RESUMO
Background/purpose: Econazole is an antifungal drug. Antifungal activity of econazole against non-dermatophyte molds was reported. Econazole inhibited Ca2+ channels and stimulated cytotoxicity in lymphoma and leukemia cells. Ca2+ cations are crucial second envoy that triggers various processes. This research was aimed to investigate action of econazole on Ca2+ levels and cytotoxicity in OC2 human oral cancer cells. Materials and methods: Cytosolic Ca2+ levels ([Ca2+]i) were detected employing fura-2 as a probe in a RF-5301PC spectrofluorophotometer (Shimadzu). Cytotoxicity was determined using 4-[3-[4-lodophenyl]-2-4(4-nitrophenyl)-2H-5-tetrazolio-1,3-benzene disulfonate] (WST-1) to detect fluorescence changes. Results: Econazole at 10-50 µmol/L provoked [Ca2+]i raises. Forty % of 50 µml//L econazole-induced signal was diminished when external Ca2+ was eliminated. The Ca2+ influx provoked by econazole was suppressed by different degrees by store-induced Ca2+ influx suppressors SKF96365 and nifedipine; GF109203X (a protein C [PKC] inhibitor); an extracellular signaling pathway (ERK) 1/2 blocker PD98059, and phospholipase A2 suppressor aristolochic acid, but was enhanced by phorbol 12-myristate 13 acetate (PMA; a PKC activator) by 18%. Without external Ca2+, econazole-caused [Ca2+]i raises were abolished by thapsigargin. In contrast, econazole partially suppressed the [Ca2+]i raises caused by thapsigargin. U73122 fell short to change econazole-caused [Ca2+]i responses. Econazole (10-70 µmol/L) elicited cytotoxicity in a dose-dependent fashion. Blockade of 50 µmol/L econazole-induced [Ca2+] rises with BAPTA/AM enhanced econazole-induced cytotoxicity by 72%. Conclusion: Econazole evoked [Ca2+]i raises and provoked cytotoxicity in a concentration-dependent manner in OC2 human oral cancer cells. In Ca2+-containing solution, BAPTA/AM enhanced 50 µmol/L econozole-induced cytotoxicity.
RESUMO
Gomisin A is a dietary lignan compound isolated from the fruit of Schisandra chinensis and has many pharmacological properties, including hepato-protective, anti-diabetic, and anti-oxidative activities. However, the benefit of gomisin A is still not well understood. The action of gomisin A is diverse. However, the effect of gomisin A on Ca2+ signaling in prostate cancer cells is unknown. Ca2+ is a pivotal second envoy that triggers and regulates cellular processes such as apoptosis, fertilization, energy transduction, secretion, and protein activation. The goal of this study was to explore the action of gomisin A on [Ca2+]i and cytotoxicity in PC3 prostate cancer cells. Gomisin A at 100-200 µM provoked [Ca2+]i raises. 20% of the response was reduced by removing external Ca2+. The Ca2+ influx provoked by gomisin A was suppressed by 20% by store-caused Ca2+ entry suppressors: econazole, SKF96365, nifedipine; also by phorbol 12-myristate 13 acetate and GF109203X. Without external Ca2+, gomisin A-caused [Ca2+]i raises were abolished by thapsigargin. In contrast, gomisin A suppressed the [Ca2+]i raises caused by thapsigargin. U73122 fell short to change gomisin A-caused [Ca2+]i responses. Gomisin A (20-100 µM) elicited cytotoxicity in a dose-associated fashion. Blockade of [Ca2+] elevations with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/acetoxy methyl failed to inhibit cytotoxicity of gomisin A. Collectively, gomisin A evoked [Ca2+]i raises and provoked cytotoxicity in a Ca2+-dissociated fashion in prostate cancer cells.
Assuntos
Lignanas , Neoplasias da Próstata , Cálcio/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular , Ciclo-Octanos , Dioxóis , Humanos , Lignanas/farmacologia , Masculino , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/metabolismo , Tapsigargina/farmacologia , Fosfolipases Tipo C/metabolismoRESUMO
Hepatotoma is the leading type of primary liver cancer in adults and third cause of death in the world. Hydroxytyrosol is a natural phenol existing in olive (Olea europaea L.). Hydroxytyrosol is the chief ingredient of olive oil, which was early deemed to be the most robust antioxidant in olive oil. Hydroxytyrosol is known to inhibit various types of cancer by different methods. This study was aimed to delineate the action of hydroxytyrosol on viability and [Ca2+]i in HepG2 hepatoma cells. Fura-2 was used to detect [Ca2+]i, and WST-1 assays were applied to explore cell cytotoxicity. Hydroxytyrosol elicited [Ca2+]i raises. Eliminating external Ca2+ diminished the Ca2+ signal by 30%. Hydroxytyrosol-evoked Ca2+ influx was diminished by 20% by three inhibitors of store-operated Ca2+ channels and by a protein kinase C activator and an inhibitor. In the absence of Ca2+, thapsigargin eradicated hydroxytyrosol-provoked [Ca2+]i raises. Suppression of phospholipase C (PLC) with U73122, a PLC inhibitor, did not inhibit hydroxytyrosol-elicited [Ca2+]i raises. Hydroxytyrosol reduced cell viability. This cytotoxic action was not reversed by preincubation with BAPTA/AM, a cytosolic Ca2+ binder. In sum, in HepG2 hepatoma cells, hydroxytyrosol elicited [Ca2+]i raises by provoking PLC-unrelated discharge of Ca2+ from ER and Ca2+ influx through PKC-sensitive store-operated Ca2+ entry. In addition, hydroxytyrosol elicited Ca2+-dissociated cytotoxicity.