Rossella Liani

Background and Aims. Patients with type 2 diabetes mellitus (T2D) exhibit enhanced platelet reactivity and an increased risk of atherothrombotic events. Although low‑dose acetylsalicylic acid (ASA) irreversibly inhibits cyclooxygenase‑1, a substantial proportion of patients with T2D show a suboptimal antiplatelet response. Oxidative stress has been proposed as a key mechanism underlying this variability. The aim of this study was to investigate the relationship between oxidative stress–related pathways, platelet function, and ASA responsiveness in patients with T2D, focusing on platelet heme oxygenase‑1 (HO‑1) and platelet‑associated microRNAs (miRNAs). Methods. Patients with T2D receiving chronic ASA therapy (T2D+ASA, n = 55), patients with T2D not receiving ASA (T2D, n = 11), and subjects without T2D (NOT2D, n = 20) were enrolled. Chronic ASA therapy consisted of 100 mg/day for at least one month. Blood and urine samples were collected for platelet isolation and biochemical analyses. Patients in the T2D+ASA group were stratified into tertiles according to platelet HO‑1 levels (Figure 1A). Platelet miRNA expression profiles were identified by next‑generation sequencing and validated by real‑time PCR. KEGG pathway enrichment analysis was performed on differentially expressed platelet‑associated miRNAs. Results. Platelet HO‑1 levels differed significantly among groups. Patients with T2D, regardless of ASA treatment, exhibited significantly lower platelet HO‑1 levels compared with NOT2D subjects (both P < 0.001), Figure 1B. Among T2D+ASA patients, lower platelet HO‑1 levels were associated with higher levels of markers of platelet activation, including serum thromboxane B₂ (first vs third tertile, P = 0.036) and urinary 11‑dehydro‑thromboxane B₂ (P = 0.030), Figure 1C-D. A total of 62 platelet‑associated miRNAs were identified. Reduced expression of hsa‑let‑7b‑5p, hsa‑miR‑98‑5p, hsa‑miR‑423‑5p, hsa‑miR‑652‑3p, and hsa‑miR‑625‑5p, and increased expression of hsa‑miR‑21‑5p, hsa‑miR‑23a‑3p, hsa‑miR‑126‑5p, hsa‑miR‑150‑5p, hsa‑miR‑151a‑3p, and hsa‑miR‑200c‑3p were observed in patients with lower HO‑1 levels, Figure 1E-F. KEGG analysis revealed enrichment of pathways related to platelet activation, fluid shear stress, and atherosclerosis. Conclusions. Patients with T2D exhibit reduced platelet HO‑1 levels independently of chronic low‑dose ASA therapy, indicating a persistent impairment of platelet antioxidant capacity that may contribute to enhanced platelet reactivity and reduced responsiveness to antiplatelet treatment. In this clinical context, lower HO‑1 levels are associated with a reduced expression of regulatory let‑7 family members, including hsa‑let‑7b‑5p and hsa‑miR‑98‑5p, suggesting a failure of compensatory mechanisms linking platelet miRNAs to redox‑sensitive pathways, potentially due to insufficient repression of the HMOX1 transcriptional inhibitor Bach1. Consistently, platelet‑associated miRNA profiling and KEGG pathway analysis indicate that low HO‑1 levels are associated with a less adaptive modulation of platelet activation and platelet–vascular signaling pathways, which may translate into suboptimal antiplatelet responses and an increased risk of atherothrombotic events.

Microangiopathy compromises the structural and functional integrity of organs and tissues in patients with type II diabetes mellitus (T2DM) negatively affecting the perceived quality of life. Nitric oxide (NO) is a multifunctional signalling molecule, acting as a vasodilator, neurotransmitter, and modulator of inflammatory processes. Patients with type II diabetes mellitus and chronic kidney disease, controlled from glycaemic status, were treated or not with pulsed electrostatic field (PESF) cycles to evaluate effect on the perfusion of peripheral tissues. Everyone was monitored for the metabolic profile, and we tested circulating NO with a commercial enzyme immunoassay kit. In addition, we tested the perceived quality of life of patients before/after a PESF cycle using a questionnaire. Patients treated with PESF were improved circulating NO levels, significant changes in systolic and diastolic blood pressure, heart rate and were more homogeneous for their metabolic profile. The questionnaire showed also a marked improvement in the perceived quality of life. The use of pulsed electrostatic fields has allowed us to observe an improvement in the metabolic, psychological, and clinical profile in patients with T2DM and chronic kidney disease whose pathological profile is strongly compromised.

