## 4 Discussion
Loliolide treatment attenuated the adipogenic suppression of AMPK phosphorylation in the presence of AMPK inhibitor compound C at the early stages of adipogenesis. Attenuation of AMPK phosphorylation indicated that loliolide suppressed the adipogenesis at the early stages via AMPK/MAPK signaling. Canonical Wnt/\(\beta\)-catenin signaling is also involved in adipogenic differentiation of MSCs. Adipogenic stimulation of cells is known to antagonize the Wnt/\(\beta\)-catenin pathway [35]. Increased degradation of \(\beta\)-catenin is observed along elevated PPAR\(\gamma\) translocation to nucleus, resulting in expression of adipogenic genes. Although Wnt signaling induces stem cells to differentiate into pre-adipocyte to commit adipogenic lineage, it also inhibits the adipocyte maturation of pre-adipocytes [36; 37]. Wnt 10b is a Wnt subunit that is expressed in MSCs, and its overexpression was shown to decrease adipogenesis [38]. Phosphorylation of \(\beta\)-catenin at Ser31, Ser41, and Thr47 facilitates its degradation and stimulates the adipogenic differentiation at the early stages of MSCs adipogenic differentiation [39]. However, Ser675 phosphorylated \(\beta\)-catenin via the activities of pKa was reported to induce its nuclear translocation and transcriptional activity [40]. In a previous report, this translocation of \(\beta\)-catenin was suggested to lead to the inhibition of PPAR\(\gamma\) activity as a result of \(\beta\)-catenin-PPAR\(\gamma\) binding [41]. The current results show that loliolide treatment not only elevated the Wnt10b levels in hBM-MSC adipocytes, but also stimulated the nuclear translocation of active \(\beta\)-catenin. [PERSON] et al. [42] showed that loliolide stimulated the dephosphorylation of \(\beta\)-catenin through enhancement of AKT/PKA-mediated phosphorylation in human dermal papilla cells, hence activating Wnt signaling. These results are in agreement with current findings, further supporting the effect of loliolide on Wnt/\(\beta\)-catenin pathway. Overall, it was suggested that loliolide inhibited the adipogenic differentiation of hBM-MSCs via activation of Wnt/\(\beta\)-catenin, which resulted in AMPK-mediated suppression of PPAR\(\gamma\) activity. Although loliolide has been reported to alter the gene expressions to exert its bioactivities, in vivo studies showing whether these effects could be observed at the phenotype levels are lacking. To the best of our knowledge, only the antioxidant effect of loliolide was observed in zebrafish models in vivo where loliolide reduced the 2,2\({}^{\prime}\)-azobis(2-amidinopropane) dihydrochloride-induced lipid peroxidation [43]. This suggested that the changes induced by loliolide at gene level might be present at phenotype level as well. Considering the potential loliolide holds, future studies to explore the effect of loliolide on adipogenesis at transcriptome and epigenome levels via in vivo models are urged. In summary, loliolide was revealed to repress adipogenic differentiation of hBM-MSCs via activating the AMPK and Wnt/\(\beta\)-catenin signaling which subsequently resulted in suppressed PPAR\(\gamma\) pathway (Figure 5). It was suggested that loliolide is a promising phytochemical with anti-adipogenic properties, although further in vivo studies are needed to better evaluate its potential and mechanism of action. Nonetheless, the current study provided important reference for future studies to develop loliolide-centered agents for prevention and treatment of obesity and related complications. **Author Contributions:** Conceptualization and methodology, F.K., J.H.O., and Y.S. ; validation, formal analysis, and investigation, J.H.O., M.-S.J., and H.K. ; resources, Y.S. and C.-S.K. ; data curation, F.K., C.-S.K., and Y.S. ; writing--original draft preparation, visualization, F.K. and J.H.O. ; supervision and project administration, C.-S.K. and Y.S. ; and funding acquisition, M.-S.J. and C.-S.K. All authors have read and agreed to the published version of the manuscript.