Vince Castranova

New Insights into the Role of Nuclear Factor-κB in Cell Growth Regulation

The nuclear factor (NF)-κB family of eukaryotic transcription factors plays an important role in the regulation of immune response, embryo and cell lineage development, cell apoptosis, cell-cycle progression, inflammation, and oncogenesis. A wide range of stimuli, including cytokines, mitogens, environmental particles, toxic metals, and viral or bacterial products, activate NF-κB, mostly through IκB kinase (IKK)-dependent phosphorylation and subsequent degradation of its inhibitor, the IκB family of proteins. Activated NF-κB translocates into the nucleus where it modulates the expression of a variety of genes, including those encoding cytokines, growth factors, acute phase response proteins, cell adhesion molecules, other transcription factors, and several cell apoptosis regulators. During the past few years, tremendous progress has been achieved in our understanding on how intracellular signaling pathways are transmitted in either a linear or a network manner leading to the activation of NF-κB and subsequent cell growth control. However, a detailed molecular mechanism of NF-κB regulating cell growth has yet to be determined. Elucidation of the relationships between NF-κB activation and cell growth will be important in developing new strategies for the treatment of various human diseases, such as chronic autoimmune disorder and cancer.

Reactive oxygen species and silica-induced carcinogenesis.

Although silica has recently been designated as a carcinogen, its mechanism of carcinogenesis is not fully understood. Recent studies suggest that free-radical reactions may play an important role in the initiation and progression of cancer. This article summarizes literature on the generation of reactive oxygen species (ROS) directly from silica and from silica-stimulated cells. It also summarizes information concerning the role of ROS in silica-induced DNA damage as well as in silica-induced cell proliferation, including the effects of silica on the activation of nuclear transcription factors, induction of growth factors and oncogene expression, redox regulation of the p53 tumor suppressor gene, induction of apoptosis, and division of damaged cells. Understanding the role of ROS in silica-mediated reactions may help develop therapeutic agents to block silica-induced free radical reactions and thus prevent or attenuate silica-induced carcinogenesis.

Role of reactive oxygen species and MAPKs in vanadate-induced G2/M phase arrest

Cell growth arrest is an important mechanism in maintaining genomic stability and integrity in response to environmental stress. Using the human lung alveolar epithelial cancer cell line A549, we investigated the role of reactive oxygen species (ROS), extracellular signal-regulated protein kinase (ERK), and p38 protein kinase in vanadate-induced cell growth arrest. Exposure of cells to vanadate led to cell growth arrest at the G(2)/M phase and caused upregulation of p21 and phospho-cdc2 and degradation of cdc25C in a time- and dose-dependent manner. Vanadate stimulated mitogen-activated protein kinases (MAPKs) family members, as determined by the phosphorylation of ERK and p38. PD98059, an inhibitor of ERK, and SB202190, an inhibitor of p38, inhibited vanadate-induced cell growth arrest, upregulation of p21 and cdc2, and degradation of cdc25C. In addition to hydroxyl radical ((*)OH) formation, cellular reduction of vanadate generated superoxide radical (O(2)(*)(-)) and hydrogen peroxide (H(2)O(2)), as determined by confocal microscopy using specific dyes. Generation of O(2)(*)(-) and H(2)O(2) was inhibited by specific antioxidant enzymes, superoxide dismutase (SOD) and catalase, respectively. ROS activate ERK and p38, which in turn upregulate p21 and cdc2 and cause degradation of cdc25C, leading to cell growth arrest at the G(2)/M phase. Specific ROS affect different MAPK family members and cell growth regulatory proteins with different potencies.

Molecular mechanism of tumor necrosis factor-alpha production in 1-->3-beta-glucan (zymosan)-activated macrophages.

The molecular details of 1→3-β-glucans, a fungal cell wall component, induced inflammatory responses are not well understood. In the present study, we conducted a systematic analysis of the molecular events leading to tumor necrosis factor (TNF)-α production after glucan stimulation of macrophages. We demonstrated that activation of nuclear factor κB (NF-κB) is essential in zymosan A (a source of 1→3-β-glucans)-induced TNF-α production in macrophages (RAW264.7 cells). Zymosan A-induced TNF-α protein production was associated with an increase in the TNF-α gene promoter activity. Activation of the TNF-α gene promoter was dependent on activation of NF-κB. Time course studies indicated that DNA binding activity of NF-κB preceded TNF-α promoter activity. Inhibition of NF-κB activation led to a dramatic reduction in both TNF-α promoter activity and TNF-α protein production in the response to zymosan A. Mutation of a major NF-κB binding site (κ3) in the gene promoter resulted in a significant decrease in the induction of the gene promoter by zymosan A, while mutation of Egr or CRE sites failed to inhibit the response to zymosan. Together, these results strongly suggest that NF-κB is involved in signal transduction of 1→3-β-glucans-induced TNF-α expression.

Antioxidant properties of aspirin: Characterization of the ability of aspirin to inhibit silica-induced lipid peroxidation, DNA damage, NF-κB activation, and TNF-α production

Electron spin resonance (ESR) was used to investigate the reaction of aspirin toward reactive oxygen species, such as hydroxyl radicals (·OH), superoxide radicals (O2-) and H2O2. The Fenton reaction (Fe(II) + H2O2 ---> FE(III) + -OH + OR) was used as a source of -OH radicals. The results show that aspirin is an efficient -OH radical scavenger with a reaction rate constant of k = 3.6 x 1010 M-1sec-1, which is faster than several well established antioxidants, such as ascorbate, glutathione and cysteine. However, aspirin is not a good scavenger for O2- or H2O2. Through its antioxidant property, aspirin exhibited a protective effect against silica-induced lipid peroxidation and DNA strand breakage. Aspirin also inhibited the activation of nuclear transcription factor-κb induced by silica, lipopolysaccharide or the transition metal, Fe(II), as demonstrated by electrophoretic mobility shift assay. The results show that aspirin functions as an antioxidant via its ability to scavenge -OH radicals. This antioxidant property may explain some of its various physiological and pharmacological actions.

