The Role of the Microbiota in the Diabetic Peripheral Artery Disease.

Vascular complications of diabetes mellitus represent a major public health problem. Although many steps forward have been made to define the causes and to find the best possible therapies, the problem remains crucial. In recent years, more and more evidences have defined a link between microbiota and the initiation, promotion, and evolution of atherosclerotic disease, even in the diabetic scenario. There is an urgency to develop the knowledge of modern medicine about the link between gut microbiota and its host's metabolic pathways, and it would be useful to understand and justify the interindividual diversity of clinical disease presentation of diabetic vascular complication even if an optimization of pharmacological treatment has been made or in the case of young patients where hypertension, dyslipidemia, and diabetes are not able to justify a very quick progress of atherosclerotic process. The aim of the present review is to gather all the best available evidence in this regard and to define a new role of the microbiota in this field, from biomarker to possible therapeutic target.

Cardiovascular biomarkers in clinical studies of type 2 diabetes.

When planning cardiovascular (CV) studies in type 2 diabetes (T2D), selection of CV biomarkers is a complex issue. Because the pathophysiology of CV disease (CVD) in T2D is multifactorial, ideally, the selected CV biomarkers should cover all aspects of the known pathophysiology of the disease. This will allow the researcher to distinguish between effects on different aspects of the pathophysiology. To this end, we discuss a host of biomarkers grouped according to their role in the pathogenesis of CVD, namely: (1) cardiac damage biomarkers; (2) inflammatory biomarkers; and (3) novel biomarkers (oxidative stress and endothelial dysfunction biomarkers). Within each category we present the best currently validated biomarkers, with special focus on the population of interest (people with T2D). For each individual biomarker, we discuss the physiological role, validation in the general population and in people with T2D, analytical methodology, modifying factors, effects of glucose-lowering drugs, and interpretation. This approach will provide clinical researchers with the information necessary for planning, conducting and interpreting results from clinical trials. Furthermore, a systematic approach to selection of CV biomarkers in T2D research will improve the quality of future research.

Expression of CD4+ CD28null T lymphocytes in children and adolescents with type 1 diabetes mellitus: Relation to microvascular complications, aortic elastic properties, and carotid intima media thickness.

BACKGROUND: Cardiovascular risk in type 1 diabetes mellitus (T1DM) is associated with endothelial dysfunction, inflammation, and altered immunity. CD4+ CD28null T-cells are a subset of long-lived cytotoxic CD4+ T-lymphocytes with proatherogenic and plaque-destabilizing properties. We hypothesized that the frequency of CD4+ CD28null T-cells may be altered in T1DM and related to vascular complications. AIM: To assess the percentage of CD4+ CD28null T-lymphocytes in children and adolescents with T1DM and their relation to vascular structure and glycemic control. METHODS: Totally 100 patients with T1DM were divided into 2 groups according to the presence of microvascular complications and compared with 50 healthy controls. CD4+ CD28null T-lymphocytes were analyzed using flow cytometry. Aortic elastic properties and carotid intima media thickness (CIMT) were assessed. RESULTS: Aortic stiffness index and CIMT were significantly higher among patients compared with healthy controls while aortic strain and distensibility were decreased. The percentage of CD4+ CD28null T-cells was significantly higher in patients with and without microvascular complications compared with controls. High frequency of CD4+ CD28null T-cells was found among patients with microalbuminuria or peripheral neuropathy. Patients with CD4+ CD28null T-cells ≥10% had higher HbA1c, urinary albumin creatinine ratio, aortic stiffness, and CIMT. CD4+ CD28null T-cells were positively correlated to HbA1c, aortic stiffness index, and CIMT. CONCLUSIONS: Changes in aortic elastic properties and increased CIMT among young patients with T1DM may enable the recognition of preclinical cardiac impairment. The correlation between CD4+ CD28null T-cells and assessed parameters of vascular structure highlights the role of altered immune response in the occurrence of diabetic vascular complications.

