Patients at risk for impaired healing may benefit from prophylactic measures aimed at improving wound repair. Several photonic devices claim to enhance repair by thermal and photochemical mechanisms. We hypothesized that laser-induced thermal preconditioning would enhance surgical wound healing that was correlated with hsp70 expression. Using a pulsed diode laser (lambda=1.85 microm, tau(p)=2 ms, 50 Hz, H=7.64 mJ cm(-2)), the skin of transgenic mice that contain an hsp70 promoter-driven luciferase was preconditioned 12 hours before surgical incisions were made. Laser protocols were optimized in vitro and in vivo using temperature, blood flow, and hsp70-mediated bioluminescence measurements as benchmarks. Biomechanical properties and histological parameters of wound healing were evaluated for up to 14 days. Bioluminescent imaging studies indicated that an optimized laser protocol increased hsp70 expression by 10-fold. Under these conditions, laser-preconditioned incisions were two times stronger than control wounds. Our data suggest that this molecular imaging approach provides a quantitative method for optimization of tissue preconditioning and that mild laser-induced heat shock may be a useful therapeutic intervention prior to surgery.

Competitive binding assays utilizing concanavalin A (ConA) have the potential to be the basis of improved continuous glucose monitoring devices. However, the efficacy and lifetime of these assays have been limited, in part, by ConA’s instability due to its thermal denaturation in the physiological environment (37 °C, pH 7.4, 0.15 M NaCl) and its electrostatic interaction with charged molecules or surfaces. These undesirable interactions change the constitution of the assay and the kinetics of its behavior over time, resulting in an unstable glucose response. In this work, poly(ethylene glycol) (PEG) chains are covalently attached to lysine groups on the surface of ConA (i.e., PEGylation) in an attempt to improve its stability in these environments. Dynamic light scattering measurements indicate that PEGylation significantly improved ConA’s thermal stability at 37 °C, remaining stable for at least 30 days. Furthermore, after PEGylation, ConA’s binding affinity to the fluorescent competing ligand previously designed for the assay was not significantly affected and remained at ∼5.4 × 106 M–1 even after incubation at 37 °C for 30 days. Moreover, PEGylated ConA maintained the ability to track glucose concentrations when implemented within a competitive binding assay system. Finally, PEGylation showed a reduction in electrostatic-induced aggregation of ConA with poly(allylamine), a positively charged polymer, by shielding ConA’s charges. These results indicate that PEGylated ConA can overcome the instability issues from thermal denaturation and nonspecific electrostatic binding while maintaining the required sugar-binding characteristics. Therefore, the PEGylation of ConA can overcome major hurdles for ConA-based glucose sensing assays to be used for long-term continuous monitoring applications in vivo.

The lifetime and efficacy of a subcutaneously implanted glucose biosensor could be greatly improved by a self-cleaning membrane capable of periodic physical removal of adhered cells associated with the foreign body reaction. Previously, we reported thermoresponsive double network nanocomposite (DNNC) membrane comprised of poly(N-isopropylacrylamide) (PNIPAAm) and embedded polysiloxane nanoparticles. When the membrane was thermally cycled above and below its volume phase transition temperature (VPTT, ~33–35 °C), the associated deswelling and reswelling, respectively, led to in vitro cell release. Herein, this membrane design was tailored to meet the specific demands of a subcutaneously implanted glucose biosensor and critical functional properties were assessed. First, N-vinylpyrrolidone (NVP) comonomer increased the VPTT to ~38 °C so that the membrane would be swollen and thus more permeable to glucose in the “off-state” (i.e. no heating) while residing in the subcutaneous tissue (~35 °C). Second, glucose diffusion kinetics though the DNNC membrane was experimentally measured in its deswollen and reswollen states. A cylindrical DNNC membrane with dimensions considered suitable for implantation (1.5×5 mm, diameter × length) was used to model the glucose diffusion lag time. In addition, the DNNC cylinder was used to observe dimensional changes associated with deswelling and reswelling. Non-cytotoxicity was confirmed and self-cleaning was assessed in vitro in terms of thermally-driven cell release to confirm the potential of the DNNC membrane to control biofouling.

A self-cleaning membrane that periodically rids itself of attached cells to maintain glucose diffusion could extend the lifetime of implanted glucose biosensors. Herein, we evaluate the functionality of thermoresponsive double network (DN) hydrogel membranes based on poly(N-isopropylacrylamide) (PNIPAAm) and an electrostatic co-monomer, 2-acrylamido-2-methylpropane sulfonic acid (AMPS). DN hydrogels are comprised of a tightly crosslinked, ionized first network [P(NIPAAm-co-AMPS)] containing variable levels of AMPS (100:0–25:75 wt% ratio of NIPAAm:AMPS) and a loosely crosslinked, interpenetrating second network [PNIPAAm]. To meet the specific requirements of a subcutaneously implanted glucose biosensor, the volume phase transition temperature is tuned and essential properties, such as glucose diffusion kinetics, thermosensitivity, and cytocompatibility are evaluated. In addition, the self-cleaning functionality is demonstrated through thermally driven cell detachment from the membranes in vitro.

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