Specific gene expression in type 1 diabetic patients with and without cardiac autonomic neuropathy.

We hypothesized that some molecular pathways might interact to initiate the process of nervous tissue destruction, promoting cardiac autonomic neuropathy (CAN) in the course of diabetes type 1 (T1D). The study group consisted of 60 T1D patients (58.33% women/41.67% men), on standard therapy. The control group consisted of twenty healthy volunteers recruited in accordance with age, gender and body weight. The presence of CAN was documented by the Ewing test method (ProSciCard apparatus). A microarray data analysis was performed using Gene Spring version 13. The microarray results for selected genes were confirmed by real-time PCR (qRT-PCR), using specific TaqMan Gene Expression Assays. Plasma IL-6 content was measured by an enzyme-linked immunosorbent assay (ELISA). The p < 0.05 value was considered as statistically significant. The microarray analysis, confirmed by qRTPCR, showed significant up-regulation of autophagy, quantity of mitochondria, quality regulatory genes (mTOR, GABARAPL2) apoptosis, ER-stress and inflammation (NFKB1, IL1b, IL1R1, SOD1), in T1D when compared to the control group. A significantly higher IL-6 protein level was observed in T1D patients, in comparison to the control group. We concluded that the observed changes in gene expression and activation of intracellular pathways give a coherent picture of the important role of oxidative stress in inflammation and the activation of apoptosis in the pathomechanism of DM. The significance of the inflammatory process, confirmed by the increased level of the inflammation biomarker IL-6 in the pathomechanisms of CAN was shown even in patients with properly treated T1D.

Epigenetic mechanism in search for the pathomechanism of diabetic neuropathy development in diabetes mellitus type 1 (T1DM).

OBJECTIVE: The aim of this study was to check the hypothesis concerning the crucial role of DNA methylation (one of the epigenetic mechanisms) within selected genes related to the destruction and regeneration of neural cells and its input in the pathogenesis of diabetic neuropathy, using a model of the DNA in peripheral blood cells. METHODS: A cross-sectional, case-control study was conducted, consisting of 24 adult Type 1 Diabetes Melitus (T1DM) patients with autonomic neuropathy (CAN), 25 T1DM patients without neuropathy and 25 matched, healthy adults acting as a control (Ctrl). The Ewing's tests, using the ProSciCard apparatus (Mewicon CATEEM-Tec GmbH), was employed to assess the severity of the patients' symptoms of autonomic neuropathy. For DNA methylation analysis, DNA material of each sample DNA after bisulfite conversion was used for the hybridization of BeadChips (Infinium Methylation EPIC Kit, Illumina), and imaged on the Illumina HiScan. The changes in the expression of selected genes were examined using real-time PCR. Probes were labeled using fluorescein amidite, FAM (Thermo Fisher Scientific). Amplification was performed using the continuous fluorescence detection 7900 HT Fast Real-Time PCR system (Thermo Fisher Scientific). The expression ratio of the target mRNA was normalized to the level of 18s RNA and compared with the control. Statistical analysis was performed using Statistica version 13.1. The statistically significant results were recognized, with a value of p < 0.05. RESULTS: Clinical analysis of the investigated groups revealed a significantly higher percentage of personal insulin pump users in the group without neuropathy. The glucose metabolic control, based on the HbA1c level analysis, was also significantly better in T1DM patients without CAN. The Bumphunter method for DNA methylation analysis showed statistically significant regions related to the genes involved in nerve regeneration ninjurin 2 (NINJ2) and functionality (BR serine/threonine kinase 2 BRSK2, claudin 4 CLDN4). When compared with T1DM patients without neuropathy, T1DM patients with neuropathy showed significantly increased methylation in the first NINJ2 axon, and a lower level of DNA methylation in the region of the first intron of BRSK2, as well as the CLDN4 5'UTR regions. The qRT-PCR results confirmed the decreased expression of NINJ2 and CLDN4 genes in patients with T1DM with CAN. CONCLUSIONS: The different DNA methylation profiles, correlating with the expression of genes related to nervous tissue development and regeneration in patients with T1DM with autonomic neuropathy provide evidence for the role of epigenetic mechanisms promoting the development of CAN, a chronic complication of T1DM.

Is treatment of type 1 diabetes mellitus (insulin therapy, metabolic control) optimal for preventing cardiovascular autonomic neuropathy?

