Xie-Bai-San increases NSCLC cells sensitivity to gefitinib by inhibiting Beclin-1 mediated autophagosome formation.

BACKGROUND: Autophagy, a cellular process involving lysosomal self-digestion, plays a crucial role in recycling biomolecules and degrading dysfunctional proteins and damaged organelles. However, in non-small cell lung cancer (NSCLC), cancer cells can exploit autophagy to survive metabolic stress and develop resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs), which reduce treatment efficacies. Currently, most studies have found that late-stage autophagy inhibitors can hinder EGFR-TKIs resistance, while research on early-stage autophagy inhibitors is still limited. PURPOSE: This study investigates the mechanism via which the Xie-Bai-San (XBS) formula enhances NSCLC cell sensitivity to gefitinib, revealing the relationship between XBS-induced cell death and the inhibition of autophagosome formation. METHODS: Cell viability was assessed using CCK-8 and EdU assays, lentivirus transfection was utilized to generate PC9 cells harboring the PIK3CA E545K mutation (referred to as PC9-M), autophagic flux was monitored using mCherry-GFP-LC3 adenovirus. Protein expression and colocalization were observed through immunofluorescence staining. The interaction between Bcl-2 and Beclin-1 in PC9-GR and PC9-M cells was determined via co-immunoprecipitation (Co-IP) assay, cell apoptosis was assessed by flow cytometry and PI staining, and overall survival analysis of lung adenocarcinoma patients was conducted using the TCGA database. In vivo experiments included a patient-derived xenograft (PDX) model with EGFR and PIK3CA mutations and subcutaneous mice xenografts of NSCLC cell lines (PC9 and PC9-GR). In addition, autophagic vesicles in mouse tumor tissues were observed via transmission electron microscopy analysis. RESULTS: XBS effectively inhibits the proliferation of gefitinib-resistant NSCLC cells and induces apoptosis both in vitro and in vivo. Mechanistically, XBS suppresses gefitinib-induced autophagic flux by inhibiting autophagy through the upregulation of p-mTOR and Bcl-2 and downregulation of Beclin-1. Additionally, XBS enhances the interaction between Bcl-2 and Beclin-1, and the overexpression of Beclin-1 promotes NSCLC cell proliferation and counteracts XBS-induced cell death, while XBS demonstrates minimal impact on autophagosome-lysosome fusion or lysosome function. CONCLUSION: This study reveals a novel role for the XBS formula in impeding autophagy initiation and demonstrates its potential as a candidate drug to counteract autophagy-induced treatment resistance in NSCLC.

Dual-Functional Laser-Guided Magnetic Nanorobot Collectives against Gravity for On-Demand Thermo-Chemotherapy of Peritoneal Metastasis.

Combining hyperthermic intraperitoneal chemotherapy with cytoreductive surgery is the main treatment modality for peritoneal metastatic (PM) carcinoma despite the off-target effects of chemotherapy drugs and the ineluctable side effects of total abdominal heating. Herein, a laser-integrated magnetic actuation system that actively delivers doxorubicin (DOX)-grafted magnetic nanorobot collectives to the tumor site in model mice for local hyperthermia and chemotherapy is reported. With intraluminal movements controlled by a torque-force hybrid magnetic field, these magnetic nanorobots gather at a fixed point coinciding with the position of the localization laser, moving upward against gravity over a long distance and targeting tumor sites under ultrasound imaging guidance. Because aggregation enhances the photothermal effect, controlled local DOX release is achieved under near-infrared laser irradiation. The targeted on-demand photothermal therapy of multiple PM carcinomas while minimizing off-target tissue damage is demonstrated. Additionally, a localization/treatment dual-functional laser-integrated magnetic actuation system is developed and validated in vivo, offering a potentially clinically feasible drug delivery strategy for targeting PM and other intraluminal tumors.

Aberrant dentato-rubro-thalamic pathway in action tremor but not rest tremor: A multi-modality magnetic resonance imaging study.

