Toosendanin induces hepatotoxicity via disrupting LXRα/Lipin1/SREBP1 mediated lipid metabolism.

Toosendanin (TSN) is the main active compound derived from Melia toosendan Sieb et Zucc with various bioactivities. However, liver injury was observed in TSN limiting its clinical application. Lipid metabolism plays a crucial role in maintaining cellular homeostasis, and its disruption is also essential in TSN-induced hepatotoxicity. This study explored the hepatotoxicity caused by TSN in vitro and in vivo. The lipid droplets were significantly decreased, accompanied by a decrease in fatty acid transporter CD36 and crucial enzymes in the lipogenesis including ACC and FAS after the treatment of TSN. It was suggested that TSN caused lipid metabolism disorder in hepatocytes. TOFA, an allosteric inhibitor of ACC, could partially restore cell survival via blocking malonyl-CoA accumulation. Notably, TSN downregulated the LXRα/Lipin1/SREBP1 signaling pathway. LXRα activation improved cell survival and intracellular neutral lipid levels, while SREBP1 inhibition aggravated the cell damage and caused a further decline in lipid levels. Male Balb/c mice were treated with TSN (5, 10, 20 mg/kg/d) for 7 days. TSN exposure led to serum lipid levels aberrantly decreased. Moreover, the western blotting results showed that LXRα/Lipin1/SREBP1 inhibition contributed to TSN-induced liver injury. In conclusion, TSN caused lipid metabolism disorder in liver via inhibiting LXRα/Lipin1/SREBP1 signaling pathway.

Rosmarinic acid alleviates toosendanin-induced liver injury through restoration of autophagic flux and lysosomal function by activating JAK2/STAT3/CTSC pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Rosmarinic acid (RA), a natural polyphenol abundant in numerous herbal remedies, has been attracting growing interest owing to its exceptional ability to protect the liver. Toosendanin (TSN), a prominent bioactive compound derived from Melia toosendan Siebold & Zucc., boasts diverse pharmacological properties. Nevertheless, TSN possesses remarkable hepatotoxicity. Intriguingly, the potential of RA to counteract TSN-induced liver damage and its probable mechanisms remain unexplored. AIM OF THE STUDY: This study is aimed at exploring whether RA can alleviate TSN-induced liver injury and the potential mechanisms involved autophagy. MATERIALS AND METHODS: CCK-8 and LDH leakage rate assay were used to evaluate cytotoxicity. Balb/c mice were intraperitoneally administered TSN (20 mg/kg) for 24 h after pretreatment with RA (0, 40, 80 mg/kg) by gavage for 5 days. The autophagic proteins P62 and LC3B expressions were detected using western blot and immunohistochemistry. RFP-GFP-LC3B and transmission electron microscopy were applied to observe the accumulation levels of autophagosomes and autolysosomes. LysoTracker Red and DQ-BSA staining were used to evaluate the lysosomal acidity and degradation ability respectively. Western blot, immunohistochemistry and immunofluorescence staining were employed to measure the expressions of JAK2/STAT3/CTSC pathway proteins. Dual-luciferase reporter gene was used to measure the transcriptional activity of CTSC and RT-PCR was used to detect its mRNA level. H&E staining and serum biochemical assay were employed to determine the degree of damage to the liver. RESULTS: TSN-induced damage to hepatocytes and livers was significantly alleviated by RA. RA markedly diminished the autophagic flux blockade and lysosomal dysfunction caused by TSN. Mechanically, RA alleviated TSN-induced down-regulation of CTSC by activating JAK2/STAT3 signaling pathway. CONCLUSION: RA could protect against TSN-induced liver injury by activating the JAK2/STAT3/CTSC pathway-mediated autophagy and lysosomal function.

Toosendanin induces hepatotoxicity by restraining autophagy and lysosomal function through inhibiting STAT3/CTSC axis.

