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OBJECTIVES Doxorubicin is a DNA-damaging agent used to treat hematological cancers. Unfortunately, drug resistance can occur by defective DNA repair activity coupled with the ability of DNA polymerases to misreplicate unrepaired DNA lesions. This study demonstrates that the efficacy of doxorubicin can be improved by using an artificial nucleoside to efficiently and selectively inhibit this activity. METHODS In vitro studies using acute lymphoblastic leukemia cell lines define the mechanism of cell death caused by combining an artificial nucleoside with doxorubicin. RESULTS Flow cytometry experiments demonstrate that combining an artificial nucleoside with doxorubicin potentiates the cell killing effects of the drug by increasing apoptosis. The potentiation effect correlates with expression of TdT, a specialized DNA polymerase overexpressed in acute lymphoblastic leukemia. Cell cycle experiments demonstrate that this combination blocks cells at S-phase prior to inducing apoptosis. Finally, the unique chemical composition of the nucleoside analog was used to visualize the replication of damaged DNA in TdT-positive cells. This represents a potential diagnostic tool to easily identify doxorubicin-resistant cancer cells. CONCLUSION Studies demonstrate that a novel artificial nucleoside improves the therapeutic efficacy of doxorubicin, thereby reducing the risk of potential side effects caused by the DNA-damaging agent.

Temozolomide is a DNA-alkylating agent used to treat brain tumors, but resistance to this drug is common. In this study, we provide evidence that efficacious responses to this drug can be heightened significantly by coadministration of an artificial nucleoside (5-nitroindolyl-2′-deoxyriboside, 5-NIdR) that efficiently and selectively inhibits the replication of DNA lesions generated by temozolomide. Conversion of this compound to the corresponding nucleoside triphosphate, 5-nitroindolyl-2′-deoxyriboside triphosphate, in vivo creates a potent inhibitor of several human DNA polymerases that can replicate damaged DNA. Accordingly, 5-NIdR synergized with temozolomide to increase apoptosis of tumor cells. In a murine xenograft model of glioblastoma, whereas temozolomide only delayed tumor growth, its coadministration with 5-NIdR caused complete tumor regression. Exploratory toxicology investigations showed that high doses of 5-NIdR did not produce the side effects commonly seen with conventional nucleoside analogs. Collectively, our results offer a preclinical pharmacologic proof of concept for the coordinate inhibition of translesion DNA synthesis as a strategy to improve chemotherapeutic responses in aggressive brain tumors. Significance: Combinatorial treatment of glioblastoma with temozolomide and a novel artificial nucleoside that inhibits replication of damaged DNA can safely enhance therapeutic responses. Cancer Res; 78(4); 1083–96. ©2017 AACR.

Background Due to the recent spread of coronavirus disease 2019, Korean nursing colleges are increasingly using virtual patient simulation to make up for a lack of available clinical practice in medical institutions. Therefore, an instrument is required to evaluate the effects of the virtual patient learning system in South Korea. Objective To assess the validity and reliability of the Korean version of the Virtual Patient Learning System Evaluation Tool (K-VPLSET). Design This is a methodological study. Settings This study was conducted via an online survey for Korean nursing students. Participants The present study included 373 participants who were 3rd and 4th year Korean nursing students. Methods After translating the English version of VPLSET into Korean, a pilot test with a cognitive interview was undertaken to ensure that the meaning of original instrument and appropriateness for Korean nursing students had been retained. The content validity of the K-VPLSET was examined by a panel of six experts. Convenience sampling was used to recruit 3rd and 4th year Korean nursing students, among whom 170 were recruited for exploratory factor analysis (EFA) and 203 for confirmatory factor analysis (CFA). SPSS version 26.0 was used for EFA, whereas AMOS version 22.0 was used for CFA. Results From the 32 initial items, the final version of the K-VPLSET ultimately included 20 items, with a Cronbach's α of 0.89. EFA identified four factors (“Nursing Knowledge Improvement,” “Clinical Competency Development,” “Confidence in Nursing Performance,” and “Nursing Care Plan Application”) that explained 56.9% of the total variance. CFA confirmed the validity of the instrument. Conclusions Our findings confirmed that the K-VPLSET is a valid and reliable instrument for assessing the effects of the virtual patient learning system, through which the quality of e-learning for Korea nursing students can be determined.

