DPP-4 Inhibitors Have Different Effects on Endothelial Low-Grade Inflammation and on the M1-M2 Macrophage Polarization Under Hyperglycemic Conditions.

PURPOSE: We explored the anti-inflammatory role of the DPP-4 inhibitor teneligliptin, using sitagliptin as comparator, in different in vitro models of low-grade inflammation (LGI), evaluating the hyperglycemia-induced endothelial inflammation, the macrophage polarization, and the endothelium-macrophage interaction. METHODS: The effects of DPP-4 and its inhibitors on macrophage polarization were evaluated in THP-1 cells by measuring mRNA expression of M1-M2 markers. HUVEC cells were used to analyze the effects of DPP-4 inhibitors on endothelial inflammation under normal and high glucose conditions. To evaluate the link between eNO and M1-M2 polarization, HUVECs were transfected with eNOS siRNA and co-cultured with THP-1 cells. The effects of DPP-4 inhibitors on macrophage polarization and eNO content were evaluated in a co-culture model of differentiated THP-1 cells + HUVECs under normal glucose (NG), high glucose (HG) and high metabolic memory (HM) conditions. RESULTS: DPP-4 regulated M1/M2 macrophage polarization. Teneligliptin reduced M1 and enhanced M2 macrophage phenotype under DPP-4 stimulation, and attenuated hyperglycemia-induced endothelial inflammation. In THP-1 cells co-cultured with eNOS depleted HUVECs, M1 markers were enhanced, while M2 reduced, indicating an important role of eNO in polarization to M2 phenotype. In the co-culture model with HUVECs exposed to HG and HM, teneligliptin reduced M1 and enhanced M2 population, by increasing eNO levels. The anti-inflammatory effects of sitagliptin were not observed in these LGI models. CONCLUSION: Teneligliptin, but not sitagliptin, has anti-inflammatory effects in the various LGI models, by promoting a switch from M1 toward M2 phenotype and by decreasing hyperglycaemia-induced endothelial inflammation, suggesting that effects for LGI are different among DPP-4 inhibitors.

Extracellular vesicle-shuttled miRNAs: a critical appraisal of their potential as nano-diagnostics and nano-therapeutics in type 2 diabetes mellitus and its cardiovascular complications.

Type 2 diabetes mellitus (T2DM) is a complex multifactorial disease causing the development of a large range of cardiovascular (CV) complications. Lifestyle changes and pharmacological therapies only partially halt T2DM progression, and existing drugs are unable to completely suppress the increased CV risk of T2DM patients. Extracellular vesicles (EV)s are membrane-coated nanoparticles released by virtually all living cells and are emerging as novel mediators of T2DM and its CV complications. As a matter of fact, several preclinical models suggest a key involvement of EVs in the initiation and/or progression of insulin resistance, ß-cell dysfunction, diabetic dyslipidaemia, atherosclerosis, and other T2DM complications. In addition, preliminary findings also suggest that EV-associated molecular cargo, and in particular the miRNA repertoire, may provide with useful diagnostic and/or prognostic information for the management of T2DM. Here, we review the latest findings showing that EV biology is altered during the entire trajectory of T2DM, i.e. from diagnosis to development of CV complications. We also critically highlight the potential of this emerging research field, by describing both preclinical and clinical observations, and the limitations that must be overcome to translate the preclinical findings into the development of EV-based nano-diagnostic and/or nano-therapeutic tools. Finally, we summarize how two lifestyle changes known to prevent or limit T2DM, i.e. diet and exercise, affect EV number and composition, with a focus on the possible role of EVs contained in food in shaping metabolic responses, a promising approach still in its infancy.

CD31+ Extracellular Vesicles From Patients With Type 2 Diabetes Shuttle a miRNA Signature Associated With Cardiovascular Complications.