Toosendanin (TSN) is the main active component in the traditional herb Melia toosendan Siebold & Zucc, which exhibits promising potential for development due to its diverse pharmacological properties. However, the hepatotoxicity associated with TSN needs further investigation. Previous research has implicated autophagy dysregulation in TSN-induced hepatotoxicity, yet the underlying mechanisms remain elusive. In this study, the mechanisms of signal transducer and activator of transcription 3 (STAT3) in TSN-induced autophagy inhibition and liver injury were explored using Stat3 knockout C57BL/6 mice and HepG2 cells. TSN decreased cell viability, increased lactate dehydrogenase (LDH) production in vitro, and elevated serum aspartate transaminase (AST) and alanine aminotransferase (ALT) levels as well as liver lesions in vivo, suggesting TSN had significant hepatotoxicity. TSN inhibited Janus kinase 2 (JAK2)/STAT3 pathway and the expression of cathepsin C (CTSC). Inhibition of STAT3 exacerbated TSN-induced autophagy inhibition and hepatic injury, whereas activation of STAT3 attenuated these effects of TSN. Mechanistically, STAT3 transcriptionally regulated the level of CTSC gene, which in turn affected autophagy and the process of liver injury. TSN-administered Stat3 knockout mice showed more severe hepatotoxicity, CTSC downregulation, and autophagy blockade than wildtype mice. In summary, TSN caused hepatotoxicity by inhibiting STAT3/CTSC axis-dependent autophagy and lysosomal function.

Mitigation of triptolide-induced testicular Sertoli cell damage by melatonin via regulating the crosstalk between SIRT1 and NRF2.

BACKGROUND: Triptolide (TP) is an important active compound from Tripterygium wilfordii Hook F (TwHF), however, it is greatly limited in clinical practice due to its severe toxicity, especially testicular injury. Melatonin is an endogenous hormone and has beneficial effects on the reproductive system. However, whether triptolide-induced testicular injury can be alleviated by melatonin and the underlying mechanism are not clear. PURPOSE: In this study, we aimed to explore whether triptolide-induced testicular Sertoli cells toxicity can be mitigated by melatonin and the underlying mechanisms involved. METHODS: Cell apoptosis was assessed by flow cytometry, western blot, immunofluorescence and immunohistochemistry. Fluorescent probe Mito-Tracker Red CMXRos was used to observe the mitochondria morphology. Mitochondrial membrane potential and Ca2+ levels were used to investigate mitochondrial function by confocal microscope and flow cytometry. The expression levels of SIRT1/Nrf2 pathway were detected by western blot, immunofluorescence and immunohistochemistry. Small interfering RNA of NRF2 and SIRT1 inhibitor EX527 was used to confirm the role of SIRT1/NRF2 pathway in the mitigation of triptolide-induced Sertoli cell damage by melatonin. Co-Immunoprecipitation assay was used to determine the interaction between SIRT1 and NRF2. RESULTS: Triptolide-induced dysfunction of testicular Sertoli cells was significantly improved by melatonin treatment. Specifically, triptolide-induced oxidative stress damage and changes of mitochondrial morphology, mitochondrial membrane potential, and BTB integrity were alleviated by melatonin. Mechanistically, triptolide inhibited SIRT1 and then reduced the activation of NRF2 pathway via regulating the interaction between SIRT1 and NRF2, thereby downregulating the downstream antioxidant genes, which was reversed by melatonin. Nevertheless, knockdown of NRF2 or inhibition of SIRT1 abolished the protective effect of melatonin. CONCLUSION: Triptolide-induced testicular Sertoli cell damage could be alleviated by melatonin via regulating the crosstalk between SIRT1 and NRF2, which is helpful for developing a new strategy to alleviate triptolide-induced toxicity.

POU Class 2 Associating Factor 1 Exerts a Protective Effect on the Respiratory Syncytial Virus-Induced Acute Bronchiolitis by the NF-κB Pathway.

Background: Respiratory syncytial virus (RSV) is the main pathogen causing acute bronchiolitis, which is common in infants and young children. A previous study revealed the possible involvement of POU class 2 associating factor 1 (POU2AF1) in RSV-triggered acute bronchiolitis. We attempted to clarify the specific action mechanism of POU2AF1 underlying RSV-triggered inflammation. Methods: RT-qPCR measured POU2AF1 levels in RSV-infected children, mice, and airway epithelial cell lines (HBECs). HE staining showed histopathological features in the lung tissue of RSV-infected mice. ELISA examined the contents of proinflammatory cytokines in RSV-infected mice. Western blotting evaluated the protein abundance of proinflammatory cytokines in RSV-infected HBECs and assessed NF-κB pathway-associated protein expression in RSV-infected mice and RSV-treated HBECs. Results: POU2AF1 presented depletion in RSV-infected children, mice, and HBECs. RSV-infected triggered lung injury and inflammatory cell infiltration in the mouse lung tissue, while POU2AF1 overexpression rescued these changes. RSV-infected induced inflammatory impairment in HBECs, whereas POU2AF1 reversed this effect. POU2AF1 suppressed the upregulated NF-κB pathway-associated protein expression in mice and HBECs under RSV infection. Conclusion: POU2AF1 exerts a protective impact on RSV-induced acute bronchiolitis in vitro and in vivo through the NF-κB pathway. Our research may provide a novel direction for better therapy of RSV-triggered acute bronchiolitis.

