Proteomics of high-density lipoprotein subfractions and subclinical atherosclerosis in type 1 diabetes mellitus: a case-control study.

BACKGROUND: Subclinical atherosclerosis is frequently observed in type 1 diabetes (T1D) although the mechanisms and markers involved in the evolution to established cardiovascular disease are not well known. High-density lipoprotein cholesterol in T1D is normal or even high, and changes in its functionality and proteomics are considered. Our aim was to evaluate the proteomics of HDL subfractions in T1D and control subjects and its association with clinical variables, subclinical atherosclerosis markers and HDL functionality. METHODS: A total of 50 individuals with T1D and 30 matched controls were included. Carotid-femoral pulse wave velocity (PWV), flow-mediated vasodilation (FMD), cardiovascular autonomic neuropathy (CAN), and ten-year cardiovascular risk (ASCVDR) were determined. Proteomics (parallel reaction monitoring) was determined in isolated HDL2 and HDL3 that were also utilized to measure cholesterol efflux from macrophages. RESULTS: Among 45 quantified proteins, 13 in HDL2 and 33 in HDL3 were differentially expressed in T1D and control subjects. Six proteins related to lipid metabolism, one to inflammatory acute phase, one to complement system and one to antioxidant response were more abundant in HDL2, while 14 lipid metabolism, three acute-phase, three antioxidants and one transport in HDL3 of T1D subjects. Three proteins (lipid metabolism, transport, and unknown function) were more abundant in HDL2; and ten (lipid metabolism, transport, protease inhibition), more abundant in HDL3 of controls. Individuals with T1D had higher PWV and ten-year ASCVDR, and lower FMD, Cholesterol efflux from macrophages was similar between T1D and controls. Proteins in HDL2 and HDL3, especially related to lipid metabolism, correlated with PWV, CAN, cholesterol efflux, HDLc, hypertension, glycemic control, ten-year ASCVDR, and statins use. CONCLUSION: HDL proteomics can be predictive of subclinical atherosclerosis in type 1 diabetes. Proteins that are not involved in reverse cholesterol transport may be associated with the protective role of HDL.

Evaluation of galectin-3 levels in patients with type 2 diabetes mellitus and chronic kidney disease / Avaliação dos níveis de galectina-3 em pacientes com diabetes mellitus tipo 2 e doença renal crônica

ABSTRACT INTRODUCTION: Type 2 diabetes mellitus (T2DM) is the most common manifestation of diabetes, accounting for about 90% of diagnosed cases. The causes of T2DM are not fully understood, but its pathogenesis is possibly associated with increased adiposity and a chronic low-grade inflammatory response. The glycoprotein galectin-3 (Gal-3) is known to play an important role in the modulation of blood glucose, adiposity, and inflammation. OBJECTIVES: The aim of this study was to evaluate Gal-3 levels in patients with T2DM and chronic kidney disease (CKD), in addition to relating them with complications and comorbidities present in these patients, comparing them to a control group. MATERIAL AND METHODS: Gal-3 was evaluated in 84 selected individuals, of which 42 had clinical and laboratory diagnosis of T2DM and CKD (treated at Santa Casa Hospital in Belo Horizonte, Minas Gerais, Brazil), and 42 individuals from the local community, with no history of diabetes (control group). RESULTS AND DISCURSION: Gal-3 levels were significantly higher (p = 0.012) in the T2DM group (15.17 ± 5.54 ng/ml) when compared to the control group (12.62 ± 3.2 ng/ml). There was a tendency for higher levels of Gal-3 in diabetic patients with hypertension (15.74 ± 5.61 ng/ml) when compared to patients without this complication (10.96 ± 2.49 ng/ml) (p = 0.069) CONCLUSION: The results suggest that Gal-3 may be involved in the pathophysiology of T2DM and still be a promising biomarker associated with hypertension in this group.

