Earticle

DNA methylation is an epigenetic mechanism of regulation, which is responsible for transcriptional silencing of tumor suppressor genes. Without any change of DNA sequence, phenotype can be differentiated by epigenetic changes, and the changes are heritable through cell cycles and generations. Therefore, heterogeneity of DNA pattern in a specific tissue provides significant biological information. However, there is no technology which can detect those heterogeneous patterns quantitatively although DNA methylation detection method in omogeneous sample is available. High-resolution capillary electrophoresis-based single strand conformation polymorphism analysis (CE-SSCP) system has great potentials in quantitative DNA methylation analysis since sequence change can be easily detected by mobility change. In this study, we developed a quantitative DNA methylation analysis method based on high-resolution CE-SSCP system combined with sodium bisulfite conversion using E-cad and p14 gene as a model. In a single reaction, a mixture of various methylation patterns could be easily analyzed and this result indicates that this technique has a huge potential as an epigenetic diagnosis system.

A cell-based electrical detection of drugs or toxicant effects is promising technology because of its simplicity and convenience with short detection time. In this study, we fabricated a neural cell chip to investigate dose-dependent toxic effects of bisphenol-A on PC12 Cells. A non-toxic plastic chamber was fixed on a gold electrode deposited silicon bottom, which was used as working electrode in three electrode systems. Electrochemical behavior of neural cancer cells were measured using cyclic voltammetry (CV) after 24h of incubation. The cathodic peak (Ipc) from CV was used as a parameter representing the effects of bisphenol-A on cells viability. The results show that bisphenol-A have stimulatory effects on neuronal cells at lower dose than 150nM but have cytotoxic effect at higher dose than 150nM. These dual effects of bisphenol-A on PC12 cells were confirmed by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and western blot analysis with maintaining similar culture condition. This developed electrochemical analysis system is a promising tool for drug screening and environmental toxicant detection. Acknowledgments: This research was supported by the Korea Science and Engineering Foundation (KOSEF) grant funded by the Korea government (MEST) (2009-0080860) and by the Nano/Bio science & Technology Program (M10536090001-05N3609-00110) of the Ministry of Education, Science and Technology (MEST) and by Ministry of Environment of the Republic of Korea as "The Eco-technopia 21 project"

AbstractWe fabricated the proein-based biomemory device by an introduction of the cytochrome c. The cytochrome c is an iron storage protein of organisms and has redox property. 11- mercaptoundecanoic acid was used as a linker material to immobilize the cytochrome c on the Au substrates. The immoibilized surface was confirmed by surface plasmon resonance spectroscopy. And the surface topography of cytochrome c layer on Au surface was investigated by scanning tunneling microscopy. The redox property of cytochrome c molecules immobilized onto Au surface using chemical linker was demonstrated by using cyclic voltammetry technique. The fabricated memory device shows the two type of memory characteristics well. Its memory effects were determined by using the chronoamperometry and open circuit potential amperometry. In these results, it could be concluded that the cytochrome c layer could be used for the construction of nanobiochip with memory function. Acknowledgments : This research was supported by the Nano/Bio Science & Technology Program (M10536090001-05N3609- 00110) of the Ministry of Education, Science and Technology (MEST), by the Korea Science and Engineering Foundation (KOSEF) grant funded by the Korea government (MEST) (2006- 05374), and by the Ministry of Knowledge Economy (MKE) and Korea Industrial Technology Foundation (KOTEF) through the Human Resource Training Project for Strategic Technology.References1. Jeong-Woo Choi, Byung-Keun Oh, Junhong Min, Young Jun Kim, Protein-based biomemory device consisting of the cysteine-modified azurin(2007), Appl. Phys. Lett., 91, 263902-1-263902-3

To identify novel genes that might be involved in tyrosine hydroxylase (TH) gene expression a preliminary microarray analysis of differential gene expression has been carried out on the olfactory bulbs (OBs) ipsilateral and contralateral to naris closure. Several developmental genes showing signifi cant differential regulation were identified based on microarrays analysis of OB mRNAs by Affymetrix GeneChip Mouse genome 430 2.0 arrays. These include Pbx1, pre-B-cell leukemia homeobox 1, Nkx2.1, also known as thyroid transcription factor 1, Evx1, the murine homologue of the even-skipped gene, and Prop1, a pituitary-specific paired-like homeodomain transcription factor. In addition, 36 ESTs for functionally uncharacterized genes were upregulated and 45 downregulated between 1.8 and 2.5 fold by naris closure. The expression of these genes and EST clones was confirmed by in situ hybridization and Northern blot techniques.

