Supplemental Table 1. Sequences of the primers and TaqMan® probes
Gene (GenBank accession) / Oligonucleotide sequence (5’-3’)LCN2 (NM_005564)
Forward / CCCAGCCCCACCTCTGA
Reverse / CTTCCCCTGGAATTGGTTGTC
TaqMan® Probe / FAM-CAAGGTCCCTCTGCAGCAGAACTTCCA-TAMRA
RARRES2 (NM_002889)
Forward / GGAAGAAACCCGAGTGCAAAG
Reverse / CGCAGAACTTGGGTCTCTATGG
TaqMan® Probe / FAM-AGGCCCAATGGGAGGAAACGGAAA-TAMRA
SPP1 (NM_000582)
Forward / CATCCAGTACCCTGATGCTACAGA
Reverse / GGCCTTGTATGCACCATTCAA
TaqMan® Probe / FAM-ACATCACCTCACACATGGAAAGCGAGGA-TAMRA
YKL40 (NM_001276)
Forward / GGACGGAGAGACAAACAGCATT
Reverse / CCTGGCTGGGCTTCCTTTAT
TaqMan® Probe / FAM-CCACCCTAATCAAGGAAATGAAGGCC-TAMRA
LCN2, lipocalin 2; RARRES2, chemerin; SPP1, osteopontin; YKL40, chitinase 3-like 1.
Supplemental Table 2. Effects of weight loss in obese patients after Roux-en-Y gastric bypass (RYGB).
Before WL / After WLn / 36 / 36
Age (years) / 42 2 / 43 3
Weight (kg) / 120.3 3.7 / 80.1 2.3***
BMI (kg/m2) / 44.7 1.1 / 31.8 0.7***
Body fat (%) / 53.1 0.9 / 36.7 1.4***
Waist (cm) / 124.4 2.6 / 95.5 1.9***
Hip (cm) / 132.5 2.4 / 106.7 1.7***
WHR / 0.94 0.01 / 0.89 0.01***
SBP (mm Hg) / 130 6 / 126 16
DBP (mm Hg) / 86 2 / 82 8
Fasting glucose (mg/dL) / 118 13 / 91 4*
Fasting insulin (U/mL) / 22.5 5.9 / 12.7 5.8***
HOMA / 7.7 3.2 / 4.4 2.6**
QUICKI / 0.301 0.011 / 0.339 0.015***
Triacylglycerols (mg/dL) / 144 11 / 90 7**
Total cholesterol (mg/dL) / 214 6 / 165 6***
LDL-cholesterol (mg/dL) / 139 5 / 93 5***
HDL-cholesterol (mg/dL) / 44 2 / 53 2**
Uric acid / 5.65 0.27 / 4.17 0.20***
Leptin (ng/mL) / 58.7 6.3 / 24.9 4.4***
CRP (mg/L) / 7.8 0.21 / 3.8 0.11
Fibrinogen (mg/dL) / 371 15 / 322 12**
Homocysteine (mol/L) / 7.9 0.6 / 8.7 0.9
AST (UI/L) / 19.8 2.3 / 17.9 1.8
ALT (UI/L) / 30.3 3.6 / 24.1 3.3
ALP (UI/L) / 119.4 7.4 / 112 6.7
-GT (UI/L) / 37.1 6.9 / 18.4 3.3***
ALP, alkaline phosphatase; ALT, alanine aminotransferase; AST, aspartate aminotransferase; BMI, body mass index; CRP, C-reactive protein; DBP, diastolic blood pressure; -GT, -glutamyltransferase; HOMA, homeostasis model assessment; QUICKI, quantitative insulin sensitivity check index; SBP, systolic blood pressure; WHR, waist-to-hip ratio; WL, weight loss. Data are mean SEM. Differences between groups were analyzed by two-tailed paired t-tests. *P<0.05, **P<0.01 and *** P<0.0001 vs before WL.
Legends for supplemental figures
Supplemental Figure 1.Tissue distribution ofWNT5A and SFRP5. Comparison of mRNA levels of (A) WNT5A and (B) SFRP5 in visceral (n=69) and subcutaneous (n=37) adipose tissue, peripheral mononuclear blood cells (PMBC) (n=66) and liver biopsies (n=60). Bars represent the mean SEM of the ratio between the gene expressionto 18S rRNA. Differences between groups were analyzed by one-way ANOVA. The expression level in VAT was assumed to be 1. Differences between groups were analyzed by one-way ANOVA followed by Tukey’s tests. *P0.05 vs VAT and ††P<0.01 vs PMBC.
Supplemental Figure 2.Gene expression levels of receptor-like tyrosine kinase (ROR)-1 (A) and ROR2 (B) in VAT of LN, obese NG and obese T2D volunteers. Bars represent the mean SEM of the ratio between the gene expression to 18S rRNA. The expression level in LN subjects was assumed to be 1 (LN: n=9; OB-NG: n=32; OB-T2D: n=28). Differences between groups were analyzed by one-way ANOVA followed by Tukey’s tests. *P<0.05 vs LN.
Supplemental Figure 3.Effect of SFRP5 (100 ng/mL) for 24 h on basal and WNT5A (100 ng/mL)-induced gene expression of inflammatory genes. Gene expression levels of (A) interleukin 6 (IL6), (B) interleukin 1 (IL1B), (C) tumor necrosis factor- (TNFA), (D) monocyte chemotactic protein 2 (CCL2), (E) matrix metalloproteinase 2 (MMP2), (F) matrix metalloproteinase 9 (MMP9), (G) lipocalin 2 (NGAL) and (H) osteopontin (SPP1) in human differentiated visceral adipocytes. Gene expression levels in unstimulated cells were assumed to be 1. Values are the mean SEM (n=10 per group). Differences between groups were analyzed by one-way ANOVA followed by Tukey’s tests. *P<0.05 vs unstimulated and †P<0.05 vs WNT5A-stimulated cells.
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