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1.
Chin J Physiol ; 64(4): 202-209, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34472451

RESUMO

Gamma-linolenic acid (GLA), a natural fatty acid obtained from oils of various vegetables and seeds, has been demonstrated as an anticancer agent. In this work, we investigated the anticancer effects of GLA on breast cancer BT-474 cells. GLA at 30 µM, a concentration reportedly within the range of circulating concentrations in clinical studies, caused apoptotic cell death. GLA caused an elevation in mitochondrial Ca2+ level and a decrease in mitochondrial membrane potential. GLA treatment depleted cyclopiazonic acid (CPA)-sensitive Ca2+ store and triggered substantial Ca2+ influx. Intracellular Ca2+ release triggered by GLA was suppressed by 3 µM xestospongin C (XeC, IP3 receptor-channel blocker) and 100 µM ryanodine (ryanodine receptor-channel blocker), suggesting that the Ca2+ release was via IP3 receptor-channel and ryanodine receptor-channel. Increased expressions of p-eIF2α and CHOP were observed in GLA-treated cells, suggesting GLA-treated cells had increased expressions of p-eIF2α and CHOP, which suggest endoplasmic reticulum (ER) stress. In addition, GLA elicited increased production of reactive oxygen species. Taken together, our results suggest a basal level of GLA induced apoptotic cell death by causing Ca2+ overload, mitochondrial dysfunction, Ca2+ store depletion, ER stress, and oxidative stress. This is the first report to show that GLA caused Ca2+ store depletion and ER stress. GLA-induced Ca2+ store depletion resulted from opening of IP3 receptor-channel and ryanodine receptor-channel.


Assuntos
Neoplasias da Mama , Ácido gama-Linolênico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Cálcio/metabolismo , Retículo Endoplasmático/metabolismo , Feminino , Humanos , Estresse Oxidativo , Ácido gama-Linolênico/metabolismo
2.
Eur J Pharmacol ; 904: 174115, 2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-33901459

RESUMO

In this report we examined the effects of lidocaine on Ca2+ homeostasis of neuronal cells using microfluorimetric measurement of cytosolic Ca2+ with fura 2 as probe. In mouse neuroblastoma N2A cells, 10 mM lidocaine caused Ca2+ release from the cyclopiazonic acid (CPA)-dischargeable pool and abolished ATP-triggered Ca2+ release. Lidocaine-triggered Ca2+ release was not affected by xestospongin C (XeC), an inositol 1,4,5-trisphosphate receptor (IP3R) inhibitor. N2A cells did not have functional ryanodine receptors (RYR) (absence of caffeine response) and we used differentiated NG108-15 cells (presence of caffeine response) for further experiments. Caffeine-triggered Ca2+ release was unaffected by a brief lidocaine exposure, but was eliminated after a prolonged treatment of lidocaine, suggesting lidocaine abolished caffeine action possibly not by interfering caffeine binding but via Ca2+ store depletion. Lidocaine-elicited Ca2+ release was unaffected by XeC or a high concentration of ryanodine, suggesting Ca2+ release was not via IP3R or RYR. Lidocaine did not affect nigericin-dischargeable lysosomal Ca2+ stores. Lastly, we observed that lidocaine suppressed CPA-induced store-operated Ca2+ influx in both N2A cells and differentiated NG108-15 cells. Our results suggest two novel actions of lidocaine in neuronal cells, namely, depletion of Ca2+ store (via an IP3R- and RYR-independent manner) and suppression of store-operated Ca2+ influx.

3.
BMC Complement Med Ther ; 20(1): 364, 2020 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-33228629

RESUMO

BACKGROUND: Arctium lappa L. root (burdock root) has long been recommended for the treatment of different diseases in traditional Chinese medicine. Burdock root possesses anti-oxidative, anti-inflammatory, anti-cancer, and anti-microbial activities. The aim of the study was to elucidate whether aqueous extract of burdock root regulates mesenchymal stem cell proliferation and differentiation. METHODS: Human bone marrow-derived mesenchymal stem cells in 2D high density culture and in 3D micromass pellets were treated with chondrogenic induction medium and chondral basal medium in the absence or presence of aqueous extract of burdock root. The chondrogenic differentiation was accessed by staining glucosaminoglycans, immunostaining SOX9 and type II collagen and immuonblotting of SOX9, aggrecan and type II collagen. RESULTS: Treatment of aqueous extract of burdock root increased the cell proliferation of hMSCs. It did not have significant effect on osteogenic and adipogenic differentiation, but significantly enhanced chondrogenic induction medium-induced chondrogenesis. The increment was dose dependent, as examined by staining glucosaminoglycans, SOX9, and type II collagen and immunobloting of SOX9, aggrecan and type II collagen in 2D and 3D cultures. In the presence of supplemental materials, burdock root aqueous extract showed equivalent chondrogenic induction capability to that of TGF-ß. CONCLUSIONS: The results demonstrate that aqueous extract of Arctium lappa L. root promotes chondrogenic medium-induced chondrogenic differentiation. The aqueous extract of burdock root can even be used alone to stimulate chondrogenic differentiation. The study suggests that the aqueous extract of burdock root can be used as an alternative strategy for treatment purposes.

