Examining the Effects of Exosomes Containing miRNA-143 on MMP-7, BAX, and BCL2 Gene Expression in 5-Fluorouracil-Treated HT-29 Colorectal Cancer Cells

Abstract

Background: One of the biggest challenges in effectively treating colorectal cancer CRC) is the resistance to chemotherapeutic agents like 5-fluorouracil 5-FU). Recently, exosomes—tiny extracellular vesicles—have gained attention for their role in carrying microRNAs miRNAs) that influence how cancer cells behave. Specifically, miRNA-143, known for its tumor-suppressive properties, is found to be downregulated in CRC. This study explores how exosomes loaded with miRNA-143 affect the expression of genes related to apoptosis and metastasis in HT-29 CRC cells that have been treated with 5-FU.

Methods: HT-29 cells were subjected to treatment with 5-FU, and exosomes were extracted using the ExoCib kit. PCR was employed to confirm the presence of miRNA-143 within the exosomes. These exosomes were then introduced to the HT-29 cells. To gauge cell viability, an MTT assay was performed, and the expression levels of the MMP-7, BAX, and BCL2 genes were measured through real-time PCR. Apoptosis was assessed via caspase-3/7 activity assays and flow cytometry with Annexin V/PI staining). Additionally, levels of intracellular reactive oxygen species ROS) were evaluated using the DCFH-DA assay.

Results: The application of exosomes loaded with miRNA-143 led to a notable decrease in HT-29 cell viability (X%) reduction, p<0.05). The expression of MMP-7 was reduced (Y-fold)), while BAX expression rose (Z-fold)), and BCL2 expression fell (W-fold)). There was a notable increase in caspase-3/7 activity, confirmed by flow cytometry, which detected a higher percentage of apoptotic cells (A%)). ROS levels were significantly elevated, indicating that oxidative stress was inducing cell death.

Conclusion: miRNA-143-loaded exosomes amplify the anti-cancer effects of 5-FU by affecting pathways related to apoptosis and metastasis in HT-29 cells. This suggests their promising potential as a complementary treatment option for colorectal cancer.