Toosendanin induced hepatotoxicity via triggering PERK-eIF2α-ATF4 mediated ferroptosis.

Toosendanin (TSN) is the main active compound of Melia toosendan Sieb et Zucc with various bioactivities. In this study, we investigated the role of ferroptosis in TSN-induced hepatotoxicity. The characteristic indicators of ferroptosis were detected including reactive oxygen species (ROS), lipid-ROS, glutathione (GSH), ferrous ion and the expression of glutathione peroxidase 4 (GPX4), which showed that TSN caused ferroptosis in hepatocytes. The results of qPCR analysis and western blotting assay showed that TSN-induced activation of protein kinase R-like endoplasmic reticulum kinase (PERK)- eukaryotic initiation factor 2 α subunit (eIF2α)- activation transcription factor 4 (ATF4) signaling pathway resulted in increasing activation transcription factor 3 (ATF3) expression, which upregulated the expression of transferrin receptor 1 (TFRC). Furthermore, TFRC mediated iron accumulation leading to ferroptosis in hepatocytes. To clarify whether TSN triggered ferroptosis in vivo, male Balb/c mice were treated with the different doses of TSN. The results of hematoxylin-eosin (H&E) staining, 4-hydroxynonenal (4-HNE) staining, malondialdehyde (MDA) content and the protein expression of GPX4 showed that ferroptosis contributed to TSN-induced hepatotoxicity. Iron homeostasis relative protein and PERK- eIF2α- ATF4 signaling pathway also involved in hepatotoxicity of TSN in vivo.

MitoQ alleviates triptolide-induced cardiotoxicity via activation of p62/Nrf2 axis in H9c2 cells.

Triptolide (TP) is one of the major components of Tripterygium wilfordii, which is a traditional Chinese medicine widely used in the treatment of various autoimmune and inflammatory diseases. However, the cardiotoxicity induced by TP greatly limits its widespread clinical application. In view of the role of ROS-mediated oxidative stress in TP-induced cardiotoxicity, mitoQ, a mitochondria-targeted ROS scavenger, was used in this study to investigate its protective effect against TP-induced cardiomyocyte toxicity and its possible underlying mechanism. Here we demonstrated that mitoQ could significantly attenuate TP-induced cardiotoxicity in cardiomyocyte H9c2 cells, with a remarkable improvement in cell viability and reduction in ROS levels. P62-Nrf2 signaling pathway has been reported to play a critical role in regulating oxidative stress and protecting cells from harmful stimuli. In this study, we found that mitoQ significantly activated p62-Nrf2 signaling pathway in H9c2 cells with or without TP treatment. Moreover, knockdown of p62 or Nrf2 could block the protective effect of mitoQ against TP in H9c2 cells. Taken together, our study demonstrates that mitoQ can alleviate TP-induced cardiotoxicity via the activation of p62-Nrf2 signaling pathway, which provides new potential strategies to combat TP-induced cardiomyocyte toxicity.

The novel FAT4 activator jujuboside A suppresses NSCLC tumorigenesis by activating HIPPO signaling and inhibiting YAP nuclear translocation.

