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Background and aim: Inflammatory diseases often result in bone loss due to persistent inflammation, which activates osteoclasts and increases bone resorption. Oxysophocarpine (OSC), a bioalkaloid extracted from the roots of Sophora japonica and other leguminous plants, has neuroprotective and anti-tumor properties. However, it is still uncertain whether OSC can effectively inhibit the differentiation of osteoclasts and bone resorption. Therefore, this study explored the potential role of OSC in osteoclast formation and inflammatory osteolysis and its underlying mechanisms. Experimental procedure: This study involved inducing primary mouse bone marrow macrophages (BMMs) into osteoclasts using macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-κB ligand (RANKL) and examined the effects of OSC on osteoclast (OC) differentiation, function, and intracellular reactive oxygen species (ROS) production. The impact of OSC on the expression of osteoclast-specific genes and inflammation-related factors was assessed using real-time quantitative PCR. Additionally, changes in oxidative stress-related factors, NF-κB, and MAPK signaling pathways were examined using western blotting. Finally, this study investigated the influence of OSC on a mouse cranial bone resorption model induced by titanium (Ti) particles in vivo. Results: OSC inhibited OC differentiation and resorption and reduces intracellular ROS levels. Moreover, OSC suppressed IL-1β, TNF-α, IL-6, and osteoclast-specific gene transcription while increasing Nrf2 and HO-1 protein expression. Furthermore, OSC inhibited the expression and autoregulation of the NFATc1 gene, ultimately leading to a reduction in Ti particle-induced bone resorption in mice. Conclusion: OSC could be regarded as an innovative medication for the treatment of osteoclast-associated inflammatory osteolytic diseases. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) is an aggressive malignancy with limited effective treatments and ranks as the second most lethal tumor. Immunotherapy has brought great hope for HCC treatment. Oxysophocarpine is a bioactive alkaloid which poses various pharmacological functions including neuroprotective, anti‐virus, anti‐convulsant, and anti‐nociception. However, there is little systematic study of Oxysophocarpine against HCC and its underlying potential and mechanism combined with immunotherapy in HCC treatment remain poorly unknown. This study was aimed to investigate whether Oxysophocarpine can distinctly suppress HCC cells and sensitize the immunotherapy of CD8+ T cells against HCC. We used HepG2, Hepa1‐6, and primary CD8+ T cells to perform in vitro assays and Hepa1‐6 subcutaneous tumor to conduct in vivo assay. Oxysophocarpine inhibited the proliferation and increased the apoptosis of HepG2 and Hepa1‐6 cells, meanwhile suppressed the migration of HepG2 and Hepa1‐6 cells. Oxysophocarpine sensitized the Lag‐3 immunotherapy effect of CD8+ T cells against HCC in vivo and in vitro by decreasing Fibrinogen‐like protein 1 (FGL1) expression through downregulating IL‐6‐mediated JAK2/STAT3 signaling, whereas Oxysophocarpine treatment had a little effect of CD8+ T cells cytotoxicity function against HCC with PD‐1, Tim‐3, or TIGIT blockade. Our studies provided preclinical basis for clinical application of Oxysophocarpine. [ABSTRACT FROM AUTHOR]

