[Antagonist of cannabinoid receptor 1 improves nerve injury caused by chronic intermittent hypoxia by inhibiting the NLRP3 inflammasome pathway in mice].

Objective

To explore the regulatory mechanism of the targeted blockade of cannabinoid receptor 1 (CB1R) in relation to the neuroinflammatory response mediated by the NOD like receptor pyrin domain-containing 3 (NLRP3) inflammasome under chronic intermittent hypoxia (CIH) conditions.

Methods

Forty adult male C57BL/6 mice were randomly divided into five groups: a normoxic control group (NC group), a CIH for 4 weeks group (4wCIH group), a CIH for 6 weeks group (6wCIH group), a CIH+CB1R antagonist AM251 intervention for 4 weeks group (4wCIH+AM251 group), and a CIH+AM251 intervention for 6 weeks group (6wCIH+AM251 group). Hematoxylin and eosin (HE) staining was used to observe the morphological changes of brain tissue. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expression of CB1R mRNA. Western blotting was used to analyze the protein levels of the NLRP3 inflammasome, and the enzyme-linked immunosorbent assay (ELISA) method was used to determine the contents of IL-1β and IL-18. Statistical analysis was performed using GraphPad Prism 9.5.0 software.

Results

(1) Compared with the NC group, the CIH group displayed pathological changes characterized by neuronal degeneration and necrosis, nuclear pyknosis, and cytoplasmic hyperchromasia. These lesions were markedly attenuated in the CIH+AM251 group compared with the CIH group. (2) The relative expression levels of CB1R mRNA in the CIH groups were significantly higher than those in the NC group (all P<0.05), with the 6w CIH group showing higher levels than the 4w CIH group (P<0.05). Conversely, the 4w and 6w CIH+AM251 groups exhibited significantly lower relative expression levels compared with the corresponding CIH groups (all P<0.05).(3) Western blot analysis showed that NLRP3 protein expression in the CIH groups was significantly higher than that in the NC group (all P<0.05), and higher in the 6w CIH group than in the 4w CIH group (P<0.05). IL-1β protein expression levels in the 4w and 6w CIH groups were significantly elevated compared with the NC group [4w: (55.45±0.75) pg/ml vs. (49.85±1.41) pg/ml; 6w: (58.03±0.95) pg/ml vs. (49.85±1.41) pg/ml, both P<0.05]. Similarly, IL-18 protein expression levels in the 4w and 6w CIH groups were significantly higher than in the NC group [4w: (55.65±1.00) pg/ml vs. (50.34±1.61) pg/ml; 6w: (58.98±0.37) pg/ml vs. (50.34±1.61) pg/ml, both P<0.05]. Conversely, IL-1β protein expression in the 4w and 6w CIH+AM251 groups was significantly lower than in the corresponding CIH groups [4w: (52.82±0.94) pg/ml vs. (55.45±0.75) pg/ml, 6w: (52.58±0.52) pg/ml vs. (58.03±0.95) pg/ml, both P<0.05], and IL-18 protein expression was also significantly reduced [4w: (53.04±0.33) pg/ml vs. (55.65±1.0) pg/ml, 6w: (55.36±0.40) pg/ml vs. (58.98±0.37) pg/ml, both P<0.05].

Conclusion

Targeted blockade of CB1R effectively attenuates CIH-induced neuroinflammatory injury by regulating the activation pathway of the NLRP3 inflammasome, with protective effects that progressively accumulate over time.