// Jung-Suk Choi 1 , Seol Kim 2 , Edward Motea 3 and Anthony Berdis 1, 2, 4, 5 1 Department of Chemistry, Cleveland State University, Cleveland, OH 44115, USA 2 Department of Biological, Geological, and Environmental Sciences, Cleveland State University, Cleveland, OH 44115, USA 3 Departments of Radiation Oncology and Pharmacology, Harold C. Simmons Comprehensive Cancer Center, UT Southwestern Medical Center, Dallas, TX 75390, USA 4 Center for Gene Regulation in Health and Disease, Cleveland State University, Cleveland, OH 44115, USA 5 Case Comprehensive Cancer Center, Cleveland, OH 44106, USA Correspondence to: Anthony Berdis, email: a.berdis@csuohio.edu Keywords: DNA damage, DNA polymerization, chemotherapy, nucleoside analogs, leukemia Received: February 24, 2017 Accepted: April 11, 2017 Published: April 19, 2017 ABSTRACT Anti-cancer agents exert therapeutic effects by damaging DNA. Unfortunately, DNA polymerases can effectively replicate the formed DNA lesions to cause drug resistance and create more aggressive cancers. To understand this process at the cellular level, we developed an artificial nucleoside that visualizes the replication of damaged DNA to identify cells that acquire drug resistance through this mechanism. Visualization is achieved using "click" chemistry to covalently attach azide-containing fluorophores to the ethynyl group present on the nucleoside analog after its incorporation opposite damaged DNA. Flow cytometry and microscopy techniques demonstrate that the extent of nucleotide incorporation into genomic DNA is enhanced by treatment with DNA damaging agents. In addition, this nucleoside analog inhibits translesion DNA synthesis and synergizes the therapeutic activity of certain anti-cancer agents such as temozolomide. The combined diagnostic and therapeutic activities of this synthetic nucleoside analog represent a new paradigm in personalized medicine.

Background: Nucleoside transport is an important therapeutic target against cancer. Results: Cell-based studies show that a metal-containing nucleoside displays anti-cancer effects and diagnostic properties against adherent cancers. Conclusion: A metal-containing nucleoside functions as a biophotonic substrate for a nucleoside transporter. Significance: A novel nucleoside with combined therapeutic and diagnostic activities provides a tool to treat cancer. Nucleoside transport is an essential process that helps maintain the hyperproliferative state of most cancer cells. As such, it represents an important target for developing diagnostic and therapeutic agents that can effectively detect and treat cancer, respectively. This report describes the development of a metal-containing nucleoside designated Ir(III)-PPY nucleoside that displays both therapeutic and diagnostic properties against the human epidermal carcinoma cell line KB3-1. The cytotoxic effects of Ir(III)-PPY nucleoside are both time- and dose-dependent. Flow cytometry analyses validate that the nucleoside analog causes apoptosis by blocking cell cycle progression at G2/M. Fluorescent microscopy studies show rapid accumulation in the cytoplasm within 4 h. However, more significant accumulation is observed in the nucleus and mitochondria after 24 h. This localization is consistent with the ability of the metal-containing nucleoside to influence cell cycle progression at G2/M. Mitochondrial depletion is also observed after longer incubations (Δt ∼48 h), and this effect may produce additional cytotoxic effects. siRNA knockdown experiments demonstrate that the nucleoside transporter, hENT1, plays a key role in the cellular entry of Ir(III)-PPY nucleoside. Collectively, these data provide evidence for the development of a metal-containing nucleoside that functions as a combined therapeutic and diagnostic agent against cancer.

Purpose: This study aimed to confirm the mediating effect of job involvement in the relationship between grit and turnover intention among nurses working at university hospitals. Methods: Participants included 437 nurses from university hospitals located in C city, Gyeongnam. Data were collected from January 8 to 19, 2018, using self-report questionnaires. Data were analyzed using the t-test, analysis of variance, Scheffe’s test, Pearson’s correlation coefficient, and multiple regression, with the SPSS/22.0 program. A mediation analysis was performed according to the Baron and Kenny, and bootstrapping methods. Results: There were significant relationships between grit and job involvement (r=.40, p

Nucleoside analogs are an important class of anticancer agent that historically show better efficacy against hematological cancers versus solid tumors. This report describes the development and characterization of a new class of nucleoside analog that displays anticancer effects against both hematological and adherent cancer cell lines. These new analogs lack canonical hydrogen-bonding groups yet are effective nucleotide substrates for several high-fidelity DNA polymerases. Permutations in the position of the non-hydrogen-bonding functional group greatly influence the kinetic behavior of these nucleosides. One particular analog designated 4-nitroindolyl-2'-deoxynucleoside triphosphate (4-NITP) is unique as it is incorporated opposite C and T with high catalytic efficiencies. In addition, this analog functions as a nonobligate chain terminator of DNA synthesis, since it is poorly elongated. Consistent with this mechanism, the corresponding nucleoside, 4-nitroindolyl-2'-deoxynucleoside (4-NIdR), produces antiproliferative effects against leukemia cells. 4-NIdR also produces cytostatic and cytotoxic effects against several adherent cancer cell lines, especially those that are deficient in mismatch repair and p53. Cell death in this case appears to occur via mitotic catastrophe, a specialized form of apoptosis. Mass spectroscopy experiments performed on nucleic acid isolated from cells treated with 4-NIdR validate that the non-natural nucleoside is stably incorporated into DNA. Xenograft mouse studies demonstrate that administration of 4-NIdR delays tumor growth without producing adverse side effects such as anemia and thrombocytopenia. Collectively, the results of in vitro, cell-based, and animal studies provide evidence for the development of a novel nucleoside analog that shows enhanced effectiveness against solid tumors.

