The activity and expression of NOS II and arginase can be stimulated in peritoneal macrophages in vitro by injecting inflammatory agents into the peritoneal cavity. A markedly different response in arginine metabolism was observed in mouse and rat macrophages. Casein treatment was a potent inducer for both enzymes. NDV vaccines induced mainly NOS II, while thioglycollate induced arginase.
Repair and replicative DNA synthesis were measured at different stages of the cell cycle in control and cadmiumtreated Chinese hamster ovary (CHO-K1) cells. Cells were synchronized by counterflow centrifugal elutriation. Elutriation resulted in five repair and four replication subphases. On Cd treatment, repair synthesis was elevated in certain subphases. Replicative subphases were suppressed by Cd treatment, with some of the peaks almost invisible. The number of spontaneous strand breaks measured by random oligonucleotide primed synthesis assay showed a cell-cycle-dependent fluctuation in control cells and was greatly increased after Cd treatment throughout the S phase. Elevated levels of the oxidative DNA damage product, 8-oxodeoxyguanosine, were observed after Cd treatment, with the highest level in early S phase, which gradually declined as damaged cells progressed through the cell cycle.Keywords: 8-oxodeoxyguanosine; elutriation; permeable cells; strand breaks; synchronization. [6,7,10±12]. Heavy metals generate oxidizing radicals through Fenton chemistry and by the Haber±Weiss reaction leading to the hypothesis that metal carcinogenesis is mediated primarily by the elevated level of free radicals (reviewed by Kasprzak [13]). According to this view, heavy-metal-induced oxidative stress can lead to different types of DNA damage as a consequence of consumption of molecular oxygen in multiple steps of incomplete O 2 reduction, ultimately producing water.Common types of oxidative damage cause changes in DNA structure, the long-term effect of which can lead to malignant transformation. Structural changes involve alterations in nucleotide bases, cross-links, strand breaks, formation of bulky DNA adducts, etc. DNA damage suppresses DNA replication at checkpoints to avoid mutagenic changes being perpetuated in the genome of the next generation of cells [14±16]. A comparison of cell cycle profiles of replicative repair synthesis in permeable cells showed opposite trends. The rates of repair synthesis and replication are inversely correlated [17].The detection of oxidative DNA damage leading to degenerative diseases associated with aging involves chromatographic immunochemical approaches, measurement of oxidized bases such as 8-hydroxyguanine, 8-hydroxyadenine, and thymine glycol as well as oxidized DNA adducts such as 8-hydroxy-2 H -deoxyguanosine and thymidine glycol. The aim of this paper is to describe how oxidative damage of Cd treatment reduces replicative, and increases repair DNA synthesis during cell cycle. Changes have been expressed as the ratio of replication/repair synthesis. Generation of DNA strand breaks and the carcinogen indicator 8-deoxyguanosine were also measured in a cell-cycle-dependent manner.
M A T E R I A L S A N D M E T H O D S
Growth conditionsChinese hamster ovary cells (CHO-K1, ATCC No. CCL61) were kept in suspension culture in spinner flasks using F12 medium supplemented with 10% fetal bovine serum at 37 8C and 5% CO 2 .
Cadmium treatmentCHO cells were treated with CdCl 2 (0.2±1 mm) 9...
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