Single- and Multi-Wall Carbon Nanotubes Versus Asbestos: Are the Carbon Nanotubes a New Health Risk to Humans?

Carbon nanotubes (CNT), since their discovery, have become one of the most promising nanomaterials in many industrial and biomedical applications. Due to their unique physicochemical properties, interest is growing in the manufacture of CNT-based products and their subsequent marketing. Since their discovery, the prospect of possible undesirable human health effects has been a focus of many scientific studies. Although CNT possess unique physical properties that include (1) nanoscale diameter, (2) a wide length distribution ranging from tens of nanometers to several micrometers, and (3) high aspect ratio, the fibrous-like shape and durability suggest that their toxic properties may be analogous to those observed with other fibrous particles, such as asbestos. The present study provides a summary of published findings on CNT bioactivity, such as the potential of CNT, especially of multi-wall carbon nanotubes (MWCNT), to activate signaling pathways modulating transcription factor activity, induce apoptosis, induce DNA damage, and initiate biological responses. Assessment of risks to human health and adoption of appropriate exposure controls is critical for the safe and successful introduction of CNT -based products for future applications.

Cobalt-mediated generation of reactive oxygen species and its possible mechanism

Electron spin resonance spin trapping was utilized to investigate free radical generation from cobalt (Co) mediated reactions using 5,5-dimethyl-1-pyrroline (DMPO) as a spin trap. A mixture of Co with water in the presence of DMPO generated 5,5-dimethylpyrroline-(2)-oxy(1) DMPOX, indicating the production of strong oxidants. Addition of superoxide dismutase (SOD) to the mixture produced hydroxyl radical (.OH). Catalase eliminated the generation of this radical and metal chelators, such as desferoxamine, diethylenetriaminepentaacetic acid or 1,10-phenanthroline, decreased it. Addition of Fe(II) resulted in a several fold increase in the .OH generation. UV and O2 consumption measurements showed that the reaction of Co with water consumed molecular oxygen and generated Co(II). Since reaction of Co(II) with H2O2 did not generate any significant amount of .OH radicals, a Co(I) mediated Fenton-like reaction [Co(I) + H2O2-->Co(II) + .OH + OH-] seems responsible for .OH generation. H2O2 is produced from O2.- via dismutation, O2.- is produced by one-electron reduction of molecular oxygen catalyzed by Co. Chelation of Co(II) by biological chelators, such as glutathione or beta-ananyl-3-methyl-L-histidine alters, its oxidation-reduction potential and makes Co(II) capable of generating .OH via a Co(II)-mediated Fenton-like reaction [Co(II) + H2O2-->Co(III) + .OH + OH-]. Thus, the reaction of Co with water, especially in the presence of biological chelators, glutathione, glycylglycylhistidine and beta-ananyl-3-methyl-L-histidine, is capable of generating a whole spectrum of reactive oxygen species, which may be responsible for Co-induced cell injury.

Sustained JNK1 activation is associated with altered histone H3 methylations in human liver cancer.

BACKGROUND/AIMS Aberrant c-Jun N-terminal kinase (JNK) activation has been linked to hepatocellular carcinoma (HCC) in mouse models. It remains unclear whether JNK activation plays an important role in human HCC and, if so, how JNK signaling contributes to the initiation or progression of HCC. METHODS The JNK activation, global gene expression, and the status of histone H3 methylations were measured in 31 primary human hepatocellular carcinoma (HCC) samples paired with the adjacent non-cancerous (ANC) tissues. RESULTS Enhanced JNK1 activation was noted in 17 out of 31 HCC samples (55%) relative to the corresponding ANC tissues, whereas JNK2 activation was roughly equal between HCC and ANC tissues. This enhancement in JNK1 activation is associated with an increased tumor size and a lack of encapsulation of the tumors. In addition, an association of JNK1 activation with the histone H3 lysines 4 and 9 tri-methylation was observed in the HCC tissues, which leads to an elevated expression of genes regulating cell growth and a decreased expression of the genes for cell differentiation and the p450 family members in HCC. CONCLUSIONS These results, thus, suggest that JNK1 plays important roles in the development of human HCC partially through the epigenetic mechanisms.

New perspectives in arsenic-induced cell signal transduction

Although the carcinogenicity of arsenic has been well established, the underlying molecular mechanisms have not yet been fully identified. Accumulating evidence indicates that the alteration of cellular signal transduction is directly related to the carcinogenesis of arsenic. This review focuses on recent advances in arsenic-induced signal transduction, including reactive oxygen species (ROS) production, tyrosine phosphorylation, MAPK signaling, NF-kappaB activation, cell cycle arrest, and apoptosis.

Effect of chlorogenic acid on hydroxyl radical

Chlorogenic acid (CGA) is considered to act as an antioxidant. However, the inhibitory effects of CGA on specific radical species are not well understood. Electron spin resonance (ESR) in combination with spin trapping techniques was utilized to detect free radicals. 5,5-Dimethyl-1-pyrroline-N-oxide (DMPO) was used as a spin trapping reagent while the Fenton reaction was used as a source of hydroxyl radical (·OH). We found that CGA scavenges ·OH in a dose-dependent manner. The kinetic parameters, IC50 and Vmax, for CGA scavenging of ·OH were 110 and 1.27 μM/sec, respectively. The rate constant for the scavenging of ·OH by CGA was 7.73 × 109 M−1 sec−1. Our studies suggest that the antioxidant properties of CGA may involve a direct scavenging effect of CGA on ·OH.