Reduction of visceral fat by liraglutide is associated with ameliorations of hepatic steatosis, albuminuria, and micro-inflammation in type 2 diabetic patients with insulin treatment: a randomized control trial.

Liraglutide, an analogue of human glucagon-like peptide 1, reduces cardiovascular events in patients with type 2 diabetes; however, it has still been unknown by which mechanisms liraglutide could reduce cardiovascular events. Type 2 diabetic patients with insulin treatment were enrolled in this randomized, open-label, comparative study. Participants were randomly assigned to liraglutide plus insulin (liraglutide group) and insulin treatment (control group) at 1:1 allocation. Primary endpoint was the change in viscera fat are (VFA, cm2) at 24 weeks. Liver attenuation index (LAI) measured by abdominal computed tomography, urinary albumin-to-creatinine ratio (ACR, mg/g), and C-reactive protein (CRP) levels, skeletal muscle index (SMI), and quality of life (QOL) related to diabetes treatment were also determined. Seventeen patients (8; liraglutide group, 9; control group, mean age 59 ± 13 years; 53% female) completed this study. Liraglutide treatment significantly reduced VFA at 24 weeks; whereas, SFA was unchanged. ACR, LAI, and CRP levels were significantly reduced by liraglutide at 24 weeks and there was no difference in SMI between the two groups. Changes in VFA from baseline to 24 weeks were significantly associated with those in LAI, albuminuria, and HbA1c. Liraglutide treatment significantly improved QOL scores associated with anxiety and dissatisfaction with treatment and satisfaction with treatment. No severe adverse events were observed in both groups. Our data suggest that liraglutide could reduce visceral adiposity in parallel with attenuation of hepatic fat accumulation, albuminuria and micro-inflammation and improve QOL related to diabetes care in insulin-treated patients with type 2 diabetes.

Plasma levels of mannan-binding lectin-associated serine proteases MASP-1 and MASP-2 are elevated in type 1 diabetes and correlate with glycaemic control.

There is increasing evidence that the complement system plays an important role in diabetes and the development of diabetic vascular complications. In particular, mannan-binding lectin (MBL) levels are elevated in diabetes patients, and diabetes patients with diabetic nephropathy have higher MBL levels than diabetes patients with normal renal function. The MBL-associated serine proteases (MASPs) MASP-1, MASP-2 and MASP-3 and MBL-associated protein MAp44 have not yet been studied in diabetes patients. We therefore measured plasma levels of MASP-1, MASP-2, MASP-3 and MAp44 in 30 children with type 1 diabetes mellitus (T1DM) and 17 matched control subjects, and in 45 adults with T1DM and 31 matched control subjects. MASP-1 and MASP-2 levels were significantly higher in children and adults with T1DM than in their respective control groups, whereas MASP-3 and MAp44 levels did not differ between patients and controls. MASP-1 and MASP-2 levels correlated with HbA1c, and MASP levels decreased when glycaemic control improved. Because MASP-1 and MASP-2 have been shown to interact directly with blood coagulation, elevated levels of these proteins may play a role in the enhanced thrombotic environment and consequent vascular complications in diabetes.

Associations between clinical characteristics and chronic complications in latent autoimmune diabetes in adults and type 2 diabetes.

BACKGROUND: This study aimed to assess the associations between clinical characteristics and chronic complications in latent autoimmune diabetes in adults (LADA) and type 2 diabetes. METHODS: This is a retrospective study. Our diabetes registry included 6975 patients aged 30-75 years old with phenotypic type 2 diabetes who underwent islet autoantibody screening between 2003 and 2012; 384 patients were identified to have LADA. Rates of chronic complications for LADA and type 2 diabetes were compared using a 1 : 2 matched design. Logistic models were fitted to identify the presence of chronic diabetic complications using clinical characteristics including gender, age, duration of diabetes, glycemic control and metabolic syndrome. RESULTS: When duration of diabetes is <5 years, the prevalence of diabetic nephropathy (nephropathy; 12.2% versus 21.8%, p = 0.018) and diabetic retinopathy (retinopathy; 8.1% versus 15.9%, p = 0.011) were significantly lower in patients with LADA than in patients with type 2 diabetes; the prevalence of nephropathy and retinopathy were comparable between both groups when duration is ≥5 years. There was no significant difference in the prevalence of macrovascular complications between groups. The areas under the receiver operating characteristic curves based on the nephropathy and retinopathy models were larger for LADA than for type 2 diabetes (0.72 versus 0.61, p = 0.013; 0.76 versus 0.68, p = 0.056). CONCLUSIONS: Patients with LADA had a lower prevalence of microvascular complications than patients with type 2 diabetes when the duration of diabetes was <5 years. Regression equation fitted by clinical characteristics can better predict the risk of microvascular complications in LADA than in type 2 diabetes.

