Genome-wide analysis of DNA methylation in subjects with type 1 diabetes identifies epigenetic modifications associated with proliferative diabetic retinopathy.

BACKGROUND: Epigenetic variation has been linked to several human diseases. Proliferative diabetic retinopathy (PDR) is a major cause of vision loss in subjects with diabetes. However, studies examining the association between PDR and the genome-wide DNA methylation pattern are lacking. Our aim was to identify epigenetic modifications that associate with and predict PDR in subjects with type 1 diabetes (T1D). METHODS: DNA methylation was analyzed genome-wide in 485,577 sites in blood from cases with PDR (n = 28), controls (n = 30), and in a prospective cohort (n = 7). False discovery rate analysis was used to correct the data for multiple testing. Study participants with T1D diagnosed before 30 years of age and insulin treatment within 1 year from diagnosis were selected based on 1) subjects classified as having PDR (cases) and 2) subjects with T1D who had had diabetes for at least 10 years when blood DNA was sampled and classified as having no/mild diabetic retinopathy also after an 8.7-year follow-up (controls). DNA methylation was also analyzed in a prospective cohort including seven subjects with T1D who had no/mild diabetic retinopathy when blood samples were taken, but who developed PDR within 6.3 years (converters). The retinopathy level was classified by fundus photography. RESULTS: We identified differential DNA methylation of 349 CpG sites representing 233 unique genes including TNF, CHI3L1 (also known as YKL-40), CHN2, GIPR, GLRA1, GPX1, AHRR, and BCOR in cases with PDR compared with controls. The majority of these sites (79 %) showed decreased DNA methylation in cases with PDR. The Natural Killer cell-mediated cytotoxicity pathway was found to be significantly (P = 0.006) enriched among differentially methylated genes in cases with PDR. We also identified differential DNA methylation of 28 CpG sites representing 17 genes (e.g. AHRR, GIPR, GLRA1, and BCOR) with P <0.05 in the prospective cohort, which is more than expected by chance (P = 0.0096). CONCLUSIONS: Subjects with T1D and PDR exhibit altered DNA methylation patterns in blood. Some of these epigenetic changes may predict the development of PDR, suggesting that DNA methylation may be used as a prospective marker of PDR.

Photocoagulation of human retinal pigment epithelium in vitro: unravelling the effects on ARPE-19 by transcriptomics and proteomics.

PURPOSE: Despite the extensive use of retinal photocoagulation for ischaemia and vascular leakage in retinal vascular disease, the molecular mechanisms behind its clinical beneficial effects are still poorly understood. One important target of laser irradiation is the retinal pigment epithelium (RPE). In this study, we aimed at identifying the isolated effects of photocoagulation of RPE at both the mRNA and protein expression levels. METHODS: Human ARPE-19 cells were exposed to photocoagulation. Gene expression and protein expression were compared to untreated cells using microarray and liquid chromatography-mass spectrometry analysis. Genes and proteins queried by microarray and mass spectrometry were subjected to the Kyoto Encyclopedia of Genes and Genomes (KEGG) database pathway analyses. RESULTS: Laser irradiation resulted in an induction of the cytoprotective heat-shock protein subfamily Hsp70 as well as in a suppression of the vascular permeability factor carbonic anhydrase 9 (CA9). These expression patterns were evident at both the mRNA and protein levels. KEGG pathway analyses revealed genes and proteins involved in cellular turnover, repair and inflammation. CONCLUSIONS: By characterizing the transcriptional and translational effects of laser coagulation on the RPE cells in culture, we have revealed responses, which might contribute to some of the beneficial effects obtained by photocoagulation for ischaemia and vascular leakage in retinal vascular disease.

GAD autoantibody affinity in adult patients with latent autoimmune diabetes, the study participants of a GAD65 vaccination trial.