RESUMEN INTRODUCCIÓN: La diabetes mellitus tipo 2 (DM2) es la forma más común de la diabetes; representa alrededor del 90% de los casos diagnosticados. Todavía no se conocen por completo las causas de la DM2, pero posiblemente su etiopatogénesis se relaciona con el aumento de adiposidad y una respuesta inflamatoria crónica de bajo grado. Se sabe que la glicoproteína galectina 3 (Gal-3) juega un papel importante en la modulación de glucemia, adiposidad e inflamación. OBJETIVOS: Evaluar los niveles de Gal-3 en pacientes con DM2 y enfermedad renal crónica, además de relacionarlos con las otras complicaciones y comorbilidades presentes en eses individuos, comparándolos con un grupo control. MATERIAL Y MÉTODO: La Gal-3 fue evaluada en 84 pacientes elegidos; entre esos, 42 poseían el diagnóstico clínico y de laboratorio de DM2 y enfermedad renal crónica (atendidos en el Hospital Santa Casa de Belo Horizonte, Minas Gerais, Brasil) y 42 eran de la comunidad local, sin historial de diabetes (grupo control). RESULTADOS Y DISCUSIÓN: Los niveles de Gal-3 fueron más altos (p = 0,012) en el grupo con DM2 (15,17 ± 5,54 ng/ml) que en el grupo control (12,62 ± 3,2 ng/ml). Hubo tendencia de mayores niveles de Gal-3 en los pacientes diabéticos con hipertensión (15,74 ± 5,61 ng/ml) que en aquellos sin esa complicación (10,96 ± 2,49 ng/ml) (p = 0,069). CONCLUSIÓN: Los resultados obtenidos apuntan que la Gal-3 puede estar involucrada en la etiología de la DM2 y aún ser un biomarcador prometedor de hipertensión en ese grupo.

RESUMO INTRODUÇÃO: O diabetes mellitus tipo 2 (DM2) é a manifestação mais comum do diabetes; representa cerca de 90% dos casos diagnosticados. As causas do DM2 ainda não foram completamente estabelecidas, mas sua patogênese está, possivelmente, relacionada com o aumento da adiposidade e uma resposta inflamatória crônica de baixo grau. Sabe-se que a glicoproteína galectina-3 (Gal-3) possui papel importante na modulação de glicemia, adiposidade e inflamação. OBJETIVOS: Avaliar os níveis de Gal-3 em pacientes com DM2 e doença renal crônica, além de relacioná-los com as demais complicações e comorbidades presentes nesses indivíduos, comparando-os com um grupo-controle. MATERIAL E MÉTODOS:: A Gal-3 foi avaliada em 84 pacientes selecionados; destes, 42 possuíam o diagnóstico clínico e laboratorial de DM2 e doença renal crônica (atendidos no Hospital Santa Casa de Belo Horizonte, Minas Gerais, Brasil), e 42 eram da comunidade local, sem histórico de diabetes (grupo-controle). RESULTADOS E DISCUSSÃO: Os níveis de Gal-3 foram significativamente mais elevados (p = 0,012) no grupo com DM2 (15,17 ± 5,54 ng/ml) quando comparados com os níveis do grupo-controle (12,62 ± 3,2 ng/ml). Houve tendência em maiores níveis de Gal-3 nos pacientes diabéticos com hipertensão (15,74 ± 5,61 ng/ml) em comparação com os pacientes sem essa complicação (10,96 ± 2,49 ng/ml) (p = 0,069). CONCLUSÃO: Os resultados obtidos sugerem que a Gal-3 pode estar envolvida na fisiopatologia do DM2 e ainda ser um promissor biomarcador associado à hipertensão nesse grupo.

Proresolving protein Annexin A1: The role in type 2 diabetes mellitus and obesity.

BACKGROUND: Annexin A1 (AnxA1) is a protein involved in inflammation resolution that might be altered in obesity-associated type 2 diabetes mellitus (DM), which is a chronic inflammatory disease. The aim of this study was to evaluate AnxA1 serum levels in individuals with and without DM stratified according to the body mass index (BMI), and the dynamic of AnxA1 expression in adipose tissue from humans with obesity and non-obesity. METHODS: Serum samples were obtained from 41 patients with DM (lean, overweight and obese) and 40 controls, and adipose tissue samples were obtained from 16 individuals with obesity (with or without DM), and 15 controls. RESULTS: DM patients showed similar AnxA1 serum levels when compared to controls. However, when the individuals were stratified according to BMI, AnxA1 levels were higher in individuals with obesity than lean or overweight, and in overweight compared to lean individuals. Moreover, AnxA1 was correlated positively with IL-6 levels. AnxA1 levels were also positively correlated with BMI, waist circumference and waist-to-hip ratio. Furthermore, higher levels of cleaved AnxA1 were observed in adipose tissue from individuals with obesity, independently of DM status. CONCLUSIONS: Enhanced levels of AnxA1 in serum of individuals with obesity suggest an attempt to counter-regulate the systemic inflammation process in this disease. However, the higher levels of cleaved AnxA1 in the adipose tissue of individuals with obesity could compromise its anti-inflammatory and proresolving actions, locally. Considering our data, AnxA1 cleavage in the adipose tissue, despite increased serum levels of this protein, and consequently the failure in inflammation resolution, suggests an important pathophysiological mechanism involved in inflammatory status observed in obesity.

