Antioxidant defense mechanisms in murine epidermis and dermis and their responses to ultraviolet light

A comprehensive comparison of antioxidant defenses in the dermis and epidermis and their response to exposure to ultraviolet (UV) irradiation has not previously been attempted. In this study, enzymic and non-enzymic antioxidants in epidermis and dermis of hairless mice were compared. Enzyme activities are presented both as units/gram of skin and units/milligram of protein; arguments are presented for the superiority of skin wet weight as a reference base. Catalase, glutathione peroxidase, and glutathione reductase (units/gram of skin) were higher in epidermis than dermis by 49%, 86%, and 74%, respectively. Superoxide dismutase did not follow this pattern. Lipophilic antioxidants (alpha-tocopherol, ubiquinol 9, and ubiquinone 9) and hydrophilic antioxidants (ascorbic acid, dehydroascorbic acid, and glutathione) were 24-95% higher in epidermis than in dermis. In contrast, oxidized glutathione was 60% lower in epidermis than in dermis. Mice were irradiated with solar light to examine the response of these cutaneous layers to UV irradiation. After irradiation with 25 J/cm2 (UVA + UVB, from a solar simulator), 10 times the minimum erythemal dose, epidermal and dermal catalase and superoxide dismutase activities were greatly decreased. alpha-Tocopherol, ubiquinol 9, ubiquinone 9, ascorbic acid, dehydroascorbic acid, and reduced glutathione decreased in both epidermis and dermis by 26-93%. Oxidized glutathione showed a slight, non-significant increase. Because the reduction in total ascorbate and catalase was much more severe in epidermis than dermis, it can be concluded that UV light is more damaging to the antioxidant defenses in the epidermis than in the dermis.     

Recurrent phototriggered summer light eruption of the axillary and inguinal area: an atypical topographic form of benign summer lucitis

INTRODUCTION This case report of benign summer light eruption emphasizes the importance of phototests in the diagnosis of photosensitive dermatoses. CASE REPORT: A 25-year-old man, phototype II, had experienced a pruriginous papulovesicular erythematous eruption of the axillary and inguinal regions each summer for 12 years. A high-dose UV phototest (40 J/cm2 x 3 days) directed on the right posterior axillary area and a whole body exposure test (4 J/cm2 UVA, 20 mJ/cm2 UVB x 3 days) were positive both clinically and histologically on day 4. DEM B was normal at 26 mJ/cm2. Iterative polychromatic phototest (DEM x 3 days) in the area usually involved (left posterior axillary region) was negative. The simple UVA (13 J/cm2) and iterative phototests performed on the back were negative. The results of the phototests led to the diagnosis of benign light eruption despite the unusual localization. DISCUSSION: The diagnosis of benign light eruption is generally clinical. Phototests are unnecessary in most cases. Benign light eruption can be triggered by high-dose iterative UVA exposure of the susceptible area or whole body phototests (UVA-UVB). These specific phototests are indicated in atypical forms or localizations in order to determine the course of benign light eruption and to uncover simulations.     

The value of a well-placed stoma

To study the cumulative influence of UV irradiations on skin matrix alterations, human skin fibroblasts were irradiated successively three-fold, at 24 h intervals, with UVA (3x5J/cm2), UVB (3x8mJ/cm2), UVA plus UVB (3x5J/cm2 and 3x8mJ/cm2) and the levels of 92 kDa gelatinase (pro-MMP9), 72 kDa gelatinase (pro-MMP2) and plasma-membrane elastase type protease were determined, following subsequent 24-h culture in 10% serum-containing medium. UV irradiations had only minor influence (1.4-fold increase for UVB) on secreted levels of pro-MMP2 and decreased the amount of plasma membrane elastase produced by cells. It did however, for UVA and UVB alone, induce a significant increase of 66 kDa activated MMP2 production: 2.5- and 1.7-fold respectively. Such enhancement was not observed when combined irradiations were administered. UV exposure possessed a much higher influence on pro-MMP9 secretion by dermal fibroblast enhancing enzyme levels by 2.5-, 6.5- and 5-fold for UVA, UVB and UVA+UVB, respectively.     

