Effects and mechanisms of astragaloside A treatment on sodium iodate-induced photoreceptor degeneration
Objective To investigate the effect of astragaloside A(AS-A)on the photoreceptor degeneration induced by sodium iodate(NaIO3)and its related mechanism.Methods Sixty healthy male C57BL/6J mice,aged 6-8 weeks,were randomly divided into normal control(NC)group,NaIO3 group,and AS-A group,with twenty mice in each group.30 min before modeling,AS-A group mice were intraperitoneally injected with 100 μl AS-A at a dose of 100 mg/kg body weight.30 min later,mice in NaIO3 group and AS-A group were intraperitoneally injected with 100 μl NaIO3 at a dose of 30 mg/kg body weight.Subsequently,AS-A group mice were administered AS-A twice daily at 12 h intervals until the end of the experiment.On day 1 post-modeling,zonula occludens-1(ZO-1)immunohistochemistry was performed to observe the structure of retinal pigment epithelium(RPE)cells;real-time quantitative polymerase chain reaction(qPCR)was conducted to detect the mRNA expression of various retinal chemokine ligand-2(Ccl2),interleukin-1 beta(Il-1β),mixed lineage kinase domain-like protein(Mlkl),receptor-interacting protein kinase 3(Ripk3),and tumor necrosis factor(Tnf).On day 3 post-modeling,immunohistochemistry was performed to observe the expression of ionized calcium binding adaptor molecule 1(Ibal)and glial fibrillary acid protein(GFAP)in the retina;TdT-mediated dUTP nick-end labeling(TUNEL)assay was used to detect photoreceptor cell death in each group.On day 4 post-modeling,fundus morphology of mice in each group was observed by fundus color photography and optical coherence tomography(OCT).Hematoxylin-eosin staining(HE)was used to observe the morphological structure of the retina in each group.Inter-group comparisons between two groups were conducted using independent samples t-test,while comparisons among three groups were performed using one-way ANOVA.Results Fundus color photography and OCT examination showed that a large number of scattered yellow-white subretinal nodular structures in the fundus of NaIO3 group mice,and a large number of strong reflection areas in the RPE layer.The number of strong reflection areas in the RPE layer was reduced in the AS-A group.Immunohistochemical analysis of ZO-1 showed that ZO-1 was largely lost on the RPE cell membrane in that NaIO3 group;whereas in the AS-A group,ZO-1 was evenly distributed on the RPE cell membrane.HE staining results showed circular black deposits were visible in the RPE layer of the NaIO3 group,and the inner and outer segments of photoreceptors were severely damaged,with a significant decrease in the number of outer nuclear layer(ONL)cell nuclei;whereas in the AS-A group,the RPE layer pigments were orderly,the inner and outer segments of photoreceptors were intact,and the number of ONL cell nuclei significantly increased.The results of TUNEL staining show that numerous TUNEL-positive cell nuclei were observed in the ONL of the retina in the NaIO3 group,while the number of TUNEL-positive cell nuclei in the ONL of the retina was significantly reduced in the AS-A group,with statistically significant differences(t=2.66,P＜0.05).The analysis of qPCR data showed that compared with the AS-A group,the relative expression levels of Mlkl,Ripk3,Ccl2,Il-1β and Tnf mRNA in the retina were significantly increased in the NaIO3 group,with statistically significant differences(F=39.18,10.66,53.51,41.40,24.13;P＜0.001).Immunohistochemical staining results showed that compared with NC group and AS-A group,the positive expression of GFAP in retina of NaIO3 group was significantly increased,and the difference was statistically significant(F=9.62,P＜0.05).Conclusion AS-A antagonizes NaIO3-induced photoreceptor degeneration in part by inhibiting photoreceptor cell death and neuroinflammation.Meanwhile,AS-A treatment protects against NaIO3-triggered perturbation of retinal homeostasis.
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