Dihydropyridines decrease X-ray-induced DNA base damage in mammalian cells.

Compounds with the structural motif of 1,4-dihydropyridine display a broad spectrum of biological activities, often defined as bioprotective. Among them are L-type calcium channel blockers, however, also derivatives which do not block calcium channels exert various effects at the cellular and organismal levels. We examined the effect of sodium 3,5-bis-ethoxycarbonyl-2,6-dimethyl-1,4-dihydropyridine-4-carboxylate (denoted here as DHP and previously also as AV-153) on X-ray-induced DNA damage and mutation frequency at the HGPRT (hypoxanthine-guanine phosphoribosyl transferase) locus in Chinese hamster ovary CHO-K1 cells. Using formamido-pyrimidine glycosylase (FPG) comet assay, we found that 1-h DHP (10nM) treatment before X-irradiation considerably reduced the initial level of FPG-recognized DNA base damage, which was consistent with decreased 8-oxo-7,8-dihydro-2'-deoxyguanosine content and mutation frequency lowered by about 40%. No effect on single strand break rejoining or on cell survival was observed. Similar base damage-protective effect was observed for two calcium channel blockers: nifedipine (structurally similar to DHP) or verapamil (structurally unrelated). So far, the specificity of the DHP-caused reduction in DNA damage - practically limited to base damage - has no satisfactory explanation.

Cell survival and chromosomal aberrations in CHO-K1 cells irradiated by carbon ions.

Chinese hamster ovary CHO-K1 cells were exposed to high LET (12)C-beam (LET: 830 keV/microm) in the dose range of 0-6 Gy and to (60)Co irradiation and the RBE value was obtained. Effects of (12)C-beam exposure on cell survival and chromosomal aberrations were calculated. The chromosomal aberration data were fitted with linear equation. The distribution of aberration in cells was examined with a standard u-test and used to evaluate the data according to Poisson probabilities. The variance to the mean ratio sigma(2)/Y and the dispersion index (u) were determined. Overdispersion was significant (p<0.05) when the value of u exceeded 1.96.

Artificial reduction in transepidermal water loss improves skin barrier function.

BACKGROUND: Artificial reduction of abnormal transepidermal water loss (TEWL) is considered to improve skin diseases associated with a defective barrier function. Treatment of the skin with moisturizers is also known to influence skin barrier function. Whether or not differences in occlusion between creams contribute to their effects on the skin barrier function is unknown. OBJECTIVES: To investigate the long-term effects of a semipermeable membrane on the skin barrier function in normal skin. In addition, the occlusive properties of two creams were studied. METHODS: The study was randomized, controlled and evaluator-blind using measurement of TEWL and skin susceptibility to sodium lauryl sulphate as indicators of skin barrier function. RESULTS: Coating of the skin with a silicone membrane for 23 h per day for 3 weeks improved skin barrier function, whereas no significant changes were found after using the membrane for 8 h per day. CONCLUSIONS: Differences between creams in terms of their effect on skin barrier function cannot be solely explained by their occlusive properties.

Changes in skin barrier function following long-term treatment with moisturizers, a randomized controlled trial.

BACKGROUND: Moisturizers are commonly used by patients with dry skin conditions as well as people with healthy skin. Previous studies on short-term treatment have shown that moisturizers can weaken or strengthen skin barrier function and also influence skin barrier recovery. However, knowledge of the effects on skin barrier function of long-term treatment with moisturizers is still scarce. OBJECTIVES: To investigate the impact of long-term treatment with moisturizers on the barrier function of normal skin, as measured by transepidermal water loss (TEWL) and susceptibility to an irritant, and to relate those effects to the composition of the designed experimental moisturizers. METHODS: Volunteers (n = 78) were randomized into five groups. Each group treated one volar forearm for 7 weeks with one of the following preparations: (i) one of three simplified creams, containing only a few ingredients in order to minimize the complexity of the system; (ii) a lipid-free gel; (iii) one ordinary cream, containing 5% urea, which has previously been shown to decrease TEWL. The lipids in the simplified creams were either hydrocarbons or vegetable triglyceride oil, and one of them also contained 5% urea. After 7 weeks, treated and control forearms were exposed for 24 h to sodium lauryl sulfate (SLS) using a patch test. TEWL, blood flow and skin capacitance of both SLS-exposed and undamaged skin were evaluated 24 h after removal of patches. Additionally, a 24-h irritancy patch test of all test preparations was performed on 11 volunteers in order to check their possible acute irritancy potential. RESULTS: Changes were found in the barrier function of normal skin after 7 weeks of treatment with the test preparations. The simplified creams and the lipid-free gel increased TEWL and skin response to SLS, while the ordinary cream had the opposite effect. One of the simplified creams also decreased skin capacitance. All test preparations were shown to be nonirritant, both by short-term irritancy patch test and by measurement of blood flow after long-term treatment. CONCLUSIONS: Moisturizers influence the skin barrier function of normal skin, as measured by TEWL and susceptibility to SLS. Moreover, the effect on skin barrier function is determined by the composition of the moisturizer. The ingredients which influence the skin barrier function need to be identified, and the mechanism clarified at the molecular level.