Psoralen-mediated photodecontamination of platelet concentrates: inactivation of cell-free and cell-associated forms of human immunodeficiency virus and assessment of platelet function in vivo

BACKGROUND: Treatment of platelet concentrates (PCs) with psoralens and broad-band ultraviolet A (UVA) radiation is being examined for the elimination of pathogens that might be present in donated blood. Previous studies have demonstrated the inactivation of cell-free viruses and the maintenance of platelet integrity with common in vitro assays. STUDY DESIGN AND METHODS: Human immunodeficiency virus (HIV) in three forms-cell-free, activity replicating, and latently infected cell lines-was added to PCs and treated with 50-microgram per mL of 4'-aminomethyl-4,5',8-trimethylpsoralen (AMT), 0.35 mM rutin, and broad- and narrow-band UVA light (320-400 nm and 360-370 nm [UVA1], respectively). The inactivation of added HIV was assessed in tissue culture; platelet hemostatic activity was assessed in thrombocytopenic rabbits. RESULTS: Each form of HIV was inactivated completely (> or = 10(5) infectious units) on treatment with 30 J per cm2 of UVA1 light. Similar results were obtained on treatment of 2.5 mL of PCs in test tubes or intact PC units (50 mL) in blood bags. Latently infected cell lines were substantially more sensitive than cell-free HIV or HIV that was actively replicating. Human platelets treated with 40 J per cm2 of UVA1 light had a fully corrected bleeding time shortly after treatment or after 5 days' storage, as assessed in thrombocytopenic rabbits. Platelet hemostatic function began to decrease with 81 J per cm2 of UVA1 light and was abolished with 113 J per cm2. At similar fluences, broad-band UVA light was more injurious to platelets than was UVA1 light. CONCLUSION: HIV transmission might be eliminated by PCs after treatment with AMT and UVA1 light and without a reduction in platelet hemostatic function.     

Digestion of 125I-labelled plasmin-derived fibrin degradation products by neutrophil lysosomal enzymes

Ultraviolet A radiation (UVA, 320-400 nm) is mutagenic and induces genomic damage to skin cells. N-acetyl-cysteine (NAC), selenium and zinc have been shown to have antioxidant properties and to exhibit protective effects against UVA cytotoxicity. The present work attempts to delineate the effect of these compounds on genomic integrity of human skin fibroblasts exposed to UVA radiation using the single cell gel electrophoresis (SCGE) or Comet assay. The cells were incubated with NAC (5 mM), sodium selenite (0.6 microM) or zinc chloride (100 microM). Then cells were embedded in low melting point agarose, and immediately submitted to UVA fluences ranging from 1 to 6J/cm2. In the Comet assay, the tail moment increased by 45% (1 J/cm2) to 89% (6J/cm2) in non-supplemented cells (p)<0.01). DNA damage was significantly prevented by NAC, Se and Zn, with a similar efficiency from 1 to 4J/cm2 (p < 0.05). For the highest UVA dose (6J/cm2), Se and Zn were more effective than NAC (p < 0.01).     

In vitro photoinhibition by psoralen and ultraviolet A radiation of human hematopoietic progenitors

The in vitro sensitivity of human hematopoietic progenitors to PUVA, 8-MOP and UVA alone was investigated. 8-MOP alone at final concentrations of 150, 200, 600 and 1,000 ng/ml did not modify colony growth of circulating and bone marrow erythroid (BFU-E), myeloid (CFU-GM) and immature (CFU-GEMM) hematopoietic progenitors obtained from normal controls. The exposure of the same progenitors to increasing doses of UVA, up to 12 J/cm2, progressively decreased hematopoietic colony growth (with estimated 50% inhibition occurring at about 5 J/cm2). In vitro PUVA treatment (8-MOP 200 ng/ml followed by UVA 5 J/cm2) caused 90% growth inhibition of circulating and bone marrow hematopoietic progenitors. In addition, the treatment completely inhibited the formation of spontaneous erythroid colonies, obtained from 5 polycythemic patients, that are considered to be a marker of this neoplastic disease. PUVA cytotoxicity was assessed by the colorimetric MTT assay. The percentage of cell death after PUVA exposure was 29 +/- 10% for both peripheral and bone marrow mononuclear cells. Our findings indicate that 8-MOP alone is not toxic to hematopoietic progenitors whereas UVA treatment determines in vitro a dose-dependent inhibition of the clonogenic capacity of normal hematopoietic cells. PUVA treatment enhances this effect, causing a quite complete inhibition of hematopoietic progenitors colony formation from normal donors and spontaneous BFU-E colony formation from polycythemic patients.     