Probucol Protects Against Atherosclerosis Through Lipid-lowering and Suppressing Immune Maturation of CD11c+ Dendritic Cells in STZ-induced Diabetic LDLR-/- Mice.

Probucol, an agent characterized by lipid-lowering and antioxidant property, retards atherosclerosis effectively. To test the hypothesis that probucol might act its antiatherosclerotic role by suppressing immune maturation of dendritic cells (DCs), 7-week-old LDLR mice were rendered diabetic with streptozotocin (STZ) and then fed either a high-fat diet only or added with 0.5% (wt/wt) probucol for 4 months, and human monocyte-derived dendritic cells were preincubated with or without probucol and stimulated by oxidized low-density lipoprotein. In STZ-induced diabetic LDLR mice, probucol treatment significantly lowered plasma total cholesterol and high-density lipoprotein-cholesterol levels; regressed aortic atherosclerotic lesions; reduced splenic CD40, CD80, CD86, MHC-II expression, and plasma IL-12p70 production; and decreased the expression of CD11c DCs within atherosclerotic lesions. In vitro, oxidized low-density lipoprotein promoted human monocyte-derived dendritic cells maturation; stimulated CD40, CD86, CD1a, HLA-DR expression; increased tumor necrosis factor-α production; and decreased IL-4 production. However, these effects were obviously inhibited by probucol pretreatment. In conclusion, our study indicated that probucol effectively retarded atherosclerosis at least partly through lipid-lowering and inhibiting immune maturation of CD11c DCs in STZ-induced diabetic LDLR mice.

Differential impact of diabetes mellitus type II and arterial hypertension on collateral artery growth and concomitant macrophage accumulation.

BACKGROUND: Diabetes mellitus type II and arterial hypertension are major risk factors for peripheral arterial disease and have been considered to reduce collateral growth (arteriogenesis). Collateral growth proceeds through different stages. Vascular proliferation and macrophage accumulation are hallmarks of early collateral growth. MATERIAL AND METHODS: We here compare the impact of arterial hypertension and diabetes mellitus type II on collateral proliferation (Brdu incorporation) and macrophage accumulation (ED 2 staining) as well as collateral vessel function (collateral conductance) in a rat model of peripheral vascular disease (femoral artery occlusion), diabetes mellitus type II (Zucker fatty diabetic rats and Zucker lean rat controls) and arterial hypertension (induced via clip placement around the right renal arteriy). We furthermore tested the impact of monocyte chemoattractant protein-1 (MCP­1) on collateral proliferation and macrophage accumulation in these models RESULTS: Diabetic animals showed reduced vascular proliferation and macrophage accumulation, which however did not translate into a change of collateral conductance. Hypertensive animals on the contrary had reduced collateral conductances without altered macrophage accumulation and only a marginal reduction in collateral proliferation. Infusion of MCP­1 only enhanced vascular proliferation in diabetic animals. CONCLUSIONS: These findings illustrate that impaired monocyte/macrophage recruitment is responsible for reduced collateral growth under diabetic conditions but not in arterial hypertension suggesting that diabetes mellitus in particular affects early stages of collateral growth whereas hypertension has its impact on later remodeling stages. Successful pro-arteriogenic treatment strategies in a patient population that presents with diabetes mellitus and arterial hypertension need to address different stages of collateral growth and thus different molecular and cellular targets simultaneously.

