Correction to: Endometrium metabolomic profiling reveals potential biomarkers for diagnosis of endometriosis at minimal-mild stages.

The authors regret that the incorrect identification of "omega-3 arachidonic acid" published in the original manuscript [1].

Endometrium metabolomic profiling reveals potential biomarkers for diagnosis of endometriosis at minimal-mild stages.

BACKGROUND: The sensitivity and specificity of non-invasive diagnostic methods for endometriosis, especially at early stages, are not optimal. The clinical diagnostic indicator cancer antigen 125 (CA125) performs poorly in the diagnosis of minimal endometriosis, with a sensitivity of 24%. Therefore, it is urgent to explore novel diagnostic biomarkers. We evaluated the metabolomic profile variation of the eutopic endometrium between minimal-mild endometriosis patients and healthy women by ultra-high-performance liquid chromatography coupled with electrospray ionization high-resolution mass spectrometry (UHPLC-ESI-HRMS). METHODS: Our study comprised 29 patients with laparoscopically confirmed endometriosis at stages I-II and 37 infertile women who underwent diagnostic laparoscopy combined with hysteroscopy from January 2014 to January 2015. Eutopic endometrium samples were collected by pipelle endometrial biopsy. The metabolites were quantified by UHPLC-ESI-HRMS. The best combination of biomarkers was then selected by performing step-wise logistic regression analysis with backward elimination. RESULTS: Twelve metabolites were identified as endometriosis-associated biomarkers. The eutopic endometrium metabolomic profile of the endometriosis patients was characterized by a significant increase in the concentration of hypoxanthine, L-arginine, L-tyrosine, leucine, lysine, inosine, omega-3 arachidonic acid, guanosine, xanthosine, lysophosphatidylethanolamine and asparagine. In contrast, the concentration of uric acid was decreased. Metabolites were filtered by step-wise logistic regression with backward elimination, and a model containing uric acid, hypoxanthine, and lysophosphatidylethanolamine was constructed. Receiver-operating characteristic (ROC) analysis confirmed the prognostic value of these parameters for the diagnosis of minimal/mild endometriosis with a sensitivity of 66.7% and a specificity of 90.0%. CONCLUSIONS: Metabolomics analysis of the eutopic endometrium in endometriosis was effectively characterized by UHPLC-ESI-HRMS-based metabolomics. Our study supports the importance of purine and amino acid metabolites in the pathophysiology of endometriosis and provides potential biomarkers for semi-invasive diagnosis of early-stage endometriosis.

Metabolic mapping of Schisandra sphenanthera extract and its active lignans using a metabolomic approach based on ultra high performance liquid chromatography with high-resolution mass spectrometry.

Schisandra sphenanthera, the dried ripe fruit of Schisandra sphenanthera Rehd. et Wils, is widely used as a restorative, tonic and nutrition in many countries. Wuzhi tablet, an ethanol extract preparation of Schisandra sphenanthera, is a well-known herbal medicine widely used in China. Our previous studies show that Wuzhi tablet and its active lignans significantly protect liver injury. However, its metabolic profile remains unknown in vivo and in vitro. In this study, ultra high performance liquid chromatography coupled with electrospray ionization high-resolution mass spectrometry based metabolomics was employed to decipher the metabolic map of Wuzhi tablet and its active lignans. Serum (2 h) and urine (24 h) samples after a 700 mg/kg single oral dose of Wuzhi tablet, and mice liver microsome samples after incubation with its active lignans were collected and analyzed. The data were further analyzed using metabolomics and metabolite identification software. In total, 33 metabolites in vivo and 34 metabolites in vitro were identified, and six among them were new metabolites. The major metabolic reactions encompassed demethylation, hydroxylation, dehydrogenation, and epoxidation. Taken together, in vitro and in vivo studies revealed the metabolic profile of Wuzhi tablet and its active lignans and demethylation and hydroxylation were their major metabolic pathways.

Schisandra sphenanthera Extract Facilitates Liver Regeneration after Partial Hepatectomy in Mice.

