Atorvastatin lowers 68Ga-DOTATATE uptake in coronary arteries, bone marrow and spleen in individuals with type 2 diabetes.

AIMS/HYPOTHESIS: Inflammation is a core component of residual cardiovascular risk in type 2 diabetes. With new anti-inflammatory therapeutics entering the field, accurate markers to evaluate their effectiveness in reducing cardiovascular disease are paramount. Gallium-68-labelled DOTATATE (68Ga-DOTATATE) has recently been proposed as a more specific marker of arterial wall inflammation than 18F-fluorodeoxyglucose (18F-FDG). This study set out to investigate whether 68Ga-DOTATATE uptake is amenable to therapeutic intervention in individuals with type 2 diabetes. METHODS: Individuals aged >50 years with type 2 diabetes underwent 68Ga-DOTATATE positron emission tomography (PET)/computed tomography (CT) at baseline and after 3 months treatment with atorvastatin 40 mg once daily. Primary outcome was the difference in coronary 68Ga-DOTATATE uptake, expressed as target-to-background ratio (TBR). The secondary outcome was difference in bone marrow and splenic uptake, expressed as the standardised uptake value (SUV). RESULTS: Twenty-two individuals with type 2 diabetes (mean age 63.2±6.4 years, 82% male, LDL-cholesterol 3.42±0.81 mmol/l, HbA1c 55±12 mmol/mol [7.2%±3.2%]) completed both 68Ga-DOTATATE PET/CT scans. The maximum TBR was -31% (95% CI -50, -12) lower in the coronary arteries, and bone marrow and splenic 68Ga-DOTATATE uptake was also significantly lower post statin treatment, with a mean percentage reduction of -15% (95% CI -27, -4) and -17% (95% CI -32, -2), respectively. CONCLUSIONS/INTERPRETATION: 68Ga-DOTATATE uptake across the cardio-haematopoietic axis was lower after statin therapy in individuals with type 2 diabetes. Therefore, 68Ga-DOTATATE is promising as a metric for vascular and haematopoietic inflammation in intervention studies using anti-inflammatory therapeutics in individuals with type 2 diabetes. TRIAL REGISTRATION: ClinicalTrials.gov NCT05730634.

Oxidative Stress in Neutrophils: Implications for Diabetic Cardiovascular Complications.

Significance: Neutrophil behavior and function are altered by hyperglycemia associated with diabetes. Aberrant activation by hyperglycemia causes neutrophils to respond with increased production of reactive oxidative species (ROS). Excess ROS, a signature of primed neutrophils, can intracellularly induce neutrophils to undergo NETosis, flooding surrounding tissues with ROS and damage-associated molecular patterns such as S100 calcium binding proteins (S100A8/A9). The cargo associated with NETosis also attracts more immune cells to the site and signals for increased immune cell production. This inflammatory response to diabetes can accelerate other associated conditions such as atherosclerosis and thrombosis, increasing the risk of cardiovascular disease. Recent Advances: As the prevalence of diabetes continues to grow, more attention has been focused on developing effective treatment options. Currently, glucose-lowering medications and insulin injections are the most widely utilized treatments. As the disease progresses, medications are usually stacked to maintain glucose at desired target levels, but this approach often fails and does not effectively reduce cardiovascular risk, even with the latest drugs. Critical Issues: Despite advances in treatment options, diabetes remains a progressive disease as glucose lowering alone has failed to abolish the associated cardiovascular complications. Future Directions: Significant interest is being generated in developing treatments that do not solely focus on glucose control but rather mitigate glucotoxicity. Several therapies have been proposed that target cellular dysfunction downstream of hyperglycemia, such as using antioxidants to scavenge ROS, inhibiting ROS production from NOX, and suppressing neutrophil release of S100A8/A9 proteins. Antioxid. Redox Signal. 36, 652-666.

Fecal microbiota transplantation in human metabolic diseases: From a murky past to a bright future?