Photoallergenicity of a fluoroquinolone antibacterial agent with a fluorine substituent at the 8-position in guinea pigs exposed to long-wavelength UV light

The 8-position of the quinolone ring of balofloxacin (BLFX), one of fluoroquinolones, was replaced with fluorine to obtain the 8-F. When an aqueous solution of bovine serum albumin (BSA) containing the 8-F was exposed to long-wavelength UV light (UVA) at a rate of 2.5 J/cm2, the absorbance of BSA at 300 nm or longer wavelengths increased markedly in comparison to that of native BSA. In addition, when a homogenate of skin tissue from Hartley guinea pigs was exposed to UVA (2.5 J/cm2) in the presence of the 8-F and then injected subcutaneously into guinea pigs, the animals produced IgG class antibody specific to the 8-F and its UVA-irradiation product. No such phenomenon, however, was observed when the parent compound, i.e. , BLFX which possesses a methoxy group at the 8-position, was used instead of the 8-F. In a subsequent experiment, the 8-F was administered either orally or topically to the shaved neck of guinea pigs and then irradiated with UVA (5 J/cm2) once daily for 5 days. When the treated animals were challenged by a combination of UVA irradiation (5 J/cm2) and either an oral or intradermal administration of the 8-F, 2 and 3 of the 5 animals showed redness and erythema on the irradiated area, respectively. However, no change was observed when BLFX was used instead of the 8-F. These results suggest that the introduction of a fluorine substituent to the 8-position of quinoline ring of fluoroquinolones induces photoallergic responses in which the fluoroquinolone or its photo-denatured product(s) act as an allergen.     

Pretherapeutic evaluation in cancer of the cardia

Two new UV lamps were investigated with respect to their antibacterial effectiveness in vitro. Propioni (n = 20 strains) and Micrococcaceae (n = 16 strains) bacteria extracted from acne patients were applied to RCM and sheep blood agar plates and irradiated with a narrow-band UVB lamp (TL 100W/10R) at a wavelength of 313 nm and a UVA1 lamp (TL 01) at a wavelength of 345-440 nm. The precisely defined energy levels were, in the case of narrow-band UVB, 0.00, 0.30, 0.50, 1.00, 2.00 and 3.00 J/cm2 and, in the case of UVA1, 0.00, 2.50, 5.00, 7.50, 10.00 and 20.00 J/cm2. UVA1 inhibited neither the growth of Propioni nor Micrococcaceae bacteria. In contrast, the growth of Micrococcaceae was inhibited already at a dosage of 0.30 J/cm2 of narrow-band UVB (P < 0.05), highly significant from 0.50 J/cm2 (P < 0.01) and to a maximum of 2.2 powers of 10 at 3.00 J/cm2 compared with non-radiated control plates. Propioni bacteria were significantly inhibited at the minimum dosage of 0.30 J/cm2 of narrow-band UVB (P < 0.01) and to a maximum of 2.8 powers of 10 at 3.00 J/cm2.     

Adverse effects of sunlight on the skin

Many people consider a summer's day pleasant: warm and bright. The sun's ultraviolet rays do not contribute to the pleasure, and are biologically mainly harmful. As UV radiation does not penetrate any deeper than our skin, this organ has to be particularly well adapted to the UV exposure. The skin exploits the UV radiation for the synthesis of vitamin D3. Our day-to-day exposure suffices for this beneficial UV effect. Excessive exposure, as in sunbathing, only contributes to the adverse effects, like sunburn and suppression of cellular immunity in the short term, and 'photoaging' and skin cancer in the long term. The UVB radiation in sunlight is mainly responsible for these harmful effects, the UVA radiation to a far lesser extent (10-20% contribution). The UVA radiation from modern tanning equipment does not differ from that in sunlight, but UVA radiation does not lead to vitamin D3 production; it rather degrades vitamin D3 and a tan offers insufficient protection against the UVB radiation in full sunlight.     

