| Summary: | 博士 === 國立成功大學 === 基礎醫學研究所 === 94 === Perinatal hypoxic-ischemic brain injury is a major cause of permanent neurological dysfunction in children. An approach to study the treatment of neonatal hypoxic-ischemic encephalopathy that allows for neuroprotection is to investigate the states of tolerance to hypoxic-ischemia. Twenty-four-hour carotid-artery-ligation preconditioning established by delaying the onset of hypoxia for 24 h after permanent unilateral carotid ligation in neonatal rats markedly diminished the cerebral injury. Although much has been learned about the ischemic preconditioning mechanisms in adult rats, the signaling mechanisms of this 24-h-carotid-artery-ligation preconditioning in neonatal rats remain unknown.
We demonstrated that carotid artery ligation 24 h before hypoxia on postnatal day 7 (P7) rat pups provided complete neuroprotection, while artery ligation 6 h produced intermediate benefit, at behavioral and pathological levels compared to ligation 1 h before hypoxia. We first showed that the 24-h-carotid-artery-ligation preconditioning was associated with a robust and sustained activation of cAMP response element-binding protein (CREB), a transcription factor that acts as a key mediator of stimulus-induced nuclear responses underlying learning and memory, survival, and synaptic plasticity of the nervous system. Intracerebroventricular infusions of antisense CREB oligodeoxynucleotides significantly reduced the 24-h-carotid-artery ligation-induced neuroprotection by decreasing CREB expression. Pharmacological activation of the cAMP-CREB signaling with rolipram 24 h prehypoxia protected rat pups at behavioral and pathological levels by sustained increased CREB phosphorylation. These findings suggest that CREB activation provides important mechanism for potential pharmacological treatment against neonatal hypoxic-ischemic brain injury.
The upstream signaling mechanisms leading to CREB activation, however, in this 24 h carotid-artery ligation preconditioning of neonatal rat brain remained unknown. We next found that vascular endothelial growth factor (VEGF)-A and VEGF receptor-2 (VEGFR-2) instead of VEGFR-1 were expressed in vessels and neurons of the P7 rat brain. Increased angiogenesis and upregulated expression of VEGF-A and VEGFR-2, but not VEGFR-1, was also found in vessels and neurons in the ipsilateral cerebral cortex 24 h after carotid artery ligation. A blockade of VEGF-A or VEGFR-2, instead of VEGFR-1, by antisense oligodeoxynucleotides decreased VEGFR-2 and pCREB expression and abolished the neuroprotective effect of carotid artery ligation preconditioning. In contrast, VEGF-A treatment or selective activation of VEGFR-2 before hypoxic-ischemia selectively upregulated VEGFR-2 and pCREB expression and provided neuroprotection against neonatal hypoxic-ischemic brain injury. Furthermore, selective activation of VEGFR-2 but not VEGFR-1 after hypoxic-ischemia also significantly protected P7 rat pups against hypoxic-ischemic brain injury. Further in vitro oxygen-glucose deprivation (OGD) study confirmed that VEGFR-2 and CREB activation was required for VEGF-A-induced neuroprotection against oxygen glucose deprivation neuronal death in differentiated H19-7 cells.
Taken together, these in vivo and in vitro evidences suggest that VEGF-A/VEGFR-2 signaling leading to CREB activation is an important event in neuroprotection against hypoxic-ischemic injury in the neonatal brain. Pharmacological activation of VEGFR-2 might be an important strategy for the treatment of neonatal hypoxic-ischemic brain injury.
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