Innovative biomarkers are needed to improve the management of patients with type 2 diabetes mellitus (T2DM). Blood circulating miRNAs have been proposed as a potential tool to detect T2DM complications, but the lack of tissue specificity, among other reasons, has hampered their translation to clinical settings. Extracellular vesicle (EV)-shuttled miRNAs have been proposed as an alternative approach. Here, we adapted an immunomagnetic bead-based method to isolate plasma CD31+ EVs to harvest vesicles deriving from tissues relevant for T2DM complications. Surface marker characterization showed that CD31+ EVs were also positive for a range of markers typical of both platelets and activated endothelial cells. After characterization, we quantified 11 candidate miRNAs associated with vascular performance and shuttled by CD31+ EVs in a large (n = 218) cross-sectional cohort of patients categorized as having T2DM without complications, having T2DM with complications, and control subjects. We found that 10 of the tested miRNAs are affected by T2DM, while the signature composed by miR-146a, -320a, -422a, and -451a efficiently identified T2DM patients with complications. Furthermore, another CD31+ EV-shuttled miRNA signature, i.e., miR-155, -320a, -342-3p, -376, and -422a, detected T2DM patients with a previous major adverse cardiovascular event. Many of these miRNAs significantly correlate with clinical variables held to play a key role in the development of complications. In addition, we show that CD31+ EVs from patients with T2DM are able to promote the expression of selected inflammatory mRNAs, i.e., CCL2, IL-1α, and TNFα, when administered to endothelial cells in vitro. Overall, these data suggest that the miRNA cargo of plasma CD31+ EVs is largely affected by T2DM and related complications, encouraging further research to explore the diagnostic potential and the functional role of these alterations.

Legacy effect of intensive glucose control on major adverse cardiovascular outcome: Systematic review and meta-analyses of trials according to different scenarios.

BACKGROUND: Early and intensive glycaemic control provides long-term protection against the development of microvascular complications, a phenomenon defined legacy effect. Whether a legacy effect of high glucose exists also on macrovascular endpoints is uncertain. METHODS AND FINDINGS: We performed a systematic review of both randomized clinical trials (RCT)s and observational studies pertinent to the research question. We searched PubMed, Embase, Scopus and the Cochrane database up to January 31th 2020. Eligibility criteria for RCTs were: 1 - efficacy assessment of intensive glucose lowering treatment vs a less-stringent/conventional treatment; 2 - the inclusion of a post-active phase, observational follow-up; 3 - enrolment of patients with T1DM, pre-diabetes, and T2DM; and 4 - data report on major adverse cardiovascular events (MACE) incidence, which was the primary endpoint of this meta-analysis. We performed multiple meta-analyses of the available RCTs according to different scenarios considering the type of diabetes, diabetes duration, the presence of previous cardiovascular events, follow-up extension, and the incidence of MACE recorded only during the observational, post-active phase of the trials. Results from observational studies reporting the association between HbA1c levels during the first year after diabetes diagnosis and subsequent MACE incidence were also collected and are reported narratively. We included data from 7 RCTs and 40,346 patients. The intensive glucose-lowering approach significantly decrease the incidence of MACE compared with conventional treatment (OR 0.86, CI 0.77-0.96; p = 0.007) when considering all the available studies, with a more consistent effect (OR 0.73, CI 0.56-0.94; p = 0.01) in the case of RCTs enrolling patients with diabetes duration <10 years, and an even more pronounced protection (OR 0.64, 48 CI 0.48, 0.86; p = 0.003) when analysing only RCTs enrolling patients without previous cardiovascular events at baseline. Considering only RCTs with a post-trial follow-up >10 years also yielded a relevant beneficial effect of the intensive approach (OR 0.71, CI 0.57, 0.88; p = 0.002). On the other hand, no effect was observed (OR 0.99, CI 0.92, 1.06; p = 0.81) when considering only the events recorded during the post-active, observational phases of the trials. Observational studies showed that HbA1c values >6.5% or 7% during the first year of diabetes diagnosis are associated with a higher incidence of late MACE with increased risk ranging from 19 up to 64%, according to the different study design and HbA1c stratification. CONCLUSIONS: These results support the recommendation regarding glucose-lowering treatment intensification in order to decrease the probability of having a macrovascular event in patients with short diabetes duration, no prevalent cardiovascular diseases, and long life-expectancy. On the other side, data from RCTs do not support the existence of a protective legacy effect on the macrovasculature beyond the period of intensive glycaemic treatment.

Extracellular vesicles circulating in young organisms promote healthy longevity.