Background/Aims: Nuclear factor erythroid 2-related factor 2 (Nrf2) is an oncogene in various types of cancers, including oral squamous cell carcinoma (OSCC). Oxysophocarpine (OSC) is a natural alkaloid that has multiple pharmacological activities. However, the biological functions and molecular mechanism underlying the effects of OSC on the growth and metastasis of OSCC are unclear. Methods: Nrf2 levels were determined in OSCC tissues and non-cancerous specimens by quantitative real-time PCR, western blotting, and immunohistochemistry (IHC) assays. The effects of OSC on OSCC cell growth and metastasis were explored (1) using 5-ethynyl-20-deoxyuridine staining and Cell Counting Kit-8, colony formation, flow cytometry, wound-healing, Transwell, and tube formation assays in vitro; and (2) by establishing a xenograft nude mouse model in vivo. The molecular mechanisms underlying the effects of OSC on the growth and metastasis of OSCC were investigated in vitro by western blotting, caspase-3 activity, and enzyme-linked immunosorbent assays, and in vivo by western blotting and IHC assays. Results: The expression levels of Nrf2 in OSCC tissues and in cell lines were much higher than in non-cancerous tissues and normal oral keratinocytes. The upregulation of Nrf2 was positively correlated with a high incidence of lymph node metastasis and advanced histological grade and TNM stage, but inversely associated with differentiation and survival of Xin Nie, MD Wenzhou Medical University, School and Hospital of Stomatology 373 College West Road, Wenzhou, Zhejiang 325035 (China) Tel. +86-577-56953758, Fax +86-577-56953758, E-Mail xinnie163@163.com R. Liu and J. Peng contributed equally to this work. [ABSTRACT FROM AUTHOR]

Oxysophocarpine (OSC), an alkaloid isolated from Sophora flavescens Ait, has been traditionally used as a medicinal agent based on the observed pharmacological effects. In this study, the direct effect of OSC against neuronal injuries induced by oxygen and glucose deprivation (OGD) in neonatal rat primary-cultured hippocampal neurons and its mechanisms were investigated. Cultured hippocampal neurons, which were exposed to OGD for 2 h followed by a 24 h reoxygenation, were used as an in vitro model of ischemia and reperfusion. 2-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and lactate dehydrogenase (LDH) assay were used to confirm neural damage and to further evaluate the protective effects of OSC. The concentration of intracellular-free calcium [Ca2+]i and mitochondrial membrane potential (MMP) were measured to determine the intracellular mechanisms and to further estimate the degree of neuronal damage. Changes in expression of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, p-ERK1/2, p-JNK1/2, and p-p38 MAPK were also observed in the in vitro model. It was shown that OSC (0.8, 2, or 5 µmol/L) significantly attenuated the increased absorbance of MTT, and the release of LDH manifests the neuronal damage by the OGD/R. Meanwhile, the pretreatment of the neurons during the reoxygenation period with OSC significantly increased MMP; it also inhibited [Ca2+]i the elevation in a dose-dependent manner. Furthermore, the pretreatment with OSC (0.8, 2, or 5 µmol/L) significantly down-regulated expressions of IL-1β, TNF-α, p-ERK1/2, p-JNK1/2, and p-p38 MAPK in neonatal rat primary-cultured hippocampal neurons induced by OGD/R injury. In conclusion, OSC displays a protective effect on OGD-injured hippocampal neurons by attenuating expression of inflammatory factors via down-regulated the MAPK signaling pathway. [ABSTRACT FROM AUTHOR]

Ulcerative colitis is an inflammation-related disease with a high recurrence risk. Oxysophocarpine (OSC) is a traditional Chinese medicine isolated from legumes and exerts vital functions on many human diseases. However, the OSC's role in ulcerative colitis has not been fully elucidated. This research aimed to investigate the OSC's impact on ulcerative colitis and its mechanisms. A mouse model of ulcerative colitis was induced by dextran sulphate sodium (DSS). The effect of OSC on ulcerative colitis was examined using Disease Activity Index detection, hematoxylin-eosin (HE) staining, and enzyme-linked immunosorbent assay (ELISA). Meanwhile, the mechanism of OSC in ulcerative colitis was assessed by immunohistochemistry assay, Western blot, HE staining, and ELISA. For the OSC's function in ulcerative colitis, OSC increased the mice weight, decreased Disease Activity Index scores, and alleviated colitis cell infiltration and epithelial cell destruction in DSS-induced ulcerative colitis. Also, OSC mitigated oxidative stress (decreased PGE2, MPO levels, and increased SOD levels) and inflammation (decreased IL-6, TNF-α and IL-1β levels) in DSS-induced ulcerative colitis. For the OSC's mechanism in ulcerative colitis, OSC inhibited the level of tumor necrosis factor receptor-associated Factor 6 (TRAF6) and the phosphorylation of nuclear factor-κB (NF-κB). TRAF6 overexpression abolished the effect of OSC on DSS-induced colon injury and its associated oxidative stress and inflammatory properties in ulcerative colitis. OSC decreased the TRAF6 level to reduce oxidative stress and inflammatory factors secretion in mice with DSS induced-ulcerative colitis. OSC ameliorated DSS-induced ulcerative colitis through repressing TRAF6/NF-κB axis, and it was reflected in low PGE2, MPO, IL-6, TNF-α, IL-1β levels, and high SOD levels. [Display omitted] • OSC alleviates intestinal injury in DSS-induced ulcerative colitis. • OSC attenuates DSS-induced oxidative stress in colonic mucosa. • OSC reduces oxidative stress and inflammation in DSS-induced ulcerative colitis by repressing TRAF6 pathway. [ABSTRACT FROM AUTHOR]