AIMS: The purpose of this study was to clarify the dentato-rubro-thalamic (DRT) pathway in action tremor in comparison to normal controls (NC) and disease controls (i.e., rest tremor) by using multi-modality magnetic resonance imaging (MRI). METHODS: This study included 40 essential tremor (ET) patients, 57 Parkinson's disease (PD) patients (29 with rest tremor, 28 without rest tremor), and 41 NC. We used multi-modality MRI to comprehensively assess major nuclei and fiber tracts of the DRT pathway, which included decussating DRT tract (d-DRTT) and non-decussating DRT tract (nd-DRTT), and compared the differences in DRT pathway components between action and rest tremor. RESULTS: Bilateral dentate nucleus (DN) in the ET group had excessive iron deposition compared with the NC group. Compared with the NC group, significantly decreased mean diffusivity and radial diffusivity were observed in the left nd-DRTT in the ET group, which were negatively correlated with tremor severity. No significant difference in each component of the DRT pathway was observed between the PD subgroup or the PD and NC. CONCLUSION: Aberrant changes in the DRT pathway may be specific to action tremor and were indicating that action tremor may be related to pathological overactivation of the DRT pathway.

Hypericin: A natural anthraquinone as promising therapeutic agent.

BACKGROUND: Hypericin is a prominent secondary metabolite mainly existing in genus Hypericum. It has become a research focus for a quiet long time owing to its extensively pharmacological activities especially the anti-cancer, anti-bacterial, anti-viral and neuroprotective effects. This review concentrated on summarizing and analyzing the existing studies of hypericin in a comprehensive perspective. METHODS: The literature with desired information about hypericin published after 2010 was gained from electronic databases including PubMed, SciFinder, Science Direct, Web of Science, China National Knowledge Infrastructure databases and Wan Fang DATA. RESULTS: According to extensive preclinical and clinical studies conducted on the hypericin, an organized and comprehensive summary of the natural and artificial sources, strategies for improving the bioactivities, pharmacological activities, drug combination of hypericin was presented to explore the future therapeutic potential of this active compound. CONCLUSIONS: Overall, this review offered a theoretical guidance for the follow-up research of hypericin. However, the pharmacological mechanisms, pharmacokinetics and structure activity relationship of hypericin should be further studied in future research.

Renal transporter OAT1 and PPAR-α pathway co-contribute to icaritin-induced nephrotoxicity.

This study aimed to investigate the potential nephrotoxicity of icaritin and the underlying mechanism by in vitro-in vivo experiment technology combined with proteomics technology. First, icaritin showed a significant cytotoxic effect on HK-2 cells, which was accompanied by increased LDH and TNF-α in the supernatant, decreased protein expressions of Bcl-2 and increased Bax and enhanced apoptosis of HK-2 cells as measured by TUNEL staining. Moreover, icaritin induced obvious tubular damage and up-regulation of BUN and CRE levels in plasma in mice. Second, intracellular uptake of icaritin was considerably higher in hOAT1-HEK293 cells than in mock-HEK293 cells, suggesting that icaritin might accumulate in renal cells via OAT1 uptake. Importantly, icaritin caused significant changes in the PPAR signaling pathway in HK2 cells through proteomic analysis. Then, in vitro and in vivo results verified that icaritin significantly downregulated the protein expression of PPAR-α as well as downregulated APOB, ACSL3, ACSL4, and upregulated 5/12/15-HETE, implying that a lipid metabolism disorder was involved in the icaritin-induced nephrotoxicity. Finally, icaritin was found to increase the accumulation of iron and LPO levels while reducing the activity of GPX4, suggesting that ferroptosis was involved in the nephrotoxicity induced by icaritin.

Comparative Inhibitory Effects of Natural Biflavones from Ginkgo against Human CYP1B1 in Recombinant Enzymes and MCF-7 Cells.

