Vitamin D3 Potentiates the Antitumor Effects of Arsenic Trioxide in Human Leukemia (hl-60) Cells

Recent studies in our laboratory have indicated that oxidative stress plays a key role in ATOinduced cytotoxicity in human leukemia (HL-60) cells. Pro-oxidants have been known to plays a role in free radical-mediated oxidative stress. Medical reports have indicated that Vitamin D3, a well-known antioxidant works synergistically with tamoxifen to inhibit breast cancer cell proliferation. Therefore, the ultimate goal of this research was to determine whether supplementation of vitamin D3 increases the activity of ATO toxicity in HL-60 cells. To accomplish this goal, HL-60 cells were treated either with 50% lethal dose of ATO alone or with ATO and different low doses of vitamin D3. Live and death cells were determined by both trypan blue exclusion test and MTT assay, respectively. The extent of oxidative cell/tissue damage was determined by measuring MDA levels by spectrophotometry. Cell apoptosis was measured by flow cytometry analysis of phosphatidylserine externalization using Annexin V assay kit. The results of MTT assay indicated that vitamin D3 exposure potentiates the toxic activity of ATO in HL-60 cells in a dose dependent manner. A similar trend was obtained with the trypan blue exclusion test. Co-administration of vitamin D3 and ATO resulted in a significant (P<0.05) increase in MDA level compared to ATO alone. A statistically significant and dose-dependent increase (p <0.05) was recorded in annexin V positive cells (apoptotic cells) with increasing doses of vitamin D3 in ATOtreated cells. This finding was confirmed by the result of DNA laddering assay showing clear evidence of DNA fragmentation in ATO and vitamin D3 treated HL-60 cells. Overall, the present study indicates that vitamin D3 potentiates the antitumor effects of ATO at least part, via oxidative stress and phosphatidylserine externalization.