RESUMEN
Acute kidney injury (AKI) with maladaptive tubular repair leads to renal fibrosis and progresses to chronic kidney disease (CKD). At present, there is no curative drug to interrupt AKI-to-CKD progression. The nuclear factor of the activated T cell (NFAT) family was initially identified as a transcription factor expressed in most immune cells and involved in the transcription of cytokine genes and other genes critical for the immune response. NFAT2 is also expressed in renal tubular epithelial cells (RTECs) and podocytes and plays an important regulatory role in the kidney. In this study, we investigated the renoprotective effect of 11R-VIVIT, a peptide inhibitor of NFAT, on renal fibrosis in the AKI-to-CKD transition and the underlying mechanisms. We first examined human renal biopsy tissues and found that the expression of NFAT2 was significantly increased in RTECs in patients with severe renal fibrosis. We then established a mouse model of AKI-to-CKD transition using bilateral ischemia-reperfusion injury (Bi-IRI). The mice were treated with 11R-VIVIT (5 mg/kg, i.p.) on Days 1, 3, 10, 17 and 24 after Bi-IRI. We showed that the expression of NFAT2 was markedly increased in RTECs in the AKI-to-CKD transition. 11R-VIVIT administration significantly inhibited the nuclear translocation of NFAT2 in RTECs, decreased the levels of serum creatinine and blood urea nitrogen, and attenuated renal tubulointerstitial fibrosis but had no toxic side effects on the heart and liver. In addition, we showed that 11R-VIVIT administration alleviated RTEC apoptosis after Bi-IRI. Consistently, preapplication of 11R-VIVIT (100 nM) and transfection with NFAT2-targeted siRNA markedly suppressed TGFß-induced HK-2 cell apoptosis in vitro. In conclusion, 11R-VIVIT administration inhibits IRI-induced NFAT2 activation and prevents AKI-to-CKD progression. Inhibiting NFAT2 may be a promising new therapeutic strategy for preventing renal fibrosis after IR-AKI.
Asunto(s)
Lesión Renal Aguda , Insuficiencia Renal Crónica , Daño por Reperfusión , Lesión Renal Aguda/metabolismo , Animales , Fibrosis , Humanos , Isquemia/metabolismo , Riñón/patología , Ratones , Ratones Endogámicos C57BL , Insuficiencia Renal Crónica/metabolismo , Reperfusión , Daño por Reperfusión/complicaciones , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Linfocitos T/metabolismoRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: The integrated effects of herbal medicines were the outcome of all of the inherent components. Currently, few studies have focused on the multicomponent interactions in an herbal medicine to elucidate its pharmacological and/or toxicological effects. In this study, an attempt was made to investigate the interaction between stilbene glucosides and the anthraquinones contained in Radix Polygoni Multiflori (RPM) and to explore the interaction's mechanism from the perspective of UDP-glucuronosyltransferase (UGT) regulation. MATERIALS AND METHODS: The extract of RPM was separated into a stilbene glucoside fraction and a emodin fraction. A rapid high-performance liquid chromatography-mass spectrometry method was developed and validated to disclose the influence of stilbene glucoside on the pharmacokinetics of emodin in rats. Drug and Statistics 2.0 was used for the estimation of the pharmacokinetic parameters. Gene expression analysis in liver and intestinal tissues was performed by a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) method. RESULTS: The analytical method appeared to be suitable for the analysis of emodin with desirable linearity, accuracy, precision and stability, and the total analysis time was less than 2 min on a short column. Glucuronide of emodin, which is the major metabolite of emodin, was determined after ß-glucuronidase hydrolysis. As the in vivo pharmacokinetic studies had indicated, the AUC, Cmax and T1/2 of emodin were increased after the stilbene glucoside treatment, and the glucuronidation of emodin was significantly inhibited. The mRNA levels from UGT1A8 and UGT1A2 were decreased by stilbene glucoside treatment. In contrast, the expression of UGT1A1, UGT1A6 and UGT1A9 mRNA was increased in the liver following treatment. CONCLUSIONS: The influence of stilbene glucoside on the pharmacokinetics of emodin may be attributed to the inhibition of UGT1A8 mRNA expression. Thus, it is important to extend this research to deepen our understanding of the pharmacological and/or toxicological effects of RPM.
