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Objective To investigate the effects of astragaloside Ⅳ (AS-Ⅳ) on axon growth inhibitory factor A (Nogo-A) and its downstream pathway protein RHO-associated coiled spiral kinase 2 (ROCK2) in experimental autoimmune encephalomyelitis (EAE) mice, and to explore the mechanism by which it promotes axon repair and regeneration. Methods EAE model was induced in C57BL/6 female mice by subcutaneous injection of myelin oligodendrocyte glycoprotein 35-55 (MOG35-55). Mice were randomly divided into EAE group and AS-Ⅳ group (n=8 per group). EAE group received intraperitoneal injection of PBS on the 3rd day post-immunization, while AS-Ⅳ group was administered AS-Ⅳ at a dosage of 30mg/(kg.d) once daily, 0.2 ml per injection, for 25 consecutive days. On the 28th day post-immunization, the expression levels of growth-associated protein 43 (GAP-43), neuronal core antigen (NeuN), microtubule associated protein 2 (MAP-2), glial fibroacidic protein (GFAP), and Iba1 in the spinal cord were detected using immunofluorescence assay. Real-time fluorescence quantitative PCR (qRT-PCR) was conducted to detect mRNA expression levels of GAP-43, Nogo-A, and Nogo receptor (NgR) genes. Western blotting was utilized to determine the expression levels of GAP-43, Nogo-A, ROCK2, phosphorylated myosin phosphatase (p-MYPT1), B-lymphoblastoma-2 (Bcl-2), and Bcl-2 associated X protein (Bax). Results Compared with EAE group, AS-Ⅳ treatment significantly reduced the positive cell expression rates of Iba1 microglia and GFAP astrocyte in spinal cord (P<0.01 and P<0.001,respectively), while it also increased the positive expression rates of NeuN and MAP-2 (P<0.001 and P<0.05, respectively). The treatment also upregulated the expression level of anti-apoptotic factor Bcl-2 (P<0.001) and downregulated the expression level of pro-apoptotic factor Bax (P<0.05), leading to an increase in Bcl-2/Bax ratio (P<0.05). Furthermore, AS-Ⅳ enhanced the expression of GAP-43 protein (P<0.05) and decreased the mRNA expression levels of neuroregeneration inhibitor Nogo receptor (NgR) and ROCK2 gene (P<0.001, P<0.05, respectively); as well as decreased the expression levels of Nogo-A, ROCK2 and p-MYPT1 proteins (P<0.05, P<0.001). Conclusion AS-IV may inhibit the activation of microglia and astrocytes and neuronal apoptosis in EAE mice by inhibiting Nogo-A and downstream pathway ROCK2, thereby promoting the expression of GAP-43, NeuN and MAP-2, alleviating neuronal damage, and facilitating axon repair and regeneration.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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