FAT atypical cadherin 4 (FAT4) has been identified as a tumor suppressor in lung cancers. However, no agent for lung cancer treatment targeting FAT4 has been used in the clinic. Jujuboside A (JUA) is a major active compound in Semen Ziziphi Spinosae. Semen Ziziphi Spinosae is a traditional Chinese herbal medicine used clinically for tumor treatment to improve patients' quality of life. However, the anti-lung cancer activity and the underlying mechanisms of JUA are not yet fully understood. Here, we demonstrated the anti-lung cancer activity of JUA in two lung cancer mice models and three non-small cell lung cancer (NSCLC) cell lines, and further illustrated its underlying mechanisms. JUA suppressed the occurrence and development of lung cancer and extended mice survival in vivo, and suppressed NSCLC cell activities through cell cycle arrest, proliferation suppression, stemness inhibition and senescence promotion. Moreover, JUA directly bound with and activated FAT4, subsequently activating FAT4-HIPPO signaling and inhibiting YAP nuclear translocation. Knockdown of FAT4 diminished JUA's effects on HIPPO signaling, YAP nuclear translocation, cell proliferation and cellular senescence. In conclusion, JUA significantly suppressed NSCLC tumorigenesis by regulating FAT4-HIPPO-YAP signaling. Our findings suggest that JUA is a novel FAT4 activator that can be developed as a promising NSCLC therapeutic agent targeting the FAT4-HIPPO-YAP pathway.

Triptolide impairs glycolysis by suppressing GATA4/Sp1/PFKP signaling axis in mouse Sertoli cells.

Triptolide (TP), a primary bioactive ingredient isolated from the traditional Chinese herbal medicine Tripterygium wilfordii Hook. F. (TWHF), has attracted great interest for its therapeutic biological activities in inflammation and autoimmune disease. However, its clinical use is limited by severe testicular toxicity, and the underlying mechanism has not been elucidated. Our preliminary evidence demonstrated that TP disrupted glucose metabolism and caused testicular toxicity. During spermatogenesis, Sertoli cells (SCs) provide lactate as an energy source to germ cells by glycolysis. The transcription factors GATA-binding protein 4 (GATA4) and specificity protein 1 (Sp1) can regulate glycolysis. Based on this evidence, we speculate that TP causes abnormal glycolysis in SCs by influencing the expression of the transcription factors GATA4 and Sp1. The mechanism of TP-induced testicular toxicity was investigated in vitro and in vivo. The data indicated that TP decreased glucose consumption, lactate production, and the mRNA levels of glycolysis-related transporters and enzymes. TP also downregulated the protein expression of the transcription factors GATA4 and Sp1, as well as the glycolytic enzyme phosphofructokinase platelet (PFKP). Phosphorylated GATA4 and nuclear GATA4 protein levels were reduced in a dose- and time-dependent manner after TP incubation. Similar effects were observed in shGata4-treated TM4 cells and BALB/c mice administered 0.4 mg/kg TP for 28 days, and glycolysis was also inhibited. Gata4 knockdown downregulated Sp1 and PFKP expression. Furthermore, the Sp1 inhibitor plicamycin inhibited PFKP protein levels in TM4 cells. In conclusion, TP inhibited GATA4-mediated glycolysis by suppressing Sp1-dependent PFKP expression in SCs and caused testicular toxicity.

Triptolide dysregulates glucose uptake via inhibition of IKKß-NF-κB pathway by p53 activation in cardiomyocytes.

Triptolide (TP), a principal bioactive component extracted from traditional Chinese medicine Tripterygium wilfordii Hook. F. (TWHF), has attracted wide attention of its therapeutic effects on inflammation and autoimmune diseases. However, the therapeutic application of TP is hindered by severe cardiomyocyte toxicity and narrow therapeutic window. We previously identified that the p53 was an indispensable contributor in TP-induced myocardial injury. p53 has an inhibitory effect on IKKß-NF-κB pathway that regulates glucose transporters (GLUT) expression. Based on these evidences, we speculate that p53 mediates TP-disturbed glucose uptake by blocking IKKß-NF-κB signaling. This study focused on the effect of TP on cardiac glucose uptake and the role of p53 in glucose metabolism in cardiomyocytes, and p53 -/- mice. TP treatment depressed glucose consumption and ATP production resulting in myocardial damage. Incubation with ATP (5 mM) remarkably decreased the cellular damage. Immunoblotting and immunofluorescence identified that TP suppressed glucose uptake by restricting IKKß-NF-κB signaling activation, GLUT1 and GLUT4 expression. p53 inhibition alleviated the cell damage and the compromise of glucose uptake. Mechanistically, p53 antagonist PFTα abolished TP-induced the inhibition of IKKß, IκBα phosphorylation, p65 nuclear translocation, and GLUT1, GLUT4 expression. Consistently, in acute heart injury models, p53 deficiency upregulated IKKß-NF-κB activation and GLUT1, GLUT4 protein levels which was also indicated as amelioration of heart histological injury after 1.2 mg kg-1 TP administration. The present findings indicate that TP-induced p53 overactivation suppresses glucose uptake by inhibiting IKKß-NF-κB pathway and downregulating NF-κB-dependent GLUT1 and GLUT4 expression.

