Fabrication and Plasma Surface Activation of Aligned Electrospun PLGA Fiber Fleeces with Improved Adhesion and Infiltration of Amniotic Epithelial Stem Cells Maintaining their Teno-inductive Potential

Abstract

Electrospun PLGA microfibers with adequate intrinsic physical features (fiber alignment and diameter) have been shown to boost teno-differentiation and may represent a promising solution for tendon tissue engineering. However, the hydrophobic properties of PLGA may be adjusted through specific treatments to improve cell biodisponibility. In this study, electrospun PLGA with highly aligned microfibers were cold atmospheric plasma (CAP)-treated by varying the treatment exposure time (30, 60, and 90 s) and the working distance (1.3 and 1.7 cm) and characterized by their physicochemical, mechanical and bioactive properties on ovine amniotic epithelial cells (oAECs). CAP improved the hydrophilic properties of the treated materials due to the incorporation of new oxygen polar functionalities on the microfibers’ surface especially when increasing treatment exposure time and lowering working distance. The mechanical properties, though, were affected by the treatment exposure time where the optimum performance was obtained after 60 s. Furthermore, CAP treatment did not alter oAECs’ biocompatibility and improved cell adhesion and infiltration onto the microfibers especially those treated from a distance of 1.3 cm. Moreover, teno-inductive potential of highly aligned PLGA electrospun microfibers was maintained. Indeed, cells cultured onto the untreated and CAP treated microfibers differentiated towards the tenogenic lineage expressing tenomodulin, a mature tendon marker, in their cytoplasm. In conclusion, CAP treatment on PLGA microfibers conducted at 1.3 cm working distance represent the optimum conditions to activate PLGA surface by improving their hydrophilicity and cell bio-responsiveness. Since for tendon tissue engineering purposes, both high cell adhesion and mechanical parameters are crucial, PLGA treated for 60 s at 1.3 cm was identified as the optimal construct.

Amnion-Derived Teno-Inductive Secretomes: A Novel Approach to Foster Tendon Differentiation and Regeneration in an Ovine Model

“…Cells orientation of AECs cells after 14 days of culture with CM CO and CM FZ under air and physoxia condition was assessed on the obtained 3D tendon-like structures using the Directionality Plugin of ImageJ (Sensini et al, 2018 ; El Khatib et al, 2020 ), to better determine the teno-inductive properties of CMs. Briefly, Plugin chops the IHC images with cells nuclei DAPI counterstained into square pieces and computes their Fourier power spectra allowing the generation of statistics data on the basis of the highest peak found represented by direction (the center of the Gaussian) and dispersion (the standard deviation of the Gaussian).…”

Amniotic Epithelial Stem Cells Counteract Acidic Degradation By-Products of Electrospun PLGA Scaffold by Improving Their Immunomodulatory Profile In Vitro

“…In previous studies, it was verified that tendon biomimetic electrospun PLGA scaffolds with highly aligned fibers and adequate fiber diameter size (ha-PLGA) possess mechanical properties that are similar to human tendons [10,13,14]. Moreover, it was demonstrated that the developed constructs are suitable for inducing the differentiation of ovine amniotic epithelial stem cells (oAECs) towards the tenogenic lineage and for boosting their immunomodulatory properties by enhancing anti-inflammatory interleukin production [10,14], crucial for tendon regeneration.…”

“…Different polymers including poly( -caprolactone) (PCL), poly(lactic acid) (PLA), and poly(lactide-co-glycolide) (PLGA) are FDA-approved polymers that have been widely applied for biomedical and in particular tendon applications [1,2,[4][5][6][7][8][9][10][11][12][13][14]. Although PLA and PCL are two biocompatible polymers, they are characterized by high hydrophobic properties that decreased cell adhesion due to the lack of cell recognition sites on their surfaces [9,15,16].…”

In Vitro Innovation of Tendon Tissue Engineering Strategies

“…On the other hand, these polymers are mostly inexpensive, can be scaled industrially, and are thermoplastic, making them moldable and allowing the development of a wide variety of different structures [317,329,330]. Synthetic materials are common in tendon tissue engineering and belong mostly to aliphatic polyesters such as polyglycolic acids (PGA), polylactic acids (PLA), and polycaprolactones (PCL), as well as their copolymers poly (lactic-co-glycolic) acids (PLGA) and poly (lactic-co-caprolactone) acids (PLCL) [329,[331][332][333][334]. Other materials are also used such as Poly (ester urethane) urea (PEUUR) [335,336], polyurethane (PU) [337], and polyethylene oxide (PEO) [338][339][340].…”

“…Different traditional techniques have been used to fabricate scaffolds for tendon regeneration including sponges [341][342][343], freeze-drying [323,324,344,345], supercritical fluid processing [346,347], extruding [348], electrochemically aligned collagen [319,349,350], and electrospinning [322,325,332,333,351,352]. Recently, 3D bioprinting has emerged as an novel technique in the field of tissue engineering aiming at fabricating organized scaffolds with complex shapes [352][353][354][355][356].…”