Objective: To evaluate the effects of progesterone on the mRNA expression of acylation stimulating protein (ASP)-receptor C5L2 in adipocytes and preadipocytes and the C5L2 protein expression on the cell surface.
Methods: Preadipocytes of the line 3T3-L1 were cultured and induced to differentiate. Progesterone of the doses 0 - 1 x 10(-6) mol/L was added into the cultured fluid of the mature 3T3-L1 adipocytes and preadipocytes overnight. RT-PCR and flow cytometry were used to detect the mRNA and protein expression of ASP receptor C5L2. Both non-progesterone treated and progesterone-treated 3T3-L1 cells were cultured with 5.0 micromol/L ASP for 4 hours, then the cell protein was extracted and the expressions of G protein (including Galphaq/11 and Gbeta) and phosphated protein kinase C (including p-PKCalpha and p-PKCzeta) were measured by Western blotting.
Results: The C5L2 protein expression level of the mature adipocytes stimulated by progesterone 1 x 10(-6) mol/L for 18 h was 36% +/- 15%, significantly downregulated compared with that of the adipocytes stimulated by progesterone 0 mol/L (46% +/- 12%, P < 0.01), with a inhibition rate of 22%. The C5L2 mRNA and protein expression levels of the preadipocytes stimulated by progesterone 1 x 10(-6) mol/L for 18 h were 0.17 +/- 0.11 and 36% +/- 16% respectively, both significantly lower than those of the preadipocytes stimulated by progesterone 0 mol/L (0.50 +/- 0.18 and 51% +/- 20% respectively, P < 0.01 and P < 0.05) with the inhibition rates of 66% and 29%respectively. The ASP-stimulated Galphaq/11, Gbeta, p-PKCalpha, and p-PKCzeta expression levels of the mature adipocytes after overnight exposure to progesterone 1 x 10(-8) and 1 x 10(-6) mol/L were suppressed dose-dependently. For example, the ASP-stimulated Galphaq/11, Gbeta, and p-PKCalpha expression levels of the progesterone 1 x 10(-6) mol/L group were significantly lower than those of the progesterone 0 mol/L group by 41%, 63%, and 49% respectively (P < 0.05 to P < 0.01. In the preadipocytes the reduction of ASP-induced Galphaq/11, Gbeta, and p-PKCzeta expression levels were observed at the concentration of progesterone as low as 1 x 10(-8) mol/L, and all the four proteins were inhibited significantly at the 1 x 10(-6) mol/L progesterone concentration.
Conclusion: Progesterone induces ASP resistance in adipocytes and preadipocytes. ASP resistance may contribute to the physiological abnormalities associated with insulin resistance induced by progesterone.