Regulation of reactive oxygen species by phytochemicals for the management of cancer and diabetes.

Cancer and diabetes mellitus are served as typical life-threatening diseases with common risk factors. Developing therapeutic measures in cancers and diabetes have aroused attention for a long time. However, the problems with conventional treatments are in challenge, including side effects, economic burdens, and patient compliance. It is essential to secure safe and efficient therapeutic methods to overcome these issues. As an alternative method, antioxidant and pro-oxidant properties of phytochemicals from edible plants have come to the fore. Phytochemicals are naturally occurring compounds, considered promising agent applicable in treatment of various diseases with beneficial effects. Either antioxidative or pro-oxidative activity of various phytochemicals were found to contribute to regulation of cell proliferation, differentiation, cell cycle arrest, and apoptosis, which can exert preventive and therapeutic effects against cancer and diabetes. In this article, the antioxidant or pro-oxidant effects and underlying mechanisms of flavonoids, alkaloids, and saponins in cancer or diabetic models demonstrated by the recent studies are summarized.

Anti-cancer activity of microbubble conjugated with Sorafenib containing liposome and IL4R-targeting peptide in kidney cancer cells.

Microbubble-based cancer treatment is a promising new approach that utilizes tiny gas-filled bubbles to deliver cancer drugs directly to tumor sites. This study aims to investigate the anti-cancer effect of the novel microbubble (MB) complex conjugated with sorafenib containing liposome and interleukin 4 receptor (IL4R) targeting peptide in kidney cancer cells. MBs were synthesized by using a solvent with an emulsion evaporation technique. To target kidney tumor cells, the produced MBs were conjugated with sorafenib (SOR) loaded liposomes and peptide ligands for (IL4RTP). The anti-cancer effect of the MB complex was accessed by WST-1 assay, confocal microscopy analysis, and western blotting analysis. The finally prepared IL4RTP (MB-Lipo(SOR)-IL4RTP) showed an average size of 1,600 nm. A498, a kidney cancer cell line that expresses IL4Rα strongly, had an uptake of the MB-Lipo(SOR)-IL4RTP when exposed to frequency ultrasonic energy. Additionally, MB-Lipo(SOR)-IL4RTP suppressed the growth of A498 cells in an IL4R-dependent manner. This cell proliferation assay results were validated by western blotting analysis of the signal transduction proteins such as FOXO3, phosphorylated Erk, total Erk, and p27. Taken together, these findings show that MB-Lipo(SOR)-IL4RTP exerts the effective targeting capacity for A498 kidney cancer cells via regulation of Erk phosphorylation as a promising ultrasound contrast and therapeutic agent for treating kidney cancers.

Simultaneous monitoring of each component on degradation of blended bioplastic using gas chromatography-mass spectrometry.

The increasing interest in bioplastics, with regard to future environmental issues, has rendered research on bioplastic biodegradation highly important. However, only a few tools directly monitor the degradation of bioplastics without measuring the levels of gaseous products, such as carbon dioxide. Classical nonquantitative methods, such as clear zone tests on solid plates, and less-sensitive weight-loss experiments in liquid media measured using a precision scale, are still employed to screen the microbial players associated with bioplastic degradation and monitor the biodegradation rates. However, the simultaneous monitoring of the degradation of each component of blended bioplastics has not been previously reported. In the present study, to provide information regarding the degradation rates and compositional changes of different bioplastics in a blend in a time-dependent manner, we simultaneously monitored and quantified the degradation of four bioplastics, polyhydroxybutyrate (PHB), polybutylene succinate (PBS), polycaprolactone (PCL), and poly(butylene adipate-co-terephthalate) (PBAT), by Bacillus sp. JY36 using gas chromatography-mass spectrometry (GC-MS) analysis after fatty acid methyl ester (FAME) derivatization. Our results demonstrate the feasibility of using the GC-MS-based method described here to obtain comprehensive data regarding blended bioplastics and their degradation. Moreover, our findings indicate that this method may support classical analytic tools for assessing bioplastic biodegradation.

6,8-Diprenylorobol induces apoptosis in human colon cancer cells via activation of intracellular reactive oxygen species and p53.

