Abstract:
Bilobalide, the only sesquiterpene compound from Ginkgo biloba leaf, exhibits numerous beneficial bioactivities, such as neuroprotective, antiinflammatory, and antioxidant activity. Certain bioactive components of Ginkgo biloba extract have previously been reported to have potential to attenuate lipid metabolism. However, the effect of bilobalide on lipid metabolism remains unclear. In this study, we used 3T3-L1 cells as the cell model to investigate the effect of bilobalide on adipogenesis and lipolysis. The results showed that bilobalide inhibited 3T3-L1 preadipocyte differentiation and intracellular lipid accumulation. The expression of several specific adipogenic transcription factors and genes was downregulated on both mRNA and protein levels in response to bilobalide treatment. By contrast, bilobalide treatment upregulated the expression of important lipolytic genes such as adipose triglyceride lipase (ATGL), hormone-sensitive lipase (HSL) and carnitine palmitoyltransferase-1α, and stimulated the phosphorylation of AMP-activated protein kinase (AMPK), acetyl-CoA carboxylase 1, and HSL and stimulated the phosphorylation of AMP-activated protein kinase (AMPK), acetyl-CoA carboxylase 1, and HSL. Furthermore, bilobalide treatment partially restored AMPK activity following its blockade by compound C. Notably, bilobalide also exerted the dose-dependent cytotoxicity specific to the mature adipocytes only, indicating its potential for regulating apoptosis in them. We then evaluated the apoptotic effects of bilobalide on 3T3-L1 mature adipocytes and elucidate the underlying mechanisms thereof. FACS assay revealed the pro-apoptotic effects of bilobalide on these cells. Bilobalide induced early apoptosis by reducing the mitochondrial membrane potential (MMP). DNA fragmentation was confirmed using TUNEL staining. Additionally, bilobalide increased the intracellular reactive oxygen species (ROS) levels and activities of Caspases 3/9. Pretreatment with NAC (an ROS scavenger) confirmed the role of ROS in inducing apoptosis. Moreover, bilobalide up- and down-regulated the expression of Bax and Bcl-2, respectively, at the mRNA and protein expression levels; upregulated the Bax/Bcl-2 ratio; triggered the release of cytochrome c from the mitochondria; and increased the protein expression of cleaved Caspase 3/9, and PARP cleavage. Taken together, these findings suggest that bilobalide exerts multiple functions including inhibiting adipogenesis, promoting lipolysis in 3T3-L1 cells by activating the AMPK pathway, and inducing apoptosis in mature 3T3-L1 adipocytes through the ROS-mediated mitochondrial pathway. Bilobalide may therefore offer a potential novel treatment for obesity.