Human leukocyte endothelial adhesion and transmigration occur in the early stage of the pathogenesis of atherosclerosis. Vascular endothelial cells are targeted by pro-inflammatory cytokines modulating many gene proteins responsible for cell adhesion, thrombosis and inflammatory responses. This study examined the potential of compound K to inhibit the pro-inflammatory cytokine TNF-α induction of monocyte adhesion onto TNF-α-activated human umbilical vein endothelial cells (HUVEC). HUVEC were cultured with 10ng/ml TNF-α with individual ginsenosides of Rb1, Rc, Re, Rh1 and compound K (CK). Ginsenosides at doses of ⩽50μM did not show any cytotoxicity. TNF-α induced THP-1 monocyte adhesion to HUVEC, and such induction was attenuated by Rh1 and CK. Consistently, CK suppressed TNF-α-induced expression of HUVEC adhesion molecules of VCAM-1, ICAM-1 and E-selectin, and also Rh1 showed a substantial inhibition. Rh1 and CK dampened induction of counter-receptors, α4/β1 integrin VLA-4 and αL/β2 integrin LFA-1 in TNF-α-treated THP-1 cells. Additionally, CK diminished THP-1 secretion of MMP-9 required during transmigration, inhibiting transendothelial migration of THP-1 cells. CK blunted TNF-α-promoted IL-8 secretion of HUVEC and CXCR1 expression of THP-1 monocytes. Furthermore, TNF-α-activated endothelial IκB phosphorylation and NF-κB nuclear translocation were disturbed by CK, and TNF-α induction of α4/β1 integrin was abrogated by the NF-κB inhibitor SN50. These results demonstrate that CK exerts anti-atherogenic activity with blocking leukocyte endothelial interaction and transmigration through negatively mediating NF-κB signaling.

The accumulation of metal ions and amyloid- β (A β ) aggregates found in the brain of patients with Alzheimer’s disease (AD) has been suggested to be involved in AD pathogenesis. To investigate metal-A β -associated pathways in AD, development of chemical tools to target metal-A β species is desired. Only a few efforts, however, have been reported. Here, we report bifunctional small molecules, N -(pyridin-2-ylmethyl)aniline ( L2-a ) and N 1 , N 1 -dimethyl- N 4 -(pyridin-2-ylmethyl)benzene-1,4-diamine ( L2-b ) that can interact with both metal ions and A β species, as determined by spectroscopic methods including high-resolution NMR spectroscopy. Using the bifunctional compound L2-b , metal-induced A β aggregation and neurotoxicity were modulated in vitro as well as in human neuroblastoma cells. Furthermore, treatment of human AD brain tissue homogenates containing metal ions and A β species with L2-b showed disassembly of A β aggregates. Therefore, our studies presented herein demonstrate the value of bifunctional compounds as chemical tools for investigating metal-A β -associated events and their mechanisms in the development and pathogenesis of AD and as potential therapeutics.

Oxidized LDL (oxLDL) has been implicated in the pathogenesis of atherosclerosis accompanying lipid-laden cell appearance, inflammatory responses, and vascular dysfunction. This study examined the potentials of polyphenol quercitrin to inhibit oxLDL induction of scavenger receptor A (SR-A) and CD36 involving activation of peroxisome proliferator-activated receptor gamma (PPARgamma). J774A1 murine macrophages were cultured with 10 microg/mL Cu(2+)-oxLDL for various times in the presence of 1-10 micromol/L quercitrin. Cu(2+)-oxLDL at the given concentration facilitated macrophage proliferation and enhanced oxLDL uptake. Quercitrin dampened oxLDL uptake and lipid accumulation elevated in macrophages exposed to oxLDL. Western blot analysis revealed that 10 microg/mL oxLDL upregulated expression of SR-A and CD36, which was rapidly abolished at the transcriptional levels by 10 micromol/L quercitrin within 4 h. Quercitrin diminished production of proinflammatory and proatherogenic vascular endothelial growth factor that augmented through the oxLDL binding to CD36. Similarly, quercitrin repressed expression of macrophage inflammatory protein-2 and monocyte chemoattractant protein-1 involved in monocyte trafficking and macropahage migration. In addition, quercitrin attenuated oxLDL-induced transcriptional activation of PPARgamma leading to CD36 induction. Furthermore, quercitrin alleviated macrophage uptake of oxLDL through interfering with PKC-PPAR signaling cascades. These results demonstrate that quercitrin blocked oxLDL uptake, cholesterol influx and lipid-laden foam cell formation through inhibiting induction of SR and VEGF linked to PKCalpha-PPARgamma-responsive pathways. Therefore, quercitrin may be an antiatherogenic agent blocking foam cell formation pertaining to induction of SR and VEGF.

The aberrant expression of matrix metalloproteinases (MMPs) has been implicated in matrix degradation leading to angiogenesis. This study examined the inhibitory effects of isoliquiritigenin (ISL) on phorbol myristate acetate (PMA)-induced MMP production and its tissue inhibitor of MMP (TIMP) in endothelial cells. No induction of either necrotic or apoptotic cell death was observed in response to a treatment with ISL ator=25 microM. ISL dose-dependently suppressed PMA-induced expression and activity of MMP-2 and membrane type 1-MMP ator=1 microM while diminishing the elevated MMP-2 transcript level. In addition, ISL inhibited PMA-triggered migration and tube formation in a dose-dependent manner. ISL further increased the TIMP production up-regulated by PMA with a biphasic effect on TIMP-2 expression. This study further attempted to investigate whether a c-Jun N-terminal kinase (JNK)- or p38 mitogen-activated protein kinase (MAPK)-responsive mechanism was responsible for the MMP production and whether ISL disturbed these signaling pathways. PMA stimulated signaling of JNK and p38 MAPK, which was dampened byor=10 microM ISL. These results demonstrate that ISL blocked JNK- or p38 MAPK-responsive pathways leading to direct MMP activation of PMA-exposed endothelial cells. Therefore, the ISL inhibition of MMP may boost a therapeutic efficacy during angiogenesis.

