Preadipocytes (3T3 L1) were used between 7 and 14 days after differentiation; they were incubated with 44 nM 3H-esterone. The medium was supplemented with 1 μM recombinant murine leptin, 10 nM recombinant human insulin, or 1 μM corticosterone for up to 72 hr. In a second series of experiments, cells were incubated for 48 hr with different concentrations of leptin, insulin or corticosterone, and compared with controls (plain medium). Cells were harvested, washed in buffer and homogenized, and protein was measured. Lipid extracts of cell homogenates were used for HPLC; the label distribution in free and acyl-estrone peaks was measured. Overall uptake of estrone (i.e., the sum of free and acyl-estrone) by cells was not affected by leptin or corticosterone, but strongly reduced by insulin. Leptin and corticosterone increased the synthesis of acyl-esterone in a dose- and time-dependent way. Insulin decreased acyl-estrone synthesis at low concentrations and with little change over time. The results suggest that control of oleoyl-estrone deposition in adipocytes is modulated in at least two distinct steps: (a) estrone uptake, affected by insulin in the physiological range; and (b) synthesis of oleoyl-estrone from cell estrone. The latter may be affected by insulin, but leptin and corticosterone enhance the process.

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