Guanxin V attenuates myocardial ischaemia reperfusion injury through regulating iron homeostasis.

CONTEXT: Guanxin V (GX), a traditional Chinese medicine formula, is safe and effective in the treatment of coronary artery disease. However, its protective effect on myocardial ischaemia reperfusion injury (MIRI) is unclear. OBJECTIVE: To investigate the cardioprotective effect of GX on MIRI and explore the potential mechanism. MATERIALS AND METHODS: Sprague-Dawley male rats were divided into Sham, MIRI and MIRI + GX groups. GX (6 g/kg) was administered to rats via intragastric administration for seven days before ischaemia reperfusion (IR) surgery. The infarct size, histopathology, serum enzyme activities, ultrastructure of the cardiac mitochondria were assessed. H9c2 cells were pre-treated with GX (0.5 mg/mL), and then exposed to hypoxia/reoxygenation (HR). The cell viability and LDH levels were measured. Network pharmacology was conducted to predict the potential mechanism. The related targets of GX were predicted using the TCMSP database, DrugBank database, etc. Finally, pharmacological experiments were used to validate the predicted results. RESULTS: In vivo, GX significantly reduced the myocardial infarct size from 56.33% to 17.18%, decreased the levels of AST (239.32 vs. 369.18 U/L), CK-MB (1324.61 vs. 2066.47 U/L) and LDH (1245.26 vs. 1969.62 U/L), and reduced mitochondrial damage. In vitro, GX significantly increased H9c2 cell viability (IC50 = 3.913 mg/mL) and inhibited the release of LDH (207.35 vs. 314.33). In addition, GX could maintain iron homeostasis and reduce oxidative stress level by regulating iron metabolism-associated proteins. CONCLUSIONS: GX can attenuate MIRI via regulating iron homeostasis, indicating that GX may act as a potential candidate for the treatment of MIRI.

Potential Plasma Metabolic Biomarkers of Tourette Syndrome Discovery Based on Integrated Nontargeted and Targeted Metabolomics Screening Plasma Metabolic Biomarkers of TS.

Objective: Tourette syndrome (TS) is a chronic neuropsychiatric disorder characterized by abnormal movements, phonations, and tics, but an accurate TS diagnosis remains challenging and indeed depends on its description of clinical symptoms. Our study was conducted to discover and verify some metabolite biomarkers based on nontargeted and targeted metabolomics. Methods: We conducted untargeted ultrahigh-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) for preliminary screening of potential biomarkers on 30 TS patients and 10 healthy controls and then performed validation experiments based on targeted ultrahigh-performance liquid chromatography triple quadrupole-MS (UHPLC/MS/MS) on 35 TS patients and 14 healthy controls. Results: 1775 differentially expressed metabolites were identified by partial least squares discriminant analysis (PLS-DA), fold-change analysis, T-test, and hierarchical clustering analysis (adjusted p value <0.05 and |logFC| > 1). TS plasma samples were found to be differentiated from healthy samples in our approach. Furthermore, aspartate and asparagine metabolism pathways were considered to be a significant enrichment pathway in TS progression based on metabolite pathway enrichment analysis. For the 8 metabolites involved in this pathway that we detected, we then performed validation experiments based on targeted UHPLC/MS/MS. The t-test, Mann-Whitney U test, and receiver operating characteristic (ROC) curve analysis were used to determine potential biomarkers. Ultimately, L-arginine and L-pipecolic acid were validated as significantly differentiated metabolites (p < 0.05), with an AUC of 70.0% and 80.3%, respectively. Conclusion: L-pipecolic acid was defined as a potential biomarker for TS diagnosis by the combined application of nontargeted and targeted metabolomic analysis.

Zi Shen Decoction Inhibits Growth and Metastasis of Lung Cancer via Regulating the AKT/GSK-3ß/ß-Catenin Pathway.

Lung cancer has become the leading cause of cancer-related death worldwide. Oxidative stress plays important roles in the pathogenesis of lung cancer. Many natural products show antioxidative activities in cancer treatment. Zi Shen decoction (ZSD) is a classic prescription for the treatment of lung disease. However, its effect on lung cancer lacks evidence-based efficacy. In this study, we investigated the anticancer effects of ZSD on lung cancer in vivo and in vitro. Our results showed that oral administration of ZSD suppressed the Lewis lung cancer (LLC) growth in a subcutaneous allograft model and promoted necrosis and inflammatory cell infiltration in the tumor tissues. Furthermore, ZSD not only inhibited tumor cell proliferation and migration but also induced cell apoptosis in lung cancer cells. PI3K/AKT signaling is well characterized in response to oxidative stress. The bioinformatics analysis and western blot assays suggested that ZSD decreased the enzyme activity of PI3K and AKT in vivo and in vitro. We also found that the AKT/GSK-3ß/ß-catenin pathway medicated anticancer effect of ZSD in lung cancer cells. In conclusion, we demonstrate for the first time that ZSD possesses antitumor properties, highlighting its potential use as an alternative strategy or adjuvant treatment for lung cancer therapy.