[Objectives] To establish the high performance liquid chromatography (HPLC) determination method for oxysophocarpine and optimize the extraction and purification technology of oxysophocarpine from Sophora alopecuroides by inverse three-phase membrane cycle. [Methods] Based on the single-factor experiment, the effects of aqueous phase and organic phase volume ratio, the concentration of sodium hydroxide, concentration of hydrochloric acid and extraction cycle time on extraction process of oxysophocarpine were investigated using orthogonal design method, to determine the optimal extraction process. [Results] The oxysophocarpine was determined by Shim-pack VP-ODS chromatographic column (4. 6 mm x 250 mm, 5 µm), mobile phase was methanol -0.2% phosphoric acid aqueous solution (7 - 93), gradient elu tion, flow rate was 1 mL/min, the sample size was 5 µm, column temperature was 30 °C, detection wavelength was 221 nm. Aqueous phase and organic phase volume ratio was 1-1, hydrochloric acid concentration was 0. 3 mol/L, sodium hydroxide concentration was 0.75 mol/L, water pump flow rate was 6 mL/min, 60 min cycle time. The extraction rate of oxysophocarpine 98.21 % within 60 min under the best experimental conditions. Oxysophocarpine has good linearity relationship within the range of 0.01 -0.7 mg/mL (r² = 0.9978, n = 6), the respective average recovery rate was 97.47% (RSD = 1.95%). [Conclusions] This extraction process is simple in operation, the organic solvent has low consumption, and can be used for extraction of alkaloid. [ABSTRACT FROM AUTHOR]

Epilepsy is one of the prevalent and major neurological disorders, and approximately one-third of the individuals with epilepsy experience seizures that do not respond well to available medications. We investigated whether oxysophocarpine (OSC) had anticonvulsant and neuroprotective property in the pilocarpine (PILO)-treated mice. Thirty minutes prior to the PILO injection, the mice were administrated with OSC (20, 40, and 80 mg/kg) once. Seizures and electroencephalography (EEG) were observed, and then the mice were killed for Nissl and Fluoro-jade B (FJB) staining. The oxidative stress was measured at 24 h after convulsion. Western blot analysis was used to examine the expressions of the Bax, Bcl-2, and Caspase-3. In this study, we found that pretreatment with OSC (40, 80 mg/kg) significantly delayed the onset of the first convulsion and status epilepticus (SE) and reduced the incidence of SE and mortality. Analysis of EEG recordings revealed that OSC (40, 80 mg/kg) significantly reduced epileptiform discharges. Furthermore, Nissl and FJB staining showed that OSC (40, 80 mg/kg) attenuated the neuronal cell loss and degeneration in hippocampus. In addition, OSC (40, 80 mg/kg) attenuated the changes in the levels of Malondialdehyde (MDA) and strengthened glutathione peroxidase and catalase activity in the hippocampus. Western blot analysis showed that OSC (40, 80 mg/kg) significantly decreased the expressions of Bax, Caspase-3 and increased the expression of Bcl-2. Collectively, the findings of this study indicated that OSC exerted anticonvulsant and neuroprotective effects on PILO-treated mice. The beneficial effects should encourage further studies to investigate OSC as an adjuvant in epilepsy, both to prevent seizures and to protect neurons in brain. [ABSTRACT FROM AUTHOR]

