Yu Hung Chen

Increased apoptotic potential and dose‐enhancing effect of gold nanoparticles in combination with single‐dose clinical electron beams on tumor‐bearing mice

High atomic number material, such as gold, may be used in conjunction with radiation to provide dose enhancement in tumors. In the current study, we investigated the dose‐enhancing effect and apoptotic potential of gold nanoparticles in combination with single‐dose clinical electron beams on B16F10 melanoma tumor‐bearing mice. We revealed that the accumulation of gold nanoparticles was detected inside B16F10 culture cells after 18 h of incubation, and moreover, the gold nanoparticles were shown to be colocalized with endoplasmic reticulum and Golgi apparatus in cells. Furthermore, gold nanoparticles radiosensitized melanoma cells in the colony formation assay (P = 0.02). Using a B16F10 tumor‐bearing mouse model, we further demonstrated that gold nanoparticles in conjunction with ionizing radiation significantly retarded tumor growth and prolonged survival compared to the radiation alone controls (P < 0.05). Importantly, an increase of apoptotic signals was detected inside tumors in the combined treatment group (P < 0.05). Knowing that radiation‐induced apoptosis has been considered a determinant of tumor responses to radiation therapy, and the length of tumor regrowth delay correlated with the extent of apoptosis after single‐dose radiotherapy, these results may suggest the clinical potential of gold nanoparticles in improving the outcome of melanoma radiotherapy. (Cancer Sci 2008; 99: 1479–1484)

Laser ablation method: Use of surfactants to form the dispersed Ag nanoparticles

Abstract The laser ablation method was employed to synthesize Ag nanoparticles. With the aid of the surfactants, SDS and CTAB, highly dispersed Ag particles were attained. Using a laser intensity of 120 mJ/pulse particle diameters of 4.2±1.9 nm were produced in SDS and 7.8±4.5 nm in CTAB. The anionic SDS and cationic CTAB exhibited differential ability in protecting Ag colloids: the particles containing SDS remained stable at least one month, while the CTAB-protected Ag precipitated completely within one week. Plausible explanations were given to such stability observations. Moreover, the changes in the ablation laser powers were found to achieve particle size control. Lesser laser intensity of 60 mJ/pulse produced larger particle sizes of 6.8±2.7 nm and 9.4±5.9 nm in SDS and CTAB, respectively.

Cathepsin L mediates resveratrol-induced autophagy and apoptotic cell death in cervical cancer cells.

Cathepsins have long been considered as housekeeping molecules. However, specific functions have also been attributed to each one of these lysosomal proteases. Squamous cell carcinoma antigen (SCCA) 1, widely expressed in various uterine cervical cells, is an endogenous cathepsin (cat) L inhibitor. In this study, we investigated whether the cat L-SCCA 1 lysosomal pathway and autophagy were involved in resveratrol (RSV)-induced cytotoxicity in cervical cancer cells. RSV induced GFP-LC3 aggregation as well as increased the presence of LC3-II and autophagosomes as was revealed by electron microscopy in cervical cancer cells. Prolonged treatment of RSV induced cytosolic translocation of cytochrome c, caspase 3 activation, and apoptotic cell death. This apoptotic effect was abrogated by trans-epoxysuccinyl- L-leucylamido - (4-guanidino)butane, an inhibitor of cat B and L, but not by pepstatin A, an inhibitor of cat D. As cervical cancer cells express little cat B, we further studied the role of cat L. RSV induced dissipation of the lysosomal membrane permeability (LMP), leakage and increased cytosolic expression and activity of cat L. Inhibition of cat L by small interference RNA (siRNA) protected cells from RSV-induced cytotoxicity. In contrast, inhibition of SCCA 1 by siRNA promoted RSV-induced cytotoxicity. Inhibition of autophagic response by wormannin (WT) or asparagine (ASP) resulted in decreased early LC3-II formation, reduced LMP, and abolishment of the increase in RSV-induced cell death. In conclusion, we have identified a new cytotoxic mechanism in which the lysosomal enzyme cat L acts as a death signal integrator in cervical cancer cells. Furthermore, SCCA 1 may play an anti-apoptotic role through anti-cat L activity.

