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1.
Peroxynitrite, a cytotoxic oxidant formed in the reaction of superoxide and nitric oxide is known to cause programmed cell death. However, the mechanisms of peroxynitrite-induced apoptosis are poorly defined. The present study was designed to characterize the molecular mechanisms by which peroxynitrite induces apoptosis in HL-60 cells, with special emphasis on the role of caspases. Peroxynitrite induced the activation of apopain/caspase-3, but not ICE/caspase-1 as measured by the cleavage of fluorogenic peptides. Considering the short half-life of peroxynitrite and the kinetics of caspase-3 activation (starting 3–4 h after peroxynitrite treatment), the enzyme is not likely to become activated directly by the oxidant. Caspase-3 activation proved to be essential for DNA fragmentation, because pretreatment of the cells with the specific tetrapeptide inhibitor DEVD-fmk completely blocked peroxynitrite-induced DNA fragmentation. Peroxynitrite-induced cytotoxicity was also significantly altered by the inhibition of caspase-3, whereas phosphatidylserine exposure was unaffected by DEVD-fmk treatment. Because many of the effects of peroxynitrite are mediated by poly(ADP-ribose) synthetase (PARS) activation, we have also investigated the effect of PARS-inhibition on peroxynitrite-induced apoptosis. We have found that PARS-inhibition modulates peroxynitrite-induced apoptotic DNA fragmentation in the HL-60 cells. The effect of the PARS inhibitors, 3-aminobenzamide and 5-iodo-6-amino-1,2-benzopyrone were dependent on the concentration of peroxynitrite used. While PARS-inhibition resulted in increased DNA-fragmentation at low doses (15 μM) of peroxynitrite, a decreased DNA-fragmentation was found at high doses (60 μM) of peroxynitrite. PARS inhibition negatively affected viability as determined by flow cytometry. These data demonstrate the crucial role of caspase-3 in mediating apoptotic DNA fragmentation in HL-60 cells exposed to peroxynitrite. 相似文献
2.
Iguchi K Usui S Ishida R Hirano K 《Apoptosis : an international journal on programmed cell death》2002,7(6):519-525
Intracellular acidification is known to be involved in the initiation phase of apoptosis. However, the necessity of intracellular acidic conditions in the execution phase of apoptosis remains unknown. In this study, we found that in HL-60 cells imidazole induces cell death, associated with intracellular acidification, caspase-3 activation and DFF-45 cleavage, but not oligonucleosomal DNA fragmentation. A caspase inhibitor prevented cell death but not intracellular acidification. When pHi was neutralized by changing from imidazole-containing medium to fresh medium, oligonucleosomal DNA fragmentation and increased caspase-3 activity was observed in the imidazole-treated HL-60 cells. Furthermore, the DNA fragmentation induced by intracellular neutralization was inhibited by caspase inhibitor treatment. These results indicate that imidazole induces caspase-dependent cell death, and suggest that maintaining pHi in the neutral range is essential for the induction of oligonucleosomal DNA fragmentation in the execution phase of apoptosis. 相似文献
3.
Munehisa Yabuki Ken Tsutsui Alan A. Horton Tamotsu Yoshioka Kozo Utsumi 《Free radical research》2013,47(6):507-514
Nitric oxide (NO) from (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (NOC-18) induces apoptosis in human leukemia HL-60 cells. This effect was prevented by the pan-caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (Z-VAD-FMK), thereby implicating caspase activity in the process. NOC-18 treatment resulted in the activation of several caspases including caspase-3, -6, -8, and -9(-like) activities and the degradation of several caspase substrates such as nuclear lamins and SP120 (hnRNP-U/SAF-A). Moreover, release of cytochrome c from mitochondria was also observed during NOC-18-induced apoptosis. This change was substantially prevented by Z-VAD-FMK, thereby suggesting that the released cytochrome c might function not only as an initiator but also as an amplifier of the caspase cascade. Bid, a death agonist member of the Bcl-2 family, was processed by caspases following exposure of cells to NOC-18, supporting the above notion. Thus, NO-mediated apoptosis in HL-60 cells involves a caspase/cytochrome c-dependent mechanism. 相似文献
4.
