Journal: Molecular Therapy

Article Title: Bone marrow mesenchymal stem cell-derived exosomal miR-34c-5p ameliorates RIF by inhibiting the core fucosylation of multiple proteins

doi: 10.1016/j.ymthe.2021.10.012

Figure Lengend Snippet: MSCs inhibited CF and pericyte activation in RIF (A) FUT8 levels were assessed using western blotting. The bottom panel shows the quantification. Scale bar, 50 μm. Data are the mean ± SD (n = 3). Control, pericytes without TGFβ1 stimulation. TGFβ1+MSCs, 48 h after TGFβ1 stimulation of pericytes, MSCs were added immediately after the TGFβ1 stimulation of pericytes. (B) FUT8 activity examined by HPLC. Data are the mean ± SD (n = 3); S indicates the peptide substrate; P is the fucosylation product. (C) Representative images of PDGFRβ (green) and LCA (red) staining, and α-SMA (green) and LCA (red) staining. The bottom panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 3). (D) Representative morphological alterations in pericytes. Scale bar, 200 μm. (E) Representative images of PDGFRβ (green) and α-SMA (red) staining. The right panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 3). α-SMA levels were assessed using western blotting. The right panel shows the quantification. Scale bar, 50 μm. Data are the mean ± SD (n = 3). TGFβ1+MSCs, 48-h TGFβ1–stimulated pericytes plus MSCs. (F) Representative images of PDGFRβ (green) and LCA (red) staining, and α-SMA (green) and LCA (red) staining. The bottom panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 6). Control, C57 mice without UUO surgery. UUO+MSCs, 7 days after mice had been injected with MSCs via the tail vein immediately after UUO surgery. (G) Representative images of PDGFRβ (green) and CD31 (red) staining, and PDGFRβ (green) and α-SMA (red) staining. The bottom panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 6). The arrows represent the pericytes leaving the endothelium. (H) Representative images of Masson's trichrome and PAS staining of UUO mouse kidney sections. The bottom panel shows the quantification. Scale bar, 200 μm. Data are the mean ± SD (n = 6).

Article Snippet: CD81–EGFR complex formation aided miR-34c-5p entry into pericytes to downregulate CF (A) (Top) Exosome and pericyte proteomics; (bottom) protein interaction network. (B) Computer simulation of CD81-EGFR molecular docking. (C) Representative images of EGFR (green) and CD81 (red) staining both in vitro (n = 3) and in vivo (n = 6).

Techniques: Activation Assay, Western Blot, Control, Activity Assay, Staining, Injection

Journal: Molecular Therapy

Article Title: Bone marrow mesenchymal stem cell-derived exosomal miR-34c-5p ameliorates RIF by inhibiting the core fucosylation of multiple proteins

doi: 10.1016/j.ymthe.2021.10.012

Figure Lengend Snippet: Exosomes accumulated in activated pericytes and obstructed kidney both in vitro and in vivo (A) (Left) Representative electron micrograph of exosomes. Scale bar, 100 μm. (Middle) Nanoparticle tracking analysis. (Right) CD63 and CD81 levels were assessed using western blotting. Scale bar, 50 μm (n = 3). (B) The exosome content of MSC-CM was detected by luciferase reporter. Data are the mean ± SD (n = 3). (C) Representative images of PKH67 (green) and PDGFRβ (red) staining in vitro . The right panel shows the quantification. Scale bar, 75 μm. Control, pericytes without TGFβ1 stimulation. 0 h, 24 h, 48 h, pericytes without TGFβ1 stimulation for 0 h, 24 h, 48 h. (D) Ex vivo UUO mouse imaging at 0 h, 6 h, 12 h, 24 h, 72 h, 168 h, and 240 h.

Article Snippet: CD81–EGFR complex formation aided miR-34c-5p entry into pericytes to downregulate CF (A) (Top) Exosome and pericyte proteomics; (bottom) protein interaction network. (B) Computer simulation of CD81-EGFR molecular docking. (C) Representative images of EGFR (green) and CD81 (red) staining both in vitro (n = 3) and in vivo (n = 6).

