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1.
J Pharm Pharmacol ; 70(6): 778-786, 2018 Jun.
Article in English | MEDLINE | ID: mdl-29516498

ABSTRACT

OBJECTIVES: To explore the apoptotic effects and underlying mechanisms of nitidine chloride (NC) in epithelial ovarian cancer. METHODS: The MTT cell proliferation assay was used to detect the inhibitory effects of different concentrations of NC (0, 0.3125, 0.625, 1.25, 2.5, 5 and 10 µg/ml) in SKOV3 ovarian carcinoma cells. The number of apoptotic cells was observed by Hoechst staining and measured by flow cytometry. Quantitative PCR was used to measure the expression of Fas, Fas-associated death domain-containing protein (FADD), caspase-8 and caspase-3. RNA interference (RNAi) was used to determine whether caspase-8 played an important role in NC-induced apoptosis. KEY FINDINGS: Nitidine chloride inhibited the proliferation of SKOV3 cells (IC50 = 2.317 ± 0.155 µg/ml) after 24 h of treatment and induced apoptosis (15.9-64.3%). Compared with the control group, a significant increase in Fas, FADD, caspase-8 and caspase-3 gene expression was observed in the NC-treated groups (P < 0.05). After silencing caspase-8 by RNAi, the antiproliferative activity and pro-apoptotic activity of NC in SKOV3 cells decreased (P < 0.05). CONCLUSIONS: Our study showed that NC induced apoptosis in SKOV3 cells by activating the Fas signalling pathway, and caspase-8 played an important role in this process.


Subject(s)
Apoptosis/drug effects , Benzophenanthridines/pharmacology , Cell Proliferation/drug effects , Signal Transduction/drug effects , fas Receptor/metabolism , Benzophenanthridines/antagonists & inhibitors , Caspase 3 , Caspase 8/biosynthesis , Caspase Inhibitors/pharmacology , Cell Line, Tumor , Dose-Response Relationship, Drug , Fas-Associated Death Domain Protein/biosynthesis , Humans , RNA, Small Interfering/pharmacology
2.
Br J Pharmacol ; 173(16): 2543-54, 2016 08.
Article in English | MEDLINE | ID: mdl-27324234

ABSTRACT

BACKGROUND AND PURPOSE: Nitidine chloride (NC), a benzophenanthridine alkaloid, has various biological properties including anticancer and analgesic activities. The aim of the present study was to evaluate the role of organic cation transporter 2 (OCT2) and multidrug and toxin extrusion 1 (MATE1) in the renal disposition and nephrotoxicity of NC. EXPERIMENTAL APPROACH: MDCK cells stably expressing human OCT2 and/or hMATE1 were used to investigate the OCT2- and MATE1-mediated transport of NC. In addition, the accumulation of NC and its potential toxicity were studied in rat primary-cultured proximal tubular (rPCPT) cells and in rats in vivo. KEY RESULTS: NC was found to be a high-affinity substrate of both OCT2 and MATE1 with high cytotoxicity in MDCK-hOCT2/hMATE1 and MDCK-hOCT2 compared to mock cells. The OCT2 inhibitors, cimetidine and (+)-tetrahydropalmatine ((+)-THP), significantly reduced NC accumulation and cytotoxicity in MDCK-hOCT2, MDCK-hOCT2/hMATE1 and rPCPT cells. Severe kidney damage with high levels of blood urea nitrogen and lactate dehydrogenase (LDH), reduced levels of alkaline phosphatase (ALP) and pathological changes were found in rats after 20 days of successive i.v. doses of NC (5 mg·kg(-1) ·day(-1) ). Concomitantly, the concentration of NC in the kidney reached similar high levels at 2 h after the last dose of the 20 day treatment as those observed at 0.5 h after a single i.v. dose of 5 mg·kg(-1) . CONCLUSIONS AND IMPLICATIONS: Our data indicate that NC-induced nephrotoxicity might be mainly attributed to OCT2-mediated extensive renal uptake and weak tubular secretion by MATE1.


Subject(s)
Benzophenanthridines/pharmacokinetics , Benzophenanthridines/toxicity , Kidney/drug effects , Kidney/metabolism , Organic Cation Transport Proteins/metabolism , Animals , Benzophenanthridines/antagonists & inhibitors , Benzophenanthridines/chemistry , Berberine Alkaloids/pharmacology , Cell Survival/drug effects , Cells, Cultured , Cimetidine/pharmacology , Dogs , Dose-Response Relationship, Drug , Humans , Madin Darby Canine Kidney Cells , Male , Organic Cation Transport Proteins/antagonists & inhibitors , Rats , Rats, Sprague-Dawley , Solute Carrier Family 22 Member 5 , Structure-Activity Relationship
3.
J Pharmacol Sci ; 127(1): 135-44, 2015 Jan.
Article in English | MEDLINE | ID: mdl-25704029

ABSTRACT

Previous studies show that several pathways are involved in sanguinarine-induced apoptotic cell death, including AKT downregulation, inhibition of NF-kB activation, mediation of ROS production, downregulation of anti-apoptosis proteins XIAP and cIAP-1, upregulation of BAX, and downregulation of BCL2. In this study, we found out that the quenching of ROS generation by N-acetyl-l-cysteine (NAC), a scavenger of ROS, reversed sanguinarine-induced apoptosis effects, also we found out that sanguinarine-induced rat hepatic stellate T6 cells (HSC-T6 cells) apoptosis was correlated with the generation of increased ROS, which was followed by the activation of caspase-8 (-3, -6, and -9), and the decreasing in the miltochondrial membrane potential (MMP) and the down-regulation of anti-apoptotic protein Bcl-2. It is not clear whether BCL2's downregulation relates to its promoter methylation and miR-15a/16-1 expression which can bind to BCL2 3'-UTR (un-translation reagon). We showed that sanguinarine-induced down regulation of BCL2 was associated with the increased methylation rate of BCL2 promotor district and the increased expression of miR-15a/16-1. HSC-T6 cells treatment with 5-Aza-2'-deoxycytidine (5'-Aza-CdR) impeded sanguinarine-induced BCL2 promotor district methylation and recovered BCL2's expression. Over expression of BCL2 using pEGFP-N1 vector decreased sanguinarine-induced HSC-T6 cells apoptotic death significantly but not completely. These observations clearly showed that BCL2 down regulation was associated with its promoter methylation and miR-15a/16-1 upregulation in sanguinarine-induced Rat HSC-T6 cells.