Effect of acetylsalicylic acid on platelet activation and oxidative profile in a set of Brazilian patients with type 2 diabetes mellitus.

Type 2 diabetes mellitus (DM2) is a metabolic disorder associated with hyperactivation of platelets, increased formation of platelet microparticles (PMPs) and oxidative stress that are related to cardiovascular complications. Acetylsalicylic acid (ASA) is an antiplatelet agent used in the prevention of atherothrombosis. The aim of this study was to evaluate the effect of ASA by means of platelet activation and oxidative profile. We collected blood samples of 81 patients with DM2 before and during ASA treatment. These samples were analyzed to determine the levels of 2,3-dinor thromboxane-B2 (2,3-dinor-TXB2), PMPs, thiobarbituric acid reactive species (TBARS) and 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT). Moreover, the relationship between the levels of 2,3-dinor-TXB2 with some clinical and laboratory variables such as glycated hemoglobin, platelet count, D dimer, low-density lipoprotein cholesterol and glycoprotein IIb/IIIa and cyclooxygenase-1 polymorphisms was evaluated. ASA intake did not change the levels of PMP, TBARS and MTT. Although a significant decrease in the levels of 2,3 dinorTXB2 (P < 0.001) in patients under ASA has been observed, an equal and satisfactory response to this drug was not found. However, the presence of PIA2 allele in GPIIIa gene may be associated with a better response to ASA intake in these patients, whereas other clinical and laboratory variables showed no association with this drug use. These findings are consistent with previous reports in the literature that patients with DM2 do not benefit in an equal way from the use of ASA for primary prevention of atherothrombotic events.

Acetylsalicylic acid therapy: influence of metformin use and other variables on urinary 11-dehydrothromboxane B2 levels.

BACKGROUND: The effect of acetylsalicylic acid (ASA) may be measured through the analysis of urinary concentrations of 11-dehydrothromboxane B2 (11-dhTXB2), a metabolite of thromboxane A2, which is a potent platelet aggregant agent. It has been suggested that metformin (an oral antidiabetic drug) could improve oxidative stress and control platelet activation in type 2 diabetic patients, potentially reducing cardiovascular risk. We determined the concentrations of urinary 11-dhTXB2 in type 2 diabetic patients taking ASA and its concentrations with metformin use and several other clinical variables (hypertension, age, gender, smoking, body mass index, insulin and statin use), considering a reduction of at least 75% in the concentrations of this marker as a target, compared to results before ASA intake. METHODS: Urinary concentrations of 11-dhTXB2 of 81 type 2 diabetic patients were measured before and at 15 days taking 100 mg of aspirin daily. RESULTS: Most patients who presented a reduction of 11-dhTXB2 above 75% were under metformin use. This reduction was achieved in 51.5% of patients taking this drug, against 20.0% in the patients who were not (p=0.027). The analysis of the other variables did not show a significant difference. The use of metformin appears to play a role in the reduction of 11-dhTXB2 concentrations in type 2 diabetic patients. CONCLUSION: According to previous reports, hyperglycemia control seems to be a determinant factor for the success of ASA therapy, given the influence of metformin in the reduction of 11-dhTXB2 concentrations.

Suppressive effect of aqueous humor from person with type 2 diabetes with or without retinopathy on reactive oxygen species generation.

