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. 2018 May 10;8:7458. doi: 10.1038/s41598-018-25883-x

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Perfusion recellularization of HEK293 cells, fibroblasts, and h-iPS-CPCs into 4-Flow cannulated hearts. (A) HEK293 cells delivered through only the SV of a 4-Flow heart perfuse through the cardiac veins and throughout the walls of the heart and distribute within the matrix proximal to the vasculature maintaining a hollow lumen (n = 3). (B) HEK293 cells delivered through both SV to the cardiac vein and AA to the coronary arteries are more thoroughly dispersed into the tissue walls leaving the vasculature hollow (orange triangles) (n = 3). (C) Perfusion culture of a primary human cardiac fibroblasts repopulated 4-Flow heart demonstrates cell survival, attachment, and invasion into the tissue (n = 3). Multiphoton imaging captures collagen fiber structures through second harmonic generation seen in cyan while autofluorescence of elastin and cells, and Hoechst nuclear staining is shown in green-yellow. (D) Multiphoton imaging of the 2-week 4-Flow heart reveals the lamellipodia extensions of the fibroblasts (yellow triangles) interacting with the decellularized matrix in three-dimension (n = 3). (E) Macroscopic observation of the recellularized heart with h-iPS-CPCs during the 3 weeks of culture indicate change in opacity (n = 3). (F) Corresponding representative image of Masson’s Trichrome staining shows cells are embedded within the collagen matrix. Histology sections were stained with Masson’s trichrome in which cell nuclei are purple and collagen fibers are blue.