6,8-Diprenylorobol is a natural compound mainly found in Glycyrrhiza uralensis fisch and Maclura tricuspidata, which has been used traditionally as food and medicine in Asia. So far, the antiproliferative effect of 6,8-diprenylorobol has not been studied yet in colon cancer. In this study, we aimed to evaluate the antiproliferative effects of 6,8-diprenylorobol in LoVo and HCT15, two kinds of human colon cancer cells. 6,8-Diprenylorobol inhibited the proliferation of LoVo and HCT15 cells in a dose- and time-dependent manner. A 40 µM of 6,8-diprenylorobol for 72 h reduced both of cell viability under 50%. After treatment of 6,8-diprenylorobol (40 and 60 µM) for 72 h, late apoptotic cell portion in LoVo and HCT15 cells were 24, 70% and 13, 90%, respectively, which was confirmed by checking DNA fragmentation in both cells. Mechanistically, 6,8-diprenylorobol activated p53 and its phosphorylated form (Ser15, Ser20, and Ser46) expression but suppressed Akt and mitogen-activated protein kinases (MAPKs) phosphorylation in LoVo and HCT15 cells. Interestingly, 6,8-diprenylorobol induced the generation of intracellular reactive oxygen species (ROS), which was attenuated with N-acetyl cysteine (NAC) treatment. Compared to the control, 60 µM of 6,8-diprenylorobol caused to increase ROS level to 210% in LoVo and HCT15, which was reduced into 161% and 124%, respectively with NAC. Furthermore, cell viability and apoptotic cell portion by 6,8-diprenylorobol was recovered by incubation with NAC. Taken together, these results indicate that 6,8-diprenylorobol has the potential antiproliferative effect against LoVo and HCT15 colon cancer cells through activation of p53 and generation of ROS.

Olfactory Receptor OR7A17 Expression Correlates with All-Trans Retinoic Acid (ATRA)-Induced Suppression of Proliferation in Human Keratinocyte Cells.

Olfactory receptors (ORs), which belong to the G-protein-coupled receptor family, have been widely studied as ectopically expressed receptors in various human tissues, including the skin. However, the physiological functions of only a few OR types have been elucidated in skin cells. All-trans retinoic acid (ATRA) is a well-known medication for various skin diseases. However, many studies have shown that ATRA can have adverse effects, resulting from the suppression of cell proliferation. Here, we investigated the involvement of OR7A17 in the ATRA-induced suppression of human keratinocyte (HaCaT) proliferation. We demonstrated that OR7A17 is expressed in HaCaT keratinocytes, and its expression was downregulated by ATRA. The ATRA-induced downregulation of OR7A17 was attenuated via RAR α or RAR γ antagonist treatment, indicating that the effects of ATRA on OR7A17 expression were mediated through nuclear retinoic acid receptor signaling. Moreover, we found that the overexpression of OR7A17 induced the proliferation of HaCaT cells while counteracting the antiproliferative effect of ATRA. Mechanistically, OR7A17 overexpression reversed the ATRA-induced attenuation of Ca2+ entry. Our findings indicated that ATRA suppresses cell proliferation through the downregulation of OR7A17 via RAR α- and γ-mediated retinoid signaling. Taken together, OR7A17 is a potential therapeutic target for ameliorating the anti-proliferative effects of ATRA.

Aspergillus oryzae-Fermented Wheat Peptone Enhances the Potential of Proliferation and Hydration of Human Keratinocytes through Activation of p44/42 MAPK.

Identifying materials contributing to skin hydration, essential for normal skin homeostasis, has recently gained increased research interest. In this study, we investigated the potential benefits and mechanisms of action of Aspergillus oryzae-fermented wheat peptone (AFWP) on the proliferation and hydration of human skin keratinocytes, through in vitro experiments using HaCaT cell lines. The findings revealed that compared to unfermented wheat peptone, AFWP exhibited an improved amino acid composition, significantly (p < 0.05) higher DPPH scavenging capability and cell proliferation activity, and reduced lipopolysaccharide-induced NO production in RAW 264.7 cells. Furthermore, we separated AFWP into eleven fractions, each ≤2 kDa; of these, fraction 4 (AFW4) demonstrated the highest efficacy in the cell proliferation assay and was found to be the key component responsible for the cell proliferation potential and antioxidant properties of AFWP. Additionally, AFW4 increased the expression of genes encoding natural moisturizing factors, including filaggrin, transglutaminase-1, and hyaluronic acid synthase 1-3. Furthermore, AFW4 activated p44/42 MAPK, but not JNK and p38 MAPK, whereas PD98059, a p44/42 MAPK inhibitor, attenuated the beneficial effects of AFW4 on the skin, suggesting that the effects of AFW4 are mediated via p44/42 MAPK activation. Finally, in clinical studies, AFW4 treatment resulted in increased skin hydration and reduced trans-epidermal water loss compared with a placebo group. Collectively, these data provide evidence that AFW4 could be used as a potential therapeutic agent to improve skin barrier damage induced by external stresses.

