Yong Taik Lim

Acute toxicity and pharmacokinetics of 13 nm-sized PEG-coated gold nanoparticles

In general, gold nanoparticles are recognized as being as nontoxic. Still, there have been some reports on their toxicity, which has been shown to depend on the physical dimension, surface chemistry, and shape of the nanoparticles. In this study, we carry out an in vivo toxicity study using 13 nm-sized gold nanoparticles coated with PEG (MW 5000). In our findings the 13 nm sized PEG-coated gold nanoparticles were seen to induce acute inflammation and apoptosis in the liver. These nanoparticles were found to accumulate in the liver and spleen for up to 7 days after injection and to have long blood circulation times. In addition, transmission electron microscopy showed that numerous cytoplasmic vesicles and lysosomes of liver Kupffer cells and spleen macrophages contained the PEG-coated gold nanoparticles. These findings of toxicity and kinetics of PEG-coated gold nanoparticles may have important clinical implications regarding the safety issue as PEG-coated gold nanoparticles are widely used in biomedical applications.

Pathobiological features of a novel, highly pathogenic avian influenza A(H5N8) virus.

The endemicity of highly pathogenic avian influenza (HPAI) A(H5N1) viruses in Asia has led to the generation of reassortant H5 strains with novel gene constellations. A newly emerged HPAI A(H5N8) virus caused poultry outbreaks in the Republic of Korea in 2014. Because newly emerging high-pathogenicity H5 viruses continue to pose public health risks, it is imperative that their pathobiological properties be examined. Here, we characterized A/mallard duck/Korea/W452/2014 (MDk/W452(H5N8)), a representative virus, and evaluated its pathogenic and pandemic potential in various animal models. We found that MDk/W452(H5N8), which originated from the reassortment of wild bird viruses harbored by migratory waterfowl in eastern China, replicated systemically and was lethal in chickens, but appeared to be attenuated, albeit efficiently transmitted, in ducks. Despite predominant attachment to avian-like virus receptors, MDk/W452(H5N8) also exhibited detectable human virus-like receptor binding and replicated in human respiratory tract tissues. In mice, MDk/W452(H5N8) was moderately pathogenic and had limited tissue tropism relative to previous HPAI A(H5N1) viruses. It also induced moderate nasal wash titers in inoculated ferrets; additionally, it was recovered in extrapulmonary tissues and one of three direct-contact ferrets seroconverted without shedding. Moreover, domesticated cats appeared to be more susceptible than dogs to virus infection. With their potential to become established in ducks, continued circulation of A(H5N8) viruses could alter the genetic evolution of pre-existing avian poultry strains. Overall, detailed virological investigation remains a necessity given the capacity of H5 viruses to evolve to cause human illness with few changes in the viral genome.

Synthesis and Characterization of a Photoluminescent Nanoparticle Based on Fullerene–Silica Hybridization

Bright lights: Fullerene-silica hybrid nanoparticles have bright photoluminescence, high photostability, and low cytotoxicity, which are assets for bioimaging agents. The origin of the photoluminescence of the nanoparticle is the C-O-Si bond (see picture).

Noninvasive imaging of dendritic cell migration into lymph nodes using near-infrared fluorescent semiconductor nanocrystals

Effective tracking of immunotherapeutic cells is essential for monitoring the migration of injected cells to the target tissue. Here we report the use of near‐infrared (NIR) ‐emitting fluorescent semiconductor nanocrystals, called quantum dots (QDs), for noninvasive in vivo tracking of dendritic cell (DC) migration into lymph nodes. The effect of QDs on DC viability and maturation was systematically investigated using MTT assays and FACS analysis. We found that the labeling of DCs with QDs had no effect on DC phenotype or maturation potential. Cytokine and migration assays revealed that there were no significant changes in either cytokine production or chemokine‐dependent migration of QD‐labeled DCs relative to unlabeled cells; in both labeled and unlabeled cells, cytokine production and migratory capacity was increased by stimulation with lipopolysaccharide. Furthermore, QDs did not influence allogenic naive T cell activation or uptake of exogenous antigens. Notably, we also demonstrated that it was possible to track QD‐labeled DCs injected into the footpad into popliteal and inguinal lymph nodes using NIR fluorescence. Taken together, our protocols establish the potential of noninvasive in vivo imaging of NIR‐emitting QDs for tracking immunotherapeutic cells.— Noh, Y.‐W., Lim, Y. T., Chung, B. H. Noninvasive imaging of dendritic cell migration into lymph nodes using near‐infrared fluorescent semiconductor nanocrystals. FASEB J. 22, 3908–3918 (2008)

