[P1-2-03] Calcitriol induces apoptosis in RAW264.7 cells
Keywords:細胞死、calcitriol、アポトーシス
Objectives: Vitamin D affects bone and mineral metabolism and immunity in our body. Epidemiological data suggest that vitamin D may also play a role in the development and progression of cancer. In addition, it has been reported that calcitriol, an active form of vitamin D, has an antiproliferative effect on malignant tumors, but it has also been reported that calcitriol suppresses cell death, suggesting that the effects of calcitriol may vary from the type of cells. In this study, we aimed to clarify the effects of calcitriol stimulation on murine leukemia monocyte-derived macrophage-like cells (RAW264.7 cells) and the pathway of cell death. Material and Methods: Cell death was measured by cytotoxicity assay kit (LDH) and Annexin V/PI staining. Western blotting and ELISA were performed to examine the pathway of cell death. The expression level of cleaved caspase 3, one of the key regulators of apoptosis, was measured by Western blotting after 12 hours of stimulation with calcitriol. The cell supernatant was collected after stimulation with calcitriol for 24 hours, and IL-1β was quantified by ELISA. Results and Discussion: LDH increased in a concentration- and time-dependent manner after stimulation with calcitriol. Annexin V/PI staining showed that calcitriol stimulation increased the number of cells stained with Annexin V and PI compared to the negative control. From these results, it is clear that calcitriol causes cell death in RAW264.7 cells. Calcitriol stimulation increased cleaved caspase 3, and ELISA showed that IL-1β was not detected in the supernatant after 24 hours of stimulation. Therefore, it is considered that calcitriol-induced cell death is due to apoptosis.