Irritation potential of bath and shower oils before and after use: a double-blind randomized study.

BACKGROUND: Difficulties in avoiding weak irritants may contribute to chronic contact dermatitis. A large variety of shower and bath oils are claimed to be suitable for use on dry skin because of their mildness and because they deposit a protective oil film on the skin. OBJECTIVES: The aim of the present study was to investigate possible differences in the irritation potential of eight shower or bath oils and to investigate whether surfactant residues may form a reservoir of irritant substance on the skin. PATIENTS AND METHODS: The study was double-blind and randomized using healthy human volunteers. The inherent capacity of the products to induce irritation was determined using conventional patch test techniques. Detection of potentially irritant residues was done by occlusion of the treated and rinsed skin area, followed by evaluation of the biological response. Instrumental measurements of transepidermal water loss and superficial skin blood flow served as indicators of the injurious effects of the products. RESULTS AND CONCLUSIONS: The results showed large differences between the products in irritant potential. Some did not irritate skin more than water, whereas others demonstrated considerably damaging effects. Moreover, the study proved the presence of barrier-impairing residues on the skin after rinsing with water. Thus, instead of protecting the skin, some formulations may induce subclinical injuries and delay skin barrier function recovery with prolonged risk for patients with eczema.

Differential anti-proliferative properties of novel hydroxydicarboxylatoplatinum(II) complexes with high or low reactivity with thiols.

We have examined the anti-proliferative effect of 13 recently synthesised platinum dicarboxylate complexes, very similar in their chemical, structural and kinetic properties to carboplatin. We used the L5178Y model: two murine lymphoma sublines, which differ in nucleotide excision repair ability and hence, in sensitivity to those platinum complexes that react with DNA. The anti-proliferative effect of the examined compounds mainly depends on the kind of amine ligand. Complexes with the primary amine (ethylenediamine) are more effective than complexes containing the tertiary amine (1-alkylimidazole). The ethylenediaminemalatoplatinum(II) complexes show a differential in vitro anti-proliferative activity in the L5178Y model; hence, it may be expected that they inflict DNA lesions that are repaired by the nucleotide excision system. The cytotoxicity of these complexes is directly correlated with reactivity with glutathione (GSH). The 1-alkylimidazole complexes are of low toxicity and moderate to low reactivity with GSH; in contrast to the ethylenediaminemalatoplatinum(II) complexes, their cytotoxicity is inversely correlated with reactivity with GSH. Two of the 1-alkylimidazole complexes, bis(1-ethylimidazole)(L-malato)platinum(II) and bis(1-propylimidazole (L-malato)platinum(II), show a considerable ability to arrest cells in G2 phase. We expect that the properties of these two groups of platinum complexes may be exploited in combined platinum complex treatment and irradiation.

Effects of topoisomerase I-targeted drugs on radiation response of L5178Y sublines differentially radiation and drug sensitive.

The murine L5178Y (LY) lymphoma sublines, LY-R (radiation resistant) and LY-S (radiation sensitive) display a difference in susceptibility to camptothecin (CPT): LY-S cells are less sensitive to killing by this inhibitor of topoisomerase I than LY-R cells. Post-treatment (CPT present until 3 h after irradiation) sensitizes only LY-S cells. In agreement with this, only in LY-S cells is the relative number of DNA-protein cross-links formed after treatment with CPT + X higher than expected for additivity of X-ray and CPT-induced damage. The pattern of changes in the labelling indices and cell cycle distribution in cells that underwent combined treatment is essentially like that seen for single-agent treatment: for LY-S cells like that for radiation, for LY-R cells like that for CPT. We found no direct relation between the patterns of cell cycle distributions and the enhancement of the lethal effect of X-irradiation by CPT post-treatment. The sublines are not markedly differentially sensitive to beta-lapachone, which modifies topoisomerase I activity, and not sensitized to X-rays by post-irradiation treatment with the drug.

Effects of 8-chloroadenosine-3',5'-monophosphate in combination with irradiation in L5178Y mouse lymphoblasts.

The effect of a new anticancer drug, 8-chloroadenosine 3',5'-monophosphate (8-Cl-cAMP-), a site selective cAMP analog, that inhibits growth of cancer cells in vitro, was examined in L5178Y (LY) murine lymphoma cells. Two LY sublines were used, grown in full Fisher's medium: LY-R, radiation resistant and LY-S, radiation sensitive. The latter was also adapted to grow in simplified medium. In the full medium conversion of 8-Cl-cAMP to 8-chloroadenosine presumably was the case of cytotoxicity. In the simplified medium this conversion was limited and the cytotoxic effect much less pronounced. Cytotoxicity was equal in LY-R and LY-S cells and it was not related to changes in the cell cycle distribution; the latter were observed in LY-S, but not in LY-R cells. There was no interaction of the drug with x-rays in LY cells grown either in full or simplified medium.