Differential regulation of P53 and Bcl-2 expression by ultraviolet A and B

The induction of apoptosis by ultraviolet (UV) radiation and other DNA damaging agents plays a critical role in monitoring the accumulation of genetic damage and the suppression of tumor development. We hypothesize that UVA and UVB induce apoptosis by modulating balances between p53 and/or bcl-2 genes. Using MCF-7 cells that express both wild-type P53 and Bcl-2 proteins, we demonstrated that UVA and UVB induced apoptosis through regulating expression of apoptosis promoting or inhibiting genes. UVA induced immediate apoptosis and downregulated bcl-2 expression. Bcl-2 expression was reduced by approximately 40% at 4 h post-150 kJ UVA irradiation per m2 with a maximum downregulation (over 70%) at 24 h. The dose-response studies revealed that significant reduction of bcl-2 expression was observed at UVA doses ranging from 50 to 200 kJ per m2; however, p53 levels were not affected by UVA. In contrast, UVB exhibited a entirely different action than UVA in that UVB substantially induced p53 expression, but had no effect on bcl-2 expression. The induction of P53 by UVB was dose and time dependent with the maximum expression at 24 h post-2 and post-4 kJ UVB irradiation per m2. Down-regulation of bcl-2 and fragmentation of DNA induced by UVA occurred earlier (approximately at 4 h) than upregulation of p53 and DNA fragmentation by UVB (12-24 h). These results suggest that UVA and UVB cause cell damage through different mechanisms and that the balances between the expression of p53 and bcl-2 may play an important role in regulating the apoptosis induced by UV irradiation.     

Inactivation of leukocytes in platelet concentrates by photochemical treatment with psoralen plus UVA

A photochemical treatment (PCT) process using a novel psoralen and long wavelength ultraviolet light (UVA, 320-400 nm) has been developed to inactivate bacteria and viruses in platelet concentrates. This study evaluated the efficacy of PCT for inactivation of leukocytes that contaminate platelet preparations. Three psoralens, 8-methoxypsoralen (8-MOP), 4'-aminomethyl 4,5', 8-trimethylpsoralen (AMT), and the novel psoralen S-59, were compared using the following four independent but complementary biological and molecular assays. (1) T-cell viability: Treatment with 150 mumol/L S-59 and 1.0 to 3.0 Joules/cm2 UVA inactivated >5.4 +/- 0.3 log10 of T cells in full-sized single-donor plateletpheresis units. Using 1.0 Joule/cm2 UVA, the lowest dose of S-59, AMT and 8-MOP required to reduce the number of T cells to the limit of detection was 0.05 micromol/L, 1.0 micromol/L, and 10.0 micromol/L, respectively. (2) Cytokine synthesis: Treatment with 1.9 Joules/cm2 UVA and 150 micromol/L S-59 or AMT completely inhibited synthesis of the cytokine IL-8 by contaminating leukocytes during 5 days of platelet storage. After treatment with 75 micromol/L 8-MOP and 1.9 Joules/cm2 UVA, only low levels of IL-8 were detected. (3) Psoralen-DNA adduct formation: The combination of 1.9 Joules/cm2 UVA and 150 micromol/L S-59, AMT, or 8-MOP induced 12.0 +/- 3.0, 6.0 +/- 0. 9, and 0.7 psoralen adducts per 1,000 bp DNA, respectively. (4) Replication competence: Polymerase chain reaction (PCR) amplification of small genomic DNA sequences (242-439 bp) after PCT was inhibited. The degree of PCR amplification inhibition correlated with the level of adduct formation (S-59 > AMT > 8-MOP). In contrast, 2,500 cGy gamma radiation, a dose that inactivates >5 log10 of T cells in blood products, had minimal effect on cytokine synthesis and did not induce sufficient DNA strand breaks to inhibit PCR amplification of the same small DNA sequences. These results demonstrate that leukocytes are sensitive to PCT with psoralens and among the psoralens tested S-59 is the most effective. Therefore, PCT has the potential to reduce the incidence of leukocyte-mediated adverse immune reactions associated with platelet transfusion.     

