Panoply™ Human GPC3 Over-expressing Stable Cell Line

Glypican-3 (GPC3) is a proteoglycan involved in regulating proliferation and survival and is associated with cancer. Here, researchers observed that GPC3 expression was inversely associated with the invasive/metastatic abilities of Hs578T, MDA-MB231, ZR-75-1, and MCF-7 human breast cancer cell lines. GPC3 silencing activated the growth, cell death resistance, migration, and invasive/metastatic abilities of MCF-7 cancer cells, while GPC3 overexpression suppressed these properties in the MDA-MB231 tumor cell line. Furthermore, GPC3 silencing enhanced the mesenchymal characteristics of MCF-7 breast cancer cells and reduced expression of the epithelial marker E-cadherin. Conversely, GPC3 overexpression induced mesenchymal-to-epithelial transition (MET) in MDA-MB231 breast cancer cells, leading to re-expression of E-cadherin and decreased expression of vimentin and N-cadherin. Although GPC3 inhibits the canonical Wnt/β-catenin pathway in breast cancer cells, this inhibition has no effect on E-cadherin expression. ZEB1, a transcriptional repressor of E-cadherin, is upregulated in GPC3-silenced MCF-7 cells but downregulated in MDA-MB231 cells when GPC3 is overexpressed. These findings suggest that GPC3 induces re-expression of E-cadherin in MDA-MB231 cells by downregulating ZEB1. Therefore, GPC3 is an important regulator of EMT in breast cancer and a potential target for procedures against breast cancer metastasis.

Although MCF-7-sh3 and control cells are morphologically similar and grow as a monolayer of epithelial polyhedral cells, researchers found that GPC3-overexpressing MDA-MB231 cells lost their fibroblast-like appearance and acquired an epithelial morphology (Figure 1A). To analyze the morphological changes induced by GPC3 in detail, F-actin organization was examined using phalloidin-FITC staining. Confocal images were processed and a graphical depiction was generated, with the x-axis representing distance through the cell and the y-axis representing fluorescence levels. The results showed that while actin in MCF-7 cells was primarily present in cortical locations (as expected for epithelial cells), GPC3 silencing induced the assembly of a small number of F-actin stress fibers (Figure 1B). Furthermore, while control cells displayed large actin stress fibers, GPC3 overexpression in MDA-MB231 cells resulted in the loss of these fibers and the re-localization of actin to a predominantly cortical distribution (Figure 1B).

Figure 1. Effect of GPC3 on cell morphology and actin cytoskeleton organization. (Castillo L F, et al., 2016)