Sensitivity to inhibitors of type II topoisomerases from mouse L5178Y lymphoma strains that are resistant or sensitive to X-radiation.

Sensitivity to topoisomerase II inhibitors was tested at the cellular and enzyme level for two strains of mouse L5178Y lymphoma cells: resistant (LY-R) and sensitive (LY-S) to X-radiation. Differences in the susceptibility to inhibitors between LY-R and LY-S cells depended on the inhibitor used and were observed for adriamycin and VP-16, but not for mitoxantrone. On the other hand, isolated enzymes displayed the same sensitivity to all inhibitors tested regardless of the cell line. These results exclude the presence of altered topoisomerase II in LY-S cells as a possible reason for the collateral sensitivity of LY-S cells to X-radiation and topoisomerase II inhibitors.

Reduced sensitivity to camptothecin of topoisomerase I from a L5178Y mouse lymphoma subline sensitive to X-radiation.

Murine L517BY (LY) lymphoma sublines, LY-R (X-radiation resistant) and LY-S (X-radiation sensitive) displayed a difference in susceptibility to camptothecin: susceptibility of LY-S cells to the alkaloid was shifted towards higher concentrations as compared to LY-R cells. A similar difference was observed at the level of genomic DNA when a number of DNA-protein cross-links was determined or single-strand breaks were revealed by the fluorescent nucleoid halo assay. Activities of topoisomerases I and II were the same in both sublines. In turn, a higher resistance to camptothecin was found for the isolated LY-S topoisomerase I in the DNA cleavage test, suggesting that an altered enzyme was responsible for the susceptibility difference observed at the cellular level. In the relaxation test the enzymes from the two sublines showed a different sensitivity to beta-lapachone, an activator of topoisomerase I, but were similarly sensitive to all inhibitors, except camptothecin.

Hydrogen peroxide induced reproductive and interphase death in two strains of L5178Y murine lymphoma differing in radiation sensitivity.

L5178Y-R (LY-R) and L5178Y-S (LY-S) cells, differing in radiation sensitivity and susceptibility to the radiosensitizing effect of benzamide (Bz) were examined for susceptibility to hydrogen peroxide. Survival and chromatid aberration frequency indicated that LY-R cells were considerably more sensitive to H2O2 than LY-S cells. So, LY strains were found to be inversely cross-sensitive to X/gamma rays and H2O2. The relative resistance to H2O2 corresponded with the previously found twofold difference in catalase activity (Jaworska et al. 1987). At higher concentrations H2O2 treatment caused interphase death, that was delayed by benzamide (Bz, 2mM), an inhibitor of poly(ADP-ribosylation), to a lesser extent in the more resistant cell subline (LY-S). From the examination of the H2O2 induced increase in the free Ca2+ concentration (with or without 2 mM Bz treatment) with the use of Fura-2 it followed, that the cells responded to the oxidative stress by Ca2+ release. The Ca2+ concentration increase was neither directly related to the killing effect of H2O2 treatment, nor did it correspond with the twofold difference in catalase activity in LY strains.

Ca2+ mobilization is related to the lethal effect of X-irradiation in L5178Y cells.

Radiation-resistant L5178Y-R (LY-R) and radiation-sensitive L5178Y-S (LY-S) murine lymphoma cells were X-irradiated and their free Ca2+ concentration was examined with Fura-2 in Ca2(+)-free salt solution. The release of free Ca2+ from intracellular stores was linear between 10 and 60 min after irradiation (1-5 Gy X-rays) and was higher in LY-S than in LY-R cells. Pre-treatment with 2 mM benzamide (Bz) further increased the concentration of free Ca2+ in LY-S cells but not in LY-R cells. In contrast with LY-R cells, LY-S cells had previously been found to be radiosensitized by continuous 2 mM Bz treatment (Szumiel et al. 1984b). Thus, there was a parallel effect of Bz on survival and on the increase in free Ca2+ concentration in LY cells. Moreover, the rates of Ca2+ release after irradiation with 1-5 Gy of X-rays with or without Bz treatment were inversely related to the respective surviving fractions of LY cells.

Benzamide-inhibitable alterations in spectral properties of chromatin accompany DNA repair in UVC-exposed L5178Y cells.

We examined the response of chromatin to increasing NaCl and MgCl2 concentrations in UVC-irradiated L5178Y (LY) R and S cells, using the spectral index method (Dixon and Burkholder 1985). We have found an alteration in chromatin properties 1 h after UVC-irradiation of repair proficient LY-S cells, but no change in repair deficient LY-R cells. The change was shown as lowered spectral index, indicating that at given Na+ and Mg++ concentrations (1 or 200 mM NaCl, 0 or 0.5 mM MgCl2) chromatin from UVC-irradiated LY-S cells was more compact than that from unirradiated ones. Benzamide treatment reversed the effect of UVC-irradiation in LY-S cells and did not change the response pattern of chromatin from LY-R cells or unirradiated LY-S cells.