Sang Wook Son

Highly Sensitive Immunoassay of Lung Cancer Marker Carcinoembryonic Antigen Using Surface-Enhanced Raman Scattering of Hollow Gold Nanospheres

A quick and reproducible surface-enhanced Raman scattering (SERS)-based immunoassay technique, using hollow gold nanospheres (HGNs) and magnetic beads, has been developed. Here, HGNs show strong enhancement effects from individual particles because hot spots can be localized on the pinholes in the hollow particle structure. Thus, HGNs can be used for highly reproducible immunoanalysis of cancer markers. Magnetic beads were used as supporting substrates for the formation of the immunocomplex. This SERS-based immunoassay technique overcomes the problem of slow immunoreaction caused by the diffusion-limited kinetics on a solid substrate because all of the reactions occur in solution. For the validation of our SERS immunoassay, a well-known lung cancer marker, carcinoembryonic antigen (CEA), was used as a target marker. According to our experimental results, the limit of detection (LOD) was determined to be 1-10 pg/mL, this value being about 100-1000 times more sensitive than the LOD of enzyme-linked immunosorbent assay. Furthermore, the assay time took less than 1 h, including washing and optical detection steps.

Surface-enhanced Raman scattering imaging of HER2 cancer markers overexpressed in single MCF7 cells using antibody conjugated hollow gold nanospheres

Antibody-conjugated hollow gold nanospheres (HGNs) have been used for the SERS imaging of HER2 cancer markers overexpressed in single MCF7 cells. SERS mapping images show that HGNs have much better homogeneous scattering properties than silver nanoparticles. The results demonstrate the potential feasibility of HGNs as highly sensitive and homogeneous sensing probes for biological imaging of cancer markers in live cells.

SERS imaging of HER2-overexpressed MCF7 cells using antibody-conjugated gold nanorods

Antibody-conjugated gold nanorods (GNRs) have been used for the targeting and imaging of specific cancer markers expressed on the surface membrane of cancer cells. GNRs with various aspect ratios were fabricated, and their surface-enhanced Raman scattering enhancement effects were evaluated. To attach the GNRs selectively onto the targets in cancer cells, specific antibodies were immobilized on the surface of GNRs using the layer-by-layer deposition method. First, Raman reporter molecules, mercaptopyridine, were attached to the surface of GNRs, and their surface charge was modified using poly(sodium 4-styrene-sulfonate) to make the surface charge negative. Then, anti-rabbit IgGs were immobilized onto the surface of the GNRs by electrostatic interactions. HER2 markers, expressed on the cell surface, were treated with anti-HER2 primary antibodies. Finally, the functionalized nanoprobes, conjugated with secondary antibodies, were attached to the markers on cancer cells by antibody-antibody interactions. In the present study, MCF7 cells overexpressing breast cancer marker HER2 were used as the optical imaging targets. Our experimental results demonstrate the potential feasibility of antibody-conjugated GNRs for the highly sensitive targeting and imaging of biomarkers expressed on the surface membrane of cancer cells.

Effect of the size and surface charge of silica nanoparticles on cutaneous toxicity

Silica nanoparticles (NPs) are widely applied in many fields, such as chemical industry, medicine, cosmetics, and agriculture. However, the hazardous effects of silica NPs exposure are not completely understood. In this study, the two different sizes (20 nm and 100 nm) and different charges (negatively charged [NC] and weakly negatively charged [WNC]) of silica NPs were used. The present study investigated the cytotoxicity and reactive oxygen species (ROS) generation of silica NPs on keratinocytes. The phototoxicity test of silica NPs was performed on skin fibroblast cells. In addition, skin irritation and skin sensitization of silica NPs were studied on HSEM and mouse skin, respectively. The cell viability of NC 20 nm silica NPs was decreased. However, there are no cytotoxicity for NC 100 nm silica NPs and WNC silica NPs (20 and 100 nm). The results for silica NPs-induced ROS generation are consistent with the cytotoxicity test by silica NPs. Further, NC and WNC silica NPs induced no phototoxicity, acute cutaneous irritation, or skin sensitization. These results suggested that silica NPs-induced ROS generation was the determinant of cytotoxicity. This study showed that the smaller size (20 nm) of silica NPs had more toxicity than the larger size (100 nm) of silica NPs for NC silica NPs. Moreover, we observed an effect of surface charge in cytotoxicity and ROS generation, by showing that the NC silica NPs (20 nm) had more toxic than the WNC silica NPs (20 nm). These findings suggested that the surface charge of silica NPs might be the important parameter for silica NPs-induced toxicity. Further study is needed to assess the effect of surface modification of nanotoxicity.

Assessment of penetration of quantum dots through in vitro and in vivo human skin using the human skin equivalent model and the tape stripping method.