Fecal microbiota transplantation (FMT) is gaining considerable traction as a therapeutic approach to influence the course of a plethora of chronic conditions, ranging from metabolic syndrome and malignancies to auto-immune and neurological diseases, and helped to establish the contribution of the gut microbiome to these conditions. Although FMT procedures have yielded important mechanistic insights, their use in clinical practice may be limited due to practical objections in the setting of metabolic diseases. While its applicability is established to treat recurrent Clostridiodes difficile, FMT is emerging in ulcerative colitis and various other diseases. A particularly new insight is that FMTs may not only alter insulin sensitivity but may also alter the course of type 1 diabetes by attenuating underlying auto-immunity. In this review, we will outline the major principles and pitfalls of FMT and where optimization of study design and the procedure itself will further advance the field of cardiometabolic medicine.

Apoptotic Ablation of Platelets Reduces Atherosclerosis in Mice With Diabetes.

OBJECTIVE: People with diabetes are at a significantly higher risk of cardiovascular disease, in part, due to accelerated atherosclerosis. Diabetic subjects have increased number of platelets that are activated, more reactive, and respond suboptimally to antiplatelet therapies. We hypothesized that reducing platelet numbers by inducing their premature apoptotic death would decrease atherosclerosis. Approach and Results: This was achieved by targeting the antiapoptotic protein Bcl-xL (B-cell lymphoma-extra large; which is essential for platelet viability) via distinct genetic and pharmacological approaches. In the former, we transplanted bone marrow from mice carrying the Tyr15 to Cys loss of function allele of Bcl-x (known as Bcl-xPlt20) or wild-type littermate controls into atherosclerotic-prone Ldlr+/- mice made diabetic with streptozotocin and fed a Western diet. Reduced Bcl-xL function in hematopoietic cells significantly decreased platelet numbers, exclusive of other hematologic changes. This led to a significant reduction in atherosclerotic lesion formation in Bcl-xPlt20 bone marrow transplanted Ldlr+/- mice. To assess the potential therapeutic relevance of reducing platelets in atherosclerosis, we next targeted Bcl-xL with a pharmacological strategy. This was achieved by low-dose administration of the BH3 (B-cell lymphoma-2 homology domain 3) mimetic, ABT-737 triweekly, in diabetic Apoe-/- mice for the final 6 weeks of a 12-week study. ABT-737 normalized platelet numbers along with platelet and leukocyte activation to that of nondiabetic controls, significantly reducing atherosclerosis while promoting a more stable plaque phenotype. CONCLUSIONS: These studies suggest that selectively reducing circulating platelets, by targeting Bcl-xL to promote platelet apoptosis, can reduce atherosclerosis and lower cardiovascular disease risk in diabetes. Graphic Abstract: A graphic abstract is available for this article.

Postprandial Glucose Spikes, an Important Contributor to Cardiovascular Disease in Diabetes?

Clinical trials investigating whether glucose lowering treatment reduces the risk of CVD in diabetes have thus far yielded mixed results. However, this doesn't rule out the possibility of hyperglycemia playing a major causal role in promoting CVD or elevating CVD risk. In fact, lowering glucose appears to promote some beneficial long-term effects, and continuous glucose monitoring devices have revealed that postprandial spikes of hyperglycemia occur frequently, and may be an important determinant of CVD risk. It is proposed that these short, intermittent bursts of hyperglycemia may have detrimental effects on several organ systems including the vasculature and the hematopoietic system collectively contributing to the state of elevated CVD risk in diabetes. In this review, we summarize the potential mechanisms through which hyperglycemic spikes may increase atherosclerosis and how new and emerging interventions may combat this.

Transient Intermittent Hyperglycemia Accelerates Atherosclerosis by Promoting Myelopoiesis.

