Cytotoxicity Study of UV-Laser-Irradiated PLLA Surfaces Subjected to Bio-Ceramisation: A New Way towards Implant Surface Modification

In this research we subjected samples of poly(L-lactide) (PLLA) extruded film to ultraviolet (193 nm ArF excimer laser) radiation below the ablation threshold. The modified film was immersed in Simulated Body Fluid (SBF) at 37 °C for 1 day or 7 days to obtain a layer of apatite ceramic (CaP) coating on the modified PLLA surface. The samples were characterized by means of optical profilometry, which indicated an increase in average roughness (Ra) from 25 nm for the unmodified PLLA to over 580 nm for irradiated PLLA incubated in SBF for 1 day. At the same time, the water contact angle decreased from 78° for neat PLLA to 35° for irradiated PLLA incubated in SBF, which suggests its higher hydrophilicity. The obtained materials were investigated by means of cell response fibroblasts (3T3) and macrophage-like cells (RAW 264.7). Properties of the obtained composites were compared to the unmodified PLLA film as well as to the UV-laser irradiated PLLA. The activation of the PLLA surface by laser irradiation led to a distinct increase in cytotoxicity, while the treatment with SBF and the deposition of apatite ceramic had only a limited preventive effect on this harmful impact and depended on the cell type. Fibroblasts were found to have good tolerance for the irradiated and ceramic-covered PLLA, but macrophages seem to interact with the substrate leading to the release of cytotoxic products.

Surface Modification of Poly(l–lactic acid) through Stereocomplexation with Enantiomeric Poly(d–lactic acid) and Its Copolymer

Poly(l–lactic acid) with high molecular weight was used to prepare PLLA films by means of the solvent casting technique. Poly(d–lactic acid) (PDLA) and poly(d–lactic acid–co–glucose) copolymer (PDLAG) with a low molecular weight were synthesized from d–lactic acid and glucose through melt polycondensation. PLLA films were immersed in PDLA or PDLAG solution to prepare surface-modified PLLA films. The modified PLLA film presented stereocomplex crystal (SC) on its surface and homogeneous crystals (HC) in its bulk. The HC structure and surface morphology of modified PLLA films were obviously damaged by PDLA or PDLAG solution. With increasing immersion time, the PLLA films modified by PDLA decreased both the HC and SC structure, while the PLLA films modified by PDLAG increased the SC structure and decreased the HC structure. Hydrophilic glucose residues of PDLAG on the surface would improve the hydrophilicity of surface-modified PLLA films. Moreover, the hydrophilicity of glucose residues and the interaction of glucose residues with lactic acid units could retard HC destruction and SC crystallization, so that PLLA films modified by PDLAG possessed lower melting temperatures of HC and SC, the crystallinity of SC and the water contact angle, compared with PDLAG–modified PLLA films. The SC structure could improve the heat resistance of modified PLLA film, but glucose residues could block crystallization to promote the thermal degradation of PLA materials. The surface modification of PLLA films will improve the thermal stability, hydrophilicity and crystallization properties of PLA materials, which is essential in order to obtain PLA-based biomaterials.

Preparation and Characterization of Poly(L-Lactide-Co-ε-Caprolactone) Copolymer for Food Packaging Application

Biodegradable poly (L-lactide-co-ε-caprolactone) (PLA-PCL) copolymers were synthesized via solution polymerization by varying the feed composition of L-lactide (LLA) and ε-caprolactone (ε-CL) (LLA/ ε-CL= 1/0, 1/1, 1/2, and 1/3). PLA-PCL film was produced by solution mixing. The films were characterized by thermal property, mechanical property, and barrier behavior tests to evaluate the effect of the PCL. The differential scanning calorimetry analyses revealed the micro-domain structure in the copolymer. The elongation at break of PLLA was improved significantly (p<0.05) in PLA-PCL copolymer while the tensile strength decreased significantly (p<0.05) with increase of PCL content. WVP of PLA-PCL films significantly decreased (p< 0.05) when compared with that of neat PLLA film. When the feed ratio of PLA-PCL copolymer increased from 1/0 to 1/3, WVP of PLA-PCL films increased from 1.85±0.15 (×10-11gm/m2sPa) to 2.83±0.26 (×10-11gm/m2sPa). The results showed that PLA-PCL copolymer can be a novel film for food packaging applications.