Endothelial apoptosis is a driving force in atherosclerosis development. Oxidized LDL promotes inflammatory and thrombotic processes and is highly atherogenic, as it stimulates macrophage cholesterol accumulation and foam cell formation. This study investigated multiple mitogen-activated protein kinase (MAPK)-responsive death/survival signaling pathways, through which flavonoids of (-)epigallocatechin gallate (EGCG) and hesperetin exerted antiapoptosis in endothelial cells exposed to oxidized LDL. EGCG and hesperetin substantially diminished the oxidized LDL-induced 2',7'-dichlorofluorecein staining, suggesting that these flavonoids inhibited intracellular accumulation of oxidized LDL-triggered reactive oxygen species and consequent apoptosis. The Western-blot data revealed that oxidized LDL upregulated c-Jun N-terminal kinase (JNK) phosphorylation, which was rapidly reversed by EGCG and hesperetin. They mitigated the consequent activation of the JNK downstream on p53 and c-Jun. Moreover, oxidized LDL increased luciferase activity of p53 in endothelial cells transfected with a p53 promoter construct, the increase of which was strikingly downregulated by EGCG and hesperetin. Surprisingly, hesperetin but not EGCG attenuated phosphorylation of p38MAPK and its downstream c-myc and signal transducers and activators of transcription (STAT)1 evoked by oxidized LDL. This study also attempted to explore a linkage of Janus kinase (JAK)2/STAT3 activation to MAPK signaling in oxidized LDL-induced endothelial apoptosis. Notably, we found that the JAK2 inhibitor substantially blocked the JNK activation. Our findings suggest that EGCG and hesperetin may act as antiatherogenic agents blocking oxidized LDL-induced endothelial apoptosis via differential cellular apoptotic machinery. These data provide evidence that the interplay between p38MAPK and JAK-STAT pathways is involved in dietary flavonoid protection against oxidized LDL through hampering MAPK-dependent pathways involving the activation of JAK2.

Ultraviolet (UV) irradiation leads to distinct changes in skin connective tissues by degradation of collagen, which is a major structural component in the extracellular matrix most likely mediated by matrix metalloproteinases (MMP), collagenases. These changes in collagenous skin tissues have been suggested to be causes of the skin wrinkling observed in premature aging of the skin. This study mimicked the action of environmental ultraviolet on skin and investigated whether (-)epigallocatechin gallate (EGCG), a bioactive catechin component of green tea, mechanistically inhibited activation of MMP-1, MMP-8, and MMP-13 and destruction of collagen in UV-B irradiated human dermal fibroblasts by modulating cellular signaling pathways. Cell viability was moderately decreased by > or = 30% in human dermal fibroblasts treated with 100 mJ/cm2 UV-B, accompanying a substantial generation of reactive oxygen species evidenced by DCF staining. Western blot analysis and immunocytochemical staining revealed that EGCG markedly suppressed collagen degradation enhanced in UV-B-exposed human dermal fibroblast. Pre-treatment of fibroblasts with EGCG also inhibited UV-B-induced production of collagenases, MMP-1, MMP-8 and MMP-13, in a dose-dependent manner. In addition, EGCG rapidly and substantially hampered UV-B irradiation-induced activation of ASK-1 and phosphorylation of MAPK, JNK, p38 MAPK, and ERK1/2, in dermal fibroblasts. These results demonstrate that EGCG has abilities to hamper UV-B-induced collagenolytic MMP production via interfering with the MAPK-responsive pathways. Therefore, EGCG may be a potential agent for the prevention and treatment of skin photoaging.

Nucleosides and their corresponding mono-, di-, and triphosphates play important roles in maintaining cellular homeostasis. In addition, perturbations in this homeostasis can result in dysfunctional cellular processes that cause pathological conditions such as cancer and autoimmune diseases. This review article discusses contemporary research areas applying nucleoside analogs to probe the mechanistic details underlying the complexities of nucleoside metabolism at the molecular and cellular levels. The first area describes classic and contemporary approaches used to quantify the activity of nucleoside transporters, an important class of membrane proteins that mediate the influx and efflux of nucleosides and nucleobases. A focal point of this section is describing how biophotonic nucleosides are replacing conventional assays employing radiolabeled substrates to study the mechanism of these proteins. The second section describes approaches to understand the utilization of nucleoside triphosphates by cellular DNA polymerases during DNA synthesis. Emphasis here is placed on describing how novel nucleoside analogs such as 5-ethynyl-2'-deoxyuridine are being used to quantify DNA synthesis during normal replication as well as during the replication of damaged DNA. In both sections, seminal research articles relevant to these areas are described to highlight how these novel probes are improving our understanding of these biological processes. This article is part of a Special Issue entitled: Physiological Enzymology and Protein Functions.