RGD peptide-induced apoptosis in human leukemia HL-60 cells requires caspase-3 activation 总被引:7,自引:0,他引:7
RGD motif-containing peptides have been used in various studies of cell adhesion and growth. We report that RGD triggered
apoptosis at a concentration of 1 mmol/L, whereas RAD-containing peptides failed to induce apoptosis in HL-60 cells. RGD-treated
cells revealed internucleosomal DNA fragmentation. Western blot reveals caspase-3 activation in RGD peptide-treated cells.
A caspase-3 inhibitor z-VAD-FMK completely blocked the apoptosis, but a caspase-1 inhibitor (Ac-YVAD-CMK) and caspase-2 inhibitor
(z-VDVAD-FMK) did not block the apoptosis, suggesting that caspase-3 might have a critical role in the execution process of
apoptosis induced by RGD. RGD peptides have been used extensively to inhibit tumor metastasis. Our results should help in
further understanding the RGD peptide-induced apoptosis, which is important since RGD peptides have a potential role in therapies
of the future.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
5.
Monocytic cell necrosis is mediated by potassium depletion and caspase-like proteases 总被引:2,自引:0,他引:2
Apoptosis is aphysiological cell death that culminates in mitochondrial permeabilitytransition and the activation of caspases, a family of cysteineproteases. Necrosis, in contrast, is a pathological cell deathcharacterized by swelling of the cytoplasm and mitochondria and rapidplasma membrane disruption. Necrotic cell death has long been opposedto apoptosis, but it now appears that both pathways involvemitochondrial permeability transition, raising the question of whatmediates necrotic cell death. In this study, we investigated mechanismsthat promote necrosis induced by various stimuli(Clostridium difficile toxins,Staphylococcus aureus alpha toxin,ouabain, nigericin) in THP-1 cells, a human monocytic cell line, and in monocytes. All stimuli induced typical features of necrosis and triggered protease-mediated release of interleukin-1 (IL-1) andCD14 in both cell types. K+depletion was actively implicated in necrosis because substituting K+ forNa+ in the extracellular mediumprevented morphological features of necrosis and IL-1 release.N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone, a broad-spectrum caspase inhibitor, prevented morphological features of necrosis, plasma membrane destruction, loss ofmitochondrial membrane potential, IL-1 release, and CD14 sheddinginduced by all stimuli. Thus, in monocytic cells, necrosis is a celldeath pathway mediated by passiveK+ efflux and activation ofcaspase-like proteases. 相似文献
6.
Brevnova EE Platoshyn O Zhang S Yuan JX 《American journal of physiology. Cell physiology》2004,287(3):C715-C722
Apoptotic cell shrinkage, an early hallmark of apoptosis, is regulated by K+ efflux and K+ channel activity. Inhibited apoptosis and downregulated K+ channels in pulmonary artery smooth muscle cells (PASMC) have been implicated in development of pulmonary vascular medial hypertrophy and pulmonary hypertension. The objective of this study was to test the hypothesis that overexpression of KCNA5, which encodes a delayed-rectifier voltage-gated K+ (Kv) channel, increases K+ currents and enhances apoptosis. Transient transfection of KCNA5 caused 25- to 34-fold increase in KCNA5 channel protein level and 24- to 29-fold increase in Kv channel current (IK(V)) at +60 mV in COS-7 and rat PASMC, respectively. In KCNA5-transfected COS-7 cells, staurosporine (ST)-mediated increases in caspase-3 activity and the percentage of cells undergoing apoptosis were both enhanced, whereas basal apoptosis (without ST stimulation) was unchanged compared with cells transfected with an empty vector. In rat PASMC, however, transfection of KCNA5 alone caused marked increase in basal apoptosis, in addition to enhancing ST-mediated apoptosis. Furthermore, ST-induced apoptotic cell shrinkage was significantly accelerated in COS-7 cells and rat PASMC transfected with KCNA5, and blockade of KCNA5 channels with 4-aminopyridine (4-AP) reduced K+ currents through KCNA5 channels and inhibited ST-induced apoptosis in KCNA5-transfected COS-7 cells. Overexpression of the human KCNA5 gene increases K+ currents (i.e., K+ efflux or loss), accelerates apoptotic volume decrease (AVD), increases caspase-3 activity, and induces apoptosis. Induction of apoptosis in PASMC by KCNA5 gene transfer may serve as an important strategy for preventing the progression of pulmonary vascular wall thickening and for treating patients with idiopathic pulmonary arterial hypertension (IPAH). potassium ion channel; pulmonary hypertension 相似文献
7.