We developed a novel carbon paste electrode (CPE) utilizing enzymeincorporated mesoporous carbon material. MSU-F-C having 25 nm inner pore size was synthesized by generally accepted method. Glucose oxidase (GOX) was entrapped in the mesoporous carbon by crosslinking with glutaldehyde to prevent leaking of enzyme. The paste formed by mixing of GOX-incorporated carbon and mineral oil was filled in 5 mm diameter tube. As a result, our GOX-mesoporous CPE showed reliable activity after 4 weeks storage in room condition, otherwise the activity of a trivial enzyme-CPE fabricated by just mixing of enzyme and carbon material was eliminated within 6 days. Also, it have shown reliable dependency according to the glucose concentration ranging from 0~100 mM and it is a reasonable detection range because a normal glucose level in human blood is about 5 mM. Our GOX-mesoporous CPE could provide a great benefit to the glucose monitoring field and it could also introduce a novel concept to improve the stability of enzyme-incorporated electrode.

In this report, we described a immunoassay method for CRP antigen detection by scFv conjugated AuNPs. Gold nanoparticles which are conjugated with anti-CRP scFvs. The functional scFv was expressed in cell-free protein synthesis system derived from an E. coli extract. And then the scFv was isolated by affinity-tag purification method. We have showed that the functional scFv was successfully prepared. And the scFvs could be used for direct detection its antigen in western blots when stained with anti-CRP scFvs. These results show that the cellfree system can produce a functional scFv and the anti-CRP scFv conjugated AuNPs could be used for detection of CRP antigen on the SPR surface. SPR measurements for detection of antigens were also possible with the use of this immobilized surface. The results suggested that the gold nanoparticle conjugation can be amplified the signal to SPR.

Endocrine disruptor chemicals (EDCs) are exogeous substances which can disrupt the physiologic function of hormones in the endocrine system. Phage display technique can be used for selecting specific peptide for detection and elimination of these chemicals. However, the traditional phage display methods, typically relying on batch adsorption/desorption procedures, provide only a single equilibrium stage for one round of selection, thus necessitating multiple rounds of repeated selection. Comparing to traditional method, phage display based on chromatographic biopanning using FPLC system provides a number of equilibrium stages, significantly increasing the screening efficiency. In our experiment, positive biopanning against Pb2+ as an endocrine disrupter chemical and negative biopanning to eliminate cross-binding peptides with non target metal ions such as Ni2+, Cu2+, Co2+, Fe3+ were performed. 17 Pb2+-affinity peptide binders have been identified and the relative affinity strength of these 17 peptide binders to Pb2+ is further determined through bead-based enzyme immunoassay (EIA) and number 3, 4, 1 and 5 peptide binders show higher affinity to Pb2+ than the others or the control peptide To the best of our knowledge, this study is the first to report Pb2+-specific peptide binders, The selective Pb2+ cognitive peptide moieties are expected to provides many potential applications including biosensors and biosorbent for detection and removal of endocrine disruptors.

Cell-based assays are increasingly being used in drug discovery, genetic analysis and single-cell analysis. Cell microarrays can be classified into two general categories: chip-based planar arrays and particle-based suspension arrays. Although chip-based planar arrays have been widely used, suspension arrays are becoming popular as an alternative approach due to several advantages over planar array. In this study, we prepared suspension microarrays of cells using various shape of SU-8 microparticles and demonstrate the potential use of this system for multiplexed cell-based assay. Different size and shape of SU-8 50 negative photoresist was micropatterned on the glass slide using typical photolithography. SU-8 microparticles were obtained by detaching micropattern by ultrasonication. Resultant microparticles were incubated with different cell types. Cells adhered on SU-8 microparticles were observed with optical microscopy and a live/dead viability/cytotoxicity fluorescence assay. In this study, three different cell types were immobilized on the three different shapes of microparticles so that each cell type was easily identified. Capability of controlling the shapes of microparticles opened the possibility of producing shape-coded suspension array, which can offer a costeffective and versatile platform for high-throughput and multiplexed cell-based assay.

