Timiri Shanmugam Prakash Srinivasan, Nair Renjith Parameshwaran, De Benedetti Arrigo, Caldito Gloria, Abreo Fleurette, Sunavala-Dossabhoy Gulshan
Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, 1501 Kings Highway, Shreveport, LA 71130, USA.
Department of Computational Biology and Bioinformatics, Louisiana State University Health Sciences Center, 1501 Kings Highway, Shreveport, LA 71130, USA.
Free Radic Biol Med. 2016 Apr;93:217-26. doi: 10.1016/j.freeradbiomed.2015.12.029. Epub 2016 Feb 15.
Accidental or medical radiation exposure of the salivary glands can gravely impact oral health. Previous studies have shown the importance of Tousled-like kinase 1 (TLK1) and its alternate start variant TLK1B in cell survival against genotoxic stresses. Through a high-throughput library screening of natural compounds, the phenolic phytochemical, gallic acid (GA), was identified as a modulator of TLK1/1B. This small molecule possesses anti-oxidant and free radical scavenging properties, but in this study, we report that in vitro it promotes survival of human salivary acinar cells, NS-SV-AC, through repair of ionizing radiation damage. Irradiated cells treated with GA show improved clonogenic survival compared to untreated controls. And, analyses of DNA repair kinetics by alkaline single-cell gel electrophoresis and γ-H2AX foci immunofluorescence indicate rapid resolution of DNA breaks in drug-treated cells. Study of DR-GFP transgene repair indicates GA facilitates homologous recombinational repair to establish a functional GFP gene. In contrast, inactivation of TLK1 or its shRNA knockdown suppressed resolution of radiation-induced DNA tails in NS-SV-AC, and homology directed repair in DR-GFP cells. Consistent with our results in culture, animals treated with GA after exposure to fractionated radiation showed better preservation of salivary function compared to saline-treated animals. Our results suggest that GA-mediated transient modulation of TLK1 activity promotes DNA repair and suppresses radiation cytoxicity in salivary gland cells.
唾液腺意外或因医疗而受到辐射会严重影响口腔健康。先前的研究已经表明类Tousled激酶1（TLK1）及其可变起始变体TLK1B在细胞抵抗基因毒性应激的存活过程中的重要性。通过对天然化合物进行高通量文库筛选，酚类植物化学物质没食子酸（GA）被鉴定为TLK1/1B的调节剂。这种小分子具有抗氧化和清除自由基的特性，但在本研究中，我们报告其在体外可通过修复电离辐射损伤来促进人唾液腺腺泡细胞NS-SV-AC的存活。与未处理的对照相比，用GA处理的受辐射细胞显示出更好的克隆形成存活率。并且，通过碱性单细胞凝胶电泳和γ-H2AX焦点免疫荧光对DNA修复动力学进行分析表明，药物处理的细胞中DNA断裂能快速修复。对DR-GFP转基因修复的研究表明，GA促进同源重组修复以建立功能性GFP基因。相反，TLK1的失活或其shRNA敲低会抑制NS-SV-AC中辐射诱导的DNA尾巴的修复以及DR-GFP细胞中的同源定向修复。与我们在培养中的结果一致，与盐水处理的动物相比，分次辐射后用GA处理的动物唾液功能保留得更好。我们的结果表明，GA介导的TLK1活性的瞬时调节可促进唾液腺细胞中的DNA修复并抑制辐射细胞毒性。
