Tendon Biomimetic Electrospun PLGA Fleeces Induce an Early Epithelial-Mesenchymal Transition and Tenogenic Differentiation on Amniotic Epithelial Stem Cells

Abstract

Background. The design of tendon biomimetic electrospun fleece with Amniotic Epithelial Stem Cells (AECs) that have shown a high tenogenic attitude may represent an alternative strategy to overcome the unsatisfactory results of conventional treatments in tendon regeneration. Methods. In this study, we evaluated AEC-engineered electrospun poly(lactide-co-glycolide) (PLGA) fleeces with highly aligned fibers (ha-PLGA) that mimic tendon extracellular matrix, their biocompatibility, and differentiation towards the tenogenic lineage. PLGA fleeces with randomly distributed fibers (rd-PLGA) were generated as control. Results. Optimal cell infiltration and biocompatibility with both PLGA fleeces were shown. However, only ha-PLGA fleeces committed AECs towards an Epithelial-Mesenchymal Transition (EMT) after 48 h culture, inducing their cellular elongation along the fibers’ axis and the upregulation of mesenchymal markers. AECs further differentiated towards tenogenic lineage as confirmed by the up-regulation of tendon-related genes and Collagen Type 1 (COL1) protein expression that, after 28 days culture, appeared extracellularly distributed along the direction of ha-PLGA fibers. Moreover, long-term co-cultures of AEC-ha-PLGA bio-hybrids with fetal tendon explants significantly accelerated of half time AEC tenogenic differentiation compared to ha-PLGA fleeces cultured only with AECs. Conclusions. The fabricated tendon biomimetic ha-PLGA fleeces induce AEC tenogenesis through an early EMT, providing a potential tendon substitute for tendon engineering research.

In Vitro Innovation of Tendon Tissue Engineering Strategies

“…AECs tenocyte differentiation was also tested on a poly(lactic-co-glycolil) acids (PLGA) electrospun tendon-mimetic scaffold. They displayed an expression of mesenchymal markers (Snail, Vimentin, αSma) after 48 h and tenogenic marker expression (Col I and Tnmd) after 28 days of culture [190].…”

“…The microstructure of the produced support matrices possesses a large specific surface area that aims to increase the cell adhesion capacity, promote protein adsorption, and present more anchoring sites for receptors on the cell membrane [363,365,366]. Such microporous architectures are in fact close to the morphology of the collagen fibers constituting the tendon ECM, allowing a biomimetic character to strongly promote cell colonization and differentiation [190,318,325,367]. The produced fibers can be adjusted and modified by varying the polymer (concentration, conductivity, solvent) and electrospinning process parameters (flow rate, voltage, collector, distance between needle and collector) [190,318,328,368,369].…”

“…Such microporous architectures are in fact close to the morphology of the collagen fibers constituting the tendon ECM, allowing a biomimetic character to strongly promote cell colonization and differentiation [190,318,325,367]. The produced fibers can be adjusted and modified by varying the polymer (concentration, conductivity, solvent) and electrospinning process parameters (flow rate, voltage, collector, distance between needle and collector) [190,318,328,368,369]. Different synthetic polymer scaffolds can be fabricated by optimizing electrospinning process parameters for tendon tissue engineering.…”

“…Randomly oriented fibers can be obtained using a ground collector while those with aligned topography can be fabricated using a rotator drum/mandrel [190,338,340,350,370] and parallel copper electrodes [301]. By using an electrospinning technique, different scaffold shapes for tendon tissue engineering have been fabricated such as aligned meshes [190,328,335,339,340,350,370], bundles [371], multilayer scaffolds [368], and stacked and braided scaffolds [166,367,372]. In the biomimetic concept of tendon-like ECM, the effect of fiber orientation and diameter size are evident on the mechanical properties of the produced electrospun scaffolds (Table 1) as well as on the cellular biological response ( Table 2).…”

“…The elastic modulus and the ultimate stress as well as the strain were three and 10 times higher than that of random fibers [371]. Moreover, Russo et al produced electrospun PLGA fleeces with randomly oriented and highly aligned fibers using a cylindrical rotator drum at a rotational speed of 100 and 1000 rpm, respectively [190], while Zhang et al [340] produced CS/polylacticacids (PLLA)/Gelatin/PEO nanofibers with randomly oriented and aligned fibers using a ground collector and rotating drum at 1000 rpm [340], respectively. The produced PLGA and CS/PLLA/Gelatin/PEO fibers with aligned fibers again showed better mechanical properties compared to the random ones [190,340].…”