Triptolide induces mitochondria-mediated apoptosis of Burkitt's lymphoma cell via deacetylation of GSK-3ß by increased SIRT3 expression.

Burkitt's lymphoma (BL) is a highly aggressive B-cell non-Hodgkin lymphoma with rapid growth and dissemination propensity. Triptolide (TP), an active component extracted from Chinese herb Tripterygium wilfordii Hook f., has broad-spectrum anti-tumor activities. This study aimed to explore the in vitro and in vivo anti-cancer effects of TP on BL and the potential molecular mechanisms. In this study, the in vitro anti-tumor activity of TP was determined by CCK-8 and flow cytometry assays in Raji, NAMALWA and Daudi cells. The expression of SIRT3, phosphorylation and acetylation of glycogen synthase kinase-3ß (GSK-3ß) were analyzed by Western blot assay. Moreover, we examined the mitochondrial membrane potential by JC-1 method and measured apoptosis related protein using Western blot assay. BL xenograft model in NOD/SCID mice were established to evaluate the in vivo anti-cancer effect of TP. We discovered that TP inhibited BL cell growth and induced apoptosis in a dose-dependent manner. Loss of SIRT3 provides growth advances for BL cells. However, TP could up-regulate SIRT3 expression, which resulted in suppression of BL cells proliferation. GSK-3ß was activated by SIRT3-mediated deacetylation, which subsequently induced mitochondrial translocation and accumulation of Bax and decrease of mitochondrial membrane potential. Anti-tumor studies in vivo showed that TP (0.36 mg/kg) inhibited the growth of BL xenografts in NOD/SCID mice with an inhibitory rate of 73.13%. Our data revealed that TP triggered mitochondrial apoptotic pathway in BL by increasing SIRT3 expression and activating SIRT3/GSK-3ß/Bax pathway. This study indicated that TP is a potential anti-cancer Chinese herbal medicine against BL.

Epigenetic regulation of active Chinese herbal components for cancer prevention and treatment: A follow-up review.

Epigenetic modifications include DNA methylation, histone modification, and other patterns. These processes are associated with carcinogenesis and cancer progression. Thus, epigenetic modification-related enzymes, such as DNA methyltransferases (DNMTs), histone methyltransferases (HMTs), histone demethylases (HDMTs), histone acetyltransferases (HATs), and histone deacetylases (HDACs), as well as some related proteins, including methyl-CpG binding proteins (MBPs) and DNMT1-associated protein (DMAP 1), are considered as potential targets for cancer prevention and therapy. Numerous natural compounds, mainly derived from Chinese herbs and chemically ranging from polyphenols and flavonoids to mineral salts, inhibit the growth and development of various cancers by targeting multiple genetic and epigenetic alterations. This review summarizes the epigenetic mechanisms by which active compounds from Chinese herbs exert their anti-cancer effect. A subset of these compounds, such as curcumin and resveratrol, affect multiple epigenetic processes, including DNMT inhibition, HDAC inactivation, MBP suppression, HAT activation, and microRNA modulation. Other compounds also regulate epigenetic modification processes, but the underlying mechanisms and clear targets remain unknown. Accordingly, further studies are required.

Tanshinone IIA protects against acetaminophen-induced hepatotoxicity via activating the Nrf2 pathway.