Cell cycle effects and concomitant p53 expression in hairless murine skin after longwave UVA (365 nm) irradiation: a comparison with UVB irradiation

Ultraviolet A (UVA, 315-400 nm) radiation is known to be a complete carcinogen, but in contrast to UVB (280-315 nm) radiation, much of the cell damage is oxygen dependent (mediated through reactive oxygen species), and the dominant premutational DNA lesion(s) remains to be identified. To investigate further the basic differences in UVA and UVB carcinogenesis, we compared in vivo cellular responses, viz. cell cycle progression and transient p53 expression in the epidermis, after UVA1 (340-400 nm) exposure with those after broadband UVB exposure of hairless mice. Using flow cytometry we found a temporary suppression of bromodeoxyuridine (BrdU) uptake in S-phase cells both after UVB and UVA1 irradiation, which only in the case of UVB is followed by an increase to well over control levels. With equally erythemogenic doses (1-2 MED), the modulation of BrdU uptake was more profound after UVB than after UVA1 irradiation. Also, a marked transient increase in the percentage of S-phase cells occurred both after UVB and after UVA1 irradiation, but this increase evolved more rapidly after UVA1 irradiation. Further, p53 expression increased both after UVB and UVA1 irradiations, with peak expression already occurring from 12 to 24 h after UVA1 exposure and around 24 h after UVB exposure. Overall, UVA1 radiation appears to have less of an impact on the cell cycle than UVB radiation, as measured by the magnitude and duration of changes in DNA synthesis and cells in S phase. These differences are likely to reflect basic differences between UVB and UVA1 in genotoxicity and carcinogenic action.     

241Am removal by DTPA vs. occurrence of skeletal malignancy

PURPOSE: The distribution of hyaluronan (HA) and the cellular response after photokeratitis induced by different ultraviolet (UV) wavelengths in the rabbit cornea was examined to help understand the mechanism of corneal injury and repair after UV damage. HA is a high molecular weight disaccharide polymer capable of binding considerable amounts of water. It is not normally found in the rabbit corneal stroma. The production of HA represents a generalized corneal response to injury. METHODS: Twenty-four albino rabbit corneas were exposed to 270, 290, and 310 nm of UV radiant energy in 8-nm full wavebands in doses producing biomicroscopically significant keratitis (three corneal thresholds for keratitis (Hc): 0.016 J/cm2 for 270 nm, 0.04 J/cm2 for 290 nm, and 0.14 J/cm2 for 310 nm) and in subkeratitis doses (0.7 Hc: 0.004 J/cm2 for 270 nm, 0.008 J/cm2 for 290 nm, and 0.03 J/cm2 for 310 nm). The rabbits exposed to 270 and 290 nm of UV radiation were sacrificed 3 days after exposure. The rabbits exposed to 310 nm of UV radiation were sacrificed 3, 7, and 14 days after exposure, respectively. The corneal tissue specimens were double stained and examined morphologically and histochemically for HA by light microscopy. RESULTS: Evaluation of corneas exposed to 270 and 290 nm of UV radiant energy in both subkeratitis and keratitis doses and those corneas exposed to 310 nm of radiant energy in subkeratitis dose showed neither stromal changes nor production of HA by corneal cells. Corneas exposed to 310 nm of UV radiant energy in keratitis dose at 3 days after exposure showed disappearance of keratocytes in entire thickness of central cornea. Cells bordering this damaged area were staining for HA. By 7 days after exposure almost the whole damaged area, except one fourth of anterior stroma, was repopulated by new keratocytes staining positive for HA. The corneal structures became normal and HA almost completely disappeared 14 days after exposure. CONCLUSIONS: A keratitis dose of 310 nm of UV light irradiation is needed to cause keratocyte damage. A keratitis dose of the shorter wavelengths does not cause keratocyte cell damage at the light microscopic level. The keratocyte production of HA appears to be a sign of cell readiness to repopulate the damaged stroma devoid of keratocytes.     