Accumulation of heavy metals is one of the major concerns for the environmental safety because of their toxicity and non-degradability. In particular, cadmium and several cadmium-containing compounds are known as carcinogens and can induce many types of cancer. Among promising strategies of cadmium removal, one emerging technology is the use of tunable, metal binding biopolymers based on elastin-like polypeptides (ELPs). An ELP consisting of the repeating pentapeptide of specific amino acids, Val-Pro-Gly-Xaa-Gly (where the “guest residue” Xaa is any amino except Proline), undergoes a reversible phase transition at a specific temperature (transition temperature, Tt). For effective elimination of cadmium, synthetic phytochelatin (EC), which is a metal-binding protein and has a repetitive motif (Glu-Cys)nGly, was fused with the ELP. We expected that a fusion protein constituting ELP and EC can detect and remove cadmium without any complex process. In this study, a fusion protein was expressed in E.coli and metal binding ability of EC to cadmium was examined quantitatively. In addition, transition temperature variation when the fusion protein binds cadmium was analyzed.

Manufacturing of DNA leads the field of nanotechnology, because of its versatility and programmability, being able to self-assemble into nanoscale structures based on specific complementarity between basese. We constructed adenosine DNA-nanosensor using aptamers, nucleic acids that bind specifically to target biomolecules such as small molecules and proteins with binding affinity as high as antibodies. The secondary structure of the aptamer undergoes a dramatic change upon binding to a target biomolecule, which renders biosensing applications possible. Here we constructed an aptamer-based nanosensor of which fluorescence signal is responsive to adenosine. The DNA nanosensor consists of two oligonucleotides; a FAM-labeled oligonucleotide carrying an adenosine aptamer sequence and its complementary one labeled with a quencher, BHQ1. Hybridization between the two oligonucleotides leads to a fluorescence-quenched duplex which undergoes adenosine-induced dissociation and accordingly results in an increase of FAM fluorescence. Detection sensitivity of the aptamer-based nanosensor depends on the total number of base paring in the duplex region as well as the length of aptamer segment in the duplex region. As a practical application, we demonstrated that the adenosine nanosensor can be used to quantify adenosine deaminase which is clinically important. It is known that mutations in the adenosine deaminase gene are involved in many diseases such as severe combined immunodeficiency, hemolytic anemia and autism. As ADA converts adenosine into inosine that does not affect the hybridization state of the nanosensor, fluorescence emission increases because of fast binding equilibrium between adenosine and the aptamer sequence embedded in the nanosensor. The higher concentration of ADA gave more rapid decrease in the fluorescent intensity, indicating that ADA concentration can be evaluated by monitoring the fluorescence intensity. Our results illustrate DNA aptamer can be exploited to design sensor materials, permitting sensitive detection of biochemicals that are difficult to be measured by conventional biochemical assay methods.

It is important to monitor or measure toxicity of environmental pollutants or new drugs. Cell chip technology has become a promising tool because high-throughput and sensitive screening is possible. In the present work, we propose "gluing" method for cell chip construction by displaying mussel adhesive proteins (MAPs) on the surface of target cells (insect Drosophila S2 and mammalian Chinese hamster ovary (CHO) DG44). Mussels use their adhesive proteins, fp-5, in order to attach on the rock in the tough sea. In the present work, we present construction of recombinant baculovirus expression vector, rBV-FP5, infection of recombinant viruses to target cells for expression and location of MAP on cell surface, and construction of cell chip using MAP-displayed infected cells. Our proposed cell chip construction method does not need any harmful chemical linkers for cell immobilization. In addition, we would construct universal recombinant baculovirus expression system which can give adhesion ability to target cells. The constructed cell chip system is applicable for environmental toxicity sensor or drug screening at a cellular level.

