Preparation, Characterization and Biological Activity of Cathelicidin-BF-30-Loaded Poly(LA-co-MA) Microspheres.

Antimicrobial peptides have a great potential to replace antibiotics in the treatment of bacterial infections. In order to improve the its stability, the antimicrobial peptides were incorporated in poly(L-lactic acid-co-D,L-mandelic acid) (LA-co-MA) microspheres prepared by a double emulsion solvent evaporation method. In this study, the microspheres obtained had a mean particle size of 2.75±0.2 µm, Encapsulation Efficiency (EE) of 92.47±1.21% and drug loading of 8.44±0.11%. The peptides were released from poly(LA-co-MA) microspheres at a constant speed and no significant initial burst effect was observed. The secondary structure and antibacterial activity of the released peptide were retained, which were compared with those of the native peptides. In addition, these BF-30-loaded microspheres presented <5% hemolysis and no toxicity for HEK293 cells even at the highest tested concentration (150 µg/mL), indicating that the poly(LA-co-MA) microspheres are promising carriers for peptides.

Characterization, Stability and Biological Activity In Vitro of Cathelicidin-BF-30 Loaded 4-Arm Star-Shaped PEG-PLGA Microspheres.

BF-30 is a single chain polypeptide of an N-segment with an α-helix from cathelicidin gene encoding, and it contains 30 amino acid residues, with a relative molecular mass and isoelectric point of 3637.54 and 11.79, respectively. Cathelicidin-BF-30 was entrapped in four-arm star-shaped poly(ethylene glycol-b-dl-lactic acid-co-glycolic acid) block copolymers (4-arm-PEG-PLGA) by a double-emulsion solvent-evaporation method. Three release phases of cathelicidin-BF-30loaded 4-arm-PEG-PLGA microspheres were observed, including an initial burst-release phase, followed by a lag phase with minimal drug release and finally a secondary zero-order release phase. The delivery system released BF-30 over more than 15 days in vitro. Furthermore, the material for preparing the microspheres has good biocompatibility and biodegradability. Additionally, based on the drug resistance of food pathogenic bacteria, the antibacterial effects of BF-30 on Shigella dysenteriae CMCC 51105 (Sh. dysenteriae CMCC 51105), Salmonella typhi (S. typhi) and Staphylococcus aureus (S. aureus) as well as the stability of the in vitro release of the BF-30-loded microspheres were studied. The α-helix secondary structure and antibacterial activity of released BF-30 were retained and compared with native peptide. These BF-30 loaded microspheres presented <10% hemolysis and no toxicity for HEK293T cells even at the highest tested concentration (150 µg/mL), indicating that they are hemocompatible and a promising delivery and protection system for BF-30 peptide.

Fabrication of flocculation-resistant pH/ionic strength/temperature multiresponsive hollow microspheres and their controlled release.

pH/ionic strength/temperature multiresponsive hollow microspheres were successfully prepared by the Ce(IV) initiated grafting polymerization of N-isopropylacrylamide (NIPAm) onto the multilayered polyelectrolyte shells encapsulating the polystyrene sulfonate (PSS) microsphere templates fabricated by the layer-by-layer assembly of chitosan (CS) and alginate (SAL), after etching the templates by dialysis. The hollow structure of the obtained multiresponsive hollow microspheres was characterized by transmission electron microscopy (TEM), which indicated that the inner diameter of the hollow microspheres was about 200 nm. The environmental responsive properties of the multiresponsive hollow microspheres were characterized with dynamic light scattering (DLS) in an aqueous system. The introduction of poly(N-isopropylacrylamide) (PNIPAm) brushes onto the pH/ionic strength dual-responsive hollow microspheres achieved temperature-responsive characteristics. It also could prevent flocculation among the obtained multiresponsive hollow microspheres in a solution with higher salt concentration. Their controlled release of drug molecules (a model hydrophobic drug, dipyridamole (DIP)) was also investigated.

A microfluidics-derived growth factor gradient in a scaffold regulates stem cell activities for tendon-to-bone interface healing.

Treatment of tendon-to-bone interface injury has long been challenging in sports medicine. The major obstacle lies with the complicated three-layer structure of the tissue that consists of a bone region with osteocytes, a tendon region with tenocytes and a transitional region with chondrocytes. Conventional tissue engineering approaches using simply biomaterial scaffolds, stem cells and combinations of them had limited abilities to reconstruct the gradient structure with normal biomechanical properties. We herein aim to construct a three-layer structure with bone marrow-derived stem cells and tendon stem cells cultured in a decellularized tendon scaffold, through application of a gradient of biological cues in the longitudinal direction of the scaffold that guides the stem cells to differentiate and remodel the extracellular matrix in response to different medium concentrations in different regions. A microfluidic chip, on which a tree-like flow pattern was implemented, was adopted to create the concentration gradient in a dichotomous manner. We screened for an optimized seeding ratio between the two stem cell types before incubation of the scaffold in the medium concentration gradient and surgical implantation. Histology and immunohistochemistry assessments, both qualitatively and semi-quantitatively, showed that the microfluidic system provided desired guidance to the seeded stem cells that the healing at 8-week post-implantation presented a similar structure to that of a normal tendon-to-bone interface, which was outstanding compared to treatments without gradient guidance, stem cells or scaffolds where chaotic and fibrotic structures were obtained. This strategy offers a potentially translational tissue engineering approach for better outcomes in tendon-to-bone healing.