Attenuation of flightless I improves wound healing and enhances angiogenesis in a murine model of type 1 diabetes.

AIMS/HYPOTHESIS: Skin lesions and ulcerations are severe complications of diabetes that often result in leg amputations. In this study we investigated the function of the cytoskeletal protein flightless I (FLII) in diabetic wound healing. We hypothesised that overexpression of FLII would have a negative effect on diabetic wound closure and modulation of this protein using specific FLII-neutralising antibodies (FnAb) would enhance cellular proliferation, migration and angiogenesis within the diabetic wound. METHODS: Using a streptozotocin-induced model of diabetes we investigated the effect of altered FLII levels through Flii genetic knockdown, overexpression or treatment with FnAb on wound healing. Diabetic wounds were assessed using histology, immunohistochemistry and biochemical analysis. In vitro and in vivo assays of angiogenesis were used to assess the angiogenic response. RESULTS: FLII levels were elevated in the wounds of both diabetic mice and humans. Reduction in the level of FLII improved healing of murine diabetic wounds and promoted a robust pro-angiogenic response with significantly elevated von Willebrand factor (vWF) and vascular endothelial growth factor (VEGF)-positive endothelial cell infiltration. Diabetic mouse wounds treated intradermally with FnAb showed improved healing and a significantly increased rate of re-epithelialisation. FnAb improved the angiogenic response through enhanced formation of capillary tubes and functional neovasculature. Reducing the level of FLII led to increased numbers of mature blood vessels, increased recruitment of smooth muscle actin-α-positive cells and improved tight junction formation. CONCLUSIONS/INTERPRETATION: Reducing the level of FLII in a wound may be a potential therapeutic approach for the treatment of diabetic foot ulcers.

Pharmacological inhibition of NOX reduces atherosclerotic lesions, vascular ROS and immune-inflammatory responses in diabetic Apoe(-/-) mice.

AIMS/HYPOTHESIS: Enhanced vascular inflammation, immune cell infiltration and elevated production of reactive oxygen species (ROS) contribute significantly to pro-atherogenic responses in diabetes. We assessed the immunomodulatory role of NADPH oxidase (NOX)-derived ROS in diabetes-accelerated atherosclerosis. METHODS: Diabetes was induced in male Apoe(-/-) mice with five daily doses of streptozotocin (55 mg kg(-1) day(-1)). Atherosclerotic plaque size, markers of ROS and immune cell accumulation were assessed in addition to flow cytometric analyses of cells isolated from the adjacent mediastinal lymph nodes (meLNs). The role of NOX-derived ROS was investigated using the NOX inhibitor, GKT137831 (60 mg/kg per day; gavage) administered to diabetic and non-diabetic Apoe(-/-) mice for 10 weeks. RESULTS: Diabetes increased atherosclerotic plaque development in the aortic sinus and this correlated with increased lesional accumulation of T cells and CD11c(+) cells and altered T cell activation in the adjacent meLNs. Diabetic Apoe(-/-) mice demonstrated an elevation in vascular ROS production and expression of the proinflammatory markers monocyte chemoattractant protein 1, vascular adhesion molecule 1 and IFNγ. Blockade of NOX-derived ROS using GKT137831 prevented the diabetes-mediated increase in atherosclerotic plaque area and associated vascular T cell infiltration and also significantly reduced vascular ROS as well as markers of inflammation and plaque necrotic core area. CONCLUSIONS/INTERPRETATION: Diabetes promotes pro-inflammatory immune responses in the aortic sinus and its associated lymphoid tissue. These changes are associated with increased ROS production by NOX. Blockade of NOX-derived ROS using the NOX inhibitor GKT137831 is associated with attenuation of these changes in the immune response and reduces the diabetes-accelerated development of atherosclerotic plaques in Apoe(-/-) mice.

Enhanced permeability responses to inflammation in streptozotocin-induced diabetic rat venules: Rho-mediated alterations of actin cytoskeleton and VE-cadherin.

