Panoply™ Human GPC3 Knockdown Stable Cell Line

Here, researchers demonstrated for the first time the bidirectional regulatory effects of GPC3 between normal cells and liver cancer cells. Treatment of LO2 and HepG2 cells with GPC3 arrested the LO2 cell cycle at the G0/G1 phase, inhibited cell proliferation, and promoted apoptosis and invasion. However, in HepG2 cells, proliferation was promoted. Silencing GPC3 inhibited the proliferation and invasion of HepG2 cells and promoted apoptosis. Subsequent experiments revealed that GPC3 was expressed in both LO2 and HepG2 exosomes, with GPC3 expression significantly higher in HepG2 exosomes than in LO2 exosomes. These findings suggest that GPC3 in exosomes may serve as a biomarker for HCC. Furthermore, HepG2 exosomes (Exo) inhibited LO2 cell proliferation and promoted apoptosis and invasion, consistent with the effects of GPC3 treatment. The researchers also found that GPC3 was also present in HepG2 exosomes silencing GPC3 (shGPC3-Exo). Its effects on LO2 cells were similar to those of HepG2 exosomes (Exo), but to a lesser extent. shGPC3-Exo promoted HepG2 cell proliferation but inhibited cell invasion. Therefore, GPC3 in Exo plays a role in the proliferation of both LO2 and HepG2 cells. Further studies demonstrated that GPC3 in liver cancer exosomes regulates the proliferation, apoptosis, and invasion of LO2 and HepG2 cells through the Wnt/β-catenin signaling pathway.

To further investigate the role of GPC3 in HCC cell growth, the researchers transfected HepG2 cells with a GPC3-targeting shRNA. Following transfection, both GPC3 mRNA and protein expression in HepG2 cells were reduced (Figures 1A-C). Knockdown of GPC3 inhibited HepG2 cell growth (Figure 1D). Flow cytometric analysis using Annexin V/PI staining also revealed an increase in apoptotic cells in HepG2 cells following GPC3 knockdown (Figure 1E). Subsequently, they tested whether cell cycle arrest affected the growth of GPC3 knockdown HepG2 cells. As shown in Figure 1F, GPC3 knockdown resulted in an increase in the proportion of cells in the G0/G1 phase of HepG2 cells, indicating a trend toward G0/G1 arrest. Next, the effect of GPC3 on HCC cell invasion was assessed using Transwell assays. The results showed that the invasive ability of GPC3 knockdown HCC cell lines was reduced (Figure 1G), suggesting that GPC3 gene silencing in HCC cells can promote cell apoptosis and cell cycle arrest, thereby inhibiting HCC cell proliferation and invasion.

Figure 1. Knockdown of GPC3 inhibited the proliferation and invasion of HCC cells. (Pu C, et al. 2021)