Angela Dardano

Type 2 diabetes mellitus is a multifactorial disease marked by progressive beta cell dysfunction; however, no reliable, easily measurable circulating biomarkers are currently available to track this decline. As microRNAs (miRNAs) tightly regulate beta cell physiology and are secreted and found in blood, we investigated whether specific circulating miRNAs reflect in vivo beta cell function in living donors. We conducted a cross-sectional study on two independent cohorts: a discovery cohort comprising 78 individuals with different levels of glucose tolerance [23 with normal glucose tolerance (NGT), 22 with impaired glucose tolerance (IGT) and 33 with type 2 diabetes] and a validation cohort comprising 158 individuals (71 non-diabetic individuals and 87 with type 2 diabetes) who were administered an oral glucose tolerance test (OGTT) and/or a mixed meal test (MMT) with measurement of glucose, insulin and C-peptide. We profiled plasma RNA by small RNA sequencing, and analysed differences between glucose tolerance groups using DESeq2. We used linear regression analyses to determine the association between miRNA abundance and clinical and metabolic measures. We validated selected miRNAs using droplet digital PCR (ddPCR). Finally, we specified a set of features that were used as an input for a LASSO model to select a biomarker signature that was able to estimate beta cell rate sensitivity (RS) values as a measure of beta cell function. We identified 11 miRNAs that were differentially expressed across the three glucose tolerance groups of the discovery cohort. Integrated analyses pinpointed a three-miRNA signature (miR-34a-5p, miR-1306-5p and miR-335-5p) that correlated with beta cell RS, which is an early indicator of insulin secretory impairment. We incorporated this panel of selected miRNAs with age and 1 h post-load glucose into a LASSO regression model that enabled discrimination of RS values (Spearman's ρ=0.43, p<0.05). We also confirmed model performance in an additional cohort of 71 non-diabetic individuals and 87 individuals with type 2 diabetes (Spearman's ρ=0.23, p<0.05). To provide proof of concept for potential clinical use, we substituted basal glucose for 1 h post-load glucose while retaining miRNA levels and age; this adaptation preserved the significance of the model in the discovery cohort (Spearman's ρ=0.46, p<0.05), and showed an interesting, but not significant, association in the validation cohort (Spearman's ρ=0.15, p=0.061). This three-miRNA signature combined with simple clinical parameters constitutes a promising biomarker for early beta cell dysfunction and type 2 diabetes progression that deserves further validation in additional cohorts.

Introduction: Sarcopenia is characterised by loss of muscle mass and strength. Although ageing is the most likely risk factor of sarcopenia, sarcopenia is prevalent even in non-elderly people. Type 2 diabetes (T2D) is a risk factor for sarcopenia, as T2D shares with sarcopenia several aetiological factors. Meanwhile, gestational diabetes mellitus (GDM) is characterised by metabolic alterations that resemble those observed in T2D, including increased insulin resistance (present even in physiologic pregnancies). Hence, GDM presents two major risk factors for sarcopenia, that is, dysglycaemia and insulin resistance. Moreover, the number of pregnancies at age >40 years is increasing, which is in an age range in which sarcopenia prevalence is already not negligible. However, data on the prevalence of sarcopenia prevalence in GDM and its effect on pregnancy outcomes are limited. Thus, this study aims to evaluate the prevalence of sarcopenia in women with GDM (and in pregnant women without GDM), identify risk factors and determine its effect on delivery and maternal and fetal outcomes. Methods and analysis: For this study, 100 each of women with and without GDM will be recruited. Women will undergo an oral glucose tolerance test within weeks 24–28 for possible GDM diagnosis (in weeks 16–18 for high-risk women). Muscle/physical performance tests will be conducted at weeks 28–32 for possible diagnosis of sarcopenia/presarcopenia. Cognitive function will also be assessed. For all women, information regarding pregnancy progression, along with any complications, will be collected. Collected data will be analysed according to the main objectives of the study: (i) determine the prevalence of sarcopenia/presarcopenia in pregnancy with and without GDM, (ii) identify factors associated with sarcopenia risk, (iii) determine the effect of sarcopenia/presarcopenia on pregnancy outcomes, (iv) explore the relationship between sarcopenia and cognitive function. Therefore, this study will provide information on sarcopenia/presarcopenia prevalence in GDM and, possibly, in pregnancy not complicated by dysglycaemia. Furthermore, the study will provide knowledge on the main factors associated with sarcopenia/presarcopenia in GDM/pregnancy. The identification of such factors will be relevant for an initial guidance for treatments that may prevent sarcopenia in GDM/pregnant women. This will become of even greater interest if sarcopenia/presarcopenia influences pregnancy outcomes, especially in GDM women. Ethics and dissemination: The study protocol has been approved by the Comitato Etico Regione Toscana - Area Vasta Nord Ovest (CEAVNO) on 25 July 2024 and by the Local Ethics Committee of the Medical University of Vienna on 17 June 2024. Participants’ enrolment began in May 2025. The results of the study will be presented at national and international conferences and in peer-reviewed journals. Trial registration number: ClinicalTrials.gov Identifier: NCT06876090 ; Registration Date: 2025-03-14