Apoptosis manifests in two major execution programs downstream of the death signal:the caspase pathway and organelle dysfunction.An important antiapoptosis factor,Bcl-2 protein,contributes in caspase pathway of apoptosis.Calcium,an important intracellular signal element in cells,is also observed to have changes during apoptosis,which maybe affected by Bcl-2 protein.We have previously reported that in Harringtonine (HT) induced apoptosis of HL-60 cells,there‘s change of intracellular calcium distribution,oving from cytoplast especially Golgi‘s apparatus to nucleus and accumulating there with the highest concentration.We report here that caspase-3 becomes activated in HT-induced apoptosis of HL-60 cells,which can be inhibited by overexpression of Bcl-2 protein.No sign of apoptosis or intracellular calcium movement from Golgi‘s apparatus to nucleus in HL-60 cells overexpressing Bcl-2 or treated with Ac-DEVD-CHO,a specific inhibitor of caspase-3.The results indicate that activated caspase-2 can promote the movement of intracellular calcium from Golgi‘s apparatus to nucleus,and the process is inhibited by Ac-DEVD-CHO(inhibitor of caspase-3),and that Bcl-2 can inhibit the movement and accumulation of intracellular calcium in nucleus through its inhibition on caspase-3.Calcium relocalization in apoptosis seems to be irreversible,which is different from the intracellular calcium changes caused by growth factor. 相似文献
8.
Byeong Mo Kim Yun Jung Choi Yong Heon Lee Young Ae Joe Sung Hee Hong 《Apoptosis : an international journal on programmed cell death》2010,15(8):982-993
Doxorubicin (Dox) is widely used to treat a variety of tumors. However, resistance to this drug is common, making successful
treatment more difficult. Previously, we introduced a novel phytosphingosine derivative, N,N-dimethyl phytosphingosine (DMPS), as a potent anticancer therapeutic agent in human leukemia cells. This study was performed
to investigate whether DMPS can sensitize HL-60/MX2, a multidrug-resistant variant of HL-60, to Dox-induced apoptosis. Low
concentrations of DMPS sensitized HL-60/MX2 cells to Dox-induced apoptosis. Combined Dox + DMPS treatment-induced apoptosis
was accompanied by the activation of caspase-8 and caspase-3 as well as PARP cleavage. Cytochrome c and AIF release were also observed in Dox + DMPS-treated HL60/MX2 cells. Pretreatment with z-VAD-fmk markedly prevented caspase-3
activation and moderately suppressed apoptosis, suggesting that Dox + DMPS-induced apoptosis is somewhat (not completely)
dependent on caspase. Cytochrome c and AIF release were not affected by pretreatment with z-VAD-fmk. The ROS scavenger NAC efficiently suppressed not only ROS
generation, but also caspase-3-mediated PARP cleavage, apoptosis, and release of cytochrome c and AIF, indicating a role of ROS in combined Dox + DMPS treatment-induced apoptotic death signaling. Taken together, these
observations suggest that DMPS may be used as a therapeutic agent for overcoming drug-resistance in cancer cells by enhancing
drug-induced apoptosis. 相似文献
9.
Meng XW Fraser MJ Feller JM Ziegler JB 《Apoptosis : an international journal on programmed cell death》2000,5(3):243-254
Single-strand DNase and poly rAase, activities characteristic of endo-exonuclease, were co-activated in nuclear fractions of HL-60 cells by caspase-3. Activation was accompanied by cleavages of large soluble polypeptides (130–185 kDa) and a 65 kDa inactive chromatin-associated polypeptide related to the endo-exonuclease of Neurospora crassa as detected on immunoblots. The major products seen in vitro were a 77 kDa soluble polypeptide and an active chromatin-associated 34 kDa polypeptide. When HL-60 cells were induced to undergo apoptosis by treating with 50 M etoposide (VP-16) for 4 hours, 77 kDa and 40 kDa polypeptides accumulated in nuclear fractions. Chromatin DNA fragmentation activity was also activated in cytosol and nuclear extract either by pre-treating the cells in vivo with VP-16 or by treating the cytosol in vitro with caspase-3 or dATP and cytochrome c. Endo-exonuclease activated by caspase-3 in cytosol-derived fractions augmented chromatin DNA fragmentation activity in vitro. Endo-exonuclease is proposed to act in vivo in conjunction with the caspase-activated DNase (CAD) to degrade chromatin DNA during apoptosis of HL-60 cells. 相似文献
10.