OBJECTIVE: Patients with latent autoimmune diabetes in adults (LADA) express autoantibodies against the 65-kDa isoform of GAD (GADA). Intervention with recombinant human GAD65 formulated with aluminium hydroxide (GAD-alum) given twice subcutaneously to LADA patients at intervals of 4 weeks was safe and did not compromise ß-cell function in a Phase II clinical trial. GADA affinity has been shown to predict progression to type 1 diabetes. Here, we asked whether GADA affinity was affected by the GAD65 antigen-specific vaccination and/or associated with ß-cell function in participants of this trial. RESEARCH DESIGN AND METHODS: GADA affinity was measured in sera of 46 LADA patients obtained prior to the first week and 20 weeks after the second injection with GAD-alum or placebo using competitive binding experiments with [125I]-labeled and unlabeled human GAD65. RESULTS: At baseline, GADA affinities ranged from 1.9 × 10(7) to 5.0 × 10(12) L/mol (median 2.8 × 10(10) L/mol) and were correlated with GADA titers (r = 0.47; P = 0.0009), fasting (r = -0.37; P = 0.01) and stimulated (r = -0.40; P = 0.006) C-peptide concentrations, and HbA1c (r = 0.39; P = 0.007). No significant changes in affinity were observed from baseline to week 24. Patients with GADA affinities in the lower first quartile (<4 × 10(9) L/mol) had better preserved fasting C-peptide concentrations at baseline than those with higher affinities (mean 1.02 vs. 0.66 nmol/L; P = 0.004) and retained higher concentrations over 30 months of follow-up (mean 1.26 vs. 0.62 nmol/L; P = 0.01). CONCLUSIONS: Intervention with GAD-alum in LADA patients had no effect on GADA affinity. Our data suggest that patients with low GADA affinity have a prolonged preservation of residual ß-cell function.

Cardiovascular disease in relation to diabetes status in immigrants from the Middle East compared to native Swedes: a cross-sectional study.

BACKGROUND: Type 2 diabetes is highly prevalent in immigrants to Sweden from Iraq, but the prevalence of cardiovascular disease (CVD) and its risk factors are not known. In this survey we aimed to compare the prevalence of CVD and CVD-associated risk factors between a population born in Iraq and individuals born in Sweden. METHODS: This population-based, cross-sectional study comprised 1,365 Iraqi immigrants and 739 Swedes (age 30-75 years) residing in the same socioeconomic area in Malmö, Sweden. Blood tests were performed and socio-demography and lifestyles were characterized. To investigate potential differences in CVD, odds ratios (ORs) with 95% confidence intervals (CIs) were estimated by multivariate logistic regression analysis with adjustment for metabolic, lifestyle and psychosocial risk factors for CVD. Outcome measures were odds of CVD. RESULTS: There were no differences in self-reported prevalence of CVD between Iraqi- and Swedish-born individuals (4.0 vs. 5.5%, OR 0.9, 95% CI 0.4-1.8). However, the prevalence of type 2 diabetes was higher in Iraqi compared to Swedish participants (8.4 vs. 3.3%, OR = 4.2, 95% CI 2.6-6.7). Moreover, among individuals with type 2 diabetes, Iraqis had a higher prevalence of CVD (22.8 vs. 8.0%, OR = 4.2, 95% CI 0.9-20.0), after adjustment for age and sex. By contrast, among those without diabetes, immigrants from Iraq had a lower prevalence of CVD than Swedes (2.2 vs. 5.5%, OR = 0.6, 95% CI 0.3-0.9).Type 2 diabetes was an independent risk factor for CVD in Iraqis only (OR = 6.8, 95% CI 2.8-16.2). This was confirmed by an interaction between country of birth and diabetes (p = 0.010). In addition, in Iraqis, type 2 diabetes contributed to CVD risk to a higher extent than history of hypertension (standardized OR 1.5 vs. 1.4). CONCLUSIONS: This survey indicates that the odds of CVD in immigrants from Iraq are highly dependent on the presence or absence of type 2 diabetes and that type 2 diabetes contributes with higher odds of CVD in Iraqi immigrants compared to native Swedes. Our study suggests that CVD prevention in immigrants from the Middle East would benefit from prevention of type 2 diabetes.

Photocoagulation of human retinal pigment epithelial cells in vitro: evaluation of necrosis, apoptosis, cell migration, cell proliferation and expression of tissue repairing and cytoprotective genes.

