Orexin A alleviates chronic cerebral hypoperfusion-induced neuroinflammation and cognitive dysfunction by inhibiting the NEK7/NLRP3 pathway

Objective: Chronic cerebral hypoperfusion (CCH)-induced neuroinflammation significantly impacts the functional prognosis of patients with vascular dementia (VaD). Microglial neuroinflammation is significantly aggravated by the chronic activation of the NLRP3 inflammasome, which has emerged as a major contributing factor. Our previous research indicated that Orexin A effectively alleviates acute inflammatory responses and neurological deficits following brain injury. However, its neuroprotective role in cognitive function recovery after CCH remains to be elucidated.

Methods: Adult male SD rats underwent permanent bilateral common carotid artery occlusion surgery for 8 weeks to establish a VaD model. Subsequently, the rats received 4 weeks of continuous intranasal Orexin A treatment (250 μg/kg). Additionally, in order to explore the potential mechanisms and neuroprotective roles of Orexin A, BV2 cells were subjected to hypoxia to simulate in vitro CCH stimulation, either with or without Orexin A pretreatment, and were then co-cultured with HT22 neurons.

Results: After 8 weeks of modeling, we noted a significant decrease in Orexin A and OXR1 expression in the hippocampus of CCH rats, which was accompanied by pronounced cognitive impairments. Furthermore, CCH exposure resulted in prolonged activation of the NLRP3 inflammasome and M1-type microglia within the hippocampus, as well as blood-brain barrier disruption and neurodegenerative changes. Orexin A treatment effectively ameliorated these alterations. In vitro experiments demonstrated hypoxia exposure promoted NLRP3 inflammasome activation in microglia, along with the release of pore-forming Gasdermin-D-NT and NINJ1, ultimately causing "bystander" neuronal Pyroptosis. Orexin A inhibited NLRP3 inflammasome activation in microglia, thereby promoting the transition of M1-type microglia to M2-type and mitigating neuronal Pyroptosis. These effects were abolished by NEK7 overexpression.

Conclusion: Our findings indicate that Orexin A restores the M1/M2 microglial balance by inhibiting the NEK7/NLRP3 pathway. This, in turn, alleviates neuroinflammation and neuronal Pyroptosis, ultimately improving cognitive dysfunction after CCH. This study enhances our understanding of the neuroprotective mechanisms of Orexin A, potentially offering a new therapeutic target for cognitive impairment following CCH.

Chronic cerebral hypoperfusion; Microglia; NEK7; NLRP3 inflammasome; Neuroregeneration; Orexin-A; Vascular dementia.