INTRODUCTION: Long-term poor metabolic control promotes the occurrence of microvascular complications, such as cardiovascular autonomic neuropathy (CAN) and atherogenic hyperlipidaemia, which translates into increased mortality in patients with type 1 diabetes mellitus (T1DM). The aim of the study was to assess the prevalence of CAN in patients with T1DM in relation to treatment method (continuous subcutaneous insulin infusion, CSII, versus multiple daily injections using pens, MDI) and metabolic control. MATERIAL AND METHODS: The study group comprised 93 adults (60 women, 33 men), mean age 31 years, with T1DM being treated at a local clinical centre from 2011 to 2015. The presence of CAN, the results of laboratory tests, and anthropometric data were analysed. The subjects were divided into two groups according to treatment method (CSII, MDI). RESULTS: The median duration of diabetes was 16 years. 61% of the subjects used MDI and 39% used CSII. 41% of the subjects presented with CAN (confirmed with the Ewing test using ProSciCard apparatus), with a significantly lower prevalence in the group of patients treated with CSII (15.4% vs. 60.4%; p < 0.001). The mean HbA1c level in the CSII-treated group was noticeably lower (7.44 ± 1.67% vs.8.55 ± 1.1%, p < 0.001), and these patients also had lower triglyceride levels (0.71 vs. 1.32 mmol/L, p < 0.001). Regardless of the treatment method, 72% of all patients under 40 years of age achieved their therapeutic target of LDL cholesterol level < 2.6 mmol/L, whereas only 13% of all those over 40 years old achieved an LDL cholesterol level < 1.8 mmol/L. CONCLUSIONS: The presented results draw attention to the high prevalence of CAN among T1DM patients. The study reveals the need for more intensive monitoring and treatment of hyperlipidaemia, despite good glycaemic control, especially in those over the age of 40 years.

[Cardiovascular autonomic neuropathy in the course of diabetes - the review of actual knowledge.] / Neuropatia autonomiczna ukladu sercowo-naczyniowego w przebiegu cukrzycy - aktualny stan wiedzy.

Diabetic neuropathy, including autonomic cardiovascular neuropathy, is the most common chronic complication of diabetes. It causes serious health and social consequences, leading to a significant reduction of life expectancy in DM patients. Its development is initially asymptomatic and therefore often underestimated, and only the early detection of diabetic neuropathy gives a real chance to stop its progression and prevent irreversible damage to the nerves. The optimal glycemic control is the most important factor preventing the development of neuropathy, inhibiting its occurrence and progression. In the advanced stage, however, only symptomatic treatment remains. The article provides an overview of current knowledge about etiopathogenesis, therapy, symptoms and the latest clinical trials on NSN and DM.

[The significance of 75 g oral glucose tolerance test (OGTT) and clinical characteristics of gestational diabetes mellitus patients requiring different therapeutic approaches].

Gestational diabetes mellitus (GDM) constitutes emerging medical problem with incidence rate on the rise all over the world. Thus, it is important to define characteristics of affected individuals. The aim of the study was to analyse the test results of oral load of 75g of glucose as a predictor of need for insulin in the treatment of gestational diabetes and to provide 2nd trimester characteristics of women eventually requiring insulin as compared with those in behavioural approach was sufficient. Material and Methods: We analysed medical records of 203 consecutive women with diagnosis of GDM (mean age 31.4+/- 4.7 years, BMI before pregnancy, 24.5+/- 4.8 kg/m2). The basis for diagnosis of GDM was the result of the OGTT with 75g of glucose, based on the Polish Diabetic Society guidelines (fasting glucose (FPG)> 5.6 mmol/l and/or glucose in 120 'of OGTT> 7.8 mmol/l). We compared patients who required insulin with those treated with diet only. Results: 82 patients (40% of the study group) required implementation of insulin while the other patients remained on diet only. Women requiring insulin therapy reported to the clinic in the earlier gestation's week (p= 0.018) and had higher BMI before pregnancy (p=0.01); also in 75g glucose OGTT obtained significantly higher FPG level (p=0.001) in compare to the diettreated group. Univariate linear regression analysis confirmed a significant, negative correlation between FPG and the week of pregnancy to implement insulin, in the studied group (R=-0.22; p=0.045). Our study showed that the 75g glucose OGTT might have predictive value in choosing insulin treatment in gestational diabetes. We showed that there are differences in the clinical picture between GDM requiring different therapeutic approaches. Our work confirmed also previous reports that higher BMI before pregnancy, an earlier week of diagnosis of the GDM are the risk factors for insulin therapy during pregnancy.