Abstract
Mature adipocytes originate from fibroblast-like precursor cells, preadipocytes, which differentiate to obtain the characteristics of adipocytes. Our aim was to investigate how differentiation of human preadipocytes affects the distribution of insulin receptors (IR) and IGF-I receptors (IGF-IR) and other cell characteristics. Preadipocytes were differentiated using indomethacine, dexamethasone, isobutyl-methylxantine (IBMX) and high concentration of insulin. Gene expression was quantified by real-time RT-PCR in preadipocytes (PA), differentiated preadipocytes (dPA) and mature adipocytes (mAD). The amount of expressed receptor protein was analyzed using receptor specific ELISAs and Western blot. We also studied DNA synthesis with radiolabeled thymidine incorporation and glucose accumulation with radiolabeled glucose. Differentiation of PA increased gene expression of IR but not IGF-IR. GLUT4, growth hormone receptor (GHR) and adiponectin appeared or increased. In PA and dPA only IR-A was expressed whereas also IR-B was detected in mAD. By Western blot and ELISA, IR and IGF-IR were detected in PA, dPA and mAD. During differentiation the ratio of IR to IGF-IR increased severalfold. In PA both the IR and the IGF-IR was phosphorylated by their own ligand at 1 nM and in dPA the activation of both receptors was stimulated by IGF-I, but not insulin, at 1 nM. Accumulation of glucose in PA was increased by insulin at 10nM and by IGF-I at 1 nM and 10nM. DNA synthesis was increased by insulin and IGF-I at 10nM. In conclusion, both IR and IGF-IR are present in human preadipocytes and adipocytes. Differentiation is characterized by an increased IR/IGF-IR ratio.