FGF21 alleviates α-II-spectrin breakdown in a model of oxygen-glucose deprivation and modulates global protein phosphorylation in hippocampal neurons.

Fibroblast growth factor 21 (FGF21) is a pleiotropic hormone that promotes hippocampal neuroprotection in animal models of ischemic brain injury, but how it works is unclear. In the first experiment we subjected isolated hippocampal neurons to oxygen-glucose deprivation (OGD) injury to test if FGF21 induced neuroprotection is mediated by its direct effects on neurons. Further, because cold stress regulates FGF21 levels in animals, we also explored if post-insult temperature (37 °C versus 33.5 °C) modified FGF21 induced neuroprotection. Experiment 1 revealed that FGF21 treatment at 37 °C decreased α-II-spectrin breakdown product-145 (SBDP-145) levels but did not affect 24 h cell survival. Intraischemic therapeutic hypothermia (TH) decreased SBDP-145 levels and increased 24 h cell survival. Co-treatment with FGF21+TH augmented the effect of cooling to increase the levels of the cold-shock proteins (CSPs) RNA-binding motif 3 (RBM3) and cold-induced RNA-binding protein (CIRBP), without further effecting survival. In Experiment 2, to obtain additional insights on FGF21 signaling responses in neurons, we measured the global phosphoproteome in hippocampal neurons after a 30 min pulse treatment with FGF21, both in normothermic cells and in cells that were subjected to hypothermia for 48 h. This revealed a reprogramming effect of temperature on FGF21 signaling responses. We also identified myristoylated alanine-rich c-kinase substrate-like protein 1 (MARCKSL1) as a novel FGF21 target. In follow-up studies in the OGD model, FGF21 treatment prevented injury induced decreases in MARCKSL1 protein levels. We conclude that FGF21 has a direct inhibitory effect on α-II-spectrin breakdown and modifies neuronal signaling responses in a temperature dependent manner in hippocampal neurons.