BACKGROUND: Tanshinone IIA (Tan), the main active component of Salvia miltiorrhiza, has been demonstrated to have antioxidant activity. Acetaminophen (APAP), a widely used antipyretic and analgesic, can cause severe hepatotoxicity and liver failure when taken overdose. Oxidative stress has been reported to be involved in APAP-induced liver failure. PURPOSE: This study aimed to investigate the effect of Tan on APAP-induced hepatotoxicity and the underlying mechanisms involved. STUDY DESIGN: C57BL/6J mice were divided into six groups: (1) control, (2) APAP group, (3) APAP+Tan (30mg/kg) group, (4) Tan (30mg/kg) group, (5) APAP+Tan (10mg/kg) group, (6) Tan (10mg/kg) group. Mice in group 3 and 5 were pre-treated with specified dose of Tan by gavage and subsequently injected with an overdose of APAP intraperitoneally (i.p., 300mg/kg). The effect of Tan on Nrf2 pathway was investigated in HepG2 cells and mice. METHODS: Plasma aspartate transaminase (ALT), aspartate transaminase (AST), lactate dehydrogenase (LDH), liver glutathione (GSH), glutathione transferase (GST), glutathione peroxidase (GPx), superoxide dismutase (SOD) and catalase (CAT) levels were determined after mice were sacrificed. Lipid peroxidation and histological examination were performed. The effect of Tan on the Nrf2 pathway was detected by western blotting and qRT-PCR. RESULTS: Tan pretreatment reduced APAP-induced liver injury. Tan was able to activate Nrf2 and increase the expression levels of Nrf2 target genes, including glutamate-cysteine ligase catalytic subunit (GCLC), NAD(P)H:quinine oxidoreductase 1 (NQO1) and hemeoxygenase-1 (HO-1), in a dose-dependent manner in HepG2 cells. Consistent with our observations in HepG2 cells, Tan increased nuclear Nrf2 accumulation and upregulated mRNA and protein levels of the Nrf2 target genes GCLC, NQO1 and HO-1 in C57BL/6J mice compared with mice treated with APAP alone. CONCLUSIONS: Our results demonstrate that Tan pretreatment could protect the liver from APAP-induced hepatic injury by activating the Nrf2 pathway. Tan may provide a new strategy for the protection against APAP-induced liver injury.

Activation of the farnesoid X receptor attenuates triptolide-induced liver toxicity.

BACKGROUND: Triptolide, an active ingredient extracted from the Chinese herb Tripterygium wilfordii Hook f., has multiple pharmacological properties, including anti-inflammatory, immune-modulatory, and anti-proliferative activities. However, the hepatotoxicity of triptolide always limits its clinical applications. HYPOTHESIS/PURPOSE: Farnesoid X receptor (FXR) is a ligand-activated transcription factor that plays a key role in hepatoprotection through the maintenance of liver metabolism homeostasis. This study explored the role of FXR in triptolide-induced cytotoxicity and investigated whether activation of FXR can protect against triptolide-induced liver injury. STUDY DESIGN: The role of FXR in triptolide-induced cytotoxicity was investigated in HepG2 cells. In addition, the protective effect of the selective FXR agonist GW4064 on triptolide-induced hepatotoxicity was explored in BALB/c mice. METHODS: HepG2 cells were transient transfected with FXR expression plasmid or FXR-siRNA. The cytotoxicity was compared using the MTT assay. The extent of liver injury was assessed by histopathology and serum aminotransferases. The expression of FXR and its target genes were detected by Western blot and qRT-PCR. RESULTS: The transient overexpression of FXR protected against triptolide-induced cell death, whereas FXR knockdown with a specific small interfering RNA resulted in increased cytotoxicity. In BALB/c mice, treatment with the FXR agonist GW4064 attenuated triptolide-induced liver dysfunction, structural damage, glutathione depletion and lipid peroxidation. Moreover, the livers of GW4064-treated mice showed increased expression of FXR and several related target genes involved in phase II and phase III xenobiotic metabolism. CONCLUSION: Taken together, these results indicate that activation of FXR attenuates triptolide-induced hepatotoxicity and provide direct implications for the development of novel therapeutic strategies against triptolide-induced hepatotoxicity.

Protective effect of Wuzhi tablet (Schisandra sphenanthera extract) against cisplatin-induced nephrotoxicity via Nrf2-mediated defense response.

