Fabrication of 3D plotted scaffold with microporous strands for bone tissue engineering.

Scaffold porosity has played a key role in bone tissue engineering aimed at effective tissue regeneration, by promoting cell attachment, proliferation, and osteogenic differentiation for new bone formation. Three-dimensional plotting systems (3DPSs) have been widely used to introduce porosity to the scaffold; however, introducing certain features in the scaffold strands that improve bone tissue regeneration remains a challenge. In this work, we fabricated bone tissue scaffolds with macro- and microporous structural features using a 3DPS and non-solvent-induced phase separation method. This approach allowed both macro- and micropores to be created in the scaffold strands. The surface morphology and mechanical and degradation properties of the perforated scaffolds were characterized carefully. Human marrow stromal cells were cultured on the scaffolds and then analyzed in vitro to assess scaffold bio-function. The highly porous scaffold exhibited mechanical properties similar to those of cancellous bone. Cell attachment, proliferation, and differentiation were significantly higher in porous scaffold compared to its nonporous counterpart. These results suggest that highly porous scaffolds have tremendous potential as a bone tissue regeneration platform.

Evaluation of Protective Immunity of Peptide Vaccines Composed of a 15-mer N-terminal Matrix Protein 2 and a Helper T-Cell Epitope Derived from Influenza A Virus.

The matrix protein 2 of influenza A virus (IFAV) has a relatively conserved ectodomain (M2e) composed of 23 amino acids, and M2e-based vaccines have been suggested to induce broad protective immunity in mice. In this study, we investigated whether N-terminal sequence of M2e (nM2e)-based vaccines with more conserved nM2e could induce influenza viral neutralizing activity. We constructed linear peptide vaccines with an nM2e sequence for PR8 virus (nM2Pr) connected to a probable 17-mer IFAV-derived helper T-cell epitope (ThE: T1, T2, or T3) at its N- or C-terminus. The peptide vaccines induced significant production of nM2e Abs regardless of either type or location of the ThE-epitope in BALB/c mice, while only T3 was effective in C57BL/6 mice. The Abs against nM2Pr-T3 elicited broader binding affinities to the nM2e peptides derived from various IFAVs than those against T3-nM2Pr. In addition, the nM2e-based vaccines efficiently protected the immunized mice from the lethal challenge of PR8 virus. These results suggest that the more conserved nM2e without cysteine will be useful for development of universal peptide vaccines than M2e.

Asymmetric liposome particles with highly efficient encapsulation of siRNA and without nonspecific cell penetration suitable for target-specific delivery.

The discovery of siRNA has been an important step in gene therapy, but the problem of delivering siRNA to a target organ limits its use as a therapeutic drug. Liposomes can be used as a nonviral vector to deliver siRNA to target cells. In this study we developed a novel method of producing asymmetric liposome particles (ALPs) with highly efficient siRNA encapsulation. Two kinds of lipid inverted micelles were prepared for the purpose of obtaining ALPs. The inner one is composed of ionizable cationic 1,2-dioleoyl-3-dimethylammonium-propane (DODAP) and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), which entrap siRNA, and the outer one is composed of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), DOPE, polyethylene glycol-1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine (PEG-PE), and cholesterol. After mixing the inverted micelles, ALPs encapsulating siRNA were obtained by solvent evaporation and dialysis. This process allowed more than 90% siRNA encapsulation as well as the negatively charged surface. The ALPs protected siRNA from ribonuclease A degradation. ALPs without any surface modification elicited almost no uptake into cells, while the surface-modified ALPs with a polyarginine peptide (R12) induced nonspecific cell penetration. The conjugation of the anti-human epidermal growth factor receptor antibody (anti-EGFR) to ALPs induces an EGFR-mediated uptake into the non-small cell lung cancer cell lines but not into NIH-3T3 cells without the receptor. The siRNA encapsulated in ALPs showed the R12- or anti-EGFR-dependent target gene silencing in NCI-H322 cells. These properties of ALPs are useful for target-specific delivery of siRNA after modification of ALPs with a target-specific ligand.