Oxidative inactivation of glutamine synthetase from the cyanobacterium Anabaena variabilis

In crude extracts of the cyanobacterium Anabaena variabilis, glutamine synthetase (GS) could be effectively inactivated by the addition of NADH. GS inactivation was completed within 30 min. Both the inactivated GS and the active enzyme were isolated. No difference between the two enzyme forms was seen in sodium dodecyl sulfate-gels, and only minor differences were detectable by UV spectra, which excludes modification by a nucleotide. Mass spectrometry revealed that the molecular masses of active and inactive GS are equal. While the Km values of the substrates were unchanged, the Vmax values of the inactive GS were lower, reflecting the inactivation factor in the crude extract. This result indicates that the active site was affected. From the crude extract, a fraction mediating GS inactivation could be enriched by ammonium sulfate precipitation and gel filtration. GS inactivation by this fraction required the presence of NAD(P)H, Fe3+, and oxygen. In the absence of the GS-inactivating fraction, GS could be inactivated by Fe2+ and H2O2. The GS-inactivating fraction produced Fe2+ and H2O2, using NADPH, Fe3+, and oxygen. Accordingly, the inactivating fraction was inhibited by catalase and EDTA. This GS-inactivating system of Anabaena is similar to that described for oxidative GS inactivation in Escherichia coli. We conclude that GS inactivation by NAD(P)H is caused by irreversible oxidative damage and is not due to a regulatory mechanism of nitrogen assimilation.     

Adhesion of leukocytes to dermal endothelial cells is induced after single-dose, but reduced after repeated doses of UVA

Approximately 20-50% of ultraviolet A (UVA) irradiation delivered to the skin surface may reach the human dermal microvascular endothelial cells (HDMEC) that play a pivotal role in cellular inflammatory tissue; however, the pathophysiologic role of HDMEC in UVA-induced skin changes is largely unknown. Based on previous in vivo and in vitro studies revealing UVA-induced expression of endothelial adhesion molecules, we studied isolated HDMEC under various conditions in order to further delineate the impact of UVA on these cells. The expression of cell adhesion molecules was determined by flow cytometry and the resulting changes of stable adhesion of leukocytes to endothelial cells were quantitated for granulocytes, lymphocytes, and monocytes using a newly developed multicellular adhesion assay. Additionally, antibody blocking experiments were performed to delineate the role of individual cell adhesion molecules in UVA-induced leukocyte adherence. High-dose polychromatic UVA (25 J per cm2, maximal emission at 375 nm) induced intercellular adhesion molecule-1 and E-selectin with different kinetics but correlating the adhesion of leukocyte subsets. This effect subsided, however, in the course of 3-6 daily applied UVA doses. Moreover, pro-inflammatory cytokine challenge by tumor necrosis factor-alpha and interleukin-1-alpha resulted in significantly weaker induction of intercellular adhesion molecule-1 and E-selectin in repeatedly UVA-exposed HDMEC. Differential quantitation of peripheral blood derived granulocytes, lymphocytes, and monocytes revealed reduced adhesion particularly of lymphocytes followed by monocytes and granulocytes compared with leukocyte adhesion to nonirradiated but cytokine-stimulated HDMEC. It is concluded that UVA substantially influences endothelial cell adhesion molecules expression and thus directly interferes with leukocyte adhesion to endothelial cells. Divergent UVA-induced effects in this respect can be attributed to the mode of UV exposure as well as to the condition of endothelial cells prior to UVA exposure.     