Effect of pH, ionic charge, and osmolality on cytochrome c-mediated caspase-3 activity 总被引:6,自引:0,他引:6
Cytochromec-mediated activation of caspase-3 is the final commonpathway for most signals that induce apoptosis. Before release of cytochrome c from mitochondria, K+ andCl efflux and intracellular acidification must occur. Wehave utilized an in vitro assay to examine the role of pH, cations,anions, and uncharged molecules on the process of cytochromec-mediated activation of procaspase-3. In this cell-freesystem, a pH above 7.4 severely suppressed the activation ofprocaspase-3 but not the activity of caspase-3. KCl, NaCl, and othersalts all inhibited caspase activation, but uncharged molecules didnot. Comparison of the inhibitory capacity of various salts suggeststhat the crucial element in causing suppression is the cation. Theinhibition of alkaline pH could be overcome by increasingconcentrations of cytochrome c, whereas the inhibition ofionic charge could not, suggesting that pH and salts affect theactivation of caspase-3 by different mechanisms. 相似文献
11.
Caspase-3-dependent and caspase-3-independent pathways leading to chromatin DNA fragmentation in HL-60 cells 总被引:1,自引:0,他引:1
Meng XW Fraser MJ Feller JM Ziegler JB 《Apoptosis : an international journal on programmed cell death》2000,5(1):61-67
Apoptosis induced by etoposide (VP-16) in HL-60 cells was confirmed to be caspase-dependent. It was fully inhibited by the broad-spectrum caspase inhibitor Z-VAD-fmk. However, the caspase-3-specific inhibitor Z-DEVD-fmk only partially inhibited apoptosis. This indicated that a second caspase is required in vivo for full activation of the apoptotic nucease CAD. Aurin tricarboxylic acid (ATA) did not inhibit VP-16-induced apoptosis. In contrast, apoptosis induced by hydroxychloroquine (HCQ) in HL-60 cells was caspase-3 independent and was fully inhibited by ATA. Thus, CAD does not appear to be involved in chromatin DNA degradation in this case. A second apoptotic nuclease is postulated to degrade the DNA, likely endo-exonuclease, an abundant nuclear enzyme that acts on both DNA and RNA and is present in latent form. HCQ, but not VP-16, stimulated DNA degradation (laddering) in isolated nuclei. This indicates that the drug can act directly in the nuclei to trigger activation of the second latent apoptotic nuclease. 相似文献
12.
Contrasting effects of NO and peroxynitrites on HSP70 expression and apoptosis in human monocytes 总被引:4,自引:0,他引:4
Adrie C Richter C Bachelet M Banzet N François D Dinh-Xuan AT Dhainaut JF Polla BS Richard MJ 《American journal of physiology. Cell physiology》2000,279(2):C452-C460
The free radicals nitric oxide(·NO) and superoxide (O2·) react to formperoxynitrite (ONOO), a highly toxic oxidant species. Inthis study we investigated the respective effects of NO andONOO in monocytes from healthy human donors. Purifiedmonocytes were incubated for 6 or 16 h with a pure NO donor(S-nitroso-N-acetyl-DL-penicillamine, 0-2 mM), an ·NO/ONOO donor(3-morpholinosydnonimine chlorhydrate, 0-2 mM) with and withoutsuperoxide dismutase (200 IU/ml), or pure ONOO. Weprovide evidence that 3-morpholinosydnonimine chlorhydrate alonerepresents a strong stress to human monocytes leading to adose-dependent increase in heat shock protein-70 (HSP70) expression, mitochondrial membrane depolarization, and cell death by apoptosis andnecrosis. These phenomena were abolished by superoxide dismutase, suggesting that ONOO, but not ·NO, was responsible forthe observed effects. This observation was further strengthened by theabsence of a stress response in cells exposed toS-nitroso-N-acetyl-DL-penicillamine. Conversely, exposure of cells to ONOO alone also inducedmitochondrial membrane depolarization and cell death by apoptosis andnecrosis. Thus ONOO formation may well explain the toxiceffect generally attributed to ·NO. 相似文献
13.