Human cytochrome P450 1B1 (CYP1B1) is an extrahepatic enzyme overexpressed in many tumors and associated with angiogenesis. Ginkgetin, isoginkgetin, sciadopitysin, and amentoflavone, the primary biflavones found in Ginkgo biloba, have excellent anti-inflammatory and anti-tumor effects. However, the effect of biflavones on CYP1B1 activities remains unknown. In this study, 7-ethoxyresorufin O-deethylation (EROD) was used to characterize the activities of CYP1 families. The impacts of four ginkgo biflavones on CYP1B1 activity and the cellular protein expression of CYP1B1 were systematically investigated. The results showed that amentoflavone with six hydroxyl substituents exhibited the most potent selective inhibitory effect on CYP1B1 activity with IC50 of 0.054 µM in four biflavones. Sciadopitysin, with three hydroxyl and three methoxy substituents, had the weakest inhibitory activity against CYP1B1. Ginkgetin and isoginkgetin, both with four hydroxyl and two methoxy substituents, showed similar inhibitory intensity towards CYP1B1 with IC50 values of 0.289 and 0.211 µM, respectively. Kinetic analysis showed that ginkgetin and amentoflavone inhibited CYP1B1 in a non-competitive mode, whereas sciadopitysin and isoginkgetin induced competitive or mixed types of inhibition. Notably, four ginkgo biflavones were also confirmed to suppress the protein expressions of CYP1B1 and AhR in MCF-7. Furthermore, molecular docking studies indicated more hydrogen bonds formed between amentoflavone and CYP1B1, which might explain the strongest inhibitory action towards CYP1B1. In summary, these findings suggested that biflavones remarkably inhibited both the activity and protein expression of CYP1B1 and the inhibitory activities enhanced with the increasing hydroxyl substitution, providing new insights into the anti-tumor potentials of biflavones.

Anti-Postmenopausal osteoporosis effects of Isopsoralen: A bioinformatics-integrated experimental study.

Isopsoralen (IPRN), which comes from the fruit of Psoralea corylifolia, has been identified as a kind of phytoestrogen and has been proven to be effective for the treatment of osteoporosis (OP). However, the mechanisms underlying IPRN's anti-OP effects, especially the anti-postmenopausal osteoporosis (PMOP) effects, remain indistinct. Thus, this study aimed to investigate the effects and mechanisms of IPRN's anti-PMOP activity. In this study, the bioinformatics results predicted that IPRN could resist PMOP by targeting EGFR, AKT1, SRC, CCND1, ESR1 (ER-α), AR, PGR, BRCA1, PTGS2, and IGF1R. An ovariectomized (OVX) mice model and a H2 O2 -induced bone marrow mesenchyml stem cells (BMSCs) model confirmed that IPRN could inhibit the bone loss induced by OVX in mice and promote the osteogenic differentiation in H2 O2 -induced BMSCs by inhibiting oxidative stress and apoptosis. Moreover, IPRN could significantly produce the above effects by upregulating ESR1. IPRN might be a therapeutic agent for PMOP by acting as an estrogen replacement agent and a natural antioxidant.

Transcatheter Arterial Embolization Combined with Anti-vascular Agent Combretastatin A4 Phosphate Inhibits Growth and Vascularization of Liver Tumor in an Animal Model.

OBJECTIVE: This study aimed to investigate the effect of combretastatin A4 phosphate (CA4P) on proliferation, migration, and capillary tube formation of human umbilical vein endothelial cells (HUVECs) and the efficacy of transcatheter arterial embolization combined with CA4P in the treatment of rabbit VX2 liver tumor. METHODS: The effects of different concentrations of CA4P on proliferation, migration and capillary tube formation of HUVECs were investigated by cell proliferation assay, wound healing assay and capillary tube formation assay, respectively. Thirty-two rabbits implanted with liver VX2 tumors were randomly divided into 4 groups. After catheterization of the left hepatic artery, the infusion was performed using normal saline (group A), CA4P aqueous solution (group B), lipiodol and polyvinyl alcohol particles (group C), and CA4P lipiodol emulsion and polyvinyl alcohol particles (group D), respectively. Half of the animals in each group were euthanized for immunohistochemical analysis to evaluate microvessel density (MVD) at 3 days post-treatment. The other half were examined by MRI and histology to evaluate tumor growth and necrosis at 7 days post-treatment. RESULTS: CA4P could inhibit the proliferation, migration, and tube formation of HUVECs in cell experiments. After interventional treatment, the level of MVD in group D was lower than that in group C (P<0.01). The tumor volume in group C or D was lower than that in group A or B (P<0.01). The tumor necrosis rate was higher in group D than in the other groups. CONCLUSION: The study suggests that CA4P could inhibit the proliferation, migration, and capillary tube formation of HUVECs, and transcatheter arterial embolization combined with CA4P could inhibit the growth of VX2 tumor and obviously induce tumor necrosis.

"Jing-Ning Granules" Can Alleviate Attention Deficit Hyperactivity Disorder in Rats by Modulating Dopaminergic D2/D1-Like Receptor-Mediated Signaling Pathways.

