Additional file 1 of An optimized exosome production strategy for enhanced yield while without sacrificing cargo loading efficiency
Creators: name: Zhang, Rongxin
name: Bu, Te
name: Cao, Ruidan
name: Li, Zhelong
name: Wang, Chen
name: Huang, Bing
name: Wei, Mengying
name: Yuan, Lijun
name: Yang, Guodong
Publisher: name: figshare
Year: 2022
Subjects: subject: Biochemistry
subject: Space Science
subject: Cell Biology
subject: Molecular Biology
subject: Pharmacology
subject: Biotechnology
subject: Chemical Sciences not elsewhere classified
subject: Immunology
subject: FOS: Clinical medicine
subject: Information Systems not elsewhere classified
subject: Mathematical Sciences not elsewhere classified
subject: Developmental Biology
subject: Cancer

description: Additional file 1: Figure S1. qPCR analysis of knockdown efficiency. A-L AML12 cells were transfected with siRNA against genes of interest and relative expression of Rab33b (A), Nfs (B), Rab9 (C), Vps18 (D), Rab22a (E), Rab35 (F), Rab24 (G), Vps39 (H), Rab14 (I), Rab4 (J), Rab11a (K) and Tfeb (L). Gapdh served as an internal reference gene. Data are expressed as mean $$\pm$$ ± SEM of three independent experiments. * p &lt; 0.05 by t-test. Figure S2. qPCR analysis of Rab31 expression in AML12 cells with Rab4 knocked-down. AML12 cells were transfected with siRab4 or siNC and expression of Rab31 was analyzed by qPCR. Gapdh as an internal reference gene. Data are expressed as mean $$\pm$$ ± SEM of three independent experiments. * p &lt; 0.05 by t-test. Figure S3. Representative flow cytometry analysis of RCMPs uptake by AML12 cells. The AML12 cells were incubated with control or DiI-labeled RCMPs for 6 h, and DiI signal was analyzed by flow cytometry. Figure S4. Characterization of ExoBooster from AML12 cells. A Representative TEM images of the indicated exosomes from AML12 cells. B Size distribution of the indicated exosomes as analyzed by NTA. C Western blot analysis of the exosome inclusive and exclusive markers in AML12 cells and derived exosomes. GAPDH served as a loading control. Data shown are representatives from triplicate experiments. Figure S5. Fluorescence microscope analysis of ExoBooster biodistribution in vivo. Lower magnification images corresponding to Fig. 4E. Scale bar = 100 μm. Figure S6. Differential expression of LDLR protein in different tissues from mice. A–F Analysis of LDLR protein expression in lung (A), kidney (C), and heart (E) by western blot. Data shown are representative of 3 different experiments. Quantification analysis of western blot results by densitometry in A, C and E, respectively. Data are expressed as mean $$\pm$$ ± SEM. ns, no signification. *, p &lt; 0.05 by one-way ANOVA. Figure S7. Ldlr@ExoBooster treatment reduces cholesterol level in Ldlr−/− mice. A–D Plasma total triglyceride (A), total cholesterol (B), LDL cholesterol (C), and HDL cholesterol (D) in Ldlr−/− mice treated as indicated. n = 5. Data are expressed as mean $$\pm$$ ± SEM. *p &lt; 0.05 by one-way ANOVA. Figure S8. Biocompatibility of indicated exosomes. A Body weight change curve in mice with indicated treatments. B H&amp;E staining in various tissues. Harvested tissues were sectioned and stained with H&amp;E. No significant histology change was observed in different tissues treated with Ldlr@ExoCtrl or Ldlr@ExoBooster. Scale bar = 100 μm. Figure S9. qPCR analysis of siRab4 sequence in exosomes as indicated. NA, not available as Ct value larger than 38. U6 as an internal control. Data are expressed as mean $$\pm$$ ± SEM of three independent experiments. Table S1. Sequences of siRNA. Table S2. Sequences of PCR primers.