AIMS: Sight-threatening diabetic retinopathy has been treated with photocoagulation for decades but the mechanisms behind the beneficial clinical effects are poorly understood. One target of irradiation and a potential player in this process is the retinal pigment epithelium (RPE). Here we establish an in vitro model for photocoagulation of human RPE cells. METHODS: ARPE-19 cells were exposed to photocoagulation and studied at various time points up to 168h. Lesion morphology, necrosis and apoptosis were investigated by light microscopy; LIVE/DEAD staining and measurements of lactate dehydrogenase activity; and TUNEL- and ELISA-based quantification of DNA fragments, respectively. Cell migration and proliferation were explored using docetaxel and mitomycin C; temporal and spatial changes in proliferation were assessed by confocal immunofluorescence of proliferating cell nuclear antigen. Gene expression was measured by qPCR. RESULTS: Photocoagulation of ARPE-19 resulted in denaturation of proteins and reproducible lesion formation. A transient peak in necrosis, followed by a peak in apoptosis was observed in cells within the lesions at 6h and 24h, respectively after photocoagulation. Cell proliferation was depressed during the first hours after photocoagulation, back to control levels at 24h and augmented in the following days. These effects were not limited to cells in the lesions, but also evident in neighbouring cells. Changes in cell proliferation during lesion repair were preceded by changes in cell migration. Altered mRNA expression of genes previously implicated in the regulation of cell proliferation (FOS, IL-1ß, IL-8, HMGA2), migration and tissue repairing (TGFBR2, ADAMTS6, TIMP3, CTGF) was observed, as well as increased expression of the alarmin IL33 and the cytoprotective gene HSPA6. CONCLUSIONS: Using a laser system and experimental settings that comply with standards used in clinical practice, we have established a suitable model for in vitro photocoagulation of human RPE cells to isolate their contribution to the beneficial effects of laser treatment.

Profile of intraocular tumour necrosis factor-α and interleukin-6 in diabetic subjects with different degrees of diabetic retinopathy.

PURPOSE: To assess and correlate the levels of inflammatory mediators in the eyes from non-diabetic and diabetic subjects without retinopathy (NDR), with non-proliferative diabetic retinopathy (NPDR) or with proliferative diabetic retinopathy (PDR) to corresponding erum levels. METHODS: The levels of interleukin 1ß, interleukin-6 (IL-6) and tumour necrosis factor-α (TNF-α) were analysed by an ELISA-mimicking technique in the vitreous from 26 diabetic subjects with active PDR and 27 non-diabetic subjects, or by a multiplex bead assay in the aqueous humour from 35 diabetic subjects with NDR/NPDR and 40 non-diabetic subjects. Intraocular protein production was estimated in vitreous specimens by calculating a vitreous/serum ratio. RESULTS: In the vitreous, IL-6 was higher in diabetic [157.5 (25.0-1401.0) pg/ml; median (min-max)] than in non-diabetic subjects [44.0 (5.0-4425) pg/ml; p = 0.021]. The vitreous/serum ratio was high (55.5:1 and 16:1, respectively), suggesting intraocular production. TNF-α was lower in diabetic [18.0 (8.0-46.0) pg/ml] than in non-diabetic subjects [22.0 (13.0-47.0) pg/ml; p = 0.034], but the vitreous/serum ratio was elevated in both groups (2:1 and 3.4:1, respectively). TNF-α levels were higher in serum from diabetic subjects [9.0 (5.0-53.0) pg/ml versus 6.7 (3.0-11.0) pg/ml; p < 0.001]. Aqueous levels of inflammatory mediators did not differ between diabetic subjects with NDR/NPDR and non-diabetic subjects despite elevated TNF-α in serum [27.8 (6.8-153.7) pg/ml versus 16.4 (4.1-42.4) pg/ml; p = 0.021]. CONCLUSION: Intraocular inflammation seems to be involved in PDR but does not seem to be prominent in early retinopathy stages, i.e. NDR or NPDR. Diabetic subjects have an overall increased inflammatory activity compared to non-diabetic subjects, as demonstrated by increased serum levels of TNF-α.

Switching from high-fat to low-fat diet normalizes glucose metabolism and improves glucose-stimulated insulin secretion and insulin sensitivity but not body weight in C57BL/6J mice.

