Vitamin D Reduces the Metastatic Potential of the MG-63 Osteosarcoma Cell Line
Osteosarcomas are the most common type of bone tumor, with metastasis to the lungs being the leading cause of death in patients with the disease. Several studies have found a link between the active vitamin D derivative, 1,25(OH)2D3, and the epithelial to mesenchymal transition (EMT), which is a major driver of cancer metastasis. Despite these studies, it is unknown whether 1,25(OH)2D3 can inhibit osteosarcoma metastasis and the mechanisms of action. To answer this question, preliminary scratch migration and RNAseq assays were performed on the osteosarcoma cell line MG-63, which revealed that 1,25(OH)2D3 inhibited migration while also repressing key EMT inducers and positive regulators (e.g. CD44 and MMP3). As a result, this study will use Western blot and confocal immunofluorescence analysis to correlate these findings at the protein expression and signaling levels. After treating MG-63 cells with 1,25(OH)2D3 for 24 and 48 hours, lysates were collected. Antibodies raised against CD44 and beta-actin were used in Western blot analysis. CD44 and MMP3 immunostaining was performed, and images were taken with a Zeiss LSM900 confocal microscope. The preliminary RNA-based findings were supported by both Western blot and immunostaining results. Furthermore, treatment with 1,25(OH)2D3 decreased the presence of MMP3 in the nucleus. These findings suggest that 1,25(OH)2D3 inhibits EMT by down regulating CD44 and MMP3 to keep MG-63 cells in a more epithelial-like non-migratory cellular phenotype.