DNA damage-induced [Zn(2+)](i) transients: correlation with cell cycle arrest and apoptosis in lymphoma cells 总被引:3,自引:0,他引:3
Smith PJ Wiltshire M Davies S Chin SF Campbell AK Errington RJ 《American journal of physiology. Cell physiology》2002,283(2):C609-C622
Reactive changes in free intracellularzinc cation concentration ([Zn2+]i) weremonitored, using the fluorescent probe Zinquin, in human lymphoma cells exposed to the DNA-damaging agent VP-16. Two-photon excitation microscopy showed that Zinquin-Zn2+ formscomplexes in cytoplasmic vesicles. [Zn2+]iincreased in both p53wt (wild type) and p53mut(mutant) cells after exposure to low drug doses. In p53mutcells noncompetent for DNA damage-induced apoptosis, elevated [Zn2+]i was maintained at higher drug doses,unlike competent p53wt cells that showed a collapse of thetransient before apoptosis. In p53wt cells, the[Zn2+]i rise paralleled an increase in p53and bax-to-bcl-2 ratio but preceded an increase in p21WAF1,active cell cycle arrest in G2, or nuclear fragmentation.Reducing [Zn2+]i, usingN,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine, caused rapid apoptosis in both p53wt andp53mut cells, although cotreatment with VP-16 exacerbatedapoptosis only in p53wt cells. This may reflectchanged thresholds for proapoptotic caspase-3 activation incompetent cells. We conclude that the DNA damage-induced transient isp53-independent up to a damage threshold, beyond which competent cellsreduce [Zn2+]i before apoptosis.Early stress responses in p53wt cells take place in anenvironment of enhanced Zn2+ availability. 相似文献
14.
Cadmium-induced ceramide formation triggers calpain-dependent apoptosis in cultured kidney proximal tubule cells 总被引:1,自引:0,他引:1
Lee WK Torchalski B Thévenod F 《American journal of physiology. Cell physiology》2007,293(3):C839-C847
A major target of cadmium (Cd2+) toxicity is the kidney proximal tubule (PT) cell. Cd2+-induced apoptosis of PT cells is mediated by sequential activation of calpains at 3–6 h and caspases-9 and -3 after 24-h exposure. Calpains also partly contribute to caspase activation, which emphasizes the importance of calpains for PT apoptosis by Cd2+. Upstream processes underlying Cd2+-induced calpain activation remain unclear. We describe for the first time that 10–50 µM Cd2+ causes a significant increase in ceramide formation by 22% (3 h) and 72% (24 h), as measured by diacylglycerol kinase assay. Inhibition of ceramide synthase with fumonisin B1 (3 µM) prevents ceramide formation at 3 h and abolishes calpain activation at 6 h, which is associated with significant attenuation of apoptosis at 3–6 h with Hoechst 33342 nuclear staining and/or 3(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) death assays. This indicates that Cd2+ enhances de novo ceramide synthesis and that calpains are a downstream target of ceramides in apoptosis execution. Moreover, addition of C6-ceramide to PT cells increases cytosolic Ca2+ and activates calpains. Apoptosis mediated by C6-ceramide at 24 h is significantly reduced by caspase-3 inhibition, which supports cross talk between calpain- and caspase-dependent apoptotic pathways. We conclude that Cd2+-induced apoptosis of PT cells entails endogenous ceramide elevation and subsequent Ca2+-dependent calpain activation, which propagates kidney damage by Cd2+. nephrotoxicity; cell signaling; cell biology and structure 相似文献
15.
A3 adenosine receptors regulate Cl- channels of nonpigmented ciliary epithelial cells 总被引:2,自引:0,他引:2
Mitchell Claire H.; Peterson-Yantorno Kim; Carre David A.; McGlinn Alice M.; Coca-Prados Miguel; Stone Richard A.; Civan Mortimer M. 《American journal of physiology. Cell physiology》1999,276(3):C659
Adenosinestimulates Cl channels ofthe nonpigmented (NPE) cells of the ciliary epithelium. We sought toidentify the specific adenosine receptors mediating this action.Cl channel activity inimmortalized human (HCE) NPE cells was determined by monitoring cellvolume in isotonic suspensions with the cationic ionophore gramicidinpresent. The A3-selective agonistN6-(3-iodobenzyl)-adenosine-5'-N-methyluronamide(IB-MECA) triggered shrinkage (apparentKd = 55 ± 10 nM). A3-selective antagonists blocked IB-MECA-triggered shrinkage, andA3-antagonists (MRS-1097, MRS-1191, and MRS-1523) also abolished shrinkage produced by 10 µMadenosine when all four known receptor subtypes are occupied. TheA1-selective agonistN6-cyclopentyladenosineexerted a small effect at 100 nM but not at higher or lowerconcentrations. The A2A agonistCGS-21680 triggered shrinkage only at high concentration (3 µM), aneffect blocked by MRS-1191. IB-MECA increased intracellularCa2+ in HCE cells and alsostimulated short-circuit current across rabbit ciliary epithelium.A3 message was detected in bothHCE cells and rabbit ciliary processes using RT-PCR. We conclude that human HCE cells and rabbit ciliary processes possessA3 receptors and that adenosinecan activate Cl channels inNPE cells by stimulating these A3 receptors. 相似文献
16.
