Tzu Sen Yang

Determining the binding mode and binding affinity constant of tyrosine kinase inhibitor PD153035 to DNA using optical tweezers

Accurately predicting binding affinity constant (K(A)) is highly required to determine the binding energetics of the driving forces in drug-DNA interactions. Recently, PD153035, brominated anilinoquinazoline, has been reported to be not only a highly selective inhibitor of epidermal growth factor receptor but also a DNA intercalator. Here, we use a dual-trap optical tweezers to determining K(A) for PD153035, where K(A) is determined from the changes in B-form contour length (L) of PD153035-DNA complex. Here, L is fitted using a modified wormlike chain model. We found that a noticeable increment in L in 1 mM sodium cacodylate was exhibited. Furthermore, our results showed that K(A)=1.18(±0.09)×10(4) (1/M) at 23±0.5°C and the minimum distance between adjacent bound PD153035≈11 bp. We anticipate that by using this approach we can determine the complete thermodynamic profiles due to the presence of DNA intercalators.

Quantifying membrane permeability of amphotericin B ion channels in single living cells

Recently, the structure-function relationships between amphotericin B (AmB) and ergosterol have been solved using synthetic techniques that require a mycosamine-mediated direct binding interaction between AmB and ergosterol to form AmB ion channels. However, studies to directly probe the AmB-induced membrane permeability changes have not been conducted. In the present work, we investigate the following fundamental question: does AmB induce concentration- and time-dependent permeability changes across ergosterol-containing membranes? Herein, we employ fluorescent dyes of known average diameter to quantify the diameters of AmB ion channels. In addition, we take a single-particle tracking approach to define the intracellular microrheology in the absence and presence of AmB ion channels. Present results show that increasing AmB concentration tends to increase the preferential accumulation of AmB ion channels in the presence of the excess membrane-embedded ergosterol. We found that AmB induces time-dependent membrane permeability; increases approaching 50% in both the velocity fluctuations and diffusion coefficients of vesicles occur on the same time scale as the efflux of potassium ions (≅30min). Furthermore, we propose a two-dimensional, semi-regular tessellation model to geometrically assess the pore size of the AmB ion channels in response to the AmB dose. This approach offers one possibility for the design of AmB ion channels with tunable aqueous pore size, which could provide an opportunity to replace damaged membrane water channels of the aquaporin family in future applications.

Probing Real-Time Response to Multitargeted Tyrosine Kinase Inhibitor 4-N-(3?-Bromo-Phenyl) Amino-6, 7-Dimethoxyquinazoline in Single Living Cells Using Biofuntionalized Quantum Dots

Recently, the quinazolinederivative, 4- N -(3′-bromo-phenyl) amino-6, 7-dimethoxyquinazoline (PD153035), has been reported not only to inhibit the epidermal growth factor receptor (EGFR) tyrosine kinase but also to bind to DNA double helical structures by intercalation . However, several important pharmacology issues such as whether PD153035 is a specific and reversible inhibitor of the EGFR tyrosine kinase should be addressed in more detail. In this study, we propose a nanotechnology-based approach to monitoring the real-time EGF-EGFR complex trafficking process and its relationship to cytoskeleton, as well as spatio-temporal cellular response to PD153035 at the single-cell level. W e utilize the biofunctionalized quntum dots (QDs) conjugated with EGF to monitor the cellular distribution of QD-EGF-EGFR complexes, which can provide a more direct access to probing the spatio-temporal distribution of EGF-EGFR complex in the absence and presence of PD153035 . We found that QD-EGF-EGFR complexes undergo retrograde transport before receptor-mediated internalization . In addition, QD-EGF-EGFR complexes colocalize with actin filaments , especially in filopodia regions. Furthermore, the cellular distribution of fluorescing QDs was strongly localized inside the cell after washing PD153035 for time period longer than 15 minutes. This observation demonstrated that PD153035 could be removed from the intracellular kinase domain, namely, PD153035 is a reversible EGFR inhibitor. We anticipate these approaches based on the platform at single-cell level could be applied to build a quick screening method for detection and treatment evaluation of many types of cancer expressed high levels of EGFR .

Oxygen-implanted induced formation of oxide layer enhances blood compatibility on titanium for biomedical applications

Titanium dioxide (TiO2) layers were prepared on a Ti substrate by using oxygen plasma immersion ion implantation (oxygen PIII). The surface chemical states, structure, and morphology of the layers were studied using X-ray photoelectron spectroscopy, X-ray diffraction, Raman microscopy, atomic force microscopy and scanning electron microscope. The mechanical properties, such as the Youngs modulus and hardness, of the layers were investigated using nanoindentation testing. The Ti(4+) chemical state was determined to be present on oxygen-PIII-treated surfaces, which consisted of nanocrystalline TiO2 with a rutile structure. Compared with Ti substrates, the oxygen-PIII-treated surfaces exhibited decreased Youngs moduli and hardness. Parameters indicating the blood compatibility of the oxygen-PIII-treated surfaces, including the clotting time and platelet adhesion and activation, were studied in vitro. Clotting time assays indicated that the clotting time of oxygen-PIII-treated surfaces was longer than that of the Ti substrate, which was associated with decreased fibrinogen adsorption. In conclusion, the surface characteristics and the blood compatibility of Ti implants can be modified and improved using oxygen PIII.