A plasmonic biosensor array by block copolymer lithography

Highly uniform and dense, hexagonal noble metal nanoparticle arrays were achieved on large-area transparent glass substrates via scalable, parallel processing of block copolymer lithography. Exploring their localized surface plasmon resonance (LSPR) characteristics revealed that the Ag nanoparticle array displayed a UV-vis absorbance spectrum sufficiently narrow and intense for biosensing application. A highly-sensitive, label-free detection of prostate cancer specific antibody (anti-PSA) with sub-ng ml−1 level detection limit (0.1∼1 ng ml−1) has been accomplished with the plasmonic nanostructure. Our approach offers a valuable route to a low-cost, manufacture-scale production of plasmonic nanostructures, potentially useful for various photonic and optoelectronic devices.

Synthesis and high performance of magnetofluorescent polyelectrolyte nanocomposites as MR/near-infrared multimodal cellular imaging nanoprobes.

Here, we describe an easy but robust chemical strategy to synthesize high-performance magnetic resonance (MR)/near-infrared (NIR) multimodal imaging nanoprobes. Poly(γ-glutamic acid) was used for the convenient phase transfer of MnFe(2)O(4) nanoparticles dispersed in organic solvents into aqueous solutions and facilitated further ionic gelation with poly(l-lysine). During the gelation process, MnFe(2)O(4) nanoparticulate satellites were encapsulated in the ionic nanocomplex, which induced synergistic magnetism and resulted in huge T(2) relaxivity (r(2)). The positively charged outer surfaces were assembled with other negatively charged NIR emitting fluorescent nanocrystals and enabled the highly efficient delivery of the magnetofluorescent polyelectrolyte nanocomposites (MagFL-PEN) into cancer cells. The enhancement of negative contrast of MagFL-PEN at 2 μg/mL concentration was similar to that of Resovist at 20 μg/mL concentration. The NIR fluorescence microscopy images of the MagFL-PEN-labeled cells even at 12.5 pM were able to be clearly observed. The labeling efficiency of MagFL-PEN was approximately 65-fold higher compared to that of the commercialized fluorescent nanocrystals, only after 3 h incubation period, even at the test concentration (100 pM). Due to the high-performance capabilities both in materials properties and cell labeling efficiency, the MagFL-PEN is expected to be used as a highly efficient MR/NIR dual-modality imaging nanoprobe in the detection of cancer cells and monitoring of therapeutic cells in vivo.

Simultaneous in vivo tracking of dendritic cells and priming of an antigen-specific immune response

We report the fabrication of a one-pot antigen system that delivers antigen to dendritic cells (DCs) and tracks their in vivo migration after injection. Multifunctional polymer nanoparticles containing ovalbumin protein, magnetic resonance imaging contrast agents (iron oxide nanoparticles), and near-infrared fluorophores (indocyanine green, ICG), MPN-OVA, were prepared using a double emulsion method. The MPN-OVA was efficiently taken up by the dendritic cells and subsequently localized in the lysosome. Flow cytometry analysis revealed an increase in the uptake of OVA antigen by MPN-OVA at 37 °C, when compared with soluble OVA protein. We found that MPN-OVA had no effect on DC surface expression of MHC class I, costimulatory (CD80, CD86) or adhesion (CD54) molecules or the ability of DCs to mature in response to LPS. Following the uptake of MPN-OVA, exogenous OVA antigen was delivered to the cytoplasm, and OVA peptides were presented on MHC class I molecules, which enhanced OVA antigen-specific cross-presentation to OT-1 T cells and CD8OVA1.3 T cell hybridoma in vitro. The immunization of mice with MPN-OVA-treated DCs induced OVA-specific CTL activity in draining lymph nodes. The presence of MPN allowed us to monitor the migration of DCs via lymphatic drainage using NIR fluorescence imaging, and the homing of DCs into the lymph nodes was imaged using MRI. This system has potential for use as a delivery system to induce T cell priming and to image DC-based immunotherapies.

Multiplexed imaging of therapeutic cells with multispectrally encoded magnetofluorescent nanocomposite emulsions.