4.
Z Naturforsch C J Biosci ; 75(3-4): 65-73, 2020 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-32092040

RESUMO

Intracellular polyamines such as spermine and spermidine are essential to cell growth in normal and especially in cancer cells. However, whether extracellular polyamines affect cancer cell survival is unknown. We therefore examined the actions of extracellular polyamines on breast cancer BT474 cells. Our data showed that spermine, spermidine, and putrescine decreased cell viability by apoptosis. These polyamines also elicited Ca2+ signals, but the latter were unlikely triggered via Ca2+-sensing receptor (CaSR) as BT474 cells have been demonstrated previously to lack CaSR expression. Spermine-elicited Ca2+ response composed of both Ca2+ release and Ca2+ influx. Spermine caused a complete discharge of the cyclopiazonic acid (CPA)-sensitive Ca2+ pool and, expectedly, endoplasmic reticulum (ER) stress. The Ca2+ influx pore opened by spermine was Mn2+-impermeable, distinct from the CPA-triggered store-operated Ca2+ channel, which was Mn2+-permeable. Spermine cytotoxic effects were not due to oxidative stress, as spermine did not trigger reactive oxygen species formation. Our results therefore suggest that spermine acted on a putative polyamine receptor in BT474 cells, causing cytotoxicity by Ca2+ overload, Ca2+ store depletion, and ER stress.


Assuntos
Neoplasias da Mama/metabolismo , Cálcio/metabolismo , Poliaminas/farmacologia , Receptores de Detecção de Cálcio/metabolismo , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica , Homeostase , Humanos , Putrescina/farmacologia , Espermidina/farmacologia , Espermina/farmacologia
5.
Fundam Clin Pharmacol ; 33(6): 604-611, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31206802

RESUMO

In a previous publication when we studied the purinergic receptor with which ATP interacted in mouse brain bEND.3 endothelial cells, we observed addition of 3 µm ARC 118925XX (ARC; selective P2Y2 antagonist) strongly suppressed ATP-triggered Ca2+ release, suggesting the response was mediated via P2Y2 receptors. We here report ARC unexpectedly promoted substantial Ca2+ influx even when ATP-triggered Ca2+ release was largely inhibited. Since this large Ca2+ influx may have important pharmacological significance, we proceeded to investigate its mechanism. ARC did not trigger intracellular Ca2+ release thus suggesting Ca2+ influx triggered by ARC was not store-operated. ARC-triggered Ca2+ influx could be blocked by 1 mm Ni2+ , a general Ca2+ channel blocker, but not by SK&F 96365, a nonselective TRP channel blocker. Unexpectedly, ARC promoted influx of Na+ and La3+ , but not Mn2+ . This is a surprising finding, since Mn2+ is conventionally used as a Ca2+ surrogate ion (as it permeates Ca2+ channel), and La3+ is classically used as a potent Ca2+ channel antagonist. Electrophysiological examination showed ARC did not stimulate any cation currents. Therefore, ARC opened, rather than a cation channel pore, an unidentified Ca2+ influx pathway which was Na+ - and La3+ -permeable but Mn2+ -impermeable.


Assuntos
Células Endoteliais/metabolismo , Furanos/farmacologia , Piperidinas/farmacologia , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Tetrazóis/farmacologia , Animais , Cálcio/metabolismo , Células Cultivadas , Manganês/metabolismo , Camundongos , Sódio/metabolismo
6.
Anticancer Res ; 39(1): 215-223, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30591461

