• The Korean Journal of Physiology and Pharmacology
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• 제5권5호
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• pp.433-441
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• 2001

The ATP-sensitive potassium $(K_{ATP})$) channel is a member of inward rectifier potassium channel (Kir) that is inhibited by intracellular ATP and functions in close relation to sulfonylurea receptors (SUR). Although the molecular mechanism and physiological function of $K_{ATP}$ channels are well understood, the expression pattern during development or treatment with the channel modulators such as glybenclamide is little known. In this work, we determined mRNA levels of a $K_{ATP}$ channel (Kir6.2) and a sulfonylurea receptor (SUR2) in rat tissues by RNase protection assay. Levels of Kir6.2 and SUR2 mRNA in the rat brain and skeletal muscle were higher in adult $(90{\sim}120\;days)$ than in neonate $(2{\sim}8\;days),$ whereas those in the heart were not much different between neonate $(2{\sim}8\;days)$ and adult $(90{\sim}120\;days).$ In addition, none of $K_{ATP}$ channel modulators (opener, pinacidil and nicorandil; blocker, glybenclamide) affected the Kir6.2 mRNA levels in the heart, brain and skeletal muscle. The results indicate that the expression of Kir and SUR genes can vary age-dependently, but the expression of Kir is not dependent on the long-term treatment of channel modulators. The effect of the channel modulators on mRNA level of SUR is remained to be studied further.

• Journal of Preventive Medicine and Public Health
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• 제32권2호
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• pp.170-176
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• 1999

본 연구는 EMT-6 세포를 이용하여 무기수은, 유기수은 및 카드뮴의 세포독성에 대한 glutathione(GSH)의 방어효과를 알아보고자 하였다. 무기수은, 유기수은 및 카드뮴을 첨가한 배양조건에서 EMT-6 세포의 세포생존율, ${NO_2}^-$ 및 ATP 생성량은 첨가한 중금속의 농도가 증가할수록 용량의존적으로 감소하였다. GSH, OTC 및 BSO를 단독 첨가한 배양조건은 세포의 세포생존율과 NO2- 및 ${NO_2}^-$ 생성량에 영향을 주지 않았다. 수은화합물 및 카드뮴과 GSH를 동시 첨가한 배양조건에서는 세포생존율이 90% 이상 유지되었고, ${NO_2}^-$ 및 ATP 생성량은 기본배양조건과 비슷한 수준으로 나타났다. $16{\mu}M$의 무기 및 유기수은과 $160{\mu}M$의 카드뮴을 첨가한 실험조건에 GSH를 동시 첨가했을 경우 방어효과는 GSH의 농도에 따라 용량의존적으로 증가하였다. 세포내에서 수은 및 카드뮴의 세포독성에 대한 GSH역할을 알아보고자 GSH, OTC, BSO 전처리 실험을 한 결과, GSH의 전처리는 이들이 세포막을 통과하지 못하기 때문에 대조군과 비슷한 양상으로 나타난 반면에 BSO를 전처리한 군에서는 세포내 GSH 농도의 감소로 수은의 세포 독성이 증가하여 대조군에 비하여 ${NO_2}^-$와 ATP 생성량이 현저히 감소하였다. 또한 세포내 GSH의 농도를 증가시키는 OTC를 전처리한 결과 수은의 독성에 대한 방어효과가 시간 및 용량 의존적으로 현저하게 증가하였다. 이러한 실험결과는 수은의 세포독성에 대한 GSH의 방어효과가 GSH 세포내 농도와 밀접한 관련이 있음을 간접적으로 보여주고 있다. 본 연구의 결과는 수은 및 카드뮴의 독성에 대한 GSH의 방어작용이 단순히 -SH기와 중금속의 결합에 의한 결과가 아니라 세포내에서 GSH 분자가 갖는 고유의 기능으로 판단되며, 특히 중금속에 의한 에너지대사의 장애를 GSH가 회복시킬 수 있음을 보여준다.

