Gamma-irradiation produces active chlorine species (ACS) in physiological solutions: Secoisolariciresinol diglucoside (SDG) scavenges ACS - A novel mechanism of DNA radioprotection

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• 作者：Om P. Mishra^a ; ¹ ; ^{mishra.o@gmail.com" class="auth_mail" title="E-mail the corresponding author} ; Anatoliy V. Popov^b ; ¹ ; ^{avpopov@mail.med.upenn.edu" class="auth_mail" title="E-mail the corresponding author} ; Ralph A. Pietrofesa^a ; ^{ralphp@mail.med.upenn.edu" class="auth_mail" title="E-mail the corresponding author} ; Melpo Christofidou-Solomidou^a ; ^{melpo@mail.med.upenn.edu" class="auth_mail" title="E-mail the corresponding author}
• 关键词：2-AP ; 2-aminopurine ; ACS ; active chlorine species ; ANOVA ; analysis of variance ; APF ; 3&prime ; -(p-aminophenyl) fluorescein ; Cl ; chlorine atom ; Cl&minus ;   ; chloride anion ; Cl2 ; chlorine ; Cl2&minus ;   ; dichloro radical anion ; ClO&minus ;   ; hypochlorite ion ; H2O2 ; hydrogen peroxide ; HOCl ; hypochlorous acid ; HPF ; 3&prime ; -(p-hydroxyphenyl) fluorescein ; &bull ; OH ; hydroxyl radical ; OC ; open-circular DNA ; PBS ; phosphate-buffered saline ; RFU ; relative fluorescence units ; ROS ; reactive oxygen
• 刊名：Biochimica et Biophysica Acta (BBA) - General Subjects
• 出版年：2016
• 出版时间：September 2016
• 年：2016
• 卷：1860
• 期：9
• 页码：1884-1897
• 全文大小：1957 K

文摘

Secoisolariciresinol diglucoside (SDG), the main lignan in whole grain flaxseed, is a potent antioxidant and free radical scavenger with known radioprotective properties. However, the exact mechanism of SDG radioprotection is not well understood. The current study identified a novel mechanism of DNA radioprotection by SDG in physiological solutions by scavenging active chlorine species (ACS) and reducing chlorinated nucleobases.

Methods

The ACS scavenging activity of SDG was determined using two highly specific fluoroprobes: hypochlorite-specific 3′-(p-aminophenyl) fluorescein (APF) and hydroxyl radical-sensitive 3′-(p-hydroxyphenyl) fluorescein (HPF). Dopamine, an SDG structural analog, was used for proton ¹H NMR studies to trap primary ACS radicals. Taurine N-chlorination was determined to demonstrate radiation-induced generation of hypochlorite, a secondary ACS. DNA protection was assessed by determining the extent of DNA fragmentation and plasmid DNA relaxation following exposure to ClO⁻ and radiation. Purine base chlorination by ClO⁻ and γ-radiation was determined by using 2-aminopurine (2-AP), a fluorescent analog of 6-aminopurine. Results: Chloride anions (Cl⁻) consumed > 90% of hydroxyl radicals in physiological solutions produced by γ-radiation resulting in ACS formation, which was detected by ¹H NMR. Importantly, SDG scavenged hypochlorite- and γ–radiation-induced ACS. In addition, SDG blunted ACS-induced fragmentation of calf thymus DNA and plasmid DNA relaxation. SDG treatment before or after ACS exposure decreased the ClO⁻ or γ-radiation-induced chlorination of 2-AP. Exposure to γ-radiation resulted in increased taurine chlorination, indicative of ClO⁻ generation. NMR studies revealed formation of primary ACS radicals (chlorine atoms (Cl^{radical dot" data-inlimg="/entities/rad" src="/sd/grey_pxl.gif" class="glyphImg imgLazyJSB">}) and dichloro radical anions (Cl₂¯^{radical dot" data-inlimg="/entities/rad" src="/sd/grey_pxl.gif" class="glyphImg imgLazyJSB">})), which were trapped by SDG and its structural analog dopamine.

Conclusion

We demonstrate that γ-radiation induces the generation of ACS in physiological solutions. SDG treatment scavenged ACS and prevented ACS-induced DNA damage and chlorination of 2-aminopurine. This study identified a novel and unique mechanism of SDG radioprotection, through ACS scavenging, and supports the potential usefulness of SDG as a radioprotector and mitigator for radiation exposure as part of cancer therapy or accidental exposure.