Supplementary Material for: <b><i>Interleukin-6</i></b> CpG Methylation and Body Weight Correlate Differently in Type 2 Diabetes Patients Compared to Obese and Lean Controls

<b><i>Background/Aims:</i></b> Diabetes mellitus type 2 (DMT2) is accompanied by systemic low-grade inflammation with elevated levels of interleukin-6 (IL-6), which is encoded by a gene <i>(IL-6)</i> previously shown to be regulated by DNA methylation. We investigated seven CpG sites in <i>IL-6</i> in individuals with DMT2, obese individuals and lean controls. Further, the DMT2 group received the glucagon-like peptide 1 agonist liraglutide. <b><i>Methods:</i></b> Blood samples were taken at the beginning of the study and after 4 months. The DNA methylation was assessed using pyrosequencing. <b><i>Results:</i></b> Methylation levels at the CpG sites -664, -628 and +13 at the first sampling time point (T1) and at -666 and -664 at the second sampling time point (T2) correlated negatively with initial body weight in the DMT2 group. We found positive correlations for the obese and the lean control group. In the obese group, CpG +27 methylation at T1 correlated with initial body weight (r = 0.685; p = 0.014). In the lean group, CpG -664 at T1 (r = 0.874; p = 0.005) and CpG -628 at T2 (r = 0.632; p = 0.050) correlated with initial body weight. <b><i>Conclusion:</i></b> These findings are an informative basis for further studies to elucidate epigenetic mechanisms underlying DMT2. Additionally, our results might provide starting points for the development of biomarkers for prevention and therapy strategies

Modulation of hyperglycemia and TNF[alfa]-mediated inflammation by helichrysum and grapefruit extracts in diabetic db/db mice

Type-2 diabetes is associated with a chronic low-grade systemic inflammation accompanied by an increased production of adipokines/cytokines by obese adipose tissue. The search for new antidiabetic drugs with different mechanisms of action, such as insulin sensitizers, insulin secretagogues and aglucosidase inhibitors, has directed the focus on the potential use of flavonoids in the management of type-2 diabetes. Thirty six diabetic male C57BL/6J db/db mice were fed a standard diet and randomly assigned into four experimental groups: non-treated control, (n ¼ 8); acarbose (5 mg per kg bw, n ¼ 8); helichrysum (1 g per kg bw, n ¼ 10) and grapefruit (0.5 g per kg bw, n ¼ 10) for 6 weeks. The mRNA expression in pancreas, liver and epididymal adipose tissue was determined by RT-PCR. DNA methylation was quantified in epididymal fat using pyrosequencing. Mice supplemented with helichrysum and grapefruit extracts showed a significant decrease in fasting glucose levels (p < 0.05). A possible mechanism of action could be the up-regulation of liver glucokinase (p < 0.05). The antihyperglycemic effect of both extracts was accompanied by decreased mRNA expression of some proinflammatory genes (monocyte chemotactic protein-1, tumor necrosis factor-a, cyclooxygenase-2, nuclear factorkappaB) in the liver and epididymal adipose tissue. The CpG3 site of TNFa, located 5 bp downstream of the transcription start site, showed increased DNA methylation in the grapefruit group compared with the non-treated group (p < 0.01). In conclusion, helichrysum and grapefruit extracts improved hyperglycemia through the regulation of glucose metabolism in the liver and reduction of the expression of proinflammatory genes in the liver and visceral fat. The hypermethylation of TNFa in adipose tissue may contribute to reduce the inflammation associated with diabetes and obesity

Modulation of hyperglycemia and TNF[alfa]-mediated inflammation by helichrysum and grapefruit extracts in diabetic db/db mice

Type-2 diabetes is associated with a chronic low-grade systemic inflammation accompanied by an increased production of adipokines/cytokines by obese adipose tissue. The search for new antidiabetic drugs with different mechanisms of action, such as insulin sensitizers, insulin secretagogues and aglucosidase inhibitors, has directed the focus on the potential use of flavonoids in the management of type-2 diabetes. Thirty six diabetic male C57BL/6J db/db mice were fed a standard diet and randomly assigned into four experimental groups: non-treated control, (n ¼ 8); acarbose (5 mg per kg bw, n ¼ 8); helichrysum (1 g per kg bw, n ¼ 10) and grapefruit (0.5 g per kg bw, n ¼ 10) for 6 weeks. The mRNA expression in pancreas, liver and epididymal adipose tissue was determined by RT-PCR. DNA methylation was quantified in epididymal fat using pyrosequencing. Mice supplemented with helichrysum and grapefruit extracts showed a significant decrease in fasting glucose levels (p < 0.05). A possible mechanism of action could be the up-regulation of liver glucokinase (p < 0.05). The antihyperglycemic effect of both extracts was accompanied by decreased mRNA expression of some proinflammatory genes (monocyte chemotactic protein-1, tumor necrosis factor-a, cyclooxygenase-2, nuclear factorkappaB) in the liver and epididymal adipose tissue. The CpG3 site of TNFa, located 5 bp downstream of the transcription start site, showed increased DNA methylation in the grapefruit group compared with the non-treated group (p < 0.01). In conclusion, helichrysum and grapefruit extracts improved hyperglycemia through the regulation of glucose metabolism in the liver and reduction of the expression of proinflammatory genes in the liver and visceral fat. The hypermethylation of TNFa in adipose tissue may contribute to reduce the inflammation associated with diabetes and obesity