Gabriella Calviello

Dietary supplementation with eicosapentaenoic and docosahexaenoic acid inhibits growth of Morris hepatocarcinoma 3924A in rats: effects on proliferation and apoptosis

The effect of individual administration of low doses of highly purified eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (1 g/kg body weight) on the growth of Morris hepatocarcinoma 3924A transplanted in ACI/T rats was investigated. Both EPA and DHA inhibited growth of the hepatocarcinoma (50% reduction of tumor weight or volume at the 19th day after transplantation for both of the n‐3 PUFA groups). EPA treatment reduced the percentage of proliferating tumor cells labeled with BUdR (10‐fold), whereas DHA did not. Conversely, DHA supplementation induced a doubling of the number of cells undergoing apoptosis (labeled by TUNEL), whereas EPA treatment was much less effective. Analysis of changes in phospholipid fatty acids in tumor‐cell membranes after both treatments with EPA and DHA showed a significant reduction in arachidonic‐acid levels. EPA and docosapentaenoic acid (DPA), its elongation product, were increased in the phospholipids from EPA‐treated animals. DHA and EPA, but not DPA, were increased in the DHA‐treated group. It is concluded from the results of the present study that the anti‐tumoral effect of EPA is related mainly to its inhibition of cell proliferation, whereas that of DHA corresponds with its induction of apoptosis. The alterations in fatty‐acid composition induced by EPA or DHA appear to be factors underlying their differential actions on cell proliferation and apoptosis. Int. J. Cancer 75:699–705, 1998.© 1998 Wiley‐Liss, Inc.

Antioxidant and prooxidant role of beta-carotene in murine normal and tumor thymocytes: effects of oxygen partial pressure

The effects of the partial pressure of oxygen (pO2) on antioxidant efficiency of beta-carotene in inhibiting radical-initiated lipid peroxidation were studied in murine normal and tumor thymocytes. At 150 mm Hg pO2 (the pressure of oxygen in normal air), beta-carotene acted as an antioxidant, inhibiting radical-induced lipid peroxidation in both normal and tumor thymocytes. At 760 mm Hg p02, beta-carotene lost its antioxidant activity in normal thymocytes and exhibited a dose-dependent prooxidant effect in tumor thymocytes. In these cells, the prooxidant effect of beta-carotene was also accompanied by an increase of endogenous alpha-tocopherol loss. beta-Carotene radical-trapping and autooxidation reactions were faster at 760 mm Hg pO2 than at 150 mm Hg pO2 in both normal and tumor thymocytes and the carotenoid was more rapidly consumed in tumor cells. These data point out a key role of the oxygen tension on the antioxidant effectiveness of beta-carotene. They also show a selective prooxidant effect of beta-carotene under 100% oxygen in tumor cells.

Antioxidant and Anti-Inflammatory Effects of Selected Natural Compounds Contained in a Dietary Supplement on Two Human Immortalized Keratinocyte Lines

Several advantages may derive from the use of dietary supplements containing multiple natural antioxidants and/or anti-inflammatory agents. At present, however, there is scarce information on the properties and potential of combined supplements. To fill the gap, the antioxidant and anti-inflammatory activities exerted by a combination of seven natural components (coenzyme Q10, krill oil, lipoic acid, resveratrol, grape seed oil, α-tocopherol, and selenium) contained in a dietary supplement used for the prevention of skin disorders were investigated in vitro. Each component was administered, alone or in combination, to human keratinocytes, and the inhibition of Reactive Oxygen Species production and lipid peroxidation as well as the ability to reduce inflammatory cytokine secretion and to modulate Nuclear Factor-κB pathway was evaluated. The combination exhibited high antioxidant activity and in specific conditions the combinations efficiency was higher than that of the most powerful components administered individually. Moreover, the combination showed remarkable anti-inflammatory properties. It reduced more efficiently than each component the secretion of Monocyte Chemoattractant Protein-1, a crucial cytokine for the development of chronic inflammation in skin, and inhibited Nuclear Factor-κB molecular pathway. Overall, our findings suggest that the combined formulation may have the potential to powerfully inhibit oxidative stress and inflammation at skin level.

