The mechanism of N-acetylserotonin regulating microglial polarization via NOD1/Rip2 pathway in rats after retinal ischemia reperfusion
10.3760/cma.j.cn511434-20231007-00408
- VernacularTitle:基于核苷酸结合的寡聚结构域1/受体相互作用蛋白2通路探讨N-乙酰-5-羟色胺调控大鼠视网膜缺血再灌注后小胶质细胞极化的机制
- Author:
Ying XU
1
;
Jianliang LIU
;
Yuze ZHAO
;
Chenxu WANG
;
Xinhao FU
;
Xiaoshuang LI
;
Xiaoli WANG
;
Yansong ZHAO
Author Information
1. 潍坊医学院附属医院眼科，潍坊 261031
- Keywords:
Retina ischemia-reperfusion injury;
N-acetylserotonin;
Retinal microglia;
Nucleotide-binding and oligomerization domain;
Animal experiment
- From:
Chinese Journal of Ocular Fundus Diseases
2024;40(4):287-295
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effect of N-acetylserotonin (NAS) on the retinal microglia polarization in retinal ischemia-reperfusion injury (RIRI) rats and explore its mechanism via nucleotide-bound oligomeric domain 1 (NOD1)/receptor interacting protein 2 (Rip2) pathway.Methods:Healthy male Sprague Dawley rats were randomly divided into Sham ( n=21), RIRI ( n=21) and NAS (injected intraperitoneally 30 min before and after modeling with NAS, 10 mg/kg, n=18) groups, using random number table. And the right eye was used experimental eye. The RIRI model of rats in RIRI group and NAS group was established by anterior chamber high intraocular pressure method. Rats in NAS group were intraperitoneally injected with 10 mg/kg NAS before and 30 min after modeling, respectively. The retinal morphology and the number of retinal ganglion cell (RGC) in each group were detected by hematoxylin-eosin staining and immunohistochemical staining. The effect of NAS on polarization of retinal microglia was detected by immunofluorescence staining. Transcriptome sequencing technology was used to screen out the differentially expressed genes between Sham and RIRI groups. Western blot and real-time quantitative polymerase chain reaction (RT-PCR) were used to examine the differentially expressed genes. Immunohistochemical staining, Western blot and RT-PCR were used to investigate the effect of NAS on the expression of NOD1 and Rip2 protein and mRNA in retinal tissue and microglia of rats. General linear regression analysis was performed to determine the correlation between the number difference of NOD1 + cells and the number difference of M1 and M2 microglia in retinal tissues of rats in NAS group and RIRI group. Results:A large number of RGC were observed in the retina of rats in Sham group. 24 h after modeling, compared with Sham group, the inner retinal thickness of rats in RIRI group was significantly increased and the number of RGC was significantly decreased. The thickness of inner retina in NAS group was significantly thinner and the number of RGC was significantly increased. Compared with Sham group, the number of retinal microglia of M1 and M2 in RIRI group was significantly increased. Compared with RIRI group, the number of M1 microglia decreased significantly and the number of M2 microglia increased significantly in NAS group. There was statistical significance in the number of M1 and M2 microglia in the retina of the three groups ( P<0.05). Transcriptome sequencing results showed that retinal NOD1 and Rip2 were important differential genes 24 h after modeling. The mRNA and protein relative expressions of NOD1 and Rip2 in retina of RIRI group were significantly higher than those of Sham group, with statistical significance ( P<0.05). The number of NOD1 + and Rip2 + cells and the relative expression of mRNA and protein in retinal microglia in RIRI group were significantly higher than those in Sham group, and NAS group was also significantly higher than that in Sham group, but lower than that in RIRI group, with statistical significance ( P<0.05). The number of Iba-1 +/NOD1 + and Iba-1 +/Rip2 + cells in retinal microglia in RIRI group was significantly increased compared with that in Sham group, and the number of Iba-1 +/Rip2 + cells in NAS group was significantly decreased compared with that in RIRI group, but still significantly higher than that in Sham group, with statistical significance ( P<0.05). Correlation analysis results showed that the difference of retinal NOD1 + and Rip2 + cells in NAS group and RIRI group was positively correlated with that of M1 microglia ( r=0.851, 0.895), and negatively correlated with that of M2 microglia ( r=-0.797, -0.819). The differences were statistically significant ( P<0.05). Conclusion:NAS can regulate the microglial polarization from M1 to M2 phenotype, the mechanism is correlated with the NOD1/Rip2 pathway.