Simultaneous monitoring of the bioconversion from lysine to glutaric acid by ethyl chloroformate derivatization and gas chromatography-mass spectrometry.

Glutaric acid is a precursor of a plasticizer that can be used for the production of polyester amides, ester plasticizer, corrosion inhibitor, and others. Glutaric acid can be produced either via bioconversion or chemical synthesis, and some metabolites and intermediates are produced during the reaction. To ensure reaction efficiency, the substrates, intermediates, and products, especially in the bioconversion system, should be closely monitored. Until now, high performance liquid chromatography (HPLC) has generally been used to analyze the glutaric acid-related metabolites, although it demands separate time-consuming derivatization and non-derivatization analyses. To substitute for this unreasonable analytical method, we applied herein a gas chromatography - mass spectrometry (GC-MS) method with ethyl chloroformate (ECF) derivatization to simultaneously monitor the major metabolites. We determined the suitability of GC-MS analysis using defined concentrations of six metabolites (l-lysine, cadaverine, 5-aminovaleric acid, 2-oxoglutaric acid, glutamate, and glutaric acid) and their mass chromatograms, regression equations, regression coefficient values (R2), dynamic ranges (mM), and retention times (RT). This method successfully monitored the production process in complex fermentation broth.

Selective ratiometric red-emission detection of In3+ in aqueous solutions and in live cells using a fluorescent peptidyl probe and metal chelating agent.

Indium has been regarded as one of the most rarely used metal ions; however, the consumption of indium has increased intensively due to its increasing use in electrodes of liquid crystal displays (LCDs). In recent years, warnings have been issued about the toxicity of indium to aquatic ecosystems and humans. Thus, the development of efficient and selective detection methods for In3+ in aquatic environments as well as in live cells is highly required. However, the selective and sensitive detection of In3+ in the presence of trivalent metal ions and other metal ions is highly challenging. In the present study, we synthesized a fluorescent probe (1) for In3+ and Al3+ based on an unnatural peptide receptor and an aggregation-induced emission fluorophore and developed a selective fluorescent detection method for In3+ in aqueous solutions and live cells using the probe and a metal chelating agent. 1 recognized In3+ and Al3+ selectively among 19 metal ions in aqueous solutions depending on pH by the enhancement of the red emission at 600 nm and decrease in the green emission at 530 nm. 1 sensitively detected In3+ and Al3+ by ratiometric response in a wide pH range (3.5-7.4), and the ratiometric response was complete within 20 seconds in an aqueous buffered solution at pH 5.0. Interestingly, the addition of EDTA to the complex of 1 with In3+ or Al3+ did not induce the Al3+-free spectrum but instead induced the In3+-free spectrum; thus, In3+ and Al3+ could be easily differentiated. The detection limit of 1 for In3+ ions was 211 nM (R2 = 0.981) in purely aqueous solutions. The fluorescence ratiometric detection method using 1 could quantify low concentrations of In3+ in ground water and tap water. Fluorescence cell image studies revealed that the probe was cell-permeable, and low concentrations of In3+ inside the cells could be recognized by the enhancement of the red emission at 600 nm. The binding mode study via NMR, IR, and CD spectroscopy revealed how the peptide receptor of 1 interacted with In3+ and resulted in the enhancement of the red emission in an aqueous solution.

Cannabidiol induces osteoblast differentiation via angiopoietin1 and p38 MAPK.

