The confluent monolayer cells were scratched using a tip, and each well was washed with phosphate buffered saline (PBS) to remove the floating cells. Various in vitro and in vivo studies have demonstrated that -mangostin has chemopreventive and chemotherapeutic potential against a wide range of cancer cell types. It shows antiproliferative, proapoptotic, antiangiogenic, and antimetastatic activities [17,18,19]. However, the low oral bioavailability of -mangostin that is due to its first-pass metabolism, poor absorption due to low water solubility, and the efflux effect of Alosetron (Hydrochloride(1:X)) P-glycoprotein, has limited its further clinical applications [20,21,22]. Therefore, designing novel -mangostin analogs is required to improve its bioavailability. The glycosylation of bioactive compounds has been considered as a promising strategy to improve their bioavailability by inducing changes in their physicochemical properties [23,24,25]. The conjugation of sugars to the compounds could facilitate biodistribution in tissues, penetration through biological membranes, metabolic Alosetron (Hydrochloride(1:X)) stabilization, receptor-binding, the stability of labile molecules, the reduction of toxicity, the modification of biological activities, and water solubility. To overcome the poor physicochemical properties of -mangostin, such as low water solubility, six novel glycoside derivatives of the natural compound have been recently produced through biocatalytic glycosylation reactions [26]. All the -mangostin glycosides exhibited an improved water solubility, and several analogs among them showed an increased antibacterial activity against Gram-positive bacteria compared to -mangostin. However, the anticancer property of the -mangostin glycosides has not yet been investigated. In our preliminary study, among six -mangostin glycosides, -mangostin 3- 0.05, ** 0.01, and *** 0.001 versus the control. Thereafter, we investigated the effect of Man-3DG and Man-6DG on the clonogenic growth of HCC cells. As shown in Figure 2B, the colony formation of HepG2, Huh7, and Hep3B cells was inhibited following the treatment with -mangostin, Man-3DG, and Man-6DG using a concentration of 10 M. However, Man-3DG and Man-6DG had a lower ability to suppress the colony formation of HCC cells compared to -mangostin. Collectively, these results indicate that the glycoside analogs of -mangostin did not exhibit a better growth inhibitory activity than -mangostin. However, the -mangostin glycoside analogs were observed to possess the antiproliferative effect against HCC cells. In subsequent studies, we focused on Hep3B cells, in which the -mangostin glycosides showed the prominent growth inhibitory effect in both assays. 2.2. Effects of -Mangostin Glycosides on the Migration of Hep3B Cells To evaluate the effects of Man-3DG and Man-6DG on the migration of Alosetron (Hydrochloride(1:X)) HCC cells, a wound-healing assay was conducted using Hep3B cells. The results showed that Man-6DG, and not Man-3DG, significantly decreased the migration of Hep3B cells at 48 h after treatment compared to the control cells, as observed for -mangostin Alosetron (Hydrochloride(1:X)) (Figure 3). Therefore, Man-6DG may have the potential to inhibit the metastasis of HCC cells. Open in a separate window Figure 3 The effects of -mangostin glycosides on the migration of Hep3B cells by a wound-healing assay. The cells were incubated in the absence or presence of -mangostin, Man-3DG, and Man-6DG (10 M) for 48 h. The cells that migrated into the gap were counted using an optical microscope. The dotted black lines indicate the edge of the gap at 0 h. Each value represents the mean Rabbit polyclonal to CapG SD from three independent experiments. ** 0.01 versus the control. 2.3. Effects of -Mangostin Glycosides on the Apoptosis of Hep3B Cells Abnormal cell cycle progression and the evasion of apoptosis are common features of cancer. Thus, induction of cell cycle arrest and apoptosis in cancer cells is considered as Alosetron (Hydrochloride(1:X)) a key cellular mechanism of action of anticancer drugs [27,28]. To determine whether -mangostin glycosides inhibit the HCC cell growth by causing arrest in a specific phase of cell cycle, we investigated the effects of Man-3DG and Man-6DG on the cell cycle distribution of Hep3B cells through flow cytometric analysis. As shown in Figure 4A, -mangostin, Man-3DG, and Man-6DG caused.