RESUMO
Renal fibrosis (RF) is a common pathological feature of chronic kidney disease (CKD), which remains a major public health problem. As now, there is still lack of chemical or biological drugs to reverse RF. Shen-shuai-yi Recipe (SSYR) is a classical Chinese herbal formula for the treatment of CKD. However, the effects and mechanisms of SSYR in treating RF are still not clear. In this study, the active constituents SSYR for treating RF were explored by UHPLC-Q-Orbitrap HRMS. Bioinformatics analyses were employed to analyze the key pharmacological targets and the core active constituents of SSYR in the treatment of RF. In experimental validation, vehicle or SSYR at doses of 2.12 g/kg/d and 4.25 g/kg/d were given by orally to unilateral ureteric obstruction (UUO) mice. 13 days after treatment, we detected the severity of renal fibrosis, extracellular collagen deposition and pre-fibrotic signaling pathways. Bioinformatics analysis suggested that signal transducer and activator of transcription 3 (STAT3) was the core target and lenticin, luteolin-7-O-rutinoside, hesperidin, kaempferol-3-O-rutinoside, and 3,5,6,7,8,3',4'-heptamethoxyflavone were the key constituents in SSYR for treating RF. SSYR significantly reduced the expressions of fibronectin (FN), α-smooth muscle actin (α-SMA), collagen-I and alleviated renal interstitial collagen deposition in UUO kidneys. In mechanism, SSYR potently blocked the phosphorylation of STAT3 and Smad3 and suppressed the expression of connective tissue growth factor (CTGF). Collectively, SSYR can ameliorate RF via inhibiting the phosphorylation of STAT3 and its downstream and reducing the collagen deposition, suggesting that SSYR can be developed as a novel medicine for treating RF.
RESUMO
Introduction: Alisol B 23-acetate (AB23A), a major bioactive constituent in the Chinese herb Zexie (Rhizoma Alismatis), has been found with multiple pharmacological activities. AB23A can be readily hydrolyzed to alisol B in mammals, but the hydrolytic pathways of AB23A in humans and the key enzymes responsible for AB23A hydrolysis are still unrevealed. This study aims to reveal the metabolic organs and the crucial enzymes responsible for AB23A hydrolysis in human biological systems, as well as to decipher the impact of AB23A hydrolysis on its biological effects. Methods: The hydrolytic pathways of AB23A in human plasma and tissue preparations were carefully investigated by using Q-Exactive quadrupole-Orbitrap mass spectrometer and LC-UV, while the key enzymes responsible for AB23A hydrolysis were studied via performing a set of assays including reaction phenotyping assays, chemical inhibition assays, and enzyme kinetics analyses. Finally, the agonist effects of both AB23A and its hydrolytic metabolite(s) on FXR were tested at the cellular level. Results: AB23A could be readily hydrolyzed to form alisol B in human plasma, intestinal and hepatic preparations, while human butyrylcholinesterase (hBchE) and human carboxylesterases played key roles in AB23A hydrolysis in human plasma and tissue preparations, respectively. It was also found that human serum albumin (hSA) could catalyze AB23A hydrolysis, while multiple lysine residues of hSA were covalently modified by AB23A, suggesting that hSA catalyzed AB23A hydrolysis via its pseudo-esterase activity. Biological tests revealed that both AB23A and alisol B exhibited similar FXR agonist effects, indicating AB23A hydrolysis did not affect its FXR agonist effect. Discussion: This study deciphers the hydrolytic pathways of AB23A in human biological systems, which is very helpful for deep understanding of the metabolic rates of AB23A in humans, and useful for developing novel prodrugs of alisol B with desirable pharmacokinetic behaviors.
