Journal: Neural Regeneration Research

Article Title: EZH2-dependent myelination following sciatic nerve injury

doi: 10.4103/NRR.NRR-D-23-02040

Figure Lengend Snippet: In situ hybridization spatial mRNA validation of Ezh2 expression in sciatic nerve of rats. (A) Schematic illustration of sciatic nerve samples obtained from normal, proximal, and distal segments of a rat sciatic nerve transection (SNT) model. Created with Adobe Illustrator CC and Adobe Photoshop CC. (B) Immunofluorescence staining of Schwann cells (SCs) on sciatic nerve or transected nerve probed for Ezh2 mRNA (green) at 4 days post-injury. All SCs were identified using S100B (Cy3, red), neurofilament 200 (NF200; Cy5, grey), and 4′,6-diamidino-2-phenylindole (DAPI; blue). Scale bars: 50 µm for low magnification, 20 µm for high magnification. (C) Percentage of SCs with Ezh2 mRNA ≥ 1 visible spots. (D) Fold change quantification of Ezh2 mRNA per SC. Ratio of number of average spots per SC were calculated for all groups, and then standardized to the normal sciatic nerve group value. SNT may induce expression of Ezh2 in SCs of both sciatic nerve stumps compared with the normal group. Data are expressed as mean ± SE ( n = 6). ** P < 0.01, vs . normal group (one-way analysis of variance followed by Bonferroni post hoc test).

Article Snippet: The rat Ezh2 probe (RNAscope TM Probe-Rn-Ezh2 cat no. 816741) and all reagents were purchased from ACDBio ( https://acdbio.com/ ).

Techniques: In Situ Hybridization, Biomarker Discovery, Expressing, Immunofluorescence, Staining

Journal: Neural Regeneration Research

Article Title: EZH2-dependent myelination following sciatic nerve injury

doi: 10.4103/NRR.NRR-D-23-02040

Figure Lengend Snippet: Ablation of Ezh2 in Schwann cells of mice leads to abnormal growth and myelination deficits in the sciatic nerve. (A) Diagram outlining the generation of Ezh2 conditional knockout ( Ezh2 fl/fl ; Mpz -Cre) mice. (B) Representative images of wild-type ( Ezh2 fl/fl ), heterozygous ( Ezh2 fl/+ ; Mpz -Cre), and homozygous conditional knockout mice ( Ezh2 fl/fl ; Mpz -Cre) at 7, 14, 21 days, and 8 weeks after birth. (C) Sciatic nerve without injury in wild-type control and Ezh2 fl/fl ; Mpz -Cre mice at 8 weeks post-birth stained for S100B (Cy5, grey), EZH2 (Cy3, red), MPZ (Alexa Fluor 488, green), and DAPI (blue). Genetic ablation of Ezh2 in Schwann cells leads to abnormal growth and deficits in the myelination of mouse sciatic nerve. Scale bars: 10 µm. DAPI: 4,6-Diamidino-2-phenylindole; MPZ: myelin protein zero.

Article Snippet: The rat Ezh2 probe (RNAscope TM Probe-Rn-Ezh2 cat no. 816741) and all reagents were purchased from ACDBio ( https://acdbio.com/ ).

Techniques: Knock-Out, Control, Staining

Journal: Neural Regeneration Research

Article Title: EZH2-dependent myelination following sciatic nerve injury

doi: 10.4103/NRR.NRR-D-23-02040

Figure Lengend Snippet: Knockout of Ezh2 in Schwann cells leads to hypomyelination of mouse sciatic nerve and suppresses remyelination post-SNC. (A) Transmission electron microscopy analysis of sciatic nerves from both Ezh2 cKO mice ( Ezh2 fl/fl ; Dhh -Cre and Ezh2 fl/fl ; Mpz -Cre) showed plenty of axons remain unmyelinated in mutant sciatic nerves. (B) Transmission electron microscopy analysis showed most axons remyelinated at the crush site of the control nerve while Ezh2 fl/fl ; Mpz -Cre mice had significantly fewer remyelinated axons than control nerve at 21 days after SNC. (C) Quantification of myelinated axons in sciatic nerves among both Ezh2 cKO mice and their wild-type littermate control mice at 8 weeks after birth and 21 days after SNC. Data are expressed as mean ± SE ( n = 3). * P < 0.05, *** P < 0.001, vs . wildtype control ( Ezh2 fl/fl ) group (one-way analysis of variance followed by Bonferroni post hoc test). (D) The percentage of myelinated axons in sciatic nerves among both Ezh2 cKO mice and wild-type control mice at 8 weeks after birth (P8W) and 21 days after SNC (21 dpi). (E) Sciatic nerve from wild-type control ( Ezh2 fl/fl ) and Ezh2 fl/fl ; Dhh -Cre P8W mice at 14 days post sciatic nerve crush (14 dpi) stained for S100B (Cy5, grey), EZH2 (Cy3, red), MPZ (Alexa Fluor 488, green), and DAPI (blue). Scale bars: 10 µm. cKO: Conditional knockout; DAPI: 4,6-diamidino-2-phenylindole; MPZ: myelin protein zero; SNC: sciatic nerve crush.

Article Snippet: The rat Ezh2 probe (RNAscope TM Probe-Rn-Ezh2 cat no. 816741) and all reagents were purchased from ACDBio ( https://acdbio.com/ ).

Techniques: Knock-Out, Transmission Assay, Electron Microscopy, Mutagenesis, Control, Staining

Journal: Neural Regeneration Research

Article Title: EZH2-dependent myelination following sciatic nerve injury

doi: 10.4103/NRR.NRR-D-23-02040

Figure Lengend Snippet: Interaction between PRC2 family members and specific myelin–related genes in a sciatic nerve crush segment and both sciatic nerve stumps post-transection. Protein–protein interaction networks of myelin-related genes showing that SOX2, insulin-like growth factor 1 (IGF1), and signal transducer and activator of transcription 3 (STAT3) were hub genes in the proximal sciatic nerve stump, while MYC was the hub gene in the distal sciatic nerve stump post-transection. Tumor necrosis factor (TNF) was the hub gene in the sciatic nerve crush segment post-crush. PRC2 complex members interacted with specific myelin-related PPI networks in a sciatic nerve crush segment and both sciatic nerve stumps post-transection. EZH2 (red) was the hub gene (blue) in sciatic nerve crush segments via interactions with TNF, interleukin (IL)-1, N-Myc downregulated gene 1 (NDRG1), Erb-B2 receptor tyrosine kinase 4 (ERBB4), chemokine (C-X-C motif) receptor 3 (CXCR3), chemokine (C-X-C motif) receptor 4 (CXCR4), hepatocyte nuclear factor-4 alpha (HNF4A), IL-12B, and IL-12 receptor subunit beta 2 (IL12RB2) (dark yellow). EZH2 (red) interacted with specific myelin-related PPI networks (blue) in proximal sciatic nerve stump post-transection via interferon beta 1 (IFNB1), IGF1, STAT3, and O-linked N-acetylglucosamine transferase (OGT) (green). EZH2 (red) interacted with specific myelin-related PPI networks (cyan) in distal sciatic nerve stump post-transection via C-X-C motif chemokine ligand 2 (CXCL2), colony stimulating factor 3 (CSF3), and MYC (dark yellow). PRC2: Polycomb repressive complex 2.

Article Snippet: The rat Ezh2 probe (RNAscope TM Probe-Rn-Ezh2 cat no. 816741) and all reagents were purchased from ACDBio ( https://acdbio.com/ ).

Techniques: