Supplementary figure 1

Supplementary figure 1. Association between the levels of circulating inflammatory proteome and diabetes complications in a cohort matched for diabetes duration, HbA1c levels, and insulin need</p

Supplementary table 1-6

Supplementary tables 1-6. Details on the statistical analyses performed on the relation between diabetes duration, insulin need, HbA1c levels, and diabetes complications with circulating immune cells (Supp table 1 & 2), PBMCs ex vivo stimulation (Supp table 3 & 4), and circulating inflammatory proteome (Supp table 5 & 6)</p

TLR-3 is functionally active in adipocytes.

<p>(a) Differentiated SGBS adipocytes were stimulated with either a TLR-3 (poly:IC 12.5μg/ml) or TLR-4 (LPS 50ng/ml) agonist. mRNA levels were measured for IL-8, MCP-1, IL-1 β, adiponectin and PPAR-γ. (b-f) SGBS adipocytes were treated with SiRNA against TLR-3 or scr SiRNA and stimulated with poly:IC. mRNA levels of (b) IL-8, (c) MCP-1, (d) IL-1β, (e) adiponectin, (f) PPAR- γ were subsequently measured. * p<0.05, ** p<0.01, *** p<0.001. Data are shown as means ± SEM.</p

Additional file 1 of The impact of prior exposure to hypoglycaemia on the inflammatory response to a subsequent hypoglycaemic episode

Additional file 1. Supplement 1: in-and exclusions criteria

TLR-3 is functionally active in adipocytes.

<p>(a) Differentiated SGBS adipocytes were stimulated with either a TLR-3 (poly:IC 12.5μg/ml) or TLR-4 (LPS 50ng/ml) agonist. mRNA levels were measured for IL-8, MCP-1, IL-1 β, adiponectin and PPAR-γ. (b-f) SGBS adipocytes were treated with SiRNA against TLR-3 or scr SiRNA and stimulated with poly:IC. mRNA levels of (b) IL-8, (c) MCP-1, (d) IL-1β, (e) adiponectin, (f) PPAR- γ were subsequently measured. * p<0.05, ** p<0.01, *** p<0.001. Data are shown as means ± SEM.</p

TLR-3 deficiency does not protect mice against metabolic abnormalities.

<p>Wild-type (WT) and TLR-3-/- mice were subjected to 16 weeks of low fat diet (LFD) or high fat diet (HFD). (a) development of the bodyweight, (b) liver weight, (c) epididymal adipose tissue weight, (d) plasma leptin levels, (e) fasting glucose levels, (f) insulin tolerance test (ITT), (g) area under the curve for ITT. * p<0.05, *** p<0.001. Number of mice per group: WT-LFD n = 10; WT-HFD n = 10; TLR-3-/-LFD n = 7; TLR-3-/-HFD n = 9. Data are shown as means ± SEM.</p

TLR-3 deficiency does not ameliorate adipose tissue inflammation.

<p>After 16 weeks of low fat diet (LFD) or high fat diet (HFD) intervention, adipose tissue of wild-type (WT) and TLR-3-/- mice was investigated for inflammatory parameters. (a) Adipose tissue of mice stained for F4/80, magnification 20x, (b) number of crown-like structures in adipose tissue. Inflammatory markers were measured (c) F480, (d) CD68, (e) MCP-1, (f) TNFα, (g) CXCL1. (h) mRNA levels of TLR-3 in WT mice fed a LFD or HFD for 16 weeks. * p<0.05, ** p<0.01. Number of mice per group: WT-LFD n = 10; WT-HFD n = 10; TLR-3-/-LFD n = 7; TLR-3-/-HFD n = 9. Data are shown as means ± SEM.</p

TLR-3 is predominantly expressed in adipocytes.

<p>(a) Biopsies from visceral- (VAT) and subcutaneous adipose tissue (SAT) were obtained from 4 healthy subjects and TLR expression was determined in stromal vascular fraction (SVF) and mature adipocytes (MA). mRNA levels of (b) TLR-3 and (c) PPAR-γ were measured during differentiation of human SGBS adipocytes. * p<0.05, ** p<0.01. Data are shown as means ± SEM.</p

TLR-3 in human adipose tissue.

<p>Subcutaneous adipose tissue samples of 80 healthy individuals were obtained. TLR-3 mRNA levels were associated with (a) BMI, (b) HOMA-IR, (c) plasma CRP-levels, (d) number of crown-like structures in adipose tissue. Association of MAP3K8 mRNA expression in human subcutaneous adipose tissue with mRNA expression of (e) IL-8, (f) MCP-1, (g) FABP4 and (h) adipocytes cell size. * p<0.05. Data are shown as means ± SEM. HOMA-IR = Homeostatic Model Assessment for insulin resistance.</p

Inflammatory profile of the adipose tissue of HFD-fed WT and MAP3K8-ko animals.

<p>MAP3K8-ko and WT mice were fed a LFD or HFD during 16 weeks. (a–f) qPCR analysis for cytokines (a) TNF-α, (b) IFNγ, (c) IL-1β, (d) CXCL-1, (e) IL-6 and (f) IL-1Ra. n = 9 mice per group. Relative phosphorylation of NFκB p65 (g) and ERK 1/2 (h) in eWAT of MAP3K8-ko and WT animals after HFD-feeding (i). * p<0.05, ** p<0.01, *** p<0.001.</p