We report on the development of an enzyme-amplified sandwich- type immunosensor based on a thin gold film sputtered on anodic porous niobium oxide (Au@Nb2O5). Redox cycling of enzymatically amplified electroactive species and a stable electrode consisting of Au@Nb2O5 are used in the electrochemical immunobiosensor for powerful signal amplification. We conclude that the novel electrochemical biosensor based on an Au@Nb2O5 electrode offers better performance than those based on conventional bulk gold or niobium oxide electrodes. Our novel approach does not require any time-consuming cleaning steps to yield reproducible signals. In addition, the adhesion of gold films on the substrate is very stable during electrochemical biosensing. Cyclic voltammetry measurements indicate that nonspecific binding of proteins to the streptavidin-modified surface is sufficiently low to be ignored in the case of our novel system. This novel biosensor offers enhanced biosensing performance with a high resolution and sensitivity.

Enzyme sensors for the on-site analysis of phenolic compounds, a class of polluting chemicals, were actively studied in the environmental fields. For the development of a microanalysis device to detect phenolic compounds, PHEMA hydrogel microparticles containing conjugates of enzyme (tyrosinase) and quantum dot as an optical biosensor were prepared by a dispersion photopolymerizatrion and the microchannel was fabricated with PDMS and filled with the particles. Using the fluorescence quenching property of quinone intermediates produced from the enzyme reaction between tyrosinase and phenol, the fluorescent responses of the quantum dot were investigated according to the phenol concentration. The activity and sensitivity of the enzyme and quantum dot were maintained when they were conjugated and encapsulated within the PHEMA hydrogel microparticles. The fluorescent response of the PHEMA hydrogel microparticles containing tyrosinase-quantum dot conjugates was affected by the concentration of phenol. These results indicate that the PHEMA hydrogel microparticles containing tyrosinasequantum dot conjugates prepared in this study have a possibility to be used as an optical biosensor to detect phenolic compounds.

Odorant perception begins from the specific interaction with olfactory receptors and odorants. Olfactory receptors, belonging to G-protein coupled receptor family, are integral membrane proteins composed of seven transmembrane spanning domains. The protein-based olfactory bioseonsors using olfactory receptors might be good tools for odorant detection. For developing protein-based olfactory sensor with high selectivity and sensitivity, olfactory receptor proteins were expressed in E. coli. Due to their complicated form, these proteins are hardly expressed in the inner membrane in E. coli and are majorly expressed as inclusion body form. Previous studies shows that the inclusion body form of olfactory receptor has function for odorant detection when it was applied to various sensing devices such as carbon nanotube, conducting nanopolymer and QCM. In this study, the olfactory receptor proteins OR520, which specifically binds to helional, are purified from E. coli using detergent and reconstituted into liposome to have more functional form. Odorant detection assay carried out using surface plasmon resonance (SPR). The inclusion body form and reconstituted form of the proteins are immobilized on the gold surface of SPR chip and the results of odorant detection were compared.

Parylene H layer was deposited on surface of protein chip to promote its immobilization efficiency. To construct immunoaffinity layer, protein chip made from glass substrate requires surface modifications such as silanization or depositing polymer film on substrates which have been developed1). Through preparing parylene H layer on glass substrate, aldehyde (-CHO) functional groups are exposed on surface of the protein chip. Aldehyde fuctional groups are make covalent bonding with most kinds of proteins and peptides which contain amine groups. Its effectiveness was determined by quantification of immobilized horseradish peroxidase (HRP) compared with polystyrene. For medical diagnostic experiment we detect antibodies against cyclic-citrullinated peptide (CCP) in sera from patients of rheumatoid arthritis (RA) on protein chip by using microarrayer and scanner. The results are compared with those of ELISA assay.

Isoflavones, a family of the major groups of phytoestrogens, are known to have anticarcinogenic, antioxidant and photoprotective effects. Although they have beneficial bioactivities for human health, low bioavailability and poor solubility have been problems in applications. To address this problems, we first have optimized large-scale conditions for enzymatic bio-conversion of crude isoflavone glycosides (≤20%) from soybean into aglycones by using a β-glucosidase. Further purification processes enabled us to obtain more than 90% purity of isoflavone aglycones analyzed by HPLC. The purified isoflavone showed to a have strong anti-inflammation activity. UV irradiation of HaCaT cells induced tumor necrotic factor-α (TNF-α) production by 3 folds but the treatment of the purified isoflavone reduced TNF-α production back to normal level. In order to make derivatives of isoflavone, which might have novel bioactivities with better solubility, acylation reactions of the purified isoflavone were first performed and the resulting isoflavone derivatives were purified for further analyses. Especially, di-palmitated isoflavone significantly reduced TNF-α production at much lower concentration than that of purified isoflavone. Further characterization of the di-palmitated isoflavone derivatives will be discussed.