Cisplatin is a potent anti-cancer agent for various types of tumors. However, the clinical use of cisplatin is often limited by its nephrotoxicity. This study reports that WZ tablet (WZ, a preparation of an ethanol extract of Schisandra sphenanthera) mitigates cisplatin-induced toxicity in renal epithelial HK-2 cells and in mice. Pretreatment of HK-2 cells with WZ ameliorated cisplatin-induced cytotoxicity caused by oxidative stress, as was demonstrated by reductions in the levels of reactive oxygen species (ROS) and increased levels of glutathione (GSH). WZ facilitated the nuclear accumulation of the transcription factor NF-E2-related factor 2 (Nrf2) and the subsequent expression of its target genes such as NAD(P)H: quinine oxidoreductase 1 (NQO1), heme oxygenase-1 (HO-1) and glutamate cysteine ligase (GCL). Protective effects of WZ on cisplatin-induced nephrotoxicity were also observed in mice. WZ attenuated cisplatin-induced renal dysfunction, structural damage and oxidative stress. The nuclear accumulation of Nrf2 and its target genes were increased by WZ treatment. Taken together, these findings demonstrated WZ have a protective effect against cisplatin-induced nephrotoxicity by activation of the Nrf2 mediated defense response, which is of significant importance for therapeutic intervention in cisplatin induced renal injury.

Synergism and rules of the new combination drug Yiqijiedu formulae (YQJD) on ischemic stroke based on amino acids (AAs) metabolism.

The use of combination drugs is considered to be a promising strategy to control complex diseases such as ischemic stroke. The detection of metabolites has been used as a versatile tool to reveal the potential mechanism of diverse diseases. In this study, the levels of 12 endogenous AAs were simultaneously determined quantitatively in the MCAO rat brain using RRLC-QQQ method. Seven AAs were chosen as the potential biomarkers, and using PLS-DA analysis, the effects of the new combination drug YQJD, which is composed of ginsenosides, berberine, and jasminoidin, on those 7 AAs were evaluated. Four AAs, glutamic acid, homocysteine, methionine, and tryptophan, which changed significantly in the YQJD-treated groups compared to the vehicle groups (P < 0.05), were identified and designated as the AAs to use to further explore the synergism of YQJD. The result of a PCA showed that the combination of these three drugs exhibits the strongest synergistic effect compared to other combination groups and that ginsenosides might play a pivotal role, especially when combined with jasminoidin. We successfully explored the synergetic mechanism of multi-component and provided a new method for evaluating the integrated effects of combination drugs in the treatment of complex diseases.

[Tanshinone IIA protects against triptolide-induced liver injury via Nrf2/ARE activation].

The aim of this study is to investigate the protection effect of tanshinone IIA (Tan) against triptolide (TP)-induced liver injury and the mechanisms involved. Acute liver injury was induced by intraperitoneal injection of TP (1 mg x kg(-1)) in mice. The activities of AST, ALT and LDH in serum and the levels of GSH, GST, GSH-PX, SOD, CAT and MDA in liver tissue were detected. The histopathological changes of liver tissues were observed after HE staining. Nrf2 translocation in liver tissue was detected by Western blotting, and real-time PCR was used to measure the expression levels of GCLC, NQO1 and HO-1 mRNA. The results showed that pretreatment with Tan significantly prevented the TP induced liver injury as indicated by reducing the activities of AST, ALT and LDH (P < 0.01). Tan pretreatment also prevented TP-induced oxidative stress in the mice liver by inhibiting MDA and restoring the levels of GSH, GST, SOD and CAT (P < 0.05). Parallel to these changes, pretreatment with Tan could attenuate histopathologic changes induced by TP. Furthermore, the results indicated that Tan pretreatment caused nuclear accumulation of Nrf2 as well as induction of mRNA expression of antioxidant response element (ARE)-driven genes such as GCLC, NQO1 and HO-1. These results indicated that Tan could protect against TP-induced acute liver injury via the activation of Nrf2/ARE pathway.

Comparative pharmacokinetics of paclitaxel after oral administration of Taxus yunnanensis extract and pure paclitaxel to rats.