Techniques: In Vitro, In Vivo, Western Blot, Luciferase, Staining, Control, Ex Vivo, Imaging

Journal: Molecular Therapy

Article Title: Bone marrow mesenchymal stem cell-derived exosomal miR-34c-5p ameliorates RIF by inhibiting the core fucosylation of multiple proteins

doi: 10.1016/j.ymthe.2021.10.012

Figure Lengend Snippet: MSCs inhibited pericyte activation and RIF mainly through secreting exosomes (A) FUT8 levels were assessed using western blotting. The bottom panel shows the quantification. Scale bar, 50 μm. Data are the mean ± SD (n = 3). Control, pericytes without TGFβ1 stimulation. TGFβ1+MSCs, 48-h TGFβ1-stimulated pericytes plus MSCs; TGFβ1+Exo, 48-h TGFβ1-stimulated pericytes plus exosomes; TGFβ1+CM, 48-h TGFβ1-stimulated pericytes plus CM; TGFβ1+CM(−)Exo, 48-h TGFβ1-stimulated pericytes plus CM(−)Exo. (B) FUT8 activity examined by HPLC. Data are the mean ± SD (n = 3); S indicates the peptide substrate; P is the fucosylation product. (C) Representative images of PDGFRβ (green) and LCA (red) staining, and α-SMA (green) and LCA (red) staining. The right panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 3). (D) Representative morphological alterations in pericytes. Scale bar, 200 μm. (E) Representative images of PDGFRβ (green) and α-SMA (red) staining. The bottom panel shows the quantification. Scale bar, 75 μm. α-SMA levels were assessed using western blotting. The right panel shows the quantification. Scale bar, 50 μm. Data are the mean ± SD (n = 6). (F) Representative images of PDGFRβ (green) and LCA (red) staining, and α-SMA (green) and LCA (red) staining. The bottom right panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 6). Control, C57 mice without UUO surgery. UUO+MSCs, 7 days after MSC injection into UUO mouse tail vein; UUO+Exo, 7 days after Exo injection into UUO mouse tail vein; UUO+CM, 7 days after CM injection into UUO mouse tail vein; UUO+CM(−)Exo, 7 days after CM(−)Exo injection into UUO mouse tail vein. (G) Representative images of PDGFRβ (green) and CD31 (red) staining, and PDGFRβ (green) and α-SMA (red) staining. The bottom panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 6). (H) Representative images of Masson's trichrome and PAS staining of UUO mouse kidney sections. The bottom panel shows the quantification. Scale bar, 200 μm. Data are the mean ± SD (n = 6).

Article Snippet: CD81–EGFR complex formation aided miR-34c-5p entry into pericytes to downregulate CF (A) (Top) Exosome and pericyte proteomics; (bottom) protein interaction network. (B) Computer simulation of CD81-EGFR molecular docking. (C) Representative images of EGFR (green) and CD81 (red) staining both in vitro (n = 3) and in vivo (n = 6).

Techniques: Activation Assay, Western Blot, Control, Activity Assay, Staining, Injection

Journal: Molecular Therapy

Article Title: Bone marrow mesenchymal stem cell-derived exosomal miR-34c-5p ameliorates RIF by inhibiting the core fucosylation of multiple proteins

doi: 10.1016/j.ymthe.2021.10.012

Figure Lengend Snippet: MSC-derived exosomes inhibited pericyte activation mainly by regulating CF (A) (Left) RT-PCR, (middle) representative images of LCA (red) staining, (right) FUT8 levels assessed by western blotting. The bottom panel shows the quantification. Data are the mean ± SD (n = 3). (B) FUT8 level was assessed by western blotting. The bottom panel shows the quantification. Scale bar, 50 μm. Data are the mean ± SD (n = 3). (C) FUT8 activity tested by HPLC. Data are the mean ± SD (n = 3); S indicates the peptide substrate; P is the fucosylation product. (D) Representative images of PDGFRβ (green) and LCA (red) staining, and α-SMA (green) and LCA (red) staining. The bottom panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 3). (E) Representative morphological alterations in pericytes. Scale bar, 200 μm. (F) Representative images of PDGFRβ (green) and α-SMA (red) staining. The bottom panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 3). α-SMA level was assessed by western blotting. The bottom panel shows the quantification. Scale bar, 50 μm. Data are the mean ± SD (n = 3).

