Sunhyae Jang

Silver nanoparticles modify VEGF signaling pathway and mucus hypersecretion in allergic airway inflammation

The anti-inflammatory action of silver nanoparticles (NPs) has been reported in a murine model of asthma in a previous study. But more specific mechanisms of silver NPs in an attenuation of allergic airway inflammation have not yet been established. Vascular and mucous changes are believed to contribute largely in pathophysiology in asthma. Among various factors related to vascular changes, vascular endothelial growth factor (VEGF) plays a pivotal role in vascular changes in asthma. Mucin proteins MUC5AC and MUC5B have been implicated as markers of goblet cell metaplasia in lung pathologies. The aim of this study was to investigate the effects of silver NPs on VEGF signaling pathways and mucus hypersecretion. Ovalbumin (OVA)-inhaled female BALBc mice were used to evaluate the role of silver NPs and the related molecular mechanisms in allergic airway disease. In this study, with an OVA-induced murine model of allergic airway disease, it was found that the increased levels of hypoxia-inducible factor (HIF)-1α, VEGF, phosphatidylinositol-3 kinase (PI3K) and phosphorylated-Akt levels, and mucous glycoprotein expression (Muc5ac) in lung tissues were substantially decreased by the administration of silver NPs. In summary, silver NPs substantially suppressed mucus hypersecretion and PI3K/HIF-1α/VEGF signaling pathway in an allergic airway inflammation.

Pitx2, a β-catenin-regulated transcription factor, regulates the differentiation of outer root sheath cells cultured in vitro

BACKGROUND Beta-catenin exerts its crucial role in hair follicle development and hair growth cycle. Although the importance of Wnt/beta-catenin is well recognized, the downstream effectors of beta-catenin have not been clearly elucidated yet. OBJECTIVE The aim of this study is to identify the beta-catenin-regulated genes in cultured human hair outer root sheath (ORS) cells. METHODS We transduced ORS cells with adenovirus harboring the expression cassette for constitutive active form of beta-catenin, then performed cDNA microarray. RESULTS Overexpression of beta-catenin led to the upregulation of hair cell differentiation markers such as keratin 16 and 17. In addition, the expression of Pitx2, a bicoid-type homeodomain transcription factor, was also increased by overexpression of beta-catenin in ORS cells cultured in vitro. To investigate the potential role of Pitx2, we made the recombinant adenovirus expressing Pitx2, then transduced into the cultured ORS cells. Interestingly, Pitx2 induced the expression of keratin 16 and 17, indicating that Pitx2 activates ORS cells towards the follicular differentiation pathway preferentially. CONCLUSION Our results implicate the potential importance of Pitx2 as a beta-catenin downstream modulator in hair growth control.

Attenuation of allergic airway inflammation and hyperresponsiveness in a murine model of asthma by silver nanoparticles

The use of silver in the past demonstrated the certain antimicrobial activity, though this has been replaced by other treatments. However, nanotechnology has provided a way of producing pure silver nanoparticles, and it shows cytoprotective activities and possible pro-healing properties. But, the mechanism of silver nanoparticles remains unknown. This study was aimed to investigate the effects of silver nanoparticles on bronchial inflammation and hyperresponsiveness. We used ovalbumin (OVA)-inhaled female C57BL/6 mice to evaluate the roles of silver nanoparticles and the related molecular mechanisms in allergic airway disease. In this study with an OVA-induced murine model of allergic airway disease, we found that the increased inflammatory cells, airway hyperresponsiveness, increased levels of IL-4, IL-5, and IL-13, and the increased NF-κB levels in lungs after OVA inhalation were significantly reduced by the administration of silver nanoparticles. In addition, we have also found that the increased intracellular reactive oxygen species (ROS) levels in bronchoalveolar lavage fluid after OVA inhalation were decreased by the administration of silver nanoparticles. These results indicate that silver nanoparticles may attenuate antigen-induced airway inflammation and hyperresponsiveness. And antioxidant effect of silver nanoparticles could be one of the molecular bases in the murine model of asthma. These findings may provide a potential molecular mechanism of silver nanoparticles in preventing or treating asthma.