RESUMO

BACKGROUND/AIM: Etomidate, an intravenous anesthetic, has been shown to have anticancer effects, including induction of cell-cycle arrest and apoptosis. However, to our knowledge, there are no reports about the anti-metastasis effects of etomidate on A549 human lung adenocarcinoma cells. MATERIALS AND METHODS: The cell viability, cell adhesion, gelatin zymography assay, transwell migration and invasion assay, and western blotting analysis were used to investigate the effects of etomidate on A549 cells. RESULTS: In our study, etomidate showed low cytotoxicity, inhibited cell adhesion, and suppressed the migration and invasion in A549 cells. The activity of matrix metallopeptidase 2 (MMP2) was reduced by 48 h treatment of etomidate. Results of western blotting analysis indicated that etomidate down-regulated the expression of protein kinase C, MMP7, MMP1, MMP9, and p-p-38, but up-regulated that of RAS, phosphoinositide 3-kinase, and phosphor-extracellular-signal related kinase after 24 and 48 h treatment, in A549 cells. CONCLUSION: Etomidate suppressed the migration and invasion of lung adenocarcinoma A549 cells via inhibiting the expression of MMP1, MMP2, MMP7 and MMP9, and provides potential therapeutic targets for lung cancer treatment.


Assuntos
Adenocarcinoma de Pulmão/tratamento farmacológico , Etomidato/farmacologia , Metaloendopeptidases/genética , Invasividade Neoplásica/genética , Células A549 , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/patologia , Apoptose/efeitos dos fármacos , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Proteínas de Neoplasias/genética
7.
J Cell Mol Med ; 22(5): 2896-2907, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29516686

RESUMO

Midazolam, a benzodiazepine derivative, is widely used for sedation and surgery. However, previous studies have demonstrated that Midazolam is associated with increased risks of congenital malformations, such as dwarfism, when used during early pregnancy. Recent studies have also demonstrated that Midazolam suppresses osteogenesis of mesenchymal stem cells (MSCs). Given that hypertrophic chondrocytes can differentiate into osteoblast and osteocytes and contribute to endochondral bone formation, the effect of Midazolam on chondrogenesis remains unclear. In this study, we applied a human MSC line, the KP cell, to serve as an in vitro model to study the effect of Midazolam on chondrogenesis. We first successfully established an in vitro chondrogenic model in a micromass culture or a 2D high-density culture performed with TGF-ß-driven chondrogenic induction medium. Treatment of the Midazolam dose-dependently inhibited chondrogenesis, examined using Alcian blue-stained glycosaminoglycans and the expression of chondrogenic markers, such as SOX9 and type II collagen. Inhibition of Midazolam by peripheral benzodiazepine receptor (PBR) antagonist PK11195 or small interfering RNA rescued the inhibitory effects of Midazolam on chondrogenesis. In addition, Midazolam suppressed transforming growth factor-ß-induced Smad3 phosphorylation, and this inhibitory effect could be rescued using PBR antagonist PK11195. This study provides a possible explanation for Midazolam-induced congenital malformations of the musculoskeletal system through PBR.


Assuntos
Condrogênese/efeitos dos fármacos , Antagonistas de Receptores de GABA-A/farmacologia , Células-Tronco Mesenquimais/metabolismo , Midazolam/farmacologia , Receptores de GABA-A/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Humanos , Isoquinolinas/farmacologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo
8.
J Physiol Sci ; 68(1): 33-41, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27873157

RESUMO

Eicosapentaenoic acid (EPA), an omega-3 fatty acid abundant in fish oil, protects endothelial cells (EC) from lipotoxicity and triggers EC NO release. The latter is related to an elevation of cytosolic Ca2+. Although EPA has been shown to cause human EC cytosolic Ca2+ elevation, the mechanism is unclear. Microfluorimetric imaging was used here to measure free cytosolic Ca2+ concentration. EPA was shown to cause intracellular Ca2+ release in mouse cerebral cortex endothelial bEND.3 cells; interestingly, the EPA-sensitive intracellular Ca2+ pool(s) appeared to encompass and was larger than the Ca2+ pool mobilized by sarcoplasmic-endoplasmic reticulum Ca2+-ATPase inhibition by cyclopiazonic acid. EPA also opened a Ca2+ influx pathway pharmacologically distinct from store-operated Ca2+ influx. Surprisingly, EPA-triggered Ca2+ influx was Ni2+-insensitive; and EPA did not trigger Mn2+ influx. Further, EPA-triggered Ca2+ influx did not involve Na+-Ca2+ exchangers. Thus, our results suggest EPA triggered unusual mechanisms of Ca2+ release and Ca2+ influx in EC.