Chronic exposure of Ultraviolet (UV) leads to a variety of skin damage such as sun-burn, pigmentation, premature aging and photocarcinogenesis. Moreover, the response of inflammation and photo-damage against UV induced skin can be aggravated by reactive oxygen species (ROS) induced by UV irradiation. Therefore, the importance of the protection effect by UV damage has been regarded continuously as well as the development of effective material that can mitigate the skin damage has been required. Ficus deltoidea, known as “Mas cotec”, has been widely used as several medications in Malaysia. Also, several studies have demonstrated that F. deltoidea has strong anti-oxidant activity. In this respect, our study focused on the photo-protective efficacy; particularly, we evaluated the antiinflammation effect of F. deltoidea extract on the UVB-induced skin cell. To investigate the anti-inflammatory effect, we examined the production of the inflammatory cytokines such as such as tumor necrotic factor-α (TNF-α), interleukin-1α (IL-1α), IL-6 by enzyme-linked immunosorbent assay (ELISA). Also, we examined the expression of COX-2 after UVB irradiation by western blot assay. We found that the amount of these inflammatory cytokines in the HaCaT keratinocytes was significantly reduced by treatment of F. deltoidea extract after UVB irradiation. Along with this result, treatment with F. deltoidea extract inhibited COX-2 expression by UVB irradiation in a dose dependent manner. These results point to the potential use of F. deltoidea extract as agent for cutaneous inflammation response

Lignins are the second most abundant three-dimensional amorphous natural nontoxic polymers, occurring in higher plants, mainly in wood. Lignin is mainly separated from wood during pulp and papermaking operations, where it serves primarily as fuel. Lignins vary in structure according to their plant source. However, because of complex and heterogenous structures of lignin, the industrial utilization of lignin has been limited. In this study, ultra-violet blocking and antioxidant effects of lignins extracted from different milled wood have been explored for more valuable applications of lignin. The tested samples were poplar (Populus albaglandulosa), yellow poplar (Liriodendron tulipifera), birch (Betula platyphylla), pitch pine (Pinus rigida), Miscanthus giganteus, and Miscanthus sinensis. The ultraviolet (UV) absorption of the milled wood lignins was observed using a spectrophotometer at a wavelength of 280 nm, 300 nm (UVB), and 340 nm (UVA). Their antioxidant capacity was determined by means of scavenging activity of free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH) and total phenol assay (Folin-Ciocalteu) method.

Citron (Citrus junos Sieb ex TANAKA) is a citrus fruit that is cultivated in northeast Asia. Citron is known for containing abundant antioxidants such as vitamin C, flavonoids, for example hesperidin and hesperetin, and terpenoids such as limononin. When mature citron is processed for tea or other beverage food products in Korea, massive amounts of seeds and pericarp are remained as waste. This study was aimed to exploit the processed remnant of Citron for developing functional cosmetic applications. Analyses of extracts of seeds and pericarp identified spots that have the same Rf values of limonin and hesperidin respetively analyzed by TLC. The two extracts did not show significant radical scavenging activities measured by DPPH(2,2-diphenyl-1- picrylhydrazyl) method. But citron extracts have much high content of phenolic compounds. Cultured human dermal fibroblasts and HaCaT keratinocytes were irradiated with 25mJ UVB and the citron extracts were added to the medium of each culture. Cellular damages caused by UVB irradiation were prevented by the addition of the citron extract. In addition, the reduction of the enhanced MMP1(Matrix Metalloproteinase) expression after irradiation of UVB in human dermal fibroblasts was observed. Also the increased level of of pro-inflammtory TNF-α in the UVB irradiated HaCaT cells were decreased. Further analyses of the effect of citron extracts on collagen production and the expression of other pro-inflammatory cytokines will be performed. In conclusion, citron extract seems to have anti-photoaging effect and are expected to be used as a potential anti-aging agent.

