Inhibition of semaphorin 4D enhances chemosensitivity by increasing 5-fluorouracile-induced apoptosis in colorectal cancer cells

Overexpression of semaphorin 4D (SEMA4D), an immune semaphorin, is found in various human malignancies, including colorectal cancer (CRC). In this study, we explored the relationship between silencing SEMA4D expression and 5-fluorouracil (5-FU) response in the colorectal cancer cell line. SW48 cells were transfected with a short interfering RNA (siRNA) in order to silence SEMA4D gene expression and then exposed to 5-FU for 48 h. The down-regulation of SEMA4D expression was confirmed by qRT-PCR and the particular concentration of 5-FU was acquired using MTT assay. Flow cytometry and western blot were used to evaluate apoptosis rate and pro- and anti-apoptotic expression levels of proteins involved in apoptosis including Bax, Bcl-2, P53, and caspase-3. Other oncogenic activities including epithelial-mesenchymal transition (EMT) process, cancer stem cell (CSC) markers, and beta-catenin pathway were investigated using qRT-PCR, and western blot. The proliferation was analyzed via colony formation test and cell invasion was assessed by transwell assay. Our data demonstrate that SEMA4D silencing results in strikingly elevated apoptosis in response to 5-FU treatment and leads to down-regulation of Bcl-2 and overexpression of Bax, P53, and caspase-3 in protein levels. Furthermore, the mRNA and protein expression levels of beta-catenin, as well as transcript expressions of CSCs and EMT markers, were remarkably diminished. However, mRNA expression of E-cadherin as an epithelial marker was significantly increased in 5-FU treatment combined with siRNA SEMA4D. This study implicates that the silencing of SEMA4D by siRNA promotes the chemosensitivity of SW48 cells to 5-FU and it may be a potential therapeutic agent for colon cancer therapy.