Assuntos
Canais de Cálcio/metabolismo , Cálcio/metabolismo , Córtex Cerebral/efeitos dos fármacos , Ácido Eicosapentaenoico/farmacologia , Células Endoteliais/efeitos dos fármacos , Adenosina Trifosfatases/metabolismo , Animais , Linhagem Celular , Córtex Cerebral/metabolismo , Células Endoteliais/metabolismo , Indóis/farmacologia , Transporte de Íons/efeitos dos fármacos , Camundongos
9.
J Cell Biochem ; 118(5): 1108-1117, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-27608291

RESUMO

Cells switch to anaerobic glycolysis when there is a lack of oxygen during brain ischemia. Extracellular pH thus drops and such acidosis causes neuronal cell death. The fate of astrocytes, mechanical, and functional partners of neurons, in acidosis is less studied. In this report, we investigated the signaling in acidosis-challenged rat cortical astrocytes and whether these signals were related to mitochondrial dysfunction and cell death. Exposure to acidic pH (6.8, 6.0) caused Ca2+ release and influx, p38 MAPK activation, and Akt inhibition. Mitochondrial membrane potential was hyperpolarized after astrocytes were exposed to acidic pH as soon as 1 h and lasted for 24 h. Such mitochondrial hyperpolarization was prevented by SC79 (an Akt activator) but not by SB203580 (a p38 inhibitor) nor by cytosolic Ca2+ chelation by BAPTA, suggesting that only the perturbation in Akt signaling was causally related to mitochondrial hyperpolarization. SC79, SB203580, and BAPTA did not prevent acidic pH-induced cell death. Acidic pH suppressed ROS production, thus ruling out the role of ROS in cytotoxicity. Interestingly, pH 6.8 caused an increase in ADP/ATP ratio and apoptosis; pH 6.0 caused a further increase in ADP/ATP ratio and necrosis. Therefore, astrocyte cell death in acidosis did not result from mitochondrial potential collapse; in case of acidosis at pH 6.0, necrosis might partly result from mitochondrial hyperpolarization and subsequent suppressed ATP production. J. Cell. Biochem. 118: 1108-1117, 2017. © 2016 Wiley Periodicals, Inc.


Assuntos
Ácidos/toxicidade , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Astrócitos/citologia , Mitocôndrias/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Apoptose , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Cálcio/metabolismo , Sobrevivência Celular , Células Cultivadas , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Ratos , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos
10.
Oncol Rep ; 35(4): 2089-96, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26781422

RESUMO

Glioblastoma multiforme (GBM) is a highly malignant devastating brain tumor in adults. Benzyl isothiocyanate (BITC) is one of the isothiocyanates that have been shown to induce human cancer cell apoptosis and cell cycle arrest. Herein, the effect of BITC on cell viability and apoptotic cell death and the genetic levels of human brain glioblastoma GBM 8401 cells in vitro were investigated. We found that BITC induced cell morphological changes, decreased cell viability and the induction of cell apoptosis in GBM 8401 cells was time-dependent. cDNA microarray was used to examine the effects of BITC on GBM 8401 cells and we found that numerous genes associated with cell death and cell cycle regulation in GBM 8401 cells were altered after BITC treatment. The results show that expression of 317 genes was upregulated, and two genes were associated with DNA damage, the DNA-damage-inducible transcript 3 (DDIT3) was increased 3.66-fold and the growth arrest and DNA-damage-inducible α (GADD45A) was increased 2.34-fold. We also found that expression of 182 genes was downregulated and two genes were associated with receptor for cell responses to stimuli, the EGF containing fibulin-like extracellular matrix protein 1 (EFEMP1) was inhibited 2.01-fold and the TNF receptor-associated protein 1 (TRAP1) was inhibited 2.08-fold. BITC inhibited seven mitochondria ribosomal genes, the mitochondrial ribosomal protein; tumor protein D52 (MRPS28) was inhibited 2.06-fold, the mitochondria ribosomal protein S2 (MRPS2) decreased 2.07-fold, the mitochondria ribosomal protein L23 (MRPL23) decreased 2.08-fold, the mitochondria ribosomal protein S2 (MRPS2) decreased 2.07-fold, the mitochondria ribosomal protein S12 (MRPS12) decreased 2.08-fold, the mitochondria ribosomal protein L12 (MRPL12) decreased 2.25-fold and the mitochondria ribosomal protein S34 (MRPS34) was decreased 2.30-fold in GBM 8401 cells. These changes of gene expression can provide the effects of BITC on the genetic level and are potential biomarkers for glioblastoma therapy.


