Fluorescence-Shadowing Nanoparticle Clusters for Real-Time Monitoring of Tumor Progression.

Monitoring tumor progression is important for elucidating appropriate therapeutic strategies in response to anticancer therapeutics. To fluorescently monitor the in vivo levels of tumor-specific enzymes, we prepared matrix metalloprotease (MMP)-responsive gold nanoparticle (AuNP) clusters to sense tumor microenvironments. Specifically, AuNPs and quantum dots (QDs) were surface-engineered with two poly(ethylene glycol) [PEG] shells and cyclooctyne moieties, respectively, for the copper-free click reaction. Upon "peeling off" of the secondary shell from the double-PEGylated AuNPs under MMP-rich conditions, shielded azide moieties of the AuNPs were displayed toward the QD, and those two particles were clicked into nanoparticle clusters. This consequently resulted in a dramatic size increase and fluorescence quenching of QDs via fluorescence energy transfer (FRET) due to the molecular proximity of the particles. We observed that FRET efficiency was modulated via changes in MMP levels and exposure time. Cancer cell numbers exhibited a strong correlation with FRET efficiency, and in vivo studies that employed solid tumor models accordingly showed that FRET efficiency was dependent on the tumor size. Thus, we envision that this platform can be tailored and optimized for tumor monitoring based on MMP levels in solid tumors.

Light-Triggered Structural Modulation of Nanofibrous Meshes to Promote Deep Penetration of Cultured Cells.

Although nanofibrous meshes are considered promising cultivation beds for maintaining cell differentiation, 3D cultivation is not possible because their nanoporous structures impede cell infiltration. To facilitate transverse cell migration across nanofibrous meshes, electrospun nanofibers are prepared with structures that vary in response to red laser light. Polyoxalate (POX), composed of oxalate linkers and oligomeric caprolactone, is synthesized and electrospun into fibrous meshes with a photosensitizer (chlorin e6, Ce6). These meshes exhibit morphological and chemical changes upon laser irradiation, and mass erosion rates of the meshes are faster after laser irradiation. Cell cultivation on POX meshes reveals that red laser effectively facilitates traverse migration of the cells without affecting cell viability. The use of light-triggered change of meshes is envisioned to promote the migration of cells during 3D matrix cultivation.

Atom transfer radical-polymerized cationic shells on gold nanoparticles for near infrared-triggered photodynamic therapy of tumor-bearing animals.

Gold nanoparticles (AuNPs) were surface-engineered with a cationic corona to enhance the incorporation of photosensitizers for photodynamic therapy (PDT). The cationic corona composed of poly(2-(dimethylamino)ethyl methacrylate) was atom transfer radical-polymerized on the surface of the AuNPs. The cationic corona of the engineered surface was characterized by dynamic light scattering, electron microscopy, Raman spectroscopy, and mass spectroscopy. Chlorin-e6 (Ce6) incorporated onto the surface-engineered AuNPs exhibited higher cell incorporation efficiency than bare AuNPs. Ce6-incorporated AuNPs were confirmed to release singlet oxygen upon NIR irradiation. Compared to Ce6, Ce6-incorporated AuNPs exhibited higher cellular uptake and cytotoxicity against cancer cells in an irradiation time-dependent manner. Near-infrared-irradiated animals administered Ce6-incorporated AuNPs exhibited higher levels of tumor suppression without noticeable body weight loss. This result was attributed to the higher localization of Ce6 at the tumor sites to induce cancer cell apoptosis. Thus, we envision that engineered AuNPs with cationic corona can be tailored to effectively deliver photosensitizers to tumor sites for photodynamic therapy.

Comparison Study of the Effects of Cationic Liposomes on Delivery across 3D Skin Tissue and Whitening Effects in Pigmented 3D Skin.

