Liu Hong-Shuang, Li Ya-Lan, Kong Jing-Wei, Zhou Man-Yu, Dong Rui-Juan, Ge Dong-Yu, Liu Jia-Jing, Peng Gui-Ying, Liao Yan
School of Traditional Chinese Medicine, Beijing University of Chinese Medicine Beijing 102488, China.
School of Life Sciences, Beijing University of Chinese Medicine Beijing 102488, China.
Zhongguo Zhong Yao Za Zhi. 2022 Apr;47(7):1913-1920. doi: 10.19540/j.cnki.cjcmm.20211029.401.
This study explored whether Sagittaria sagittifolia polysaccharides(SSP) activates the nuclear factor erythroid-2-related factor2(Nrf2)/heme oxygenase-1(HO-1) signaling pathway to protect against liver damage jointly induced by multiple heavy metals. First, based on the proportion of dietary intake of six heavy metals in rice available in Beijing market, a heavy metal mixture was prepared for inducing mouse liver injury and HepG2 cell injury. Forty male Kunming mice were divided into five groups: control group, model group, glutathione positive control group, and low-and high-dose SSP groups, with eight mice in each group. After 30 days of intragastric administration, the liver injury in mice was observed by HE staining. In the in vitro experiment, MTT assay was conducted to detect the effects of SSP at 0.25, 0.5, 1, and 2 mg·mL~(-1) on HepG2 cell survival at different time points. The content of alanine transaminase(ALT) and aspartate aminotransferase(AST) in the 48-h cell culture fluid was measured using micro-plate cultivation method, followed by the detection of the change in reactive oxygen species(ROS) content by flow cytometry. The mRNA expression levels of Nrf2 and HO-1 in cells were determined by RT-PCR, and their protein expression by Western blot. HE staining results showed that compared with the model group, the SSP administration groups exhibited significantly alleviated inflammatory cell infiltration and fatty infiltration in the liver, with better outcomes observed in the high-dose SSP group. In the in vitro MTT assay, compared with the model group, SSP at four concentrations all significantly increased the cell survival rate, decreased the ALT, AST, and ROS content(P<0.05), and down-regulated Nrf2 and HO-1 mRNA and protein expression(P<0.05). SSP significantly improves inflammatory infiltration in the liver tissue of mice exposed to a variety of heavy metals and corrects the liver fat degeneration, which may be related to its regulation of the Nrf2/HO-1 signaling pathway, reduction of ROS, and alleviation of oxidative damage.
本研究探讨了慈姑多糖（SSP）是否通过激活核因子红细胞2相关因子2（Nrf2）/血红素加氧酶-1（HO-1）信号通路来预防多种重金属联合诱导的肝损伤。首先，根据北京市场大米中六种重金属的膳食摄入量比例，制备重金属混合物以诱导小鼠肝损伤和HepG2细胞损伤。将40只雄性昆明小鼠分为五组：对照组、模型组、谷胱甘肽阳性对照组以及低剂量和高剂量SSP组，每组8只。灌胃给药30天后，通过苏木精-伊红（HE）染色观察小鼠的肝损伤情况。在体外实验中，采用MTT法检测0.25、0.5、1和2mg·mL⁻¹的SSP在不同时间点对HepG2细胞存活率的影响。使用微孔板培养法测定48小时细胞培养液中丙氨酸转氨酶（ALT）和天冬氨酸转氨酶（AST）的含量，随后通过流式细胞术检测活性氧（ROS）含量的变化。通过逆转录聚合酶链反应（RT-PCR）测定细胞中Nrf2和HO-1的mRNA表达水平，并用蛋白质免疫印迹法检测其蛋白质表达。HE染色结果显示，与模型组相比，SSP给药组肝脏中的炎性细胞浸润和脂肪浸润明显减轻，高剂量SSP组效果更佳。在体外MTT实验中，与模型组相比，四个浓度的SSP均显著提高细胞存活率，降低ALT、AST和ROS含量（P<0.05），并下调Nrf2和HO-1的mRNA及蛋白质表达（P<\0.05）。SSP可显著改善暴露于多种重金属的小鼠肝组织中的炎性浸润并纠正肝脏脂肪变性，这可能与其对Nrf2/HO-1信号通路的调节、ROS的减少以及氧化损伤的减轻有关。