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Olea , Apoptose , Cálcio/metabolismo , Sinalização do Cálcio , Carcinoma Hepatocelular/tratamento farmacológico , Linhagem Celular Tumoral , Sobrevivência Celular , Etanol , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Olea/metabolismo , Fenóis , Álcool Feniletílico/análogos & derivados , Fosfolipases Tipo C/metabolismoRESUMO
Thioridazine, belonging to first-generation antipsychotic drugs, is a prescription used to treat schizophrenia. However, the effect of thioridazine on intracellular Ca2+ concentration ([Ca2+]i) and viability in human liver cancer cells is unclear. This study examined whether thioridazine altered Ca2+ signaling and viability in HepG2 human hepatocellular carcinoma cells. Ca2+ concentrations in suspended cells were measured using the fluorescent Ca2+-sensitive dye fura-2. Cell viability was examined by WST-1 assay. Thioridazine at concentrations of 25-100 µM induced [Ca2+]i rises. Ca2+ removal reduced the signal by 20%. Thioridazine (100 µM) induced Mn2+ influx suggesting of Ca2+ entry. Thioridazine-induced Ca2+ entry was inhibited by 20% by protein kinase C (PKC) activator (phorbol 12-myristate 13 acetate) and inhibitor (GF109203X) and by three inhibitors of store-operated Ca2+ channels: nifedipine, econazole, and SKF96365. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin (TG) abolished thioridazine-evoked [Ca2+]i rises. On the other hand, thioridazine preincubation completely inhibited the [Ca2+]i rises induced by TG. Furthermore, U73122 totally suppressed the [Ca2+]i rises induced by thioridazine via inhibition of phospholipase C (PLC). Regarding cytotoxicity, at 30-80 µM, thioridazine reduced cell viability in a concentration-dependent fashion. This cytotoxicity was not prevented by preincubation with 1,2-bis (2-aminophenoxy) ethane-N, N, N', N'-tetraacetic acid-acetoxymethyl ester (BAPTA/AM) (a Ca2+ chelator). To conclude, thioridazine caused concentration-dependent [Ca2+]i rises in HepG2 human hepatoma cells by inducing Ca2+ release from the endoplasmic reticulum via PLC-associated pathways and Ca2+ influx from extracellular medium through PKC-sensitive store-operated Ca2+ entry. In addition, thioridazine induced cytotoxicity in a Ca2+-independent manner.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Apoptose , Cálcio , Sinalização do Cálcio , Morte Celular , Linhagem Celular Tumoral , Sobrevivência Celular , Humanos , Tioridazina , Fosfolipases Tipo CRESUMO
Tectorigenin, a traditional Chinese medicine, is isolated from the flower of plants such as Pueraria thomsonii Benth. It is an O-methylated isoflavone, a type of flavonoid. Previous studies have shown that tectorigenin evoked various physiological responses in different models, but the effect of tectorigenin on cytosolic-free Ca2+ levels ([Ca2+]i) and cytotoxicity in renal tubular cells is unknown. Our research explored if tectorigenin changed Ca2+ signal transduction and viability in Madin-Darby Canine Kidney (MDCK) renal tubular cells. [Ca2+]iin suspended cells were measured by applying the fluorescent Ca2+-sensitive probe fura-2. Viability was explored by using water-soluble tetrazolium-1 as a fluorescent dye. Tectorigenin at concentrations of 5-50 µM induced [Ca2+]irises. Ca2+ removal reduced the signal by approximately 20%. Tectorigenin (50 µM) induced Mn2+ influx suggesting of Ca2+ entry. Tectorigenin-induced Ca2+ entry was inhibited by 10% by three inhibitors of store-operated Ca2+ channels, namely, nifedipine, econazole, and SKF96365. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin inhibited 83% of tectorigenin-evoked [Ca2+]irises. Conversely, treatment with tectorigenin abolished thapsigargin-evoked [Ca2+]irises. Inhibition of phospholipase C with U73122 inhibited 50% of tectorigenin-induced [Ca2+]irises. Tectorigenin at concentrations between 10 and 60 µM killed cells in a concentration-dependent fashion. Chelation of cytosolic Ca2+ with 1,2-bis (2-aminophenoxy)ethane-N, N, N', N'-tetraacetic acid/acetoxy methyl did not reverse tectorigenin's cytotoxicity. Our data suggest that, in MDCK cells, tectorigenin evoked [Ca2+]irises and induced cell death that was not associated with [Ca2+]irises. Therefore, tectorigenin may be a Ca2+-independent cytotoxic agent for kidney cells.