Background: Attention deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder characterized by attention deficit, hyperactivity, and impulsivity. Jing-Ning Granules (JNG) is a traditional Chinese medicine (TCM) that can alleviate ADHD. Although JNG is commonly used for the effective treatment of ADHD and has obtained the national invention patent, the exact mechanism of action remains unclear. Objective: In this study, we examined the effect and mechanism of JNG in spontaneously hypertensive rats (SHRs). We hypothesized that JNG affects dopaminergic D2/D1-like receptors and related pathways. Materials and Methods: Six rat groups were used in the experiment: Wistar-Kyoto rats (WKY, control group) and five SHR groups, including a model group; atomoxetine (ATX, positive control) group; and low, medium, and high-dose JNG groups. The corresponding treatments were daily administered to each group for 6 weeks. A behavioral test, including a step-down test and open field test (OFT), was carried out at the end of treatment. After the behavioral test, all animals were sacrificed, and the brain tissue was collected and analyzed ex vivo; histopathological analysis was performed to assess the pathological changes of the hippocampus; expression of D1-like and D2-like receptors, sensor protein calmodulin (CaM), protein kinase A (PKA), and calcium/calmodulin-dependent serine/threonine protein kinase (CaMKII) in the striatum and hippocampus was measured by western blot and real-time quantitative PCR (RT-PCR); cyclic adenosine monophosphate (cAMP) levels in the striatum were analyzed using an enzyme-linked immunosorbent assay (ELISA), while the level of Ca2+ in the striatum was analyzed by a calcium kit. Results: Our results showed that ATX or JNG could ameliorate the hyperactive/impulsive behavior and cognitive function of ADHD by promoting neuroprotection. Mechanistically, ATX or JNG could prompt the expressions of Dl-like and D2-like receptors and improve the mRNA and protein levels of cAMP/PKA and Ca2+/CAM/CAMKII signaling pathways. Conclusion: These results indicate that JNG can produce therapeutic effects by regulating the balance of D2/D1-like receptor-mediated cAMP/PKA and Ca2+/CaM/CaMKII signaling pathways.

Clinical efficacy and psychological improvement in elderly patients with knee arthritis after comprehensive traditional Chinese medicine care.

OBJECTIVE: To analyze the effect of comprehensive traditional Chinese medicine (TCM) care on the clinical efficacy and psychological improvement of elderly patients with knee arthritis. METHODS: Retrospective analysis was conducted on 114 patients with knee osteoarthritis who underwent arthroscopic minimally invasive surgery in our hospital from January 2018 to January 2022. Among them, 55 patients received routine nursing served as the control group (CG), and the remaining 59 patients received comprehensive TCM care as the observation group (OG). Patients were re-examined two weeks after discharge from the hospital, and the knee joint function recovery effect, pain score, nursing satisfaction, as well as the changes in adverse mood, hospitalization expenses and length of hospitalization during the treatment were compared between two groups. Logistic regression analysis was used to identify factors affecting the curative effect of patients. RESULTS: Compared with the CG, the OG held a significantly better clinical efficacy and lower knee joint score and visual analogue scale (VAS) score (P<0.05). After intervention, the OG showed markedly higher SF-36 score as well as notably declined scores of anxiety and depression than the CG (P<0.05). Quadriceps peak torque ratio (H/Q), as well as satisfaction of patients on nursing in the OG was comparatively better than those of the CG (P<0.05). For hospitalization costs and length of stay, the OG has proven to be more economical and effective with lower cost in both two indexes (P<0.05). Age, course of disease, and nursing program were risk factors affecting the efficacy of treatment in patients (P<0.05). CONCLUSION: Comprehensive TCM care markedly improved the clinical efficacy of the elderly with knee arthritis, relieved patients' pain, and improved knee function and quality of life, as well as reduced patients' anxiety and economic pressure.

Total flavonoids of Rhizoma drynariae promote angiogenesis and osteogenesis in bone defects.

