The Transcriptome Analysis Of ATP Inhibit Proliferation Of 3T3-L1 Preadipocytes

Adipose tissue in animal is not only the largest energy storage reservoir, but also an important endocrine organ. It is mainly composed of adipocyte. The hypertrophy and hyperplasia of adipocyte could contribute to obesity, diabetes mellitus type 2 and cardiovascular disease. The mature adipocte is differentiated from preadipocyte. Therefore, the investigation on the mechanism of pre-adipocyte proliferation for is very important for the treatment of metabolic diseases such as obesity. ATP is not only a kind of important energy substance, but also crucial signal molecular to participate in various physiological process. However, its action on proliferation of preadipocytes and the underlying mechanism have not been clarified. To explore above problem, MTT assay and LDH release ratio were used to evaluate the effect of ATP on proliferation and death of 3T3-L1 preadipocytes. Meanwhile, RNA-Seq was used to find out differentially expressed genes (DEGs) between ATP treatment and non-ATP treatment at 6,12,24 and 48 hours and to reveal the molecular mechanisms of ATP affect the 3T3-L1 preadipocytes proliferation in transcriptome level. The main results are as follow:(1) Extracellular ATP inhibited the proliferation of 3T3-L1 preadipocytes and induced intracellular LDH release(2) The results of RNA-Seq displayed that there are 2995,3969,5462,5628 DEGs between ATP and non-ATP treatment cells in different time points(6,12,24 and 48h),respectively. Among these DEGs,1188 genes which co-expressed in all of different time points were specifically expressed in ATP-treatment cells.(3) Short Time-series Expression Miner (STEM) software was applied to analysis the expression profiles of co-expressed DEGs,and four significant profiles (profiles 16,19,17, and 15) were significantly clustered, the four profiles include 391, 367.202,132 genes, respectively.(4) Gene ontology (GO) analysis demonstrate that DEGs were enriched in proliferation and cell death related categories. Especially, DEGs enriched in cell apoptosis and death related categories were clustered in profiles 16 by STEM software, and their expression displayed a linear increase trend.(5) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that the co-expressed DEGs were prominently enriched in cancer related pathway, ECM receptor interaction, MAPK and focal adhesion pathway.Taken together, these results indicate that extracellular ATP can inhibit the proliferation of 3T3-L1 preadipocytes, through upregulation of genes which is involved in apoptosis and cell death.