Background/objectives Diabetic subjects are at increased risk of subtle cognitive impairment since the disease early stages and of dementia later in life. In animal models, glucagon-like peptide-1 receptor agonizts (GLP1-RAs) have been shown to exert neuroprotective effects, expecially in the memory domain. We assessed whether treatment with a GLP1-RA might affect cognitive functions in type 2 diabetic subjects independently on the weight loss it might induce. Subjects/methods Forty metformin-treated obese subjects with prediabetes or newly diagnosed type 2 diabetes mellitus, received liraglutide (1.8 mg/d) (n = 20) or lifestyle counseling (dietary intervention and exercise training) (n = 20) until achieving a modest and comparable weight loss (−7% of initial body weight). Interventions/methods A detailed neuropsychological assessment before and after weight loss was completed in 16 patients per arm, who were administered a total of seven psychological tests, thus assessing three composite domain z-scores for attention, memory, and executive control. Results After comparable weight loss and superimposable glycemic control and insulin sensitivity, a significant increase in short term memory (mean Digit Span Z score from −0.06 to 0.80, p = 0.024) and memory composite z-score (mean memory z-score from −0.67 to 0.032, p = 0.0065) was observed in the liraglutide exposed subjects (between group p = 0.041 and p = 0.033, respectively). Conclusions Liraglutide might slow down memory function decline in diabetic patients in early, and possibly preclinical stages of the disease.

Background: Impaired glucose tolerance (IGT) is associated with increased cardiovascular morbidity and mortality. Enhanced thromboxane (TX)‐dependent platelet activation plays a pivotal role in atherothrombosis and characterizes type 2 diabetes mellitus (DM). Whether this also pertains to IGT is currently unknown.We investigated whether TXA₂‐dependent platelet activation, as reflected by 11‐dehydro‐TXB₂ (TXM) urinary excretion, is comparably abnormal in IGT as in DM, is persistent over long‐term follow‐up, changes as a function of metabolic disease progression, and is influenced by food intake.Methods: We prospectively investigated subjects with IGT (n = 48) and two control groups with DM diagnosed either less than 12 months (n = 60) or 12 months or more (n = 58).Results: Baseline TXM excretion was comparable between subjects with IGT and DM, with no evidence of a circadian variation. During a 36‐month follow‐up, urinary TXM excretion was stable over time in the DM groups, while tended to increase in subjects with IGT. Increasing urinary TXM excretion over time was observed in the subjects who progressed to diabetes vs nonprogressors.Conclusions: We conclude that TXA₂‐dependent platelet activation was at least as high in IGT as in patients with DM and further increased over time, especially in those who progressed to overt diabetes.

Objectives: The aim of this study was to evaluate the circulating levels of CD40 ligand (CD40 L), Dickkopf‐1 (DKK‐1) and P‐selectin, their relationships and their contributions to cardiovascular risk in subjects with HIV infection.Methods: The study population included 80 HIV‐infected patients, 14 (17.5%) of whom had diabetes mellitus (DM) and 32 (40.0%) of whom had arterial hypertension (AH). The HIV‐infected patients were compared with a control group with similar demographic and clinical features. CD40L, DKK‐1 and P‐selectin levels were measured using an enzyme‐linked immunosorbent assay.Results: The HIV‐infected patients showed higher levels of all the cardiovascular disease (CVD) markers. Both serum CD40L and DKK‐1 were significantly higher in HIV‐infected patients than in the HIV‐negative controls (P < 0.001), while soluble P‐selectin showed no significant between‐group difference (P = 0.133), reflecting the role of HIV infection in CVD. In the HIV‐infected group, patients with DM showed lower levels of CD40L and DKK‐1 in comparison with the nondiabetic patients and patients with AH (P < 0.05, with Bonferroni correction). In contrast, patients with AH showed higher levels of CD40L and DKK‐1 in comparison to patients without DM or AH (P < 0.05, with Bonferroni correction). Patients with AH showed higher levels of CD40L and DKK‐1 than patients with DM (P < 0.05, with Bonferroni correction).Conclusions: In this study, we found that HIV‐infected patients displayed significantly higher circulating levels of both CD40L and DKK‐1, which were linearly and directly correlated, when compared to HIV‐negative patients. The presence of diabetes was associated with lower levels of both CD40L and DKK‐1, whereas the presence of hypertension was associated with higher levels of CD40L.

