A Genome-Wide Association Study of Diabetic Kidney Disease in Subjects With Type 2 Diabetes

dentification of sequence variants robustly associated with predisposition to diabetic kidney disease (DKD) has the potential to provide insights into the pathophysiological mechanisms responsible. We conducted a genome-wide association study (GWAS) of DKD in type 2 diabetes (T2D) using eight complementary dichotomous and quantitative DKD phenotypes: the principal dichotomous analysis involved 5,717 T2D subjects, 3,345 with DKD. Promising association signals were evaluated in up to 26,827 subjects with T2D (12,710 with DKD). A combined T1D+T2D GWAS was performed using complementary data available for subjects with T1D, which, with replication samples, involved up to 40,340 subjects with diabetes (18,582 with DKD). Analysis of specific DKD phenotypes identified a novel signal near GABRR1 (rs9942471, P = 4.5 x 10(-8)) associated with microalbuminuria in European T2D case subjects. However, no replication of this signal was observed in Asian subjects with T2D or in the equivalent T1D analysis. There was only limited support, in this substantially enlarged analysis, for association at previously reported DKD signals, except for those at UMOD and PRKAG2, both associated with estimated glomerular filtration rate. We conclude that, despite challenges in addressing phenotypic heterogeneity, access to increased sample sizes will continue to provide more robust inference regarding risk variant discovery for DKD.Peer reviewe

Activation of immune responses against the basement membrane component collagen type IV does not affect the development of atherosclerosis in ApoE-deficient mice

Oxidation of low-density lipoprotein (LDL) in the arterial extracellular matrix results in malondialdehyde (MDA)-modifications of surrounding matrix proteins. We have recently demonstrated an association between high levels of autoantibodies against MDA-modified collagen type IV and risk for development of myocardial infarction. Collagen type IV is an important component of the endothelial basement membrane and influences smooth muscle cell function. We hypothesized that immune responses against collagen type IV could contribute to vascular injury affecting the development of atherosclerosis. To investigate this possibility, we induced an antibody-response against collagen type IV in apolipoprotein E (Apo E)-deficient mice. Female ApoE −/− mice on C57BL/6 background were immunized with α1α2 type IV collagen chain peptides linked to the immune-enhancer PADRE, PADRE alone or PBS at 12 weeks of age with three subsequent booster injections before the mice were killed at 23 weeks of age. Immunization of PADRE alone induced autoantibodies against PADRE, increased IL-4 secretion from splenocytes and reduced SMC content in the subvalvular plaques. Immunization with peptides of α1α2 type IV collagen chains induced a strong IgG1antibody response against collagen type IV peptides without affecting the distribution of T cell populations, plasma cytokine or lipid levels. There were no differences in atherosclerotic plaque development between collagen α1α2(IV)-PADRE immunized mice and control mice. Our findings demonstrate that the presence of antibodies against the basement membrane component collagen type IV does not affect atherosclerosis development in ApoE −/− mice. This suggests that the association between autoantibodies against collagen type IV and risk for myocardial infarction found in humans does not reflect a pathogenic role of these autoantibodies

Expansion of CD25+ Innate Lymphoid Cells Reduces Atherosclerosis

Biopharmaceutic

High levels of IgM against methylglyoxal-modified apolipoprotein B100 are associated with less coronary artery calcification in patients with type 2 diabetes

Objective: Advanced glycation end products (AGE) have been implicated in diabetic vascular complications through activation of pro-inflammatory genes. AGE-modified proteins are also targeted by the immune system resulting in the generation of AGE-specific autoantibodies, but the association of these immune responses with diabetic vasculopathy remains to be fully elucidated. The aim of this study was to determine whether antibodies against apolipoprotein B100 modified by methylglyoxal (MGO-apoB100) are associated with coronary atherosclerosis in patients with type 2 diabetes. Methods. We measured antibodies against MGO-apoB100 in plasma from 497 type 2 diabetic patients without clinical signs of cardiovascular disease. Severity of coronary disease was assessed as coronary artery calcium (CAC) imaging. Immunoglobulin (Ig)M and IgG levels recognizing MGO-apoB100 were determined by enzyme-linked immunosorbent assay. Results. Anti-MGO-apoB100 IgM antibody levels were higher in subjects with a low to moderate CAC score (≤400 Agatston units) than in subjects with a high score (>400 Agatston units; 136.8 ± 4.4 vs. 101.6 ± 7.4 arbitrary units (AU), P < 0.0001) and in subjects demonstrating no progression of CAC during 30 months of follow-up (136.4 ± 5.7 vs. 113.9 ± 6.2 AU in subjects with progression, P < 0.0001). Subjects with a family history of premature myocardial infarction had lower levels of anti-MGO-apoB100 IgM. Female subjects had higher levels of anti-MGO-apoB100 antibodies and lower CAC than men. Accordingly, high levels of IgM against MGO-apoB100 are associated with less severe and a lower risk of progression of coronary disease in subjects with type 2 diabetes. Conclusions. Although conclusions regarding causal relationships based on epidemiological observations need to be made with caution, our findings suggest the possibility that anti-MGO-apoB100 IgM may be protective in diabetic vasculopathy