Dietary polyunsaturated fatty acids as inducers of apoptosis: implications for cancer

It has recently become clear the role played by alterations in apoptosis during the development of several chronic diseases (i.e. inflammatory, neurodegenerative and neoplastic pathologies). For this reason, the research for possible therapeutic strategies involving the modulation of the apoptotic pathways has attracted considerable interest in the past few years. In particular, it has been shown that apoptosis may be induced or inhibited by a variety of nutritional compounds providing health benefits. The aim of this review is to examine the ability of different dietary polyunsaturated fatty acids (PUFAs) to induce apoptosis, especially in the cancer field. The molecular effects of different PUFAs found in dairy products, meat, fish, vegetable seeds and oils, and known to affect the incidence and progression of cancer and other chronic diseases, will be analyzed. To this aim, our effort will concentrate in critically reviewing the published works concerning the effects of: (a) the n-6 PUFAs γ-linolenic acid, arachidonic acid, and conjugated linoleic acid; (b) the n-3 PUFAs eicosapentaenoic acid and docosahexaenoic acid on the apoptotic process. We will also pay attention to the recent findings regarding the possible role of PUFAs as regulators of the endoplasmic reticulum stress-pathway of apoptosis.

Prooxidant effects of β-carotene in cultured cells

There is a growing body of interest on the role of beta-carotene and other carotenoids in human chronic diseases, including cancer. While epidemiological evidence shows that people who ingest more dietary carotenoids exhibit a reduced risk for cancer, results from intervention trials indicate that supplemental beta-carotene enhances lung cancer incidence and mortality among smokers. A possible mechanism which can explain the dual role of beta-carotene as both a beneficial and a harmful agent in cancer as well as in other chronic diseases is its ability in modulating intracellular redox status. beta-Carotene may serve as an antioxidant or as a prooxidant, depending on its intrinsic properties as well as on the redox potential of the biological environment in which it acts. This review summarizes the available evidence for a prooxidant activity of beta-carotene in cultured cells, focusing on biochemical and molecular markers of oxidative stress, which have been reported to be enhanced by the carotenoid.

beta-carotene at high concentrations induces apoptosis by enhancing oxy-radical production in human adenocarcinoma cells.

This is the first report demonstrating a relationship between apoptosis induction and changes of intracellular redox potential in the growth-inhibitory effects of high concentrations of beta-carotene in a tumor cell line. beta-Carotene inhibited the growth of human WiDr colon adenocarcinoma cells in a dose- and time-dependent manner, induced apoptosis, and blocked Bcl-2 expression. These effects were accompanied by an enhanced production of intracellular reactive oxygen species (ROS). The addition of the antioxidant alpha-tocopherol blocked both the pro-oxidant and the growth-inhibitory effects of the carotenoid. These findings suggest that beta-carotene may act as an inductor of apoptosis by its pro-oxidant properties.

n-3 Polyunsaturated Fatty Acids and the Prevention of Colorectal Cancer: Molecular Mechanisms Involved

Increasing evidence supports the hypothesis that nutrition habits play a critical role in the incidence and growth of colorectal cancer. Among dietary factors, fish-derived n-3 polyunsaturated fatty acids (PUFAs) have gained particular interest, since epidemiological studies have shown a reduced incidence of this cancer in populations consuming high levels of fish. Also a variety of experimental studies and different clinical trials substantiated the beneficial role of n-3 PUFAs. Such an anti-neoplastic activity has been related to the regulatory effects exhibited by n-3 PUFAs on cell proliferation and apoptosis. Anti-angiogenic and anti-metastatic effects have been also reported for these fatty acids. Finally, it has been suggested that they may act as adjuvant therapeutic agents sensitizing tumors, including colon cancer, to different anti-neoplastic drugs. Several molecular mechanisms have been hypothesized to explain their anti-neoplastic action and, in particular, the modulating effect on the expression of several proteins involved in the regulation of cell cycle and apoptosis, such as Bcl-2, Bax, c-Myc seem to play a central role. Their inhibitory action has been also recently suggested for the molecular pathways driven by COX-2 and beta-catenin, known to play a major role in the development and progression of colon cancer. The aim of the present review is to analyze the anti-neoplastic effect of n-3 PUFAs towards colon cancer, and examine the molecular mechanisms involved.