OBJECTIVES: Environmental factors such as a high-fat diet contribute to type 2 diabetes and obesity. This study examined glycemia, insulin sensitivity, and ß-cell function after switching from a high-fat diet to a low-fat diet in mice. METHODS: C57BL/6J mice were fed a high-fat diet or low-fat diet for 18 months, after which mice on the high-fat diet either maintained this diet or switched to a low-fat diet for 4 weeks. Body weight and glucose and insulin responses to intraperitoneal glucose were determined. Insulin secretion (insulinogenic index: the 10-minute insulin response divided by the 10-minute glucose level) and insulin sensitivity (1 divided by basal insulin) were determined. RESULTS: After 18 months on a high-fat diet, mice had glucose intolerance, marked hyperinsulinemia, and increased body weight compared to mice on a low-fat diet (P < 0.001). Switching from a high-fat diet to low-fat diet normalized glucose tolerance, reduced but not normalized body weight (P < 0.001), increased insulin secretion (248 ± 39 vs 141 ± 46 pmol/mmol; P = 0.028) and improved but not normalized insulin sensitivity (3.2 ± 0.1 vs 1.0 ± 0.1 [pmol/L]; P = 0.012). CONCLUSION: Switching from a high-fat diet to low-fat diet normalizes glucose tolerance and improves but not normalizes insulin secretion and insulin sensitivity. These effects are more pronounced than the reduced body weight.

HLA-DQB1 genotypes and islet cell autoantibodies against GAD65 and IA-2 in relation to development of diabetes post partum in women with gestational diabetes mellitus.

AIMS: To study HLA-DQB1 genes and islet cell autoantibodies against glutamic acid decarboxylase 65 (GADA) and insulinoma antigen-2 (IA-2A) in relation to diabetes post partum in mothers with diagnosed gestational diabetes mellitus (GDM). METHODS: During 2003-2004, women undergoing a 75 g oral glucose tolerance test (OGTT) during pregnancy were invited to participate in the Mamma Study. Cut-off level defining GDM was a 2-h capillary blood glucose of 7.8 mmol/L. 1-2 years after delivery a 75 g OGTT was performed, GADA and IA-2A were measured and HLA-DQB1 genes analysed. Data were available for 452 mothers with previous GDM and 168 randomly selected control subjects. RESULTS: HLA-DQB1*0602 was negatively associated with GDM (p=0.033) and with development of diabetes post partum (p=0.017), whereas high risk HLA were not associated with GDM or with diabetes. The presence of GADA post partum was positively associated with diabetes post partum (p=0.0009), but not with impaired glucose tolerance. CONCLUSIONS: Mothers with GDM and HLA-DQB1*0602 were less likely to develop diabetes after pregnancy, and type 1 diabetes associated high risk HLA genes did not predict type 1 diabetes post partum. Additionally, GADA were positively associated with diabetes development.

High prevalence of type 2 diabetes in Iraqi and Swedish residents in a deprived Swedish neighbourhood--a population based study.

BACKGROUND: Immigrants from the Middle-East are at high risk of developing type 2 diabetes (T2D). The aim of the present survey was to measure, in a single deprived neighbourhood, the prevalence rates of impaired fasting glucose (IFG), impaired glucose tolerance (IGT) and T2D in residents originating from Iraq and to compare them to those in residents born in Sweden. An additional aim was to identify metabolic, lifestyle and socioeconomic risk factors associated with IFG/IGT and T2D in these residents. METHODS: The study was conducted February 1'st to March 31'st 2010. Men and women aged 45 to 65 years of Swedish or Iraqi origin, living in the neighbourhood of Rosengård, Malmö, Sweden, were randomly selected from the census register. Each participant signed a written informed consent form, underwent a physical examination and an oral glucose tolerance test (OGTT), provided blood samples and filled in a questionnaire. A total of 175 subjects participated (Swedish origin n = 79, Iraqi origin n = 96), reflecting an overall response rate of almost 60%. RESULTS: In total, 21.9% and 19.0% of the Iraqi and Swedish participants, respectively, suffered from T2D, while 24.0% of the Iraqi participants and 25.3% of the Swedish participants had IFG/IGT. There were no significant differences in prevalence rates relating to country of origin.Obesity (BMI ≥30 kg/m2) and sedentary leisure time physical activity were highly prevalent in both groups, while a family history of diabetes was more prevalent in participants from Iraq (49.2%) than in those from Sweden (22.8%) (p = 0.001).Being obese or having a sedentary leisure time were, independently associated with T2D (OR 5.43 (95% CI 2.10-14.02) and 2.89 (95% CI 1.03-8.10) respectively), while economic difficulties were independently associated with IFG/IGT (OR 2.55 (95% CI 1.06-6.15)) after adjustment for the confounding effects of other common risk factors for T2D. CONCLUSIONS: This study reveals a high prevalence of T2D, independently of country of origin (Iraq or Sweden), in a socially vulnerable area and additionally presents a risk factor profile that is markedly different from that of Sweden in general.

