Controlling Supramolecular Structures of Drugs by Light.

Controlling physicochemical properties of light-unresponsive drugs, by light, prima facie, a paradox approach. We expanded light control by ion pairing light-unresponsive salicylate or ibuprofen to photoswitchable azobenzene counterions, thereby reversibly controlling supramolecular structures, hence the drugs' physicochemical and kinetic properties. The resulting ion pairs photoliquefied into room-temperature ionic liquids under ultraviolet light. Aqueous solutions showed trans-cis-dependent supramolecular structures under a light with wormlike aggregates decomposing into small micelles and vice versa. Light control allowed for permeation through membranes of cis-ibuprofen ion pairs within 12 h in contrast to the trans ion pairs requiring 72 h. In conclusion, azobenzene ion-pairing expands light control of physicochemical and kinetic properties to otherwise light-unresponsive drugs.

Photo-induced structural modification of silk gels containing azobenzene side groups.

Azobenzene modification of Bombyx mori silkworm silk creates a photo-responsive 'azosilk' biomaterial, allowing for 3D laser patterning. Written regions fluoresce, and become fluid-filled raised 'micro-blisters' with a 10-fold photo-softening effect of the modulus. Patterning is facile and versatile, with potential applications as soft tunable materials for dynamic cell guidance and microfluidics.

Photoresponsive retinal-modified silk-elastin copolymer.

The chimeric proteins, silk-elastin-like protein polymers (SELPs), consist of repeating units of silk and elastin to retain the mechanical strength of silk, while incorporating the dynamic environmental sensitivity of elastin. A retinal-modified SELP was prepared, modified, and studied for photodynamic responses. The protein was designed, cloned, expressed, and purified with lysine present in the elastin repeats. The purified protein was then chemically modified with the biocompatible moiety retinal via the lysine side chains. Structural changes with the polymer were assessed before and after retinal modification using Fourier transform infrared spectroscopy and circular dichroism spectroscopy. Optical studies and spectral analysis were performed before and after retinal modification. The random-coil fraction of the protein increased after retinal modification while the ß-sheet fraction significantly decreased. Birefringence of the modified protein was induced when irradiated with a linearly polarized 488 nm laser light. Retinal modification of this protein offers a useful strategy for potential use in biosensors, controlled drug delivery, and other areas of biomedical engineering.

Osteoblastic differentiation and stress response of human mesenchymal stem cells exposed to alternating current electric fields.

BACKGROUND: Electric fields are integral to many biological events, from maintaining cellular homeostasis to embryonic development to healing. The application of electric fields offers substantial therapeutic potential, while optimal dosing regimens and the underlying mechanisms responsible for the positive clinical impact are poorly understood. METHODS: The purpose of this study was to track the differentiation profile and stress response of human bone marrow derived mesenchymal stem cells (hMSCs) undergoing osteogenic differentiation during exposure to a 20 mV/cm, 60 kHz electric field. Morphological and biochemical changes were imaged using endogenous two-photon excited fluorescence (TPEF) and quantitatively assessed through eccentricity calculations and extraction of the redox ratio from NADH, FAD and lipofuscin contributions. Real time reverse transcriptase-polymerase chain reactions (RT-PCR) were used to track osteogenic differentiation markers, namely alkaline phosphatase (ALP) and collagen type 1 (col1), and stress response markers, such as heat shock protein 27 (hsp27) and heat shock protein 70 (hsp70). Comparisons of collagen deposition between the stimulated hMSCs and controls were examined through second harmonic generation (SHG) imaging. RESULTS: Quantitative differences in cell morphology, as described through an eccentricity ratio, were found on days 2 and days 5 (p < 0.05) in samples exposed to the electric field. A delayed but two fold increase in ALP and col1 transcript was detected by week 2 (p < 0.05) in differentiating hMSCs exposed to an electric field in comparison to the nonstimulated controls. Upregulation in stress marker, hsp27, and type 1 collagen deposition were correlated with this response. Increases in NADH, FAD, and lipofuscin were traced in the stimulation group during the first week of field exposure with differences statistically significant on day 10 (p < 0.05). Changes in hsp27 expression correlate well with changes in lipofuscin detected in the stimulation group, suggesting a connection with oxidative stress. Both differentiation factors and electrical stimulation improved hMSC differentiation potential to bone based on calcium deposition on day 28. CONCLUSIONS: Electrical stimulation is a useful tool to improve hMSC osteogenic differentiation, while heat shock proteins may reveal underlying mechanisms, and optical non-invasive imaging may be used to monitor the induced morphological and biochemical changes.

Supercontinuum trap stiffness measurement using a confocal approach.

We report a novel method for characterizing the stiffness of white light supercontinuum tweezers, in which the nonlinear photonic crystal fiber used for supercontinuum generation is also utilized as an effective confocal pinhole to track the motion of a trapped bead and as a scan head to realize rapid scanning of the optical trap. By measuring the phase of the bead's motion in following the trap, a lateral stiffness value of about 7.9 µN/m was obtained with supercontinumm power of about 75 mW. Our technique can potentially allow for trap stiffness calibration along an arbitrary direction in three dimensions.

Bioactive silk protein biomaterial systems for optical devices.