Parabiosis experiments in mice demonstrated that a young environment could partially rejuvenate multiple tissues of old organisms. However, the circulating mediators responsible of such effect have been elusive so far. Novel results suggest that extracellular vesicles isolated from plasma of young mice increase lifespan in old mice. Here we integrate these findings in a larger framework, advancing the hypothesis that circulating vesicles may mediate the beneficial effect of a young milieu on ageing.

The Unique Pharmacological and Pharmacokinetic Profile of Teneligliptin: Implications for Clinical Practice.

Teneligliptin is a dipeptidyl peptidase-4 (DPP-4) inhibitor that was approved for the treatment of type 2 diabetes mellitus (T2DM) in Japan and Korea and is being researched in several countries. Teneligliptin is a potent, selective, and long-lasting DPP-4 inhibitor with a t½ of approximately 24 h and unique pharmacokinetic properties: it is metabolized by cytochrome P450 (CYP) 3A4 and flavin-containing monooxygenase 3 (FMO3), or excreted from the kidney in an unchanged form. Because of its multiple elimination pathways, dose adjustment is not needed in patients with hepatic or renal impairment, and it is considered to have a low potential for drug-drug interactions. Clinical studies and postmarketing surveillance show that teneligliptin, administered as monotherapy and/or in combination with antihyperglycemic agents, is effective and well tolerated in T2DM patients, including in elderly patients and those with renal impairment. Furthermore, teneligliptin has antioxidative properties, which induce the antioxidant cascade, as well as ·OH scavenging properties. In addition, it has shown endothelial protective effects in several non-clinical and clinical studies. From its unique profile and clinical data, teneligliptin represents a potential therapeutic option in a wide variety of patients, including elderly diabetic patients and those with renal impairment. The fixed-dose combination (FDC) tablet of teneligliptin and canagliflozin has been approved in Japan; this is the first FDC tablet of a DPP-4 inhibitor and sodium glucose co-transporter 2 inhibitor in Japan, and the third globally. The FDC tablet may also provide additional prescribing and adherence benefits.

Pleiotropic effects of metformin: Shaping the microbiome to manage type 2 diabetes and postpone ageing.

Metformin is the first-choice therapy to lower glycaemia and manage type 2 diabetes. Continuously emerging epidemiological data and experimental models are showing additional protective effects of metformin against a number of age-related diseases (ARDs), e.g., cardiovascular diseases and cancer. This evidence has prompted the design of a specific trial, i.e., the Targeting Aging with Metformin (TAME) trial, to test metformin as an anti-ageing molecule. However, a unifying or prevailing mechanism of action of metformin is still debated. Here, we summarize the epidemiological data linking metformin to ARD prevention. Then, we dissect the deeply studied mechanisms of action explaining its antihyperglycemic effect and the putative mechanisms supporting its anti-ageing properties, focusing on studies using clinically pertinent doses. We hypothesize that the molecular observations obtained in different models with metformin could be indirectly mediated by its effect on gut flora. Novel evidence suggests that metformin reshapes the human microbiota, promoting the growth of beneficial bacterial species and counteracting the expansion of detrimental bacterial species. In turn, this action would influence the balance between pro- and anti-inflammatory circulating factors, thereby promoting glycaemic control and healthy ageing. This framework may reconcile diverse observations, providing information for designing further studies to elucidate the complex interplay between metformin and the metabiome harboured in mammalian body compartments, thereby paving the way for innovative, bacterial-based therapeutics to manage type 2 diabetes and foster a longer healthspan.

Increases in circulating levels of ketone bodies and cardiovascular protection with SGLT2 inhibitors: Is low-grade inflammation the neglected component?