Bone defects are difficult to heal, which conveys a heavy burden to patients' lives and their economy. The total flavonoids of Rhizoma drynariae (TFRD) can promote the osteogenesis of distraction osteogenesis. However, the dose effect is not clear, the treatment period is short, and the quality of bone formation is poor. In our study, we observed the long-term effects and dose effects of TFRD on bone defects, verified the main ingredients of TFRD in combination with network pharmacology for the first time, explored its potential mechanism, and verified these findings. We found that TFRD management for 12 weeks regulated osteogenesis and angiogenesis in rats with 4-mm tibial bone defects through the PI3K/AKT/HIF-1α/VEGF signaling pathway, especially at high doses (135 mg kg-1  d-1 ). The vascularization effect of TFRD in promoting human umbilical vein endothelial cells was inhibited by PI3K inhibitors. These results provide a reference for the clinical application of TFRD.

The Effect of Hemoglobin Concentration on Hyperbaric Oxygen and Non-hyperbaric Oxygen in the Treatment of Hypertensive Intracerebral Hemorrhage After Operation at the High Altitude.

Background: The prognosis of hypertensive intracerebral hemorrhage (HICH) is poor at high altitudes. The objective of this study was to explore whether hyperbaric oxygen (HBO) can improve the results of computed tomography perfusion (CTP) imaging and the neurological function of patients with HICH, and influence the hemoglobin concentration. Method: The patients with HICH were treated with puncture and drainage. Twenty-one patients (51.22% of 41 patients in total) were treated with HBO after the operation, and the other patients received conventional treatment. CTP was performed twice, and all indices were measured. Scatter plots were used to determine the effect of hemoglobin concentration on CTP imaging. Receiver operating characteristic (ROC) curves were plotted to analyze the effects of hemoglobin concentration and hematoma volume on recovery results. The patients were followed up for 6 months. Results: Forty-one patients with HICH were treated with puncture and drainage. In total, 21 were treated with HBO after the operation, and 20 received conventional treatment as the control group. No significant differences in the CBV and CBF values of the two groups were noted before treatment. After 10 days, the values of CBV and CBF in the HBO group were significantly higher than those in the control group. A scatter diagram showed there was no significant in the HBO group, but significant correlation for the CBV and CBF values in the control group's hematoma center and margin. The ROC curves showed that hematoma volume had an influence on prognosis of the control group. The Glasgow Coma Scale (GOS) scores of the HBO group were significantly higher than those of the control group (p < 0.05). Conclusions: HBO therapy can improve the postoperative CBV and CBF values of patients with HICH and ameliorate their prognoses. There was no significant correlation between HBO group and hemoglobin concentration on admission.

Protective effect of Rhein against vancomycin-induced nephrotoxicity through regulating renal transporters and Nrf2 pathway.

Vancomycin (VCM)'s nephrotoxicity limits its application and therapeutic efficiency. The aim of this study was to determine the protective effect of rhein against VCM-induced nephrotoxicity (VIN). VIN models were established in rats and NRK-52E cells. Rhein up-regulated the expressions of renal organic anion transporter (Oat) 1, Oat3, organic cation transporter 2 (Oct2), multidrug resistance-associated protein 2 (Mrp2), mammal multidrug and toxin extrusion proteins 1 (Mate 1) and P-glycoprotein (P-gp) to facilitate the efflux of plasma creatinine, blood urea nitrogen (BUN), and plasma indoxyl sulfate. Rhein increased the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) to regulate the expression of Mrp2, P-gp, and Mate 1. The increased level of superoxide dismutase (SOD), decreased level of malondialdehyde (MDA) and reduced number of apoptosis cells were observed after treatment of rhein. Rhein decreased the number of apoptosis cells as well as increased the expression of B-cell lymphoma-2 (Bcl-2) and decreased expressions of Bcl-2-like protein 4 (Bax). ML385, as a typical inhibitor of Nrf2, reversed the protective effects of rhein in cells. Rhein oriented itself in the site of Keap1, inhibiting the Keap1-Nrf2 interaction. Rhein ameliorated VIN mainly through regulating the expressions of renal transporters and acting on Nrf2 pathway.

Coptis chinensis polysaccharides dynamically influence the paracellular absorption pathway in the small intestine by modulating the intestinal mucosal immunity microenvironment.