S100A8 and S100A9 are messengers in the crosstalk between epidermis and dermis modulating a psoriatic milieu in human skin

S100A8 and S100A9 are members of the S100A8 protein family that exist as homodimers and heterodimers in neutrophils, monocytes, and macrophages. Recent studies have shown the pivotal roles of S100A8 and S100A9 in the propagation of inflammation and keratinocyte proliferation in psoriasis. We found significant up-regulation of S100A8 and S100A9 secretion from keratinocytes in psoriatic lesions. To mimic the in vivo secretory conditions of S100A8 and S100A9 from psoriatic epidermal keratinocytes, we used the culture medium (CM) of S100A8 and S100A8/A9 adenovirus-transduced keratinocytes to investigate the functions of S100A8 and S100A9. We detected increased levels of various pro-inflammatory cytokines in the CM, including IL-8 and TNF-α, which are involved in aggravating psoriatic skin lesions, and IL-6 and members of the CXCL family of pro-angiogenic cytokines. The CM increased immune cell migration and increased angiogenesis in human umbilical vein endothelial cells. In conclusion, we found that the upregulated production of S100A8 and S100A9 by psoriatic epidermal keratinocytes activated adjacent keratinocytes to produce several cytokines. Moreover, S100A8 and S100A9 themselves function as pro-angiogenic and chemotactic factors, generating a psoriatic milieu in skin.

Establishment of type II 5α-reductase over-expressing cell line as an inhibitor screening model

Dihydrotestosterone (DHT) is the most potent male hormone that causes androgenetic alopecia. The type II 5alpha-reductase is an enzyme that catalyzes the conversion of testosterone (T) to DHT, therefore it can be expected that specific inhibitors for type II 5alpha-reductase may improve the pathophysiologic status of androgenetic alopecia. In this study, we attempted to establish the reliable and convenient screening model for type II 5alpha-reductase inhibitors. After transfection of human cDNA for type II 5alpha-reductase into HEK293 cells, the type II 5alpha-reductase over-expressing stable cells were selected by G418 treatment. RT-PCR and Western blot analyses confirmed that type II 5alpha-reductase gene was expressed in the stable cells. In in vitro enzymatic assay, 10 microg of stable cell extract completely converted 1 microCi (approximately 0.015 nmol) of T into DHT. The type II 5alpha-reductase activity was inhibited by finasteride in a dose-dependent manner, confirming the reliability of screening system. In cell culture condition, 2 x 10(5) of stable cells completely converted all the input T (approximately 0.03 nmol) into DHT by 4h incubation, demonstrating that the stable cell line can be used as a cell-based assay system. Using this system, we selected the extracts of Curcumae longae rhizoma and Mori ramulus as the potential inhibitors for type II 5alpha-reductase. These results demonstrate that the type II 5alpha-reductase over-expressing stable cell line is a convenient and reliable model for screening and evaluation of inhibitors.

The Role of Nkx2.5 in Keratinocyte Differentiation

BACKGROUND Nkx2.5 is a homeodomain-containing nuclear transcription protein that has been associated with acute T-lymphoblastic leukemia. In addition, Nkx2.5 has an essential role in cardiomyogenesis. However, the expression of Nkx2.5 in the skin has not been investigated. OBJECTIVE In an attempt to screen the differentially regulated genes involved in keratinocyte differentiation, using a cDNA microarray, we identified Nkx2.5 as one of the transcription factors controlling the expression of proteins associated with keratinocyte differentiation. METHODS To investigate the expression of Nkx2.5 during keratinocyte differentiation, we used a calcium-induced keratinocyte differentiation model. RESULTS RT-PCR and Western blot analysis revealed that the expression of Nkx2.5, in cultured human epidermal keratinocytes, increased with calcium treatment in a time-dependent manner. In normal skin tissue, the expression of Nkx2.5 was detected in the nuclei of the keratinocytes in all layers of the epidermis except the basal layer by immunohistochemistry. In addition, the expression of Nkx2.5 was significantly increased in psoriasis and squamous cell carcinoma, but was barely detected in atopic dermatitis and basal cell carcinoma. CONCLUSION These results suggest that Nkx2.5 may play a role in the change from proliferation to differentiation of keratinocytes and in the pathogenesis of skin disease with aberrant keratinocyte differentiation.

