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In this study, the protective functions of bacteria-free conditioned media from Bifidobacteriumand Lactobacillusspecies against ultraviolet radiation-induced skin ageing and associated cellular damage were investigated. The effects of ultraviolet radiation-induced reactive oxygen species production were suppressed by all conditioned media; particularly, the loss of cell viability and downregulation of collagen gene expression were significantly reversed by the conditioned media from B. longumand B. lactis. Further exa mination of potential anti-pigmentation effects revealed that the B. lactis-derived conditioned media significantly inhibited tyrosinase activity and alpha-melanocyte-stimulating hormone-induced melanin production in human epidermal melanocytes. Further, the conditioned media suppressed the phosphorylation of extracellular signal- related kinase, which functions as an upstream regulator of melanogenesis. Therefore, B. lactis-derived conditioned media can potentially protect against cellular damage involved in skin-ageing processes.

We present a mold-free high-resolution nanopatterning technology named piezo-actuated one-axis vibrational patterning (POP) that enables continuous and scalable fabrication of micro- and nanopatterns with precisely programmable periods and dimensions. POP utilizes the piezoelectric stack-actuated high-precision uniaxial vibration of a flat, pattern-free rigid tool edge to conduct sub-50 nm-periodic indentations on various compliant substrates laterally fed underneath. By controlling the tool vibration frequency, tool temperature, and substrate feed rate and by combining sequential tool strokes along multiple directions, diverse functional micro- and nanopatterns with variable periods and depths and multidimensional profiles can be continuously created without resorting to mold prefabrication. With its simple but universal principle, excellent scalability, and versatile processability, POP can be practically applied to many functional devices particularly requiring large-area micro- and nanopatterns with specifically designed periods and dimensions.

We have rationally designed and developed a fully automated desktop furnace system that enables programmable chemical vapour deposition growth of carbon nanotubes with controlled height, density, and pattern architecture. Comprising several essential components involving a heating furnace, mass flowmeters, and computer controller, the developed system realizes controlled and practical carbon nanotube growth without resort to expensive and ponderous instruments. By programming, modifying, and loading the reusable recipes in the developed system, systematic and reproducible growths of carbon nanotubes with desired morphology and dimension can be performed. Growth results with controlled height, density, and pattern are demonstrated through the actual operations, confirming the validity and usefulness of the developed system towards various practical applications.

In this research, an electrochemical PSA (prostate specific antigen) immunosensor based on Nafion/reduced graphene oxide (rGO)/aldehyde methyl pyridine (CHO-MP) composite electrode was newly developed by using noncovalent functionalization of rGO and CHO-MP. Due to improper binding stability of the immobilized antibody, immunosensors may show false positive results. The proposed composite material can provide suitable microenvironment for the immobilized anti-PSA and improve the conductivity of electrode. Furthermore, the used nafion can improve the mechanical and chemical behaviors of the developed immunosensor. Physical characterization of rGO/CHO-MP was observed through XPS, FTIR, and FE-SEM. Under the optimized conditions, the fabricated immunosensor showed a linear range of 0.005- 90 ng mL[?]1, detection limit of 1.6 pg mL[?]1, excellent selectivity, stability, and reproducibility. Lastly, human serum samples were analyzed and the recovery range was obtained as 95.8% to 107.9% with an RSD of 5.1% (n = 5). These results showed that the as-fabricated sensor is highly promising in clinical applications.

Dermal papilla (DP) is a pivotal part of hair follicle, and the smaller size of the DP is related with the hair loss. In this study, we investigated the effect of titrated extract of Centella asiatica(TECA) on hair growth inductive property on 3D spheroid cultured human DP cells (HDP cells). Significantly increased effect of TECA on cell viability was only shown in 3D sphered HPD cells, not in 2D cultured HDP cells. Also, TECA treatment increased the sphere size of HDP cells. The luciferase activity of STAT reporter genes and the expression of STAT-targeted genes, SOCS1 and SOCS3, were significantly decreased. Also, TECA treatment increased the expression of the hair growth-related signature genes in 3D sphered HDP cells. Furthermore, TECA led to downregulation of the level of phosphorylated STAT proteins in 3D sphered HDP cells. Overall, TECA activates the potential of hair inductive capacity in HDP cells.