Diabetes is a progressive disease that often leads to microvascular complications. This study investigates the impact of diabetes on microvessel permeability under basal and inflammatory conditions. Streptozotocin-induced diabetic rats were used to mimic type 1 diabetes. Parallel experiments were conducted in intact mesenteric venules in normal rats and diabetic rats experiencing hyperglycemia for 2-3 wk. Microvessel permeability was determined by measuring hydraulic conductivity (Lp). The correlated changes in endothelial intracellular Ca(2+) concentration ([Ca(2+)]i), adherens junctions, and cytoskeleton F-actin were examined with fluorescence imaging. Diabetic vessels showed moderately increased basal Lp, but upon platelet-activating factor (PAF) exposure, these vessels showed an ~10-fold higher Lp increase than the normal vessels. Concomitantly, we observed higher increases in endothelial [Ca(2+)]i, enhanced stress fiber formation, vascular endothelial-cadherin separation, and larger gap formation between endothelial cells than those occurring in normal vessels. PAF receptor staining showed no significant difference between normal and diabetic vessels. The application of Rho kinase inhibitor Y27632 did not affect PAF-induced increases in endothelial [Ca(2+)]i but significantly reduced PAF-induced Lp increases by 90% in diabetic vessels. The application of both Y27632 and nitric oxide (NO) synthase inhibitor attenuated PAF-induced Lp increases more than using one inhibitor alone. Our studies indicate that diabetic conditions prime endothelial cells into a phenotype with increased susceptibility to inflammation without altering receptor expression and that the increased Rho activation and NO production play important roles in exaggerated permeability increases when diabetic vessels were exposed to inflammatory mediators, which may account for the exacerbated vascular dysfunction when diabetic patients are exposed to additional inflammation.

Islet transplantation in type 1 diabetes: hype, hope and reality - a clinician's perspective.

The ß-cell replacement by islet transplantation is an attractive approach for normalizing blood glucose without hypoglycaemia in patient with type 1 diabetes mellitus (T1D). A pioneer study by the Edmonton group more than a decade ago showed that alloislet transplantation may result in insulin independence for at least 1 year after transplantation. This breakthrough excited researchers, physicians and patients, who felt that the ultimate goal of cure for T1D was at hand. Longer follow-up of patients who underwent islet transplantation showed less favourable results, with only approximately 10% of the patients remaining insulin-free 5 years after transplantation. In the last few years, progress has been made, and the success rate of islet transplantation has steadily increased. Important hurdles, however, related to limited tissue supply and need for life-long immunosuppressive drugs have yet to be overcome. Herein, we review recent achievements in islet transplantation and the challenges that still need to be addressed before this procedure can become a standard therapy for T1D.

PKCß promotes vascular inflammation and acceleration of atherosclerosis in diabetic ApoE null mice.

OBJECTIVE: Subjects with diabetes mellitus are at high risk for developing atherosclerosis through a variety of mechanisms. Because the metabolism of glucose results in production of activators of protein kinase C (PKC)ß, it was logical to investigate the role of PKCß in modulation of atherosclerosis in diabetes mellitus. APPROACH AND RESULTS: ApoE(-/-) and PKCß(-/-)/ApoE(-/-) mice were rendered diabetic with streptozotocin. Quantification of atherosclerosis, gene expression profiling, or analysis of signaling molecules was performed on aortic sinus or aortas from diabetic mice. Diabetes mellitus-accelerated atherosclerosis increased the level of phosphorylated extracellular signal-regulated kinase 1/2 and Jun-N-terminus kinase mitogen-activated protein kinases and augmented vascular expression of inflammatory mediators, as well as increased monocyte/macrophage infiltration and CD11c(+) cells accumulation in diabetic ApoE(-/-) mice, processes that were diminished in diabetic PKCß(-/-)/ApoE(-/-) mice. In addition, pharmacological inhibition of PKCß reduced atherosclerotic lesion size in diabetic ApoE(-/-) mice. In vitro, the inhibitors of PKCß and extracellular signal-regulated kinase 1/2, as well as small interfering RNA to Egr-1, significantly decreased high-glucose-induced expression of CD11c (integrin, alpha X 9 complement component 3 receptor 4 subunit]), chemokine (C-C motif) ligand 2, and interleukin-1ß in U937 macrophages. CONCLUSIONS: These data link enhanced activation of PKCß to accelerated diabetic atherosclerosis via a mechanism that includes modulation of gene transcription and signal transduction in the vascular wall, processes that contribute to acceleration of vascular inflammation and atherosclerosis in diabetes mellitus. Our results uncover a novel role for PKCß in modulating CD11c expression and inflammatory response of macrophages in the development of diabetic atherosclerosis. These findings support PKCß activation as a potential therapeutic target for prevention and treatment of diabetic atherosclerosis.