Ultraviolet (UV) A radiation is one of the most important environmental factors involved in the skin aging 1). However, the expression pattern of skin-aging related proteins and skin-aging mechanism in UVA irradiated-fibroblasts are yet to be investigated. Skin-aging mechanism of induced-fibroblast by UVA radiation was studied using proteome analysis. The fibroblasts were cultured in Dulbecco's Modified Eagle's Medium (DMEM) with or without UVA irradiation. The proteome of cultured human fibroblasts were profiled using 2D-PAGE 2). Isoelectric focusing was carried out using commercial immobilized dry strips 3-10, 24cm. SDS-PAGE was performed using 25 x 21cm, 12 % polyacrylamide gel. Gels were stained with silver nitrate, analyzed by image master 2D platinum 7. In conclusion, we could identify that many proteins affect the skin aging, were up or down-regulated by UVA irradiation.

Silk is a natural protein fiber composed of fibroin and sericin (1). Silk protein, which has a good absorbency and is also known to promote fibroblast growth (2), can be used for many cosmetic applications (3). Previous method to prepare a silk powder for a cosmetic material comprises the step of solubilizing the rigid silk fiber in alkali solution. To avoid such a destructive chemical process and prepare a natural cosmetic silk material, we utilized a silk protein in silk grand of silkworm, which physical characteristic is rather softer than silk fiber from a cocoon because it has not yet been fully oxidized. In this study, we extracted a silk protein in silk grand of Bombyx mori with aqueous protease solution, oxidized it with tyrosinase solution in emulsion in oil phase, and demonstrated to prepare a micro silk bead for the potential application of cosmetic materials.

Skin color is determined by the amount of melanin. Tyrosinase is a key enzyme for melanin synthesis (melanogenesis). Its transcription is started by MITF and E-box binding. MITF (Microphthalmia-associated transcription factor) is the basic helix-loop-helix-zip transcription factor that regulates gene expression by E-box binding. To screen depigmenting agents by a HTS (high-throughput screening) system, an EMSA (Electrophoretic Mobility Shift Assay) was performed. Based on the computer-simulated structure, 10 compounds isolated from natural plants and microbial sources were selected and were investigated as MITF-DNA binding inhibitors. First, we performed a cell test to check if the compounds have depigmenting activity. From the results of the melanin assay and MTT assay, 3 compounds exhibited depigmenting activity in a dose dependent manner without toxicity. Among these 3 compounds, we choose compound #7 for further experiments. To check its inhibitory activity against MITF-DNA binding, an EMSA (Electrophoretic Mobility Shift Assay) was performed. As a result, intensities of MITF-DNA binding bands were dose-dependently reduced by compound #7. In addition, to investigate the MITF and tyrosinase protein level, western blot was performed by using B16F10 melanoma cell extract. As a result, tyrosinase protein was down-regulated by compound #7 in a dose dependent manner but the MITF protein not was reduced. Thus, compound #7 which was isolated from a natural plant can be used as an inhibitor of MITF-DNA binding for depigmenting agents.

Recently all the nations are competitively focusing on developing biorefinery technologies. Biorefinery is defined as a bioprocess to produce high-valued industrial chemicals and energy using natural biomasses which are present enormously in nature. Cassava is a good alternative biomass source which could be used to produce various biorefinery products. The roots of cassava are mainly composed of 20∼25% starches and contain plenty of valuable nutrients such as vitamin C and calcium. Recently, it has been reported that a newly-developed super cassavas is being tested to be used as a carbon and energy source in biorefinery industries. Our research team intended to develop an economic bioprocess for mass production of exopolysaccharides(EPS) using cassava as a carbon source in liquid mycelial fermentations of Schizophyllum commune [1]. This product has the potent capability of maintaining high moisturized conditions on epidermal tissue, and also having positive effects on the restoration of epidermal tissue. First of all, medium optimization process was carried out in order to find an optimal composition of the production medium containing the cassava biomass. Approximately 2~3 fold increase in EPS productivity was observed in the shake flask cultures performed with the optimized production medium [2]. For the purpose of enhancing the EPS production, intensive strain improvement program was also carried out by obtaining large amounts of protoplasts for the isolation of single colonies. Mass screening for high-yielding producers was possible through the formation of the protoplasts of S. commune because high concentration of protoplasts(5x106 protoplasts/ml) could be recovered with efficient cell wall disrupting enzymes. Notably, the protoplast regeneration rate was as high as 98% when RPA medium containing 1.2M sorbitol was used as a regeneration medium.