Article Snippet: CD81–EGFR complex formation aided miR-34c-5p entry into pericytes to downregulate CF (A) (Top) Exosome and pericyte proteomics; (bottom) protein interaction network. (B) Computer simulation of CD81-EGFR molecular docking. (C) Representative images of EGFR (green) and CD81 (red) staining both in vitro (n = 3) and in vivo (n = 6).

Techniques: Derivative Assay, Activation Assay, Reverse Transcription Polymerase Chain Reaction, Staining, Western Blot, Activity Assay

Journal: Molecular Therapy

Article Title: Bone marrow mesenchymal stem cell-derived exosomal miR-34c-5p ameliorates RIF by inhibiting the core fucosylation of multiple proteins

doi: 10.1016/j.ymthe.2021.10.012

Figure Lengend Snippet: Exosomes downregulated CF by delivering miR-34c-5p during pericyte activation and RIF (A) (Left) Heatmap of exosomal miRNA and (right) histogram of miR-34c-5p at 0 h, 24 h, and 48 h. Data are the mean ± SD (n = 3). (B) Luciferase reaction between FUT8 and miR-34c-5p. Data are the mean ± SD (n = 3). (C) Representative morphological alterations in pericytes. Scale bar, 200 μm. Also shown are representative images of PDGFRβ (green) and α-SMA (red) staining, and α-SMA (green) and LCA (red) staining in vitro . The right panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 3). (D) Representative images of Masson's trichrome and PAS staining of UUO mouse kidney sections. Scale bar, 200 μm. Also shown are representative images of PDGFRβ (green) and α-SMA (red) staining, and α-SMA (green) and LCA (red) staining in vivo . The right panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 6).

Article Snippet: CD81–EGFR complex formation aided miR-34c-5p entry into pericytes to downregulate CF (A) (Top) Exosome and pericyte proteomics; (bottom) protein interaction network. (B) Computer simulation of CD81-EGFR molecular docking. (C) Representative images of EGFR (green) and CD81 (red) staining both in vitro (n = 3) and in vivo (n = 6).

Techniques: Activation Assay, Luciferase, Staining, In Vitro, In Vivo

Journal: Molecular Therapy

Article Title: Bone marrow mesenchymal stem cell-derived exosomal miR-34c-5p ameliorates RIF by inhibiting the core fucosylation of multiple proteins

doi: 10.1016/j.ymthe.2021.10.012

Figure Lengend Snippet: CD81–EGFR complex formation aided miR-34c-5p entry into pericytes to downregulate CF (A) (Top) Exosome and pericyte proteomics; (bottom) protein interaction network. (B) Computer simulation of CD81-EGFR molecular docking. (C) Representative images of EGFR (green) and CD81 (red) staining both in vitro (n = 3) and in vivo (n = 6). The right panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD. (D) Representative images of α-SMA (green), LCA (red), and miR-34c-5p (fam) staining in vitro . The right panel shows the quantification. Scale bar, 75 μm. (E) Representative images of α-SMA (green), LCA (red), and miR-34c-5p (fam) staining in vivo . The right panel shows the quantification. Scale bar, 75 μm. Data are the mean ± SD (n = 6).

Article Snippet: CD81–EGFR complex formation aided miR-34c-5p entry into pericytes to downregulate CF (A) (Top) Exosome and pericyte proteomics; (bottom) protein interaction network. (B) Computer simulation of CD81-EGFR molecular docking. (C) Representative images of EGFR (green) and CD81 (red) staining both in vitro (n = 3) and in vivo (n = 6).

Techniques: Staining, In Vitro, In Vivo