Liver regeneration after surgical liver resection is crucial for the restoration of liver mass and the recovery of liver function.Schisandra sphenanthera extract (Wuzhi tablet, WZ) is a preparation of an extract from the dried ripe fruit of Schisandra sphenanthera Rehd. et Wils, a traditional hepatoprotective herb. Previously, we found that WZ could induce liver regeneration-related genes against acetaminophen-induced liver injury. However, whether WZ can directly facilitate liver regeneration after liver resection remains unknown. We investigated whether WZ has potential in promoting liver regeneration after a partial hepatectomy (PHX) in mice. Remnant livers were collected 1, 1.5, 2, 3, 5, 7, and 10 days after PHX. Hepatocyte proliferation was assessed using the Ki-67 labeling index. Western blot analysis was performed on proteins known to be involved in liver regeneration. The results demonstrated that WZ significantly increased the liver-to-body weight ratio of mice after PHX but had no effect on that of mice after a sham operation. Additionally, the peak hepatocyte proliferation was observed at 1.5 days in PHX/WZ-treated mice but at 2 days in PHX/saline-treated mice, as evidenced by the Ki-67 positive ratio. Furthermore, WZ significantly increased the protein expression of ligand-induced phosphorylation of epidermal growth factor receptor and up-regulated cyclin D1, cyclin D-dependent kinase 4, phosphorylated retinoblastoma, and proliferating cell nuclear antigen protein expression and down-regulated the expression of cell cycle inhibitors p21 and p27 in the regenerative process after PHX. These results demonstrate that WZ significantly facilitates hepatocyte proliferation and liver regeneration after PHX.

Hepatoprotective Effects of Schisandra sphenanthera Extract against Lithocholic Acid-Induced Cholestasis in Male Mice Are Associated with Activation of the Pregnane X Receptor Pathway and Promotion of Liver Regeneration.

We previously reported that the ethanol extract of Schisandra sphenanthera [Wuzhi (WZ) tablet] significantly protects against acetaminophen-induced hepatoxicity. However, whether WZ exerts a protective effect against cholestasis remains unclear. In this study, the protective effect of WZ on lithocholic acid (LCA)-induced intrahepatic cholestasis in mice was characterized and the involved mechanisms were investigated. WZ pretreatment (350 mg/kg) with LCA significantly reversed liver necrosis and decreased serum alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase activity. More importantly, serum total bile acids and total bilirubin were also remarkably reduced. Quantitative reverse-transcription polymerase chain reaction and Western blot analysis showed that hepatic expression of pregnane X receptor (PXR) target genes such as CYP3A11 and UDP-glucuronosyltransferase (UGT) 1A1 were significantly increased by WZ treatment. Luciferase assays performed in LS174T cells illustrated that WZ extract and its six bioactive lignans could all activate human PXR. In addition, WZ treatment significantly promoted liver regeneration via inhibition of p53/p21 to induce cell proliferation-associated proteins such as cyclin D1 and proliferating cell nuclear antigen. In conclusion, WZ has a protective effect against LCA-induced intrahepatic cholestasis, partially owing to activation of the PXR pathway and promotion of liver regeneration.

Targeted bile acids and gut microbiome profiles reveal the hepato-protective effect of WZ tablet (Schisandra sphenanthera extract) against LCA-induced cholestasis.

Cholestasis is caused by the obstacle of bile formation or secretion and can develop into severe liver diseases. We previously reported the ethanol extract of Schisandra sphenanthera (Wuzhi tablet, WZ) can significantly protect against lithocholic acid (LCA)-induced intrahepatic cholestasis in mice, partially due to the activation of PXR pathway and promotion of liver regeneration. However, the effect of WZ on the bile acids profile and gut microbiome in cholestastic mice remain unknown. In this study, the effect of WZ against LCA-induced liver injury was evaluated and its effect on the bile acids metabolome and gut microbiome profiles in cholestastic mice was further investigated. Targeted metabolomics analysis was performed to examine the change of bile acids in the serum, liver, intestine and feces. The change of intestinal flora were detected by the genomics method. Targeted metabolomics analysis revealed that WZ enhanced the excretion of bile acids from serum and liver to intestine and feces. Genomics analysis of gut microbiome showed that WZ can reverse LCA-induced gut microbiome disorder to the normal level. In conclusion, WZ protects against LCA-induced cholestastic liver injury by reversing abnormal bile acids profiles and alteration of gut microbiome.

Lipid Profiling of Peri-implantation Endometrium in Patients With Premature Progesterone Rise in the Late Follicular Phase.