In this study, we report the potential of cannabidiol, one of the major cannabis constituents, for enhancing osteoblastic differentiation in U2OS and MG-63 cells. Cannabidiol increased the expression of Angiopoietin1 and the enzyme activity of alkaline phosphatase in U2OS and MG-63. Invasion and migration assay results indicated that the cell mobility was activated by cannabidiol in U2OS and MG-63. Western blotting analysis showed that the expression of tight junction related proteins such as Claudin1, Claudin4, Occuludin1, and ZO1 was increased by cannabidiol in U2OS and MG-63. Alizarin Red S staining analysis showed that calcium deposition and mineralization was enhanced by cannabidiol in U2OS and MG-63. Western blotting analysis indicated that the expression of osteoblast differentiation related proteins such as distal-less homeobox 5, bone sialoprotein, osteocalcin, type I collagen, Runt-related transcription factor 2 (RUNX2), osterix (OSX), and alkaline phosphatase was time dependently upregulated by cannabidiol in U2OS and MG-63. Mechanistically, cannabidiol-regulated osteoblastic differentiation in U2OS and MG-63 by strengthen the protein-protein interaction among RUNX2, OSX, or the phosphorylated p38 mitogen-activated protein kinase (MAPK). In conclusion, cannabidiol increased Angiopoietin1 expression and p38 MAPK activation for osteoblastic differentiation in U2OS and MG-63 suggesting that cannabidiol might provide a novel therapeutic option for the bone regeneration.

Effects of hepatocyte growth factor gene-transfected mesenchymal stem cells on dimethylnitrosamine-induced liver fibrosis in rats.

Nowadays, transplantation of human mesenchymal stem cells (MSCs) has emerged as a potential cellular therapy for liver cirrhosis. Hepatocyte growth factor (HGF) plays an important role in the regeneration of the liver. The objective of the study was to investigate the therapeutic effect of HGF-transfected human umbilical cord blood-derived MSCs on dimethylnitrosamine (DMN)-induced liver fibrosis in rats. HGF-transfected MSCs were transplanted into rats with DMN-induced liver fibrosis. H2O2-induced cytotoxicity, apoptosis and intracellular reactive oxygen species were reduced in HGF-transfected MSCs in HGF-transfected MSCs. Pro-apoptotic proteins, such as cleaved poly (ADP-ribose) polymerase and cleaved caspase-3, were decreased in HGF-transfected MSCs. Biochemical analysis showed that the levels of aspartate aminotransferase and alanine aminotransferase were decreased after transplantation of HGF-transfected MSCs in rat fibrosis. Trichrome staining showed that HGF-transfected MSCs reduced liver damage. Taken together, our study indicated that HGF-transfected MSCs have therapeutic effects on DMN-induced liver fibrosis in rats.

Selective red-emission detection for mercuric ions in aqueous solution and cells using a fluorescent probe based on an unnatural peptide receptor.

The selective ratiometric red-emission detection of Hg2+ ions in aqueous buffered solutions and live cells is still a significant challenge. In the present study, we synthesized a fluorescent probe (1) based on an unnatural peptide receptor containing sulfonamide groups with an aggregation-induced emission and twisted internal charge transfer (TICT)-active fluorophore, cyanostilbene. 1 exhibited a highly selective ratiometric response to Hg2+ among 14 metal ions tested by ratiometric red-emission at 600 nm, with a clear isoemissive point in purely aqueous solution containing 1% DMSO. The ratiometric response for Hg2+ ions was complete within 3 min and the ratiometric responses induced by Hg2+ ions did not suffer considerable interference from the other metal ions. The ratiometric response was complete for less than 7 µM Hg2+ and 1 had a potent binding affinity (7.42 × 10-6 M, R2 = 0.98) for Hg2+ and a nanomolar detection limit. 1 detected Hg2+ ions by ratiometric responses in aqueous buffered solutions over a wide range of pH (5.5-11.5). Binding mode studies using TEM, NMR, IR, and a mass spectrometer revealed that the sulfonamide groups of the unnatural peptide receptor played an important role in the complexation of Hg2+ and in the complexation-induced nano-sized aggregates, which resulted in a significant increase in emissions at 600 nm and a decrease in emissions at 535 nm. 1 quantified micro-molar concentrations (0-6 µM) of Hg2+ in tap water and groundwater by ratiometric detection. Furthermore, 1 passed through the lipid membranes of live cells and detected intracellular Hg2+ ions at 2 µM by a ratiometric red-emission change.