Silk-based biomaterial systems have been previously explored for a variety of medical and nonmedical materials needs. The unique biophysical features of silks provide options to generate highly tailored structures and morphologies with this unique family of fibrous proteins. To exploit these features, we have optimized the all aqueous processing of silk fibroin into novel surface nanopatterned protein materials. We have exploited control of this nanomorphology to optimize the optical features of these silk protein systems. We demonstrate control of surface morphology down to 125 nm, with fidelity over large length scales. This surface nanopatterning allows the silk protein to be formed into diffractive optics such as diffraction gratings, pattern generators, and lenses due to novel aqueous processing into optically clear materials via control of beta sheet crystallinity. Further, we incorporate biological components, such as hemoglobin and the enzyme peroxidase, during the process of forming the silk diffraction gratings. The ambient processing of the silk protein in water, in combination with these bioactive components, allows these entrained molecules to retain activity and provide added functions and selectivity to the optically active silk films. Thus, combinations of biochemical and optical readout is feasible and provides in a single, disposable/all degradable element with both spectral discrimination and biological function. These new surface nanopatterned, bioactive silk protein-based material systems offer a unique combination of features potentially useful for a range of biosensor needs, particularly when considered in concert with the remarkable mechanical properties of these proteins, their biocompatibility, and controllable biodegradation.

Simple fabrication technique for rapid prototyping of seamless cylindrical microchannels in polymer substrates.

This article presents a simple and novel method for the fabrication of cylindrical microchannels in polymer or biopolymer substrates. This process results in highly regular, cylindrical microchannels, suitable both to the transport of liquid and the transmission of light. This method eliminates issues associated with positioning tolerances characteristic of conventional fabrication techniques, such as soft lithography. Such devices hold great promise in the field of biomedical engineering by providing a true cylindrical profile for microflow studies and vascular modeling, as well as the ability to optically analyze injected biological samples. An evaluation of these devices is performed by real time optical imaging of blood flow.

Terahertz parametric generation photonic band gap structure with negligible structural dispersion in the optical range.

We present a photonic band gap (PBG) structure (or nonlinear photonic crystal) design for terahertz (THz) wave parametric generation, whose component materials have a small refractive index difference in the near infrared and a large index difference for THz waves. The structural dispersion of such a PBG structure is strong in the THz range but negligible in the optical range. The former allows the phase-matched pump wavelength to be placed in the near infrared to eliminate two-photon absorption of the pump and signal beams. The latter leads to a crystal layer fabrication tolerances of a few micrometers and traditional polishing methods are suitable for device fabrication. The added design flexibility also allows the use of the most efficient crystal orientations.

High-resolution nanomechanical analysis of suspended electrospun silk fibers with the torsional harmonic atomic force microscope.

Atomic force microscopes have become indispensable tools for mechanical characterization of nanoscale and submicron structures. However, materials with complex geometries, such as electrospun fiber networks used for tissue scaffolds, still pose challenges due to the influence of tension and bending modulus on the response of the suspended structures. Here we report mechanical measurements on electrospun silk fibers with various treatments that allow discriminating among the different mechanisms that determine the mechanical behavior of these complex structures. In particular we were able to identify the role of tension and boundary conditions (pinned versus clamped) in determining the mechanical response of electrospun silk fibers. Our findings show that high-resolution mechanical imaging with torsional harmonic atomic force microscopy provides a reliable method to investigate the mechanics of materials with complex geometries.

Cascaded nonlinear difference-frequency generation of enhanced terahertz wave production.

Cascaded nonlinear optical interactions are analyzed for their potential to overcome quantum-defect related limitations on the efficiency of terahertz wave difference-frequency generation. The dispersion of ZnTe permits phase-matched production of a series of Stokes lines from two initial near-infrared beams. As the pump beams run down the Stokes ladder, the number of terahertz photons continually increases. A potential improvement by a factor of 5 is demonstrated in a 0.26-cm-long crystal by use of 25-MW/mm2 pumps at a wavelength of 824 nm.

Measuring microscopic viscosity with optical tweezers as a confocal probe.

We demonstrate, what is to the best our knowledge, a new method for studying the motion of a particle trapped by optical tweezers; in this method the trapping beam itself is used as a confocal probe. By studying the response of the particle to periodic motion of the tweezers, we obtain information about the medium viscosity, particle properties, and trap stiffness. We develop the mathematical model, demonstrate experimentally its validity for our system, and discuss advantages of using this method as a new form of scanning photonic force microscopy for applications in which a high spatial and temporal resolution of the medium viscosity is desired.

Microscopic flow measurements with optically trapped microprobes.

The use of optical tweezers to measure micrometer-resolution velocity fields in fluid flow is demonstrated as an extension of a scanning confocal viscosity microscope. This demonstration is achieved by detection of the motion of an optically trapped microsphere in an oscillating laser trap. The technique is validated by comparison with an independent video-based measurement and applied to obtain a two-dimensional map of the flow past a microscopic wedge. Since the velocity is measured simultaneously with the trap relaxation time, the technique requires no fluid-dependent calibration and is independent of the trap stiffness and the particle size.

Imaging microscopic viscosity with confocal scanning optical tweezers.

The techniques of confocal microscopy and optical tweezers have shown themselves to be powerful tools in biological and medical research. We combine these methods to develop a minimally invasive instrument that is capable of making hydrodynamic measurements more rapidly than is possible with other devices. This result leads to the possibility of making scanning images of the viscosity distribution of materials around biopolymer-producing cells. 100 x 100 images can be taken with 0.5-microm spatial resolution in 3 min. An image of the viscosity distribution around a pullulan-producing cell of Aureobasidium pullulans is shown as an example.