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Rapamycin inhibits human adipocyte differentiation in primary culture
1Department of Medicine, Loeb Health Research Institute, Ottawa Hospital, University of Ottawa, Ontario, Canada.
Objective:
The immunosuppressant drug rapamycin, has been reported to inhibit 3T3-L1 adipocyte differentiation by interfering with critical postconfluent mitoses that are required early on for successful differentiation of this cell line (clonal expansion phase). In contrast to the murine 3T3-L1 preadipocyte cell line, human preadipocytes in primary culture do not undergo clonal expansion during differentiation. We investigated whether rapamycin could inhibit human adipocyte differentiation.
Research Methods And Procedures:
The effect of rapamycin on the induction of differentiation of human preadipocytes in primary culture into adipocytes was measured using Oil Red O staining and glycerol phosphate dehydrogenase activity.
Results:
We have observed that rapamycin severely curtails human adipocyte differentiation of both omental and abdominal subcutaneous preadipocytes (to 14% and 19% of standard differentiation, respectively). The rapamycin-mediated inhibition of human adipocyte differentiation could be reversed in the presence of excess amounts of FK-506, which displaces rapamycin from its intracellular receptor, FKPB12. Measurement of cytosolic protein and [3H]thymidine incorporation into DNA confirmed the absence of proliferation during differentiation of human preadipocytes in primary culture.
Discussion:
Our data indicate that rapamycin exerts important negative regulatory effects on adipogenesis in human preadipocytes, through a mechanism that does not depend on interruption of clonal expansion.
Insights
Rapamycin significantly inhibits human adipocyte differentiation, a process independent of clonal expansion. This effect on adipogenesis was reversible with FK-506.
Area of Science:
- Cell Biology
- Endocrinology
- Pharmacology
Background:
- Rapamycin inhibits 3T3-L1 adipocyte differentiation by affecting clonal expansion.
- Human preadipocytes lack clonal expansion during differentiation.
- Investigating rapamycin's effect on human adipogenesis is crucial.
Purpose of the Study:
- To determine if rapamycin inhibits human adipocyte differentiation.
- To explore the mechanism of rapamycin's action in human preadipocytes.
Main Methods:
- Primary human omental and abdominal subcutaneous preadipocytes were cultured.
- Rapamycin's effect on differentiation was assessed using Oil Red O staining and glycerol phosphate dehydrogenase activity.
- Reversal of inhibition was tested with FK-506; proliferation was measured via [3H]thymidine incorporation.
Main Results:
- Rapamycin significantly reduced human adipocyte differentiation (14% omental, 19% abdominal).
- The inhibitory effect was reversed by FK-506, indicating FKBP12 involvement.
- Human preadipocyte differentiation occurred without significant proliferation.
Conclusions:
- Rapamycin negatively regulates human adipogenesis.
- The mechanism does not involve the interruption of clonal expansion.
- Rapamycin's action on human adipogenesis is distinct from its effect on murine 3T3-L1 cells.