Topical vascular organoid therapy promotes microvascular regeneration and functional recovery in porcine ischemic cardiomyopathy

Figures

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  Figure 1. Generation and delivery of SMC-EPC-based vascular organoids (A) Graphical summary of the study. Vascular organoids generated using EPCs and SMCs were transplanted into the ischemic heart disease model pig. (B) Flow cytometry for CD31 on cultured EPCs. (C) Flow cytometry for αSMA and SM22α on cultured SMCs. (D) Transplantation of EPC-SMC-based vascular organoid into the pig via left thoracotomy.

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  Figure 2. Cardiac magnetic resonance imaging (MRI) outcomes (A–C) Changes in left ventricular ejection fraction (LVEF) (A), left ventricular end-diastolic volume index (LVEDVI) (B), and left ventricular mass index (LVMI) (C) following treatment. (D) Representative delayed-enhancement MRI (DEMRI) images illustrate infarct area extension at the level of the papillary muscles. (E) Representative end-systolic MRI images and changes in wall thickness at the mid-anterior wall. (F) Functional assessment, including representative images of circumferential strain analysis. (G) Representative end-systolic MRI images of the left ventricle (LV) and changes in the sphericity index. Bars represent the mean ± standard error (SE), with each dot indicating an individual animal (n = 4 animals per group, except D, where n = 3 for the vascular organoid group). Student’s t test.

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  Figure 3. Histological and immunohistochemical (IHC) staining Representative images of the left ventricular border zone (n = 4 animals per group). (A and B) H&E staining of the border zone in the control group (A) and vascular organoid group (B). (C–J) Immunohistochemical staining in the control group (C, E, G, I) and vascular organoid group (D, F, H, J). (C, D) Troponin I and human nuclear antigen (HNA) staining; HNA-positive cells are visible in the vascular organoid group (D). (E and F) αSMA and vWF staining; αSMA+vWF+ cells were observed in the vascular organoid group (F), with high-magnification images of the boxed area shown in (a) and (b). (G and H) αSMA and HNA staining; αSMA+HNA+ cells were detected in pig heart (H), with high-magnification images in (c) and (d). (I and J) vWF and HNA staining; vWF+HNA+ cells indicate the migration of transplanted EPCs from the vascular organoid are shown in (J), with high-magnification images in (e) and (f).

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  Figure 4. Microvascular density and maturation analysis (A) The number of vWF+ cells in the border zone and infarct area. (B) The αSMA+vWF+ cell counts in the border zone and infarct area. (C) The vascular maturation index, calculated as the ratio of αSMA+vWF+ cells to total vWF+ cells. Data are presented as the mean ± standard error (SE). n = 3 animals per group. Student’s t test.

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  Figure 5. Differential gene expression and pathway analysis (A) Results of differentially expressed gene analysis using RNA-sequencing data are summarized in a volcano plot. (B) Hallmark angiogenesis-related leading-edge genes (LEGs); fold change of enriched genes in the vascular organoid group compared to controls is shown. (C) Heatmap of angiogenesis-related LEGs. n = 3 animals per group. BZ, border zone. VO, vascular organoid.