Flavones have been classified as anti-atherogenic agents that inhibit monocyte adhesion to stimulated endothelium, possibly by blocking induction of cell adhesion molecules (CAM). This anti-atherogenic feature of these flavonoids appears to be related to their chemical structures. Flavones may interfere with key signaling events involved in endothelial cell activation by inflammatory mediators. This study examined the effects of flavones on the induction of CAM and the translocation and DNA binding of nuclear factor-kappa B (NF-kappa B) in TNF-alpha-activated human umbilical vein endothelial cells (HUVEC). The effects of flavones, luteolin and apigenin, on adhesion of THP-1 monocytes to the TNF-alpha-activated HUVEC, protein expression and mRNA levels of vascular cell adhesion molecule-1 (VCAM-1), intracellular cell adhesion molecule-1 (ICAM-1) and E-selectin, and nuclear appearance and DNA binding activity of NF-kappa B were determined. Flavanols, flavonols, and flavanones were used for comparison. TNF-alpha significantly induced HUVEC protein expression of VCAM-1, ICAM-1, and E-selectin with increasing mRNA levels. Luteolin and apigenin inhibited the TNF-alpha-induced upregulation of THP-1 adhesion and VCAM-1 expression; these inhibitory effects were dose-dependent. The flavones at doses of > or =25 micromol/L almost completely abolished the increased CAM protein and mRNA regardless of their anti-oxidative activity. With the exception of the flavonol quercetin, flavonoids had no such effect; quercetin substantially attenuated the CAM induction. The flavones inhibited nuclear translocation and DNA binding activity of the NF-kappa B-containing binding site in the promoter region of the CAM genes in TNF-alpha-activated HUVEC. The inhibition of endothelial CAM induction by flavones is mediated by their interference with the NF-kappa B-dependent transcription pathway. Thus, the flavones may hamper initial atherosclerotic events involving endothelial CAM induction.

Oxidative injury induces cellular and nuclear damage that leads to apoptotic cell death. Agents or antioxidants that can inhibit production of reactive oxygen species can prevent apoptosis. We tested the hypothesis that flavonoids can inhibit H 2 0 2 -induced apoptosis in human umbilical vein endothelial cells. A 30-min pulse treatment with 0.25 mmol/L H 2 0 2 decreased endothelial cell viability within 24 h by ∼40% (P < 0.05) with distinct nuclear condensation and DNA fragmentation. In the H 2 0 2 apoptosis model, the addition of 50 μmol/L of the flavanol (-)epigallocatechin gallate and the flavonol quercetin, which have in vitro radical scavenging activity, partially (P < 0.05) restored cell viability with a reduction in H 2 0 2 -induced apoptotic DNA damage. In contrast, the flavones, luteolin and apigenin, at the nontoxic dose of 50 μmol/L, intensified cell loss (P < 0.05) after exposure to H 2 0 2 and did not protect cells from oxidant-induced apoptosis. The flavanones, hesperidin and naringin, did not have cytoprotective effects. The antioxidants, (-)epigallocatechin gallate and quercetin, inhibited endothelial apoptosis, enhanced the expression of bcl-2 protein and inhibited the expression of bax protein and the cleavage and activation of caspase-3. Therefore, flavanols and flavonols, in particular (-)epigallocatechin gallate and quercetin, qualify as potent antioxidants and are effective in preventing endothelial apoptosis caused by oxidants, suggesting that flavonoids have differential antiapoptotic efficacies. The antiapoptotic activity of flavonoids appears to be mediated at the mitochondrial bcl-2 and bax gene level. J. Nutr. 133: 985-991, 2003.

Fundamental study of enzymatic nucleoside transport suffers for lack of optical probes that can be tracked noninvasively. Nucleoside transporters are integral membrane glycoproteins that mediate the salvage of nucleosides and their passage across cell membranes. The substrate recognition site is the deoxyribose sugar, often with little distinction among nucleobases. Reported here are nucleoside analogues in which emissive, cyclometalated iridium(III) complexes are "clicked" to C-1 of deoxyribose in place of canonical nucleobases. The resulting complexes show visible luminescence at room temperature and 77 K with microsecond-length triplet lifetimes. A representative complex is crystallographically characterized. Transport and luminescence are demonstrated in cultured human carcinoma (KB3-1) cells.