Assuntos
Sinalização do Cálcio , Animais , Apoptose , Cálcio , Linhagem Celular Tumoral , Sobrevivência Celular , Cães , Isoflavonas , Fosfolipases Tipo CRESUMO
Literature has shown that diosgenin, a naturally occurring sapogenin, inducedcytotoxic effects in many cancer models. This study investigated the effect of diosgenin on intracellular Ca2+ concentration ([Ca2+ ]i) and cytotoxicity in PC3 human prostate cancer cells. Diosgenin (250-1000 µM) caused [Ca2+ ]i rises which was reduced by Ca2+ removal. Treatment with thapsigargin eliminated diosgenin-induced [Ca2+ ]i increases. In contrast, incubation with diosgeninabolished thapsigargin-caused [Ca2+ ]i increases. Suppression of phospholipase C with U73122 eliminated diosgenin-caused [Ca2+ ]i increases. Diosgenin evoked Mn2+ influx suggesting that diosgenin induced Ca2+ entry. Diosgenin-induced Ca2+ influx was suppressed by PMA, GF109203X, and nifedipine, econazole, or SKF96365. Diosgenin (250-600 µM) concentration-dependently decreased cell viability. However, diosgenin-induced cytotoxicity was not reversed by chelation of cytosolic Ca2+ with BAPTA/AM. Together, diosgenin evoked [Ca2+ ]i increases via Ca2+ release and Ca2+ influx, and caused Ca2+ -non-associated deathin PC3 cells. These findings reveal a newtherapeutic potential of diosgenin for human prostate cancer.
Assuntos
Antineoplásicos/farmacologia , Cálcio/metabolismo , Diosgenina/farmacologia , Neoplasias da Próstata/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Masculino , Células PC-3 , Sapogeninas/farmacologia , Fosfolipases Tipo C/antagonistas & inibidoresRESUMO
Glyburide is an agent commonly used to treat type 2 diabetes and also affects various physiological responses in different models. However, the effect of glyburide on Ca2+ movement and its related cytotoxicity in prostate cancer cells is unclear. This study examined whether glyburide altered Ca2+ signalling and viability in PC3 human prostate cancer cells and investigated those underlying mechanisms. Intracellular Ca2+ concentrations ([Ca2+ ]i ) in suspended cells were measured by using the fluorescent Ca2+ -sensitive dye fura-2. Cell viability was examined by WST-1 assay. Glyburide at concentrations of 100-1000 µM induced [Ca2+ ]i rises. Ca2+ removal reduced the signal by approximately 60%. In Ca2+ -containing medium, glyburide-induced Ca2+ entry was inhibited by 60% by protein kinase C (PKC) activator (phorbol 12-myristate 13 acetate, PMA) and inhibitor (GF109203X), and modulators of store-operated Ca2+ channels (nifedipine, econazole and SKF96365). Furthermore, glyburide induced Mn2+ influx suggesting of Ca2+ entry. In Ca2+ -free medium, inhibition of phospholipase C (PLC) with U73122 significantly inhibited glyburide-induced [Ca2+ ]i rises. Treatment with the endoplasmic reticulum (ER) Ca2+ pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) abolished glyburide-evoked [Ca2+ ]i rises. Conversely, treatment with glyburide abolished BHQ-evoked [Ca2+ ]i rises. Glyburide at 100-500 µM decreased cell viability, which was not reversed by pretreatment with the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA/AM). Together, in PC3 cells, glyburide induced [Ca2+ ]i rises by Ca2+ entry via PKC-sensitive store-operated Ca2+ channels and Ca2+ release from the ER in a PLC-dependent manner. Glyburide also caused Ca2+ -independent cell death. This study suggests that glyburide could serve as a potential agent for treatment of prostate cancer.
Assuntos
Sinalização do Cálcio/efeitos dos fármacos , Cálcio/metabolismo , Glibureto/farmacologia , Hipoglicemiantes/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Diabetes Mellitus Tipo 2/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Humanos , Masculino , Células PC-3 , Neoplasias da Próstata/metabolismo , Proteína Quinase C/metabolismoRESUMO
Chlorzoxazone is a skeletal muscle relaxant. However, the effect of chlorzoxazone on intracellular Ca2+ concentrations ([Ca2+]i) in oral cancer cells is unclear. This study examined whether chlorzoxazone altered Ca2+ signaling and cell viability in OC2 human oral cancer cells. [Ca2+]iin suspended cells was measured using the fluorescent Ca2+-sensitive dye fura-2. Cell viability was examined by water-soluble tetrazolium-1 assay. Chlorzoxazone (250-1000 µM) induced [Ca2+]irises in a concentration-dependent manner. Ca2+ removal reduced the signal by approximately 50%. Mn2+ has been shown to enter cells through similar mechanisms as Ca2+ but quenches fura-2 fluorescence at all excitation wavelengths. Chlorzoxazone (1000 µM) induced Mn2+ influx, suggesting that Ca2+ entry occurred. Chlorzoxazone-induced Ca2+ entry was inhibited by 20% by inhibitors of store-operated Ca2+ channels and protein kinase C (PKC) modulators. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin (TG) inhibited chlorzoxazone-evoked [Ca2+]irises by 88%. Conversely, treatment with chlorzoxazone-suppressed TG-evoked [Ca2+]irises 75%. Chlorzoxazone induced [Ca2+]irises by exclusively releasing Ca2+ from the endoplasmic reticulum. Inhibition of phospholipase C (PLC) with U73122 did not alter chlorzoxazone-induced [Ca2+]irises. PLC activity was not involved in chlorzoxazone-evoked [Ca2+]irises. Chlorzoxazone at 200-700 µM decreased cell viability, which was not reversed by pretreatment with Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/acetoxy methyl. In sum, in OC2 cells, chlorzoxazone induced [Ca2+]irises by evoking PLC-independent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via PKC-sensitive store-operated Ca2+ entry. Chlorzoxazone also caused Ca2+-independent cell death. Since [Ca2+]irises play a triggering or modulatory role in numerous cellular phenomena, the effect of chlorzoxazone on [Ca2+]iand cell viability should be taken into account in other in vitro studies.
