Absence of an adipogenic effect of rosiglitazone on mature 3T3-L1 adipocytes: increase of lipid catabolism and reduction of adipokine expression

Abstract

Aims/hypothesis: The thiazolidinedione (TZD) rosiglitazone is a peroxisome proliferator-activated receptor-gamma agonist that induces adipocyte differentiation and, hence, lipid accumulation. This is in apparent contrast to the long-term glucose-lowering, insulin-sensitising effect of rosiglitazone. We tested whether the action of rosiglitazone involves specific effects on mature adipocytes, which are different from those on preadipocytes.

Materials and methods: Differentiated mature 3T3-L1 adipocytes were used as an in vitro model. Transcriptomics, proteomics and assays of metabolism were applied to assess the effect of rosiglitazone in different insulin and glucose conditions.

Results: Rosiglitazone does not induce an increase, but rather a decrease in the lipid content of mature adipocytes. Analysis of transcriptome data, confirmed by quantitative RT-PCR and measurements of lipolysis, indicates that an altered energy metabolism may underlie this change. The pathway analysis shows a consistent picture dominated by lipid catabolism. In addition, we confirmed at both mRNA level and protein level that rosiglitazone represses adipokine expression and production, except for genes encoding adiponectin and apolipoprotein E. Moreover, transcriptome changes indicate that a general repression of genes encoding secreted proteins occurs.

Conclusions/interpretation: Our findings suggest that the change of adiposity as seen in vivo reflects a shift in balance between the different effects of TZDs on preadipocytes and on mature adipocytes, while the changes in circulating adipokine levels primarily result from an effect on mature adipocytes.

Fig. 1

Oil Red O staining (ORO) for lipid content in 3T3-L1 adipocytes. Data are expressed as mean±SD. n = 5 for high glucose and insulin (control condition) (IG) and rosiglitazone with high insulin and glucose condition (RosiIG); n = 3 for basal and rosiglitazone with low insulin and glucose condition (Rosi). Blank bars: absorbance, relative value; filled bars: absorbance, relative value corrected for cell numbers. ** p < 0.01 compared with IG condition before cell number correction (t test), *** p < 0.001 compared with IG condition after cell number correction (t test)

Fig. 2

Regulated canonical pathways by rosiglitazone. Microarray data analysed by Ingenuity Pathway Analysis. Blank bars, with low insulin/glucose; filled bars, with high insulin/glucose. Bold line, threshold

Fig. 3

Regulation pattern by rosiglitazone of genes encoding extracellular proteins and of genes encoding intracellular proteins or proteins of unknown location. a, c: extracellular pattern; b, d: intracellular pattern. a, b: rosiglitazone with low insulin/glucose; c, d: rosiglitazone with high insulin/glucose

Fig. 4

The effect of rosiglitazone on the secretion of complement C3 as measured by 1D gel analysis. The marked bands (arrows) were identified as C3. One representative image is shown. A sample treated by Brefeldin A (+BFA) was used to verify secretion

Fig. 5

The effect of rosiglitazone on the secretion of adipsin as measured by 2D gel analysis. A section of one representative 2D gel image is shown for an individual adipsin spot (squares) from a low glucose and insulin control condition (Basal), b rosiglitazone with low insulin and glucose condition (Rosi), c high glucose and insulin control condition (IG) and d rosiglitazone with high insulin and glucose condition (RosiIG). e Total quantity of adipsin in the culture medium of 3T3-L1 adipocytes expressed in arbitrary units (AU)

Fig. 6

The effect of rosiglitazone on the secretion of adiponectin as measured by western blotting. Data are expressed as mean ± SD (n = 3) and one representative western blot image is shown. *p < 0.05 compared with Basal condition (t test); **p < 0.01 compared with IG condition (t test). Basal, low glucose and insulin control condition; Rosi, rosiglitazone with low insulin and glucose condition; IG, high glucose and insulin control condition; RosiIG, rosiglitazone with high insulin and glucose condition

Fig. 7

The effect of rosiglitazone on the culture medium glycerol levels. Data are expressed as mean±SD (n = 3). *p < 0.05 compared with IG condition (t test); **p < 0.01 compared with Basal condition (t test). Basal, low glucose and insulin control condition; Rosi, rosiglitazone with low insulin and glucose condition; IG, high glucose and insulin control condition; RosiIG, rosiglitazone with high insulin and glucose condition