Subject(s)
Apoptosis/drug effects , Benzophenanthridines/pharmacology , Isoquinolines/pharmacology , MicroRNAs/metabolism , Promoter Regions, Genetic/drug effects , Proto-Oncogene Proteins c-bcl-2/metabolism , Animals , Azacitidine/analogs & derivatives , Azacitidine/pharmacology , Benzophenanthridines/antagonists & inhibitors , Cell Line , Cell Survival/drug effects , Decitabine , Down-Regulation , Isoquinolines/antagonists & inhibitors , Membrane Potential, Mitochondrial/drug effects , Methylation/drug effects , Rats , Reactive Oxygen Species/metabolism , Up-Regulation
4.
Biol Chem ; 393(7): 647-58, 2012 Jul.
Article in English | MEDLINE | ID: mdl-22944669

ABSTRACT

We show that the plant quaternary benzo[c]phenanthridine alkaloid sanguilutine (SL) is a strong inducer of caspase-independent non-apoptotic death in human melanoma cells. Necrostatin-1, a specific inhibitor of necroptosis, completely reversed the cytotoxic effect of SL, suggesting that necroptosis was a predominant type of cell death induced by SL in these cells. In addition, we showed that SL can trigger an autophagic response, as confirmed by GFP-LC3 puncta formation and LC3-II accumulation. Interestingly, we observed a significant decrease in the viability of melanoma cells treated with combination of autophagy inhibitors (3-methyladenine, bafilomycin-A1 and LY294002) and SL. Our results further indicated that autophagy may serve as a pro-survival mechanism, delaying the induction of necroptosis in melanoma cells. The ability of SL to induce caspase-independent non-apoptotic cell death (necroptosis) suggests its possible therapeutic potential in the treatment of apoptosis-resistant melanoma tumours. Furthermore, SL might serve as a useful tool for studying the mechanisms of necroptosis and autophagy induction and the interplay between these two processes.


Subject(s)
Antineoplastic Agents/pharmacology , Autophagy/drug effects , Benzophenanthridines/pharmacology , Melanoma/pathology , Adenine/analogs & derivatives , Adenine/pharmacology , Antineoplastic Agents/antagonists & inhibitors , Benzophenanthridines/antagonists & inhibitors , Caspases/metabolism , Cell Line, Tumor , Cell Survival/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Humans , Imidazoles/pharmacology , Indoles/pharmacology , Microtubule-Associated Proteins/metabolism , Necrosis/chemically induced , Proto-Oncogene Proteins c-bcl-2/metabolism
5.
Toxicol Lett ; 205(3): 285-92, 2011 Sep 10.
Article in English | MEDLINE | ID: mdl-21722720

ABSTRACT

In this study, we examined the cytotoxic effects of sanguinarine, a phytoalexin with antimicrobial, anti-oxidant, anti-inflammatory and pro-apoptotic effects, on the blastocyst stage of mouse embryos, subsequent embryonic attachment and outgrowth in vitro and in vivo implantation via embryo transfer. Blastocysts treated with 0.5-2 µM sanguinarine exhibited significantly increased apoptosis and a corresponding decrease in total cell number. Notably, the implantation success rates of blastocysts pretreated with sanguinarine were lower than that of their control counterparts. Moreover, in vitro treatment with 0.5-2 µM sanguinarine was associated with increased resorption of post-implantation embryos and decreased fetal weight. Our results collectively indicate that sanguinarine induces apoptosis and retards early post-implantation development in vitro and in vivo. In addition, sanguinarine induces apoptotic injury effects on mouse blastocysts through intrinsic and extrinsic apoptotic signaling processes to impair sequent embryonic development. However, the extent to which sanguinarine exerts teratogenic effects on early human development is not known at present, and further studies are required to establish effective protection strategies against its cytotoxic effects.


Subject(s)
Apoptosis/drug effects , Benzophenanthridines/toxicity , Blastocyst/drug effects , Embryonic Development/drug effects , Isoquinolines/toxicity , Sesquiterpenes/toxicity , Teratogens/toxicity , Animals , Benzophenanthridines/antagonists & inhibitors , Blastocyst/pathology , Blastocyst Inner Cell Mass/drug effects , Blastocyst Inner Cell Mass/pathology , Caspase Inhibitors , Cell Proliferation/drug effects , Cysteine Proteinase Inhibitors/pharmacology , Cysteine Proteinase Inhibitors/therapeutic use , Dose-Response Relationship, Drug , Ectogenesis/drug effects , Embryo Implantation/drug effects , Embryo Loss/chemically induced , Embryo Loss/drug therapy , Embryo Transfer , Female , Fetal Weight/drug effects , Isoquinolines/antagonists & inhibitors , Mice , Mice, Inbred ICR , Random Allocation , Sesquiterpenes/antagonists & inhibitors , Signal Transduction/drug effects , Phytoalexins
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