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INTRODUCTION: Zhen-wu-tang (ZWT) is a traditional Chinese medicine (TCM) formula for the treatment of several kidney diseases. However, due to the complexity of the TCM formula, there is a lack of accurate knowledge of the chemical constituents of ZWT and its bioactive components, as well as in vivo metabolic pathway studies. OBJECTIVES: The chemical composition of ZWT and its bioactive components along with the metabolic pathways were investigated by a combination of chemical profiling and serum pharmacochemistry. METHODS: High-resolution ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry was used to identify the chemical components of ZWT and its bioactive components and metabolites in vivo. RESULTS: As a result, a total of 110 chemical components were identified from ZWT solution, mainly amino acids, alkaloids, gingerols, monoterpene glycosides and terpenoids, and so on. In addition, 24 prototype components and 36 metabolites were detected in rat plasma. Meanwhile, 8 prototype components were detected in rat kidney tissue but no metabolites. Interestingly, 4 of the 28 bioactive components were detected in both plasma and renal tissue, which were atractylenolide III, trimethoxyaconitane, methyl gallate, and paeoniflorin. The metabolic pathways mainly involved Phases I and/or II metabolic reactions such as hydrolysis, oxidation, reduction and hydration, methylation/demethylation, sulphation, glucuronidation, acetylation, and glutathione conjugation. CONCLUSION: Overall, the present study has comprehensively elucidated the chemical composition of ZWT and its potential bioactive components and metabolites, which provides a basis for the basic study of its pharmacodynamic substances and a reference for the study of the bioactive components of TCM formulae.
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BACKGROUND: Macrophages M1 polarization involved in the process of renal inflammatory injury, is a well-established hallmark of chronic kidney disease (CKD). Paeoniflorin (PF), a water-soluble monoterpene glycoside extracted from Paeonia lactiflora, revealed renal anti-inflammatory activities in our previous study. However, the potential molecular mechanism of PF on CKD remains unknown. PURPOSE: The present study aims to investigate the regulation of PF on macrophage polarization in CKD. METHODS: A CKD model was established by cationic bovine serum albumin and a murine macrophage cell line RAW264.7 induced with lipopolysaccharide (LPS) were used to clarify the underlying mechanisms of PF in CKD. RESULTS: Results showed that PF exhibited favorable protective effects on CKD model mice by promoting renal function, ameliorating renal pathological injury and podocyte damage. Furthermore, PF inhibited the infiltration of M1 macrophage marker CD68 and iNOS in kidney tissue, but increased the proportion of M2 macrophage marker CD206. In RAW264.7 cells stimulated with LPS, the levels of cytokines including IL-6, IL-1ß, TNF-α, MCP-1 were lessened under PF treatment, while the levels of Arg1, Fizz1, IL-10 and Ym-1 were augmented. These results indicated that PF promoted macrophage polarization from M1 to M2 in vivo and in vitro. More importantly, PF repaired the damaged mitochondria through increasing mitochondrial membrane potential and reducing ROS accumulation. The mitophagy-related proteins PINK1, Parkin, Bnip3, P62 and LC3 were up-regulated by PF, accompanied by the incremental expressions of Krüppel-like transcription factor 4 (KLF4). Moreover, the promotion of mitophagy and inhibition of M1 macrophage polarization owing to PF were reversed by mitophagy inhibitor Mdivi-1 or silencing KLF4. CONCLUSION: Overall, PF suppressed renal inflammation by promoting macrophage polarization from M1 to M2 and inducing mitophagy via regulating KLF4. It is expected to provide a new strategy for exploring the effects of PF in treating CKD.