RATIONALE: Treatment efficacy for diabetes mellitus is largely determined by assessment of HbA1c (glycated hemoglobin A1c) levels, which poorly reflects direct glucose variation. People with prediabetes and diabetes mellitus spend >50% of their time outside the optimal glucose range. These glucose variations, termed transient intermittent hyperglycemia (TIH), appear to be an independent risk factor for cardiovascular disease, but the pathological basis for this association is unclear. OBJECTIVE: To determine whether TIH per se promotes myelopoiesis to produce more monocytes and consequently adversely affects atherosclerosis. METHODS AND RESULTS: To create a mouse model of TIH, we administered 4 bolus doses of glucose at 2-hour intervals intraperitoneally once to WT (wild type) or once weekly to atherosclerotic prone mice. TIH accelerated atherogenesis without an increase in plasma cholesterol, seen in traditional models of diabetes mellitus. TIH promoted myelopoiesis in the bone marrow, resulting in increased circulating monocytes, particularly the inflammatory Ly6-Chi subset, and neutrophils. Hematopoietic-restricted deletion of S100a9, S100a8, or its cognate receptor Rage prevented monocytosis. Mechanistically, glucose uptake via GLUT (glucose transporter)-1 and enhanced glycolysis in neutrophils promoted the production of S100A8/A9. Myeloid-restricted deletion of Slc2a1 (GLUT-1) or pharmacological inhibition of S100A8/A9 reduced TIH-induced myelopoiesis and atherosclerosis. CONCLUSIONS: Together, these data provide a mechanism as to how TIH, prevalent in people with impaired glucose metabolism, contributes to cardiovascular disease. These findings provide a rationale for continual glucose control in these patients and may also suggest that strategies aimed at targeting the S100A8/A9-RAGE (receptor for advanced glycation end products) axis could represent a viable approach to protect the vulnerable blood vessels in diabetes mellitus. Graphic Abstract: A graphic abstract is available for this article.

Higher Plasma Methylglyoxal Levels Are Associated With Incident Cardiovascular Disease and Mortality in Individuals With Type 2 Diabetes.

OBJECTIVE: Methylglyoxal (MGO) is a reactive dicarbonyl compound and a potential key player in diabetic cardiovascular disease (CVD). Whether plasma MGO levels are associated with CVD in type 2 diabetes is unknown. RESEARCH DESIGN AND METHODS: We included 1,003 individuals (mean ± SD age 59.1 ± 10.5 years, 69.3% male, and 61.6% with prior CVD) with type 2 diabetes from the Second Manifestations of ARTerial disease cohort (SMART). We measured plasma MGO levels and two other dicarbonyls (glyoxal [GO] and 3-deoxyglucosone [3-DG]) at baseline with mass spectrometry. Median follow-up of CVD events was 8.6 years. Data were analyzed with Cox regression with adjustment for sex, age, smoking, systolic blood pressure, total cholesterol, HbA1c, BMI, prior CVD, and medication use. Hazard ratios are expressed per SD Ln-transformed dicarbonyl. RESULTS: A total of 287 individuals suffered from at least one CVD event (n = 194 fatal events, n = 146 myocardial infarctions, and n = 72 strokes); 346 individuals died, and 60 individuals underwent an amputation. Higher MGO levels were associated with total (hazard ratio 1.26 [95% CI 1.11-1.42]) and fatal (1.49 [1.30-1.71]) CVD and with all-cause mortality (1.25 [1.11-1.40]), myocardial infarction (1.22 [1.02-1.45]), and amputations (1.36 [1.05-1.76]). MGO levels were not apparently associated with stroke (1.03 [0.79-1.35]). Higher GO levels were significantly associated with fatal CVD (1.17 [1.00-1.37]) but not with other outcomes. 3-DG was not significantly associated with any of the outcomes. CONCLUSIONS: Plasma MGO and GO levels are associated with cardiovascular mortality in individuals with type 2 diabetes. Influencing dicaronyl levels may therefore be a target to reduce CVD in type 2 diabetes.

Dietary intake of advanced glycation endproducts is associated with higher levels of advanced glycation endproducts in plasma and urine: The CODAM study.