Despite the significance of Alzheimer’s disease, the link between metal-associated amyloid-β (metal–Aβ) and disease etiology remains unclear. To elucidate this relationship, chemical tools capable of specifically targeting and modulating metal–Aβ species are necessary, along with a fundamental understanding of their mechanism at the molecular level. Herein, we investigated and compared the interactions and reactivities of the green tea extract, (−)-epigallocatechin-3-gallate [(2 R ,3 R )-5,7-dihydroxy-2-(3,4,5-trihydroxyphenyl)-3,4-dihydro-2 H -1-benzopyran-3-yl 3,4,5-trihydroxybenzoate; EGCG], with metal [Cu(II) and Zn(II)]–Aβ and metal-free Aβ species. We found that EGCG interacted with metal–Aβ species and formed small, unstructured Aβ aggregates more noticeably than in metal-free conditions in vitro. In addition, upon incubation with EGCG, the toxicity presented by metal-free Aβ and metal–Aβ was mitigated in living cells. To understand this reactivity at the molecular level, structural insights were obtained by ion mobility-mass spectrometry (IM-MS), 2D NMR spectroscopy, and computational methods. These studies indicated that ( i ) EGCG was bound to Aβ monomers and dimers, generating more compact peptide conformations than those from EGCG-untreated Aβ species; and ( ii ) ternary EGCG–metal–Aβ complexes were produced. Thus, we demonstrate the distinct antiamyloidogenic reactivity of EGCG toward metal–Aβ species with a structure-based mechanism.

Nucleoside transporters play important physiological roles by regulating intra- and extra-cellular concentrations of purine and pyrimidine (deoxy)nucleosides. This review describes the biological function and activity of the two major families of membrane nucleoside transporters that exist in mammalian cells. These include equilibrative nucleoside transporters that transport nucleosides in a gradient-dependent fashion and concentrative nucleoside transporters that import nucleosides against a gradient by coupling movement with sodium transport. Particular emphasis is placed on describing the roles of nucleoside transport in normal physiological processes, including inflammation, cardiovascular function and nutrient transport across the blood–brain barrier. In addition, the role of nucleoside transport in pathological conditions such as cardiovascular disease and cancer are discussed. The potential therapeutic applications of manipulating nucleoside transport activities are discussed, focusing on nucleoside analogs as anti-neoplastic agents. Finally, we discuss future directions for the development of novel chemical entities to measure nucleoside transport activity at the cellular and organismal level.

Amyloid-β (Aβ) peptides and their metal-associated aggregated states have been implicated in the pathogenesis of Alzheimer's disease (AD). Although the etiology of AD remains uncertain, understanding the role of metal-Aβ species could provide insights into the onset and development of the disease. To unravel this, bifunctional small molecules that can specifically target and modulate metal-Aβ species have been developed, which could serve as suitable chemical tools for investigating metal-Aβ-associated events in AD. Through a rational structure-based design principle involving the incorporation of a metal binding site into the structure of an Aβ interacting molecule, we devised stilbene derivatives (L1-a and L1-b) and demonstrated their reactivity toward metal-Aβ species. In particular, the dual functions of compounds with different structural features (e.g., with or without a dimethylamino group) were explored by UV-vis, X-ray crystallography, high-resolution 2D NMR, and docking studies. Enhanced bifunctionality of compounds provided greater effects on metal-induced Aβ aggregation and neurotoxicity in vitro and in living cells. Mechanistic investigations of the reaction of L1-a and L1-b with Zn(2+)-Aβ species by UV-vis and 2D NMR suggest that metal chelation with ligand and/or metal-ligand interaction with the Aβ peptide may be driving factors for the observed modulation of metal-Aβ aggregation pathways. Overall, the studies presented herein demonstrate the importance of a structure-interaction-reactivity relationship for designing small molecules to target metal-Aβ species allowing for the modulation of metal-induced Aβ reactivity and neurotoxicity.

Metal ions associated with amyloid-β (Aβ) species have been suggested to be involved in neurodegeneration leading to the progression of Alzheimer's disease (AD). The role of metal-involved Aβ species in AD neuropathogenesis, however, is not fully elucidated. In order to advance this understanding and contribute to the therapeutic development for AD, the rational structure-based design of small molecules that specifically target metal ions surrounded by Aβ species has recently received increased attention. To date, only a few compounds have been fashioned for this purpose. Herein, we report the design strategy, synthesis, characterization, and reactivity of new bifunctional IMPY derivatives K1 and K2. Using UV-vis and high-resolution two-dimensional (2D) NMR spectroscopy, the bifunctionality of K1 and K2 (metal chelation and Aβ interaction) was confirmed. These bifunctional IMPY derivatives showed preferential reactivity toward metal-induced Aβ aggregation over metal-free conditions in both in vitro inhibition and disaggregation experiments. Taken together, this study provides another example of a bifunctional small molecule framework that can target metal ions associated with Aβ species.

Ellagic acid, a polyphenol compound present in berries and pomegranate, has received attention as an agent that may have potential bioactivities preventing chronic diseases. This study examined photoprotective effects of ellagic acid on collagen breakdown and inflammatory responses in UV (ultraviolet)-B irradiated human skin cells and hairless mice. Ellagic acid attenuated the UV-B-induced toxicity of HaCaT keratinocytes and human dermal fibroblasts. Non-toxic ellagic acid markedly prevented collagen degradation by blocking matrix metalloproteinase production in UV-B-exposed fibroblasts. Anti-wrinkle activity of ellagic acid was further investigated in hairless mice exposed to UV-B, in which it attenuated UV-B-triggered skin wrinkle formation and epidermal thickening. Topical application of 10 micromol/l ellagic acid diminished production of pro-inflammatory cytokines IL-1beta and IL-6, and blocked infiltration of inflammatory macrophages in the integuments of SKH-1 hairless mice exposed to UV-B for 8 weeks. In addition, this compound mitigated inflammatory intracellular cell adhesion molecule-1 expression in UV-B-irradiated keratinocytes and photoaged mouse epidermis. These results demonstrate that ellagic acid prevented collagen destruction and inflammatory responses caused by UV-B. Therefore, dietary and pharmacological interventions with berries rich in ellagic acid may be promising treatment strategies interrupting skin wrinkle and inflammation associated with chronic UV exposure leading to photoageing.