Recent clinical trials have demonstrated a strong cardiovascular (CV) protective effect of sodium/glucose cotransporter (SGLT) 2 inhibitors, a recently introduced class of hypoglycaemic agents. The improvement in glycated haemoglobin and other conventional risk factors explains only a portion of the observed reduction in CV risk. A relevant feature of SGLT2-inhibitor-treated diabetic patients is the increase in circulating levels of ketone bodies, which has been proposed to mediate part of the beneficial effects of this class of drugs, mainly through their bioenergetic properties. However, ketone bodies are emerging as potent anti-inflammatory molecules, and inflammation is a recognized risk factor for the development of CV events. In this framework, we hypothesize that, through their unique mechanism of action and by increasing circulating ketone bodies, SGLT2 inhibitors indirectly target the IL-1ß pathway and thus produce a consistent amelioration of low-grade inflammation, a clinically relevant phenomenon in diabetic patients with high CV risk. This attenuation could slow the progression of CV disease and especially the atherosclerotic process, which is sensitive to environmental changes, even over a short time period. To test this conceptual structure, it would be necessary to measure circulating pro-inflammatory molecules in patients treated with SGLT inhibitors. The addition of inflammatory markers to the list of clinical data measured in FDA-requested, large CV outcome trials could provide supplementary information regarding potential secondary effects of new anti-hyperglycaemic drugs, considering that the inflammatory process is an often neglected cornerstone of CV diseases.

Novel insights into the regulation of miRNA transcriptional control: implications for T2D and related complications.

In recent years, epigenetics has emerged as an important form of biological regulation involving chromatin control of gene expression. The mechanisms of this fine-tuned regulation are susceptible to changes forced by environmental stimuli and nutritional factors and may be potentially reversible. Dysregulation of epigenetic processes has important consequences for the pathogenesis of complex and multifactorial diseases such as type 2 diabetes (T2D) and vascular complications. Along with DNA methylation (DNA-me), histone modifications and RNA-based mechanisms as the major epigenetic controllers, small non-coding RNAs known as microRNAs (miRNAs) have their own important implications for the pathogenesis of diabetes. There is increasing evidence supporting the role of miRNAs in modulating gene expression, cumulatively contributing to epigenetic gene silencing by acting either on the methylation status of the cells or in alternative roles. Although significant progress has been made in the characterization of miRNA functions, most miRNA promoters have not yet been characterized, and the transcriptional regulation of miRNAs remains elusive. The present work is centred on the new biological insights pertaining to the epigenetics-miRNA regulatory axis, focusing on the development of T2D and cardiovascular complications, and the ability of these mechanisms to interact in a network of DNA-me regulation. The genomic organization of inter- and intragenic miRNA genes is discussed, and the mutual connections between pre-mRNA splicing and miRNA biogenesis are summarized, along with the discovery of novel miRNA transcriptional regulation sites.

Teneligliptin enhances the beneficial effects of GLP-1 in endothelial cells exposed to hyperglycemic conditions.

High-glucose-induced oxidative stress contributes to cardiovascular endothelial damage in diabetes. Glucagon-like peptide 1 (GLP-1) is beneficial to endothelial cells, but its effects are diminished when cells are continuously exposed to high glucose. Teneligliptin is a dipeptidyl peptidase-4 (DPP-4) inhibitor that prevents oxidative stress, apoptosis and the metabolic memory effect. We explored the potential additive effects of Teneligliptin and GLP-1 in hyperglycemia-damaged endothelial cells. Human umbilical vein endothelial cells (HUVECs) were exposed to normal-glucose (5 mmol/L) or high-glucose (HG, 25 mmol/L) for 21 days, or to HG for 14 days followed by normal-glucose for 7 days (HM). These cells were continually treated with Teneligliptin 3.0 µmol/L, alone or in combination with an acute dose of GLP-1 50 nmol/L. DPP-4 was upregulated under hyperglycemic conditions, but Teneligliptin reduced DPP-4 expression and activity. Simultaneous Teneligliptin and GLP-1 synergistically increased the antioxidant response and reduced ROS levels in HG- and HM-exposed HUVECs. Concurrent treatment also enhanced cell proliferation, reduced apoptotic gene expression and ameliorated endoplasmic reticulum stress in HG- and HM-exposed HUVECs. Thus, long-term Teneligliptin treatment reduced DPP-4 levels and activity in HUVECs exposed to chronic hyperglycemia. Moreover, Teneligliptin enhanced the beneficial effects of GLP-1 on oxidative stress, proliferation, apoptosis and endoplasmic reticulum homeostasis.