CONTEXT: Late follicular phase elevation in serum progesterone (P) during controlled ovarian hyperstimulation negatively affects the outcome of assisted reproductive technology by contributing to endometrial-embryo asynchrony. There are still no data on lipid metabolite alterations during this process. OBJECTIVES: To investigate alterations in the lipid profile during the window of implantation in patients with premature P rise. DESIGN: Lipidomic variations in the endometrium were evaluated by ultrahigh-performance liquid chromatography coupled with electrospray ionization high-resolution mass spectrometry. SETTING: University assisted reproductive medicine unit. PATIENTS OR OTHER PARTICIPANTS: Forty-three patients undergoing in vitro fertilization/intracytoplasmic sperm injection because of a tubal factor or male factor infertility were included in this study. The patients were divided into a high P group (P ≥ 1.5 ng/mL, 15 patients) and a normal P group (P < 1.5 ng/mL, 28 patients) on the day of human chorionic gonadotropin administration. INTERVENTIONS: The endometrial tissues were obtained by Pipelle biopsy 7 days after human chorionic gonadotropin administration. MAIN OUTCOME MEASURES: Alterations in lipid metabolites. RESULTS: A total of 1026 ions were identified, and 25 lipids were significantly upregulated. The endometrial lipid profile was characterized by substantial increases in the concentrations of phosphatidylcholine, phosphatidylethanolamine, lysophosphatidylcholine, diacylglycerol, ceramide, phosphatidylinositol, and phosphatidylserine in patients with a premature P rise at the end of the follicular phase. The correlation analysis between P levels and lipids showed a stronger negative correlation between phosphatidylethanolamine or phosphatidylserine and P levels. CONCLUSIONS: Premature P elevation disrupts the lipid homeostasis of the endometrium during the peri-implantation period. The altered lipid levels may impair endometrial receptivity and early embryo implantation.

Discovery of Phosphatidic Acid, Phosphatidylcholine, and Phosphatidylserine as Biomarkers for Early Diagnosis of Endometriosis.

The sensitivity and specificity of clinical diagnostic indicators and non-invasive diagnostic methods for endometriosis at early stage is not optimal. Previous studies demonstrated that abnormal lipid metabolism was involved in the pathological development of endometriosis. Our cross-sectional study included 21 patients with laparoscopically confirmed endometriosis at stage I-II and 20 infertile women who underwent diagnostic laparoscopy combined with hysteroscopy from January 2014 to January 2015. Eutopic endometrium was collected by pipelle endometrial biopsy. Lipid metabolites were quantified by ultra-high performance liquid chromatography coupled with electrospray ionization high-resolution mass spectrometry (UHPLC-ESI-HRMS). Lipid profiles of endometriosis patients at early stage (I-II) was characterized by a decreased concentration of phosphatidylcholine (18:1/22:6), (20:1/14:1), (20:3/20:4), and phosphatidylserine (20:3/23:1) and an increased concentration of phosphatidic acid (25:5/22:6) compared with control. The synthesized predicting strategy with 5 biomarkers has a specificity of 75.0% and a sensitivity of 90.5%. Lipid profile of eutopic endometrium in endometriosis was effectively characterized by UHPLC-ESI-HRMS-based metabolomics. Our study demonstrated the alteration of phosphatidic acid, phosphatidylcholine, phosphatidylserine metabolites in endometriosis and provided potential biomarkers for semi-invasive diagnose of endometriosis at early stage.

Schisandrol B promotes liver regeneration after partial hepatectomy in mice.

Liver regeneration is a vital process of recovery after liver damage, which is a promising clinical strategy after partial hepatectomy (PHx). Schisandrol B (SolB), one of the bioactive ingredients from Schisandra sphenanthera, displays significant hepato-protection effects against drug-induced liver injure in mice. However, the effect of SolB on liver regeneration after PHx remains unclear. Here, we showed that SolB treatment promoted liver mass restoration and increased the number of proliferative hepatocytes following PHx. SolB treatment significantly improved the levels of growth factors (HGF and EGF) and cytokines (IL-6), which further activated STAT3/Akt/MAPK signaling pathways and induced the expression of several the protein of cell cycle core. Overall, this study is the first to demonstrate the role of SolB in promoting liver regeneration during PHx challenge, which provide a clinically relevant argument for using SolB to facilitate liver recovery after undergoing PHx or liver transplantation.

Schisandra sphenanthera extract (Wuzhi Tablet) protects against chronic-binge and acute alcohol-induced liver injury by regulating the NRF2-ARE pathway in mice.