Background: Diabetes-associated neurodegeneration is amplified by methylglyoxal (MGO)-driven dicarbonyl stress linking hyperglycemia to neuronal insulin resistance and maladaptive neuroinflammation. We tested the neuroprotective activity of MNP-021, a non-electrophilic TRPA1 modulator, in neurons and glial cells in vitro . Methods: SH-SY5Y neurons were pretreated with MNP-021 and challenged with MGO, then profiled by high-content imaging, RNA-seq, Seahorse OCR/ECAR, glycolytic stress assays and AKT/ERK/CREB immunoblotting ± insulin. In parallel, HMC3 glial cells were treated with MNP-021, exposed to LPS/TNF-α or Aβ _(25–35) and tested for viability and inflammatory markers by ELISA and qRT-PCR. Results: MGO increased nucleus-to-cytoplasm area ratio by 49% and dysregulated glucose handling, increasing 2-NBDG uptake by ∼25%, with GLUT1/GLUT4 membrane redistribution; MNP-021 normalized morphology, uptake, and transporter localization without cytotoxicity up to 10 µM. RNA-seq identified 754 MGO-deregulated genes, including ISR/metabolic nodes (GCK, SESN2, PHGDH/PSAT1, PCK2); MNP-021 buffered stress-induced transcription with limited baseline effects, remodeled mitochondrial redox readouts consistent with controlled ROS signaling, while improving mitochondrial content/architecture and blunting stress-evoked compensatory glycolysis. MNP-021 restored pro-survival signaling (pAKT/pERK and nuclear pCREB), including insulin responsiveness during MGO exposure. MNP-021 reduced IL-6/TNF-α release while increasing IL-10 and ARG1 (∼1.9-fold vs LPS/TNF-α) in HMC3 glial cells, shifting them toward a pro-resolving IL-10/ARG1 program with reduced Aβ _(25–35)- evoked cytokine release with GLP-1 remaining very low (≤10 pg/mL) and not significantly increased in this system. Conclusions: MNP-021 coordinates transcriptomic restraint, transporter-level glucose handling, mitochondrial resilience, and pro-survival/pro-resolving signaling across neuron–microglia compartments, supporting TRPA1-tuned small-molecule modulation as a candidate strategy against dicarbonyl-linked neuro-metabolic stress.

Aims/hypothesis: Type 2 diabetes mellitus (T2D) is a multifactorial disease marked by progressive beta cell dysfunction; however, no reliable, easily measurable and non-invasive circulating biomarkers are currently available to track this decline. Since microRNAs (miRNAs) tightly regulate beta cell physiology and are secreted and found in blood, we investigated whether specific circulating miRNAs may reflect in vivo beta cell function in living donors. Methods: We conducted a cross-sectional study on two independent cohorts: a discovery cohort composed by 78 individuals with different glucose tolerance [23 with normal glucose tolerance (NGT), 22 with impaired glucose tolerance (IGT), and 33 with T2D] and a validation cohort composed by 158 subjects [71 non-diabetic (ND) and 87 with T2D] who were administered an oral glucose tolerance test (OGTT) and/or a mixed meal test (MMT) with measurement of glucose, insulin and C-peptide. Plasma RNA was profiled by small RNA sequencing and differences between glucose tolerance groups were analysed using DESeq2. Linear regression analyses were used to associate miRNA abundance to clinical and metabolic measures. Droplet Digital PCR (ddPCR) was used to validate selected miRNAs. A LASSO model was used to select a set of features predictive of beta cell function. Results: We identified eleven differentially expressed miRNAs across the three glucose tolerance groups of the discovery cohort. Integrated analyses pinpointed a three-miRNAs signature (miR-34a-5p, miR-1306-5p and miR-335-5p) each significantly correlated with beta cell rate sensitivity (RS), an early indicator of insulin secretory impairment. Incorporating this panel with age and 1-hour post-load glucose into a LASSO regression model enabled accurate prediction of RS (Spearman ρ=0.43, p<0.05). Model performance was confirmed in the validation cohort of 71 ND and 87 T2D subjects (Spearman ρ=0.23, p<0.05). To streamline its clinical use, we substituted fasting glucose for 1-hour post-load glucose while retaining miRNA levels and age; this adaptation preserved predictive power (Spearman ρ=0.46, p<0.05 discovery; ρ=0.15, p=0.06 validation). Conclusion/interpretation: This three-miRNAs signature combined with simple clinical parameters constitutes a promising, non-invasive biomarker for early beta cell dysfunction and T2D progression. Research in context: What is already known about this subject?: Beta cell dysfunction is a key event in Type 2 Diabetes (T2D) pathogenesis Circulating microRNAs (miRNAs) are biomarkers linked to T2D onset and T2D-associated metabolic abnormalities What is the key question?: Can specific circulating miRNAs serve as reliable biomarkers of beta cell dysfunction across different stages of glucose tolerance and Type 2 Diabetes (T2D)? What are the new findings?: Small RNA profiling of plasma samples from deep metabolically characterized NGT, IGT, and T2D subjects revealed circulating miRNAs significantly associated with in-vivo beta cell function, reflecting key mechanisms underlying pancreatic beta cell physiology A specific set of three circulating miRNAs- miR-34a-5p, miR-1306-5p and miR-335-5p - resulted linked to beta cell dysfunction This three-miRNA signature can predict early alterations in insulin secretion, representing a promising non-invasive biomarker for the early identification of individuals at risk of developing T2D How might this impact on clinical practice in the foreseeable future?: Integrating circulating miRNA profiling with routine baseline clinical parameters could provide a simpler, non-invasive, and more accessible method to identify individuals at risk of beta cell dysfunction and T2D. Implementing miRNA-based diagnostics could facilitate personalized monitoring of disease progression. Graphical

