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. 2023 Jun 8;14:3345. doi: 10.1038/s41467-023-38582-7

Fig. 5. Engineered AAVs can efficiently transduce cultured human brain microvascular endothelial cells and ex vivo macaque and human brain slices.

Fig. 5

A (Left) Representative images of AAV (AAV2, AAV9, AAV-DJ, PHP.eB, PHP.V1, BR1, X1, X1.1, X1.2, X1.3, X1.4, X1.5)-mediated eGFP expression (green) in human brain microvascular endothelial cells (HBMECs). (AAVs packaged with ssAAV:CAG-eGFP, n = 6 per condition, 1-day expression). (Right) Percentage of cells transduced by the AAVs. In the condition of MOI:3E4, one-way ANOVA nonparametric Kruskal–Wallis test and multiple comparisons with uncorrected Dunn’s test are reported (***P = 0.0004 for AAV9 versus AAV-X1, ***P = 0.0002 for AAV9 versus AAV-X1.1, ****P = 3.3e-5 for AAV9 versus AAV-X1.2, **P = 0.002 for AAV9 versus AAV-X1.3). In the condition of MOI:3E3, one-way ANOVA nonparametric Kruskal–Wallis test and multiple comparisons with uncorrected Dunn’s test are reported (**P = 0.0082 for AAV9 versus AAV-X1, ***P = 0.0004 for AAV9 versus AAV-X1.1, ***P = 0.0001 for AAV9 versus AAV-X1.2). n = 6 per group, each data point is the mean of three technical replicates, mean ± SEM is plotted. B Illustration of AAV testing in ex vivo macaque and human brain slices. Created with BioRender.com. C DNA and RNA levels in southern pig-tailed macaque brain slices for AAVs, with DNA and RNA levels normalized to AAV9. N = 12 independent slices were examined over three pig-tailed macaque cases. Box plots represent the median, the first quartiles and third quartiles, with whiskers drawn at the 1.5 IQR value. Points outside the whiskers are outliers. D Representative images of AAV-mediated CAG-FXN-HA expression in ex vivo southern pig-tailed macaque brain slices. N = 12 independent slices were examined over three pig-tailed macaque cases. The tissues were co-stained with antibodies against HA (green) and NeuN (magenta). E DNA and RNA level in human brain slices for AAVs, with DNA and RNA levels normalized to AAV9. N = 5 independent slices were examined over two human cases. Box plots represent the median, the first quartiles and third quartiles, with whiskers drawn at the 1.5 IQR value. Points outside the whiskers are outliers. F RNA log enrichment of AAVs across three pig-tailed macaques, one rhesus macaque, and two human brain cases. Source data are provided as a Source Data file.