Assuntos
Neoplasias Encefálicas/genética , Ciclo Celular/efeitos dos fármacos , Perfilação da Expressão Gênica/métodos , Glioblastoma/genética , Isotiocianatos/farmacologia , Apoptose , Neoplasias Encefálicas/tratamento farmacológico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA , Proteínas da Matriz Extracelular/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioblastoma/tratamento farmacológico , Proteínas de Choque Térmico HSP90/genética , Humanos , Fator de Transcrição CHOP/genética
11.
Pharmacol Rep ; 67(6): 1049-54, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26481521

RESUMO

BACKGROUND: Hinokiol is a naturally occurring diterpenoid compound isolated from plants such as Taiwania cryptomerioides. Anti-oxidation, anti-cancer, and anti-inflammation effects of this compound have been reported. It is not yet known if hinokiol affects neurons or neuronal ion channel activities. We reported here that hinokiol inhibited voltage-gated Na(+) channels (VGSC) in neuronal cells and we characterized the mechanisms of block. METHODS: The effects of hinokiol on Na(+) channels were examined using the voltage-clamp (whole-cell mode) technique. RESULTS: VGSC was blocked by hinokiol in a concentration-dependent and state-dependent manner in neuroblastoma N2A cells: IC(50) are 11.3 and 37.4µM in holding potentials of -70 and -100 mV, respectively. In the presence of hinokiol there was a 13-mV left shift in steady-state inactivation curves; however, activation gating was not altered. VGSC inhibition by hinokiol did not require channel opening and was thus considered to be closed-channel block. In the presence of hinokiol, since the degree of block did not enhance with stimulation frequency, block by hinokiol thus did not exhibit use-dependence. Recovery from channel inactivation was not significantly affected in the presence of hinokiol. In addition, hinokiol also inhibited VGSC of differentiated neuronal NG108-15 cells and rat hippocampal CA1 neurons. CONCLUSION: Our results therefore suggest hinokiol inhibited VGSC in a closed-channel block manner and such inhibition involved intensification of channel inactivation.


Assuntos
Abietanos/farmacologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Bloqueadores do Canal de Sódio Disparado por Voltagem/farmacologia , Canais de Sódio Disparados por Voltagem/metabolismo , Animais , Região CA1 Hipocampal/efeitos dos fármacos , Região CA1 Hipocampal/fisiologia , Linhagem Celular , Relação Dose-Resposta a Droga , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Ratos
12.
J Physiol Sci ; 65(2): 171-7, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25617267

RESUMO

Astrocytes have multiple functions such as provision of nourishment and mechanical support to the nervous system, helping to clear extracellular metabolites of neurons and modulating synaptic transmission by releasing gliotransmitters. In excitable cells, voltage-gated K(+) (Kv) channels serve to repolarize during action potentials. Astrocytes are considered non-excitable cells since they are not able to generate action potentials. There is an abundant expression of various Kv channels in astrocytes but the functions of these Kv channels remain unclear. We examined whether these astrocyte Kv channels regulate astrocyte "excitability" in the form of cytosolic Ca(2+) signaling. Electrophysiological examination revealed that neonatal rat cortical astrocytes possessed both delayed rectifier type and A-type Kv channels. Pharmacological blockade of both delayed rectifier Kv channels by TEA and A-type Kv channels by quinidine significantly suppressed store-operated Ca(2+) influx; however, TEA alone or quinidine alone did not suffice to cause such suppression. TEA and quinidine together dramatically enhanced current injection-triggered membrane potential overshoot (depolarization); either drug alone caused much smaller enhancements. Taken together, the results suggest both delayed rectifier and A-type Kv channels regulate astrocyte Ca(2+) signaling via controlling membrane potential.


Assuntos
Astrócitos/metabolismo , Astrócitos/fisiologia , Cálcio/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Animais , Astrócitos/efeitos dos fármacos , Células Cultivadas , Eletrofisiologia/métodos , Ativação do Canal Iônico/efeitos dos fármacos , Ativação do Canal Iônico/fisiologia , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/fisiologia , Bloqueadores dos Canais de Potássio/farmacologia , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
13.
Environ Toxicol ; 30(12): 1416-22, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24910415