Analysis of DNA fragments by microchip electrophoresis fabricated on poly(methyl methacrylate) substrates using a wire-imprinting method

Microfluidic devices were fabricated on poly(methyl methacrylate) (PMMA) substrate using two small‐diameter (79 μm) wires to create a cross impression in plastics softened by low‐temperature heating. The resulting channels had a rounded shape and 75 μm in depth. The variability of the channel dimensions was found to be less than 6% from different locations of the same channel and less than 10% between chips. Moreover, the fabricated PMMA chip appeared to sustain an electric field strength up to 300 V/cm without significant Joule heating. The function of resulting devices for electrophoretic injection and separation of a DNA size marker, HaeIII digest of ΦX174, was also characterized. Results indicated that all of the 11 DNA fragments of the size marker could be identified in less than 3 min with relative standard deviations less than 0.4% and 8% for migration time and peak area, respectively. Moreover, with the use of near infrared (IR) dye, fluorescence signals of the higher molecular weight fragments (> 603 bp in length) could be detected at total DNA concentrations as low as 0.1 μg/mL (S/N = 4.2). In conclusion, the performance of wire‐imprinted devices on PMMA substrate were comparable to those fabricated by other professional means.

Galectin-1 Binds to Influenza Virus and Ameliorates Influenza Virus Pathogenesis

ABSTRACT Innate immune response is important for viral clearance during influenza virus infection. Galectin-1, which belongs to S-type lectins, contains a conserved carbohydrate recognition domain that recognizes galactose-containing oligosaccharides. Since the envelope proteins of influenza virus are highly glycosylated, we studied the role of galectin-1 in influenza virus infection in vitro and in mice. We found that galectin-1 was upregulated in the lungs of mice during influenza virus infection. There was a positive correlation between galectin-1 levels and viral loads during the acute phase of viral infection. Cells treated with recombinant human galectin-1 generated lower viral yields after influenza virus infection. Galectin-1 could directly bind to the envelope glycoproteins of influenza A/WSN/33 virus and inhibit its hemagglutination activity and infectivity. It also bound to different subtypes of influenza A virus with micromolar dissociation constant (Kd ) values and protected cells against influenza virus-induced cell death. We used nanoparticle, surface plasmon resonance analysis and transmission electron microscopy to further demonstrate the direct binding of galectin-1 to influenza virus. More importantly, we show for the first time that intranasal treatment of galectin-1 could enhance survival of mice against lethal challenge with influenza virus by reducing viral load, inflammation, and apoptosis in the lung. Furthermore, galectin-1 knockout mice were more susceptible to influenza virus infection than wild-type mice. Collectively, our results indicate that galectin-1 has anti-influenza virus activity by binding to viral surface and inhibiting its infectivity. Thus, galectin-1 may be further explored as a novel therapeutic agent for influenza.

Current progress in dengue vaccines

Dengue is one of the most important emerging vector-borne viral diseases. There are four serotypes of dengue viruses (DENV), each of which is capable of causing self-limited dengue fever (DF) or even life-threatening dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). The major clinical manifestations of severe DENV disease are vascular leakage, thrombocytopenia, and hemorrhage, yet the detailed mechanisms are not fully resolved. Besides the direct effects of the virus, immunopathological aspects are also involved in the development of dengue symptoms. Although no licensed dengue vaccine is yet available, several vaccine candidates are under development, including live attenuated virus vaccines, live chimeric virus vaccines, inactivated virus vaccines, and live recombinant, DNA and subunit vaccines. The live attenuated virus vaccines and live chimeric virus vaccines are undergoing clinical evaluation. The other vaccine candidates have been evaluated in preclinical animal models or are being prepared for clinical trials. For the safety and efficacy of dengue vaccines, the immunopathogenic complications such as antibody-mediated enhancement and autoimmunity of dengue disease need to be considered.

