• Biomolecules & Therapeutics
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• v.12 no.3
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• pp.165-174
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• 2004

The aim of the present study was to investigate the effect of FCCP (carbonyl cyanide p-trifluoromethoxyphenyIhydrazone), which is a potent mitochondrial uncoupler, on secretion of catecholamines (CA) from the perfused model of the rat adrenal gland and to establish the mechanism of its action. The perfusion of FCCP (3 ${\times}$ $10^{-5}$ M) into an adrenal vein of for 90 min resulted in great increases in CA secretions. Tachyphylaxis to CA-releasing effect of FCCP was not observed by repeated perfusion of it. The CA-releasing effects of FCCP were depressed by pre-treatment with pirenzepine, chlorisondamine, nicardipine, TMB-8, and the perfusion of EGTA plus $Ca^{2+}$-free medium. In the presence of FCCP (3 ${\times}$ $10^{-5}$ M), the CA secretory responses induced by Ach (5.32 ${\times}$ $10^{-3}$ M), and DMPP ($10^{-4}$ M) were significantly enhanced. Furthermore, the perfusion of CCCP (3 ${\times}$ $10^{-5}$ M), a similar mitochondrial uncoupler, into an adrenal vein for 90 min also caused an increased response in CA secretion. Taken together these experimental results indicate that FCCP causes the CA secretion the perfused rat adrenal medulla in a calcium-dependent fashion. It is suggested that this facilitatory effects of FCCP may be mediated by cholinergic receptor stimulation, which is relevant to both stimulation of the $Ca^{2+}$ influx and $Ca^{2+}$ release from cytoplasmic $Ca^{2+}$ stores.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.8
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• pp.3653-3656
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• 2012

Background: A number of effective prevention measures have been introduced in attempts to substantially reduce both the incidence and mortality due to many kinds of cancer. The search for new anti-cancer compounds in foods or in plant medicines is one realistic and promising approach to prevention. Chinese medicines provide a rich pool of novel and efficacious agents for cancer prevention and treatment. Previously it was demonstratrated that hyperin extracted from the Manchurian rhododendron leaf reduces the proliferation of many cancer cells. The present study was carried out to evaluate its effects on human endometrial cancer cell viability and apoptosis and to investigate its mechanisms of action in RL952 cells. Methods: Cell viability was measured using the MTT assay. Intracellular calcium ions were detected using laser-scanning confocal microscopy. The effects of hyperin on apoptosis related proteins in RL952 cells were examined using Western blot analysis. Results: The growth of RL952 cells was inhibited by treatment with hyperin. OD values of caspase-3 and caspase-9 were increased and expression of bcl-2 was increased and bax was decreased in protein levels in RL952 cells after 24 h of hyperin treatment, Moreover, intracellular calcium accumulation occurred in hyperin-treated cells. Conclusion: These results suggest that hyperin may play an important role in tumor growth suppression by inducing apoptosis in human endometrial cells via a $Ca^{2+}$-related mitochondrion apoptotic pathway in RL952 cells.

• The Journal of Korean Obstetrics and Gynecology
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• v.19 no.2
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• pp.92-106
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• 2006

Purpose : To address the ability of Olibanum to induce cell death, we investigated the effect of olibanum on cell apoptosis. Twenty-four hours later, apoptosis occurred following olibanum exposure in a dose-dependent manner. Methods : We culture HeLa cell which is human metrocarcinoma cell in D-MEM included 10% fetal bovine serum(Hyclone Laboratories) below $37^{\circ}C$, 5% CO2. Then we observed apoptosis of log phage cell which is changed cultivation liquid 24 Hours periodically. Results : The treatment of BAPTA-AM regulated olibanum-induced apoptosis in HeLa human cervical carcinoma cells. The 24 hr-earlier -thapsigargin-pretreated cell showed the resistance against olibanum-induced apoptosis and the Ru360-mitochondrial uniporter-inhibited olibanum-induced apoptosis, too. It means that olibanum leads to the accumulation of calcium and the resultant apoptosis in HeLa cells. Immunoblotting data also shows that the expression of GRP78, ER stress marker protein, was induced by the olibanum. Bcl-2, anti-apototic protein, was decreased and that the expression of Bax, pro-apoptotic protein, was increased by the addition of olibanum. Interestingly, the olibanum increased the activity of caspase-8 as well as calpain cysteine pretense in HeLa cervical carcinoma cells. Calpain inhibitor-calpastatin as well as caspase-8C/A expression abrogated olibanum-induced apoptosis in the carcinoma cells. The inhibition of caspase-8 regulated olibanum-induced calpain activation but the inhibition of calpain did not have any effect on the caspase-8 activation in HeLa human cervical carcinoma cells. Conclusion : We conclude that olibanum induces the accumulation of calcium and the resultant apoptosis in which caspase-8 and calpain are involved.