Ultraviolet (UV) radiation is a major inducer of skin aging and accumulated exposure to UV radiation increases DNA damage in skin cells, including dermal fibroblasts. In the present study, we developed a novel DNA repair regulating material discovery (DREAM) system for the high-throughput screening and identification of putative materials regulating DNA repair in skin cells. First, we established a modified lentivirus expressing the luciferase and hypoxanthine phosphoribosyl transferase (HPRT) genes. Then, human dermal fibroblast WS-1 cells were infected with the modified lentivirus and selected with puromycin to establish cells that stably expressed luciferase and HPRT (DREAM-F cells). The first step in the DREAM protocol was a 96-well-based screening procedure, involving the analysis of cell viability and luciferase activity after pretreatment of DREAM-F cells with reagents of interest and post-treatment with UVB radiation, and vice versa. In the second step, we validated certain effective reagents identified in the first step by analyzing the cell cycle, evaluating cell death, and performing HPRT-DNA sequencing in DREAM-F cells treated with these reagents and UVB. This DREAM system is scalable and forms a time-saving high-throughput screening system for identifying novel anti-photoaging reagents regulating DNA damage in dermal fibroblasts.

Accumulating evidence indicates that reactive oxygen species (ROS) are an important etiological factor for the induction of dermal papilla cell senescence and hair loss, which is also known alopecia. Arctiin is an active lignin isolated from Arctium lappaand has anti-inflammation, anti-microbial, and anti-carcinogenic effects. In the present study, we found that arctiin exerts anti-oxidative effects on human hair dermal papilla cells (HHDPCs). To better understand the mechanism, we analyzed the level of hydrogen peroxide (H2O2)-induced cytotoxicity, cell death, ROS production and senescence after arctiin pretreatment of HHDPCs. The results showed that arctiin pretreatment significantly inhibited the H2O2-induced reduction in cell viability. Moreover, H2O2-induced sub-G1 phase accumulation and G2 cell cycle arrest were also downregulated by arctiin pretreatment. Interestingly, the increase in intracellular ROS mediated by H2O2was drastically decreased in HHDPCs cultured in the presence of arctiin. This effect was confirmed by senescence associated-beta galactosidase (SA-β-gal) assay results; we found that arctiin pretreatment impaired H2O2-induced senescence in HHDPCs. Using microRNA (miRNA) microarray and bioinformatic analysis, we showed that this anti-oxidative effect of arctiin in HHDPCs was related with mitogen-activated protein kinase (MAPK) and Wnt signaling pathways. Taken together, our data suggest that arctiin has a protective effect on ROS-induced cell dysfunction in HHDPCs and may therefore be useful for alopecia prevention and treatment strategies.

Histone deacetylase (HDAC) inhibitors represent a promising group of anticancer agents. Treatment of cancer cells with HDAC blockers, such as suberoylanilide hydroxamic acid (SAHA), leads to the activation of apoptosis-promoting genes. To enhance proapoptotic efficiency, SAHA has been used in conjunction with radiation, kinase inhibitors, and cytotoxic drugs. In the present study, we show that at the suboptimal dose of 250 µM, sulindac [2-[6-fluoro-2-methyl-3-[(4-methylsulfinylphenyl)methylidene]inden-1-yl]-acetic acid] significantly enhances SAHA-induced growth suppression and apoptosis of A549 human non-small cell lung cancer cells, primarily via enhanced collapse of the mitochondrial membrane potential, release of cytochrome c, and caspase activation. Furthermore, sulindac/SAHA cotreatment induced marked down-regulation of survivin at both the mRNA and protein levels and stimulated the production of reactive oxygen species (ROS), which were blocked by the antioxidant N-acetyl-L-cysteine. Overexpression of survivin was associated with reduced sulindac/SAHA-induced apoptosis of A549 cells, whereas suppression of survivin levels with antisense oligonucleotides or small interfering RNA further sensitized cells to sulindac/SAHA-induced cell death. Our results collectively demonstrate that sulindac/SAHA-induced apoptosis is mediated by ROS-dependent down-regulation of survivin in lung cancer cells.

Abstract: YBa2Cu3Ox (Y123) superconducting films were fabricated on Cu substrates using a simple screen-printing method, from Cu-free powders (Y2O3 and BaCO3). In the process, CuO, which causes superconducting properties of Y123 films to deteriorate, was formed on the film surface. By varying the atomic ratio of Y to Ba (Y:Ba = 1:1∼1:4), the ratio needed to prevent CuO formation was found for the film surface that had been heat-treated at 980∘C for 17 s. The film, with the ratio of Y to Ba (Y:Ba = 1:1), is reheat-treated at 930∘C for 9 min 30 s to form a superconducting Y123 phase. It was possible to prevent CuO formation by controlling the ratio of Cu-free powders in the mixture and to fabricate YBCO superconductors on Cu substrates using a two-step heat-treatment.

