Wu Shaohua, Tian Chen, Tu Zhengwei, Guo Jianghong, Xu Feifei, Qin Weida, Chang Huajie, Wang Zhiyun, Hu Tong, Sun Xiao, Ning Hongxin, Li Yiliang, Gou Wenfeng, Hou Wenbin
Tianjin Key Laboratory of Radiation Medicine and Molecular Nuclear Medicine, Institute of Radiation Medicine, Peking Union Medical College & Chinese Academy of Medical Sciences, Tianjin, 300192, China.
Tianjin Key Laboratory of Acute Abdomen Disease Associated Organ Injury and ITCWM Repair, Tianjin NanKai Hospital, Tianjin, 300100, China.
J Ethnopharmacol. 2023 Jul 15;311:116428. doi: 10.1016/j.jep.2023.116428. Epub 2023 Mar 28.
Irradiation-induced intestinal injury (RIII) often occurs during radiotherapy in patients, which would result in abdominal pain, diarrhea, nausea, vomiting, and even death. Engelhardia roxburghiana Wall. leaves, a traditional Chinese herb, has unique anti-inflammatory, anti-tumor, antioxidant, and analgesic effects, is used to treat damp-heat diarrhea, hernia, and abdominal pain, and has the potential to protect against RIII.
To explore the protective effects of the total flavonoids of Engelhardia roxburghiana Wall. leaves (TFERL) on RIII and provide some reference for the application of Engelhardia roxburghiana Wall. leaves in the field of radiation protection.
The effect of TFERL on the survival rate of mice was observed after a lethal radiation dose (7.2 Gy) by ionizing radiation (IR). To better observe the protective effects of the TFERL on RIII, a mice model of RIII induced by IR (13 Gy) was established. Small intestinal crypts, villi, intestinal stem cells (ISC) and the proliferation of ISC were observed by haematoxylin and eosin (H&E) and immunohistochemistry (IHC). Quantitative real-time PCR (qRT-PCR) was used to detect the expression of genes related to intestinal integrity. Superoxide dismutase (SOD), reduced glutathione (GSH), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in the serum of mice were assessed. In vitro, cell models of RIII induced by IR (2, 4, 6, 8 Gy) were established. Normal human intestinal epithelial cells HIEC-6 cells were treated with TFERL/Vehicle, and the radiation protective effect of TFERL on HIEC-6 cells was detected by clone formation assay. DNA damage was detected by comet assay and immunofluorescence assay. Reactive oxygen species (ROS), cell cycle and apoptosis rate were detected by flow cytometry. Oxidative stress, apoptosis and ferroptosis-related proteins were detected by western blot. Finally, the colony formation assay was used to detect the effect of TFERL on the radiosensitivity of colorectal cancer cells.
TFERL treatment can increase the survival rate and time of the mice after a lethal radiation dose. In the mice model of RIII induced by IR, TFERL alleviated RIII by reducing intestinal crypt/villi structural damage, increasing the number and proliferation of ISC, and maintaining the integrity of the intestinal epithelium after total abdominal irradiation. Moreover, TFERL promoted the proliferation of irradiated HIEC-6 cells, and reduced radiation-induced apoptosis and DNA damage. Mechanism studies have found that TFERL promotes the expression of NRF2 and its downstream antioxidant proteins, and silencing NRF2 resulted in the loss of radioprotection by TFERL, suggesting that TFERL exerts radiation protection by activating the NRF2 pathway. Surprisingly, TFERL reduced the number of clones of colon cancer cells after irradiation, suggesting that TFERL can increase the radiosensitivity of colon cancer cells.
Our data showed that TFERL inhibited oxidative stress, reduced DNA damage, reduced apoptosis and ferroptosis, and improved IR-induced RIII. This study may offer a fresh approach to using Chinese herbs for radioprotection.
辐射诱导的肠道损伤（RIII）在患者放疗期间经常发生，这会导致腹痛、腹泻、恶心、呕吐，甚至死亡。黄杞叶是一种传统中药，具有独特的抗炎、抗肿瘤、抗氧化和镇痛作用，用于治疗湿热腹泻、疝气和腹痛，并且具有预防RIII的潜力。
探讨黄杞叶总黄酮（TFERL）对RIII的保护作用，并为黄杞叶在辐射防护领域的应用提供一些参考。
通过电离辐射（IR）给予致死剂量（7.2 Gy）的辐射后，观察TFERL对小鼠存活率的影响。为了更好地观察TFERL对RIII的保护作用，建立了由IR（13 Gy）诱导的RIII小鼠模型。通过苏木精-伊红（H&E）染色和免疫组织化学（IHC）观察小肠隐窝、绒毛、肠干细胞（ISC）以及ISC的增殖情况。采用定量实时聚合酶链反应（qRT-PCR）检测与肠道完整性相关基因的表达。评估小鼠血清中的超氧化物歧化酶（SOD）、还原型谷胱甘肽（GSH）、白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）。在体外，建立由IR（2、4、6、8 Gy）诱导的RIII细胞模型。将正常人类肠上皮细胞HIEC-6细胞用TFERL/溶剂处理，通过克隆形成试验检测TFERL对HIEC-6细胞的辐射防护作用。通过彗星试验和免疫荧光试验检测DNA损伤。通过流式细胞术检测活性氧（ROS）、细胞周期和凋亡率。通过蛋白质免疫印迹法检测氧化应激、凋亡和铁死亡相关蛋白。最后，采用克隆形成试验检测TFERL对结肠癌细胞放射敏感性的影响。
TFERL治疗可提高致死剂量辐射后小鼠的存活率和存活时间。在由IR诱导的RIII小鼠模型中，TFERL通过减少肠道隐窝/绒毛结构损伤、增加ISC的数量和增殖以及在全腹照射后维持肠上皮的完整性来减轻RIII。此外，TFERL促进了受辐射的HIEC-6细胞的增殖，并减少了辐射诱导的凋亡和DNA损伤。机制研究发现，TFERL促进NRF2及其下游抗氧化蛋白的表达，并且沉默NRF2导致TFERL失去辐射防护作用，这表明TFERL通过激活NRF2途径发挥辐射防护作用。令人惊讶的是，TFERL减少了照射后结肠癌细胞的克隆数量，这表明TFERL可增加结肠癌细胞的放射敏感性。
我们的数据表明，TFERL抑制氧化应激，减少DNA损伤，减少凋亡和铁死亡，并改善IR诱导的RIII。本研究可能为利用中药进行辐射防护提供一种新方法。
