ORIGINAL ARTICLE Comparative expression analysis of isolated human adipocytes and the human adipose cell lines LiSa-2 and PAZ6 EA van Beek 1 , AH Bakker 2 , PM Kruyt 3 , C Vink 1 , WH Saris 2 , NLW Franssen-van Hal 1 and J Keijer 1 1 RIKILTFInstitute of Food Safety, Wageningen UR, Wageningen, The Netherlands; 2 Maastricht University, Maastricht, The Netherlands and 3 Gelderse Vallei Hospital, Ede, The Netherlands Objective: To obtain insight in the extent to which the human cell lines LiSa-2 and PAZ6 resemble isolated primary human adipocytes. Design: A combination of cDNA subtraction (representative difference analysis; RDA) and cDNA microarray analysis was used to select adipose specific genes to compare isolated (pre-)adipocytes with (un)differentiated LiSa-2 and PAZ6 cells. Measurements: RDA was performed on adipose tissue against lung tissue. A total of 1400 isolated genes were sequenced and cDNA microarray technology was used for further adipose related gene selection. 30 genes that were found to be enriched in adipose tissue were used to compare isolated human adipocytes and LiSa-2 and PAZ6 cells in the differentiated and undifferentiated states. Results: RDA and microarray analysis resulted in the identification of adipose enriched genes, but not in adipose specific genes. Of the 30 most differentially expressed genes, as expected, most were related to lipid metabolism. The second category consisted of methyltransferases, DNMT1, DNMT3a, RNMT and SHMT2, of which the expression was differentiation dependent and higher in differentiated adipocytes. Using the 30 adipose expressed genes, it was found that isolated adipocytes on one hand, and PAZ6 and LiSa-2 adipocytes on the other, differ primarily in lipid metabolism. Furthermore, LiSa-2 cells seem to be more similar to isolated adipocytes than PAZ6 cells. Conclusion: The LiSa-2 cell line is a good model for differentiated adipocytes, although one should keep in mind that the lipid metabolism in these cells deviates from the in vivo situation Furthermore, our results imply that methylation may have an important function in terminal adipocyte differentiation. International Journal of Obesity (2008) 32, 912–921; doi:10.1038/ijo.2008.10; published online 19 February 2008 Keywords: adipose tissue; adipocyte; cDNA microarray; stromal vascular cells; subtractive hybridization; RDA Introduction Adipose tissue is a dispersed tissue that has many functions in the human body. Its primary function is the storage of lipids. Its capacity to expand is essential to maintain healthy levels of circulating lipids in times of abundant food supply and provides a source of energy in times of need. Adipose tissue was shown to be essential for glucose and lipid homeostasis (reviewed in Waki and Tontonoz 1 ), since too little 2,3 as well as too much adipose tissue 4 results in insulin resistance and dyslipidemia. Therefore adipose tissue and adipocytes are subjected to studies to understand the mechanisms of the development of obesity-associated diseases. A widely used approach to study adipocyte physiology is to use cell lines. The most widely used cell lines are 3T3-L1 and its close relatives. 3T3-L1 is a mouse fibroblast cell line that can be differentiated in vitro. Common adipogenic markers like FABP4 and adiponectin were first identified in 3T3F42a 5,6 and the importance of these molecules has been shown in mice and human. Although the use of 3T3-L1 has been of great importance for the understanding of adipose physiology, not all mechanisms identified in 3T3-L1 and its relatives, or for that matter in mice are the same in humans. Therefore for improved extrapolation to humans it would be worthwhile to study human cell lines. However, only few human adipose cell lines exist. One cell line is PAZ6. 7 This is a human brown fat derived adipose cell line that can be Received 5 September 2007; revised 30 November 2007; accepted 10 January 2008; published online 19 February 2008 Correspondence: Dr J Keijer, Food Bioactives, RIKILTFInstitute of Food Safety, PO Box 230, Wageningen 6700AE, The Netherlands. E-mail: jaap.keijer@wur.nl International Journal of Obesity (2008) 32, 912–921 & 2008 Nature Publishing Group All rights reserved 0307-0565/08 $30.00 www.nature.com/ijo