We found for the first time that β-catenin phosphorylation was transiently increased, and total protein of β-catenin degraded after stroke in both the penumbra and core of the normothermic brain; moderate hypothermia increased β-catenin phosphorylation at 24 h to 48 h after stroke, and blocked degradation of total β-catenin in the penumbra.
We have demonstrated in this study that hypothermia prevented stroke-induced β-catenin degradation in the ischemic penumbra but not in the core; this action might contribute to the protective effect of hypothermia. β-catenin is not only a transcription factor in the Akt pathway [9
], but is also associated with multiple adhesion molecules such as α-catenin, p-catenin and cadherin to form cell-cell adhesion, which is fundamental for constructing and maintaining multicellular organisms [10
]. In the brain, β-catenin plays a critical role in the formation of synapses by associating with the cell adhesion proteins [12
]. Therefore, β-catenin degradation after stroke may down-regulate pro-survival gene expression activated by β-catenin and dismantle cell-cell adhesion and connection, thus aggravating ischemic injury. Indeed, β-catenin degradation has been shown to increase while its overexpression blocks apoptosis in vitro
], and protein levels of β-catenin decrease in patient brains with Alzheimer’s disease [4
]. β-catenin degradation may also play critical roles in pathological processes after stroke. We found in this study that β-catenin phosphorylation transiently increased at 5h in the normothermic brain, and this hyper-phosphorylation was accompanied by a later degradation of total β-catenin. This correlation suggests that β-catenin phosphorylation progresses to degradation [9
]. Furthermore, β-catenin might be phosphorylated by GSK3 β activity, as levels of phosphorylated GSK3 β transiently decreased from 5h to 24h, indicating increased-activity during this period after stroke.
We have previously reported that hypothermia blocks ischemic injury without inhibiting increased-GSK3β activity [14
]; this result contradicts with the earlier results showing that GSK3β activity leads to neuronal death [2
]. In our prior study, however, hypothermia blocks nuclear translocation of β-catenin [14
], suggesting that hypothermia may block molecular signaling downstream of GSK3β. The current study further supports this conclusion by showing that hypothermia could stop the degradation of β-catenin. Nevertheless, it is yet not clear how hypothermia retarded β-catenin’s degradation. As we have discussed, in the penumbra of normothermic brain, transient increases in β-catenin phosphorylation at 5h appear to correlate with β-catenin degradation at later time points, which is consistent with the notion that β-catenin phosphorylation results its degradation [9
]. However, in the penumbra of hypothermic brains, phosphorylated β-catenin accumulated from 5 to 48 h after stroke while total β-catenin was not degraded, which contradicts the aforementioned hypothesis. A possible reason is that hypothermia blocks transportation of phosphorylated β-catenin into the proteosome and thus stops β-catenin degradation. Nevertheless, in the ischemic core where infarction was not attenuated by hypothermia, β-catenin degradation was not blocked by hypothermia either, suggesting that β-catenin degradation correlates with ischemic injury.
Transient reduction in GSK3β phosphorylation after stroke in normothermic brains appears to contradict the result of our previous study [14
], in which GSK3 β phosphorylation persistently decreased. This inconsistency might be due to sample collection. In the previous study, the sample was dissected from a region lateral to the midline of the brain; by contrast, in the current study, the sample was collected from a more dorsal region. Nevertheless, hypothermia showed a similar effect by reducing additional GSK3β phosphorylation.
Intra-ischemic hypothermia has long term effects on multiple cell signaling pathways after stroke. As we recently reviewed [16
], intra-ischemic hypothermia not only blocks excitatory neurotransmitter, glutamate release and inhibits intracellular Calcium increase during stroke, it also improves ATP recovery after reperfusion, inhibits free radical generation, inflammatory response and blood–brain barrier permeability. In addition, intra-ischemic hypothermia has multiple effects on necrotic and apoptotic pathways. As consistent with previous investigations, here we provide further evidence that intra-ischemic hypothermia enhances β-catenin phosphorylation and protein levels from 5 to 48 h after stroke.
In conclusion, we demonstrated that β-catenin degraded after stroke in normothermic rats, whereas moderate hypothermia inhibited its degradation. Inhibiting β-catenin degradation might contribute to the protective effect of moderate hypothermia.