing interest

      LM declares no conflicts of interest. MR has been on scientific advisory boards of Bristol-Myers Squibb, Eli Lilly, Gilead Sciences, Fishawack Group, NovoNordisk, Servier Laboratories, Target Pharmasolutions, and Terra Firma and receives investigator-initiated support from Boehringer Ingelheim , Nutricia/Danone , Sanofi–Aventis .

      Acknowledgements

      The research of the authors is supported by grants from the German Federal Ministry of Health (BMG), Ministry of Culture and Science of the State North Rhine-Westfalia (MKW NRW), Federal Ministry of Education and Research (BMBF) to German Center for Diabetes Research (DZD e.V.), European Funds for Regional Development ( EFRE-0400191 ), EUREKA Eurostars-2 (E! 113230 DIA-PEP ), German Research Foundation , Schmutzler Stiftung , German Diabetes Association and DZD e. V .

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