Extended Data Figure 8. Molecular correlates of miR-199a transduction.

a, Real-time PCR quantification of the ratio between α- and β-myosin heavy chain mRNA in sham, AAV6-Control- and AAV6-miR-199a-injected pig hearts, at 12 and 30 days after surgery in the H (border zone) and L (remote zone) cardiac sectors. Data are mean±SEM; the number of animals per group and time point is indicated. ns: not significant; *P<0.05 vs. AAV6-Control at the same time point; two-way ANOVA with Bonferroni post-hoc. b,c, Lectin immunofluorescence images (b) of sham, AAV6-Control- and AAV6-miR-199a-treated pig sections, 30 days after MI and vector administration along with quantification (c) of CM cross-sectional area (μm²). Data are mean±SEM; the number of analysed animals is indicated. ns: not significant. One-way ANOVA with Bonferroni post-hoc. Scale bar: 50 µm. d, Low and high magnification (insets) representative images of infarcted hearts injected with AAV6-Control or AAV6-miR-199a after immunohistochemistry to detect desmin (which is essential for maintaining structural and functional integrity of myocytes ⁴⁰ and was expressed at normally high levels), myogenin (which coordinates skeletal myogenesis and repair ⁴¹ and was not expressed), endothelin-B receptor (which selectively stained arterioles smooth muscle cells) and Wilms' tumour protein 1 (Wt1, which was expressed at low levels in the vascular endothelium, but not in myocytes). Analysis was performed in at least 7 high-resolution images acquired from at least 8 different regions of the heart of 3 pigs per group. Scale bar: 100 µm. e, Real-time PCR quantification of the levels of ANP and BNP in sham, AAV6-Control- and AAV6-miR-199a-injected pig hearts, at 12 and 30 days after surgery. Data are mean±SEM; the number of animals per group and time point is indicated. ns: not significant; *P<0.05 vs. AAV6-Control at the same time point. One-way ANOVA with Bonferroni post-hoc. f, Representative sections of pig hearts treated with AAV6-Control and AAV6-miR-199a at day 30 after infarction and vector injection stained with FITC-lectin to visualize vessels and with an anti-α-SMA antibody to detect smooth muscle cells, along with quantification of lectin-positive vessels. No significant difference between the two MI groups was detected in capillary density at either 12 or 30 days. Data are mean±SEM; the number of animals per group is indicated. Analysis was performed in at least 7 high-resolution images acquired from at least 8 different regions of the heart. *P<0.05. t-test, two-sided. Scale bar: 100 µm.