BACKGROUND: Traditional Chinese Medicine decoctions (TCMDs) can be used to prepare outstanding pharmaceutical preparations by the patient themselves. Small molecular active ingredients and macromolecular polysaccharides are inevitably co-existed in TCMDs. Different from the pharmacological synergies among small molecules, the macromolecular polysaccharides in TCMDs might contribute to disease treatment in several ways, although it is frequently overlooked. HYPOTHESIS/PURPOSE: This study proposes that the oral bioavailability of the water-insoluble alkaloids of Coptis chinensis Franch. (Ranunculaceae) (C. chinensis) decoction may be attributed to the co-existing C. chinensis polysaccharides (CCPs) dynamically influencing the small intestine microenvironment and regulating the modulation of the paracellular absorption pathway. METHODS: First, the effects of CCPs on the oral bioavailability of the main active ingredient of C. chinensis, berberine, were evaluated in vivo. Next, a series of in situ experimental models of intestinal perfusion and models of isolated jejunal mucosa, Caco-2 cell monolayer membranes, and microfold-like cells were established to assess the correlation among CCPs, intestinal mucosal immunity, and paracellular absorption in the small intestine. RESULTS: It was observed that CCPs could be endocytosed by the microfold cells on the surface of Peyer's patches, allowing CCPs to activate the lymphocytes, modulate the balance of Th1/Th2, control the secretion of immune effectors IFN-γ and IL-4, and finally regulate the tight junctions in the intestinal epithelial cells. This was a dynamic process with the movement of CCPs in the gastrointestinal tract that altered the flora distribution and functioning of the TLR/NF-κB signal pathway in the small intestine. CONCLUSION: The dynamical regulation of CCP on the immune microenvironment of small intestine is responsible for its promotion on the health controlling effects of C. chinensis in traditional dosage forms of decoction.

Design and characterization of gambogic acid-loaded mixed micelles system for enhanced oral bioavailability.

The aim of this study was to develop a gambogic acid-loaded mixed micelles (GA-M) system, using Kolliphor HS15 and lecithin, for enhancement of oral bioavailability. GA-M was prepared using the thin film hydration method, and particle size and zeta potential indexes were used to determine the optimized formulation was optimized with taking particle size, zeta potential as indexes. The optimal GA-M system had a mean particle size in the nanometer range (87.22 ± 0.68 nm) and zeta potential greater than 20 mV in magnitude (-21.63 ± 1.69 mV) at a 1:1 proportion of HS15: lecithin. Additionally, the carriers had a high entrapment efficiency (98.32 ± 3.52%) and drug loading (4.68 ± 0.17%). Furthermore, the in vitro GA release characteristics followed first-order kinetics, suggesting that the release of the molecule was achieved both by medium diffusion and structural erosion. Transport elucidation in Caco-2 cells demonstrated that the efflux ratio of encapsulated GA was dramatically decreased from 1.42 to 0.76, and pharmacokinetic studies showed that the oral bioavailability of GA-M was 2.3 times higher than that of free GA, indicating that HS15/lecithin mixed micelles could promote absorption in the gastrointestinal tract. Overall, these results present a micelle system suitable for oral delivery, with increased solubility and oral bioavailability of GA.

Unraveling the Structure-Dependent Inhibitory Effects of Ginsenoside Series Compounds on Human Cytochrome P450 1B1.

BACKGROUND: Cytochrome P450 1B1(CYP1B1) is an extrahepatic P450 isoenzyme that can participate in processes of undermining the effectiveness and safety of anti-cancer therapy. Ginsenosides are the main active ingredients in ginseng, which possesses rich pharmacological activities, including anti-cancer activity and organ protection. However, the effect of ginsenosides on the activity of CYP1B1 remains unclear. OBJECTIVE: The present study aimed to investigate the inhibitory effect of ginsenosides on CYP1B1 and reveal the structure-inhibitory activity relationship. METHODS: Firstly, recombinant CYP1B1 and EROD reactions were used to evaluate the inhibitory effect of ginsenosides. Secondly, molecular docking was used to simulate the interactions between ginsenosides and CYP1B1. Finally, the structure-inhibitory activity relationship was analyzed. RESULTS: The ginsenosides, Rb2, Rd, and Rg3, significantly inhibited CYP1B1; the ginsenoside Rd showed the strongest inhibition effect, with a Ki value of 47.37 µM in non-competitive mode. Notably, ginsenoside Rd formed hydrogen bonds with two key amino acid residues of CYP1B1, and one bond was between the glycosyl in position 20 and ALA330, which also made ginsenoside Rd close to the heme iron of CYP1B1. In contrast, ginsenosides, Rb2 and Rg3, which showed weaker inhibition, interacted with only one CYP1B1 residue by the hydrogen bond, which was far away from the heme iron. Finally, the structure-inhibitory activity relationship analysis demonstrated that the number of glycosyls in position 20 and the type of sapogenins in the ginsenoside structure are the key factors determining inhibitory activity. Meanwhile, ALA330 was a vital amino acid in the potent inhibition of CYP1B1 by ginsenosides. CONCLUSION: A structure-dependent inhibitory effect on CYP1B1 was revealed for ginsenosides, among which ginsenoside Rd showed the strongest inhibition due to its mono-glycosyl in position 20 of the ginsenoside parent structure. These findings would provide evidence for the synthesis of novel CYP1B1 inhibitors to augment the anti-cancer therapeutic effect.