Protective effects of hepatocyte growth factor gene overexpression against hydrogen peroxide-induced apoptosis in mesenchymal stem cells.

Hepatocyte growth factor (HGF) has recently been reported to exhibit antioxidant and antiapoptotic effects. Therefore, we investigated the effect of overexpression of HGF gene in H2 O2 -treated mesenchymal stem cells (MSCs). HGF-overexpression increased the cell viability from 50% to 84%, decreased the population of apoptotic cells from 20% to 16%, and decreased the intracellular reactive oxygen species (ROS) levels from 127% to 100% in cells treated with H2 O2 . HGF suppression decreased the cell viability from 58% to 36%, increased the population of apoptotic cells from 23 to 81%, and increased the intracellular ROS levels from 181% to 240% in cells exposed to H2 O2 . HGF-overexpression also reduced the expression levels of proapoptotic proteins in MSCs treated with H2 O2 . Phosphorylation of extracellular signal-regulated kinases, c-Jun amino-terminal kinases, and p38, which was induced by H2 O2 , decreased in MSCs overexpressing the HGF gene. Taken together, our results suggest that HGF has a protective effect on H2 O2 -induced apoptosis in MSCs.

Esculetin induces cell cycle arrest and apoptosis in human colon cancer LoVo cells.

We investigated the anti-cancer effects of ESC in human colon cancer LoVo cells. Cell counting assay results showed that ESC inhibited the proliferation of LoVo cells. Cell cycle arrest results showed that cell cycle was arrested during the G0/G1 phase in the ESC-treated LoVo cells. Western blot results showed that the cell cycle inhibitory proteins p53, p27, and p21 were increased, and cyclin D1, the cell cycle progressive protein, was decreased. Sp1 is a transcription factor regulating cell proliferation, was decreased in the ESC-treated LoVo cells. Annexin V/propidium iodide staining results showed that ESC induces apoptosis in LoVo cells. Western blot results showed that Bax, cleaved caspase -3, -7, -9, and poly(ADP-ribose) polymerase, which are proapoptotic proteins, were increased and the antiapoptotic protein Bcl-2 was decreased. Taken together, ESC induced apoptosis and has an anti-cancer effect in LoVo cells.

Tomentosin Displays Anti-Carcinogenic Effect in Human Osteosarcoma MG-63 Cells via the Induction of Intracellular Reactive Oxygen Species.

Tomentosin is a natural sesquiterpene lactone extracted from various plants and is widely used as a medicine because it exhibits essential therapeutic properties. In this study, we investigated the anti-carcinogenic effects of tomentosin in human osteosarcoma MG-63 cells by performing cell migration/viability/proliferation, apoptosis, and reactive oxygen species (ROS) analysis assays. MG-63 cells were treated with various doses of tomentosin. After treatment with tomentosin, MG-63 cells were analyzed using the MTT assay, colony formation assay, cell counting assay, wound healing assay, Boyden chamber assay, zymography assay, cell cycle analysis, FITC Annexin V apoptosis assay, terminal deoxynucleotidyl transferase dUTP nick end labeling assay, western blot analysis, and ROS detection analysis. Our results indicated that tomentosin decreased cell viability and migration ability in MG-63 cells. Moreover, tomentosin induced apoptosis, cell cycle arrest, DNA damage, and ROS production in MG-63 cells. Furthermore, tomentosin-induced intracellular ROS decreased cell viability and induced apoptosis, cell cycle arrest, and DNA damage in MG-63 cells. Taken together, our results suggested that tomentosin exerted anti-carcinogenic effects in MG-63 cells by induction of intracellular ROS.

Ratiometric red-emission fluorescence detection of Al3+ in pure aqueous solution and live cells by a fluorescent peptidyl probe using aggregation-induced emission.