OBJECTIVE: To explore the complementary effects of a combination of dipeptidyl peptidase 4 and sodium–glucose cotransporter 2 inhibitors added to metformin on hormonal and metabolic responses to meal ingestion. RESEARCH DESIGN AND METHODS: Forty-five patients (age 58 ± 8 years; HbA1c 58 ± 6 mmol/mol; BMI 30.7 ± 3.2 kg/m2) with type 2 diabetes uncontrolled with metformin were evaluated at baseline and 3 and 28 days after 5 mg saxagliptin (SAXA), 10 mg dapagliflozin (DAPA), or 5 mg saxagliptin plus 10 mg dapagliflozin (SAXA+DAPA) using a mixed-meal tolerance test (MMTT) spiked with dual-tracer glucose to assess glucose metabolism, insulin secretion, and sensitivity. RESULTS: At day 3, fasting and mean MMTT glucose levels were lower with SAXA+DAPA (−31.1 ± 1.6 and −91.5 ± 12.4 mg/dL) than with SAXA (−7.1 ± 2.1 and −53 ± 10.5 mg/dL) or DAPA (−17.0 ± 1.1 and −42.6 ± 10.0 mg/dL, respectively; P < 0.001). Insulin secretion rate (SAXA+DAPA +75%; SAXA +11%; DAPA +3%) and insulin sensitivity (+2.2 ± 1.7, +0.4 ± 0.7, and +0.4 ± 0.4 mg ⋅ kg−1⋅ min−1, respectively) improved with SAXA+DAPA (P < 0.007). Mean glucagon-like peptide 1 (GLP-1) was higher with SAXA+DAPA than with SAXA or DAPA. Fasting glucagon increased with DAPA and SAXA+DAPA but not with SAXA. Fasting endogenous glucose production (EGP) increased with SAXA+DAPA and DAPA. During MMTT, EGP suppression was greater (48%) with SAXA+DAPA (vs. SAXA 44%; P = 0.02 or DAPA 34%; P = 0.2). Metabolic clearance rate of glucose (MCRglu) increased more with SAXA+DAPA. At week 4, insulin secretion rate, β-cell glucose sensitivity, and insulin sensitivity had further increased in the SAXA+DAPA group (P = 0.02), with no additional changes in GLP-1, glucagon, fasting or MMTT EGP, or MCRglu. CONCLUSIONS: SAXA+DAPA provided superior glycemic control compared with DAPA or SAXA, with improved β-cell function, insulin sensitivity, GLP-1 availability, and glucose clearance.