Thromboxane (TX)-dependent platelet activation and lipid peroxidation, as reflected in vivo by the urinary excretion of 11-dehydro-TXB₂ and 8-iso-prostaglandin (PG)F_2α, play a key role in atherothrombosis in obesity and type 2 diabetes mellitus (T2DM) since the earlier stages. Thirty-five metformin-treated obese subjects with prediabetes or newly-diagnosed T2DM were randomized to the glucagon-like peptide receptor agonist (GLP-RA) liraglutide (1.8 mg/day) or lifestyle counseling until achieving a comparable weight loss (−7% of initial body weight), to assess whether changes in subcutaneous (SAT) and visceral (VAT) adipose tissue distribution (MRI), insulin sensitivity (Matsuda Index) and beta-cell performance (multiple sampling OGTT beta-index), with either intervention, might affect TX-dependent platelet activation, lipid peroxidation and inflammation. At baseline, Ln-8-iso-PGF_2α (Beta = 0.31, p = 0.0088), glycosylated hemoglobin (HbA1c) (Beta = 2.64, p = 0.0011) Ln-TNF-α (Beta = 0.58, p = 0.0075) and SAT (Beta = 0.14, p = 0.044) were significant independent predictors of 11-dehydro-TXB₂. After achievement of the weight loss target, a comparable reduction in U-11-dehydro-TXB₂ (between-group p = 0.679) and 8-iso-PGF-_2α (p = 0.985) was observed in both arms in parallel with a comparable improvement in glycemic control, insulin sensitivity, SAT, high-sensitivity C-reactive protein (hs-CRP). In obese patients with initial impairment of glucose metabolism, the extent of platelet activation is related to systemic inflammation, isoprostane formation and degree of glycemic control and abdominal SAT. Successful weight loss, achieved with either lifestyle changes or an incretin-based therapy, is associated with a significant reduction in lipid peroxidation and platelet activation.

Resistin is an adipokine that promotes inflammation and insulin resistance by targeting several cells including platelets. We hypothesised that in type 2 diabetes (T2DM), resistin may foster in vivo oxidative stress, thromboxane-dependent platelet activation and platelet-derived inflammatory proteins release, key determinants of atherothrombosis. A cross-sectional comparison of circulating resistin, sCD40L, as a marker of platelet-mediated inflammation, asymmetric dimethylarginine (ADMA), endothelial dysfunction marker, Dickkopf (DKK)-1, reflecting the inflammatory interaction between platelets and endothelial cells, and urinary 8-iso-PGF_2α and 11-dehydro-TxB₂, reflecting in vivo lipid peroxidation and platelet activation, respectively, was performed between 79 T2DM patients and 30 healthy subjects. Furthermore, we investigated the effects of the α-glucosidase inhibitor acarbose and the PPARγ agonist rosiglitazone, targeting hyperglycaemia or insulin resistance, versus placebo, in 28 and 18 T2DM subjects, respectively. Age- and gender-adjusted serum resistin levels were significantly higher in patients than in controls. HOMA (β=0.266, p=0.017) and 11-dehydro-TXB₂ (β=0.354, p=0.002) independently predicted resistin levels. A 20-week treatment with acarbose was associated with significant reductions (p=0.001) in serum resistin, DKK-1, urinary 11-dehydro-TXB₂ and 8-iso-PGF_2α with direct correlations between the change in serum resistin and in other variables. A 24-week rosiglitazone treatment on top of metformin was associated with significant decreases in resistin, DKK-1, 11-dehydro-TXB₂ and 8-iso-PGF_2α, in parallel with HOMA decrease. In conclusion, resistin, antagonising insulin action in part through PPARγ activation, may favour insulin resistance and enhance oxidative stress, endothelial dysfunction and platelet activation. The adipokine-platelet interactions may be involved in platelet insulin resistance and their consequent pro-aggregatory phenotype in this setting.