A Magnetically Responsive Hydrogel System for Controlling the Timing of Bone Progenitor Recruitment and Differentiation Factor Deliveries.

The sequence and timing of growth factor delivery plays a crucial role in bone regeneration. While a variety of biomaterial scaffolds have been developed to provide multiple growth factor deliveries, there still exists a strong need for on-demand control over sequential delivery profiles to optimize regenerative outcomes. One particular growth factor, bone morphogenetic protein-2 (BMP-2), has established effects in the osteodifferentiation process; however, the optimal timing of its delivery is not yet known. Here, we investigate the effect of the timing of BMP-2 delivery on osteodifferentiation on both 2D and 3D cell cultures in vitro. It was shown that immediate BMP-2 delivery inhibited mouse mesenchymal stem cell (mMSC) proliferation and therefore resulted in suboptimal levels of mMSC osteodifferentiation (as measured by alkaline phosphatase activity) compared to mMSC cultures exposed to delayed BMP-2 delivery (4 day delay). Because of this, we aimed to develop a biomaterial system capable of rapidly recruiting mMSCs and exposing them to BMP-2 in a delayed manner (i.e., after a strong mMSC population has been established). This biomaterial system consisted of (i) an outer porous gelatin compartment that could be loaded with an mMSC recruitment factor (stromal cell-derived factor 1-α (SDF-1α)) for rapid establishment of a 3D mMSC culture and (ii) an inner ferrogel compartment that could deliver BMP-2 in an immediate or delayed manner, depending on when magnetic stimulation was applied. It was shown that the outer compartment was able to recruit and harbor mMSCs and that the rapidity of this recruitment could be enhanced by loading the compartment with SDF-1α. The inner ferrogel compartment enabled magnetically triggered release of BMP-2 where the timing of release could be remotely controlled from immediate to a delay of up to 11 days. This hydrogel system provides controllability over the timing between bone progenitor recruitment and osteodifferentiation factor release and can thus potentially enhance therapies that require new bone growth by optimizing the timing of these deliveries.

Pulsatile Chemotherapeutic Delivery Profiles Using Magnetically Responsive Hydrogels.

Pulsatile chemotherapeutic delivery profiles may provide a number advantages by maximizing the anticancer toxicity of chemotherapeutics, reducing off-target side effects, and combating adaptive resistance. While these temporally dynamic deliveries have shown some promise, they have yet to be clinically deployed from implantable hydrogels, whose localized deliveries could further enhance therapeutic outcomes. Here, several pulsatile chemotherapeutic delivery profiles were tested on melanoma cell survival in vitro and compared to constant (flatline) delivery profiles of the same integrated dose. Results indicated that pulsatile delivery profiles were more efficient at killing melanoma cells than flatline deliveries. Furthermore, results suggested that parameters like the duration of drug "on" periods (pulse width), delivery rates during those periods (pulse heights), and the number/frequency of pulses could be used to optimize delivery profiles. Optimization of pulsatile profiles at tumor sites in vivo would require hydrogel materials capable of producing a wide variety of pulsatile profiles (e.g., of different pulse heights, pulse widths, and pulse numbers). This work goes on to demonstrate that magnetically responsive, biphasic ferrogels are capable of producing pulsatile mitoxantrone delivery profiles similar to those tested in vitro. Pulse parameters such as the timing and rate of delivery during "on" periods could be remotely regulated through the use of simple, hand-held magnets. The timing of pulses was controlled simply by deciding when and for how long to magnetically stimulate. The rate of release during pulse "on" periods was a function of the magnetic stimulation frequency. These findings add to the growing evidence that pulsatile chemotherapeutic delivery profiles may be therapeutically beneficial and suggest that magnetically responsive hydrogels could provide useful tools for optimizing and clinically deploying pulsatile chemotherapeutic delivery profiles.

Switchable Release of Entrapped Nanoparticles from Alginate Hydrogels.

Natural biological processes are intricately controlled by the timing and spatial distribution of various cues. To mimic this precise level of control, the physical sizes of gold nanoparticles are utilized to sterically entrap them in hydrogel materials, where they are subsequently released only in response to ultrasound. These nanoparticles can transport bioactive factors to cells and direct cell behavior on-demand.

Biphasic ferrogels for triggered drug and cell delivery.

Ferrogels are an attractive material for many biomedical applications due to their ability to deliver a wide variety of therapeutic drugs on-demand. However, typical ferrogels have yet to be optimized for use in cell-based therapies, as they possess limited ability to harbor and release viable cells. Previously, an active porous scaffold that exhibits large deformations and enhanced biological agent release under moderate magnetic fields has been demonstrated. Unfortunately, at small device sizes optimal for implantation (e.g., 2 mm thickness), these monophasic ferrogels no longer achieve significant deformation due to a reduced body force. A new biphasic ferrogel, containing an iron oxide gradient, capable of large deformations and triggered release even at small gel dimensions, is presented in this study. Biphasic ferrogels demonstrate increased porosity, enhanced mechanical properties, and potentially increased biocompatibility due to their reduced iron oxide content. With their ability to deliver drugs and cells on-demand, it is expected that these ferrogels will have wide utility in the fields of tissue engineering and regenerative medicine.

Cationic peptide exposure enhances pulsed-electric-field-mediated membrane disruption.

BACKGROUND: The use of pulsed electric fields (PEFs) to irreversibly electroporate cells is a promising approach for destroying undesirable cells. This approach may gain enhanced applicability if the intensity of the PEF required to electrically disrupt cell membranes can be reduced via exposure to a molecular deliverable. This will be particularly impactful if that reduced PEF minimally influences cells that are not exposed to the deliverable. We hypothesized that the introduction of charged molecules to the cell surfaces would create regions of enhanced transmembrane electric potential in the vicinity of each charged molecule, thereby lowering the PEF intensity required to disrupt the plasma membranes. This study will therefore examine if exposure to cationic peptides can enhance a PEF's ability to disrupt plasma membranes. METHODOLOGY/PRINCIPAL FINDINGS: We exposed leukemia cells to 40 µs PEFs in media containing varying concentrations of a cationic peptide, polyarginine. We observed the internalization of a membrane integrity indicator, propidium iodide (PI), in real time. Based on an individual cell's PI fluorescence versus time signature, we were able to determine the relative degree of membrane disruption. When using 1-2 kV/cm, exposure to >50 µg/ml of polyarginine resulted in immediate and high levels of PI uptake, indicating severe membrane disruption, whereas in the absence of peptide, cells predominantly exhibited signatures indicative of no membrane disruption. Additionally, PI entered cells through the anode-facing membrane when exposed to cationic peptide, which was theoretically expected. CONCLUSIONS/SIGNIFICANCE: Exposure to cationic peptides reduced the PEF intensity required to induce rapid and irreversible membrane disruption. Critically, peptide exposure reduced the PEF intensities required to elicit irreversible membrane disruption at normally sub-electroporation intensities. We believe that these cationic peptides, when coupled with current advancements in cell targeting techniques will be useful tools in applications where targeted destruction of unwanted cell populations is desired.