Vascular cellular adhesion molecule-1 (VCAM-1) expression in mice retinal vessels is affected by both hyperglycemia and hyperlipidemia.

BACKGROUND: Inflammation has been proposed to be important in the pathogenesis of diabetic retinopathy. An early feature of inflammation is the release of cytokines leading to increased expression of endothelial activation markers such as vascular cellular adhesion molecule-1 (VCAM-1). Here we investigated the impact of diabetes and dyslipidemia on VCAM-1 expression in mouse retinal vessels, as well as the potential role of tumor necrosis factor-α (TNFα). METHODOLOGY/PRINCIPAL FINDINGS: Expression of VCAM-1 was examined by confocal immunofluorescence microscopy in vessels of wild type (wt), hyperlipidemic (ApoE(-/-)) and TNFα deficient (TNFα(-/-), ApoE(-/-)/TNFα(-/-)) mice. Eight weeks of streptozotocin-induced diabetes resulted in increased VCAM-1 in wt mice, predominantly in small vessels (<10 µm). Diabetic wt mice had higher total retinal TNFα, IL-6 and IL-1ß mRNA than controls; as well as higher soluble VCAM-1 (sVCAM-1) in plasma. Lack of TNFα increased higher basal VCAM-1 protein and sVCAM-1, but failed to up-regulate IL-6 and IL-1ß mRNA and VCAM-1 protein in response to diabetes. Basal VCAM-1 expression was higher in ApoE(-/-) than in wt mice and both VCAM-1 mRNA and protein levels were further increased by high fat diet. These changes correlated to plasma cholesterol, LDL- and HDL-cholesterol, but not to triglycerides levels. Diabetes, despite further increasing plasma cholesterol in ApoE(-/-) mice, had no effects on VCAM-1 protein expression or on sVCAM-1. However, it increased ICAM-1 mRNA expression in retinal vessels, which correlated to plasma triglycerides. CONCLUSIONS/SIGNIFICANCE: Hyperglycemia triggers an inflammatory response in the retina of normolipidemic mice and up-regulation of VCAM-1 in retinal vessels. Hypercholesterolemia effectively promotes VCAM-1 expression without evident stimulation of inflammation. Diabetes-induced endothelial activation in ApoE(-/-) mice seems driven by elevated plasma triglycerides but not by cholesterol. Results also suggest a complex role for TNFα in the regulation of VCAM-1 expression, being protective under basal conditions but pro-inflammatory in response to diabetes.

Evaluation of the aldose reductase inhibitor fidarestat on ischemia-reperfusion injury in rat retina.

This study evaluated the effects of retinal ischemia-reperfusion (IR) injury and pre-treatment with the potent and specific aldose reductase inhibitor fidarestat on apoptosis, aldose reductase and sorbitol dehydrogenase expression, sorbitol pathway intermediate concentrations, and oxidative-nitrosative stress. Female Wistar rats were pre-treated with either vehicle (N-methyl-D-glucamine) or fidarestat, 32 mg kg(-1) d(-1) for both, in the right jugular vein, for 3 consecutive days. A group of vehicle- and fidarestat-treated rats were subjected to 45-min retinal ischemia followed by 24-h reperfusion. Ischemia was induced 30 min after the last vehicle or fidarestat administration. Retinal IR resulted in a remarkable increase in retinal cell death. The number of TUNEL-positive nuclei increased 48-fold in the IR group compared with non-ischemic controls (p<0.01), and this increase was partially prevented by fidarestat. AR expression (Western blot analysis) increased by 19% in the IR group (p<0.05), and this increase was prevented by fidarestat. Sorbitol dehydrogenase and nitrated protein expressions were similar among all experimental groups. Retinal sorbitol concentrations tended to increase in the IR group but the difference with non-ischemic controls did not achieve statistical significance (p=0.08). Retinal fructose concentrations were 2.2-fold greater in the IR group than in the non-ischemic controls (p<0.05). Fidarestat pre-treatment of rats subjected to IR reduced retinal sorbitol concentration to the levels in non-ischemic controls. Retinal fructose concentrations were reduced by 41% in fidarestat-pre-treated IR group vs. untreated ischemic controls (p=0.0517), but remained 30% higher than in the non-ischemic control group. In conclusion, IR injury to rat retina is associated with a dramatic increase in cell death, elevated AR expression and sorbitol pathway intermediate accumulation. These changes were prevented or alleviated by the AR inhibitor fidarestat. The results identify AR as an important therapeutic target for diseases involving IR injury, and provide the rationale for development of fidarestat and other AR inhibitors.