Asthma is a high-incidence disease in the world. Oxysophocarpine (OSC), a quinolizidine alkaloid displays various pharmacological functions including anti-inflammation, neuroprotective, anti-virus and antioxidant. Here, we established mice and cell asthmatic model to explore the effects of OSC for asthma treatment. Mice were sensitized and challenged with ovalbumin (OVA) and treated with OSC before challenge. Enzyme-linked immuno sorbent assay (ELISA), hematoxylin and eosin (H&E), periodic acid-schiff (PAS), tolonium chloride staining and immunohistochemical assay were performed. OSC treatment inhibited inflammatory cell infiltration and mucus secretion in the airway, reduced IgE level in mouse serum and decreased IL-4, IL-5 production in bronchoalveolar lavage fluid (BALF). OSC also reduced the spleen index to regulate immune function. Meanwhile, NCI-H292 cells were induced by lipopolysaccharide (LPS) to simulate airway epithelial injury. OSC pretreatment decreased the IL-6 and IL-8 cytokine levels, mucin 5 AC expression, and mucin 5 AC mRNA level in the cell model. Further, OSC suppressed the phosphorylation of c-Jun N-terminal kinase (JNK), and activator protein 1 (AP-1, Fos and Jun). These findings revealed that OSC alleviated bronchial asthma associated with JNK/AP-1 signaling pathway. [Display omitted] [ABSTRACT FROM AUTHOR]

Oxysophocarpine (OSC), an active and toxic quinolizidine alkaloid, is highly valued in Sophora flavescens Ait. and Subprostrate sophora Root. OSC is used to treat inflammation and hepatitis for thousands of years in China. This study aims to investigate the CYP450-mediated reduction responsible for metabolizing OSC and to evaluate the absorption and metabolism of OSC in rat in situ. Four metabolites were identified, with sophocarpine (SC) as the major metabolite. SC formation was rapid in human and rat liver microsomes (HLMs and RLMs, respectively). The reduction rates in the liver are two fold higher than in the intestine, both in humans and rats. In HLMs, inhibitors of CYP2C9, 3A4/5, 2D6, and 2B6 had strong inhibitory effects on SC formation. Meanwhile, inhibitors of CYP3A and CYP2D6 had significant inhibition on SC formation in RLMs. Human recombinant CYP3A4/5, 2B6, 2D6, and 2C9 contributed significantly to SC production. The permeability in rat intestine and the excretion rates of metabolites were highest in the duodenum (p < 0.05), and the absorbed amount of OSC in duodenum and jejunum was concentration-dependent. The metabolism could be significantly decreased by CYP3A inhibitor ketoconazole. In conclusion, the liver was the main organ responsible for OSC metabolism. First-pass metabolism via CYP3A4/5, 2B6, 2D6, and 2C9 may be the main reason for the poor OSC bioavailability. [ABSTRACT FROM AUTHOR]

Abstract: Matrine (MT)- and oxymatrine (OMT)-imprinted monodisperse polymers have been prepared by precipitation polymerization. The prepared molecularly imprinted polymers (MIPs) for MT and OMT, MIPMT and MIPOMT, were monodispersed microspheres of 3.3 and 3.9μm in diameter, respectively. Binding experiments and Scatchard analyses revealed that two classes of binding sites were formed on MIPMT and MIPOMT. In addition to shape recognition, ionic and hydrophobic interactions seemed to affect the retention and recognition of MT and OMT on MIPMT and MIPOMT, respectively, in low acetonitrile content, and ionic and hydrophilic interactions affected these properties in high acetonitrile content. MIPMT was used to selectively extract MT and sophocarpine (13,14-dehydromatrine) from Sophora flavescens root, while MIPOMT was used to extract OMT and oxysophocarpine (13,14-dehydrooxymatrine). [Copyright &y& Elsevier]

