Nanopatterned acellular valve conduits drive the commitment of blood-derived multipotent cells

Abstract

Considerable progress has been made in recent years toward elucidating the correlation among nanoscale topography, mechanical properties, and biological behavior of cardiac valve substitutes. Porcine TriCol scaffolds are promising valve tissue engineering matrices with demonstrated self-repopulation potentiality. In order to define an in vitro model for investigating the influence of extracellular matrix signaling on the growth pattern of colonizing blood-derived cells, we cultured circulating multipotent cells (CMC) on acellular aortic (AVL) and pulmonary (PVL) valve conduits prepared with TriCol method and under no-flow condition. Isolated by our group from Vietnamese pigs before heart valve prosthetic implantation, porcine CMC revealed high proliferative abilities, three-lineage differentiative potential, and distinct hematopoietic/endothelial and mesenchymal properties. Their interaction with valve extracellular matrix nanostructures boosted differential messenger RNA expression pattern and morphologic features on AVL compared to PVL, while promoting on both matrices the commitment to valvular and endothelial cell-like phenotypes. Based on their origin from peripheral blood, porcine CMC are hypothesized in vivo to exert a pivotal role to homeostatically replenish valve cells and contribute to hetero- or allograft colonization. Furthermore, due to their high responsivity to extracellular matrix nanostructure signaling, porcine CMC could be useful for a preliminary evaluation of heart valve prosthetic functionality.

Keywords:

• blood-derived multipotent cells
• self-repopulation potential
• guided tissue engineering
• TriCol decellularization procedure
• ECM nanostructure signaling

Supplementary material

Figure S1 Data from one-step RT-PCR analysis were analyzed using ImageLab software.

Notes: The quantification of amplicon density band was measured for each target gene considering three independent experiments performed in triplicate. Expression levels were reported as ratio within target and housekeeping (GAPDH) gene expression level. Statistical significance was calculated by the Wilcoxon test comparing the expression profile of 1) pCMC grown on polystyrene dishes and control native valves (AV, *P≤0.05) and (PV, ●P≤0.05), or 2) pCMC cultured on valve leaflets against samples cultured for 7 days (D7) (▲P≤0.05).

Abbreviations: RT-PCR, reverse transcription–polymerase chain reaction; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; pCMC, porcine circulating multipotent cells; MMP, matrix metalloproteinase; FBN1, fibrillin; EMILIN1, emilin 1; COL1A1, alpha-1 type I collagen; COL3A1, alpha-1 type III collagen; COL4A1, alpha-1 type IV collagen; TNC, tenascin C; CALD1, caldesmon 1; VEGFA, vascular endothelial growth factor A.

Acknowledgments

This study was supported by financial grants from the “Provincia di Padova”, Italy, and Foundation for Biology and Regenerative Medicine, Tissue Engineering and Signaling (TES) ONLUS, Padova, Italy. The authors thank Annachiara Martinelli for technical support. Maria Teresa Conconi and Gerosa Gino are co-senior authors.

Author contributions

RDL: conception and design, data analysis and interpretation, manuscript writing, final approval of manuscript; SB, AM, AT, SS: collection and/or assembly of data; TB: collection and/or assembly of data, manuscript writing; PA, LI, AG: provision of study material or animals; PPP: financial support, administrative support, final approval of manuscript; MTC: collection and/or assembly of data, final approval of manuscript; GG: provision of study material or animals, final approval of manuscript. All authors contributed toward data analysis, drafting and critically revising the paper and agree to be accountable for all aspects of the work.

Disclosure

The authors report no conflicts of interest in this work.