Charged phospholipids are employed to formulate liposomes with different surface charges to enhance the permeation of active ingredients through epidermal layers. Although 3D skin tissue is widely employed as an alternative to permeation studies using animal skin, only a small number of studies have compared the difference between these skin models. Liposomal delivery strategies are investigated herein, through 3D skin tissue based on their surface charges. Cationic, anionic, and neutral liposomes are formulated and their size, zeta-potential, and morphology are characterized using dynamic light scattering and cryogenic-transmission electron microscopy (cryo-TEM). A Franz diffusion cell is employed to determine the delivery efficiency of various liposomes, where all liposomes do not exhibit any recognizable difference of permeation through the synthetic membrane. When the fluorescence liposomes are applied to 3D skin, considerable fluorescence intensity is observed at the stratum cornea and epithelium layers. Compared to other liposomes, cationic liposomes exhibit the highest fluorescence intensity, suggesting the enhanced permeation of liposomes through the 3D skin layers. Finally, the ability of niacinamide (NA)-incorporated liposomes to suppress melanin transfer in pigmented 3D skin is examined, where cationic liposomes exhibit the highest degree of whitening effects.

Preparation of Stretchable Nanofibrous Sheets with Sacrificial Coaxial Electrospinning for Treatment of Traumatic Muscle Injury.

Traumatic muscle injury with massive loss of muscle volume requires intramuscular implantation of proper scaffolds for fast and successful recovery. Although many artificial scaffolds effectively accelerate formation and maturation of myotubes, limited studies are showing the therapeutic effect of artificial scaffolds in animal models with massive muscle injury. In this study, improved myotube differentiation is approved on stepwise stretched gelatin nanofibers and applied to damaged muscle recovery in an animal model. The gelatin nanofibers are fabricated by a two-step process composed of co-axial electrospinning of poly(ɛ-caprolactone) and gelatin and subsequent removal of the outer shells. When stepwise stretching is applied to the myoblasts on gelatin nanofibers for five days, enhanced myotube formation and polarized elongation are observed. Animal models with volumetric loss at quadriceps femoris muscles (>50%) are transplanted with the myotubes cultivated on thin and flexible gelatin nanofiber. Treated animals more efficiently recover exercising functions of the leg when myotubes and the gelatin nanofiber are co-implanted at the injury sites. This result suggests that mechanically stimulated myotubes on gelatin nanofiber is therapeutically feasible for the robust recovery of volumetric muscle loss.

Coaxial Electrospun Nanofibers with Different Shell Contents to Control Cell Adhesion and Viability.

Electrospun nanofibers are widely employed as cell culture matrices because their biomimetic structures resemble a natural extracellular matrix. However, due to the limited cell infiltration into nanofibers, three-dimensional (3D) construction of a cell matrix is not easily accomplished. In this study, we developed a method for the partial digestion of a nanofiber into fragmented nanofibers composed of gelatin and polycaprolactone (PCL). The PCL shells of the coaxial fragments were subsequently removed with different concentrations of chloroform to control the remaining PCL on the shell. The swelling and exposure of the gelatin core were manipulated by the remaining PCL shells. When cells were cultivated with the fragmented nanofibers, they were spontaneously assembled on the cell sheets. The cell adhesion and proliferation were significantly affected by the amount of PCL shells on the fragmented nanofibers.

Self-assembled cell sheets composed of mesenchymal stem cells and gelatin nanofibers for the treatment of full-thickness wounds.

Cell sheet engineering has attracted great attention because thin layers of tissue can be easily transplanted to defect sites. Wound-dressing materials are required to support fast re-epithelization, both with keratinocytes and fibroblasts, to enhance the prognosis and therapeutic outcomes. We prepared self-assembled cell sheets composed of adipocyte-derived stem cells (ADSCs) and surface-engineered nanofibrils (NFs). NFs were surface-engineered with multilayers of gelatin so that the cell sheets could spontaneously assemble within 3 days in cell culture plates. Dorsal wounds transplanted with the cell sheets exhibited higher wound-healing rates when a high concentration of gelatin was immobilized on the surfaces of the NFs. Histochemical staining revealed that those with gelatin-immobilized NFs showed a higher expression of cytokeratin and collagen in the re-epithelized epidermis. Keratinocytic differentiation of the epidermis was molecularly evidenced by the higher expression of keratinocyte-specific genes.