Assuntos
Sinalização do Cálcio , Neoplasias Bucais , Apoptose , Cálcio , Linhagem Celular Tumoral , Sobrevivência Celular , Clorzoxazona , Humanos , Fosfolipases Tipo CRESUMO
Malathion, one of commonly used organophosphate insecticides, has a wide range of toxic actions in different models. However, the effect of this compound on Ca2+ homeostasis and its related cytotoxicity in glial cells is elusive. This study examined whether malathion evoked intracellular Ca2+ concentration ([Ca2+]i) rises and established the relationship between Ca2+ signaling and cytotoxicity in normal human astrocytes, rat astrocytes and human glioblastoma cells. The data show that malathion induced concentration-dependent [Ca2+]i rises in Gibco® Human Astrocytes (GHA cells), but not in DI TNC1 normal rat astrocytes and DBTRG-05MG human glioblastoma cells. In GHA cells, this Ca2+ signal response was reduced by removing extracellular Ca2+. In Ca2+-free medium, pretreatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin abolished malathion-induced [Ca2+]i rises. Conversely, incubation with malathion abolished thapsigargin-induced [Ca2+]i rises. Inhibition of phospholipase C (PLC) with U73122 also blocked malathion-induced [Ca2+]i rises. In Ca2+-containing medium, malathion-induced [Ca2+]i rises was inhibited by store-operated Ca2+ channel blockers (2-APB, econazole or SKF96365) and the protein kinase C (PKC) inhibitor GF109203X. Malathion (5-25⯵M) concentration-dependently caused cytotoxicity in GHA, DI TNC1 and DBTRG-05MG cells. This cytotoxic effect was partially prevented by prechelating cytosolic Ca2+ with BAPTA-AM (a selective Ca2+ chelator) only in GHA cells. Together, in GHA but not in DI TNC1 and DBTRG-05MG cells, malathion induced [Ca2+]i rises by inducing PLC-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via PKC-sensitive store-operated Ca2+ channels. Furthermore, malathion induced Ca2+-associated cytotoxicity, suggesting that Ca2+ chelating may have a protective effect on malathion-induced cytotoxicity in normal human astrocytes.
Assuntos
Cálcio/metabolismo , Malation/farmacologia , Animais , Sinalização do Cálcio/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Quelantes , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Humanos , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , RatosRESUMO
Terfenadine, an antihistamine used for the treatment of allergic conditions, affected Ca2+-related physiological responses in various models. However, the effect of terfenadine on cytosolic free Ca2+ levels ([Ca2+]i) and its related physiology in renal tubular cells is unknown. This study examined whether terfenadine altered Ca2+ signaling and caused cytotoxicity in Madin-Darby canine kidney (MDCK) renal tubular cells. The Ca2+-sensitive fluorescent dye fura-2 was used to measure [Ca2+]i. Cell viability was measured by the fluorescent reagent 4-[3-[4-lodophenyl]-2-4(4-nitrophenyl)-2H-5-tetrazolio-1,3-benzene disulfonate] water soluble tetrazolium-1 (WST-1) assay. Terfenadine at concentrations of 100-1000 µM induced [Ca2+]i rises concentration dependently. The response was reduced by approximately 35% by removing extracellular Ca2+. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) partly inhibited terfenadine-evoked [Ca2+]i rises. Conversely, treatment with terfenadine abolished BHQ-evoked [Ca2+]i rises. Inhibition of phospholipase C (PLC) with U73122 inhibited 95% of terfenadine-induced Ca2+ release. Terfenadine-induced Ca2+ entry was supported by Mn2+-caused quenching of fura-2 fluorescence. Terfenadine-induced Ca2+ entry was partly inhibited by an activator of protein kinase C (PKC), phorbol 12-myristate 13 acetate (PMA) and by three modulators of store-operated Ca2+ channels (nifedipine, econazole, and SKF96365). Terfenadine at 200-300 µM decreased cell viability, which was not reversed by pretreatment with the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA/AM). Together, in MDCK cells, terfenadine induced [Ca2+]i rises by evoking PLC-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via PKC-sensitive store-operated Ca2+ entry. Furthermore, terfenadine caused cell death that was not triggered by preceding [Ca2+]i rises.