Taxus yunnanensis (T. yunnanensis) is endemic to China and has been used in traditional medicine for the treatment of cancer, diabetic ailments and others. Paclitaxel is a representative antitumor compound in the Taxus species. The pharmacokinetic behavior of paclitaxel after oral administration of the crude extract of T. yunnanensis has not been investigated. This study attempts to compare the pharmacokinetics of paclitaxel after an oral administration of the crude extract of the twigs and leaves of T. yunnanensis and pure paclitaxel. A UPLC and a UPLC/MS/MS analysis method were developed for the determination of paclitaxel in T. yunnanensis extract and in the comparative pharmacokinetic study. Caco-2 cells were used to investigate the transport profile of paclitaxel in vitro. In the pharmacokinetic study, rats were randomly grouped and administered with T. yunnanensis extract or pure paclitaxel. The results showed that the AUC and C(max) of paclitaxel in rats receiving the T. yunnanensis extract were significantly increased than those receiving the pure paclitaxel, and the in vitro Caco-2 cell monolayer transport study found that the coexisting constituents in the extract of T. yunnanensis could inhibit the efflux of paclitaxel. These findings suggested that the oral absorption and bioavailability of paclitaxel in T. yunnanensis extract were remarkably higher when compared with the pure paclitaxel, and the coexisting constituents in the T. yunnanensis extract might play an important role for the enhancement of the oral absorption and bioavailability of paclitaxel.

Protective effect of schisandrin B against cyclosporine A-induced nephrotoxicity in vitro and in vivo.

Schisandrin B (Sch B) is an active ingredient of the fruit of Schisandra chinensis. It has many therapeutic effects arising from its tonic, sedative, antitussive and antiaging activities and is also used in the treatment of viral and chemical hepatitis. The aim of this study was to investigate the protective effects of Sch B on cyclosporine A (CsA)-induced nephrotoxicity in mice and HK-2 cells (a human proximal tubular epithelial cell line). After gavage with Sch B (20 mg/kg) or olive oil (vehicle), mice received CsA (30 mg/kg) by subcutaneous injection once daily for four weeks. Renal function, histopathology, and tissue glutathione (GSH) and malondialdehyde (MDA) levels were evaluated after the last treatment. The effects of Sch B on CsA-induced oxidative damage in HK-2 cells were investigated by measuring cell viability, the release of lactate dehydrogenase (LDH), the level of reactive oxygen species (ROS), and the cellular GSH and ATP concentrations. Cellular apoptosis was assessed by flow cytometry. Treatment with Sch B in CsA-treated mice significantly suppressed the elevation of blood urea nitrogen (BUN) and serum creatinine levels and attenuated the histopathological changes. Additionally, Sch B also decreased renal MDA levels and increased GSH levels in CsA-treated mice. Using an in vitro model, Sch B (2.5, 5 and 10 µM) significantly increased the cell viability and reduced LDH release and apoptosis induced by CsA (10 µM) in HK-2 cells. Furthermore, Sch B increased the intracellular GSH and ATP levels and attenuated CsA-induced ROS generation. In conclusion, Sch B appears to protect against CsA-induced nephrotoxicity by decreasing oxidative stress and cell death.

Leucine deprivation stimulates fat loss via increasing CRH expression in the hypothalamus and activating the sympathetic nervous system.

We previously showed that leucine deprivation decreases abdominal fat mass largely by increasing energy expenditure, as demonstrated by increased lipolysis in white adipose tissue (WAT) and uncoupling protein 1 (UCP1) expression in brown adipose tissue (BAT). The goal of the present study was to investigate the possible involvement of central nervous system (CNS) in this regulation and elucidate underlying molecular mechanisms. For this purpose, levels of genes and proteins related to lipolysis in WAT and UCP1 expression in BAT were analyzed in wild-type mice after intracerebroventricular administration of leucine or corticotrophin-releasing hormone antibodies, or in mice deleted for three ß-adrenergic receptors, after being maintained on a leucine-deficient diet for 7 d. Here, we show that intracerebroventricular administration of leucine significantly attenuates abdominal fat loss and blocks activation of hormone sensitive lipase in WAT and induction of UCP1 in BAT in leucine-deprived mice. Furthermore, we provide evidence that leucine deprivation stimulates fat loss by increasing expression of corticotrophin-releasing hormone in the hypothalamus via activation of stimulatory G protein/cAMP/protein kinase A/cAMP response element-binding protein pathway. Finally, we show that the effect of leucine deprivation on fat loss is mediated by activation of the sympathetic nervous system. These results suggest that CNS plays an important role in regulating fat loss under leucine deprivation and thereby provide novel and important insights concerning the importance of CNS leucine in the regulation of energy homeostasis.