Wang Q Wang L Feng YH Li X Zeng R Gorodeski GI 《American journal of physiology. Cell physiology》2004,287(5):C1349-C1358
Normal human ectocervical epithelial (hECE) cells undergo apoptosis in culture. Baseline apoptosis could be increased by shifting cells to serum-free medium and blocked by lowering extracellular calcium. Treatment with the ATPase apyrase attenuated baseline apoptosis, suggesting that extracellular ATP and purinergic mechanisms control the apoptosis. Treatment with ATP and the P2X7 receptor analog 2'-3'-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate (BzATP) increased apoptosis significantly, in a time- and dose-related manner. The threshold of ATP effect was 0.5 µM in hECE cells and 1 µM in CaSki cancer cells. The apoptotic effect of BzATP was additive in part to that of tumor necrosis factor (TNF)-, and it could be attenuated by lowering extracellular calcium and by treatment with the caspase-9 inhibitor Leu-Glu-His-Asp-O-methyl-fluoromethylketone (LEHD-FMK). Treatment with BzATP activated caspase-9, and, in contrast to TNF-, it had only a mild effect on caspase-8. Both BzATP and TNF- activated caspase-3, suggesting that BzATP activates predominantly the mitochondrial apoptotic pathway. Both hECE and CaSki cells secrete ATP into the extracellular fluid, and mean ATP activity in conditioned medium was 0.5 µM, which is in the range of values that suffice to activate the P2X7 receptor. On the basis of these findings we propose a novel autocrine-paracrine mechanism of cervical cell apoptosis that operates by P2X7 receptor control of cytosolic calcium and utilizes the mitochondrial apoptotic pathway. cervix; epithelium; ATP; 2'-3'-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate 相似文献
17.
The role of protein kinase C-beta (PKC-beta) in apoptosis induced by tumor necrosis factor (TNF)-alpha and anti-Fas monoclonal antibody (mAb) in the human myeloid HL-60 leukemia cell line was studied by using its variant HL-525, which is deficient in PKC-beta. In contrast to the parental HL-60 cells, HL-525 is resistant to TNF-alpha-induced apoptosis but sensitive to anti-Fas mAb-induced apoptosis. Both cell types expressed similar levels of the TNF-receptor I, whereas the Fas receptor was detected only in HL-525 cells. Transfecting the HL-525 cells with an expression vector containing PKC-beta reestablished their susceptibility to TNF-alpha-induced apoptosis. The apoptotic effect of TNF-alpha in HL-60 and the transfectants was abrogated by fumonisin, an inhibitor of ceramide generation, and by the peptide Ac-YVAD-BoMK, an inhibitor of caspase-1 and -4. Supplementing HL-525 cells with exogenous ceramides bypassed the PKC-beta deficiency and induced apoptosis, which was also restrained by the caspase-1 and -4 inhibitor. The apoptotic effect of anti-Fas mAb in HL-525 cells was abrogated by the antioxidants N-acetylcysteine and glutathione and by the peptide z-DEVD-FMK, an inhibitor of caspase-3 and -7. We suggest that TNF-alpha-induced apoptosis involves PKC-beta and then ceramide and, in turn, caspase-1 and/or -4, whereas anti-Fas mAb-induced apoptosis utilizes reactive oxygen intermediates and, in turn, caspase-3 and/or -7. 相似文献
18.