Fingernails and toenails are made of a tough protein called keratin and have many different parts. Nails can dry out, just like skin. They can also peel, break and be infected. Toe infections, for instance, can be caused or exacerbated by dirty socks, specific types of aggressive exercise, tight footwear, and walking unprotected in an unclean environment. Artificial nails, also known as fake nails, fashion nails, nail enhancements, or nail extensions, are coverings placed over fingernails as fashion accessories. Some artificial nails attempt to mimic the look of real fingernails as closely as possible, while other designs may deliberately stray in look from real fingernails. Artificial nails are made from a wide variety of materials. The purpose of this paper is to analyze the effect of polyglutamic acid (PGA) on the recovery of defect of injured nails caused by artificial nail treatments. PGA is a polymer of the amino acid glutamic acid (GA). Gamma PGA (the form where the peptide bonds are between the amino group of GA and the carboxyl group at the end of the GA side chain) is a major constituent of the Japanese food natto. Gamma PGA is formed by bacterial fermentation. The scope of this dissertation included the electron microscopy analysis of nail plates and cross-sections before and after the treatments. In this study, we have shown that an artificial nail treatment produces very harmful effects although it was just once. PGA of three different molecular weights showed the positive effect on nail condition all together. Especially PGA with molecular weight of 5,000 Da (BLS 5) represents the best recovery effect. On the other hand, the control untreated group did not show the recovery appearance. For the term of use of PGA, 4 weeks was better than 2 weeks. The initial recovery rate (2 wk) is much faster than the latter one (4 wk). PGA effects were noticeable more on nail section than on nail plate according the SEM data. As a summary, PGA could be used as an ingredient in nail guard and mush work should be done before preparing the commercial product.

Microphthalmia-associated transcription factor (MITF), a basic Helix-Loop Helix (HLH) transcription factor with a leucine zipper, is a key regulatory protein for the pigmenting process in melanocytes. Binding of MITF to E-box caused transcription of several pigmenting genes including the tyrosinase gene. The purpose of this study was to analyze the interaction of MITF and its DNA binding site (M-box) by using surface plasmon resonance (SPR) imaging. We developed DNAprotein interaction arrays, coated with polypeptide poly(L-lysine) with a bifunctional coupling property that permits efficient immobilization of 1,4-phenylenediisothiocyanate on gold surfaces. DNA arrays on modified gold surfaces were fabricated, and then used for the analysis of DNA-MITF interaction. It is expected that SPR imaging can be used as a powerful tool to study DNA-protein interactions.

We reported the isolation and identification of a new compound, 1-(2-hydroxy-3,4,5,6-tetramethoxyphenyl)-1-methoxy-3-phenylpropane; (Jeju-Erythrane) from the bark of Lindera erythrocarpa. Also, we investigated the effects of Jeju-Erythrane on alpha melanocytestimulating hormone (MSH)-induced melanogenesis in mouse B16F10 melanoma cells. The new compound inhibited dose dependently the tyrosinase activity and melanin content in B16F10 cells. The new compound showed inhibitory effect on Tyrosinase and TRP-1 gene transcription but not inhibited TRP-2 gene transcription. These results suggest that new compound of L. erythrocarpa could be used as functional biomaterial in developing a skin whitening agent.

Vitiligo is one of the most psychologically devastating skin diseases. characterized by areas of depigmented skin resulting from loss of epidermal melanocytes. The prevalence of this disease varies from 0.1% to 2% in various global populations. It is known that the existance of inactive melanoblasts in hair root follicles provide a melanocyte source for repigmentation in Vitiligo. Differentiation and migration of these melanocytes precursors from the outer root sheath of hair follicles into clinically depigmented epidermis is crucial for the repigmentation in Vitiligo therapy. An effective treatment regime has been proposed that would stimulate the inactive melanoblasts in the hair follicle to divide, proliferate and migrate upward along the surface of outer root sheath to the nearby epidermis. Melanoblasts are defined as unpigmented but committed precursors of melanocytes. The characteristic phenotype of melanoblasts includes a lack of expression of tyrosinase and no DOPA staining or melanin production, Thus, those cells provide an ideal model for studing the effect of natural herbs on differentiation of unpigmented melanoblasts to pigmented melanocytes, and to characterize the mechanisms involved.To evaluate the potential of herbal candidates in treatment of Vitiligo, we studied its effects on the differentiation, proliferation and migration of melanoblast cells. Crude herbal extract was added to the complete RPMI medium, and its effect on differentiation, proliferation and migration of melanoblasts were investigated. Selected herbal candidates induced 67% to 70% melanin induction and differentiation. Transwell migration assay showed the potential of these candidates to induce direct migration of treated cells. The findings of present study are significant in the direction of developing safer strategies for Vitiligo treatment.