PLLA-Nanodiamond Composites and Their Application in Bone Tissue Engineering

Nanodiamond (ND) is an attractive nanomaterial for reinforcement of polymers [1] due to the ND’s superior mechanical and chemical properties, and low biotoxicity. A novel composite material has been produced for bone scaffolds utilizing the biodegradable polymer, poly(L-lactic acid) (PLLA), and octadecylamine-functionalized nanodiamond (ND-ODA) [2]. Composites were prepared by admixing to a PLLA/chloroform solution chloroform suspensions of ND-ODA at concentrations of 0, 1, 3, 5, 7, and 10 (w/w). Dispersion of ND-ODA in the composites was studied by transmission electron microscopy (TEM). The lower-resolution TEM micrograph of 1% wt ND-ODA/PLLA film (Fig. 1a) shows nanodiamond particles dispersed in PLLA film on amorphous carbon support. Due to long hydrocarbon chains of ODA the ND-ODA particles have good wettability with the PLLA so there is no segregation of ND-ODA and PLLA, and the polymer completely surrounds the particles. The high-resolution TEM image (Fig. 1b) shows ND crystals with attached organic material that can be ODA or PLLA. Nanoindentation tests show that the mechanical strength of ND-ODA/PLLA composites improves upon addition of ND (Table 1). Even at low concentrations (1% wt) the ND-ODA increased the hardness of the composite by 60% and Young’s modulus by 20% over neat PLLA. Based on our preliminary observations, we conclude that further additions of ND-ODA resulted in smaller changes with subsequent saturation in the mechanical properties at ∼7% wt (see Table 1). ND is relatively novel nanomaterial. Establishing its biocompatibility requires further studies, especially for modified ND. We studied the biocompatibility of 5–10nm ND and ND-ODA in experiments with a murine osteoblast cell line (7F2, from ATCC). Incubation of a cultured osteoblasts with 1–100μg/ml of ND or ND-ODA particles for 4 hours did not show much influence on the cell viability (Fig. 2), as inferred from an alamarBlue™ assay. To test the feasibility of ND-ODA/PLLA as a matrix material supporting cell growth, osteoblasts were cultured on the composites for 6 days. The attactment and proliferation of 7F2 cells on the scaffolds were assessed, respectively, by fluorescent nuclear staining with Hoechst 33258 and the alamarBlueTM assay. Our results showed that the addition of ND-ODA had only a negligibly small effect on cell proliferation, which is indicative of good biocompatibility of the composites (Fig. 3). The morphology of 7F2 cells growing on all ND-ODA/PLLA composite scaffolds was assessed by SEM. The data (not shown) confirm that the osteoblasts spread on the scaffolds similar to their spreading on TCP (tissue culture plastic). To summarize, the improved mechanical properties of the PLLA/ND-ODA composites and their good biocompatibility suggest that these materials may be suitable for applications in musculoskeletal tissue engineering.

In Vitro Chondrocyte Culture Using Hydrophilized PLLA Scaffold in Bioreactor System

Nonporous PLLA film and porous PLLA scaffolds were prepared and then grafted with acrylic acid (AA) using in situ direct plasma treatment to obtain PLLA-g-PAA. Chondrocytes isolated from rabbit knee articular cartilages were cultivated in Dulbecco’s modified eagle medium- F12 (DMEM-F12) containing 10% fetal bovine serum (FBS) and 1% antibiotics and passaged twice before cell seeding. Once seeded on either PLLA films or scaffolds, they were placed in a bioreactor system and an intermittent hydrodynamic pressure (IHP) was applied in 3 bars, while turned on for 2 min and off for 28 min during 15-day culture. AA grafting to PLLA surface was confirmed from various surface analyses. From WST-1 assay, chondrocyte proliferation was significantly improved with dynamic IHP for PLLA and PLLA-g-PAA scaffolds as compared to static culture. This study indicates that IHP may have significant influence on chondrocytes behavior in 3D culture environment.

Immobilization of Chitosan on the Plasma-Activated Poly-L-Lactic Acid Film Surface Using Evaporated Acrylic Acid as the Intermediate

Nerve bridging is to suture a biomaterial-made conduit and to overpass the damaged nerve end to end with microsurgery. Poly L-lactide (PLLA) is an excellent biomaterial that has biocompatible, biodegradable and good mechanical properties; it is thus potential to be engineered as nerve conduits and manufactured as scaffolds for nerve tissue replacement. On the other hand, chitosan provides cell affinity and considerably promotes nerves regeneration. This study is to apply plasma processing for PLLA film modification, graft the plasma-modified film with vaporized acrylic acid (AAc) monomers and then immobilize chitosan by amide bonding on the pAAc-grafted surface. This work using plasma-activation and subsequent evaporation of AAc greatly avoids PLLA thermal cracking and remaining the PLLA film in good mechanical properties. Surface morphologies are evaluated by Nano Focus. Electron Spectroscopy for Chemical Analysis (ESCA) and Attenuated Total Reflectance Fourier-Transform Infrared Spectroscopy (ATR-FTIR) are respectively employed for determining elements’ functionalities and chemical structures. Moreover, biological functionalities of the chitosan-immobilized PLLA films are thereafter assessed by antibacterial test and in vitro fibroblastic cell growth assay. The result exhibits that chitosan is immobilized on the modified PLLA films, which is plasma-activated subsequent to the evaporation of AAc. The process does not induce thermal cracking. In vitro fibroblastic cell growth assay on the chitosan-immobilized PLLA films has demonstrated that fibroblast cells on the surface become circular in shape. It decreases cell growth rate and the development of scar tissues, which may thereafter promote the effect of nerve repairing.