Publisher Summary This chapter discusses the role of free radicals and catalytic metal ions in human disease. The importance of transition metal ions in mediating oxidant damage naturally leads to the question as to what forms of such ions might be available to catalyze radical reactions in vivo . The chapter discusses the metabolism of transition metals, such as iron and copper. It also discusses the chelation therapy that is an approach to site-specific antioxidant protection. The detection and measurement of lipid peroxidation is the evidence most frequently cited to support the involvement of free radical reactions in toxicology and in human disease. A wide range of techniques is available to measure the rate of this process, but none is applicable to all circumstances. The two most popular are the measurement of diene conjugation and the thiobarbituric acid (TBA) test, but they are both subject to pitfalls, especially when applied to human samples. The chapter also discusses the essential principles of the peroxidation process. When discussing lipid peroxidation, it is essential to use clear terminology for the sequence of events involved; an imprecise use of terms such as initiation has caused considerable confusion in the literature. In a completely peroxide-free lipid system, first chain initiation of a peroxidation sequence in a membrane or polyunsaturated fatty acid refers to the attack of any species that has sufficient reactivity to abstract a hydrogen atom from a methylene group.

Role of free radicals and catalytic metal ions in human disease: an overview.

Flavonoids are nearly ubiquitous in plants and are recognized as the pigments responsible for the colors of leaves, especially in autumn. They are rich in seeds, citrus fruits, olive oil, tea, and red wine. They are low molecular weight compounds composed of a three-ring structure with various substitutions. This basic structure is shared by tocopherols (vitamin E). Flavonoids can be subdivided according to the presence of an oxy group at position 4, a double bond between carbon atoms 2 and 3, or a hydroxyl group in position 3 of the C (middle) ring. These characteristics appear to also be required for best activity, especially antioxidant and antiproliferative, in the systems studied. The particular hydroxylation pattern of the B ring of the flavonoles increases their activities, especially in inhibition of mast cell secretion. Certain plants and spices containing flavonoids have been used for thousands of years in traditional Eastern medicine. In spite of the voluminous literature available, however, Western medicine has not yet used flavonoids therapeutically, even though their safety record is exceptional. Suggestions are made where such possibilities may be worth pursuing.

The Effects of Plant Flavonoids on Mammalian Cells:Implications for Inflammation, Heart Disease, and Cancer

If carcinogenesis occurs by somatic evolution, then common components of the cancer phenotype result from active selection and must, therefore, confer a significant growth advantage. A near-universal property of primary and metastatic cancers is upregulation of glycolysis, resulting in increased glucose consumption, which can be observed with clinical tumour imaging. We propose that persistent metabolism of glucose to lactate even in aerobic conditions is an adaptation to intermittent hypoxia in pre-malignant lesions. However, upregulation of glycolysis leads to microenvironmental acidosis requiring evolution to phenotypes resistant to acid-induced cell toxicity. Subsequent cell populations with upregulated glycolysis and acid resistance have a powerful growth advantage, which promotes unconstrained proliferation and invasion.

Why do cancers have high aerobic glycolysis

Phenolics are broadly distributed in the plant kingdom and are the most abundant secondary metabolites of plants. Plant polyphenols have drawn increasing attention due to their potent antioxidant properties and their marked effects in the prevention of various oxidative stress associated diseases such as cancer. In the last few years, the identification and development of phenolic compounds or extracts from different plants has become a major area of health- and medical-related research. This review provides an updated and comprehensive overview on phenolic extraction, purification, analysis and quantification as well as their antioxidant properties. Furthermore, the anticancer effects of phenolics in-vitro and in-vivo animal models are viewed, including recent human intervention studies. Finally, possible mechanisms of action involving antioxidant and pro-oxidant activity as well as interference with cellular functions are discussed.

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Plant phenolics: extraction, analysis and their antioxidant and anticancer properties.

The relationship between antioxidant activity and antimutagenicity of various tea extracts (green tea, pouchong tea, oolong tea, and black tea) was investigated. All tea extracts exhibited markedly antioxidant activity and reducing power, especially oolong tea, which inhibited 73.6% peroxidation of linoleic acid. Tea extracts exhibited a 65-75% scavenging effect on superoxide at a dose of 1 mg and 30 - 60% scavenging effect on hydrogen peroxide at a dose of 400 microgram. They scavenged 100% hydroxyl radical at a dosage of 4 mg except the black tea. Tea extracts also showed 50 - 70% scavenging effect on alpha, alpha-diphenyl-beta-picrylhydrazyl radical. The antioxidant activity and the scavenging effects on active oxygen decreased in the order semifermented tea > nonfermented tea > fermented tea. Tea extracts showed strong antimutagenic action against five indirect mutagens, i.e., AFB1, Trp-P-1, Glu-P-1, B[a]P, and IQ, especially oolong and pouchong teas. The antioxidant effect of tea extracts was well correlated to their antimutagenicity in some cases but varied with the mutagen and antioxidative properties.