The proapoptotic tumor necrosis factor–related apoptosis inducing ligand (TRAIL) receptors death receptor (DR) 4 and DR5 are attractive targets to develop the receptorspecificagonistic monoclonal antibodies (mAb) as anticancer agents because of their tumor-selective cell death–inducing activity. Here, we report a novel agonistic mAb, AY4, raised against human DR4 in mice. ELISA analysis revealed that AY4 specifically bound to DR4 withoutcompetition with TRAIL for the binding. Despite distinct binding regions of AY4 on DR4 from those of TRAIL, AY4 as a single agent induced caspase-dependent apoptotic cell death of several tumor types through the extrinsic and/or intrinsic pathways without substantial cytotoxicity to normal human hepatocytes. Further, the AY4-sensitive cells followed the same cell death characteristics classified as type I and type II cells by the response to TRAIL,suggesting that the cell death profiles in responses to DR4 and/or DR5 stimulation are determined by the downstream signaling of the receptor rather than the kind of receptor. Noticeably, AY4 efficiently induced cell death of Jurkat cells, which have been reported to be resistant to other anti-DR4 agonistic mAbs, most likely due to the unique epitope property of AY4. In vivo administration of AY4 significantly inhibited tumor growth of human non–small cell lung carcinoma preestablished in athymic nude mice. Conclusively, our results provide further insight into the DR4-mediated cell death signaling and potential use of AY4 mAb as an anticancer therapeutic agent, particularly for DR4-responsive tumor types.

In this study, biodegradable poly(L-lactic acid) (L-PLA) microparticles containing gemcitabine (2,2-difluorodeoxycytidine, dFdC) were prepared by a supercritical process, called aerosol solvent extraction system (ASES). ASES is a method based on solvent extraction using supercritical carbon dioxide for the preparation of microparticles and microspheres. Gemcitabine is an effective antitumor agent in the treatment of several solid tumors. However, it has problems such as short plasma half-life, rapid metabolism, and low selectivity towards tumor tissue. The influence of operating parameters such as temperature, pressure, drug/polymer feed ratio, and polymer molecular weight on the morphology and release characteristics was studied in detail. The drug recovery, entrapment efficiency, and drug release profiles were determined by HPLC assays. The drug recovery was found to be about 70~80%. The entrapment efficiency decreased with increasing temperature and drug/polymer ratio. The ASES-processed gemcitabine-loaded microparticles showed a relatively high initial burst due to the poor affinity between gemcitabine and L-PLA.

Influenza virus hemagglutinin (HA) is expressed as a precursor protein, HA0, and then proteolytically cleaved to produce two disulfide-linked polypeptide chains, HA1 and HA2. The HA2 protein segment, an influenza virus glycoprotein, contains two membrane-interacting hydrophobic peptide sequences: an N-terminal "fusion peptide" (residues 1-22), which interacts with the target membrane, and a C-terminal transmembrane domain (residues 186-221) that passes through the viral membrane. This study aims to develop biodegradable microparticles containing both HA1 and truncated HA2 proteins for vaccination with enhanced biosafety. The HA2 ectodomain (residues 23-185) truncated with two membraneinteracting sequences was expressed in Escherichia coli. This segment of HA2 contains changes in three rare Arg codons (Arg-123, Arg-124, and Arg-127) and the mutation of Cys-137, which forms a disulfide bond with HA1 residue 14, to Ser. The full-length HA1 polypeptide segment was expressed separately in Escherichia coli with mutation of Cys-14, which forms a disulfide bond with HA2 residue 137, to Ser. Both purified HA1 and truncated HA2 segments were microencapsulated in poly(D,L-lacticco-glycolic acid) (PLGA) using a W/O/W double emulsion technique. Antigen release from microspheres was determined in vitro using a hemagglutinin-specific ELISA.