Brette F Lacampagne A Sallé L Findlay I Le Guennec JY 《American journal of physiology. Cell physiology》2003,285(2):C310-C318
Inactivation of the L-type Ca2+ current (ICaL) was studied in isolated guinea pig ventricular myocytes with different ionic solutions. Under basal conditions, ICaL of 82% of cells infused with Cs+-based intracellular solutions showed enhanced amplitude with multiphasic decay and diastolic depolarization-induced facilitation. The characteristics of ICaL in this population of cells were not due to contamination by other currents or an artifact. These phenomena were reduced by ryanodine, caffeine, cyclopiazonic acid, the protein kinase A inhibitor H-89, and the cAMP-dependent protein kinase inhibitor. Forskolin and isoproterenol increased ICaL by only 60% in these cells. Cells infused with either N-methyl-D-glucamine or K+-based intracellular solutions did not show multiphasic decay or facilitation under basal conditions. Isoproterenol increased ICaL by 200% in these cells. In conclusion, we show that multiphasic inactivation of ICaL is due to Ca2+-dependent inactivation that is reversible on a time scale of tens of milliseconds. Cs+ seems to activate the cAMP-dependent protein kinase pathway when used as a substitute for K+ in the pipette solution. L-type calcium current; calcium-dependent inactivation; facilitation; phosphorylation; cesium 相似文献
19.
Characterization of norepinephrine-evoked inward currents in interstitial cells isolated from the rabbit urethra 总被引:6,自引:0,他引:6
Sergeant GP Thornbury KD McHale NG Hollywood MA 《American journal of physiology. Cell physiology》2002,283(3):C885-C894
Freshly dispersedinterstitial cells from the rabbit urethra were studied by using theperforated-patch technique. When cells were voltage clamped at 60 mVand exposed to 10 µM norepinephrine (NE) at 80-s intervals, eitherlarge single inward currents or a series of oscillatory inward currentsof diminishing amplitude were evoked. These currents were blocked byeither phentolamine (1 µM) or prazosin (1 µM), suggesting that theeffects of NE were mediated via 1-adrenoceptors.NE-evoked currents were depressed by the blockers ofCa2+-activated Cl currents, niflumic acid (10 µM), and 9-anthracenecarboxylic acid (9-AC, 1 mM). The reversalpotential of the above currents changed in a predictable manner whenthe Cl equilibrium potential was altered, againsuggesting that they were due to activation of a Clconductance. NE-evoked currents were decreased by 10 µM cyclopiazonic acid, suggesting that they were dependent on store-releasedCa2+. Inhibition of NE-evoked currents by the phospholipaseC inhibitor 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate(100 µM) suggested that NE releases Ca2+ via an inositol1,4,5-trisphosphate (IP3)-dependent mechanism. Theseresults support the idea that stimulation of1-adrenoceptors releases Ca2+ from anIP3-sensitive store, which in turn activatesCa2+-activated Cl current in freshlydispersed interstitial cells of the rabbit urethra. This elevates slowwave frequency in these cells and may underlie the mechanismresponsible for increased urethral tone during nerve stimulation. 相似文献
20.
Mitsui Toshiaki; Loboda Tadeusz; Itoh Akira; Ikarashi Taro 《Plant & cell physiology》1999,40(8):884-893
Sugar starvation-induced synthesis and extracellular liberationof -amylase molecules in suspension-cultured cells of rice (Oryzasativa L.) required Ca2+, although the level of translatable-amylase mRNA was not affected in the presence of Ca2+. Sugardepletion markedly stimulated Ca2+ uptake by rice cells andsucrose supplementation reduced it. Immunohistochemical andelectron probe microanalyzer studies indicated an apparent resemblancebetween the distribution pattern of Ca2+ and that of -amylasemolecules induced in the sugar-depleted cells. Ca2+ uptake wasreduced by sucrose, maltose, fructose, and glucose similarlyat more than 5 mM, but was unaffected by mannitol (88 mM), 6-deoxy-D-glucose(10 mM), and 3-O-methyl-D-glucose (10 mM). Furthermore, an effectiveCa2+ channel blocker, La3+ significantly inhibited the Ca2+uptake and the synthesis and extracellular liberation of -amylasemolecules in the absence of sucrose, while a general P-typeATPase inhibitor, vanadate greatly stimulated both in the presenceof sucrose. We concluded that, by controlling the Ca2+ uptake,metabolic sugars regulate the protein synthesis and posttranslationalsecretory processes of -amylase molecules in rice cells.
4 Invited research fellow of the Japan Society for the Promotionof Science. Present address: Plant Physiology Department, WarsawAgricultural University, Rakowiecka Str. 26/30 02-528 Warsaw,Poland. 相似文献