Dendritic Cell KLF2 Expression Regulates T Cell Activation and Proatherogenic Immune Responses

Dendritic cells (DCs) have been implicated as important regulators of innate and adaptive inflammation in many diseases, including atherosclerosis. However, the molecular mechanisms by which DCs mitigate or promote inflammatory pathogenesis are only partially understood. Previous studies have shown an important anti-inflammatory role for the transcription factor Krüppel-like factor 2 (KLF2) in regulating activation of various cell types that participate in atherosclerotic lesion development, including endothelial cells, macrophages, and T cells. We used a pan-DC, CD11c-specific cre-lox gene knockout mouse model to assess the role of KLF2 in DC activation, function, and control of inflammation in the context of hypercholesterolemia and atherosclerosis. We found that KLF2 deficiency enhanced surface expression of costimulatory molecules CD40 and CD86 in DCs and promoted increased T cell proliferation and apoptosis. Transplant of bone marrow from mice with KLF2-deficient DCs into Ldlr−/− mice aggravated atherosclerosis compared with control mice, most likely due to heightened vascular inflammation evidenced by increased DC presence within lesions, enhanced T cell activation and cytokine production, and increased cell death in atherosclerotic lesions. Taken together, these data indicate that KLF2 governs the degree of DC activation and hence the intensity of proatherogenic T cell responses.FWN – Publicaties zonder aanstelling Universiteit Leide

Blockade of Tim-1 and Tim-4 Enhances Atherosclerosis in Low-Density Lipoprotein Receptor-Deficient Mice

Objective - T cell immunoglobulin and mucin domain (Tim) proteins are expressed by numerous immune cells, recognize phosphatidylserine on apoptotic cells, and function as costimulators or coinhibitors. Tim-1 is expressed by activated T cells but is also found on dendritic cells and B cells. Tim-4, present on macrophages and dendritic cells, plays a critical role in apoptotic cell clearance, regulates the number of phosphatidylserine-expressing activated T cells, and is genetically associated with low low-density lipoprotein and triglyceride levels. Because these functions of Tim-1 and Tim-4 could affect atherosclerosis, their modulation has potential therapeutic value in cardiovascular disease. Approach and Results - ldlr-/- mice were fed a high-fat diet for 4 weeks while being treated with control (rat immunoglobulin G1) or anti-Tim-1 (3D10) or -Tim-4 (21H12) monoclonal antibodies that block phosphatidylserine recognition and phagocytosis. Both anti-Tim-1 and anti-Tim-4 treatments enhanced atherosclerosis by 45% compared with controls by impairment of efferocytosis and increasing aortic CD4+T cells. Consistently, anti-Tim-4-treated mice showed increased percentages of activated T cells and late apoptotic cells in the circulation. Moreover, in vitro blockade of Tim-4 inhibited efferocytosis of oxidized low-density lipoprotein-induced apoptotic macrophages. Although anti-Tim-4 treatment increased T helper cell (Th)1 and Th2 responses, anti-Tim-1 induced Th2 responses but dramatically reduced the percentage of regulatory T cells. Finally, combined blockade of Tim-1 and Tim-4 increased atherosclerotic lesion size by 59%. Conclusions - Blockade of Tim-4 aggravates atherosclerosis likely by prevention of phagocytosis of phosphatidylserine-expressing apoptotic cells and activated T cells by Tim-4-expressing cells, whereas Tim-1-associated effects on atherosclerosis are related to changes in Th1/Th2 balance and reduced circulating regulatory T cells