Substrate-dependent Inhibition of Hypericin on Human Carboxylesterase 2: Implications for Herb-drug Combination.

BACKGROUND: Hypericin is the main active ingredient of St. John's wort, a Chinese herb commonly used for treating depression. Previous studies shown that hypericin can strongly inhibit human cytochrome P450 (CYP) enzyme activities; however, its potential interactions that inhibit human carboxylesterases 2 (hCE2) are unclear. PURPOSE: This study aimed to investigate the inhibitory effect of hypericin on hCE2. METHODS: The inhibition mechanism of hypericin on hCE2 was studied by using N-(2-butyl-1,3-dioxo-2,3-dihydro- 1H-phenalen-6-yl)-2-chloroacetamide (NCEN). The type of inhibition of hypericin on hCE2 and the corresponding inhibition constant (Ki) value were determined. The inhibition of hypericin on hCE2 in living cells was discussed. The risk of herb-drug interactions (HDI) of hypericin in vivo was predicted by estimating the area under the drug concentration-time curve (AUC) in the presence or absence of hypericin. To understand the inhibition mechanism of hypericin on the activity of hCE2 in-depth, molecular docking was performed. RESULTS: The half-maximal inhibitory concentration (IC50) values of hypericin against the hydrolysis of NCEN and irinotecan (CPT-11) were calculated to be 26.59 µM and 112.8 µM, respectively. Hypericin inhibited the hydrolysis of NCEN and CPT-11. Their Ki values were estimated as 10.53 µM and 81.77 µM, respectively. Moreover, hypericin distinctly suppressed hCE2 activity in living cells. In addition, the AUC of hCE2 metabolic drugs with metabolic sites similar to NCEN was estimated to increase by up to 5 % in the presence of hypericin. More importantly, the exposure of CPT-11 in the intestinal epithelium was predicted to increase by 2 % - 69 % following the oral coadministration of hypericin. Further, molecular simulations indicated that hypericin could strongly interact with ASP98, PHE307, and ARG355 to form four hydrogen bonds within hCE2. CONCLUSION: These findings regarding the combination of hypericin-containing herbs and drugs metabolized by hCE2 are of considerable clinical significance.

Activation of PGC-1α via isoliquiritigenin-induced downregulation of miR-138-5p alleviates nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD), a metabolic disease, has received wide attention worldwide. However, there is no approved effective drug for NAFLD treatment. In the study, H&E and Oil Red O staining were employed to detect liver histopathological changes and the accumulation of lipid droplets. Quantitative real-time PCR, Western blot, bioinformatics, luciferase assay, immunofluorescence staining, reactive oxygen species (ROS), and siRNA were used to further elucidate the mechanism of isoliquiritigenin (ISL) against NAFLD. The results showed that ISL significantly reduced the liver-to-body weight ratios and biochemical index. And the staining results showed that ISL remarkedly ameliorated liver histopathological changes of NAFLD. Furthermore, ISL significantly increased the levels of PPARα, CPT1α, and ACADS, which were involved in lipid metabolism, and inhibited the ROS, TNF-α, IL-1ß, and IL-6 expression by activating PGC-1α. Bioinformatics and luciferase assay analysis confirmed that miR-138-5p might bind to PGC-1α mRNA in NAFLD. Importantly, the expression of miR-138-5p was increased in the NAFLD, which was significantly decreased by ISL. In addition, the miR-138-5p inhibitor also promoted lipid metabolism and inhibited inflammatory response in NAFLD via PGC-1α activation. The above results demonstrate that ISL alleviates NAFLD through modulating miR-138-5p/PGC-1α-mediated lipid metabolism and inflammatory reaction in vivo and in vitro.