Background: The mechanism through which sodium-glucose cotransporter 2 inhibitors (SGLT2i) prevent the incidence of heart failure and/or affect cardiac structure and function remains unclear. Methods: The EMPA-HEART trial is aimed at verifying whether empagliflozin improves myocardial contractility (left ventricle global longitudinal strain, LV-GLS) and/or cardiopulmonary fitness (peak oxygen uptake, VO2peak) in subjects with type 2 diabetes (T2D) without heart disease. Patients with T2D, normal LV systolic function (2D-Echo EF > 50%), and no heart disease were randomized to either empagliflozin 10 mg or sitagliptin 100 mg for 6 months and underwent repeated cardiopulmonary exercise tests with echocardiography and determination of plasma biomarkers. Results: Forty-four patients completed the study, 22 per arm. Despite comparable glycaemic control, modest reductions in body weight (− 1.6; [− 2.7/− 0.5] kg, p = 0.03) and plasma uric acid (− 1.5; [− 2.3/− 0.6], p = 0.002), as well as an increase in haemoglobin (+ 0.7; [+ 0.2/+ 1.1] g/dL, p = 0.0003) were evident with empagliflozin. No difference was detectable in either LV-GLS at 1 month (empagliflozin vs sitagliptin: + 0.44; [− 0.10/+ 0.98]%, p = 0.11) and 6 months of therapy (+ 0.53; [− 0.56/+ 1.62]%), or in VO_2peak (+ 0.43; [− 1.4/+ 2.3] mL/min/kg, p = 0.65). With empagliflozin, the subgroup with baseline LV-GLS below the median experienced a greater increase (time*drug p < 0.05) in LV-GLS at 1 month (+ 1.22; [+ 0.31/+ 2.13]%) and 6 months (+ 2.05; [+ 1.14/+ 2.96]%), while sitagliptin induced a modest improvement in LV-GLS only at 6 months (+ 0.92; [+ 0.21/+ 0.62]%). Conclusions: Empagliflozin has neutral impact on both LV-GLS and exercise tolerance in subjects with T2D and normal left ventricular function. However, in patients with subclinical dysfunction (LV-GLS < 16.5%) it produces a rapid and sustained amelioration of LV contractility. Trial registration EUDRACT Code 2016-002225-10

Background/Objectives The link between obesity and brain function is a fascinating but still an enigmatic topic. We evaluated the effect of Roux-en-Y gastric bypass (RYGB) on peripheral glucose metabolism, insulin sensitivity, brain glucose utilization and cognitive abilities in people with obesity. Subjects/Methods Thirteen subjects with obesity (F/M 11/2; age 44.4 ± 9.8 years; BMI 46.1 ± 4.9 kg/m²) underwent 75-g OGTT during a [18F]FDG dynamic brain PET/CT study at baseline and 6 months after RYGB. At the same timepoints, cognitive performance was tested with Mini Mental State Examination (MMSE), Montreal Cognitive Assessment (MoCA), Trail making test (TMT) and Token test (TT). Glucose, insulin, C-peptide, GLP-1, GIP, and VIP levels were measured during OGTT. Leptin and BDNF levels were measured before glucose ingestion. Results RYGB resulted in significant weight loss (from 46.1 ± 4.9 to 35.3 ± 5.0 kg/m²; p < 0.01 vs baseline). Insulin sensitivity improved (disposition index: from 1.1 ± 0.2 to 2.9 ± 1.1; p = 0.02) and cerebral glucose metabolic rate (CMRg) declined in various brain areas (all p ≤ 0.01). MMSE and MoCA score significantly improved (p = 0.001 and p = 0.002, respectively). TMT and TT scores showed a slight improvement. A positive correlation was found between CMRg change and HOMA-IR change in the caudate nucleus (ρ = 0.65, p = 0.01). Fasting leptin decreased (from 80.4 ± 13.0 to 16.1 ± 2.4 ng/dl; p = 0.001) and correlated with CMRg change in the hippocampus (ρ = 0.50; p = 0.008). CMRg change was correlated with cognitive scores changes on the TMT and TT (all p = 0.04 or less). Conclusions Bariatric surgery improves CMRg directly related to a better cognitive testing result. This study highlights the potential pleiotropic effects of bariatric surgery. Trial registry number NCT03414333.

We report the case of a 32-year-old woman with a history of increased sleep need and difficulty waking up; the diagnosis of idiopathic hypersomnia was hypothesized. During ambulatory polysomnography (PSG), the patient presented an episode characterized by loss of consciousness and jerking of the four limbs. A video-PSG monitoring was performed and the patient showed unresponsiveness and drowsiness at 7 a.m. During the episode, EEG showed theta–delta diffuse activity, and blood glucose level was 32 mg dl⁻¹. The diagnosis of insulinoma was then assumed; CT scan showed a hypodense mass into the pancreatic tail, and a partial pancreasectomy was performed. The described symptoms disappeared, and 5 years later the findings of a complete clinical and neurophysiological examination were negative. The clinical picture of insulinoma presenting with paroxysmal disorders has been previously described; however, whereas hypersomnia is uncommon, in the current case it represents the main symptom. Clinicians should keep in mind that neuroglycopenia should be considered in the differential diagnosis of patients with hypersomnia, particularly if the clinical scenario does not conform to standard criteria.