• The Korean Journal of Physiology and Pharmacology
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• 제25권1호
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• pp.69-77
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• 2021

Propofol infusion syndrome characterized by rhabdomyolysis, metabolic acidosis, kidney, and heart failure has been reported in long-term propofol use for sedation. It has been reported that intracellular adenosine triphosphate (ATP) is reduced in rhabdomyolysis. The study aims to investigate the protective effect of ATP against possible skeletal muscle damage of propofol in albino Wistar male rats biochemically and histopathologically. PA-50 (n = 6) and PA-100 (n = 6) groups of animals was injected intraperitoneally to 4 mg/kg ATP. An equal volume (0.5 ml) of distilled water was administered intraperitoneally to the P-50, P-100, and HG groups. One hour after the administration of ATP and distilled water, 50 mg/kg propofol was injected intraperitoneally to the P-50 and PA-50 groups. This procedure was repeated once a day for 30 days. The dose of 100 mg/kg propofol was injected intraperitoneally to the P-100 and PA-100 groups. This procedure was performed three times with an interval of 1 days. Our experimental results showed that propofol increased serum CK, CK-MB, creatinine, BUN, TP I, ALT, AST levels, and muscle tissue MDA levels at 100 mg/kg compared to 50 mg/kg and decreased tGSH levels. At a dose of 100 mg/kg, propofol caused more severe histopathological damage compared to 50 mg/kg. It was found that ATP prevented propofol-induced muscle damage and organ dysfunction at a dose of 50 mg/kg at a higher level compared to 100 mg/kg. ATP may be useful in the treatment of propofol-induced rhabdomyolysis and multiple organ damage.

• BMB Reports
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• 제28권1호
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• pp.79-82
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• 1995

Exposure of spleen lymphocytes to 2-chloroethylethyl sulfide (CEES) leads to a reduction of the intracellular ATP level, followed by a decrease in cell viability. Addition of nicotinamide, an inhibitor of poly(ADP-ribose) polymerase (PADPRP), restores both ATP level and viability, indicating that an activation of PADPRP is responsible for the cytotoxicity of CEES. The involvement of a $Ca^{2+}$-mediated process in cytotoxicity is suggested. Verapamil, EGTA, trifluoperazine, and butacaine exhibit a partial protection (20 to 58%) against the cytotoxicity of CEES. Investigation of the causative role of proteolytic degradation in cell death indicate that pepstatin and leupeptin exert a substantial protective effect (60 to 70%), suggesting the involvement of lysosomal destabilization in CEES-induced cytotoxicity. Also, lysosomotropic agents markedly decrease the cytotoxicity. Lysosomal labilization may be a mechanism for the cytotoxicity of CEES.

• 대한소아치과학회지
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• 제35권1호
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• pp.47-56
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• 2008

리스페리돈(risperidone)은 세계적으로 가장 널리 처방되고 있는 정신분열증 치료제로서 소아자폐증의 선택약물로 FDA 승인을 받았으며 틱장애, 뚜렛장애의 치료제로도 쓰이고 있다. 치과와 관련된 리스페리돈의 이상반응으로 구강건조가 보고되고 있으며 그 기전은 밝혀지지 않은 상태이다. 본 연구의 목적은 리스페리돈이 타액분비 기전의 중요한 요소인 세포내 칼슘농도에 미치는 영향을 세포수준에서 밝히고자 하는 것이다. 세포내 칼슘농도를 측정하기 위해 Human salivary gland cell line(HSG)에 Fura-2/AM을 세포내로 부하한 뒤 340 및 380 nm의 파장으로 교대로 여기시킬 때 방출되는 형광강도를 500 nm 파장에서의 비율로 측정하였다. 각 실험 후 형광강도의 비율을 실제 세포내 칼슘농도로 보정하기 위한 calibration 실험을 시행하였다. 카바콜, ATP, 히스타민을 처리하여 세포내 칼슘농도의 변화를 측정하고 리스페리돈의 전처리가 이에 미치는 효과를 비교하였으며 다음과 같은 결과를 얻었다. 1. HSG에서 카바콜, ATP, 히스타민 처리로 인해 세포내 칼슘농도가 증가하였으며 리스페리돈을 전처리한 경우 카바콜과 ATP의 작용에는 영향을 주지 않았으나 히스타민의 작용을 억제하였다. 2. HSG의 세포내 칼슘 변화에 미치는 히스타민의 효과는 농도의존적인 양상을 보였으며 50% 유효농도($EC_{50}$)는 $3.3{\pm}0.5\;{\mu}M$이었다. 3. 히스타민에 의한 HSG에서 칼슘 변화에 미치는 리스페리돈의 저해 효과는 농도의존적인 양상을 보였으며 대조군의 효과를 50% 억제하는 농도($IC_{50}$)는 $104.4{\pm}14\;nM$로 리스페리돈의 적정혈중농도 이하에 해당되었다. 4. 리스페리돈은 히스타민에 의한 소포체에서의 칼슘 유리와 세포 밖 칼슘 유입을 모두 유의성 있게 억제하였다(p<0.05). 항정신병 약물은 장기간 복용하고 적정혈중농도가 계속 유지되기 때문에 이러한 약물이 타액분비감소를 일으킬 경우 다발성우식증 등 심각한 치과적 질환을 야기할 수 있으므로 이에 대한 예방 및 치료방안이 필요하리라 사료된다.

