Figure 4.
Figure 4. miR669a overexpression rescues cardiac commitment of Sgcb-null cardiac progenitors in vivo. (A) Hematoxylin and eosin (H&E) staining of heart sections after ischemia/reperfusion. (B) Sirius red specifically stains scare tissue in infarcted hearts. (C) H&E-stained coronal sections of infarcted sham-operated hearts (left) and infarcted hearts transplanted with Sgcb-null cells (H4 KO/GFP; middle) or with Sgcb-null cells transduced with LVmiR669a (H4 KO/GFP/miR669; right). (D–M) Immunofluorescence analysis on infarcted hearts of sham-operated mice (D, G, G′, and K), infarcted hearts after intraventricular transplantation of 5 × 105 H4 KO/GFP cells (E, H, H′, and L); infarcted hearts after intraventricle transplantation of 5 × 105 H4 KO/GFP/miR669 (F, I, I′, and M). Heart sections of untreated (D) and treated (E and F) mice were stained with anti-Cx43 and anti-GFP. H4 KO/GFP transplanted hearts (E) showed large GFP-positive and Cx43-negative cardiac areas. On the contrary, Cx43-positive cells were homogenously distributed in the sham-operated (D) and in H4 KO/GFP/miR669 transplanted hearts (F). In the inset (F), there is shown a higher magnification of GFP-positive cells expressing Cx43 that are integrated with the surviving cardiomyocytes. Heart sections of untreated (G) and treated (H and I) mice were stained with anti-MyHC and anti-Cx43. High magnification of the highlighted areas in G, H, and I are reported in G′, H′, and I′, respectively. H4 KO/GFP transplanted hearts showed the presence of polynucleated muscle fibers that did not express Cx43, as highlighted by the asterisk in H and arrows in H′. In the inset (H), higher magnification is shown of skeletal MyHC-positive cells in a H4 KO/GFP transplanted heart. On the contrary, sham-operated (G and G′) and H4 KO/GFP/miR669 transplanted hearts (I and I′) showed Cx43 distributed all over the cardiac muscle. Heart sections from untreated (K) and treated (L and M) mice were analyzed for MyoD expression. MyoD-positive nuclei (arrows) are highly expressed in H4 KO/GFP transplanted heart sections (L), whereas they were completely absent in sham-operated heart sections (K) and extremely reduced in H4 KO/GFP/miR669 heart sections (M). Bars: (A, B, and D–I) 50 µm; (G′–M) 100 µm.

miR669a overexpression rescues cardiac commitment of Sgcb-null cardiac progenitors in vivo. (A) Hematoxylin and eosin (H&E) staining of heart sections after ischemia/reperfusion. (B) Sirius red specifically stains scare tissue in infarcted hearts. (C) H&E-stained coronal sections of infarcted sham-operated hearts (left) and infarcted hearts transplanted with Sgcb-null cells (H4 KO/GFP; middle) or with Sgcb-null cells transduced with LVmiR669a (H4 KO/GFP/miR669; right). (D–M) Immunofluorescence analysis on infarcted hearts of sham-operated mice (D, G, G′, and K), infarcted hearts after intraventricular transplantation of 5 × 105 H4 KO/GFP cells (E, H, H′, and L); infarcted hearts after intraventricle transplantation of 5 × 105 H4 KO/GFP/miR669 (F, I, I′, and M). Heart sections of untreated (D) and treated (E and F) mice were stained with anti-Cx43 and anti-GFP. H4 KO/GFP transplanted hearts (E) showed large GFP-positive and Cx43-negative cardiac areas. On the contrary, Cx43-positive cells were homogenously distributed in the sham-operated (D) and in H4 KO/GFP/miR669 transplanted hearts (F). In the inset (F), there is shown a higher magnification of GFP-positive cells expressing Cx43 that are integrated with the surviving cardiomyocytes. Heart sections of untreated (G) and treated (H and I) mice were stained with anti-MyHC and anti-Cx43. High magnification of the highlighted areas in G, H, and I are reported in G′, H′, and I′, respectively. H4 KO/GFP transplanted hearts showed the presence of polynucleated muscle fibers that did not express Cx43, as highlighted by the asterisk in H and arrows in H′. In the inset (H), higher magnification is shown of skeletal MyHC-positive cells in a H4 KO/GFP transplanted heart. On the contrary, sham-operated (G and G′) and H4 KO/GFP/miR669 transplanted hearts (I and I′) showed Cx43 distributed all over the cardiac muscle. Heart sections from untreated (K) and treated (L and M) mice were analyzed for MyoD expression. MyoD-positive nuclei (arrows) are highly expressed in H4 KO/GFP transplanted heart sections (L), whereas they were completely absent in sham-operated heart sections (K) and extremely reduced in H4 KO/GFP/miR669 heart sections (M). Bars: (A, B, and D–I) 50 µm; (G′–M) 100 µm.

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