Bilayer electrospun fibers aimed to be used for skin tissue engineering applications were fabricated for enhanced cell attachment and proliferation. Different ratios of PHBV-PLLA (70:30, 80:20, and 90:10 w/w) blends were electrospun on previously formed electrospun PHBV membranes to produce their bilayers. The fabricated electrospun membranes were characterized with FTIR, which conformed to the characteristic peaks assigned for both PHBV and PLLA. The surface morphology was evaluated using SEM analysis that showed random fibers with porous morphology. The fiber diameter and pore size were measured in the range of 0.7 ± 0.1 µm and 1.9 ± 0.2 µm, respectively. The tensile properties of the bilayers were determined using an electrodynamic testing system. Bilayers had higher elongation at break (44.45%) compared to the monolayers (28.41%) and improved ultimate tensile strength (7.940 MPa) compared to the PHBV monolayer (2.450 MPa). In vitro cytotoxicity of each of the scaffolds was determined via culturing MC3T3 (pre-osteoblastic cell line) on the membranes. Proliferation was evaluated using the Alamar Blue assay on days 3, 7, and 14, respectively. SEM images of cells cultured on membranes were taken in addition to bright field imaging to visually show cell attachment. Fluorescent nuclear staining performed with DAPI was imaged with an inverted fluorescent microscope. The fabricated bilayer shows high mechanical strength as well as biocompatibility with good cell proliferation and cell attachment, showing potential for skin substitute applications.
Keywords: PHBV; PLLA; bilayer composites; polymeric membranes; skin regeneration.