Anti-proliferative effect of hibernating American bullfrog, Lithobates catesbeianus, blood plasma on the THP-1 cells

Hibernation is a remarkable process that allows an organism to go dormant for long periods of time during the winter, when food is scarce and living conditions can be lethal. Much is unknown about the physiology behind hibernation, and most research comes from mammalian models. For example, it is known that during hibernation many of the cells in the body are impeded from going though mitosis but the mechanism that caused this was unknown. It was recently discovered that 13-lined ground squirrels (Ictidomys tridecemlineatus) and woodchucks (Marmota monax) have a protein, alpha-2-macroglobulin, in their blood plasma, which has significant anti-proliferative effects on spleen cells (Sieckman et al., 2014). The goal of my study was to determine if an ectothermic organism, the American bullfrog, Lithobates catesbeianus, also actively suppresses mitosis while hibernating and if so, determine if it is caused by a component in their blood plasma. Cellular assays using Human THP-1 cells were conducted by incubating cells in different treatments of plasma collected from active and hibernating bullfrogs. After incubation, the cells were either counted to determine if the rate of cell growth had been inhibited or stained with a dye that attaches to DNA and run through a flow cytometer. Results of the cell count assays were inconclusive regarding how the rate of growth had been affected by hibernating plasma treatment. Flow cytometry assays found that the majority of cells in the hibernating plasma treatment were in the M phase of the cell cycle, instead of the G0 phase. This suggests that the majority of cells had replicated their DNA, but mitosis was halted before the cell divided. This may be because there is substantial cell death throughout hibernation and by stopping mitosis just after DNA replication, the cells are better prepared for proliferation when the animal is exiting hibernation