• Archives of Pharmacal Research
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• 제28권7호
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• pp.810-815
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• 2005

Previously, we have shown that astrocytes deprived of glucose became highly vulnerable to peroxynitrite, and adenosine and its metabolites attenuated the gliotoxicity via the preservation of cellular ATP level. Here, we found that adenosine and related metabolites prevented the disruption of mitochondrial transmembrane potential (MTP) in glucose-deprived rat primary astrocytes exposed to 3-morpholinosydnonimine (SIN-1), a peroxynitrite releasing agent. Exposure to glucose deprivation and SIN-1(2h) significantly disrupted MTP in astrocytes, and adenosine prevented it in dose-dependent manner with an $EC_{50}\;of\;5.08{\mu}M$. Adenosine also partially prevented the cell death by myxothiazol, a well-known inhibitor of mitochondrial respiration. Blockade of adenosine deamination or intracellular transport with erythro-9-(-hydroxy-3-nonyl)adenosine (EHNA) or S-(4-nitrobenzyl)-6-thioinosine (NBTI), respectively, completely reversed the protective effect of adenosine. Other purine nucleos(t)ides including inosine, guanosine, ATP, ADP, AMP, ITP, and GTP also showed similar protective effects. This study indicates that adenosine and related purine nucleos(t)ides may protect astrocytes from peroxynitrite-induced mitochondrial dysfunction.

• Molecules and Cells
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• 제38권2호
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• pp.138-144
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• 2015

Raloxifene is a selective estrogen receptor modulator (SERM) that binds to the estrogen receptor (ER), and exhibits potent anti-tumor and autophagy-inducing effects in breast cancer cells. However, the mechanism of raloxifene-induced cell death and autophagy is not well-established. So, we analyzed mechanism underlying death and autophagy induced by raloxifene in MCF-7 breast cancer cells. Treatment with raloxifene significantly induced death in MCF-7 cells. Raloxifene accumulated GFP-LC3 puncta and increased the level of autophagic marker proteins, such as LC3-II, BECN1, and ATG12-ATG5 conjugates, indicating activated autophagy. Raloxifene also increased autophagic flux indicators, the cleavage of GFP from GFP-LC3 and only red fluorescence-positive puncta in mRFP-GFP-LC3-expressing cells. An autophagy inhibitor, 3-methyladenine (3-MA), suppressed the level of LC3-II and blocked the formation of GFP-LC3 puncta. Moreover, siRNA targeting BECN1 markedly reversed cell death and the level of LC3-II increased by raloxifene. Besides, raloxifene-induced cell death was not related to cleavage of caspases-7, -9, and PARP. These results indicate that raloxifene activates autophagy-dependent cell death but not apoptosis. Interestingly, raloxifene decreased the level of intracellular adenosine triphosphate (ATP) and activated the AMPK/ULK1 pathway. However it was not suppressed the AKT/mTOR pathway. Addition of ATP decreased the phosphorylation of AMPK as well as the accumulation of LC3-II, finally attenuating raloxifene-induced cell death. Our current study demonstrates that raloxifene induces autophagy via the activation of AMPK by sensing decreases in ATP, and that the overactivation of autophagy promotes cell death and thereby mediates the anti-cancer effects of raloxifene in breast cancer cells.

• Toxicological Research
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• 제22권4호
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• pp.397-402
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• 2006

Lipid mediator such as lysophosphatidic acid (LPA) plays an important role in inflammation and wound heating, has been recently reported to induce influx of extracellular calcium into erythrocytes. This elevation in intracellular calcium level may cause destruction of membrane asymmetry and procoagulant microvesicle formation. Thus, we investigated if the lipid mediator could induce microvesicle formation as a result of extracellular calcium influx in human erythrocytes. Treatment with lipid mediator to erythrocytes resulted in microvesicle generation In a concentration-, time-dependent manner. Microvesicles formed expressed procoagulant phosphatidylserine (PS) on their surface membrane significantly as well. LPA did not affect the band 3 phosphorylation which is involved in morphological change in erythrocytes. Pretreatment with suramin did not inhibit LPA-induced microvesicle generation, suggesting microvesicle generation was not receptor-dependent pathway. Depletion of intracellular ATP levels in erythrocytes was suggested to be one of the mechanism for these events.

