Association of adipocyte genes with ASP expression: a microarray analysis of subcutaneous and omental adipose tissue in morbidly obese subjects

Abstract

Background: Prevalence of obesity is increasing to pandemic proportions. However, obese subjects differ in insulin resistance, adipokine production and co-morbidities. Based on fasting plasma analysis, obese subjects were grouped as Low Acylation Stimulating protein (ASP) and Triglyceride (TG) (LAT) vs High ASP and TG (HAT). Subcutaneous (SC) and omental (OM) adipose tissues (n = 21) were analysed by microarray, and biologic pathways in lipid metabolism and inflammation were specifically examined.

Methods: LAT and HAT groups were matched in age, obesity, insulin, and glucose, and had similar expression of insulin-related genes (InsR, IRS-1). ASP related genes tended to be increased in the HAT group and were correlated (factor B, adipsin, complement C3, p < 0.01 each). Differences between LAT and HAT group were almost exclusively in SC tissue, with little difference in OM tissue. Increased C5L2 (p < 0.01), an ASP receptor, in HAT suggests a compensatory ASP pathway, associated with increased TG storage.

Results: HAT adipose tissue demonstrated increased lipid related genes for storage (CD36, DGAT1, DGAT2, SCD1, FASN, and LPL), lipolysis (HSL, CES1, perilipin), fatty acid binding proteins (FABP1, FABP3) and adipocyte differentiation markers (CEBPalpha, CEBPbeta, PPARgamma). By contrast, oxidation related genes were decreased (AMPK, UCP1, CPT1, FABP7). HAT subjects had increased anti-inflammatory genes TGFB1, TIMP1, TIMP3, and TIMP4 while proinflammatory PIG7 and MMP2 were also significantly increased; all genes, p < 0.025.

Conclusion: Taken together, the profile of C5L2 receptor, ASP gene expression and metabolic factors in adipose tissue from morbidly obese HAT subjects suggests a compensatory response associated with the increased plasma ASP and TG.

Figure 1

ASP generation related genes tend to be increased in HAT group. (A) Expression of genes related to ASP generation in SC adipose tissue from LAT (hatched bars) and HAT (solid bars) as assessed by microarray where LAT expression is set at 100%. Correlations are shown between (B) C3 and factor B (FB), (C) FB and adipsin (Factor D), and (D) C3 and adipsin in SC adipose tissue from both LAT and HAT subjects. Data is expressed as means ± SEM where * p < 0.025, ** p < 0.01, *** p < 0.001, where R values represent Spearman coefficients, and lines are based on linear regression.

Figure 2

C5L2 expression is increased in HAT group while similar immune receptors are unaltered. (A) C5L2 gene expression in SC and OM adipose tissue from control non-obese (NO), LAT, and HAT subjects as assessed by ΔΔ real time RT-PCR for C5L2 relative to the housekeeping gene B2M. 2-way ANOVA indicates significant differences (p < 0.0001) with a difference between groups (p < 0.0001) but no significant difference between tissues (pNS). Post-hoc test indicates that HAT was significantly increased vs LAT in SC (p < 0.0.01) and OM (p < 0.001), and HAT was significantly increased vs NO (non-obese) in both SC (p < 0.05) and OM ((p < 0.001) where * p < 0.05, ** p < 0.01 and *** p < 0.001 for HAT vs LAT and + p < 0.05, ++ p < 0.01 and +++ p < 0.001 for HAT vs NO. There were no significant differences between LAT and NO. (B) Gene expression of immune receptor genes related to C5L2 in SC adipose tissue from LAT (hatched bars) and HAT (solid bars) as assessed by microarray where LAT expression is set at 100%. No significant differences between the groups were found for any of the related receptors. Data is expressed as means ± SEM.

Figure 3

Intracellular lipid synthesis genes in HAT vs LAT in SC adipose tissue. Gene expression of intracellular lipid synthesis genes (A and B) in SC adipose tissue from LAT (hatched bars) and HAT (solid bars) as assessed by microarray where LAT expression is set at 100%. Correlation in SC adipose tissue from both LAT and HAT subjects between the ASP triad of genes (C3, factor B and adipsin) and (C) DB1 presented for C3 (open circles), FB (solid circles) and adipsin (stars). R values for C3, FB, and FD respectively are (C) 0.548, 0.639, 0.509. (D) Correlations between DGAT2 and GLUT4, R = -0.891. Correlations between GLUT4 and DGAT2 and (E) SCD1 and (F) DGAT1 shown as GLUT4 (solid circles) and DGAT2 (stars). R values (Spearman coefficients) for GLUT4 and DGAT2 respectively are (E) -0.952, 0.903 and (F) -0.964, 0.952. Lines are based on linear regression. Data is expressed as means ± SEM where * p < 0.025, ** p < 0.01 and *** p < 0.001.

Figure 4

Expression of Lipolytic genes in LAT vs HAT. Gene expression of lipolytic genes (A) LPL and LIPC, (B) PNLIP and EL and (E) CES1, MGLL, HSL, perilipin and (F) adiponutrin in SC adipose tissue from LAT (hatched bars) and HAT (solid bars) as assessed by microarray where LAT expression is set at 100%. Correlation between the ASP triad of genes and (C) LPL and (D) LIPC are shown for C3 (open circles), FB (solid circles), and adipsin (stars). R values (Spearman coefficients) for C3, FB, and adipsin respectively are (C) 0.430, 0.600, 0.564 and (D) 0.673, 0.842, 0.709. Correlations between lipid metabolism genes GLUT4 and DGAT2 with (G) HSL and (H) perilipin are shown for GLUT4 (solid circles) and DGAT2 (stars). R values (Spearman coefficients) for GLUT4 and DGAT2 respectively are (G) -0.952, 0.915 and (H) -0.564, 0.515. Lines are based on linear regression. Data is expressed as means ± SEM where * p < 0.025, ** p < 0.01 and *** p < 0.001.

Figure 5

Expression of CEBPs and PPARs in LAT vs HAT. Gene expression of differentiation related gene families, CEBPs and PPARs (A and B) in SC adipose tissue from LAT (hatched bars) and HAT (solid bars) as assessed by microarray where LAT expression is set at 100%. Correlations between the ASP triad and CEBPγ are shown in (C) with R values (Spearman coefficients) for C3, FB, and adipsin are 0.527, 0.709, and 0.698 respectively. Correlations for CEBPα (D) and PPARδ (E) with GLUT4 and DGAT2 respectively are (D) -0.915, 0.830 and (E) 0.758, -0.673. Lines are based on linear regression. Data is expressed as means ± SEM where * p < 0.025, ** p < 0.01 and *** p < 0.001.