Laser-induced alloying Au–Pd and Ag–Pd colloidal mixtures: the formation of dispersed Au/Pd and Ag/Pd nanoparticlesElectronic supplementary information (ESI) available: TEM images of the molar ratios 2∶1 for both Au–Pd and Ag–Pd colloids. See http://www.rsc.org/suppdata/jm/b2/b200587e/.

Laser irradiation of colloidal mixtures consisting of Au–Pd and Ag–Pd were conducted to synthesize Au/Pd and Ag/Pd alloy nanoparticles. Compositional ratios of 1∶1 and 2∶1 were prepared in both the Au/Pd and Ag/Pd systems. The compositions, the particle sizes, and the morphology were measured as a function of the laser irradiation period in order to monitor the changes from the two-component mixtures into the alloy colloids. The average particle sizes of the 1∶1 and 2∶1 metal ratios were 3.9 ± 1.2 nm and 4.3 ± 1.1 nm, respectively, for Au/Pd and 7.3 ± 2.0 nm and 5.4 ± 2.0 nm, respectively, for Ag/Pd. However, Ag/Pd with a 2∶1 ratio had difficult in forming dispersed spherical particles and the expected compositions. In addition to EDX analysis, X-ray diffraction measurements were carried out to confirm the formation of the alloy nanoparticles.

A new approach for the formation of alloy nanoparticles: laser synthesis of gold-silver alloy from gold-silver colloidal mixtures†

A new methodology has been developed to synthesize ngold–silver alloy nanoparticles by laser irradiation of mixtures nconsisting of gold and silver nanoparticles.

Dual drug-eluting stents coated with multilayers of hydrophobic heparin and sirolimus

Polymer coatings for stents are considered one of the key factors that lead to adverse cardiac events after coronary arterial stenting. This study presents a dual drug-eluting stent (DES) that is coated with multilayers of Duraflo heparin and sirolimus but containing no other organic polymers. The hydrophobic Duraflo heparin coating was used to improve the hemocompatibility of the stent and serve as a drug reservoir for the controlled release of sirolimus, thus avoiding inflammatory reactions induced by the conventional polymers. The Duraflo heparin and sirolimus were coated layer-by-layer onto the stent surface using a homemade spray-coating device. The drug loading amount can be easily controlled by adjusting the numbers of layers applied and the concentration of the drug solution, indicating the developed coating process is reproducible and well-controlled. After balloon expansion, the coating did not crack or peel off, which demonstrates that the sirolimus/Duraflo heparin coating layers tightly adhere to the stent surface. The activated partial thromboplastin time (APTT) assay showed that the Duraflo heparin coating significantly prolonged the APTT from 27.3 ± 0.3 s to 69.7 ± 6.2 s, demonstrating the anticoagulant ability of the coated stents. The dual DES exhibited a nearly linear sustained-release profile of Duraflo heparin and an initial burst release followed by a slow release of sirolimus. Less than 15% of heparin was released from the DES within 14 days, indicating the stent can maintain its antithrombotic surface for a long time. Because of the layer-by-layer structure, the most outer layer of Duraflo heparin coating may act as a diffusion barrier to retard sirolimus release from the stent. These results confirm that the dual DESs enable simultaneous delivery of antithrombotic and antiproliferative drugs and have potential for the treatment of coronary artery disease.

Pharmacokinetic applications of capillary electrophoresis.

This review briefly discusses the use of capillary electrophoretic (CE) methods for the investigations of different aspects of pharmacokinetics. In most investigations, CE was the method of choice because of its unique features, including high resolving power for chiral or metabolite separation, small sample volume for pediatric pharmacokinetics or for cell‐based investigations, in situ microdialysis sampling for rapid eliminations, low UV wavelength detection for nonderivatized analytes, fast and simplified sample processing for existing methods that require tedious sample preparation, or as a second method for verifications. Moreover, instrumental aspects of CE‐based assays for pharmacokinetic studies, such as different modes of CE methods for analyzing biological samples, sample stacking for increasing detection sensitivity, and coupling techniques with microdialysis and mass spectrometry, are also discussed in this review. Furthermore, the advantages and limitations of CE methods as well as the future outlook for pharmacokinetic studies are summarized.