Simultaneous determination of nine constituents by validated UFLC-MS/MS in the plasma of cough variant asthma rats and its application to pharmacokinetic study after oral administration of Huanglong cough oral liquid.

Huanglong cough oral liquid (HL), an important traditional Chinese medicine prescription for treating pediatric cough variant asthma (CVA) in Nanjing hospital of traditional Chinese medicine for many years. In this study, a selective, accurate and sensitive ultra fast liquid chromatography extreme resolution coupled with mass spectrometer (UFLC-MS/MS) method was established and validated for the simultaneous determination of nine constituents including morusin, ephedrine, praeruptorin A, praeruptorin B, luteolin, rosmarinic acid, quercetin, amygdalin, caffeic acid in CVA rat plasma sensitized and challenged with ovalbumin and cinnamaldehyde. Plasma samples were prepared by protein precipitation with four-fold amount of methanol. UFLC separation was performed on a Thermo Scientific AcclaimTM RSLC 120 C18 column (2.1 mm × 100 mm, 2.2 µm) with mobile phase containing methanol and 0.1% formic acid-water by gradient elution in 8.1 min at total flow of 0.3 mL/min. The determination of target compounds in plasma was operated by multiple reaction monitoring (MRM) mode with positive and negative electrospray ionization (ESI) source. The correlation coefficients (r) of all compounds were from 0.9930 to 0.9994 in the linear range. Lower limit of quantification (LLOQ, ng/mL) was 0.81, 2.01, 2.11, 1.17, 1.04, 0.89, 0.67, 1.45 and 0.59 for morusin, ephedrine, praeruptorin A, praeruptorin B, luteolin, rosmarinic acid, quercetin, amygdalin and caffeic acid, respectively. Intra- and inter-day accuracy and precision, extraction recovery, matrix effect, carryover effect, dilution integrity, and stability were within the limits specified. The established method was effectively applied to a pharmacokinetic study of the nine compounds in CVA rat plasma following oral administration HL exact (7.5, 15, 30 g/kg).

Elaboration of the Comprehensive Metabolic Profile of Salvianolic Acid A in Vivo and in Vitro Using UFLC-Q/TOF-MS.

Salvianolic acid A (Sal A) has a wide range of pharmacological activities. To date, there have been no systematic and detailed metabolite research data of Sal A after oral administration in vitro and in vivo. In this study, a rapid and systematic method based on ultrafast liquid chromatography-quadrupole-time-of-flight mass spectrometry was developed to detect metabolites of Sal A in vitro (human liver microsome, human intestinal microbiota, artificial gastric, and intestinal juice) and in vivo (urine, plasma, feces, and various organs collected after oral administration of Sal A to normal rats and pseudo-germ-free rats). A total of 26 metabolites of Sal A were characterized. These metabolites were formed through extensive metabolic reactions, such as hydroxylation, hydrogenation, and glucuronidation reactions. This study provides novel possibility for exploring the potential biological mechanism of Sal A, and aids the promotion of clinical application.

Triptolide Induces hepatotoxicity via inhibition of CYP450s in Rat liver microsomes.

BACKGROUND: Triptolide (TP), an active constituent of Tripterygium wilfordii, possesses numerous pharmacological activities. However, its effects on cytochrome P450 enzymes (CYP450s) in rats remain unexplored. METHODS: In this study, the effects of triptolide on the six main CYP450 isoforms (1A2, 2C9, 2C19, 2D6, 2E1, and 3A) were investigated both in vivo and in vitro. We monitored the body weight, survival proportions, liver index, changes in pathology, and biochemical index upon TP administration, in vivo. Using a cocktail probe of CYP450 isoform-specific substrates and their metabolites, we then carried out in vitro enzymatic studies in liver microsomal incubation systems via ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Finally, we verified our results at the messenger ribonucleic acid (mRNA) and protein level through quantitative real-time polymerase chain reaction (RT-qPCR), western blotting, and immunohistochemical detection. RESULTS: The in vivo toxicity study confirmed that Sprague-Dawley (SD) rats exhibited dose-dependent hepatotoxicity after intragastric administration of TP [200, 400, and 600 µg/(kg.day)] for 28 days. In case of the CYP450 isoforms 3A, 2C9, 2C19, and 2E1, the in vitro metabolic study demonstrated a decrease in the substrate metabolic rate, metabolite production rate, and Vmax, with an increase in the Km value, compared with that observed in the control group. Additionally, a TP dose-dependent decrease in the mRNA levels was observed in the four major isoforms of CYP3A subfamily (3A1/3A23, 3A2, 3A9, and 3A62) and CYP2C9. A similar effect was also observed with respect to the protein levels of CYP2C19 and CYP2E1. CONCLUSIONS: This study suggests that TP can cause hepatotoxicity by reducing the substrate affinity, activity, and expression at the transcriptional and protein levels of the CYP450 isoforms 3A, 2C9, 2C19, and 2E1. TP also has the potential to cause pharmacokinetic drug interactions when co-administered with drugs metabolized by these four isoforms. However, further clinical studies are needed to evaluate the significance of this interaction.