Unrepaired cyclobutane pyrimidine dimers do not prevent proliferation of UV-B-irradiated cultured human fibroblasts

Mutagenic and carcinogenic UV-B radiation is known to damage DNA mostly through the formation of bipyrimidine photoproducts, including cyclobutane dimers (CPD) and (6-4) photoproducts ((6-4) PP). Using high-performance liquid chromatography coupled to tandem mass spectrometry, we investigated the formation and repair of thymine-thymine (TT) and thymine-cytosine (TC) CPD and (6-4) PP in the DNA of cultured human dermal fibroblasts. A major observation was that the rate of repair of the photoproducts did not depend on the identity of the modified pyrimidines. In addition, removal of CPD was found to significantly decrease with increasing applied UV-B dose, whereas (6-4) PP were efficiently repaired within less than 24 h, irrespective of the dose. As a result, a relatively large amount of CPD remained in the genome 48 h after the irradiation. Because the overall applied doses (<500 J m(-2)) were chosen to induce moderate cytotoxicity, fibroblasts could recover their proliferation capacities after transitory cell cycle arrest, as shown by 5-bromo-2'-deoxyuridine (BrdUrd) incorporation and flow cytometry analysis. It could thus be concluded that UV-B-irradiated cultured primary human fibroblasts normally proliferate 48 h after irradiation despite the presence of high levels of CPD in their genome. These observations emphasize the role of CPD in the mutagenic effects of UV-B.     

Molecular responses to stress induced in normal human caucasian melanocytes in culture by exposure to simulated solar UV

Melanocytes play a central role in the response of skin to sunlight exposure. They are directly involved in UV-induced pigmentation as a defense mechanism. However, their alteration can lead to melanoma, a process where the role of sun overexposure is highly probable. The transformation process whereby UV damage may result in melanoma initiation is poorly understood, especially in terms of UV-induced genotoxicity in pigmented cells, where melanin can act either as a sunscreen or as a photosensitizer. The aim of this study was to analyze the behavior of melanocytes from fair skin under irradiation mimicking environmental sunlight in terms of spectral power distribution. To do this, normal human Caucasian melanocytes in culture were exposed to simulated solar UV (SSUV, 300-400 nm). Even at relatively high doses (until 20 min exposure, corresponding to 12 kJ/m2 UV-B and 110 kJ/m2 UV-A), cell death was limited, as shown by cell viability and low occurrence of apoptosis (caspase-3 activation). Moreover, p53 accumulation was three times lower in melanocytes than in unpigmented cells such as fibroblasts after SSUV exposure. However, an important fraction of melanocyte population was arrested in G2-M phase, and this correlated well with a high induction level of the gene GADD45, 4 h after exposure. Among the genes involved in DNA repair, gene XPC was the most inducible because its expression increased more than two-fold 15 h after a 20 min exposure, whereas expression of P48 was only slightly increased. In addition, an early induction of Heme Oxygenase 1 (HO1) gene, a typical response to oxidative stress, was also observed for the first time in melanocytes. Interestingly, this induction remained significant when melanocytes were exposed to UV-A radiation only (320-400 nm), and stimulation of melanogenesis before irradiation further increased HO1 induction. These results were obtained with normal human cells after exposure to SSUV radiation, which mimicked natural sunlight. They provide new data related to gene expression and suggest that melanin in light skin could contribute to sunlight-induced genotoxicity and maybe to melanocyte transformation.     

Mid-infrared trace gas detection using continuous-wave difference frequency generation in periodically poled RbTiOAsO4

A tunable mid-infrared continuous-wave (cw) spectroscopic source in the 3.4–4.5 μm region is reported, based on difference frequency generation (DFG) in a quasi-phase-matched periodically poled RbTiOAsO₄ (PPRTA) crystal, DFG power levels of 10 μW were generated at approximately 4 μm in a 20-mm long PPRTA crystal by mixing two cw single-frequency Ti:Al₂O₃ lasers operating near 713 nm and 871 nm, respectively, using a laser pump power of 300 mW. A quasi-phase-matched infrared wavelength-tuning bandwidth (FWHM) of ∼12 cm^-1 and a temperature tuning rate of 1.02 cm^-1/°C were achieved. Experimental details regarding the feasibility of trace gas detection based on absorption spectroscopy of CO₂ in ambient air using this DFG radiation source are also described. Received: 23 October 2000 / Revised version: 22 January 2001 / Published online: 27 April 2001     

Terahertz spectroscopy applied to the measurement of strengths and self-broadening coefficients for high-J lines of OCS

A spectrometer operating in the 100-2000 GHz range and allowing for absolute line strength measurements has been developed. The continuous wave terahertz radiation is generated by mixing two Ti:Sapphire laser beams in a vertically integrated low temperature grown GaAs (LTG-GaAs) photomixer. Pure rotational lines of ¹⁶O¹²C³²S in the ground vibrational state have been considered for J values up to 90. Observed self-broadening parameters are in agreement with those deduced from infrared experiments. For the first time in the submillimeter range, absolute line strengths have been determined, allowing for a determination of the electric dipole moment in good agreement with the value previously obtained from Stark effect measurements.     

Sterols associated with small unilamellar vesicles (SUVs): intrinsic mobility role for 1H NMR detection

Small unilamellar vesicles (SUVs) of phospholipids are often used as a membrane model system for studying the interaction of molecules. When using NMR under the standard liquid‐state conditions, SUV phospholipid proton spectra can be recorded, exhibiting sharp signals. This is not only because of the fast vesicular tumbling but also because of the combination of this tumbling with the individual motion of the lipids inside the bilayer. This appears evident because addition of cholesterol is responsible of broader resonances because of the slowing down of the lipid motion. On the other hand, no ¹H signal is detected for cholesterol in the bilayer. This lack of detection of the inserted molecules explains why generally SUVs are not considered as a good model for NMR studies under the standard liquid‐state conditions. Here, we use two other sterols in order to demonstrate that an increase of the molecular mobility inside the bilayer could allow the detection of their proton resonances. For desmosterol and lanosterol, which show higher mobility inside the bilayer, with increasing lateral diffusion rates, ¹H sterol signals are detected in contrast to cholesterol. For the fast diffusing lanosterol, no significant improvement in detection is observed using deuterated lipids, demonstrating that homonuclear dipolar coupling is fully averaged out. Furthermore, in the case of low mobility such as for cholesterol, the use of a fast magic angle spinning probe is shown to be efficient to recover the full proton spectrum. Copyright © 2014 John Wiley & Sons, Ltd.     

Difference-frequency laser spectroscopy detection of acetylene trace constituent

5 sbandpectrum. Spectroscopic detection of 28 ppmm/Hz^1/2 was performed. This corresponds to a minimum detectable mixing ratio of ∼1 ppb in conjunction with a 100-m White cell and balanced detection. Received: 3 April 1998/Revised version: 5 June 1998