The effect of autoimmunity on the development time of microvascular complications in patients with type 1 diabetes mellitus.

BACKGROUND: Type 1 diabetes mellitus (DM) is an autoimmune disease with chronic complications that is becoming more frequent as life expectancy of diabetics has increased owing to improved methods of detection and better management. In this study, we investigated whether the presence of autoimmunity can be used in predicting the development time of microvascular complications. MATERIAL AND METHODS: Our study included 52 patients with type 1 diabetes mellitus (DM). The subjects had developed microvascular complications and they had been tested for anti-GAD (glutamic acid decarboxylase) antibodies and/or islet-cell antibodies (ICA). In the assessment of microvascular complications, we used ocular fundus examination, electromyography (EMG), and 24-h urine microalbuminuria tests. RESULTS: Of the patients included in the study, 30 were female and 22 were male. Of all patients characterized for the existence of diabetic complications, 36 of 52 had both diabetic retinopathy and diabetic nephropathy, 5 patients had diabetic neuropathy, and 11 patients had diabetic retinopathy only. At the diagnosis of diabetes, 20 in 52 patients tested negative for autoantibodies (anti-GAD and anti-ICA), while 32 of 52 tested positive for anti-GAD and/or anti-ICA. The mean HbA1C level of autoantibody-negative patients was 7.7%, while antibody-positive patients had slightly higher HbA1c levels (7.9%). However, this difference was not statistically significant (p>0.05). The mean development time of microvascular complications in autoantibody-positive patients was calculated as 11: 40±6.46 years, and in patients with negative autoimmunity results it was 10.91±6.70 years. CONCLUSIONS: The presence of diabetes-related autoantibodies (DRAs) in patients with type 1 diabetes mellitus does not have a significant effect on the development time of diabetic microvascular complications.

Role of epigenetic mechanisms in the vascular complications of diabetes.

Diabetes and metabolic disorders are leading causes of micro- and macrovascular complications. Furthermore, efforts to treat these complications are hampered by metabolic memory, a phenomenon in which prior exposure to hyperglycemia predisposes diabetic patients to the continued development of vascular diseases despite subsequent glycemic control. Persistently increased levels of oxidant stress and inflammatory genes are key features of these pathologies. Biochemical and molecular studies showed that hyperglycemia induced activation of NF-κB, signaling and actions of advanced glycation end products and other inflammatory mediators play key roles in the expression of pathological genes. In addition, epigenetic mechanisms such as posttranslational modification of histones and DNA methylation also play central roles in gene regulation by affecting chromatin structure and function. Recent studies have suggested that dysregulation of such epigenetic mechanisms may be involved in metabolic memory leading to persistent changes in the expression of genes associated with diabetic vascular complications. Further exploration of these mechanisms by also taking advantages of recent advances in high throughput epigenomics technologies will greatly increase our understanding of epigenetic variations in diabetes and its complications. This in turn can lead to the development of novel new therapies.

[Effects of physical stimulation of spontaneous arteriolar vasomotion on microcirculation and the immune system in diabetes and impaired wound healing]. / Wirkungen einer physikalischen Stimulierung der spontanen arteriolären Vasomotion auf Mikrozirkulation und Immunsystem bei Diabetes und Wundheilungsstörungen.

