Panoply™ Human MSLN Knockdown Stable Cell Line

Cancer-associated fibroblasts (CAFs), known for promoting colorectal cancer (CRC) progression and metastasis, have emerged as a promising therapeutic target. However, the remarkable heterogeneity of CAFs and their complex interactions with tumor cells pose a significant challenge to the development of precise and effective therapeutic strategies. Here, researchers discovered significant spatial colocalization between CTHRC1+ CAFs and a distinct malignant epithelial cell subtype, both located within an epithelial-mesenchymal transition (EMT)-active microenvironment. Results indicate that CTHRC1+ CAFs, as the primary source of WNT5A, promote epithelial-mesenchymal transition (EMT) and enhance tumor cell invasiveness by upregulating WNT5A expression in adjacent mesenchymal lymph nodes (MSLNs). This signaling pathway is crucial for CRC progression and metastasis. Therefore, targeting the CTHRC1+ CAF-WNT5A-MSLN signaling pathway offers a promising therapeutic strategy for patients with advanced CRC. These studies provide new insights into the role of CAFs in CRC progression and offer potential avenues for developing targeted therapies to block this pathway.

Here, researchers established stable MSLN knockdown in HCT116 cells and stable MSLN overexpression in DLD-1 cells. Scratch-healing and transwell assays demonstrated that MSLN overexpression enhanced the migration and invasion capabilities of DLD-1 cells (Figure 1A, C), while MSLN knockdown in HCT116 cells had the opposite effect (Figure 1B, D). Furthermore, in MSLN knockdown cells, E-cadherin levels were increased, while N-cadherin and Vimentin expression was decreased, while MSLN overexpression enhanced EMT activity in DLD-1 cells (Figure 1I). Next, the researchers investigated whether CTHRC1+ fibroblasts affect the invasion and EMT activity of CRC cells by co-culturing CRC cells with CAFs and performing transwell and scratch-healing assays. The results showed that overexpression of MSLN or addition of rhWNT5A in DLD-1 cells reversed the effects of CAFs-shCTHRC1 on DLD-1 cell migration, invasion, and EMT activity (Figure 1E, G, J). Conversely, MSLN knockdown in HCT116 cells or treatment with the WNT5A inhibitor Box5 reversed the enhanced effects of CAFs-CTHRC1 on HCT116 cell migration, invasion, and EMT activity (Figure 1F, H, J). These results suggest that CTHRC1+CAFs regulate WNT5A, which in turn modulates MSLN expression, thereby enhancing CRC cell migration, invasion, and EMT activity, ultimately driving tumor progression.

Figure 1. CTHRC1+ CAF regulates MSLN expression via WNT5A to impact migration, invasion and EMT activity in CRC cells. (Lu Y, et al., 2025)