Abstract A18: Silver nanoparticles induce apoptosis by regulation of cyclic AMP response element-binding protein in lung cancer cells

Recently, much effort has been devoted to the development of biomedical applications for nanoparticles (NPs). Silver NPs have been shown various therapeutic effects such as antimicrobial, antioxidant, and anti-inflammatory effects. Recently silver NPs have been shown to induce the apoptotic pathway in vitro. But exact mechanism is not yet established. We conducted this study to determine the mechanisms of silver NPs on cellular apoptosis in non-small cell lung cancer (NSCLC) cell lines using cyclic AMP response element-binding protein (CREB) activity which has been implicated to contribute an important pathobiologic role in lung carcinogenesis and is considered a potential therapeutic target for NSCLC. We hypothesized that constitutively increased CREB and its related signaling pathways can be blocked by silver NPs. To determine cytotoxic effect of silver NPs, several NSCLC cell lines were treated with various concentrations for different times. Among these lung cancer cell lines, H520, human lung squamous carcinoma cell lines, was the most sensitive. FACS analysis also showed that silver NPs induced cell death of H520 cells. Treating H520 cells with silver NPs (100, 200, or 500 M) inhibited CREB activation by blocking the activity of extracellular signal kinase/ribosomal s6 kinase and also by blocking the activity of phosphatidylinositol 3-kinase/Akt. We subsequently confirmed the expression of Bax and Bcl-2 in H520 cells was substantially regulated by silver NPs. We also demonstrated that silver NPs induced the cleavage of apoptosis-related proteins, poly (ADP-ribose) polymerase and caspase-3. In this study, we suggest that silver NPs induce apoptosis in H520 cells by regulation of CREB signaling pathways and may be useful as a therapeutic strategy for cancer. Citation Information: Cancer Prev Res 2010;3(12 Suppl):A18.

Abstract 4785: Silver nanoparticles induce apoptosis in squamous lung cancer cell by regulation of cyclic AMP response element-binding protein: regulating its upstream oxidative stress-dependent Ras signaling pathways

Recently, much effort has been devoted to the development of biomedical applications (drug delivery system and as therapeutic drugs themselves) for nanoparticles (NPs). Silver NPs have been shown various therapeutic effects such as antimicrobial, antioxidant, and anti-inflammatory effects. Recently silver NPs have been shown to induce cytotoxicity in cancer cells. But exact mechanism is not yet established. We conducted this study to determine the mechanisms of silver NPs on cellular apoptosis in non-small cell lung cancer (NSCLC) cell lines using cyclic AMP response element-binding protein (CREB) activity which has been implicated to contribute an important pathobiologic role in lung carcinogenesis and is considered a potential therapeutic target for NSCLC. We hypothesized that constitutively increased CREB and its related signaling pathways can be blocked by silver NPs and it might be by regulation of oxidative stress in tumorigenesis. In H520 cell, human lung squamous cancer cell line, reactive oxygen species (ROS) were increased. In western blot analysis, Ras signaling molecule in which ROS is involved was constitutively overexpressed and the levels of pErk, pRsk, pAkt and pCREB were increased in H520 cell. Incubation of the H520 cells with silver NPs (at dose of 100 or 200 μM) substantially suppressed the ROS in H520 cell and Ras-Erk-Rsk-CREB and Ras-PI3K-Akt signaling pathway in a dose-dependent manner. We subsequently demonstrated that silver NPs inhibited the caspase-dependent pathway, determined by increased levels of PARP (poly (ADP-ribose) polymerase) cleavage. In this study, we suggest that silver NPs induce apoptosis in human lung cancer cells by attenuation of oxidative stress in lung cancer cell line and ROS-dependent Ras signaling which involves CREB signaling pathways and may be useful as a therapeutic strategy for cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4785. doi:10.1158/1538-7445.AM2011-4785