Crosstalk of LncRNA HOTAIR and SP1-mediated repression of PDK1 contributes to ß-Elemene-inhibited proliferation of hepatocellular carcinoma cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Hepatocellular carcinoma (HCC) is a liver malignancy which lacks effective treatment and has a poor prognosis. ß-Elemene refers to a natural Curcuma wenyujin-derived single molecular entity, which exhibits various biological activities, and is especially well-known for it's antitumor properties. AIM OF THE RESEARCH: LncRNA HOTAIR, SP1, and PDK1 have displayed oncogenic roles in many tumors, participating in the initiation and progression of cancers by mediating multiple signaling pathways. However, there are only a few reports about their roles and mutual relationship in the growth of HCC cells. Therefore, this study aimed to investigate the expression of LncRNA HOTAIR, SP1, and PDK1 and their interaction with ß-Elemene in HCC cells. MATERIALS AND METHODS: MTT, a Colony formation assay, and flow cytometry were employed to evaluate the growth of HCC and LO2 cells under ß-Elemene. LncRNA HOTAIR, SP1 and PDK1 plasmids were transfected into HCC cells by a transient transfection assay, and the expression and interaction of LncRNA HOTAIR, SP1 and PDK1 were assessed via qRT-PCR and western blotting. RESULTS: ß-Elemene suppressed HCC cell growth through the downregulation of LncRNA HOTAIR, SP1 and PDK1. The results demonstrated a reciprocal interaction among LncRNA HOTAIR, SP1 and PDK1. Exogenous overexpression LncRNA HOTAIR or SP1 eliminated the suppressive effects of ß-Elemene on them, and both of which regulated PDK1 expression in HCC cells. Additionally, exogenously overexpressed SP1 or LncRNA HOTAIR prevented ß-Elemene inhibition of the protein-level expression of PDK1, whereas overexpressing PDK1 had no effect on SP1, though it still weakened the inhibition of cell growth and LncRNA HOTAIR expression by ß-Elemene. CONCLUSION: ß-Elemene suppresses HCC cell proliferation via through the regulation of LncRNA HOTAIR, SP1, PDK1 and their interaction.

Isoliquiritigenin-mediated miR-23a-3p inhibition activates PGC-1α to alleviate alcoholic liver injury.

BACKGROUND: Alcoholic liver disease (ALD), one of the most prevalent forms of liver disease, has received wide attention worldwide. However, limited efficient and appropriate therapeutic agents were responded to ALD. Isoliquiritigenin (ISL), a flavonoid isolated from liquorice, possesses multiple pharmacological activities. PURPOSE: The current study investigated the hepatoprotective effect of ISL against ALD and further elucidate the involvement of miR-23a-3p/peroxisome proliferative activated receptor-γ coactivator 1 alpha (PGC-1α) in vivo and in vitro experiments. STUDY DESIGN AND METHODS: In the study, H&E and Oil Red O staining were employed to detect liver histopathological changes and the accumulation of lipid droplets. Quantitative real-time PCR, bioinformatics, luciferase assay, immunofluorescence staining, reactive oxygen species (ROS), Western blot, and siRNA were used to further explore the mechanism of ISL protection. RESULTS: ISL significantly reduced the liver-to-body weight ratios and biochemical index. The staining results showed that ISL remarkedly ameliorated the histopathological changes in the liver. Furthermore, ISL promoted fatty acid metabolism via induction in the expression of PGC-1α-target genes PPARα, CPT1α, and ACADs, and inhibited the ROS, TNF-α, IL-1ß, and IL-6 expression. Bioinformatics and Luciferase assay analysis confirmed that miR-23a-3p might bind to PGC-1α mRNA in ALD. Significantly, the expression of miR-23a-3p was increased in the ALD, which was significantly decreased by ISL. In addition, the miR-23a-3p inhibitor also promoted lipid metabolism in ALD via PGC-1α activation. CONCLUSIONS: We first demonstrated that ISL could alleviate ALD, and further verified that ISL exerted protective effects through modulating miR-23a-3p/PGC-1α-mediated lipid metabolism in vivo and in vitro.