Here, we describe the fabrication of multispectrally encoded nanoprobes, perfluorocarbon (PFC)/quantum dots (QDs) nanocomposite emulsions, which could provide both multispectral MR and multicolor optical imaging modalities. Our strategy exploited the combination of the multispectral MR properties of four different PFC materials and the multicolor emission properties of three different colored CdSe/ZnS QDs. The PFC/QDs nanocomposite emulsions were fabricated by exchanging hydrophobic ligands coated onto CdSe/ZnS QDs using 1H,1H,2H,2H-perfluorooctanethiol, which renders the QDs to be dispersible in the PFC liquids. To provide biocompatibility, the PFC liquids containing QDs were emulsified into aqueous solutions with the aid of phospholipids. The distinct (19)F-based MR images of PFC/QDs nanocomposite emulsions were obtained by selective excitation of the nanocomposite emulsions with magnetic resonance frequency of each PFC, while a specific fluorescence image of them could be selected using appropriate optical filters. The uptake of PFC/QDs nanocomposite emulsions was high in phagocytic cells such as macrophages (90.55%) and dendritic cells (85.34%), while it was low in nonphagocytic T cells (33%). We have also shown that the nanocomposite emulsions were successfully applied to differentially visualize immunotherapeutic cells (macrophages, dendritic cells, and T cells) in vivo. The PFC/QDs nanocomposite emulsions are expected to be a promising multimodality nanoprobe for the multiplexed detection and imaging of therapeutic cells both in vitro and in vivo.

Programmed nanoparticles for combined immunomodulation, antigen presentation and tracking of immunotherapeutic cells

We report programmed nanoparticles (pNPs) that can tailor the immunotherapeutic function of primary bone marrow-derived dendritic cells (BMDCs) by ex vivo combined immunomodulation and track the in vivo migration of them after injection into body. Because DCs are the most effective antigen-presenting cells (APCs) that are able to present the antigens to T cells that contribute to tumor rejection, the maturation and monitoring of therapeutic DCs are essential for the efficient cancer immunotherapy. For combined immunomodulation of DCs, poly (lactic-co-glycolic acid) (PLGA) NPs containing both small interfering RNA (siRNA) for the knock-down of immune-suppressor gene (signal transducer and activator of transcription-3, STAT3) of DCs and an immune response modifier (imiquimod, R837) for the activation of DCs through the toll-like receptor 7 (TLR7) were developed. To deliver tumor antigen-specific information to DCs ex vivo and track the migration of DCs in vivo, another type of PLGA NPs containing tumor model antigen (ovalbumin, OVA) and near-infrared (NIR) fluorophores (indocyanine green, ICG) were also fabricated. These pNPs were taken up efficiently by DCs and various cytokines were expressed in matured DCs. DCs treated with pNPs also efficiently presented antigen-peptide to CD8 OVA 1.3 T cells through cross-presentation. Immunization of mice with these pNPs-treated DCs induced OVA-specific cytotoxic T lymphocytes (CTL) activity against the EG7-OVA tumor model and inhibited tumor growth efficiently. In addition, the migration of PLGA NPs-treated DCs to lymph nodes was monitored by NIR imaging technique. These multifunctional pNPs represent a promising technology for the combined immunomodulation and antigen-specific tumor therapy.

Quantum dot‐based protein micro‐ and nanoarrays for detection of prostate cancer biomarkers

In this article, we demonstrate the fabrication and detection of cancer protein biochips consisting of micro‐ and nanoarrays whereby pegylated quantum dots (QDs) conjugated to antibodies (Abs) of prostate specific antigens (PSA) were used for the detection of clinical biomarkers such as PSA. BSA which acts as an efficient blocking layer in microarrays, tends to show an interaction with QDs. In view of this fact, we investigated two series of samples which were fabricated in the presence and absence of BSA blocking layer. Variation in the incubation time required for the antigen–antibody interaction to take place, different proteins as controls and the effect of bare QDs on these microarrays, were the three main parameters which were studied in these two series. Samples fabricated in the absence of BSA blocking layer exhibited an extremely high specificity in the detection of cancer proteins and were also marked by negligible nonspecific binding effects of QDs, in stark contrast to the samples fabricated using BSA as a blocking layer. Fabrication of nanoarrays of QD‐conjugated PSA Abs having a spot size of nearly 900 nm has also been demonstrated. Thus, we show the potential offered by QDs in in vitro analysis of cancer biomarker imaging.