Effects of moderate and deep hypothermia on RNA-binding proteins RBM3 and CIRP expressions in murine hippocampal brain slices

Therapeutic hypothermia has emerged as an effective neuroprotective therapy for cardiac arrest survivors. There are a number of purported mechanisms for therapeutic hypothermia, but the exact mechanism still remains to be elucidated. Although hypothermia generally down-regulates protein synthesis and metabolism in mammalian cells, a small subset of homologous (>70%) cold-shock proteins (RNA-binding motif protein 3, RBM3 and cold-inducible RNA-binding protein, CIRP) are induced under these conditions. In addition, RBM3 up-regulation in neuronal cells has recently been implicated in hypothermia-induced neuroprotection. Therefore, we compared the effects of moderate (33.5 °C) and deep (17 °C) hypothermia with normothermia (37 °C) on the regulation of RBM3 and CIRP expressions in murine organotypic hippocampal slice cultures (OHSC), hippocampal neuronal cells (HT-22), and microglia cells (BV-2).Moderate hypothermia resulted in significant up-regulation of both RBM3 and CIRP mRNA in murine OHSC, but deep hyporthermia did not. RBM3 protein regulation was also significantly up-regulated by 33.5 °C, but no significant up-regulation of CIRP protein was observed in the OHSC. Additionally, OHSC exposed to 17 °C for 24 h were positive for Propidium Iodide (PI) immunostaining, indicating the onset of cell death. Similarly, RBM3 gene expression in a HT-22 neuronal cells mono-culture and direct co-culture of HT-22 neuronal cells with BV-2 microglia cells were also up-regulated at 33.5 °C but only in the co-culture at 17 °C. No significant up-regulation of RBM3 nor CIRP gene expression were observed in a BV-2 mono-culture at either temperature.We observed that RBM3 mRNA and protein expressions in murine OHSC, as well as in mono-culture of HT-22 neuronal cells and direct co-culture of HT-22 neuronal cells with BV-2 microglia cells were significantly up-regulated by exposure to moderate hypothermia. These findings further support the implication of RBM3 as a potential effector for hypothermia-induced neuroprotection.

► RBM3 up-regulation has been implicated in hypothermia-induced neuroprotection. ► Moderate hypothermia up-regulates RBM3 and CIRP in brain slices and HT22 cells. ► Hypothermia has no effect on RBM3 and CIRP expressions in BV-2 microglia cells. ► Deep hypothermia leads to cell death in organotypic hippocampal slice cultures. ► Deep hypothermia reduces viability in HT-22 neuronal and BV-2 microglia cells.