Nuclear factor of activated T cells regulates osteopontin expression in arterial smooth muscle in response to diabetes-induced hyperglycemia.

OBJECTIVE: Hyperglycemia is a recognized risk factor for cardiovascular disease in diabetes. Recently, we reported that high glucose activates the Ca(2+)/calcineurin-dependent transcription factor nuclear factor of activated T cells (NFAT) in arteries ex vivo. Here, we sought to determine whether hyperglycemia activates NFAT in vivo and whether this leads to vascular complications. METHODS AND RESULTS: An intraperitoneal glucose-tolerance test in mice increased NFATc3 nuclear accumulation in vascular smooth muscle. Streptozotocin-induced diabetes resulted in increased NFATc3 transcriptional activity in arteries of NFAT-luciferase transgenic mice. Two NFAT-responsive sequences in the osteopontin (OPN) promoter were identified. This proinflammatory cytokine has been shown to exacerbate atherosclerosis and restenosis. Activation of NFAT resulted in increased OPN mRNA and protein in native arteries. Glucose-induced OPN expression was prevented by the ectonucleotidase apyrase, suggesting a mechanism involving the release of extracellular nucleotides. The calcineurin inhibitor cyclosporin A or the novel NFAT blocker A-285222 prevented glucose-induced OPN expression. Furthermore, diabetes resulted in higher OPN expression, which was significantly decreased by in vivo treatment with A-285222 for 4 weeks or prevented in arteries from NFATc3(-/-) mice. CONCLUSIONS: These results identify a glucose-sensitive transcription pathway in vivo, revealing a novel molecular mechanism that may underlie vascular complications of diabetes.

Exposure to p,p'-DDE: a risk factor for type 2 diabetes.

BACKGROUND: Persistent organic pollutants (POPs), such as PCBs, DDT and dioxins have in several cross-sectional studies shown strong associations with type 2 diabetes mellitus. Reversed causality can however not be excluded. The aim of this case-control study was to evaluate whether POPs concentration is a risk factor for type 2 diabetes. METHODOLOGY/PRINCIPAL FINDINGS: A case-control study was performed within a well-defined cohort of women, age 50-59 years, from the Southern part of Sweden. Biomarkers for POP exposure, 2,2',4,4',5,5'-hexachlorobiphenyl (CB-153) and 1,1-dichloro-2,2-bis (p-chlorophenyl)-ethylene (p,p'-DDE) were analyzed in stored serum samples, which were collected at the baseline examination when the cohort was established. For 107 out of the 371 cases, serum samples were stored at least three years before their type 2 diabetes was diagnosed. In this data set, CB-153 and p,p'-DDE were not associated with an increased risk to develop type 2 diabetes. However, when only the cases (n = 39) that were diagnosed more than six years after the baseline examination and their controls were studied, the women in the highest exposed quartile showed an increased risk to develop type 2 diabetes (OR of 1.6 [95% 0.61, 4.0] for CB-153 and 5.5 [95% CI 1.2, 25] for p,p'-DDE). CONCLUSIONS/SIGNIFICANCE: The results from the present case-control study, including a follow-up design, confirms that p,p'-DDE exposure can be a risk factor for type 2 diabetes.

Tumor necrosis factor-alpha does not mediate diabetes-induced vascular inflammation in mice.