Alcohol abuse leads to alcoholic liver disease and no effective therapy is currently available. Wuzhi Tablet (WZ), a preparation of extract from Schisandra sphenanthera that is a traditional hepato-protective herb, exerted a significant protective effect against acetaminophen-induced liver injury in our recent studies, but whether WZ can alleviate alcohol-induced toxicity remains unclear. This study aimed to investigate the contribution of WZ to alcohol-induced liver injury by using chronic-binge and acute models of alcohol feeding. The activities of ALT and AST in serum were assessed as well as the level of GSH and the activity of SOD in the liver. The expression of CYP2E1 and proteins in the NRF2-ARE signaling pathway including NRF2, GCLC, GCLM, HO-1 were measured, and the effect of WZ on NRF2 transcriptional activity was determined. We found that both models resulted in liver steatosis accompanied by increased transaminase activities, but that liver injury was significantly attenuated by WZ. WZ administration also inhibited CYP2E1 expression induced by alcohol, and elevated the level of GSH and the activity of SOD in the liver. Moreover, the NRF2-ARE signaling pathway was activated by WZ and the target genes were all upregulated. Furthermore, WZ significantly activated NRF2 transcriptional activity. Collectively, our study demonstrates that WZ protected against alcohol-induced liver injury by reducing oxidative stress and improving antioxidant defense, possibly by activating the NRF2-ARE pathway.

p53-mediated regulation of bile acid disposition attenuates cholic acid-induced cholestasis in mice.

BACKGROUND AND PURPOSE: The tumour suppressor p53 is traditionally recognized as a surveillance molecule to preserve genome integrity. Recent studies have demonstrated that it contributes to metabolic diseases. Here, we investigated the role of p53 in the regulation of bile acid disposition and cholestasis. EXPERIMENTAL APPROACH: The bile acid disposition-related gene expression profile affected by p53 activation was assessed in mouse primary hepatocytes with p53 depletion and in Trp53-null mice. Dual luciferase reporter assay was used to detect the transcriptional activities of target genes. Anticholestatic effects of p53 activator doxorubicin were investigated in a 0.5% cholic acid-fed mouse model of cholestasis. Changes in bile acids were evaluated using metabolomics analysis. KEY RESULTS: Doxorubicin-mediated p53 activation induced Cyp2b10, Sult2a1 and Abcc2/3/4 expression in mice in vitro and in vivo. ABCC3 and CYP2B6 (human orthologue of Cyp2b10) were identified as direct p53 target genes. Doxorubicin attenuated cholic acid-induced cholestasis in mice, as demonstrated by shrunken gall bladder, decreased serum total bile acid and total bilirubin levels and alkaline phosphatase activity. Targeted metabolomics analysis revealed that doxorubicin enhanced the excretion of bile acid metabolites from serum and liver to intestine and faeces. Up-regulation of Cyp2b10, Sult2a1 and Abcc2/3/4 expression was further confirmed in cholestatic mice. Cholic acid-induced cholestatic injury was aggravated in p53-deficient mice and levels of bile acid in intestine and faeces were decreased. CONCLUSIONS AND IMPLICATIONS: Our findings suggest a novel role of p53 in promoting bile acid disposition and alleviating cholestatic syndrome, which provides a potential therapeutic target for cholestasis.

Schisandrol B protects against cholestatic liver injury through pregnane X receptors.

BACKGROUND AND PURPOSE: Currently, ursodeoxycholic acid and obeticholic acid are the only two FDA-approved drugs for cholestatic liver diseases. Thus, new therapeutic approaches need to be developed. Here we have evaluated the anti-cholestasis effects of Schisandrol B (SolB), a bioactive compound isolated from Schisandra sphenanthera. EXPERIMENTAL APPROACH: Hepatoprotective effect of SolB against intrahepatic cholestasis, induced by lithocholic acid (LCA), was evaluated in mice. Metabolomic analysis and gene analysis were used to assess involvement of pregnane X receptor (PXR). Molecular docking, cell-based reporter gene analysis and knockout mice were used to demonstrate the critical role of the PXR pathway in the anti-cholestasis effects of SolB. KEY RESULTS: SolB protected against LCA-induced intrahepatic cholestasis. Furthermore, therapeutic treatment with SolB decreased mortality in cholestatic mice. Metabolomics and gene analysis showed that SolB accelerated metabolism of bile acids, promoted bile acid efflux into the intestine, and induced hepatic expression of the PXR-target genes Cyp3a11, Ugt1a1, and Oatp2, which are involved in bile acid homeostasis. Mechanistic studies showed that SolB activated human PXR and up-regulated PXR target genes in human cell lines. Additionally, SolB did not protect Pxr-null mice from liver injury induced by intrahepatic cholestasis, thus providing genetic evidence that the effect of SolB was PXR-dependent. CONCLUSION AND IMPLICATIONS: These findings provide direct evidence for the hepatoprotective effects of SolB against cholestasis by activating PXR. Therefore, SolB may provide a new and effective approach to the prevention and treatment of cholestatic liver diseases.