Antioxidant activity of various tea extracts in relation to their antimutagenicity

The plasma-membrane association and transforming activity of the ras oncoprotein p21 are dependent upon posttranslational farnesylation. Farnesyl synthesis and p21 ras farnesylation are inhibited by hydroxymethylglutaryl-CoA reductase inhibitors such as lovastatin. In this study, we examined whether lovastatin could reverse the transformed phenotype of a v-Ha-ras-transformed rat liver epithelial cell line (WB-ras cells) and if changes were associated with the enhancement of gap-junctional intercellular communication (GJIC). WB-ras cells grow in soft agar, have reduced GJIC, and are highly tumorigenic. Membrane association of p21 ras in these cells was inhibited after in vitro treatment with lovastatin (0.1-0.5 microM) for 48 h. Concomitantly, the cells displayed a more normal morphology, decreased growth in soft agar, and enhanced GJIC. These changes were prevented by cotreatment with mevalonic acid. The morphology and GJIC of rat liver epithelial cells transformed with other oncogenes (src, neu, and raf/myc) were not affected by lovastatin. Intrahepatic WB-ras tumors were induced in male rats by intraportal-vein injection of WB-ras cells. The size and DNA labeling index of these tumors were decreased approximately 75% by administration of lovastatin (5 mg/kg orally twice daily for 2 wk). These results suggest that lovastatin reversed the transformed phenotype of WB-ras cells by inhibiting p21 ras plasma membrane association. Furthermore, the concomitant enhancement of GJIC in lovastatin-treated cells suggests a role for reduced GJIC in the expression of the transformed phenotype.

Reversal of ras-induced inhibition of gap-junctional intercellular communication, transformation, and tumorigenesis by lovastatin.

Gap junction-mediated intercellular communication in untreated and phenobarbital-treated C57BL/6 × C3H F1 mouse hepatocytes was evaluated by microinjection of fluorescent Lucifer Yellow CH dye Intercellular communication (dye coupling) was detected in untreated hepatocytes after 05 h in culture, reached a maximum level in 24- and 48-h-old cultures (852%), and then decreased over the next 72 h Phenobarbital (20–500 µg/ml) decreased dye coupling in a dose-related manner when added to freshly plated cultures This inhibitory effect was evident during 05–12 h of treatment but was not seen in cultures treated for 24 h Phenobarbital also decreased dye coupling within 30 min when added to established (24-h-old) hepatocyte cultures This effect was maximal after 2 h treatment In these cultures, dye coupling recovered within 15 min after removal of the promoter Hepatocytes, pretreated with phenobarbital for 24 h, did not show inhibition of dye coupling after reapplication of phenobarbital Thus, phenobarbital inhibited mouse hepatocyte dye coupling rapidly and reversibly, and the cells became refractory to the inhibitory effect after prolonged treatment

https://cancerres.aacrjournals.org/content/canres/48/9/2519.full.pdf

Kinetics of Phenobarbital Inhibition of Intercellular Communication in Mouse Hepatocytes

The role of altered ras oncoprotein (Ras) farnesylation and membrane association in the growth inhibitory effects of several monoterpenes (limonene, perillic acid, perillyl alcohol, menthol, pinene and cineole) was investigated in rat liver epithelial cells. All of the above compounds except cineole inhibited the growth of viral Ha-ras-transformed rat liver epithelial cells (WB-ras cells) at concentrations of 0.25-2.5 mM. These cells, however, were not necessarily more sensitive to these compounds compared to non-transformed and viral raf-transformed rat liver epithelial cells. Growth inhibition by limonene, perillic acid and pinene was only partially restored (20-50%) by supplementing the culture medium with 2 mM mevalonic acid. Western blot analyses of cytosolic and membranous fractions of WB-ras cells treated with monoterpenes indicated no change in Ras distribution. In contrast, lovastatin, a potent inhibitor of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase and Ras farnesylation, specifically reduced WB-ras cell growth and increased cytosolic levels of Ras. Thus, monoterpene-induced growth inhibition of rat liver epithelial cells was dissimilar to lovastatin and did not appear to involve altered Ras plasma membrane association.

Randall J. Ruch

Papers

Reversal of ras-induced inhibition of gap-junctional intercellular communication, transformation, and tumorigenesis by lovastatin.

Kinetics of Phenobarbital Inhibition of Intercellular Communication in Mouse Hepatocytes

Growth inhibition of rat liver epithelial tumor cells by monoterpenes does not involve Ras plasma membrane association.

Strain and species effects on the inhibition of hepatocyte intercellular communication by liver tumor promoters.

Comparative effects of phenobarbital, DDT, and lindane on mouse hepatocyte gap junctional intercellular communication.