OBJECTIVE: Vascular inflammation is a key feature of both micro- and macrovascular complications in diabetes. Several lines of evidence have implicated the cytokine tumor necrosis factor (TNF) alpha as an important mediator of inflammation in diabetes. In the present study we evaluated the role of TNF alpha in streptozotocin (STZ)-induced diabetes on vascular inflammation in C57BL/6 wild-type and apoE-/- mice. METHODS AND RESULTS: Diabetes increased the expression of vascular cell adhesion molecule (VCAM)-1 in cerebral arteries 150 m in diameter as well as the macrophage accumulation in aortic root atherosclerotic plaques in apoE-/- mice. A more pronounced vascular inflammatory response was observed in diabetic TNF alpha-deficient apoE-/- mice. These mice were also characterized by increased accumulation of IgG and IgM autoantibodies in atherosclerotic lesions. Diabetes also increased VCAM-1 expression and plaque formation in apoE-competent TNF alpha -/- mice, whereas no such effects were observed in C57BL/6 wild-type mice. CONCLUSIONS: The present findings suggest that TNF alpha does not mediate diabetic-induced vascular inflammation in mice and reveal an unexpected protective role for TNF alpha. These effects are partly attributable to a direct antiinflammatory role of TNF alpha, but may also reflect a defective development of the immune system in these mice.

The aldose reductase inhibitor fidarestat suppresses ischemia-reperfusion-induced inflammatory response in rat retina.

Recent studies suggest that increased aldose reductase (AR) activity plays an important role in ischemia-reperfusion injury in the retina. The mechanisms are not completely understood, but may be linked to inflammation. In the present study, we investigated whether the AR inhibitor fidarestat suppressed the retinal inflammatory response induced by ischemia-reperfusion in a rat model. The inflammatory response was manifested by increased gene expression of tumor necrosis factor-alpha and intercellular adhesion molecule-1 (ICAM-1) as well as elevated protein levels of soluble ICAM-1. This response was partially suppressed by the AR inhibitor fidarestat. The findings may reveal beneficial effects of AR inhibition on retinal inflammation associated with ischemia-reperfusion and are in agreement with recent developments in pharmacological research suggesting that pathological conditions other than diabetes may benefit from AR inhibitors.

Inflammatory activity increases with haemoglobin A1c in patients with acute coronary syndrome.

OBJECTIVE: To study the relationship between inflammation, diabetes and HbA(1)c levels in patients with acute coronary syndrome (ACS). DESIGN: Single-centre cross-sectional study comprising 688 consecutive patients with ACS (108 with diabetes) and 341 with stable coronary artery disease (SCAD) (51 with diabetes). High-sensitive C-reactive protein (hsCRP), albumin and fibrinogen concentrations, erythrocyte sedimentation rates (ESR) and leukocyte counts were measured. RESULTS: hsCRP, fibrinogen and ESR levels were higher and albumin lower in ACS patients. ESR was higher, albumin lower and hsCRP borderline significantly higher (p=0.053) in ACS patient with diabetes compared to those without. All inflammatory markers were associated with HbA(1)c in all 688 ACS patients as well as in 540 non-diabetic ACS patients with normal HbA(1)c. In multivariate analyses, all inflammatory markers were independently associated with HbA(1)c in the entire ACS group, regardless of diabetes being present or not. When non-diabetic ACS patients were analyzed separately, only ESR and leukocyte counts were independently correlated with HbA(1)c. CONCLUSIONS: Patients with ACS had increased inflammatory activity, which increased with HbA(1)c levels in patients who neither had a history of diabetes nor HbA(1)c above normal, and was further exaggerated in the presence of diabetes.

Coeliac disease-specific tissue transglutaminase autoantibodies are associated with osteoporosis and related fractures in middle-aged women.

