Salidroside inhibits the invasion and migration of colorectal cancer cells by regulating MMP-12 and WNT signaling pathway

Abstract

Colorectal cancer (CRC) is a prevalent and highly lethal malignancy, with current therapeutic efficacy limited by the tumor's high invasiveness and metastatic potential. Matrix metalloproteinases (MMPs) and the WNT (Wingless/Integrated) signaling pathway play key roles in the invasion and metastasis of CRC. Salidroside, a natural compound, has demonstrated inhibitory effects in several cancers, but its precise molecular mechanism in CRC cells remains unclear. This study aims to investigate the antitumor effect of salidroside on CRC and its molecular mechanism in influencing epithelial-mesenchymal transition (EMT) by regulating MMP-12 and the WNT signaling pathway. The effects of salidroside on CRC cell proliferation, migration, and invasion were evaluated through in vitro experiments using HCT-116 and SW620 cell lines. The antitumor effects of salidroside were validated using CCK-8, wound healing, and Transwell assays. Expression changes of MMP-12, WNT signaling-related proteins (e.g., β-catenin, GSK-3β), and EMT markers (e.g., E-cadherin, Vimentin) after salidroside treatment were measured by qRT-PCR and Western Blot. Additionally, bioinformatics analysis was performed using TCGA and GEO databases in combination with the BEST online tool to identify differentially expressed genes, followed by GSEA enrichment analysis. Salidroside showed significant antiproliferative and inhibitory effects on the migration and invasion of CRC cells. In vitro experiments demonstrated that salidroside significantly inhibited CRC cell proliferation and reduced their migration and invasion capabilities. qRT-PCR and Western Blot analyses showed that salidroside significantly downregulated MMP-12 expression and led to changes in the expression of WNT signaling and EMT-related proteins, specifically downregulating β-catenin, upregulating E-cadherin, and downregulating Vimentin. Furthermore, bioinformatics analysis indicated that MMP-12 plays a crucial role in salidroside-mediated CRC inhibition, further supporting its potential as a key target. In conclusion, salidroside suppresses CRC invasion and migration by downregulating MMP-12 and modulating the WNT signaling pathway, thereby inhibiting the EMT process. These findings suggest that salidroside holds potential as a therapeutic agent for CRC, offering a novel approach to CRC treatment.

Keywords: MMP-12; Salidroside; WNT signaling pathway; colon cancer; invasion; migration.

Figure 1

Time-dependent effect of different concentrations of salidroside on CRC cell proliferation and viability. A. CCK-8 analysis of the time-dependent effect of salidroside on cell viability (12 h, 24 h, 48 h). B. Cell proliferation assay showing the effect of different salidroside concentrations on HCT-116 and SW620 cells. Note: CRC, colorectal cancer; nsP > 0.05, **P < 0.01, ****P < 0.0001.

Figure 2

Inhibitory effect of salidroside IC50 on migration and invasion abilities of HCT-116 and SW620 cells. A. Wound healing assay showing the effect of salidroside IC50 treatment on HCT-116 and SW620 cell migration. B. Transwell invasion assay evaluating the effect of salidroside IC50 on cell invasion. Note: CRC, colorectal cancer; ***P < 0.001, ****P < 0.0001.

Figure 3

Differential gene analysis under salidroside intervention. A. TCGA CRC samples. B. TCGA breast cancer samples. C. MCF7 cells from the GSE85871 dataset following salidroside intervention. D. GSE110225 CRC samples compared to controls. Note: CRC, colorectal cancer.

Figure 4

Volcano plots of differential gene expression. A. TCGA CRC samples showing upregulated and downregulated genes. B. TCGA breast cancer samples showing differential gene expression. C. Differential gene expression in MCF7 cells from the GSE85871 dataset following salidroside intervention. D. GSE110225 CRC samples. Note: CRC, colorectal cancer.

Figure 5

Intersection analysis of differential genes in MCF7 cells and multiple cancer datasets. A. Venn diagram showing the intersection of highly expressed genes in MCF7 cells treated with salidroside and low-expressed genes in TCGA/GEO datasets. B. Venn diagram showing the intersection of low-expressed genes in MCF7 cells treated with salidroside and highly expressed genes in TCGA/GEO datasets. C. PPI analysis of 17 co-expressed genes, with four key genes identified. Note: CRC, colorectal cancer.

Figure 6

KEGG enrichment analysis revealing the regulatory effect of salidroside on the WNT signaling pathway.

Figure 7

Effect of salidroside and MMP-12 overexpression/inhibition on CRC. A. Screening of MMP-12 expression in CRC tissues using the BEST online database. B. qRT-PCR analysis of MMP-12 expression in different treatment groups. C. Western blot analysis of MMP-12 protein levels in different treatment groups. D. Relative expression of MMP-12 mRNA in transfected cells. Note: IC50 = 66.5 μM, CRC, colorectal cancer; MMP-12, matrix metalloproteinase-12; ****P < 0.0001.

Figure 8

Regulation of Cyclin D1, c-Myc, and β-catenin protein expression by salidroside and MMP-12. A. Western Blot analysis of Cyclin D1, c-Myc, and β-catenin protein levels in HCT-116 cells under different treatment conditions: Control, Salidroside (IC50), si-NC, si-MMP-12, pcDNA-NC, and pcDNA-MMP-12. B. Western Blot analysis of Cyclin D1, c-Myc, and β-catenin protein levels in SW620 cells under the same treatment conditions as in 8A. Note: IC50 = 66.5 μM, CRC, colorectal cancer; MMP-12, matrix metalloproteinase-12; nsP > 0.05, ****P < 0.0001.

Figure 9

Effect of salidroside on EMT marker E-cadherin and Vimentin expression through MMP-12 regulation. A. Western Blot analysis of E-cadherin and Vimentin protein levels in HCT-116 cells under different treatment conditions: Control, Salidroside (IC50), si-NC, si-MMP-12, pcDNA-NC, and pcDNA-MMP-12. B. Western Blot analysis of E-cadherin and Vimentin protein levels in SW620 cells under the same treatment conditions as in 9A. Note: IC50 = 66.5 µM, CRC, colorectal cancer; MMP-12, matrix metalloproteinase-12; nsP > 0.05, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Figure 10

Synergistic effect of MMP-12 regulation and salidroside on the migration and invasion abilities of HCT-116 and SW620 cells. A. Wound healing assay showing the effect of MMP-12 overexpression or knockdown combined with salidroside treatment on cell migration. B. Transwell invasion assay showing the effect of different treatments on cell invasion abilities. Note: IC50 = 66.5 μM, CRC, colorectal cancer; MMP-12, matrix metalloproteinase-12; nsP > 0.05, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.