• Journal of Life Science
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• v.18 no.9
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• pp.1177-1185
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• 2008

In the present work, we show that spermine (spm)-induced cytotoxicity is due to the mitochondrial-dependent pathway triggered by the intracellular $Ca^{2+}$ increase in MCF-7 human breast cancer cells. Spm induced the intracellular $Ca^{2+}$ increase in a dose-dependent manner in the medium containing 1.5 mM $Ca^{2+}$. Even in the $Ca^{2+}$-free medium, spm could induce a minor $Ca^{2+}$ increase in a dose-dependent fashion, suggesting a probable leak from the internal storage. The cytotoxic effect of $Ca^{2+}$ could be further proved by using either BAPTA or ionophore. Spm-induced $Ca^{2+}$ increase led to the release of cytochrome c from mitochondria into the cytosol and the change of mitochondrial membrane potential. In MCF-7 cells, caspase-7 plays a key role in the downstream of apoptosis because caspase-3 is absent. In the cells treated with spm, the cleavage of caspase-7 and -12 was increased almost two-fold. The level of anti-apoptotic Bcl-2 protein decreased to 35% of the control; however, the cells showed increased expression of pro-apoptotic Bax protein about two-fold in response to spm. These results imply that the apoptotic signaling pathway activated by spm is likely to be mediated via the mitochondrial-dependent pathway.

• Animal cells and systems
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• v.8 no.2
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• pp.97-103
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• 2004

The cytotoxic effect of iron was examined in peritoneal macrophage to determine contributing factors by iron injection to rat. Viability was reduced by 24% by the iron-overload and by 30% by short-term iron addition. Total iron was increased by 45% in the iron-overloaded with remarkable elevation (9 to 14 fold) in the presence of $FeSO_4$. Free calcium was also increased by 19% in control and 44% in iron-overloaded group due to additional $FeSO_4$ NO and MDA were increased by 40% and 136%, respectively, with significant reduction (37%) of NAD(P)H. RCR and cytochrome c oxidase activity were lowered approximately by 10% with reduction of mitochondrial membrane potential. Addition of iron was frequently associated with altered distribution of mitochondria of high membrane potential in the iron-overloaded macrophage. These results suggest altered mitochondria with high NO and low NAD(P)H due to iron.

• Animal cells and systems
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• v.14 no.4
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• pp.245-252
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• 2010

Iron uptake in mitochondria and fractionated mitochondria compartments was studied to understand iron transport in heart mitochondria. The inner membrane is most active in iron uptake. Mitochondrial uptake was dependent on iron concentration and the amount of mitochondria. Iron transport was inversely proportional to pH in the range of 6.0 to 8.0. Iron transport reached a maximum after 30 min of incubation at $37^{\circ}C$. Iron uptake was inhibited by 1 mM ATP and stimulated by 1 mM ADP. The oxidative phosphorylation inhibitor oligomycin inhibited iron uptake, but rotenone and antimycin A did not. The divalent ions $Mg^{2+}$, $Cu^{2+}$, $Mn^{2+}$, and $Zn^{2+}$ suppressed iron uptake at $10\;{\mu}M$ and stimulated it at 1 mM. The divalent ion $Ca^{2+}$ stimulated iron uptake at $10\;{\mu}M$ and suppressed it at 1 mM, competing with iron. The uptake of calcium was stimulated by 10 to $1000\;{\mu}M$ ATP, while iron uptake was stimulated reciprocally by 10 to $1000\;{\mu}M$ ADP, suggesting that these ions have movements similar to those of ATP and ADP.

• The Korean Journal of Physiology and Pharmacology
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• v.19 no.5
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• pp.389-399
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• 2015

Zinc has been considered as a vital constituent of proteins, including enzymes. Mobile reactive zinc ($Zn^{2+}$) is the key form of zinc involved in signal transductions, which are mainly driven by its binding to proteins or the release of zinc from proteins, possibly via a redox switch. There has been growing evidence of zinc's critical role in cell signaling, due to its flexible coordination geometry and rapid shifts in protein conformation to perform biological reactions. The importance and complexity of $Zn^{2+}$ activity has been presumed to parallel the degree of calcium's participation in cellular processes. Whole body and cellular $Zn^{2+}$ levels are largely regulated by metallothioneins (MTs), $Zn^{2+}$ importers (ZIPs), and $Zn^{2+}$ transporters (ZnTs). Numerous proteins involved in signaling pathways, mitochondrial metabolism, and ion channels that play a pivotal role in controlling cardiac contractility are common targets of $Zn^{2+}$. However, these regulatory actions of $Zn^{2+}$ are not limited to the function of the heart, but also extend to numerous other organ systems, such as the central nervous system, immune system, cardiovascular tissue, and secretory glands, such as the pancreas, prostate, and mammary glands. In this review, the regulation of cellular $Zn^{2+}$ levels, $Zn^{2+}$-mediated signal transduction, impacts of $Zn^{2+}$ on ion channels and mitochondrial metabolism, and finally, the implications of $Zn^{2+}$ in health and disease development were outlined to help widen the current understanding of the versatile and complex roles of $Zn^{2+}$.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.2
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• pp.233-239
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• 2017