Abstract: Although the sera used in animal cell culture media provide the macromolecules, nutrients, hormones, and growth factors necessary to support cell growth, it could also be an obstacle to the production of recombinant proteins in animal cell culture systems used in many sectors of the biotechnology industry. For this reason, many research groups, including our laboratory, have been trying to develop serum-free media (SFM) or serum-supplemented media (SSM) for special or multi-purpose cell lines. The Chinese hamster ovary (CHO) cell, for example, is frequently used to produce proteins and is especially valuable in the large-scale production of pharmaceutically important proteins, yet information about its genome is lacking. Also, SFMs have only been evaluated by comparing growth patterns for cells grown in SFMs with those grown in SSM or by measuring the titer of the target protein obtained from cells grown in each type of medium. These are not reliable methods of obtaining the type of information needed to determine whether an SFM should be replaced with an SSM. We carried out a cDNA microarray analysis to evaluate MED-3, an SFM developed in our laboratory, as a CHO culture medium. When CHO cells were cultured in MED-3 instead of an SSM, several genes associated with cell growth were down-regulated, although this change diminished over time. We found that the insulin-like growth factor (IGF) gene was representative of the proteins that were down-regulated in cells cultured in MED-3. When several key supplements-including insulin, transferrin, ethanolamine, and selenium-were removed from MED-3, theIGF expression was consistently down-regulated and cell growth decreased proportionately. Based on these results, we concluded that when an SFM is used as a culture medium, it is important to supplement it with substances that can help the cells maintain a high level ofIGF expression. The data presented in this study, therefore, might provide useful information for the design and development of SFM or SSM, as well as for the design of genome-based studies of CHO cells to determine how they can be used optimally for protein production in pharmaceutical and biomedical research.

The proliferation and survival of a B cell population is necessarily tightly controlled to prevent the arisal of malignancy or autoimmunity. The expansion or elimination of a B cell clone is determined through a complex interaction of the tumour necrosis factor receptor/nerve growth factor receptor family members CD40 and Fas, which are expressed on the B cell surface, with their respective physiological ligands (CD40L and FasL) expressed on the surface of CD4+T cells. The regulation of B cell growth by signals transduced through CD40 and Fas contributes to the maintenance of peripheral tolerance and likely takes place and in the germinal centres (GC) of secondary lymphoid tissues. In this study, we investigate the relationship between the expression of Fas and B cell survival following engagement of CD40 and Fas in the Epstein–Barr virus-genome-negative Ramos–Burkitt lymphoma (Ramos-BL) B cell line model of GC B lymphocyte selection during maturation of the humoral immune response. We now present evidence that Ramos-BL B cells constitutively express both Fas and FasL on their surface and that expression of Fas, but not FasL, is enhanced following ligation of CD40. Coligation of CD40 and Fas, triggers for growth inhibition, activation of the interleukin-1β-converting enzyme, now caspase, family member CPP32 (caspase-3) but not Ich-1L(caspase-2), cleavage of its death substrate poly(ADP-ribose) polymerase, and apoptosis from the G1phase of cell cycle; engagement of Fas alone fails to trigger for growth inhibition and apoptosis but enhances AgR-mediated cellular death. This CD40-potentiated Fas-triggered growth inhibition and apoptosis occurs in the presence of CD40-induced expression of the anti-apoptotic proteins Bcl-xLand Bcl-2. Taken together, these data indicate that ligation of CD40 facilitates efficient coupling of Fas to the caspase-mediated pathway of apoptosis.