Physiological effects of ultraviolet light on Dictyostelium discoideum spore germination

The spore germination in Dictyostelium discoideum consists of four stages: activation, postactivation lag, swelling and emergence. Ultraviolet irradiation (total fluence of 250 J/m(2)) of spores at any time prior to late spore swelling allows full swelling, but inhibits the emergence of myxamoebae. In the case of freshly activated spores, a UV exposure time of 30 s (total fluence of 50 J/m(2)) is sufficient to reduce emergence to about 6% when measured after 24 h of incubation. This same fluence results in about 10% viability as measured by plaque forming ability. Experiments utilizing "fractionated exposures' result in the same percentage inhibition of emergence as that found for "single exposures' provided the total fluence is equivalent. The higher fluences (250 J/m(2)) which completely prevent emergence, do not affect the endogenous oxygen uptake of spores during swelling. Ultraviolet light irradiated spores respond to the same activation and deactivation treatments as control unirradiated spores. Ultraviolet irradiation after late spore swelling allows emergence to occur in only a small fraction of the population. This fraction of cells which can emerge after UV treatment is said to have passed a "competence point', which is believed to be the time when all the events necessary for emergence have been completed. Though the sites of UV inactivation in spores can only be postulated at present, it is apparent that the initial stages of germination (activation, postactivation lag and spore swelling) occur independently of the UV sensitive sites. The final stage of germination (emergence), however, is dependent on UV sensitive functions.     

Determination of Ag ions in water, urine and blood

Isolated human polymorphonuclear leukocytes were submitted to long wave ultraviolet light (UVA) with and without preincubation of the cells with 8-methoxypsoralen (8-MOP). Leukocyte chemotaxis was determined in modified Boyden chambers using caseine as the attractant. Combined treatment (8-MOP + UVA) significantly inhibited the chemotactic activity with 8-MOP concentrations above 0.1 mug/ml and 2 J/cm(2). However, with high dosage of UVA (5 J/cm(2)) with and without 8-MOP still 25-30% cells migrated through the filters. Also, cell viability as determined by trypan blue exclusion was only moderately affected by combined treatment. The results indicate that these nonreplicating cells are comperatively insensitive to UVA and 8-MOP.     

Radiation sensitivity of an Escherichia coli mutant lacking NADP+-dependent isocitrate dehydrogenase

Ionizing radiation induces the production of reactive oxygen species, which play an important causative role in radiation damage. NADP+-dependent isocitrate dehydrogenase (ICDH) in Escherichia coli produces NADPH, an essential reducing equivalent for the antioxidant system. The protective role of ICDH against ionizing radiation in E. coli was investigated in wild-type and ICDH-deficient strains. Upon exposure to ionizing radiation, the viability was lower and the lipid peroxidation was higher in mutant cells compared to wild-type cells. Activities of key antioxidant enzymes such as superoxide dismutase, catalase, glutathione reductase, and glucose-6-phosphate dehydrogenase were decreased by irradiation in both cells. Results suggest that ICDH plays an important role as an antioxidant enzyme in cellular defense against ionizing radiation.     

Quinolone antibiotic photodynamic production of 8-oxo-7, 8-dihydro-2'-deoxyguanosine in cultured liver epithelial cells

To study the basis for the phototoxicity of quinolones, a class of synthetic antibacterials, the photodynamic ability to mediate 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) formation in cultured cells was measured for lomefloxacin (LMX), which is strongly associated with clinical phototoxicity in humans, and ciprofloxacin (CFX), which has few reports of phototoxicity. Adult rat liver (ARL-18) cells were exposed to the quinolones in the presence of UVA and DNA was extracted and analyzed by HPLC with electrochemical detection. Low levels of 8-oxo-dG were found in the DNA of nonirradiated ARL-18 cells and this was increased up to 6-fold in the presence of either LMX (50-400 microM) or up to 3.6-fold in the presence of CFX (50-400 microM) and UVA (20 J/cm2) when compared to the UVA control. Comparing separate experiments with LMX and CFX, LMX produced greater levels of 8-oxo-dG either after dark exposure or after UVA exposure at 20 J/cm2. Also, LMX and CFX were both shown to photodegrade in the presence of UVA, and it was determined that UVA photoinstability alone does not reflect phototoxic potential. These data suggest that the photodynamic potential of LMX and CFX to produce 8-oxo-dG may relate to their human clinical phototoxicity profile. We suggest that the observed clinical phototoxicity is mediated through a UVA photodynamic effect on the quinolone to form reactive oxygen species in the presence of molecular oxygen. The findings indicate that 8-oxo-dG formation can serve as a marker for the potential phototoxicity of new quinolones.