Assuntos
Apoptose/efeitos dos fármacos , Sinalização do Cálcio/efeitos dos fármacos , Antagonistas não Sedativos dos Receptores H1 da Histamina/farmacologia , Túbulos Renais/patologia , Terfenadina/farmacologia , Animais , Sobrevivência Celular , Cães , Túbulos Renais/efeitos dos fármacos , Túbulos Renais/metabolismo , Células Madin Darby de Rim CaninoRESUMO
OBJECTIVE: Colchicine, extracted from plants of the genus Colchicum, is a commonly prescribed drug for inflammatory diseases. It has been shown that colchicine affected various physiological responses in different models. However, the effect of colchicine on cytosolic free Ca2+ levels ([Ca2+]i) and its related physiology in human oral cancer cells is unknown. This study examined whether colchicine altered Ca2+ homeostasis and caused cytotoxicity in OC2 human oral cancer cells. METHODS: The Ca2+-sensitive fluorescent dye fura-2 was used to measure [Ca2+]i. Cell viability was measured by the fluorescent reagent 4-[3-[4-lodophenyl]-2-4(4-nitrophenyl)-2H-5-tetrazolio-1,3-benzene disulfonate] water soluble tetrazolium-1 (WST-1) assay. RESULTS: Colchicine at concentrations of 250-650 µM induced [Ca2+]i rises concentration-dependently. The response was reduced by approximately 40% by removing extracellular Ca2+. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin inhibited colchicine-evoked [Ca2+]i rises. Conversely, treatment with colchicine inhibited thapsigargin-evoked [Ca2+]i rises. Inhibition of phospholipase C (PLC) with U73122 abolished colchicine-induced Ca2+ release. In Ca2+-containing medium, colchicine-induced Ca2+ entry was supported by Mn2+-caused quenching of fura-2 fluorescence and the entry was partly inhibited by protein kinase C (PKC) modulators (phorbol 12-myristate 13 acetate, PMA; and GF109203X) and by three modulators of store-operated Ca2+ channels (nifedipine, econazole and SKF96365). Colchicine at 250-650 µM decreased cell viability, which was not reversed by pretreatment with the Ca2+chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA/AM). CONCLUSIONS: In OC2 cells, colchicine induced [Ca2+]i rises by evoking PLC-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via PKC-sensitive store-operated Ca2+ entry. Furthermore, colchicine caused cell death that was not triggered by preceding [Ca2+]i rises.
Assuntos
Neoplasias Bucais , Alcaloides , Apoptose , Cálcio , Sinalização do Cálcio , Linhagem Celular Tumoral , Sobrevivência Celular , Colchicina , HumanosRESUMO
Malathion is one of the most widely used organophosphorus insecticides in agriculture. However, malathion may be involved in the etiology of human brain dysfunction. Induction of ROS has been proposed as a mechanism of malathion-induced poisoning cases, but there are few data regarding the effects of malathion on oxidative stress-associated neurotoxicity in human glial cells. The aim was to explore the mechanism underlying effects of malathion on neurotoxicity in Gibco® Human Astrocytes (GHA cells) and evaluate the protective effects of the antioxidant (N-acetylcysteine, NAC). Cell viability was measured by the cell proliferation reagent (WST-1). Antioxidant enzymes (glutathione peroxidase and catalase) were measured by an ELISA reader. Cell cycle distribution and ROS productions were detected by flow cytometry. Cell cycle-related protein levels (cyclin E1, CDK2, cyclin A2, CDK1/CDC2, or cyclin B1) and apoptotic protein levels (Bcl-2, Bax, and cleaved caspase-9/caspase-3) were analyzed by Western blotting. In GHA cells, treatment with malathion (10-25 µM) for 24 h concentration-dependently induced cytotoxicity and cell cycle arrest. In terms of oxidative stresses, malathion elevated intracellular ROS levels, but reduced glutathion and antioxidant enzyme levels. Treatment with NAC (5 µM) reversed malathion-induced oxidative stress responses, and prevented malathion-evoked apoptosis by regulating apoptotic protein expressions. Together, in GHA cells, NAC mediated inhibition of malathion-activated mitochondrial apoptotic pathways that involved cell cycle arrest and ROS responses. These data provide further insights into the molecular mechanisms behind malathion poisoning, and might suggest that NAC with its protective effects may be a potential compound for prevention of malathion-induced brain injury.
Assuntos
Acetilcisteína/farmacologia , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Astrócitos/efeitos dos fármacos , Malation/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Apoptose/fisiologia , Astrócitos/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Citoproteção/efeitos dos fármacos , Citoproteção/fisiologia , Relação Dose-Resposta a Droga , Humanos , Inseticidas/toxicidade , Compostos Organofosforados/toxicidade , Estresse Oxidativo/fisiologiaRESUMO
Timolol is a medication used widely to treat glaucoma. Regarding Ca2+ signaling, timolol was shown to modulate Ca2+-related physiology in various cell types, however, the effect of timolol on Ca2+ homeostasis and cell viability has not been explored in human prostate cancer cells. The aim of this study was to explore the effect of timolol on intracellular Ca2+ concentrations ([Ca2+]i) and viability in PC3 human prostate cancer cells. Timolol at concentrations of 100-1000 µM induced [Ca2+]i rises. The Ca2+ signal in Ca2+-containing medium was reduced by removal of extracellular Ca2+ by approximately 75%. Timolol (1000 µM) induced Mn2+ influx suggesting of Ca2+ entry. Timolol-induced Ca2+ entry was partially inhibited by three inhibitors of store-operated Ca2+ channels: nifedipine, econoazole and SKF96365, and by a protein kinase C (PKC) activator (phorbol 12-myristate 13 acetate [PMA]) or an inhibitor (GF109203X). In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin abolished timolol-evoked [Ca2+]i rises. Conversely, treatment with timolol abolished thapsigargin-evoked [Ca2+]i rises. Inhibition of phospholipase C (PLC) with U73122 abolished timolol-induced [Ca2+]i rises. Timolol at concentrations between 200 and 600 µM killed cells in a concentration-dependent fashion. Chelation of cytosolic Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/AM (BAPTA/AM) did not reverse cytotoxicity of timolol. Together, in PC3 cells, timolol induced [Ca2+]i rises by evoking Ca2+release from the endoplasmic reticulum in a PLC-dependent manner, and Ca2+ influx via PKC-regulated store-operated Ca2+ entry. Timolol also caused cell death that was not linked to preceding [Ca2+]i rises.
