Conclusion
These observations illustrated that knockdown of UBD partially suppressed porcine intramuscular and subcutaneous preadipocyte adipogenesis through the Akt/mTOR signalling and inhibited cell proliferation, suggesting the essential role of UBD in the differentiation of preadipocytes.
Methods
Intramuscular and subcutaneous preadipocytes were isolated from the longissimus dorsi and neck subcutaneous deposits of Chinese native Guanzhong Black piglets (3-5 days old), respectively. Lentivirus with short hairpin RNA (shRNA) for UBD was applied to knockdown UBD expression. We used real-time PCR and Western blot analysis to detect gene expression. Lipid droplets were dyed with Oil Red O, and cell proliferation was assessed using flow cytometry, 5-ethynyl-2'-deoxyuridine incorporation and cell counting assays.
Results
Lipogenesis through the Akt/mTOR pathway was inhibited when preadipocytes were transfected with UBD shRNA. The expression of adipogenic genes and the number of lipid droplets were obviously diminished. Moreover, repression of UBD attenuated cell proliferation. UBD downregulation resulted in cell cycle arrest because of a decreased proportion of S-phase cells, and the expression of positive cell proliferation markers was significantly decreased.