Oleanolic acid attenuates obstructive cholestasis in bile duct-ligated mice, possibly via activation of NRF2-MRPs and FXR antagonism.

Obstructive cholestasis is characterized by impairment of hepatic canalicular bile efflux and there are no clinically effective drugs to cure except surgeries. Previously we revealed that oleanolic acid (OA) protected against lithocholic acid (LCA)-induced intrahepatic cholestasis in mice. Cholestasis caused by LCA is characterized by segmental bile duct obstruction, whether OA possesses the beneficial effect on completed obstructive cholestasis induced by bile duct ligation (BDL) remains unknown. In this study, we demonstrated that BDL-induced mice liver pathological change, and increase in serum levels of ALT, AST and ALP were all significantly reduced by OA (20 mg/kg, i.p.). Meanwhile, OA also lowered total bilirubin and total bile acids levels in serum, as well as total bile acids level in liver, in contrast, urinary total bile acids output was remarkably up-regulated by OA. Gene expression analysis showed that OA caused significant increased mRNA expression of MRP3 and MRP4 located at hepatic basolateral membrane, and restoration of MRP2 and BSEP located at hepatic cannalicular membrane. Furthermore, significant NRF2 protein accumulation in nucleus was also observed in OA treated mice. In mice primary cultured hepatocytes, the effects of OA on MRP2, MRP3 and MRP4 expression were directly proved to be mediated via NRF2 activation, and BSEP downregulation induced by OA was in part due to FXR antagonism. Luciferase assay performed in Hep G2 cells also illustrated that OA was a partial FXR antagonist. Taken together, we conclude that OA attenuates obstructive cholestasis in BDL mice, possibly via activation of NRF2-MRPs and FXR antagonism.

Simultaneous determination of seven taxoids in rat plasma by UPLC-MS/MS and pharmacokinetic study after oral administration of Taxus yunnanensis extracts.

A rapid, sensitive and reliable method has been developed and validated for the simultaneous determination of seven taxoids including 10-deacetylbaccatin III (10-DAB III), baccatin III, 5-epi-canadensene, taxinine M, 10-deacetyltaxol (10-DAT), cephalomannine and paclitaxel in rat plasma using docetaxel as the internal standard (IS). The plasma samples were pretreated by liquid-liquid extraction with methyl tert-butyl ether. The chromatographic separation was achieved on a C18 column (50 mm × 2.1 mm, 1.8 µm, Waters, USA) with a gradient elution program consisting of methanol and water (containing 0.1% formic acid) at a flow rate of 0.2 mL/min. Detection was performed under the selected reaction monitoring (SRM) scan using an electrospray ionization (ESI) in the positive ion mode. The mass transitions were as follows: m/z 567.4→444.9 for 10-DAB III, m/z 609.0→549.3 for baccatin III, m/z 617.4→496.9 for 5-epi-canadensene, m/z 709.6→649.3 for taxinine M, m/z 834.8→307.9 for 10-DAT, m/z 854.5→285.4 for cephalomannine, m/z 876.8→307.3 for paclitaxel and m/z 830.8→549.6 for IS, respectively. All calibration curves exhibited good linearity (r(2)>0.99) over a wide concentration range for all components. The intra-day and inter-day precisions at three different levels were both less than 14.3% in terms of relative standard deviation (RSD) and the accuracies ranged from -8.3% to 14.8% in terms of relative error (RE). The extraction recoveries of the seven compounds ranged from 62.5% to 100.5%. The developed method was successfully applied to the pharmacokinetic study of the seven taxoids in rat plasma after oral administration of the crude extract of the twigs and leaves of Taxus yunnanensis.