Olga Brossa

Effects of 4-Hydroxynonenal, A Product of Lipid Peroxidation, on Cell Proliferation and Ornithine Decarboxylase Activity

4-hydroxynonenal (HNE) is one of the major breakdown products of cellular lipid peroxidation. Its effects on proliferation, ornithine decarboxylase (ODC) activity and DNA synthesis have been investigated in leukemic cell lines. The cells were incubated for 1 hour with different aldehyde concentrations, then washed and resuspended in medium with fresh foetal calf serum. HNE concentrations ranging from 10(-5) to 10(-6) M significantly inhibited ODC activity when induced by addition of fresh foetal calf serum both in K562 and HL-60 cells. 3H-Thymidine incorporation in K562 cells was also inhibited from 6 to 12 hours after the treatment. The same HNE concentrations did not inhibit ODC activity when added to cytosol, thus a direct action on the enzyme can be excluded. Moreover, HNE did not affect the half-life of ODC, so that a specific effect on ODC synthesis may be supposed. These data indicate a reduction of proliferative capacity of the cells and are consistent with the possibility that HNE, at concentrations close to those found in normal cells, plays a role in the control of cell proliferation.

IN HEPATOMA CELL LINES RESTORED LIPID PEROXIDATION AFFECTS CELL VIABILITY INVERSELY TO ALDEHYDE METABOLIZING ENZYME ACTIVITY

Hepatoma cells are less susceptible to oxidative stress than normal hepatocytes: the decreased content of polyunsaturated fatty acids in such cells decreases their capability to undergo lipid peroxidation (Gravela et al., 1975; Canuto et al., 1991; Cheeseman et al., 1988; Feo et al., 1975; Canuto et al., 1994; Masotti et al., 1988). A consequence of this is the reduced production of aldehydes inside the cells (Poli et al., 1986). Depending on the quantity present, aldehydes have several effects on the cells. In particular, the effects of 4-hydroxynonenal (4-HNE), an important aldehyde produced by lipid peroxidation, have been studied (Esterbauer et al., 1991). At μM concentrations, the effects are positive: adenylate cyclase and phospholipase C are stimulated (Paradisi et al., 1985; Garramone et al., 1988) and differentiation is induced in HL-60 cells (Barrera et al., 1991). At higher concentrations, 4-HNE is cytotoxic (Esterbauer et al., 1991; Canuto et al., 1995).

Increased susceptibility to transglutaminase of eye lens proteins exposed to activated oxygen species produced in the glucose-glucose oxidase reaction.

In order to test whether a mild oxidative stress could promote the transglutaminase damaging effect on eye lens proteins, total lens soluble proteins and purified beta L-crystallin have been exposed to H2O2 slowly produced by the glucose-glucose oxidase reaction. Soon after the pretreatment, the substrate capacity of the lens proteins for an exogenous transglutaminase has been evaluated. Exposure to the oxidative stress increased the susceptibility of the lens proteins to transglutaminase. When ferrous ions were added to the preincubation medium, in order to convert the H2O2 into the hydroxyl radical, the increase was more evident.

Phenobarbital stimulation of cytochrome P-450 and aminopyrine N-demethylase in hyperplastic liver nodules during LD-ethionine carcinogenesis

Microsomes isolated from hyperplastic liver nodules and hepatomas, induced by DL-ethionine, exhibited a reduced cytochrome P-450 content and aminopyrine N-demethylase activity when compared to the organelles of control and surrounding non-nodular liver. Phenobarbital administration to rats caused an increase of microsomal protein, cytochrome P-450 and aminopyrine N-demethylase in all tissue tested. In the hepatoma the rise of cytochrome P-450 and aminopyrine N-demethylase/g of tissue was very low and it is compensated by a slight increase of microsomal protein. In hyperplastic nodules as well as in control and surrounding livers, cytochrome P-450 and aminopyrine N-demethylase increased more than microsomal protein. However, the phenobarbital-induced stimulation was significantly lower in hyperplastic nodules than in control and surrounding livers.

The effect of 4-hydroxy-2, 3-trans-pent-en-1-al and maleylaldehyde on some mitochondrial functions

Abstract Low concentrations of HPE and MLA inhibited state 3 respiration of rat liver mitochondria in the presence of different NAD+-dependent substrates. MLA appeared to be more active than HPE. High aldehyde concentrations inhibited the state 3 respiration with succinate. The restraint of succinate oxidation by HPE and MLA and of glutamate plus malate oxidation by MLA correlated with the inhibition of succinate and glutamate dehydrogenase activites, respectively. HPE inhibited glutamate dehydrogenase at concentrations higher than those affecting glutamate oxidation. Malate dehydrogenase activity was slightly sensitive to HPE and MLA. Both aldehydes inhibited NADH oxidation by freeze-thawed mitochondria. These results suggest the existence of a site particularly sensitive to aldehydes in the electron transport chain between the specific NAD+-linked dehydrogenases and ubiquinone.

Studies on ornithine decarboxylase from liver, kidney and tumoral tissues during activity decay following cycloheximide administration

We have studied the activity, thiol-dependency and Km of ornithine decarboxylase (ODC) from the following sources: liver of rats subjected to partial hepatectomy or administered thioacetamide, the rat 3924A Morris hepatoma, the rat AH130 Yoshida ascites hepatoma, a mouse transplantable mammary carcinoma and kidney of rats administered testosterone propionate. In order to detect possible changes occurring during in vivo ageing of this enzyme we inhibited protein synthesis with cycloheximide. A gradual decrease of Km during ageing was observed in ODC from liver.