BACKGROUND: Whether and to what extent the complementary use of a biorhythm-defined physical stimulation of insufficient spontaneous arteriolar vasomotion contributes to increasing the therapeutic success of established treatment concepts were examined. MATERIALS AND METHODS: In a placebo-controlled study on a biometrically defined sample of older diabetes patients with impaired wound healing, measurements of representative features of the functional status of the microcirculation and the immune system were investigated using high-resolution methods (intravital microscopy, reflective spectrometry, white light spectroscopy combined with laser Doppler microflow measurements). The stimulation signal corresponding to physiological spontaneous arteriolar vasomotion was transmitted using an electromagnetic alternating field of low magnetic flux density. RESULTS: During the 27-day treatment and observation period, a complementary treatment effect of the applied biorhythm-defined physical vasomotion stimulation could be detected.

Thrombospondins: old players, new games.

PURPOSE OF REVIEW: Thrombospondins (TSPs) are secreted extracellular matrix (ECM) proteins from TSP family, which consists of five homologous members. They share a complex domain structure and have numerous binding partners in ECM and multiple cell surface receptors. Information that has emerged over the past decade identifies TSPs as important mediators of cellular homeostasis, assigning new important roles in cardiovascular pathology to these proteins. RECENT FINDINGS: Recent studies of the functions of TSP in the cardiovascular system, diabetes and aging, which placed several TSPs in a position of critical regulators, demonstrated the involvement of these proteins in practically every aspect of cardiovascular pathophysiology related to atherosclerosis: inflammation, immunity, leukocyte recruitment and function, function of vascular cells, angiogenesis, and responses to hypoxia, ischemia and hyperglycemia. TSPs are also critically important in the development and ultimate outcome of the complications associated with atherosclerosis--myocardial infarction, and heart hypertrophy and failure. Their expression and significance increase with age and with the progression of diabetes, two major contributors to the development of atherosclerosis and its complications. SUMMARY: This overview of recent literature examines the latest information on the newfound functions of TSPs that emphasize the importance of ECM in cardiovascular homeostasis and pathology. The functions of TSPs in myocardium, vasculature, vascular complications of diabetes, aging and immunity are discussed.

Blood immune cell profiling in adults with longstanding type 1 diabetes is associated with macrovascular complications.

Aims/hypothesis: There is increasing evidence for heterogeneity in type 1 diabetes mellitus (T1D): not only the age of onset and disease progression rate differ, but also the risk of complications varies markedly. Consequently, the presence of different disease endotypes has been suggested. Impaired T and B cell responses have been established in newly diagnosed diabetes patients. We hypothesized that deciphering the immune cell profile in peripheral blood of adults with longstanding T1D may help to understand disease heterogeneity. Methods: Adult patients with longstanding T1D and healthy controls (HC) were recruited, and their blood immune cell profile was determined using multicolour flow cytometry followed by a machine-learning based elastic-net (EN) classification model. Hierarchical clustering was performed to identify patient-specific immune cell profiles. Results were compared to those obtained in matched healthy control subjects. Results: Hierarchical clustering analysis of flow cytometry data revealed three immune cell composition-based distinct subgroups of individuals: HCs, T1D-group-A and T1D-group-B. In general, T1D patients, as compared to healthy controls, showed a more active immune profile as demonstrated by a higher percentage and absolute number of neutrophils, monocytes, total B cells and activated CD4+CD25+ T cells, while the abundance of regulatory T cells (Treg) was reduced. Patients belonging to T1D-group-A, as compared to T1D-group-B, revealed a more proinflammatory phenotype characterized by a lower percentage of FOXP3+ Treg, higher proportions of CCR4 expressing CD4 and CD8 T cell subsets, monocyte subsets, a lower Treg/conventional Tcell (Tconv) ratio, an increased proinflammatory cytokine (TNFα, IFNγ) and a decreased anti-inflammatory (IL-10) producing potential. Clinically, patients in T1D-group-A had more frequent diabetes-related macrovascular complications. Conclusions: Machine-learning based classification of multiparameter flow cytometry data revealed two distinct immunological profiles in adults with longstanding type 1 diabetes; T1D-group-A and T1D-group-B. T1D-group-A is characterized by a stronger pro-inflammatory profile and is associated with a higher rate of diabetes-related (macro)vascular complications.

