The traditional Chinese herbal medicine Eupatorium fortunei Turcz. (E. fortunei) has been widely adopted to treat nausea, diabetes, siriasis, and poor appetite. However, E. fortunei contains multiple pyrrolizidine alkaloids (PAs). This study aimed to investigate the hepatotoxicity of total alkaloids in E. fortunei (EFTAs) and identify the toxic mechanisms of EFTAs on hepatocytes. Liquid chromatography with a tandem mass spectrometry assay with reference standards indicated that EFTAs mainly consisted of eight PAs whose content accounted for 92.38% of EFTAs. EFTAs markedly decreased mouse body and liver weights and increased the contents of AST and ALT. The histopathological assays demonstrated that, after exposition to EFTAs, the structures of hepatocytes were damaged and the fibrosis and apoptosis in hepatocytes were accelerated. Moreover, EFTAs increased the serum level of inflammatory cytokines and aggravated circulating oxidative stress. A combination of hepatic proteomics and metabolomics was used to investigate the toxic mechanisms of EFTAs. The study revealed that EFTAs seriously disrupted glycerophospholipid metabolism by upregulating the contents of lysophosphatidylglycerol acyltransferase 1 and phosphatidylinositol and downregulating the contents of choline/ethanolamine kinase beta, choline-ethanolamine phosphotransferase 1, phospholipase D4, 1-acylglycerophosphocholine, phosphatidylcholine, and dihydroxyacetone phosphate in the liver, resulting in detrimental inflammation, fibrosis, and apoptosis. This study revealed that EFTAs induced severe hepatotoxicity by disrupting glycerophospholipid metabolism.

Although doxorubicin (DOX) is a highly efficient antineoplastic chemotherapeutic drug, it greatly increases the risk of heart damage. Salvianolic acid A (SAA), a water-soluble active component from traditional Chinese medicine Salvia miltiorrhiza Bunge, is shown to exert various cardioprotective effects. However, the potential of SAA in protecting DOX-induced cardiotoxicity (DIC) and its underlying mechanisms have yet to be elucidated. In the current study, we aimed to identify the targets and investigate the cardioprotective mechanism of SAA improving DIC. we established DIC mice to evaluate the cardioprotective effects of SAA by pharmacodynamics, myocardial metabolomic and protomics analysis. Multipe protein-small molecule interaction strategies including surface plasmon resonance, microscale thermophoresis and cellular thermal shift assay, combined with target-depleted cell and animal models were applied for identifying potential targets of SAA. For clinical translation, we also investigated cardioprotective activities of SAA in DOX-treated lewis lung carcinoma-bearing mice. SAA significantly alleviated cardiomyocyte apoptosis and oxidative damage, and improved echocardiographic parameters in DIC mice. In addition, myocardium metabolomic and proteomics analysis revealed that SAA preserved hearts by modulating l-Glutamic acid, l-Aspartic acid, citrate and isocitrate and restoring glutamic-oxaloacetic transaminase 2 (GOT2) expression. With the help of multipe protein-small molecule interaction strategies, GOT2 was identified as one of the targets of SAA, and a strong affinity was characterized with a dissociation constant of 1.712μM. SAA ameliorated DOX-induced oxidative damage, mitochondrial respiration suppression, mitochondrial membrane potential loss and NADH levels, which were validated in GOT2 knockdown H9C2 cells. In GOT2 depleted zebrafish, SAA did not display a protective effect against DIC, while in lewis lung carcinoma-bearing mice, SAA not only improved DIC, but also exerted combined anti-tumor effects with DOX. SAA targets GOT2 to alleviate myocardial oxidative stress by regulating malate-aspartate NADH shuttle, thereby protecting against DIC. Our findings highlighted the therapeutic potential of activating GOT2 in mitigating DIC and the possibility of SAA and DOX-based combination therapy.

Although the cardiovascular-kidney-metabolic (CKM) syndrome framework offers a structured approach to risk stratification, the identification of individual