Signals transduced through CD40 rescue cells of the Ramos–Burkitt lymphoma (Ramos-BL) B cell line from surface immunoglobulin M (sIgM)-triggered growth arrest and apoptosis. This study investigates whether protein tyrosine kinase (PTK) activity and tyrosine phosphorylation on p95vavand on the p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3 kinase) play a role in the regulation of Ramos-BL B cell survival. The PTK inhibitor herbimycin A (HA) triggers significant growth arrest prior to apoptosis from the G1-phase of the cell cycle, indicating that tyrosine phosphorylation of key proteins is critical for Ramos-BL cell cycle progression and survival. Indeed, signals transduced through CD40 fail to rescue Ramos-BL B cells from HA-triggered growth arrest and apoptosis. Since Vav and PI3 kinase are intimately involved in the regulation of cellular growth, their tyrosine phosphorylation status was determined in unstimulated and anti-IgM- and anti-CD40-treated Ramos-BL B cells: Vav and p85 are devoid of tyrosine-phosphorylated epitopes in control cells whereas p85, but not Vav, is significantly phosphorylated following ligation of sIgM and anti-CD40 triggers tyrosine phosphorylation on both proteins. Thus, tyrosine-phosphorylated Vav may be a critical effector of CD40-mediated survival. As tyrosine-phosphorylated PI3 kinase is common to both sIgM-triggered death and CD40-triggered survival pathways, its lipid kinase activity was correlated with tyrosine phosphorylation on p85: Ramos-BL B cells exhibit high basal levels of PI3 kinase activity, determined by immunoprecipitation with anti-p85 and32P incorporation into phosphatidylinositol, which is not significantly affected by stimulation with anti-IgM but which is elevated by 36 ± 2.9% following ligation of CD40. Thus, tyrosine phosphorylation on p85 correlates with the CD40-triggered increase in PI3 kinase activity but not with basal levels nor with sIgM-triggered levels of enzymatic activity: these data suggest the presence of different PI3 kinase isoforms or the existence of multiple regulatory pathways for the same PI3 kinase isotype in Ramos-BL B cells.

The new and growing family of interleukin‐1β‐converting enzyme (ICE) cysteine proteases are now recognised to be major effectors of cellular death by apoptosis. Like other members of this family, the CPP32/Yama proform is activated by processing to its active heterodimeric enzyme or apopain when it likely contributes to the process of apoptosis by cleaving poly(ADP‐ribose) polymerase (PARP) and thereby inhibiting much of its DNA repair activity. Apoptosis plays a fundamental role in the regulation of the immune system where it is involved in the selection of both T and B lymphocytes bearing antigen receptor (AgR) for non‐self. Cells of the Ramos Epstein‐Barr virus (EBV)‐genome‐negative Burkitt lymphoma (BL) B cell line (Ramos‐BL) can be triggered into growth arrest and apoptosis by treating with the calcium ionophore ionomycin or by crosslinking their surface AgR with antibodies directed against immunoglobulin (Ig)M (anti‐IgM). Ionomycin‐ and AgR‐triggered growth arrest and apoptosis are arrested by signals transduced through the surface CD40 of Ramos‐BL B cells. Both ionomycin and anti‐IgM trigger activation of CPP32 and cleavage of PARP prior to the onset of apoptosis; this process is abrogated by treatment with anti‐CD40 and is independent of Bcl‐2 expression. A tripeptide inhibitor of ICE family cysteine proteases, Z‐Val‐Ala‐Asp‐fluoromethylketone (zVAD‐fmk) inhibits ionomycin‐ and AgR‐triggered CPP32 activation, PARP cleavage and apoptosis, but not growth arrest, in Ramos‐BL B cells. Thus, in this report we demonstrate that in a physiological system, activation of endogenous members of the ICE family, including CPP32, and cleavage of the death substrate PARP act as major effectors of apoptotic death.

In this research, an electrochemical PSA (prostate specific antigen) immunosensor based on Nafion/reduced graphene oxide (rGO)/aldehyde methyl pyridine (CHO-MP) composite electrode was newly developed by using noncovalent functionalization of rGO and CHO-MP. Due to improper binding stability of the immobilized antibody, immunosensors may show false positive results. The proposed composite material can provide suitable microenvironment for the immobilized anti-PSA and improve the conductivity of electrode. Furthermore, the used nafion can improve the mechanical and chemical behaviors of the developed immunosensor. Physical characterization of rGO/CHO-MP was observed through XPS, FTIR, and FE-SEM. Under the optimized conditions, the fabricated immunosensor showed a linear range of 0.005- 90 ng mL−1, detection limit of 1.6 pg mL−1, excellent selectivity, stability, and reproducibility. Lastly, human serum samples were analyzed and the recovery range was obtained as 95.8% to 107.9% with an RSD of 5.1% (n = 5). These results showed that the as-fabricated sensor is highly promising in clinical applications.