Additional file 1 of Microglia-derived exosomes modulate myelin regeneration via miR-615-5p/MYRF axis
Creators: name: Ji, Xiao-Yu
name: Guo, Yu-Xin
name: Wang, Li-Bin
name: Wu, Wen-Cheng
name: Wang, Jia-Qi
name: He, Jin
name: Gao, Rui
name: Rasouli, Javad
name: Gao, Meng-Yuan
name: Wang, Zhen-Hai
name: Xiao, Dan
name: Zhang, Wei-Feng
name: Ciric, Bogoljub
name: Zhang, Yuan
name: Li, Xing
Publisher: name: figshare
Year: 2024
Subjects: subject: Medicine
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: Molecular Biology
subject: Neuroscience
subject: Pharmacology
subject: Immunology
subject: FOS: Clinical medicine
subject: Biological Sciences not elsewhere classified
subject: Developmental Biology
subject: Cancer

description: Additional file 1: Figure S1. The map of plasmid. A MYRF 3′UTR-Luciferase vector. B Mut MYRF 3′UTR-Luciferase vector. Figure S2. The expression of MBP and Iba1 in MS Lesion. A Immunofluorescence staining of MBP and Iba1 in brain of NAWM and MS lesion and B, C MBP fluorescence intensity and the number of Ibal+ cells per field. Scale bar = 100 μm. All data are represented by mean ± SEM (n = 5, each group). One-way ANOVA was used to determine P values (B, C). ****p &lt; 0.0001. Figure S3. The expression of PDGFRα and MYRF. A After co-incubation of OPCs with supernatant, the expression of PDGFRα and MYRF and B the percentage of PDGFRα+MYRF+ cells were detected by immunofluorescence staining. C Immunofluorescence staining to determine the PDGFRα and MYRF expression intensity after OPCs treatment with EXOs/EXOs-LPS, and D the percentage of PDGFRα + MYRF + cells. One-way ANOVA was used to determine P values. *P &lt; 0.05, **P &lt; 0.01, ****P &lt; 0.0001. Scale bar = 50 μm. Figure S4. Expression of miR-615-5p in EXOs/EXOs-LPS. A, B After exosomes were captured by CD81/CD9 chip, miR-615-5p was detected in EXOs/EXOs-LPS by Exoview. In addition, surface markers CD63 and CD9 of EXOs/EXOs-LPS were detected by Exoview. C, D ExoView detected the average colocalization percent of EXOs or EXOs-LPS. Figure S5. The co-expression of miR-615-5p and PDGFRα, APC, GFAP and NeuN. A Immunofluorescence staining and FISH detected the expression of PDGFRα and miR-615-5p in EAE/naïve spinal cords, and B the number of PDGFRα+miR-615-5p+ cells per field. C Immunofluorescence staining and FISH detected the expression of APC and miR-615-5p in EAE/naïve spinal cords, and D the number of APC +miR-615-5p+ cells per field. E Immunofluorescence staining and FISH detected the expression of GFAP and miR-615-5p in EAE/naïve spinal cords, and F the number of GFAP+miR-615-5p+ cells per field. G Immunofluorescence staining and FISH detected the expression of NeuN and miR-615-5p in EAE/naïve spinal cords, and H the number of NeuN +miR-615-5p+ cells per field. Scale bar = 50 μm. All data are represented by mean ± SEM (n = 5, each group). One-way ANOVA was used to determine P values. Figure S6. Expression of miR-615-5p. A The expression of miR-615-5p was detected by qRT-PCR in the brain and spinal cords of naïve/EAE mice. B The expression of miR-615-5p was detected by qRT-PCR in SM and SM-LPS. C The expression of miR-615-5p was detected by qRT-PCR in microglia, activated microglia, OPCs, and activated OPCs. D Raw264.7 was stimulated with 100 ng/ml LPS. FISH detected the expression of miR-615-5p in Raw264.7 and Raw264.7-LPS, and E the miR-615-5p fluorescence intensity. All data are mean ± SEM. t-tests were used to determine P values (B, E). One-way ANOVA was used to determine P values (C). Two-way ANOVA was used to determine p values (A). *P &lt; 0.05, **P &lt; 0.01, ***P &lt; 0.001, ****P &lt; 0.0001. One representative of three independent experiments is shown. Figure S7. Expression of miR-615-5p target genes. A–F After OPCs were incubated with SM-derived supernatant/SM-LPS-derived supernatant, qRT-PCR was used to detect the other target genes of miR-615-5p, including Plp2, Tcf7l2, Cdk5, Sox6. T-test was used to determine P values. **P &lt; 0.01, ****P &lt; 0.0001. One representative of three independent experiments is shown. Figure S8. Antagonistic miR-615-5p inhibited the expression of GFAP and APP. A Immunofluorescence staining of GFAP and APP in spinal cords of naïve and EAE mice, and B, C the number of GFAP+ cells and APP+ cells per field. Scale bar = 100 μm. All data are represented by mean ± SEM (n = 10, each group). One-way ANOVA was used to determine P values (B, C). ****p &lt; 0.0001. One representative of three independent experiments is shown. Table S1. Primers for PCR. Table S2. Primers for qRT-PCR. Table S3. miR-615-5p target gene network expression table.