RESUMO

Various sedative agents, including dexmedetomidine (dex), induce immunosuppression, and enhance infection progression. However, there was no information on how anesthetic affects local and systemic cellular immune function. We conducted this study to examine the impact of dex on the differentiation and function of immune cells at site of inflammation and in peripheral blood during endotoxemia of mice. In BALB/c mice with and without endotoxemia, we evaluated the influence of two dosages of 5 and 50 mcg/kg/h intravenous dex on immune cells: including number of T cells (CD3), B cells (CD19), natural killer cells (CD8a), monocytes (CD11b), and macrophages (Mac-3) in peripheral blood, the activities of macrophages in peripheral blood and in peritoneal lavage, and proliferation of B and T cells and of natural killer cells activity in the spleen. Endotoxemia increased the number of CD3 T cells, CD 19 B cells and macrophages in the peripheral blood, augmented macrophage activity in the peritoneum, and increased T cell proliferation and natural killer cell activity in the spleen. Further administration of 5 mcg/kg/h dex attenuated systemic increase in number of T cells, B cells, and macrophages during endotoxemia and 50 mcg/kg/h dex significantly attenuated the increase in activity of macrophages in the peripheral blood during endotoxemia. In the peritoneum, however, 5 mcg/kg/h dex preserved and 50 mcg/kg/h dexmedetomidine enhanced the activity of macrophages during endotoxemia. Increased in proliferation of T cells in spleen during endotoxemia was attenuated by both doses of dex. Last, 50 mcg/kg/h dex enhanced natural killer cells activity during endotoxemia. While preserving the effects of endotoxemia on macrophage's activity in the infection site and natural killer cell's activity in the spleen, dex decreased systemic fulminant immune reaction in endotoxemia, by attenuating the augmented response in the number of T cells, B cells and macrophages, activity of macrophages in the peripheral blood, and proliferation of T cells in spleen during endotoxemia.


Assuntos
Dexmedetomidina/farmacologia , Hipnóticos e Sedativos/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Animais , Linfócitos B/citologia , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Proliferação de Células/efeitos dos fármacos , Endotoxemia/metabolismo , Endotoxemia/patologia , Escherichia coli/metabolismo , Células Matadoras Naturais/citologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Lipopolissacarídeos/toxicidade , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Monócitos/citologia , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Fagocitose/efeitos dos fármacos , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia
14.
Eur J Pharmacol ; 724: 152-60, 2014 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-24374009

RESUMO

Midazolam (MDL) was known to act through stimulation of benzodiazepine receptors (GABA). Whether midazolam affects ion currents and membrane potential in neurons remains largely unclear. Electrophysiological studies of midazolam actions were performed in differentiated motor neuron-like (NSC-34 and NG108-15) cells. Midazolam suppressed the amplitude of delayed rectifier K(+) current (IK(DR)) in a time- and concentration-dependent manner with an IC50 value of 10.4 µM. Addition of midazolam was noted to enhance the rate of IK(DR) inactivation. On the basis of minimal binding scheme, midazolam-induced block of IK(DR) was quantitatively provided with a dissociation constant of 9.8 µM. Recovery of IK(DR) from inactivation in the presence of midazolam was fitted by a single exponential. midazolam had no effect on M-type or erg-mediated K(+) current in these cells. Midazaolam (30 µM) suppressed the peak amplitude of voltage-gated Na(+) current (INa) with no change in the current-voltage relationships of this current. Inactivation kinetics of INa remained unaltered in the presence of this agent. In current-clamp configuration, midazolam (30 µM) prolonged the duration of action potentials (APs) and reduce AP amplitude. Similarly, in differentiated NG108-15 cells, the exposure to midazolam also suppressed IK(DR) with a concomitant increase in current inactivation. Midazolam can act as an open-channel blocker of delayed-rectifier K(+) channels in these cells. The synergistic blocking effects on IK(DR) and INa may contribute to the underlying mechanisms through which midazolam affects neuronal function in vivo.


Assuntos
Canais de Potássio de Retificação Tardia/antagonistas & inibidores , Midazolam/farmacologia , Neurônios Motores/efeitos dos fármacos , Bloqueadores dos Canais de Potássio/farmacologia , Animais , Linhagem Celular , Canais de Potássio de Retificação Tardia/fisiologia , Potenciais da Membrana/efeitos dos fármacos , Metadona/farmacologia , Camundongos , Neurônios Motores/fisiologia
15.
Eur J Pharmacol ; 721(1-3): 49-55, 2013 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-24113522

RESUMO

Imperatorin is a naturally occurring furocoumarin compound isolated from plants such as Angelica archangelica and Cnidium monnieri. It has multiple pharmacological effects including anticonvulsant effects. Here we determined the effects of imperatorin on voltage-gated Na(+) channels (VGSC) using whole-cell patch clamp techniques in differentiated neuronal NG108-15 cells. We showed that imperatorin inhibited VGSC; such inhibition did not show state-dependence. Imperatorin caused a left shift in the steady-state inactivation curve without affecting activation gating. The inhibition of VGSC by imperatorin displayed a mild frequency-dependence. Imperatorin was also shown to inhibit VGSC and action potential amplitude without affecting voltage-gated K(+) channels in rat hippocampal CA1 neurons. In conclusion, our results suggest that imperatorin dampens neuronal excitability by inhibiting VGSC.