Prooxidant activity of β-carotene under 100% oxygen pressure in rat liver microsomes

The effects of the partial pressure of oxygen (pO2 on antioxidant efficiency of β-carotene in inhibiting 2,2′-azobis(2-amidinopropane) (AAPH)-induced lipid peroxidation are investigated in rat liver microsomal membranes. The rate of peroxyl radicals generated by thermolysis of AAPH at 37°C is markedly higher at 150 than 760 mm Hg pO2. At 150 mm Hg pO2 β-carotene acts as an antioxidant, inhibiting 2,2′-azobis(2-amidinopropane) (AAPH)-induced Malondialdehyde (MDA) formation. At 760 mm Hg pO2, it loses its antioxidant activity and shows a prooxidant effect, increasing lipid peroxidation products, α-Tocopherol prevents the prooxidant effect of β-carotene in a dose-dependent manner. Our data provide the first evidence of a prooxidant effect of β-carotene under 100% oxygen pressure in a biological membrane model and point out the existence of cooperative interactions between β-carotene and α-tocopherol.

Cell proliferation’ differentiation’ and apoptosis are modified by n−3 polyunsaturated fatty acids in normal colonic mucosa

Supplementation with low doses of eicosapentaenoic (EPA) or docosahexaenoic (DHA) acid was used here to investigate changes in epithelial proliferation’ differentiation’ and apoptosis in normal rat colonic mucosa. ACI/T rats received by oral administration low doses of purified EPA or DHA ethyl esters (1g/kg body weight) and colonic mucosa was analyzed for cell proliferation’ differentiation’ and apoptosis. n−3 Polyunsaturated fatty acid incorporation into membrane phospholipids was investigated as reflections of fatty acid metabolism. Both EPA and DHA suppressed colonocyle proliferation and increased the numbers of differentiating and apoptotic cells without modification of the crypt morphology and the number of cells per crypt columns. A significant incorporation of the supplemented fatty acids into total phospholipids was observed. This enrichment was accompanied by a decreased content in arachidonic acid. The observation that EPA and DHA do not alter crypt morphology although they modify cell turnover in normal colonic mucosa suggests a possible use of these fatty acids as dietary chemopreventive agents.

Low-dose eicosapentaenoic or docosahexaenoic acid administration modifies fatty acid composition and does not affect susceptibility to oxidative stress in rat erythrocytes and tissues.

In view of the promising future for use of n-3 polyunsaturated fatty acids (PUFA) in the prevention of cancer and cardiovascular diseases, it is necessary to ensure that their consumption does not result in detrimental oxidative effects. The aim of the present work was to test a hypothesis that low doses of eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) do not induce harmful modifications of oxidative cell metabolism, as modifications of membrane fatty acid composition occur. Wistar rats received by gavage oleic acid, EPA, or DHA (360 mg/kg body weight/day) for a period of 1 or 4 wk. Fatty acid composition and α-tocopherol content were determined for plasma, red blood cell (RBC) membranes, and liver, kidney, lung, and heart microsomal membranes. Susceptibility to oxidative stress induced by tert-butylhydroperoxide was measured in RBC. EPA treatment increased EPA and docosapentaenoic acid (DPA) content in plasma and in all the membranes studied. DHA treatment mainly increased DHA content. Both treatments decreased arachidonic acid content and n-6/n-3 PUFA ratio in the membranes, without modifying the Unsaturation Index. No changes in tissue α-tocopherol content and in RBC susceptibility to oxidative stress were induced by either EPA or DHA treatment. The data suggest that EPA and DHA treatments can substantially modify membrane fatty acids, with-out increasing susceptibility to oxidative stress, when administered at low doses. This opens the possibility for use of low doses of n-3 PUFA for chemoprevention without risk of detrimental secondary effects.