Intracellular calcium ($Ca^{2+}$) oscillation is an initial event in digestive enzyme secretion of pancreatic acinar cells. Reactive oxygen species are known to be associated with a variety of oxidative stress-induced cellular disorders including pancreatitis. In this study, we investigated the effect of hydrogen peroxide ($H_2O_2$) on intracellular $Ca^{2+}$ accumulation in mouse pancreatic acinar cells. Perfusion of $H_2O_2$ at $300{\mu}M$ resulted in additional elevation of intracellular $Ca^{2+}$ levels and termination of oscillatory $Ca^{2+}$ signals induced by carbamylcholine (CCh) in the presence of normal extracellular $Ca^{2+}$. Antioxidants, catalase or DTT, completely prevented $H_2O_2$-induced additional $Ca^{2+}$ increase and termination of $Ca^{2+}$ oscillation. In $Ca^{2+}$-free medium, $H_2O_2$ still enhanced CCh-induced intracellular $Ca^{2+}$ levels and thapsigargin (TG) mimicked $H_2O_2$-induced cytosolic $Ca^{2+}$ increase. Furthermore, $H_2O_2$-induced elevation of intracellular $Ca^{2+}$ levels was abolished under sarco/endoplasmic reticulum $Ca^{2+}$ ATPase-inactivated condition by TG pretreatment with CCh. $H_2O_2$ at $300{\mu}M$ failed to affect store-operated $Ca^{2+}$ entry or $Ca^{2+}$ extrusion through plasma membrane. Additionally, ruthenium red, a mitochondrial $Ca^{2+}$ uniporter blocker, failed to attenuate $H_2O_2$-induced intracellular $Ca^{2+}$ elevation. These results provide evidence that excessive generation of $H_2O_2$ in pathological conditions could accumulate intracellular $Ca^{2+}$ by attenuating refilling of internal $Ca^{2+}$ stores rather than by inhibiting $Ca^{2+}$ extrusion to extracellular fluid or enhancing $Ca^{2+}$ mobilization from extracellular medium in mouse pancreatic acinar cells.

• The Korean Journal of Zoology
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• v.15 no.4
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• pp.163-182
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• 1972

The effects of caffeine on the ATPase activity and Ca uptake of the fragmented sarcoplasmic reticulum isolated from rabbit skeletal muscle were studied. The ATPase activity of the heavy fraction (2,000-8,000xG) was stimulated by caffeine while that of other lighter fractions was not. It is suggested that the enhancement of the ATPase by the caffeine treatment. The Ca uptake of the heavy and middle (10,000-20,000xG) fractions was inhibited by caffeine when measured at the medium Ca concentration higher than 200 nmoles/mg protein, while only that of the heavy fraction was inhibited when measured at the Ca concentration below 200 nmoles/mg protein. Experiments with dicumarol suggested that caffeine inhibits the Ca uptake of the mitochondria as well as that of the sarcoplasmic reticulum and that the inhibition of the Ca uptake by caffeine in the low Ca concentration in the heavy fraction is due to the inhibition of the mitochondrial Ca uptake by caffeine. It appeared highly probable that the potentiation of muscle contraction caused by caffeine is solely due to the inhibition of the Ca uptake by and to the release of the accumulated Ca from the sarcoplasmic reticulum.

• Proceedings of the Korean Society of Applied Pharmacology
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• 2002.07a
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• pp.236-236
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• 2002

Pyruvate dehydrogenase phosphatase (PDP) is a mitochondrial protein serine/threonine phosphatase that catalyzes the dephosphorylation and concomitant reactivation of the pyruvate dehydrogenase componant of the pyruvate dehydrogenase complex (PDC). PDP consists of a Mg$\^$+2/ -dependent and Ca$\^$+2)-stimulated catalytic subunit (PDPc) of Mr 52,600 and a FAD-containing regulatory subunit (PDPr) of Mr 95.600. Catalytic subunit of pyruvate dehydrogenase phosphatase (PDPc) has been suggested to have three major functional domains such as dihydrolipoamide acetyltransferase(E$_2$)-binding domain, regulatory subunit of PDP(PDPr)-binding domain, and calcium-binding domain. In order to identify functional domains, recombinant catalytic subunit of pyruvate dehydrogenase phosphatase (rPDPc) was expressed in E. coli JM101 and purified to near homogeneity using the unique property of PDPc: PDPm binds to the inner lipoyl domain (L$_2$) of E$_2$ of pyruvate dehydrogenase complex (PDC) in the presence of Ca$\^$+2/, not under EGTA. PDPc was limited-proteolysed by trypsin, chymotrypsin, Arg-C, and elastase at pH7.0 and 30$^{\circ}C$ and N-terminal analysis of the fragment was done. Chymotrypsin, trypsin, and elastase made two major framents: N-terminal large fragment, approx. 50kD and C-terminal small fragment, approx. 0 kDa. Arg-C made three major fragments: N-terminal fragment, approx. 35 kD, and central fragment, approx. 15 kD, and C-terminal fragment, approx. 10 kD. This study strongly suggest that PDPc consists of three major functional domains. However, further study should be necessary to identify the functional role.