Fig. 5: DNA damage and immune responses in HSPCs upon RNA-mediated base editing and RNP-mediated Cas9 treatment. | Nature Communications

Fig. 5: DNA damage and immune responses in HSPCs upon RNA-mediated base editing and RNP-mediated Cas9 treatment.

From: Base-editing-mediated dissection of a γ-globin cis-regulatory element for the therapeutic reactivation of fetal hemoglobin expression

Fig. 5

a C-G to T-A or A-T to G-C base-editing efficiency, calculated by the EditR software in mobilized HD HSPC samples subjected to Sanger sequencing 6 days post-transfection. Data are expressed as mean ± SEM (n = 3 biologically independent experiments, 3 donors). b C-G to T-A or A-T to G-C base-editing efficiency, calculated by the EditR software in mobilized HD HSPC samples subjected to Sanger sequencing 12h-, 24h-, 48h-, and 6 days post-transfection. Data are expressed as mean ± SEM (n = 2–3 biologically independent experiments, 2–3 donors). c Frequency of InDels, measured by TIDE analysis, for control, base- and Cas9-edited samples. Data are expressed as mean ± SEM (n = 3 biologically independent experiments, 3 donors). ***P = 0.0005 (ordinary one-way ANOVA with Dunnett correction for multiple comparisons). d Frequency of the 4.9-kb deletion, measured by ddPCR, for control, base- and Cas9- edited samples. Data are expressed as mean ± SEM (n = 3 biologically independent experiments, 3 donors). ****P ≤ 0.0001 (ordinary one-way ANOVA with Dunnett correction for multiple comparisons). e RT-qPCR analysis of genes activated by RNA stimuli, 24 h post-transfection in HD HSPCs. TNF-α, IL-6, IL-12, IFN-α, IFN-β, TLR7, TLR8, RIG-I mRNA expression was normalized to GAPDH mRNA. LPS-activated macrophages were used as a positive control. Data are expressed as mean ± SEM (n = 3 biologically independent experiments, 3 donors). ****P ≤ 0.0001 (ordinary one-way ANOVA with Dunnett correction for multiple comparisons). f RT-qPCR analysis of CDKN1 (p21), 24, 48 and 72 h post-transfection in HD HSPCs. CDKN1 mRNA expression was normalized to GAPDH mRNA. Data are expressed as mean ± SEM (n = 3 biologically independent experiments, 3 donors). *P = 0.0292 for 24 h, or P = 0.0259 for 48 h, or P = 0.0131 for 72 h; **P = 0.0019; ****P ≤ 0.0001 (ordinary one-way ANOVA with Dunnett correction for multiple comparisons). Source data are provided as a Source Data file.

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