Quantum dots (QDs) are rapidly emerging as an important class of nanoparticles (NPs) with potential applications in medicine. However, little is known about penetration of QDs through human skin. This study investigated skin penetration of QDs in both in vivo and in vitro human skin. Using the tape stripping method, this study demonstrates for the first time that QDs can actually penetrate through the stratum corneum (SC) of human skin. Transmission electron microscope (TEM) and energy diverse X-ray (EDX) analysis showed accumulation of QDs in the SC of a human skin equivalent model (HSEM) after dermal exposure to QDs. These findings suggest possible transdermal absorption of QDs after dermal exposure over a relatively long period of time.

Up-regulation of TNF-alpha secretion by cigarette smoke is mediated by Egr-1 in HaCaT human keratinocytes

Please cite this paper as: Up‐regulation of TNF‐alpha secretion by cigarette smoke is mediated by Egr‐1 in HaCaT human keratinocytes. Experimental Dermatology 2010; 19: e206–e212.

Assessment of dermal toxicity of nanosilica using cultured keratinocytes, a human skin equivalent model and an in vivo model.

Assessments of skin irritation potentials are important aspects of the development of nanotechnology. Nanosilica is currently being widely used for commercial purposes, but little literature is available on its skin toxicity and irritation potential. This study was designed to determine whether nanosilica has the potential to cause acute cutaneous toxicity, using cultured HaCaT keratinocytes (CHK), a human skin equivalent model (HSEM), and invivo model. Nanosilica was characterized by scanning electron microscopy. We evaluated the cytotoxic effects of nanosilica on CHKs and the HSEM. In addition, we also investigated whether two commercially available nanosilicas with different sizes (7 and 10-20 nm) have different effects. To confirm invitro results, we evaluated the irritation potentials of nanosilicas on rabbit skin. Nanosilicas reduced the cell viabilities of CHKs in a dose-dependent manner. However, the HSEM revealed no irritation at 500 microg/ml of nanosilica. Furthermore, this result concurred with Draize skin irritation test findings. The present study data indicate that nanosilica does not cause acute cutaneous irritation. Furthermore, this study shows that the HSEM used provides more useful screening data than the conventional cell culture model on the relative toxicities of NPs.

Analysis for the potential of polystyrene and TiO2 nanoparticles to induce skin irritation, phototoxicity, and sensitization.

The human skin equivalent model (HSEM) is well known as an attractive alternative model for evaluation of dermal toxicity. However, only limited data are available on the usefulness of a HSEM for nanotoxicity testing. This study was designed to investigate cutaneous toxicity of polystyrene and TiO2 nanoparticles using cultured keratinocytes, a HSEM, and an animal model. In addition, we also evaluated the skin sensitization potential of nanoparticles using a local lymph node assay with incorporation of BrdU. Findings from the present study indicate that polystyrene and TiO2 nanoparticles do not induce phototoxicity, acute cutaneous irritation, or skin sensitization. Results from evaluation of the HSEMs correspond well with those from animal models. Our findings suggest that the HSEM might be a useful alternative model for evaluation of dermal nanotoxicity.

ZnO nanoparticles induce TNF-α expression via ROS-ERK-Egr-1 pathway in human keratinocytes

BACKGROUND The area of nanotechnology continues to expand rapidly and zinc oxide (ZnO) nanoparticles (NPs) are widely being used in cosmetics and sunscreens. Although ZnO-NPs are considered materials that can potentially cause skin inflammation, the underlying mechanisms remain elusive. OBJECTIVE The aim of this study was to investigate the signaling pathways of a cutaneous inflammatory response induced by ZnO-NPs. ZnO-NPs increased the early growth response-1 (Egr-1) expression, promoter activity and its nuclear translocation in HaCaT cells. METHODS HaCaT cells and primary keratinocytes were exposed to ZnO NPs over a range of doses and time course. Protein levels and mRNA levels of Egr-1 and mitogen-activated protein kinase (MAPK) were measured by Western blot and ELISA, respectively. As an in vivo study, ZnO-NPs were applicated on mouse skin, and immunohistochemical stain with TNF-α and Egr-1 was done. RESULTS ZnO-NPs activated extracellular signal-regulated kinase (ERK) of MAPK pathways. The up-regulation of Egr-1 expression by ZnO-NPs stimulation was found to be inhibited by an ERK inhibitor, but by neither c-Jun-N-terminal kinase (JNK) nor p38 inhibitor. Antioxidative N-acetyl-cysteine (NAC) strongly inhibited the level of Egr-1 and phosphorylated ERK expression in ZnO-NPs treated cells. ZnO NPs also increased tumor necrosis factor (TNF)-α expression and secretion, which were inhibited by the blockade of Egr-1 expression. CONCLUSIONS The present study demonstrated that ZnO-NPs might induce inflammatory response via ROS-ERK-Egr-1 pathway in human keratinocytes.

Influence of surface charge of gold nanorods on skin penetration

The skin plays an important role as a protective barrier against toxic environments and also is a route of drug administration. In spite of evidence for and interest in the skin penetration of nanoparticles, no study has examined the effect of nanoparticle surface charge on percutaneous absorption. In this study, we investigated the effect of surface charges of gold nanorods (GNs) on skin penetration.