Association of the systemic immuno-inflammation index, neutrophil-to-lymphocyte ratio, and platelet-to-lymphocyte ratio with diabetic microvascular complications.

Background: The systemic immuno-inflammation index (SII), neutrophil-to-lymphocyte ratio (NLR), and platelet-to-lymphocyte ratio (PLR) are widely used and have been shown to be predictive indicators of various diseases. Diabetic nephropathy (DN), retinopathy (DR), and peripheral neuropathy (DPN) are the most prominent and common microvascular complications, which have seriously negative impacts on patients, families, and society. Exploring the associations with these three indicators and diabetic microvascular complications are the main purpose. Methods: There were 1058 individuals with type 2 diabetes mellitus (T2DM) in this retrospective cross-sectional study. SII, NLR, and PLR were calculated. The diseases were diagnosed by endocrinologists. Logistic regression and subgroup analysis were applied to evaluate the association between SII, NLP, and PLR and diabetic microvascular complications. Results: SII, NLR, and PLR were significantly associated with the risk of DN [odds ratios (ORs): 1.52, 1.71, and 1.60, respectively] and DR [ORs: 1.57, 1.79, and 1.55, respectively] by multivariate logistic regression. When NLR ≥2.66, the OR was significantly higher for the risk of DPN (OR: 1.985, 95% confidence interval: 1.29-3.05). Subgroup analysis showed no significant positive associations across different demographics and comorbidities, including sex, age, hypertension, HbA1c (glycated hemoglobin), and dyslipidemia. Conclusion: This study found a positive relationship between NLR and DN, DR, and DPN. In contrast, SII and PLR were found to be only associated with DN and DR. Therefore, for the diagnosis of diabetic microvascular complications, SII, NLR and PLR are highly valuable.

An unbalanced monocyte polarisation in peripheral blood and bone marrow of patients with type 2 diabetes has an impact on microangiopathy.

AIM/HYPOTHESIS: Monocytes/macrophages play important roles in adipose and vascular tissues and can be polarised as inflammatory M1 or anti-inflammatory M2. We sought to analyse monocyte polarisation status in type 2 diabetes, which is characterised by chronic inflammation. METHODS: We enrolled 60 individuals without diabetes and 53 patients with type 2 diabetes. We quantified standard monocyte subsets defined by cluster of differentiation (CD)14 and CD16. In addition, based on the phenotype of polarised macrophages in vitro, we characterised and quantified more definite M1 (CD68(+)CCR2(+)) and M2 (CX3CR1(+)CD206(+)/CD163(+)) monocytes. We also analysed bone marrow (BM) samples and the effects of granulocyte-colony stimulating factor (G-CSF) stimulation in diabetic and control individuals. RESULTS: We found no alterations in standard monocyte subsets (classical, intermediate and non-classical) when comparing groups. For validation of M1 and M2 phenotypes, we observed that M2 were enriched in non-classical monocytes and had lower TNF-α content, higher LDL scavenging and lower transendothelial migratory capacity than M1. Diabetic patients displayed an imbalanced M1/M2 ratio compared with the control group, attributable to a reduction in M2. The M1/M2 ratio was directly correlated with waist circumference and HbA1c and, among diabetic patients, M2 reduction and M1/M2 increase were associated with microangiopathy. A decrease in M2 was also found in the BM from diabetic patients, with a relative M2 excess compared with the bloodstream. BM stimulation with G-CSF mobilised M2 macrophages in diabetic but not in healthy individuals. CONCLUSIONS/INTERPRETATION: We show that type 2 diabetes markedly reduces anti-inflammatory M2 monocytes through a dysregulation in bone-marrow function. This defect may have a negative impact on microangiopathy.