Asunto(s)
Emodina/farmacocinética , Glucósidos/farmacología , Glucuronosiltransferasa/genética , Polygonum , Estilbenos/farmacología , Animales , Regulación hacia Abajo , Interacciones Farmacológicas , Emodina/sangre , Glucurónidos/sangre , Mucosa Intestinal/metabolismo , Hígado/metabolismo , Masculino , Extractos Vegetales , Raíces de Plantas , ARN Mensajero/metabolismo , Ratas , Ratas WistarRESUMEN
The emodin-involved hepatotoxicity has been gaining increasing attention. The purpose of the present study was to evaluate the cytotoxicity of emodin on cultured human liver cells (L-02) and predict the possible relation between its cytotoxicity and cellular toxicokinetics. Cell viability and cell damage were assessed by Cell Counting Kit-8 (CCK-8) assay and phase-contrast microscopy, respectively. Cytotoxicity tests demonstrated a concentration- and time-dependent toxic effect of emodin on L-02 cells. Furthermore, emodin at concentration of 30µM led to a significant apoptosis in a time-dependent manner supported by the morphological changes of drug-treated cells. In addition, to elucidate the toxicokinetic characteristics of emodin, a highly sensitive and selective liquid chromatography-mass spectrometry (LC-MS) method was employed and validated for detecting the dynamic alteration of emodin in cells and cell culture media. The proposed method appeared to be suitable for the analysis of emodin with desirable linearity (r(2)>0.99), and satisfying precision being less than 8.7%. The range of recoveries of this method was 90.2-101.9%. The preliminary cellular toxicokinetic study revealed a time-dependent intracellular accumulation of emodin, which was consistent with its in vitro toxic effects. These findings confirmed the cytotoxicity of emodin against L-02 cells and displayed the cytotoxic manner of emodin in terms of its cellular uptake and accumulation in L-02 cells.
Asunto(s)
Cromatografía Liquida/métodos , Medios de Cultivo/química , Emodina/análisis , Emodina/toxicidad , Espectrometría de Masas/métodos , Apoptosis/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Emodina/farmacocinética , Humanos , Hígado/efectos de los fármacos , Microscopía de Contraste de Fase/métodos , FarmacocinéticaRESUMEN
UNLABELLED: Ten promising jujubes were analyzed for textural characteristic (hardness), antioxidant activity, sugar, organic acid, and phenolic profiles. Total phenolic content (TP) measured by Folin-Ciocalteu method ranged from 275.6 to 541.8 mg of gallic acid equivalents per 100 g fresh weight (mg GAE/100 g FW). Four sugars (rhamnose, fructose, sucrose, and glucose), 3 organic acids (malic, citric, and succinic acids), and 11 phenolic compounds (gallic acid, protocatechuic acid, cinnamic acid, chlorogenic acid, caffeic acid, ferulic acid, ellagic acid, catechin, epicatechin, rutin, and quercetin) were identified and quantified by high-performance liquid chromatography in jujube fruits. The results showed that ascorbic acid and proanthocyanidins contents were positively correlated with hardness, and antioxidant activity was well correlated with TP content. Among 10 jujube cultivars, Zizyphus jujuba cv. Qingjianmuzao is good for direct consumption containing high levels of total soluble solids, total sugars, fructose, and glucose, while Zizyphus jujuba cv. Jiaxianmuzao could be an important dietary source of natural antioxidants. PRACTICAL APPLICATION: Genotype is the main factor influencing the composition of bioactive compounds in jujubes. Zizyphus jujuba cv. Qingjianmuzao is good for direct consumption, while Zizyphus jujuba cv. Jiaxianmuzao could be an important dietary source of natural antioxidants for prevention of diseases caused by oxidative stress.