BACKGROUND & AIMS: Advanced glycation endproducts (AGEs) are formed by the reaction between reducing sugars and proteins. AGEs in the body have been associated with several age-related diseases. High-heat treated and most processed foods are rich in AGEs. The aim of our study was to investigate whether dietary AGEs, are associated with plasma and urinary AGE levels. METHODS: In 450 participants of the Cohort on Diabetes and Atherosclerosis Maastricht study (CODAM study) we measured plasma and urine concentrations of the AGEs Nε-(carboxymethyl)lysine (CML), Nε-(1-carboxyethyl)lysine (CEL) and Nδ-(5-hydro-5-methyl-4-imidazolon-2-yl)-ornithine (MG-H1) using UPLC-MS/MS. We also estimated dietary intake of CML, CEL and MG-H1 with the use of a dietary AGE database and a food frequency questionnaire (FFQ). We used linear regression to investigate the association between standardized dietary AGE intake and standardized plasma or urinary AGE levels, after adjustment for age, sex, glucose metabolism status, waist circumference, kidney function, energy- and macro-nutrient intake, smoking status, physical activity, alcohol intake, LDL-cholesterol and markers of oxidative stress. RESULTS: We found that higher intake of dietary CML, CEL and MG-H1 was associated with significantly higher levels of free plasma and urinary CML, CEL and MG-H1 (ßCML = 0.253 (95% CI 0.086; 0.415), ßCEL = 0.194 (95% CI 0.040; 0.339), ßMG-H1 = 0.223 (95% CI 0.069; 0.373) for plasma and ßCML = 0.223 (95% CI 0.049; 0.393), ßCEL = 0.180 (95% CI 0.019; 0.332), ßMG-H1 = 0.196 (95% CI 0.037; 0.349) for urine, respectively). In addition, we observed non-significant associations of dietary AGEs with their corresponding protein bound plasma AGEs. CONCLUSION: We demonstrate that higher intake of dietary AGEs is associated with higher levels of AGEs in plasma and urine. Our findings may have important implications for those who ingest a diet rich in AGEs.

Higher Plasma Methylglyoxal Levels Are Associated With Incident Cardiovascular Disease in Individuals With Type 1 Diabetes: A 12-Year Follow-up Study.

Methylglyoxal (MGO), a major precursor for advanced glycation end products, is increased in diabetes. In diabetic rodents, inhibition of MGO prevents cardiovascular disease (CVD). Whether plasma MGO levels are associated with incident CVD in people with type 1 diabetes is unknown. We included 159 individuals with persistent normoalbuminuria and 162 individuals with diabetic nephropathy (DN) from the outpatient clinic at Steno Diabetes Center. We measured MGO at baseline and recorded fatal and nonfatal CVD over a median follow-up of 12.3 years (interquartile range 7.6-12.5 years). Data were analyzed by Cox regression, with adjustment for sex, age, HbA1c, DN, diabetes duration, smoking, systolic blood pressure, antihypertensive medication, and BMI. During follow-up, 73 individuals suffered at least one CVD event (36 fatal and 53 nonfatal). Higher MGO levels were associated with total, fatal, and nonfatal incident CVD (hazard ratios [HRs] 1.47 [95% CI 1.13-1.91], 1.42 [1.01-1.99], and 1.46 [1.08-1.98], respectively). We observed a similar trend for total mortality (HR 1.24 [0.99-1.56]). This study shows for the first time in our knowledge that plasma MGO levels are associated with cardiovascular events in individuals with type 1 diabetes. MGO may explain, at least in part, the increased risk for CVD in type 1 diabetes.

Plasma advanced glycation end products are associated with incident cardiovascular events in individuals with type 2 diabetes: a case-cohort study with a median follow-up of 10 years (EPIC-NL).

Experimental data suggest a role for advanced glycation end products (AGEs) in cardiovascular disease (CVD), particularly in type 2 diabetes (T2DM). However, epidemiological evidence of an association between high plasma AGEs and increased cardiovascular risk remains inconclusive. Therefore, in a case-cohort study comprising 134 cardiovascular case subjects and a random subcohort of 218 individuals (including 65 cardiovascular case subjects), all with T2DM and nested in the European Prospective Investigation into Cancer and Nutrition in the Netherlands (EPIC-NL) study, plasma levels of protein-bound Nε-(carboxymethyl)lysine, Nε-(carboxyethyl)lysine, and pentosidine were measured with liquid chromatography. AGEs were loge-transformed, combined in a z-score, and the association with incident cardiovascular events was analyzed with Cox proportional hazard regression, adapted for case-cohort design (Prentice method). After multivariable adjustment (sex, age, cohort status, diabetes duration, total cholesterol to HDL-cholesterol ratio, smoking, systolic blood pressure, BMI, blood pressure-, cholesterol- and glucose-lowering treatment, prior cardiovascular events, and triglycerides), higher plasma AGE z-scores were associated with higher risk of incident cardiovascular events in individuals without prior cardiovascular events (hazard ratio 1.31 [95% CI: 1.06-1.61]). A similar trend was observed in individuals with prior cardiovascular events (1.37 [0.63-2.98]). In conclusion, high plasma AGEs were associated with incident cardiovascular events in individuals with T2DM. These results underline the potential importance of AGEs in development of CVD.