Inflammageing and metaflammation: The yin and yang of type 2 diabetes.

Type 2 diabetes mellitus (T2DM) is characterised by chronic low-grade inflammation, recently referred to as 'metaflammation', a relevant factor contributing to the development of both diabetes and its complications. Nonetheless, 'canonical' anti-inflammatory drugs do not yield satisfactory results in terms of prevention of diabetes progression and of cardiovascular events, suggesting that the causal mechanisms fostering metaflammation deserve further research to identify new druggable targets. Metaflammation resembles ageing-induced low-grade inflammation, previously referred to as inflammageing, in terms of clinical presentation and the molecular profile, pointing to a common aetiology for both conditions. Along with the mechanisms proposed to fuel inflammageing, here we dissect a plethora of pathological cascades triggered by gluco- and lipotoxicity, converging on candidate phenomena possibly explaining the enduring pro-inflammatory program observed in diabetic tissues, i.e. persistent immune-system stimulation, accumulation of senescent cells, epigenetic rearrangements, and alterations in microbiota composition. We discuss the possibility of harnessing these recent discoveries in future therapies for T2DM. Moreover, we review recent evidence regarding the ability of diets and physical exercise to modulate selected inflammatory pathways relevant for the diabetic pathology. Finally, we examine the latest findings showing putative anti-inflammatory mechanisms of anti-hyperglycaemic agents with proven efficacy against T2DM-induced cardiovascular complications, in order to gain insights into quickly translatable therapeutic approaches.

Short-term sustained hyperglycaemia fosters an archetypal senescence-associated secretory phenotype in endothelial cells and macrophages.

Diabetic status is characterized by chronic low-grade inflammation and an increased burden of senescent cells. Recently, the senescence-associated secretory phenotype (SASP) has been suggested as a possible source of inflammatory factors in obesity-induced type 2 diabetes. However, while senescence is a known consequence of hyperglycaemia, evidences of SASP as a result of the glycaemic insult are missing. In addition, few data are available regarding which cell types are the main SASP-spreading cells in vivo. Adopting a four-pronged approach we demonstrated that: i) an archetypal SASP response that was at least partly attributable to endothelial cells and macrophages is induced in mouse kidney after in vivo exposure to sustained hyperglycaemia; ii) reproducing a similar condition in vitro in endothelial cells and macrophages, hyperglycaemic stimulus largely phenocopies the SASP acquired during replicative senescence; iii) in endothelial cells, hyperglycaemia-induced senescence and SASP could be prevented by SOD-1 overexpression; and iiii) ex vivo circulating angiogenic cells derived from peripheral blood mononuclear cells from diabetic patients displayed features consistent with the SASP. Overall, the present findings document a direct link between hyperglycaemia and the SASP in endothelial cells and macrophages, making the SASP a highly likely contributor to the fuelling of low-grade inflammation in diabetes.

The "Metabolic Memory" Theory and the Early Treatment of Hyperglycemia in Prevention of Diabetic Complications.

Several epidemiological and prospective studies suggest that an early intensive control of hyperglycaemia is able to decrease the risk of diabetic micro- and macro-vascular complications. A growing body of experimental evidence supports the concept that the risk for diabetes complications may be linked to oxidative stress, non-enzymatic glycation of proteins, epigenetic changes, and chronic inflammation, laying the foundation for the "metabolic memory" theory. From a clinical point of view, this theory supports the need for a very early aggressive treatment, with the goal of normalizing metabolic control as soon as possible. It may also prove beneficial to introduce therapeutic agents that are able to reduce reactive species and glycation, in addition to presenting better control of glucose levels in patients with diabetes, in order to minimize long-term diabetes complications. In this review, we evaluate the effect of glucose intake and metabolism in the light of this theory.

The dipeptidyl peptidase-4 (DPP-4) inhibitor teneligliptin functions as antioxidant on human endothelial cells exposed to chronic hyperglycemia and metabolic high-glucose memory.