Fruits of bog blueberry (Vaccinium uliginosum L.) are rich in anthocyanins that contribute pigmentation. Anthocyanins have received much attention as agents with potentials preventing chronic diseases. This study investigated the capacity of anthocyanin-rich extract from bog blueberry (ATH-BBe) to inhibit photoaging in UV-B-irradiated human dermal fibroblasts. BBe anthocyanins were detected as cyanidin-3-glucoside, petunidin-3-glucoside, malvidin-3-glucoside, and delphinidin3-glucoside. ATH-BBe attenuated UV-B-induced toxicity accompanying reactive oxygen species (ROS) production and the resultant DNA damage responsible for activation of p53 and Bad. Preincubation of ATH-BBe markedly suppressed collagen degradation via blunting production of collagenolytic matrix metalloproteinases (MMP). Additionally, ATH-BBe enhanced UV-B-downregulated procollagen expression at transcriptional levels. We next attempted to explore whether ATH-BBe mitigated the MMP-promoted collagen degradation through blocking nuclear factor kappaB (NF-kappaB) activation and MAPK-signaling cascades. UV-B radiation enhanced nuclear translocation of NF-kappaB, which was reversed by treatment with ATH-BBe. The UV-B irradiation rapidly activated apoptosis signal-regulating kinase-1 (ASK-1)-signaling cascades of JNK and p38 mitogen-activated protein kinase (p38 MAPK), whereas ATH-BBe hampered phosphorylation of c-Jun, p53, and signal transducers and activators of transcription-1 (STAT-1) linked to these MAPK signaling pathways. ATH-BBe diminished UV-B augmented-release of inflammatory interleukin (IL)-6 and IL-8. These results demonstrate that ATH-BBe dampens UV-B-triggered collagen destruction and inflammatory responses through modulating NF-kappaB-responsive and MAPK-dependent pathways. Therefore, anthocyanins from edible bog blueberry may be protective against UV-induced skin photoaging.

Oxidized LDL is highly atherogenic, as it stimulates foam cell formation and promotes inflammatory and thrombotic processes. The present study elucidated whether the antioxidants quercetin and its rutinoside rutin exert antiapoptosis in endothelial cells exposed to Cu(2+)-oxidized LDL. Quercetin and rutin inhibited the oxidized LDL-induced endothelial toxicity at nontoxic doses of/=25 muM with an inhibition of intracellular oxidant accumulation. These effects accompanied disappearance of apoptotic bodies and suppression of caspase-3 activation. Additionally, condensed nuclei vanished in oxidized LDL-exposed cells treated with quercetin and rutin. This study further explored whether such effects were achieved by redox manipulation via JAK2-STAT3-responsive death/survival signaling pathways involving multiple MAPK. Unlike quercetin, rutin blocked the activation of oxidized LDL-induced JNK and p38 MAPK as well as the upstream ASK1 phosphorylation. Quercetin dose-dependently attenuated the JAK2 phosphorylation evoked by oxidized LDL, whereas rutin abolished the JAK signaling accompanying nuclear transactivation of STAT3 and enhanced the JAK activity-inhibiting SOCS3 expression. Conversely, oxidized LDL-induced IL-6 release was minimal for the JAK2 activation, although this effect was counteracted by quercetin and rutin. These results suggest that quercetin and rutin inhibit Cu(2+)-oxidized LDL-induced endothelial apoptosis through modulating JAK2-STAT3 pathways and that rutin may modulate a signaling crosstalk between JAK2 and MAPK.

Diabetes can cause a wide variety of vascular complications and endothelial dysfunction. In this study, human vascular endothelial cells were exposed to 5.5 mM and 33 mM glucose for 5 d in the absence and presence of 1 to 20 mug/mL roasted licorice (Glycyrrhiza inflata Bat.) ethanol extracts (rLE). Caspase-3 activation and Annexin V staining revealed that high glucose induced endothelial apoptotic toxicity with a generation of reactive oxygen species (ROS) and these effects were reversed by rLE at >/=1 mug/mL in a dose-dependent manner. Cytoprotective rLE substantially reduced high glucose-induced expression of endothelial nitric oxide synthase (eNOS), and hence attenuated the formation of peroxynitrite radicals derived from NO. In addition, rLE suppressed expression of PKCbeta2 and activation of NADPH oxidase subunit of p22phox promoted by high glucose. However, rLE