Glyoxalase 1 overexpression does not affect atherosclerotic lesion size and severity in ApoE-/- mice with or without diabetes.

AIMS: Advanced glycation end-products (AGEs) and their precursors have been associated with the development of atherosclerosis. We recently discovered that glyoxalase 1 (GLO1), the major detoxifying enzyme for AGE precursors, is decreased in ruptured human plaques, and that levels of AGEs are higher in rupture-prone plaques. We here investigated whether overexpression of human GLO1 in ApoE(-/-) mice could reduce the development of atherosclerosis. METHODS AND RESULTS: We crossed C57BL/6 ApoE(-/-) mice with C57BL/6 GLO1 overexpressing mice (huGLO1(+/-)) to generate ApoE(-/-) (n = 16) and ApoE(-/-) huGLO1(+/-) (n = 20) mice. To induce diabetes, we injected a subset with streptozotocin (STZ) to generate diabetic ApoE(-/-) (n = 8) and ApoE(-/-) huGLO1(+/-) (n = 13) mice. All mice were fed chow and sacrificed at 25 weeks of age. The GLO1 activity was three-fold increased in huGLO1(+/-) aorta, but aortic root lesion size and phenotype did not differ between mice with and without huGLO1(+/-) overexpression. We detected no differences in gene expression in aortic arches, in AGE levels and cytokines, in circulating cells, and endothelial function between ApoE(-/-) mice with and without huGLO1(+/-) overexpression. Although diabetic mice showed decreased GLO1 expression (P < 0.05) and increased lesion size (P < 0.05) in comparison with non-diabetic mice, GLO1 overexpression also did not affect the aortic root lesion size or inflammation in diabetic mice. CONCLUSION: In ApoE(-/-) mice with or without diabetes, GLO1 overexpression did not lead to decreased atherosclerotic lesion size or systemic inflammation. Increasing GLO1 levels does not seem to be an effective strategy to reduce glycation in atherosclerotic lesions, likely due to increased AGE formation through GLO1-independent mechanisms.

Higher levels of advanced glycation endproducts in human carotid atherosclerotic plaques are associated with a rupture-prone phenotype.

AIMS: Rupture-prone atherosclerotic plaques are characterized by inflammation and a large necrotic core. Inflammation is linked to high metabolic activity. Advanced glycation endproducts (AGEs) and their major precursor methylglyoxal are formed during high metabolic activity and can have detrimental effects on cellular function and may induce cell death. Therefore, we investigated whether plaque AGEs are increased in human carotid rupture-prone plaques and are associated with plaque inflammation and necrotic core formation. METHODS AND RESULTS: The protein-bound major methylglyoxal-derived AGE 5-hydro-5-methylimidazolone (MG-H1) and N(ε)-(carboxymethyl)lysine (CML) were measured in human carotid endarterectomy specimens (n = 75) with tandem mass spectrometry. MG-H1 and CML levels were associated with rupture-prone plaques, increased protein levels of the inflammatory mediators IL-8 and MCP-1 and with higher MMP-9 activity. Immunohistochemistry showed that AGEs accumulated predominantly in macrophages surrounding the necrotic core and co-localized with cleaved caspase-3. Intra-plaque comparison revealed that glyoxalase-1 (GLO-1), the major methylglyoxal-detoxifying enzyme, mRNA was decreased (-13%, P < 0.05) in ruptured compared with stable plaque segments. In line, in U937 monoctyes, we found reduced (GLO-1) activity (-38%, P < 0.05) and increased MGO (346%, P < 0.05) production after stimulation with the inflammatory mediator TNF. Direct incubation with methylglyoxal increased apoptosis up to two-fold. CONCLUSION: This is the first study showing that AGEs are associated with human rupture-prone plaques. Furthermore, this study suggests a cascade linking inflammation, reduced GLO-1, methylglyoxal- and AGE-accumulation, and subsequent apoptosis. Thereby, AGEs may act as mediators of the progression of stable to rupture-prone plaques, opening a window towards novel treatments and biomarkers to treat cardiovascular diseases.