Dipeptidyl peptidase-4 inhibitors are widely used in type 2 diabetes. Endothelium plays a crucial role maintaining vascular integrity and function. Chronic exposure to high glucose drives to endothelial dysfunction generating oxidative stress. Teneligliptin is a novel dipeptidyl peptidase-4 inhibitor with antioxidant properties. This study is aimed to verify a potential protective action of teneligliptin in endothelial cells exposed to high glucose. Human umbilical vein endothelial cells were cultured under normal (5 mmol/L) or high glucose (25 mmol/L) during 21 days, or at high glucose during 14 days followed by 7 days at normal glucose, to reproduce the high-metabolic memory state. During this period, different concentrations of teneligliptin (0.1, 1.0 and 3.0 µmol/L) or sitagliptin (0.5 µmol/L) were added to cells. Ribonucleic acid and protein expression were assessed for antioxidant response, proliferation, apoptosis and endoplasmic reticulum stress markers. Teneligliptin promotes the antioxidant response in human umbilical vein endothelial cells, reducing ROS levels and inducing Nrf2-target genes messenger ribonucleic acid expression. Teneligliptin, but not sitagliptin, reduces the expression of the nicotine amide adenine dinucleotide phosphate oxidase regulatory subunit P22 -phox , however, both blunt the high glucose-induced increase of TXNIP. Teneligliptin improves proliferation rates in human umbilical vein endothelial cells exposed to high glucose, regulating the expression of cell-cycle inhibitors markers (P53, P21 and P27), and reducing proapoptotic genes (BAX and CASP3), while promotes BCL2 expression. Teneligliptin ameliorates high glucose-induced endoplasmic reticulum stress reducing the expression of several markers (BIP, PERK, ATF4, CHOP, IRE1a and ATF6). Teneligliptin has antioxidant properties, ameliorates oxidative stress and apoptotic phenotype and it can overcome the metabolic memory effect, induced by chronic exposure to high glucose in human endothelial cells.

"Inflammaging" as a Druggable Target: A Senescence-Associated Secretory Phenotype-Centered View of Type 2 Diabetes.

Aging is a complex phenomenon driven by a variety of molecular alterations. A relevant feature of aging is chronic low-grade inflammation, termed "inflammaging." In type 2 diabetes mellitus (T2DM), many elements of aging appear earlier or are overrepresented, including consistent inflammaging. T2DM patients have an increased death rate, associated with an incremented inflammatory score. The source of this inflammation is debated. Recently, the senescence-associated secretory phenotype (SASP) has been proposed as the main origin of inflammaging in both aging and T2DM. Different pathogenic mechanisms linked to T2DM progression and complications development have been linked to senescence and SASP, that is, oxidative stress and endoplasmic reticulum (ER) stress. Here we review the latest data connecting oxidative and ER stress with the SASP in the context of aging and T2DM, with emphasis on endothelial cells (ECs) and endothelial dysfunction. Moreover, since current medical practice is insufficient to completely suppress the increased death rate of diabetic patients, we propose a SASP-centered view of T2DM as a futuristic therapeutic option, possibly opening new prospects by moving the attention from one-organ studies of diabetes complications to a wider targeting of the aging process.

The Possible Role of Flavonoids in the Prevention of Diabetic Complications.

Type 2 diabetes mellitus is a disease that affects many metabolic pathways. It is associated with insulin resistance, impaired insulin signaling, ß-cell dysfunction, abnormal glucose levels, altered lipid metabolism, sub-clinical inflammation and increased oxidative stress. These and other unknown mechanisms lead to micro- and macro-complications, such as neuropathy, retinopathy, nephropathy and cardiovascular disease. Based on several in vitro animal models and some human studies, flavonoids appear to play a role in many of the metabolic processes involved in type 2 diabetes mellitus. In this review, we seek to highlight the most recent papers focusing on the relationship between flavonoids and main diabetic complications.

Oscillating glucose induces microRNA-185 and impairs an efficient antioxidant response in human endothelial cells.

