In the past decade, modular scaffolds prepared by assembling biocompatible and biodegradable building blocks (e.g. microspheres) have found promising applications in tissue engineering (TE) towards the repair/regeneration of damaged and impaired tissues. Nevertheless, to date this approach has failed to be transferred to the clinic due to technological limitations regarding microspheres patterning, a crucial issue for the control of scaffold strength, vascularization and integration in vivo. In this work, we propose a robust and reliable approach to address this issue through the fabrication of polycaprolactone (PCL) microsphere-based scaffolds with in-silico designed microarchitectures and high compression moduli. The scaffold fabrication technique consists of four main steps, starting with the manufacture of uniform PCL microspheres by fluidic emulsion technique. In the second step, patterned polydimethylsiloxane (PDMS) moulds were prepared by soft lithography. Then, layers of 500 µm PCL microspheres with geometrically inspired patterns were obtained by casting the microspheres onto PDMS moulds followed by their thermal sintering. Finally, three-dimensional porous scaffolds were built by the alignment, stacking and sintering of multiple (up to six) layers. The so prepared scaffolds showed excellent morphological and microstructural fidelity with respect to the in-silico models, and mechanical compression properties suitable for load bearing TE applications. Designed porosity and pore size features enabled in vitro human endothelial cells adhesion and growth as well as tissue integration and blood vessels invasion in vivo. Our results highlighted the strong impact of spatial patterning of microspheres on modular scaffolds response, and pay the way about the possibility to fabricate in silico-designed structures featuring biomimetic composition and architectures for specific TE purposes.
Tissue engineering (TE) pursues the ambitious goal to heal damaged tissues. One of the most successful TE approaches relies on the use of scaffolds specifically designed and fabricated to promote tissue growth. During regeneration the guidance of biological events may be essential to sustain vasculature neoformation inside the engineered scaffold. In this context, one of the most effective strategies includes the incorporation of vasculature forming cells, namely endothelial cells (EC), into engineered constructs. However, the most common EC sources currently available, intended as primary cells, are affected by several limitations that make them inappropriate to personalized medicine. Human induced Pluripotent Stem Cells (hiPSC), since the time of their discovery, represent an unprecedented opportunity for regenerative medicine applications. Unfortunately, human induced Pluripotent Stem Cells-Endothelial Cells (hiPSC-ECs) still display significant safety issues. In this work, we reviewed the most effective protocols to induce pluripotency, to generate cells displaying the endothelial phenotype and to perform an efficient and safe cell selection. We also provide noteworthy examples of both in vitro and in vivo applications of hiPSC-ECs in order to highlight their ability to form functional blood vessels. In conclusion, we propose hiPSC-ECs as the preferred source of endothelial cells currently available in the field of personalized regenerative medicine.
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