ChREBP is an essential transcription factor for lipogenesis. Its physiological role in adipose tissue has been studied only to a small extent and the control of its expression remains unknown in human adipocytes. We have studied ChREBP mRNA and protein expression levels in the liver and the omental (OM) and subcutaneous (SC) adipose tissues from obese and lean subjects, as well as in human differentiated preadipocytes. Liver and OM and SC adipose tissue biopsies were obtained from lean and obese patients. Human preadipocytes were isolated from the adipose tissues from obese patients and differentiated under adipogenic conditions. ChREBP expression levels were quantified by RT-PCR and Western blot analysis. We found opposing results in terms of ChREBP regulation in the liver and adipose samples. ChREBP increased in the liver from obese compared to lean subjects, whereas the expression decreased in both adipose tissues. The mRNAs of other adipogenic markers were checked in these tissues. The pattern of FASN was similar to the one for ChREBP, ADCY3 decreased in both adipose tissues from obese patients, AP2 decreased only in OM adipose tissue of obese patients and ATGL did not change. The levels of ChREBP mRNA and protein showed dramatic increases during the differentiation of human OM and SC preadipocytes. In conclusion, ChREBP expression has an opposite regulation in the liver and adipose tissue from obese subjects which is compatible with the increased hepatic lipogenesis and decreased adipocytic lipogenesis found in these patients. The dramatic increase of ChREBP mRNA and protein levels during preadipocyte differentiation suggests a role in adipogenesis.

Housekeeping genes frequently used in gene expression studies are highly regulated in human adipose tissue. To ensure a correct interpretation of results, it is critical to select appropriate reference genes. Subcutaneous (SC) and omental (OM) adipose tissue expression was analyzed from lean and obese subjects using whole genome complementary DNA (cDNA) microarrays to identify stably expressed genes and commercial TaqMan low density arrays (LDAs), with 16 common control genes. The best candidate gene from microarrays analysis was F‐box and leucine‐rich repeat protein‐10 (FBXL10) (fold‐change 10−3 P < 0.01), an ubiquitous nucleolar protein evolutionarily conserved. Hypoxanthine phosphoribosyltransferase 1 (HPRT1) and importin 8 (IPO8), were the best reference genes among the 16 genes in the LDAs with coefficient of variation (CV) of 4.51 and 4.55%, respectively. However, when the LDAs data were further analyzed by the geNorm and NormFinder softwares, IPO8, a nuclear protein mediating import of proteins, was the first and the third better reference gene, respectively. IPO8 and FBXL10 were further validated by real‐time PCR in additional OM and SC fat samples and primary cultured preadipocytes. According to their CV, IPO8 resulted more suitable than FBXL10 in both adipose tissue depots and SC preadipocytes, whereas FBXL10 performed better than IPO8 in OM cultured preadipocytes. Both genes expression levels did not change throughout adipogenesis. Thus, we provide clear evidence that IPO8 and FBXL10 are good candidates to use as reference genes in gene expression studies in human OM and SC adipose tissues as well as differentiated primary preadipocytes.

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