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

Abstract

Electrospun poly(lactic-co-glycolic acid) (PLGA) scaffolds with highly aligned fibers (ha-PLGA) represent promising materials in the field of tendon tissue engineering (TE) due to their characteristics in mimicking fibrous extracellular matrix (ECM) of tendon native tissue. Among these properties, scaffold biodegradability must be controlled allowing its replacement by a neo-formed native tendon tissue in a controlled manner. In this study, ha-PLGA were subjected to hydrolytic degradation up to 20 weeks, under di-H2O and PBS conditions according to ISO 10993-13:2010. These were then characterized for their physical, morphological, and mechanical features. In vitro cytotoxicity tests were conducted on ovine amniotic epithelial stem cells (oAECs), up to 7 days, to assess the effect of non-buffered and buffered PLGA by-products at different concentrations on cell viability and their stimuli on oAECs’ immunomodulatory properties. The ha-PLGA scaffolds degraded slowly as evidenced by a slight decrease in mass loss (14%) and average molecular weight (35%), with estimated degradation half-time of about 40 weeks under di-H2O. The ultrastructure morphology of the scaffolds showed no significant fiber degradation even after 20 weeks, but alteration of fiber alignment was already evident at week 1. Moreover, mechanical properties decreased throughout the degradation times under wet as well as dry PBS conditions. The influence of acid degradation media on oAECs was dose-dependent, with a considerable effect at 7 days’ culture point. This effect was notably reduced by using buffered media. To a certain level, cells were able to compensate the generated inflammation-like microenvironment by upregulating IL-10 gene expression and favoring an anti-inflammatory rather than pro-inflammatory response. These in vitro results are essential to better understand the degradation behavior of ha-PLGA in vivo and the effect of their degradation by-products on affecting cell performance. Indeed, buffering the degradation milieu could represent a promising strategy to balance scaffold degradation. These findings give good hope with reference to the in vivo condition characterized by physiological buffering systems.

Tendon 3D Scaffolds Establish a Tailored Microenvironment Instructing Paracrine Mediated Regenerative Amniotic Epithelial Stem Cells Potential

“…Accordingly, the 3D scaffolds’ average diameter size was within the range of that of tendon collagen fibers (1–20 µm) where tenocytes reside [ 44 , 45 , 46 , 47 ]. Similarly to what was previously observed in PLGA fleeces [ 15 , 16 , 17 , 34 ], SEM analyses demonstrated that PLGA 3D scaffolds exhibited a good fiber alignment and arrangement, which could be considered desirable features to mimic tendon morphology [ 48 , 49 , 50 ].…”

“…This time span was selected according to previous studies in which it was demonstrated that AECs engineered on highly aligned constructs differentiated towards the tenogenic lineage and modulated IL-10 and IL-12 gene expression profile already after 48 h of culture [ 15 , 16 , 17 ]. As a support, it has been evidenced that 48 h represents the initial time contact between the scaffold and the host tissue, as well as, the early inflammatory phase of tendon healing [ 26 , 34 , 53 , 54 ].…”

“…PLGA fleeces were fabricated as previously reported [ 16 , 17 , 34 ] using a commercial E-Spintronic electrospinning apparatus (Erich Huber, Gerlinden, Germany), equipped with a controllable climate machine and metallic cylindrical drum rotating collector (diameter = 120 mm and circumference = 380 mm). In detail, 12% wt/wt PLGA solution was prepared in HFIP and loaded into a polypropylene syringe of 5 mL connected to a stainless-steel needle (inner diameter = 0.8 mm) with a 35 cm PTFE tube (Intra Special Catheters, Rehlingen-Siersburg, Germany).…”

“…The resulting PLGA fleeces were obtained by electrospinning 250 µL of PLGA solution. Rectangular pieces (10 mm × 7 mm) [ 16 , 17 , 34 ] were obtained from the PLGA fleeces with aligned microfibers and the width of the strips was rolled up manually to obtain 3D tendon-like scaffolds (3D scaffolds).…”

Advanced Nanofiber-Based Scaffolds for Achilles Tendon Regenerative Engineering

“…The MSCs are pluripotent non-hematopoietic adult stem cells generated from the germinal layer of mesoderm ( Pillai et al, 2017 ). The MSCs can be found in bone marrow, adipose, periosteum, skin, blood vessel walls, tendons, muscles, peripheral circulation, umbilical cord blood, periodontal ligament, and tooth tissue ( Wang et al, 2013 ; Chen J. et al, 2021 ; El Khatib et al, 2021 ). A range of active molecules (e.g., BMP-12/7 and growth factor) can promote further expansion and tendinogenic differentiation ( Tang and Lane, 2012 ; Zhou et al, 2019a ; Perucca Orfei et al, 2019 ).…”