Background: Insights into the biology of chemotherapy resistance may suggest novel strategies to treat Acute Myeloid Leukemia (AML). We examined the role of hyperbaric oxygen (HBO) in AML sensitivity to chemotherapy, oxygen consumption, reactive oxygen species (ROS) generation, glycolytic pathway activation, and gene expression.
Methods: Human AML cell lines HL-60 and MV-411 were treated with cytarabine followed by HBO (100% oxygen at 2.5 atmosphere absolutes for 2 hours in an in vitro HBO chamber) or normobaric culture conditions. At 24 hours, cells were analyzed for proliferation, apoptosis and oxygen consumption. Studies of HBO- treated cells without chemotherapy included CellROX Oxidative Stress assay and real-time PCR with primer sets specific for human hexokinase2 and phosphofructokinase to examine HBO effects on glycolysis. Gene expression profiling by the Human Transcriptome Array 2.0 (Affymetrix) examined differences in gene expression following HBO in HL-60 cells and AML patient samples.
Results: Leukemia cells treated with HBO and cytarabine demonstrated reduced proliferation and increased late apoptosis confirming increased sensitivity to cytarabine. Treatment with HBO plus cytarabine resulted in increased oxygen consumption measured by the seahorse assay at 24 hours post-HBO in MV-411 cells compared to cytarabine alone. Significant increases in ROS levels were observed at 24 hours post-HBO for both cell lines when compared to non-treated cells. Glycolysis pathway components human hexokinase 2 and phosphofructokinase were upregulated in HL60 and MV411 cells treated with HBO compared to controls. Gene expression profiling showed that approximately 200 genes in coding regions were upregulated in HBO-treated HL-60 cells; specific piRNA clusters were downregulated in both AML cell line HL60 and an AML bone marrow sample when treated with HBO.
Conclusions: HBO treatment significantly increases the sensitivity to cytarabine in AML cell lines in vitro, an effect associated with increased late apoptosis and changes in cellular respiration. Increased ROS, upregulation of glycolysis-related enzymes and changes in gene expression profiling provide insight into potential mechanisms of HBO in AML.