AIM: To evaluate the antioxidant capacity and concentrations of vascular endothelial growth factor (VEGF) and interleukin 6 (IL-6) in aqueous humor from patients with type 2 diabetes mellitus (T2DM) with or without retinopathy. METHODS: Aqueous humor was obtained during elective cataract surgery from T2DM patients with or without retinopathy and from healthy subjects. Reducing response was evaluated by MTT dye reduction and the generation of reactive oxygen species (ROS) was determined by chemiluminescence assay. Granulocytes were treated with phorbol dibutyrate (PDB)-stimulated. Cytokines were quantified by ELISA. RESULTS: Antioxidant capacity of aqueous humor from patients with retinopathy was greater (P<0.05) than that of healthy controls or persons with diabetes without retinopathy. ROS production in PDB (protein kinase C activator)-stimulated granulocytes from T2DM patients with or without retinopathy was inhibited by autologous aqueous humor. Concentrations of VEGF and IL-6 were similar in aqueous humor from healthy controls and from patients without retinopathy, but lower (P<0.05) than those from T2DM patients with retinopathy. Plasma levels of VEGF and IL-6 were similar (P>0.05) in healthy controls and in T2DM patients with and without retinopathy. CONCLUSION: Aqueous humor from T2DM patients with retinopathy exhibits elevated antioxidant activity with significant suppressive effect on ROS production and enhanced levels of locally secreted VEGF and IL-6 in comparison with T2DM patients without retinopathy. These results suggest an inflammatory profile in the absence of typical oxidative stress for T2DM patients with retinopathy, possibly resulting from the compensatory antioxidant response detected in the aqueous humor improving the ocular redox state.

-1131T>C and SW19 polymorphisms in APOA5 gene and lipid levels in type 2 diabetic patients.

Type 2 diabetes mellitus is a metabolic, vascular, and neuropathic disease with a high risk of atherosclerotic events due to dyslipidemic states. Polymorphisms in Apolipoprotein A5 gene (APOA5) have been associated with increased triglyceride levels in many different populations. This study aimed to identify the frequencies of the APOA5 -1131T>C and SW19 polymorphisms and evaluate their effects on lipid levels in patients with type 2 diabetes. Genotyping of APOA5 -1131T>C and SW19 polymorphisms was performed by PCR-RFLP in 146 diabetic patients and in controls (n = 173), from 30 to 80 years of age. Diabetic patients were divided into two groups: patients not treated with lipid lowering drugs (group G1; n = 62) and those treated with lipid lowering drugs (group G2, n = 84). Lipids and lipoproteins were determined enzymatically. Among participants not treated with lipid-lowering drugs (diabetics G1 and controls; n = 235), the -1131C was associated with lower LDLc levels (p = 0.015). In the diabetic patients, the 19W allele was associated with higher triglyceride levels (p = 0.004). In G1 diabetic patients, the combined analysis of APOA5 -1131T>C and SW19 polymorphisms showed that [TC or CC] + SS carriers presented lower total cholesterol levels than did other genotype combinations (p = 0.049). It could therefore be concluded that APOA5 -1131T>C and SW19 polymorphisms influence lipid levels in type 2 diabetic patients.

Oxidizing and reducing responses in type 1 diabetic patients determined up to 5 years after the clinical onset of the disease.

Oxidative stress has been suggested to be the mediator of hyperglycaemia-induced diabetic complications. For this study we asked whether a significant imbalance between oxidizing and plasmatic reducing responses could be observed in DM1 patients receiving intensive therapy up to 5 years following the clinical onset of the disease. A total of 16 type 1 diabetic patients (DM1) without complications and 13 non-diabetic subjects were enrolled. Clinical and biochemical parameters were compared in the two populations studied. Moreover, reactive oxygen species (ROS) generation by granulocytes and the total plasma antioxidant status were simultaneously evaluated. Granulocytes-ROS derived and plasma antioxidant status were determined by chemiluminescence assay and tetrazolium dye reduction, respectively. Type 1 diabetic patients were receiving intensive therapy by multiple daily injections. In comparison with healthy individuals, DM1 patients exhibited an increase in ROS generation whilst plasma antioxidant status was unaltered and appeared to be sufficient to prevent the onset of typical oxidative stress. The clinical characteristics and the remaining biochemical parameters studied were similar for the two groups, except for a significantly decreased plasmatic level of uric acid in DM1 patients. This study suggests that the absence of complications in DM1 patients up to 5 years after onset of the disease may be associated with the oxidizing and reducing balance which need to be maintained in order to prevent or delay the onset of oxidative stress. The effective diabetic control involves evaluation of the oxidizing/antioxidant balance besides glycaemic control.