OBJECTIVE: To investigate whether the serological marker for coeliac disease, tissue transglutaminase autoantibody (tTGAb), is associated with decreased bone mass density (BMD) and increased frequency of fractures in middle-aged women screened for osteoporosis. MATERIAL AND METHODS: The study comprised 6480 women (mean age 56 years, range 50-64) who answered a number of questionnaires and who underwent dual X-ray absorptiometry of the wrist bone. Serum samples were analysed for tTGAb using radioligand binding assays. A tTGAb level of >4 U/ml was used to determine a positive value and a level of >17 U/ml was used as an alternative discrimination of high levels. RESULTS: A tTGAb level >4 U/ml was found among 90/6480 (1.4%) women and correlated with lower BMD (multiple linear regression coefficient -382.1; 95% CI = - 673.6-90.7, p=0.011) and with fracture frequency (r=0.18, p=0.023). The 59 women with tTGAb levels >or=17 U/ml had a lower BMD (0.41+/-0.08 g/cm(2) versus 0.44+/-0.08 g/cm(2), p=0.001) and a lower T-score (-1.40+/-1.28 versus -0.90+/-1.40, p=0.003) as well as a higher prevalence of osteoporosis (13.4% versus 6.5%, p=0.008) compared with the remaining 6421 women with tTGAb levels <17 U/ml. Furthermore, fracture frequency was more pronounced in women with tTGAb levels >or=17 U/ml, among whom 19/59 (32.2%) had fractures during the study period compared with 1204/6421 (18.8%) among women with tTGAb levels <17 U/ml (p=0.009). CONCLUSIONS: High levels of tTGAb indicating coeliac disease are associated with lower BMD and higher fracture frequency in women between 50 and 64 years of age. Osteometry is therefore warranted in middle-aged women detected with tTGAb.

Nonhormonal drug use and its relation to androgens in perimenopausal women: a population-based study of Swedish women. The Women's Health in the Lund Area Study.

OBJECTIVE: To outline the prevalence of nonhormonal drug use in middle-aged women and to assess plausible associations between serum androgen levels and variables associated to health such as drug use and planned visits to healthcare units. METHODS: This was a population-based study of women aged 50 to 59 years (n = 6,893). Women were divided into three groups according to their menopause status: premenopausal (PM), postmenopausal without hormone therapy (PM0), and postmenopausal with hormone therapy (PMT). Data regarding current drug use and healthcare visits were collected from a questionnaire. RESULTS: The overall prevalence of nonhormonal drug use was 36.4% in all women. In the PM, PM0, and PMT groups, these percentages were 28.3%, 35.3%, and 39.3%, respectively, and the differences between them were statistically significant (P < 0.01). In all women, the most common medication used was for cardiovascular conditions (12.0%), followed by those for asthma (4.0%) and pain (3.7%). The number of drugs used by all women and women in the PM0 and the PMT groups were negatively associated with the serum levels of androstenedione (P < 0.05). In the postmenopausal groups, the number of visits to healthcare units was negatively associated to the levels of serum testosterone and androstenedione (P < 0.05). CONCLUSIONS: Hormone therapy in postmenopausal women seems to be associated with increased use of nonhormonal pharmacotherapy, rendering higher prevalence of such drugs in middle-aged women. Postmenopausal women with lower serum testosterone and a higher number of office visits used medications for cardiovascular problems and depression more than other medications. Whether this is an effect related to the hormone therapy itself or to experiencing more perimenopausal symptoms in this group of women is still unclear.

Increased retinopathy occurrence in type 1 diabetes patients with increased serum levels of the advanced glycation endproduct hydroimidazolone.

PURPOSE: We aimed to investigate associations between serum levels of the advanced glycation endproduct methylglyoxal-derived hydroimidazolone (MG-H1) and retinopathy in a sample of patients with type 1 diabetes. METHODS: We conducted a cross-sectional study in a Scandinavian ophthalmology outpatient clinic on 61 randomly selected patients with type 1 diabetes. Blood samples and retinal photographs were taken at the same visit. Serum levels of hydroimidazolone immunoreactivity were determined using an immunoassay, and levels of retinopathy were determined from seven standard field stereo photographs of each eye according to the ETDRS method. Results were compared between patients with and without retinopathy. RESULTS: Hydroimidazolone quartiles were significantly associated with retinopathy (p = 0.013). The most profound increase in occurrence of retinopathy was observed from the lowest to the second-lowest hydroimidazolone quartile. Adjusted for duration of diabetes using logistic regression, a significant difference in the presence of retinopathy was found when comparing the lowest quartile with the rest (p = 0.022). CONCLUSIONS: In our patients with type 1 diabetes, serum levels of hydroimidazolone were found to be associated with retinopathy. This is in keeping with findings in a larger sample of patients with type 2 diabetes.