BACKGROUND: Intracellular antioxidant response to high glucose is mediated by Cu/Mn-superoxide dismutases (SOD-1/SOD-2), catalase (CAT) and glutathione peroxidases (GPx), particularly glutathione peroxidase-1 (GPx-1). Although oscillating glucose can induce a more deleterious effect than high glucose on endothelial cells, the mechanism by which oscillating glucose exerts its dangerous effects is incompletely understood; however, the involvement of oxidative damage has been generally accepted. In this study we sought to determine whether oscillating glucose differentially modulates antioxidant response, and to elucidate the potential regulatory mechanisms exerted by the microRNA-185 (miR-185). METHODS: Human endothelial cells were exposed for 1 week to constant and oscillating high glucose. SOD-1, SOD-2, CAT and GPx-1, as well as two markers of oxidative stress [8-hydroxy-2'-deoxyguanosine (8-OHdG) and the phosphorylated form of H2AX (γ-H2AX)] were measured at the end of the experiment. Intracellular miR-185 was measured and loss-of function assays were performed in HUVEC. Bioinformatic tool was used to predict the link between miR-185 on 3'UTR of GPx-1 gene. Luciferase assay was performed to confirm the binding on HUVEC. RESULTS: After exposure to constant high glucose SOD-1 and GPx-1 increased, while in oscillating glucose SOD-1 increased and GPx-1 did not. SOD-2 and CAT remained unchanged under both conditions. A critical involvement of oscillating glucose-induced miR-185 in the dysregulation of endogenous GPx-1 was found. Computational analyses predict GPx-1 as miR-185's target. HUVEC cultures were used to confirm glucose's causal role on the expression of miR-185, its target mRNA and protein and finally the activation of antioxidant response. In vitro luciferase assays confirmed computational predictions targeting of miR-185 on 3'-UTR of GPx-1 mRNA. Knockdown of miR-185, using anti-miR-185 inhibitor, was accompanied by a significant upregulation of GPx-1 in oscillating glucose. 8-OHdG and γ-H2AX increased more in oscillating glucose than in constant high glucose. CONCLUSIONS: Glucose oscillations may exert more deleterious effects on the endothelium than high glucose, likely due to an impaired response of GPx-1, coupled by the upregulation of miR-185.

The simultaneous control of hyperglycemia and GLP-1 infusion normalize endothelial function in type 1 diabetes.

BACKGROUND: To test the effect of normoglycemia and glucagon-like peptide-1 (GLP-1), alone or in combination, on the possible normalization of endothelial function in type 1 diabetes. METHODS: Fifteen people with type 1 diabetes participated in three experiments: reaching and maintaining normoglycemia for 4h; reaching and maintaining hyperglycemia plus GLP-1 infusion for 4h; and reaching and maintaining normoglycemia for 4h with simultaneous infusion of GLP-1. RESULTS: Both normoglycemia and GLP-1 infusion restored endothelial function and decreased and plasma 8-iso prostaglandin F2α levels. However, only the combination of normoglycemia and GLP-1 was able to normalize endothelial function. CONCLUSIONS: This study confirms that long-lasting hyperglycemia in type 1 diabetes induces a permanent alteration which contributes to maintaining endothelial dysfunction even when glycemia is normalized, and that in the presence of normoglycemia, GLP-1 can contribute to normalizing endothelial function.

Dietary Betaine Supplementation Increases Fgf21 Levels to Improve Glucose Homeostasis and Reduce Hepatic Lipid Accumulation in Mice.

Identifying markers of human insulin resistance may permit development of new approaches for treatment and prevention of type 2 diabetes. To this end, we analyzed the fasting plasma metabolome in metabolically characterized human volunteers across a spectrum of insulin resistance. We demonstrate that plasma betaine levels are reduced in insulin-resistant humans and correlate closely with insulin sensitivity. Moreover, betaine administration to mice with diet-induced obesity prevents the development of impaired glucose homeostasis, reduces hepatic lipid accumulation, increases white adipose oxidative capacity, and enhances whole-body energy expenditure. In parallel with these beneficial metabolic effects, betaine supplementation robustly increased hepatic and circulating fibroblast growth factor (Fgf)21 levels. Betaine administration failed to improve glucose homeostasis and liver fat content in Fgf21(-/-) mice, demonstrating that Fgf21 is necessary for betaine's beneficial effects. Together, these data indicate that dietary betaine increases Fgf21 levels to improve metabolic health in mice and suggest that betaine supplementation merits further investigation as a supplement for treatment or prevention of type 2 diabetes in humans.