Abstract: In this paper, the enthalpies of dissolution of oxysophocarpine in water were measured using a RD496-2000 Calvet microcalorimeter at 298.15K under atmospheric pressure, showing that the dissolution process was an endothermic and exothermic oscillation behavior. A series of thermodynamics functions and the half-life were obtained by thermodynamic and kinetic methods. The results show that this work not only provide a simple method for the determination of the half-life for a drug but also offer a theoretical reference for the clinical application of resveratrol. [Copyright &y& Elsevier]

Abstract: A rapid, sensitive and selective high-performance liquid chromatography tandem mass spectrometric method (HPLC-MS) has been developed and validated for the simultaneous determination of matrine (MT), oxymatrine (OMT) and oxysophocarpine (OSP) in rat plasma after oral administration of Sophora flavescens Ait. extract using pseudoephedrine hydrochloride as an internal standard (I.S.). The three analytes were extracted from the plasma samples by liquid–liquid extraction with chloroform. The chromatographic separation was accomplished on a Kromasil C18 column (150mm×4.6mm). Detection was performed on a single quadrupole mass spectrometer by selected ion monitoring (SIM) mode via electrospray ionization (ESI) source. The total run time was 12min between injections. The assay had a lower limit of quantification of 1.0ng/ml for MT, 2.0ng/ml for OMT and 2.0ng/ml for OSP using 200μl of plasma. The calibration curves were linear in the measured range. The overall precision and accuracy for all concentrations of quality controls and standards was better than 15%. The proposed method enables unambiguous identification and quantification of MT, OMT and OSP in vivo. This was the first report on determination of the major quinolizidine alkaloids in rat plasma after oral administration of Sophora flavescens Ait. extract. The results provided a meaningful basis for evaluating the clinical applications of the herbal medicine. [Copyright &y& Elsevier]

Hepatocellular carcinoma (HCC) is an aggressive solid tumor with restricted therapeutics. Lenvatinib is the second approved frontline drug for advanced HCC, however lenvatinib-resistant cases have been reported in clinical. Overexpression of fibroblast growth factor receptor (FGFR1) has been found to be associated with advanced HCC. This study was aimed to investigate the relationship between FGFR1 overexpression and lenvatinib resistance, and explore the potential candidate that can sensitize lenvatinib against FGFR1-overexpressed HCC. Development of FGFR1 overexpression was accomplished in Hep3B and HepG2 cell lines by pCDH-FGFR1 lentiviral vector. In vitro, cell proliferation, colony formation, cell migration and cell apoptosis assays were used to explore the effect of lenvatinib and Oxysophocarpine. In vivo, BALB/c nude mice were burdened with subcutaneous FGFR1-overexpressed Hep3B tumor to assess the therapeutic effect of lenvatinib and Oxysophocarpine. qRT-PCR and western blotting were further used to identify the underlying mechanism. Here, we revealed that overexpressed FGFR1 and its downstream AKT/mTOR and ERK signaling activation could induce lenvatinib resistance in HCC. In vivo and in vitro results showed Oxysophocarpine inhibited the proliferation and induced the apoptosis of FGFR1-overexpressed HCC cells. Oxysophocarpine could further sensitize FGFR1-overexpressed HCC cells to lenvatinib treatment. Mechanism studies revealed that Oxysophocarpine downregulated FGFR1 expression along with downstream AKT/mTOR and ERK signaling to sensitize lenvatinib against FGFR1-overexpressed HCC. These data collectively provided evidence that FGFR1 overexpression could be a potential cause of lenvatinib resistance and Oxysophocarpine could be an ideal combined therapy with lenvatinib in HCC treatment. [ABSTRACT FROM AUTHOR]