Hyperglycemia is a causal factor in the development of diabetic vascular complications including impaired vascular smooth muscle contractility and increased cell proliferation. The present study was designed to investigate the effects of Sasa borealis water-extract (SBwE) on chronic hyperglycemia-induced oxidative stress and apoptosis in human umbilical endothelial cells (HUVEC). HUVEC were cultured in 5.5 m M low glucose, 5.5 m M glucose plus 27.5 m M mannitol as an osmotic control, or 33 m M high glucose for 5 days in the absence and presence of 1–30 μg/ ml SBwE. Caspase-3 activation and Annexin V staining revealed chronic high glucose–induced endothelial apoptotic toxicity with a generation of oxidants detected by DCF-fluorescence, and these effects were reversed by SBwE at ≥1 μg/ml in a dose-dependent manner. Cytoprotective SBwE substantially reduced the sustained high glucose–induced expression of endothelial nitric oxide synthase and attenuated the formation of peroxynitrite radicals. The suppressive effects of SBwE were most likely mediated through blunting activation of PKCβ2 and NADPH oxidase promoted by high glucose. In addition, this bamboo extract modulated the high glucose–triggered mitogen-activated protein kinase–dependent upregulation of heat-shock proteins. Our results suggest that SBwE suppressed these detrimental effects caused by PKC-dependent peroxynitrite formation via activation of NADPH oxidase and induction of nitric oxide synthase and heat-shock protein family that may be essential mechanisms responsible for increased apoptotic oxidative stress in diabetic vascular complications. Moreover, the blockade of high glucose–elicited heat-shock protein induction appeared to be responsible for SBwE-alleviated endothelial apoptosis. Therefore, SBwE may be a therapeutic agent for the prevention and treatment of diabetic endothelial dysfunction and related complications.

Oxidative modification of LDL is causally involved in the development of atherosclerosis and occursin vivoin the blood as well as within the vascular wall. The present study attempted to explore whether polyphenolic flavonoids influence monocyte-endothelium interaction and lectin-like oxidised LDL receptor 1 (LOX-1) expression involved in the early development of atherosclerosis. The flavones luteolin and apigenin inhibited THP-1 cell adhesion onto oxidised LDL-activated human umbilical vein endothelial cells (HUVEC), while the flavanols of ( − )epigallocatechin gallate and (+)catechin, the flavonols of quercetin and rutin, and the flavanones of naringin, naringenin, hesperidin and hesperetin did not have such effects. Consistently, Western blot analysis revealed that the flavones at 25 μmdramatically and significantly abolished HUVEC expression of vascular cell adhesion molecule-1 and E-selectin evidently enhanced by oxidised LDL; these inhibitory effects were exerted by drastically down regulating mRNA levels of these cell adhesion molecules. In addition, quercetin and luteolin significantly attenuated expression of LOX-1 protein up regulated in oxidised LDL-activated HUVEC with a fall in transcriptional mRNA levels of LOX-1. In addition, quercetin and luteolin clearly blunted oxidised LDL uptake by HUVEC treated with oxidised LDL. The results demonstrate that the flavones luteolin and apigenin as well as quercetin were effective in the different initial steps of atherosclerosis process by inhibiting oxidised LDL-induced endothelial monocyte adhesion and/or oxidised LDL uptake. Therefore, certain flavonoids qualify as anti-atherogenic agents in LDL systems, which may have implications for strategies attenuating endothelial dysfunction-related atherosclerosis.

Numerous polyphenolic compounds have been found to inhibit adhesion and migration of leukocytes to sites of inflammation that are partly regulated by the expression of cell adhesion molecules (CAM) such as vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and platelet endothelial cell adhesion molecule-1 (PECAM-1). Licorice root extracts have been used in traditional Chinese, Tibetan, and Indian medicine for the treatment of pulmonary diseases and inflammatory processes. Expression of CAM proteins was examined in human umbilical vein endothelial cells (HUVEC) treated with a licorice component (isoliquiritigenin, 18beta-glycyrrhetinic acid, glycyrrhizin, formononetin, or ononin) and exposed to TNF-alpha. The involvement of NF-kappaB in the transcriptional control of CAM proteins was assessed by degradation of IkappaBalpha and nuclear translocation of NF-kappaB using Western blotting techniques and immunocytochemical staining. At nontoxicor =10 microM, isoliquiritigenin blocked the induction of VCAM-1 and E-selectin on activated HUVEC and markedly interfered with THP-1 monocyte adhesion to TNF-alpha-activated endothelial cells. Isoliquiritigenin abolished TNF-alpha-induced mRNA accumulation of VCAM-1 and E-selectin. Additionally, immunocytochemical staining revealed that isoliquiritigenin attenuated PECAM-1 expression induced by TNF-alpha. In contrast, other components recognized in licorice, 18beta-glycyrrhetinic acid, glycyrrhizin, formononetin, and ononin did not down-regulate the expression of VCAM-1 and/or PECAM-1 activated by TNF-alpha, implying that these components are inactive in modulating adhesion of leukocytes to stimulated endothelial cells. Isoliquiritigenin downregulated CAM proteins in TNF-alpha-activated HUVEC at the transcriptional levels by blocking degradation of IkappaBalpha and nuclear translocation of NF-kappaB. These results demonstrate that the induction blockade of VCAM-1 and E-selectin by isoliquiritigenin was directly mediated by its interference with the CAM mRNA transcription through NF-kappaB-dependent mechanisms under inflammatory conditions.