Assuntos
Agonistas dos Canais de Cálcio/toxicidade , Canais de Cálcio/efeitos dos fármacos , Sinalização do Cálcio/efeitos dos fármacos , Cálcio/metabolismo , Próstata/efeitos dos fármacos , Timolol/toxicidade , Canais de Cálcio/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Humanos , Cinética , Masculino , Células PC-3 , Próstata/metabolismo , Próstata/patologia , Proteína Quinase C/metabolismo , Fosfolipases Tipo C/metabolismoRESUMO
Niflumic acid, a drug used for joint and muscular pain, affected Ca²âº signaling in different models. However, the effect of niflumic acid on Ca²âº homeostasis and Ca²âº-related physiology in human osteosarcoma cells is unknown. This study examined the effect of niflumic acid on cytosolic free Ca²âº concentrations ([Ca²âº]i) in MG63 human osteosarcoma cells. Intracellular Ca²âº concentrations in suspended cells were monitored by using the fluorescent Ca²âº-sensitive dye fura- 2. Cell viability was examined by using 4-[3-[4-lodophenyl]-2-4(4-nitrophenyl)-2H-5-tetrazolio- 1,3-benzene disulfonate] water soluble tetrazolium-1 (WST-1). In MG63 cells, niflumic acid at concentrations of 250-750 µM evoked [Ca²âº]i rises concentration-dependently. Niflumic acid-evoked Ca²âº entry was confirmed by Mn²âº-induced quenching of fura-2 fluorescence. This entry was inhibited by nifedipine, econazole, SKF96365, the protein kinase C (PKC) activator phorbol 12-myristate 13 acetate (PMA), but was not affected by the PKC inhibitor GF109203X. In Ca²âº- free medium, treatment with the endoplasmic reticulum Ca²âº pump inhibitor thapsigargin (TG) inhibited niflumic acid-evoked [Ca²âº]i rises. Conversely, treatment with niflumic acid abolished TG-evoked [Ca²âº]i rises. Inhibition of phospholipase C (PLC) with U73122 also partly reduced niflumic acid-evoked [Ca²âº]i rises. Niflumic acid killed cells at 200-500 µM in a concentration-dependent fashion. Chelating cytosolic Ca²âº with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/ AM (BAPTA/AM) did not reverse niflumic acid-induced cytotoxicity. Collectively, our data suggest that in MG63 cells, niflumic acid induced [Ca²âº]i rises by evoking PLC-dependent Ca²âº release from the endoplasmic reticulum, and Ca²âº entry via PKC-sensitive store-operated Ca²âº entry. Niflumic acid also induced Ca²âº-independent cell death.
Assuntos
Neoplasias Ósseas , Osteossarcoma , Apoptose , Cálcio , Sinalização do Cálcio , Linhagem Celular Tumoral , Sobrevivência Celular , Humanos , Ácido Niflúmico , Fosfolipases Tipo CRESUMO
Exposure to insecticides has been found to have deleterious effects on human health. Lambda-cyhalothrin (LCT), a mixture of isomers of cyhalothrin, is a pyrethroid insecticide routinely used in pest control programs. LCT was reported to cause neurotoxic effects in various models. However, the mechanism of underlying effect of LCT on cytotoxicity in normal human brain cells is still elusive. This study examined whether LCT affected Ca2+ homeostasis and Ca2+-related physiology in Gibco® Human Astrocytes (GHA cells), and explored whether BAPTA-AM (1,2-bis(2-aminophenoxy)ethane-N,N,N'N'-tetraacetic acid), a selective Ca2+ chelator, has protective effects on LCT-treated GHA cells. The data show that LCT (10-15 µM) concentration-dependently induced cytotoxicity in both GHA cells and DI TNC1 normal rat astrocytes but only induced intracellular Ca2+ concentration ([Ca2+]i) rises in GHA cells. In terms of Ca2+ signaling in GHA cells, LCT-induced [Ca2+]i rises were reduced by removing extracellular Ca2+ and were inhibited by store-operated Ca2+ channel modulators (2-APB, econazole or SKF96365). In Ca2+-free medium, pretreatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin abolished LCT-induced [Ca2+]i rises. Conversely, incubation with LCT abolished thapsigargin-induced [Ca2+]i rises. Regarding cytotoxicity, LCT evoked apoptosis by regulating apoptotic protein expressions (Bax, BCl-2, cleaved caspase-9/-3). This apoptotic response was significantly inhibited by prechelating cytosolic Ca2+ with BAPTA-AM. Together, in GHA cells, LCT induced [Ca2+]i rises by inducing Ca2+ entry via store-operated Ca2+ channels and Ca2+ release from the endoplasmic reticulum. Moreover, BAPTA-AM has a protective effect on inhibiting LCT-activated mitochondrial apoptotic pathway. This study provided new insights into the molecular protective mechanism of LCT-induced cytotoxicity in normal human astrocytes.
