DNA strand break dependence on Tris and arginine scavenger concentrations under ultra-soft X-ray irradiation: the contribution of secondary arginine radicals

Abstract

In this study, we used a bench-top cold-cathode ultra-soft X-ray (USX) generator to expose aqueous DNA plasmid solutions to low-LET radiation under various scavenging conditions. Single- and double-strand breaks were assessed using classic gel electrophoresis quantification of linear, circular and supercoiled plasmid DNA topologies. With their very low penetration range in water, USX can only interact with matter up to short distances, of the order of 50 μm. We validated a stirring procedure which makes it possible to expose 100 µL of aqueous samples (2 mm thick). The scavenging of OH radicals by Tris buffer was studied at ambient temperature under aerobic conditions and compared to data gathered in the literature. A very good agreement was found with the rare data dealing with DNA plasmid exposed to Al Kα photons at low temperature (T ≤ 277 K), which therefore validated the experimental procedure. The yields for DNA single-strand breaks determined during this study enabled the ratio of indirect to direct effects to be determined at 96.2%, in good agreement with the value of 97.7% stemming from a study based on γ-ray irradiation of frozen solutions of plasmid DNA. Then, arginine was used both to create a "biological-like" chemical environment around the DNA plasmids and as an OH radical scavenger, in vitro. Although arginine has a greater scavenging (protecting) power than Tris, surprisingly, it led to higher rates of strand breakage. Based on the specific binding modes of arginine to DNA, we suggest that the side effects observed are due to the presence of arginine near to, but also inside, the DNA double helix.

The contribution of hydrogen peroxide to the radiosensitizing effect of gold nanoparticles

“…The irradiation technique was described in details elsewhere [31]. Briefly described, the aqueous solution has a 2 mm thickness and 8 mm diameter (total volume of 100 µL), the irradiation was generated with a cold cathode source (1.5 keV, Al Kα).…”

“…Briefly described, the aqueous solution has a 2 mm thickness and 8 mm diameter (total volume of 100 µL), the irradiation was generated with a cold cathode source (1.5 keV, Al Kα). The irradiation of DNA was performed at controlled constant room temperature (295 K) and at constant 98% RH (humidity) [31]. Irradiation of 100 µL of plasmid aqueous solution was performed using a cylindrical 8 mm-diameter sample holder under constant stirring rate of 110 RPM.…”

Complexation des acides aminés basiques arginine, histidine et lysine avec l’ADN plasmidique en solution aqueuse : participation à la capture de radicaux sous irradiation X à 1,5 keV

“…Sur cette figure, nous avons présenté aussi les valeurs expérimentales obtenues dans un travail antérieur [12] en cours d'édition et qui sont relatives aux taux de cassures simple. plasmide -1 .…”

“…L'agitation permet d'assurer un dépôt homogène de la dose dans les 100 μL de solution (2 mm d'épaisseur). L'efficacité de l'agitation sur le dépôt de dose dans la solution aqueuse a été vérifiée de deux façons complémentaires [12]. D'une part, l'utilisation d'ADN plasmidique comme sonde lors d'expériences d'irradiation à durée fixe mais avec des vitesses d'agitation (ω) variant de 0 à 120 révolutions.minute -1 a montré que les taux de cassures simple et double brin croissent avec la vitesse d'agitation et atteignent un palier pour ω > ~80 rev.min -1 .…”

“…Les solutions dosimétriques sont préparées fraîchement avant l'expérience, la mesure de dose est effectuée à l'aide d'un spectrophotomètre UV-visible (CARY 100 Scan, VARIAN). L'ensemble du protocole de mesure ainsi que la méthode sont détaillés dans une publication récente de notre groupe [12]. …”