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Abstract：
BACKGROUND: Polygonum multiflorum (PM), a widely used traditional Chinese medicine herb, is divided into two forms, namely raw polygonum multiflorum (RPM) and polygonum multiflorum praeparata (PMP), according to the processing procedure. Emerging data has revealed the differential hepatotoxicity of RPM and PMP, however, its potential mechanism is still unclear.
METHODS: In our study, we investigated the differential hepatotoxicity of RPM and PMP exerted in C57BL/6 mice. First, sera were collected for biochemical analysis and HE staining was applied to examine the morphological alternation of the liver. Then we treated L02 cells with 5 mg / mL of RPM or PMP. The CCK8 and EdU assays were utilized to observe the viability and proliferation of L02 cells. RNA sequencing was performed to explore the expression profile of L02 cells. Western blotting was performed to detect the expression level of ferroptosis-related protein. Flow cytometry was used to evaluate ROS accumulation.
RESULTS: In our study, a significant elevation in serum ALT, AST and TBIL levels was investigated in the RMP group, while no significant differences were observed in the PMP group, compared to that of the CON group. HE staining showed punctate necrosis, inflammatory cell infiltration and structural destruction can be observed in the RPM group, which can be significantly attenuated after processing. In addition, we also found RPM could decrease the viability and proliferation capacity of L02 cells, which can be reversed by ferroptosis inhibitor. RNA sequencing data revealed the adverse effect of PM exerted on the liver is closely associated with ferroptosis. Western blotting assay uncovered the protein level of GPX4, HO-1 and FTL was sharply decreased, while the ROS content was dramatically elevated in L02 cells treated with RPM, which can be partially restored after processing.
CONCLUSIONS: The hepatotoxicity induced by RPM was significantly lower than the PMP, and its potential mechanism is associated with ferroptosis.
METHODS: In our study, we investigated the differential hepatotoxicity of RPM and PMP exerted in C57BL/6 mice. First, sera were collected for biochemical analysis and HE staining was applied to examine the morphological alternation of the liver. Then we treated L02 cells with 5 mg / mL of RPM or PMP. The CCK8 and EdU assays were utilized to observe the viability and proliferation of L02 cells. RNA sequencing was performed to explore the expression profile of L02 cells. Western blotting was performed to detect the expression level of ferroptosis-related protein. Flow cytometry was used to evaluate ROS accumulation.
RESULTS: In our study, a significant elevation in serum ALT, AST and TBIL levels was investigated in the RMP group, while no significant differences were observed in the PMP group, compared to that of the CON group. HE staining showed punctate necrosis, inflammatory cell infiltration and structural destruction can be observed in the RPM group, which can be significantly attenuated after processing. In addition, we also found RPM could decrease the viability and proliferation capacity of L02 cells, which can be reversed by ferroptosis inhibitor. RNA sequencing data revealed the adverse effect of PM exerted on the liver is closely associated with ferroptosis. Western blotting assay uncovered the protein level of GPX4, HO-1 and FTL was sharply decreased, while the ROS content was dramatically elevated in L02 cells treated with RPM, which can be partially restored after processing.
CONCLUSIONS: The hepatotoxicity induced by RPM was significantly lower than the PMP, and its potential mechanism is associated with ferroptosis.
摘要：
背景：何首乌（PM），一种广泛使用的中草药，分为两种形式，即生何首乌（RPM）和何首乌（PMP），根据处理程序。新兴数据揭示了RPM和PMP的不同肝毒性，然而,其潜在机制尚不清楚。
方法：在我们的研究中，我们研究了在C57BL/6小鼠中施加的RPM和PMP的不同肝毒性。首先,收集血清进行生化分析，并应用HE染色检查肝脏的形态变化。然后我们用5mg/mLRPM或PMP处理L02细胞。利用CCK8和EdU测定来观察L02细胞的活力和增殖。进行RNA测序以探索L02细胞的表达谱。蛋白质印迹法检测铁凋亡相关蛋白的表达水平。流式细胞术用于评估ROS积累。
结果：在我们的研究中，血清ALT显著升高，在RMP组中研究了AST和TBIL水平，虽然在PMP组中没有观察到显着差异，与CON组相比。HE染色显示为点状坏死，在RPM组中可以观察到炎症细胞浸润和结构破坏，处理后可以显着减弱。此外,我们还发现RPM可以降低L02细胞的活力和增殖能力，可以通过铁凋亡抑制剂逆转。RNA测序数据显示，PM对肝脏的不利影响与铁凋亡密切相关。Westernblotting检测发现GPX4、HO-1和FTL蛋白水平急剧下降，而ROS含量在用RPM处理的L02细胞中显著升高，处理后可以部分恢复。
结论：RPM引起的肝毒性明显低于PMP，其潜在机制与铁凋亡有关。
方法：在我们的研究中，我们研究了在C57BL/6小鼠中施加的RPM和PMP的不同肝毒性。首先,收集血清进行生化分析，并应用HE染色检查肝脏的形态变化。然后我们用5mg/mLRPM或PMP处理L02细胞。利用CCK8和EdU测定来观察L02细胞的活力和增殖。进行RNA测序以探索L02细胞的表达谱。蛋白质印迹法检测铁凋亡相关蛋白的表达水平。流式细胞术用于评估ROS积累。
结果：在我们的研究中，血清ALT显著升高，在RMP组中研究了AST和TBIL水平，虽然在PMP组中没有观察到显着差异，与CON组相比。HE染色显示为点状坏死，在RPM组中可以观察到炎症细胞浸润和结构破坏，处理后可以显着减弱。此外,我们还发现RPM可以降低L02细胞的活力和增殖能力，可以通过铁凋亡抑制剂逆转。RNA测序数据显示，PM对肝脏的不利影响与铁凋亡密切相关。Westernblotting检测发现GPX4、HO-1和FTL蛋白水平急剧下降，而ROS含量在用RPM处理的L02细胞中显著升高，处理后可以部分恢复。
结论：RPM引起的肝毒性明显低于PMP，其潜在机制与铁凋亡有关。