Assuntos
Astrócitos/efeitos dos fármacos , Sinalização do Cálcio/efeitos dos fármacos , Ácido Egtázico/análogos & derivados , Inseticidas/toxicidade , Mitocôndrias/efeitos dos fármacos , Nitrilas/toxicidade , Piretrinas/toxicidade , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Astrócitos/metabolismo , Cálcio/metabolismo , Quelantes de Cálcio/farmacologia , Sinalização do Cálcio/fisiologia , Células Cultivadas , Relação Dose-Resposta a Droga , Ácido Egtázico/farmacologia , Humanos , Mitocôndrias/metabolismo , RatosRESUMO
Captopril, an angiotensin-converting enzyme (ACE) inhibitor, induced different Ca²âº signaling responses in various cell models. However, the effect of captopril on Ca²âº homeostasis and cell viability in hepatoma cells is unknown. This study examined whether captopril altered Ca²âº homeostasis and viability in HepG2 human hepatoma cells. Intracellular Ca²âº concentrations in suspended cells were monitored by using the fluorescent Ca²âº-sensitive dye fura-2. Cell viability was examined by using 4-[3-[4-lodophenyl]-2-4(4-nitrophenyl)-2H-5-tetrazolio-1,3-benzene disulfonate] water soluble tetrazolium-1 (WST-1). Captopril at concentrations of 500-3000 µM induced [Ca²âº]i rises in a concentration-dependent manner. Ca²âº removal reduced the signal by approximately 15%. Mn²âº has been shown to enter cells through similar mechanisms as Ca²âº but quenches fura-2 fluorescence at all excitation wavelengths. Captopril (3000 µM)-induced Mn²âº influx indirectly suggested that captopril evoked Ca²âº entry. Captopril-induced Ca²âº entry was inhibited by 15% by a protein kinase C (PKC) activator (phorbol 12-myristate 13 acetate, PMA) and an inhibitor (GF109203X) and three inhibitors of store-operated Ca²âº channels: nifedipine, econazole and SKF96365. In Ca²âº-free medium, treatment with the endoplasmic reticulum Ca²âº pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) abolished captopril-evoked [Ca²âº]i rises. Conversely, treatment with captopril abolished BHQ-evoked [Ca²âº]i rises. Inhibition of phospholipase C (PLC) with U73122 inhibited 70% of captopril-induced [Ca²âº]i rises. Captopril at concentrations between 150-550 µM killed cells in a concentration-dependent fashion. Chelation of cytosolic Ca²âº with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/AM (BAPTA/AM) did not reverse captopril's cytotoxicity. Together, in HepG2 human hepatoma cells, captopril induced [Ca²âº]i rises and caused cell death that was not triggered by preceding [Ca²âº]i rises.
Assuntos
Carcinoma Hepatocelular , Homeostase , Neoplasias Hepáticas , Apoptose , Cálcio , Sinalização do Cálcio , Captopril , Linhagem Celular Tumoral , Sobrevivência Celular , Humanos , Fosfolipases Tipo CRESUMO
OBJECTIVE: Magnolol, a polyphenol compound from herbal medicines, was shown to alter physiology in various cell models. However, the effect of magnolol on Ca2+ homeostasis and its related physiology in oral cancer cells is unclear. This study examined whether magnolol altered Ca2+ signaling and cell viability in OC2 human oral cancer cells. METHODS: Cytosolic Ca2+ concentrations ([Ca2+]i) in suspended cells were measured by using the fluorescent Ca2+-sensitive dye fura-2. Cell viability was examined by 4-[3-[4-lodophenyl]-2-4(4-nitrophenyl)-2H-5-tetrazolio-1,3-benzene disulfonate] water soluble tetrazolium-1 (WST-1) assay. RESULTS: Magnolol at concentrations of 20-100⯵M induced [Ca2+]i rises. Ca2+ removal reduced the signal by approximately 50%. Magnolol (100⯵M) induced Mn2+ influx suggesting of Ca2+ entry. Magnolol-induced Ca2+ entry was partially suppressed by protein kinase C (PKC) regulators, and inhibitors of store-operated Ca2+ channels. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) abolished magnolol-evoked [Ca2+]i rises. Conversely, treatment with magnolol abolished BHQ-evoked [Ca2+]i rises. Inhibition of phospholipase C (PLC) with U73122 partially inhibited magnolol-induced [Ca2+]i rises. Magnolol at 20-100⯵M decreased cell viability, which was not reversed by pretreatment with the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA/AM). CONCLUSIONS: Together, in OC2 cells, magnolol induced [Ca2+]i rises by evoking partially PLC-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via PKC-sensitive store-operated Ca2+ entry. Magnolol also caused Ca2+-independent cell death. Therefore, magnolol-induced cytotoxicity may not be involved in activation mechanisms associated with intracellular Ca2+ mobilization in oral cancer cells.
