Positive interaction between the amniotic epithelial stem cells and electrospun poly(lactide-coglycolide), poly(ε-caprolactone), poly(lactic acid)

INTRODUCTION: Amniotic epithelial stem cells (AECs) retain great potential in cell-based regenerative medicine for their plasticity, immune tolerance and anti-inflammatory properties, thus, ideal for allo and xeno-transplantation settings [1,2]. Their performance would be increased by creating AECs engineered scaffolds allowing the tissue regeneration, mechanical and structural preservation. PLGA, PCL, and PLA nonwovens electrospun scaffolds were tested for oAECs’ biocompatibility. In particular, it was studied the correlation between biopolymer characteristics and AECs’ attachment, spatial organization and proliferation. METHODS: PCL, PLA and PLGA scaffolds were prepared by electrospinning using the apparatus EC-CLI by IME Technologies (The Netherlands). The physical and morphological characterization was performed by: X-ray diffraction (Bruker D8), Scanning Electron Microscopy (SEM; ZEISS, EVO), Contact-Angle (DataPhysics Instruments GmbH). Ovine AECs (oAECs) were isolated by enzymatic digestion and cultured for 48h on all analyzed scaffolds. It was assessed cell adhesion (SEM), survival (using membranes, PKH26, and nuclei, Hoechst33342, vital dyes), spatial organization, proliferation index (detecting and quantifying Ki- 67), and DNA quantification (QubitTM DNA HS Assay, Life Technologies, Thermo Fisher Scientific Inc.). All data were obtained in triplicate and evaluated by T-test (p<0.05). RESULTS: All produced cylindrical fibres were randomly orientated with a mean pore size of about 6 μm for PLA, 8 μm for PCL and 14 μm for PLGA. Structural analysis showed that electrospun PLA and PLGA scaffolds were prevalently amorphous, whereas PCL scaffold showed a discrete level of crystallinity. Contact angle analyses showed samples similar wettability. In fact, regardless the type of polymer, they ranged between 125 and 136°. oAECs seeded on PLA, PCL and PLGA spread within the whole surface and especially around the microfibres. Although, the type of biomaterial influenced oAECs spatial distribution efficiency, and among them PLGA showed the best result. In fact, oAECs coated almost all microfibers and no nuclear pyknosis was observed. Cell proliferation index was significantly higher in PLGA (about 20%) compared to PCL and PLA (about 15 and 10%, respectively: p<0.05). DNA quantification demonstrated a different oAECs growth rate, in fact DNA quantity was significantly higher in PLGA (350.66±1.17pg/μl) compared to PCL and PLA (248±1.44pg/μl and 198.66±1.31pg/μl, respectively: p<0.05). DISCUSSION & CONCLUSIONS: these results demonstrate that oAECs and electrospun PCL, PLA and PLGA scaffolds are biocompatible. This positive interaction can be attributed to the geometrical features, including scaffold pore size and biopolymer properties which support oAECs cell adhesion, spatial organization, survival and proliferation. Indeed, the samples having higher percentage of higher pore size (i.e. PLGA) exhibited the highest cell viability in terms of cell migration and proliferation. Thus, hydrophilic nature of these biopolymers seems to mediates the first contact with cells, whereas pore size distribution appears to have an important positive effect on cell diffusion and activity. In conclusion, the investigated biocompatible electrospun matrices, in particular PLGA, represent promising bioscaffolds in AECs-based regenerative medicine