Assuntos
Furocumarinas/farmacologia , Neurônios/citologia , Neurônios/efeitos dos fármacos , Bloqueadores do Canal de Sódio Disparado por Voltagem/farmacologia , Canais de Sódio Disparados por Voltagem/metabolismo , Potenciais de Ação/efeitos dos fármacos , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Relação Dose-Resposta a Droga , Feminino , Masculino , Ratos , Ratos Sprague-Dawley
16.
Oncol Rep ; 30(5): 2304-10, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24008596

RESUMO

Propofol is one of the most widely clinically used intravenous anesthetic, and it induces apoptosis in human and murine leukemia cell lines. Yet, whether propofol causes DNA damage and affects the mRNA expression of repair-associated genes in cancer cells remains undetermined. In the present study, we investigated the effects of propofol on DNA damage and associated mRNA gene expression in RAW264.7 cells. Comet assay and DNA gel electrophoresis were used to evaluate DNA damage in RAW264.7 cells and propofol-inhibited cell growth in vitro. The results revealed a longer DNA tail and DNA fragmentation. Real-time PCR assay was used to examine mRNA gene expression of DNA damage and DNA repair-associated genes. Following exposure to propofol for 48 h, a decrease in the mRNA expression of DNA-PK, BRCA1, MGMT and p53 was noted in the RAW264.7 cells. Results from the western blotting indicated that p53, MGMT, 14-3-3-σ, BRCA1 and MDC1 proteins were decreased while p-p53 and p-H2A.X(S140) were increased in the RAW264.7 cells following exposure to propofol. In conclusion, exposure to propofol caused DNA damage and inhibited mRNA expression and protein levels of repair-associated genes in RAW264.7 cells.


Assuntos
Reparo do DNA/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Leucemia/patologia , Propofol/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ensaio Cometa , Dano ao DNA/efeitos dos fármacos , Humanos , Leucemia/genética , Macrófagos/efeitos dos fármacos , Camundongos , Monócitos/efeitos dos fármacos , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/genética
17.
Cell Physiol Biochem ; 32(2): 402-16, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23988522

RESUMO

BACKGROUND/AIMS: The objective of this study is to examine the current signals in response to large hyperpolarizations with the aid of principal component analysis (PCA) to search for or even predict current fluctuations related to membrane electroporation-induced current (I(MEP)). METHODS: The characteristics of principal eigenvalues generated for I(MEP) and the current signals at 10 sec prior to the start of initial I(MEP) (I(Pre)) were examined. As membrane hyperpolarizations were applied at 0.1 Hz, the appearance of I(MEP) coincided with the higher principal eigenvalues extracted in PCA. RESULTS: Subsequent addition of LaCl3 (100 µM) greatly reduced I(MEP) and associated principal eigenvalues. In real-time analysis for a single frame (i.e, 300 msec), in response to large hyperpolarization, multiple runs of heralded minuscule inward currents (Imin) occurring before large rise in current amplitudes were detected. With PCA, such heralded Imin was noted to coincide with the extreme principal eigenvalues. The duration of Imin together with large principal eigenvalues was influenced by different levels of membrane hyperpolarization. In GH3 cells, palmitoyl-L-carnitine (PALCAR), a long-chain acylcarnitine, effectively increased the I(MEP) amplitude with an EC50 value of 2.4 µM. However, in PALCAR-treated cells, the Imin together with higher principal eigenvalues disappeared, while in isoflurane-treated cells, Imin occurring before large rise of current amplitude remained intact. Similarly, the PCA analysis from I(Pre) in RAW 264.6 macrophages showed the presence of herald Imin accompanied by the extreme principal eigenvalues. CONCLUSION: It is clear from this study that these large principal eigenvalues are representative of MEP-associated formation of electropores. Therefore, different compositions around the surface membrane of cells may alter the appearance of Imin followed by I(MEP) emergence.