The development of a fluorescence method for the selective ratiometric detection of Al3+ ions in pure aqueous solutions and live cells is still a significant challenge. In the present study, we synthesized a new type of fluorescent probe using an Al3+-triggered self-assembly based on the dipeptide receptor and an aggregation-induced emission fluorophore. The fluorescent probe (1) bearing cyanostilbene with excitation by visible light detected Al3+ ions sensitively in pure aqueous buffered solution by ratiometric red-emission at 600 nm. 1 provided a highly selective ratiometric detection of Al3+ among 16 metal ions in aqueous solution. 1 exhibited sensitive ratiometric response to Al3+ in aqueous buffered solutions at pH ranging from 5 to 7.4. The detection limit (145 nM, R2 = 0.999) for Al3+ ions in pure aqueous solution was much lower than the maximum allowable level of Al3+ in drinking water demanded by the Environmental Protection Agency (EPA). The probe provided an efficient approach to detect low concentrations of Al3+ in ground water, tap water, and live cells by ratiometric red-emissions at 600 nm. The binding study using dynamic light scattering, NMR, IR, and TEM revealed that the complex between 1 and Al3+ self-assembled to form nanoparticles, resulting in the enhancement of the emission at 600 nm and a concomitant decrease in the emission at 535 nm.

Anti-melanogenic effects of resorcinol are mediated by suppression of cAMP signaling and activation of p38 MAPK signaling.

In this study, we investigated the inhibitory mechanisms of resorcinol in B16F10 mouse melanoma cells. We found that resorcinol reduced both the melanin content and tyrosinase activity in these cells. In addition, resorcinol suppressed the expression of melanogenic gene microphthalmia-associated transcriptional factor (MITF) and its downstream target genes tyrosinase, tyrosinase-related protein (TRP)-1, and TRP-2. In addition, we found that resorcinol reduced intracellular cAMP levels and protein kinase A (PKA) activity, and increased phosphorylation of the p38 mitogen-activated protein kinase (MAPK). Resorcinol was also found to directly inhibit tyrosinase activity. However, resorcinol-induced decrease in melanin content, tyrosinase activity, and tyrosinase protein levels were attenuated by SB203580, a p38 MAPK inhibitor. Taken together, these data indicate that anti-melanogenic activity of resorcinol is be mediated through the inhibition of cAMP signaling and activation of p38 MAPK, indicating that resorcinol may be a possible ameliorating agent in the treatment of hyperpigmentation skin disorders.

Negative Cellular Effects of Urban Particulate Matter on Human Keratinocytes Are Mediated by P38 MAPK and NF-κB-dependent Expression of TRPV 1.

Urban particulate matter (UPM) exerts negative effects on various human organs. Transient receptor potential vanilloid 1 (TRPV1) is a polymodal sensory transducer that can be activated by multiple noxious stimuli. This study aimed to explore the effects of the UPM 1648a on the expression of TRPV1, and its regulatory mechanisms in HaCaT cells. UPM enhanced TRPV 1 promoter-luciferase reporter activity. UPM also increased expression of the TRPV 1 gene as evidenced by increased mRNA and protein levels of TRPV 1. In addition, elucidation of the underlying mechanism behind the UPM-mediated effects on TRPV 1 expression revealed that UPM can upregulate expression of the TRPV1 gene by activating activator protein-1 (AP-1) and nuclear factor kappa B (NF-κB). The UPM treatment also altered Ca2+ influx and cell proliferation, as well as production of interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α), and interleukin-1ß (IL-1ß). In addition, these UPM-induced effects were attenuated by SB203580 and ammonium pyrrolidinedithiocarbamate (PDTC). However, SP600125 and PD98059 did not alter the UPM-induced effects. Taken together, these findings indicate that UPM upregulates expression of the TRPV 1 gene, which is mediated by the p38 mitogen-activated protein kinase (MAPK) and NF-κB signaling pathways and suggest that UPM is a potential irritant that can induce skin processes such as aging and inflammatory responses.

CK2α/CSNK2A1 Phosphorylates SIRT6 and Is Involved in the Progression of Breast Carcinoma and Predicts Shorter Survival of Diagnosed Patients.