Assuntos
Compostos de Bifenilo/farmacologia , Cálcio/metabolismo , Homeostase/efeitos dos fármacos , Lignanas/farmacologia , Neoplasias Bucais/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Linhagem Celular Tumoral/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Citosol/metabolismo , Retículo Endoplasmático/metabolismo , Fura-2/farmacologia , Humanos , Manganês/metabolismo , Proteína Quinase C/efeitos dos fármacos , Proteína Quinase C/metabolismo , Sais de Tetrazólio , Fosfolipases Tipo C/efeitos dos fármacosRESUMO
Carvacrol, a monoterpenic phenol compound, has been shown to possess various biological effects in different models. However, the effect of carvacrol on intracellular Ca²âº and its related physiology in human prostate cancer is unknown. This study explored the effect of carvacrol on cytosolic free Ca²âº levels ([Ca²âº]i) and viability in PC3 human prostate cancer cells. Fura-2, a Ca²âº- sensitive fluorescent dye, was used to assess [Ca²âº]i. Cell viability was measured by the detecting reagent WST-1. Carvacrol at concentrations of 200-800 µM caused [Ca²âº]i rises in a concentration-dependent manner. Removal of extracellular Ca²âº reduced carvacrol's effect by approximately 60%. Carvacrol-induced Ca²âº entry was confirmed by Mn²âº entry-induced quench of fura-2 fluorescence, and was inhibited by approximately 30% by nifedipine, econazole, SKF96365, and the protein kinase C (PKC) inhibitor GF109203X. In Ca²âº-free medium, treatment with the endoplasmic reticulum Ca²âº pump inhibitor thapsigargin (TG) abolished carvacrol-induced [Ca²âº]i rises. Treatment with carvacrol also abolished TG-induced [Ca²âº]i rises. Carvacrol-induced Ca²âº release from the endoplasmic reticulum was abolished by inhibition of phospholipase C (PLC). Carvacrol killed cells at concentrations of 200-600 µM in a concentration-dependent fashion. Chelating cytosolic Ca²âº with BAPTA/AM did not prevent carvacrol's cytotoxicity. Together, in PC3 cells, carvacrol induced [Ca²âº]i rises by inducing PLC-dependent Ca²âº release from the endoplasmic reticulum and Ca²âº entry via PKC-sensitive store-operated Ca²âº channels and other unknown channels. Carvacrol also induced Ca²âº-dissociated cell death.
Assuntos
Cálcio/metabolismo , Monoterpenos/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cimenos , Humanos , Masculino , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Fosfolipases Tipo C/fisiologiaRESUMO
Propofol (2,6-diisopropylphenol), one of the extensively and commonly used anesthetic agents, has been shown to affect the biological behavior of various models. Previous researches have shown that propofol-induced cytotoxicity might cause anticancer effect in different cells. However, the mechanisms underlying the effect of propofol on cytotoxicity is still elusive in human glioblastoma cells. The aims of this study were to evaluate effects of propofol on cytotoxicity, cell cycle distribution and ROS production, and establish the relationship between oxidative stress and cytotoxicity in GBM 8401 human glioblastoma cells and DI TNC1 rat astrocytes. Propofol (20-30 µM) concentration-dependently induced cytotoxicity, cell cycle arrest, and increased ROS production in GBM 8401 cells but not in DI TNC1 cells. In GBM 8401 cells, propofol induced G2/M phase cell arrest, which affected the CDK1, cyclin B1, p53, and p21 protein expression levels. Furthermore, propofol induced oxygen stresses by increasing O2- and H2 O2 levels but treatment with the antioxidant N-acetylcysteine (NAC) partially reversed propofol-regulated antioxidative enzyme levels (superoxide dismutase, catalase, and glutathione peroxidase). Most significantly, propofol induced apoptotic effects by decreasing Bcl-2 but increasing Bax, cleaved caspase-9/caspase-3 levels, which were partially reversed by NAC. Moreover, the pancaspase inhibitor Z-VAD-FMK also partially prevented propofol-induced apoptosis. Together, in GBM 8401 cells but not in DI TNC1 cells, propofol activated ROS-associated apoptosis that involved cell cycle arrest and caspase activation. These findings indicate that propofol not only can be an anesthetic agent which reduces pain but also has the potential to be used for the treatment of human glioblastoma.
Assuntos
Antineoplásicos/farmacologia , Astrócitos/efeitos dos fármacos , Propofol/farmacologia , Acetilcisteína/farmacologia , Clorometilcetonas de Aminoácidos/farmacologia , Animais , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Astrócitos/citologia , Caspase 3/metabolismo , Caspase 9/metabolismo , Inibidores de Caspase/farmacologia , Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Glioblastoma , Humanos , Ratos , Espécies Reativas de Oxigênio/metabolismoRESUMO
Researches have been conducted to explore the biological effect of gastrodin, a natural compound extracted from the rhizome of Gastrodia elata Blume, in different models. However, the effects of gastrodin on cytotoxicity, cell cycle distribution and oxidative stress in glia cells have not been explored. The aim of this study was to investigate the cytotoxic effect of gastrodin and its mechanisms in DBTRG-05MG human glioblastoma cells and CTX TNA2 rat astrocytes. In DBTRG-05MG cells but not in CTX TNA2 cells, gastrodin (20-30 µM) induced cytotoxicity, G2/M phase cell cycle arrest and apoptosis. Regarding oxidative stress, gastrodin (20-30 µM) elevated intracellular ROS levels but reduced GSH levels. Treatment with the antioxidant NAC (10 µM) partially reversed gastrodin-altered antioxidant enzymes levels. Furthermore, gastrodin induced mitochondria-associated apoptosis. The apoptotic effects evoked by gastrodin were partially inhibited by the antioxidant NAC and the pancaspase inhibitor Z-VAD-FMK. Together, in DBTRG-05MG cells, but not in CTX TNA2 cells, gastrodin activated ROS-associated mitochondrial apoptotic pathways that involved cell cycle arrest. These data provide insight into the molecular mechanisms governing the ability of gastrodin to induce cytotoxicity in human glioblastoma cells and further suggest that gastrodin is a new potential agent for the treatment of human gliblasoma.