Assuntos
Membrana Celular/fisiologia , Fenômenos Eletrofisiológicos , Eletroporação , Macrófagos/fisiologia , Linhagem Celular Tumoral , Células Cultivadas , Humanos , Análise de Componente Principal
18.
Eur J Pharmacol ; 704(1-3): 41-8, 2013 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-23420002

RESUMO

During prolonged depolarization, voltage-gated K(+) (Kv) channels display C-type inactivation, a process which is due to selectivity filter destabilization and serves to limit K(+) flux. Here we reported that coumarsabin, a coumarin derivative isolated from Juniperus Sabina, could hasten C-type inactivation and thus cause block of Kv channels in neuronal NG108-15 cells and Kv1.2 channels heterologously expressed in lung epithelial H1355 cells. In NG108-15 cells, extracellular, but not intracellular, coumarsabin (30 µM) strongly speeded up Kv current decay and caused a left-shift in the steady-state inactivation curve. Coumarsabin inhibited end-of-pulse Kv currents with an IC50 of 13.4 µM. The kinetics and voltage-dependence of activation were not affected by coumarsabin. The degree of block by coumarsabin was not enhanced by a reduction in intracellular K(+) concentration. Data reveal that coumarsabin was a closed channel blocker and it displayed a frequency-independent mode of inhibition. Coumarsabin did not accelerate current decay in a Kv1.2 mutant (V370G) defective in C-type inactivation. Taken together, our data suggest that Kv channel inhibition by coumarsabin did not appear to result from a direct obstruction of the outer pore but relied on C-type inactivation.


Assuntos
Cumarínicos/farmacologia , Ativação do Canal Iônico/efeitos dos fármacos , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana/fisiologia , Potenciais de Ação/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Região CA1 Hipocampal/citologia , Linhagem Celular Tumoral , Juniperus , Camundongos , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Folhas de Planta , Ratos , Ratos Sprague-Dawley
19.
Sci Technol Adv Mater ; 14(5): 054401, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27877605

RESUMO

Stem cells are known for their potential to repair damaged tissues. The adhesion, growth and differentiation of stem cells are likely controlled by the surrounding microenvironment which contains both chemical and physical cues. Physical cues in the microenvironment, for example, nanotopography, were shown to play important roles in stem cell fate decisions. Thus, controlling stem cell behavior by nanoscale topography has become an important issue in stem cell biology. Nanotechnology has emerged as a new exciting field and research from this field has greatly advanced. Nanotechnology allows the manipulation of sophisticated surfaces/scaffolds which can mimic the cellular environment for regulating cellular behaviors. Thus, we summarize recent studies on nanotechnology with applications to stem cell biology, including the regulation of stem cell adhesion, growth, differentiation, tracking and imaging. Understanding the interactions of nanomaterials with stem cells may provide the knowledge to apply to cell-scaffold combinations in tissue engineering and regenerative medicine.

20.
Anticancer Res ; 32(11): 4833-42, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23155249

RESUMO

Metastasis is a major cause of death of patients with malignant tumors. Matrix metalloproteinases (MMPs) are important for the migration and invasion of various types of cancer cell. Propofol is a known anesthetic agent, widely used for short-term anesthesia and for longer-term sedation. Propofol inhibits the proliferation of a variety of tumor cells, but there is no available information regarding propofol-inhibited migration and invasion of tumor cells in vitro. In this study, we investigated the effects of propofol on the migration and invasion of human lung carcinoma A549 cells. Wound healing assay and Boyden chamber assays indicated that propofol inhibited the migration and invasion of A549 cells in vitro. Gelatin zymographic analysis showed the inhibitory effect of propofol on the activation of expression MMP-2. Western blot analysis also indicated that propofol suppressed the protein expiration of growth factor receptor-bound protein 2 (GRB2), Jun N-terminal kinases 1/2 (p-JNK1/2), p-p38, MMP-2 and MMP-9 in A549 cells. Results from real-time PCR assay also showed that propofol inhibited the mRNA gene expression of MMP-2, -7 and -9, and enhanced that of tissue inhibitor of metalloproteinase 1 (TIMP1) and TIMP2 in A549 cells. Taken together, these data show that propofol inhibits MMP-2 and -9 mRNA and protein expressions, resulting in suppression of lung cancer cell invasion and migration in vitro.


Assuntos
Adenocarcinoma/metabolismo , Antineoplásicos/farmacologia , Movimento Celular/efeitos dos fármacos , Neoplasias Pulmonares/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Propofol/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Adenocarcinoma de Pulmão , Western Blotting , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Regulação para Baixo , Ativação Enzimática/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Humanos , Invasividade Neoplásica , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/efeitos dos fármacos
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