Hybrid micro/nanostructural surface offering improved stress distribution and enhanced osseointegration properties of the biomedical titanium implant

OBJECTIVES The aim of the present study was to investigate the surface characteristic, biomechanical behavior, hemocompatibility, bone tissue response and osseointegration of the optimal micro-arc oxidation surface-treated titanium (MST-Ti) dental implant. MATERIALS AND METHODS The surface characteristic, biomechanical behavior and hemocompatibility of the MST-Ti dental implant were performed using scanning electron microscope, finite element method, blood dripping and immersion tests. The mini-pig model was utilized to evaluate the bone tissue response and osseointegration of the MST-Ti dental implant in vivo. Data were analyzed by analysis of variance using the Students t-test (P ≤ 0.05). RESULTS The hybrid volcano-like micro/nanoporous structure was formed on the surface of the MST-Ti dental implant. The hybrid volcano-like micro/nanoporous surface played an important role to improve the stress transfer between fixture, cortical bone and cancellous bone for the MST-Ti dental implant. Moreover, the MST-Ti implant was considered to have the outstanding hemocompatibility. In vivo testing results showed that the bone-to-implant contact (BIC) ratio significantly altered as the implant with micro/nanoporous surface. After 12 weeks of implantation, the MST-Ti dental implant group exhibited significantly higher BIC ratio than the untreated dental implant group. In addition, the MST-Ti dental implant group also presented an enhancing osseointegration, particularly in the early stages of bone healing. CONCLUSION It can be concluded that the micro-arc oxidation approach induced the formation of micro/nanoporous surface is a promising and reliable alternative surface modification for Ti dental implant applications.

Optical Tweezers-Assisted Cross-Correlation Analysis for a Non-intrusive Fluid Temperature Measurement in Microdomains

An image-based approach to predict the fluid temperature in microfluidic flow cell is presented. We apply Fourier-based cross-correlation processing to determine the lateral displacement of the optically trapped bead; therefore, both the mean square displacement (MSD) and the diffusion coefficient (D) can be obtained. On the other hand, applying the Stokes–Einstein equation, together with Faxens law correction, the theoretical relation showed that D is proportional to (T/η), where T and η are temperature and temperature-dependent fluid viscosity, respectively. Hence, the fluid temperature can be determined by MSD-based thermometry.

Disinfection effects of undoped and silver-doped ceria powders of nanometer crystallite size

Being endowed with an ability of capturing and releasing oxygen, the ceria surface conventionally assumes the role of catalyzing redox reactions in chemistry. This catalytic effect also makes possible its cytotoxicity toward microorganisms at room temperature. To study this cytotoxicity, we synthesized the doped and undoped ceria particles of 8–9 nm in size using an inexpensive precipitation method and evaluated their disinfecting aptitudes with the turbidimetric and plate count methods. Among the samples being analyzed, the silver-doped ceria exhibits the highest sterilization ability, yet the undoped ceria is the most intriguing. The disinfection effect of undoped ceria is moderate in magnitude, demanding a physical contact between the ceria surface and bacteria cell wall, or the redox catalysis that can damage the cell wall and result in the cell killing. Evidently, this effect is short-range and depends strongly on dispersion of the nanoparticles. In contrast, the disinfection effects of silver-doped ceria reach out several millimeters since it releases silver ions to poison the surrounding microorganisms. Additionally, the aliovalent silver substitution creates more ceria defects. The synergetic combination, silver poisoning and heterogeneous redox catalysis, lifts and extends the disinfecting capability of silver-doped ceria to a superior level.

Research of StemBios cell therapy on dental implants containing nanostructured surfaces: Biomechanical behaviors, microstructural characteristics, and clinical trial

Purpose:The aim of the present study was to examine the osseointegration in low-density bone tissue for SLAffinity-treated implants with StemBios (SB) cell therapy. Materials and Methods:The morphologies of SLAffinity-treated surfaces were characterized using scanning electron microscopy. In the animal model, implants were installed in the mandibular canine-premolar area of 12 miniature pigs. Each pig received 3 implants of machine, sand blasted, large grit, and acid etched, and SLAffinity-treated implants. In the clinical trial, 10 patients received 1 SLAffinity-treated implant in the maxilla in the posterior area and 1 patient with low bone tissue density received 2 SLAffinity-treated implants with SB cell therapy. Resonance frequency analysis and computed tomography were assessed monthly over the first 3 months after implant placement. Results:The results demonstrated that surface treatment significantly affected early osseointegration in patients who received SB cell therapy. SB cell therapy transferred the stress caused by the implant more uniformly, and the stress decreased with healing time. SLAffinity-treated implants also proved clinically successful after the 3 months. Conclusion:The SLAffinity treatments enhanced osseointegration significantly, especially at early stages of bone tissue healing with SB cell therapy.

Single-molecule manipulation and detection platform for studying cancer cell chemotaxis

Abstract Chemotaxis of cancer cells is an essential component of tumor dissemination. The chemotactic response is comprised of three separate steps, including chemosensing, polarization and locomotion. We present an innovative approach on chemotaxis assay to address cancer cell chemotaxis. We applied a high-resolution optical tweezers system to manipulate epidermal growth factor (EGF)-coated beads positioned close to the filopodia, to locally stimulate HT29 cells expressing the EGF receptor (EGFR). We demonstrated that membrane protrusion at the leading edge induced by an EGF chemotaxis occurred at about 30∼40 s. In addition, the present observation revealed that the locomotion of HT29 cell depended on whether the HT29 cell sensed the presence of the chemoattractant EGF. We anticipate the proposed approach based on optical tweezers, together with the platform at single-cell level, could be applied to build a quick screening method for detection and treatment evaluation of many types of cancer during chemotaxis.