Recently, the roles of sirtuins (SIRTs) in tumorigenesis have been of interest to oncologists, and protein kinase CK2 α1 (CSNK2A1) has been shown to be involved in tumorigenesis by phosphorylating various proteins, including SIRT1. Therefore, we evaluated the roles of CSNK2A1, SIRT6, and phosphorylated SIRT6 and their relationships in breast carcinoma. Nuclear expression of CSNK2A1 and SIRT6 predicted shorter overall survival and relapse-free survival by multivariate analysis. Inhibition of CSNK2A1 decreased the proliferative and invasive activity of cancer cells. In addition, CSNK2A1 was bound to SIRT6 and phosphorylated SIRT6; evidence for this is provided from immunofluorescence staining, co-immunoprecipitation of CSNK2A1 and SIRT6, a glutathione S-transferase pull-down assay, an in vitro kinase assay, and transfection of mutant CSNK2A1. Knockdown of SIRT6 decreased the proliferation and invasiveness of cancer cells. Overexpression of SIRT6 increased proliferation, but mutation at the Ser338 phosphorylation site of SIRT6 inhibited the proliferation of MCF7 cells. Moreover, both knockdown of SIRT6 and a mutation at the phosphorylation site of SIRT6 decreased expression of matrix metallopeptidase 9, ß-catenin, cyclin D1, and NF-κB. Especially, SIRT6 expression was associated with the nuclear localization of ß-catenin. This study demonstrates that CSNK2A1 and SIRT6 are indicators of poor prognosis for breast carcinomas and that CSNK2A1-mediated phosphorylation of SIRT6 might be involved in the progression of breast carcinoma.

Inhibition of collagenase and melanogenesis by ethanol extracts of Orostachys japonicus A. Berger: possible involvement of Erk and Akt signaling pathways in melanoma cells.

Orostachys japonicus is an herb that contains several functional components and has traditionally been used to treat various diseases in Asia. In this study, bioactive components from different parts of the O. japonicus plant were investigated, and the contents of the functional components in ethanol extracts of O. japonicus cultivated in Korea and China were compared. The antioxidant effects of O. japonicus ethanol extracts were investigated using Raw 264.7 cells. It was found that 2,2-diphenyl-1-picrylhydrazyl radical-scavenging activity was significantly decreased in the cells treated with the extracts. Moreover, the novel inhibitory functions of O. japonicus extracts on collagenase, elastase, and tyrosinase were established. We also found that O. japonicus extracts strongly inhibited melanin synthesis in B16F10 melanoma cells by decreasing MITF protein levels and activating the Erk and Akt signaling pathways. Thus, these findings would be useful for developing new cosmetic and pharmaceutical formulations based on O. japonicus extracts.

Potential of l-thyroxine to differentiate osteoblast-like cells via Angiopoietin1.

Angiogenesis is closely associated with osteoblast differentiation. Previously, we demonstrated that bone formation can be accelerated by treatment with COMP-Angiopoietin1, a known angiogenic factor. Angiopoietin1 (Ang1) is a specific growth factor that generates stable and mature vasculature through the Tie2 receptor. In this study, we aimed to identify a novel drug that can activate endogenous Ang1 expression as a pharmacological treatment for bone formation. Therefore, Ang1 expression was examined in U2OS osteoblast-like cells treated with 770 drugs from a library of Food and Drug Administration (FDA)-approved drugs by using ELISA for Ang1. l-thyroxine was selected as a novel drug candidate. l-Thyroxine is a synthetic form of the hormone thyroxine, which is used to treat patients with hypothyroidism. Enzyme-linked immunosorbent assays (ELISAs) were performed to test whether Ang1 is induced in a dose-dependent manner in human osteoblast-like cell lines, U2OS and MG63. The effects of l-thyroxine on osteoblast differentiation and mineralization were evaluated by alkaline phosphatase (ALP) activity and Alizarin red s staining. To determine the molecular mechanism, the expression of proteins related to bone formation and differentiation, such as type I collagen (COL1A1), osteocalcin (OC), bone sialoprotein (BSP), distal-less homeobox 5 (Dlx5), Runt-related transcription factor 2 (Runx2), osterix (OSX), and ALP, was tested by Western blotting analysis. Consequently, l-thyroxine induced Ang1 expression in a dose-dependent manner in both U2OS and M63 cells, which was confirmed by ELISA and Western blotting. Also, l-thyroxine activated ALP activity in U2OS and MG63 cells as well as ALP expression. Furthermore, l-thyroxine enhanced the expression of COL1A1, Runx2, OC, BSP, Dlx5, and OSX mRNA and proteins. Taken together, we demonstrated that l-thyroxine increased Ang1 expression and induces bone formation, differentiation, and mineralization in U2OS and MG63 cell lines, which suggests that l-thyroxine could be a potential bone production agent.