• Nutrition Research and Practice
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• 제16권2호
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• pp.147-160
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• 2022

BACKGROUND/OBJECTIVES: Patients with chronic kidney disease (CKD) have a high concentration of uremic toxins in their blood and often experience muscle atrophy. Indoxyl sulfate (IS) is a uremic toxin produced by tryptophan metabolism. Although an elevated IS level may induce muscle dysfunction, the effect of IS on physiological concentration has not been elucidated. Additionally, the effects of ursolic acid (UA) on muscle hypertrophy have been reported in healthy models; however, it is unclear whether UA ameliorates muscle dysfunction associated with chronic diseases, such as CKD. Thus, this study aimed to investigate whether UA can improve the IS-induced impairment of mitochondrial biogenesis. MATERIALS/METHODS: C2C12 cells were incubated with or without IS (0.1 mM) and UA (1 or 2 μM) to elucidate the physiological effect of UA on CKD-related mitochondrial dysfunction and its related mechanisms using real-time reverse transcription-polymerase chain reaction, western blotting and enzyme-linked immunosorbent assay. RESULTS: IS suppressed the expression of differentiation marker genes without decreasing cell viability. IS decreased the mitochondrial DNA copy number and ATP levels by downregulating the genes pertaining to mitochondrial biogenesis (Ppargc1a, Nrf1, Tfam, Sirt1, and Mef2c), fusion (Mfn1 and Mfn2), oxidative phosphorylation (Cycs and Atp5b), and fatty acid oxidation (Pdk4, Acadm, Cpt1b, and Cd36). Furthermore, IS increased the intracellular mRNA and secretory protein levels of interleukin (IL)-6. Finally, UA ameliorated the IS-induced impairment in C2C12 cells. CONCLUSIONS: Our results indicated that UA improves the IS-induced impairment of mitochondrial biogenesis by affecting differentiation, ATP levels, and IL-6 secretion in C2C12 cells. Therefore, UA could be a novel therapeutic agent for CKD-induced muscle dysfunction.

• 대한수의학회지
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• 제35권2호
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• pp.245-254
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• 1995

The density of ATP-sensitive potassium($K_{APT}$) channels, that open as intracellular ATP concentration falls below a critical level, is very high in skeletal muscle surface membrane and those high density may imply that $K_{ATP}$ channels have very important physiological roles. To elucidate a role of $K_{ATP}$ in relation to fatigue, the modulating effects of potassium channel openers and blockers on the fatigue velocity(FV) of mouse extensor hallucis longus muscle(EHL) were investigated in vitro. Twitch contraction was induced by an electrical field stimulation (EFS: 24-48V, 20ms, 0.2-4Hz) and resulting contraction force was isometrically recorded. The twitch forces were gradually decreased to 25% of initial contraction force(ICF) in $37.52{\pm}1.55sec$($mean{\pm}s.e.m.$, n=135), indicating the fatigue phenomena. The mean velocity for development of the fatigue was measured during the period that twitch force decreased to half($FV_{0/0.5}$) and during the period from half to 25%($FV_{0.5/0.25}$) of ICF. The fatigue was induced once every one hour and the tissue response was stable for up to 4 hours. In control condition, ICF was $5.8{\pm}0.12g$ (n=144) and decreased to 50% of ICF with the mean fatigue velocity of $0.182{\pm}0.006g/sec$($FV_{0/0.5}$, n=135) and from 50% to 25% of ICF with $0.084{\pm}0.004g/sec$($FV_{0.5/0.25}$, n=135). Cromakalim($50{\mu}M$) significantly increased $FV_{0.5/0.25}$(n=4). Glibenclamide($IC_{50}>50{\mu}M$), $Ba^{2+}$($IC_{50}=10{\mu}M$), 4-aminopyridine($FV_{0/0.5}$, $IC_{50}=0.5mM$; $FV_{0.5/0.25}$, $IC_{50}=2mM$) decreased both $FV_{0/0.5}$ and $FV_{0.5/0.25}$ concentration-dependently up to 75%. $TEA^+$(30mM), E-4031($10{\mu}M$), tolbutamide(1mM) decreased $FV_{0.5/0.25}$, but apamin(300nM) and $TEA^+$(10mM) showed no significant effects. Our results suggest that activation of the $K_{ATP}$ channels may be major cause of $K^+$ outflux during development of the fatigue and the isolated EHL muscle could be an useful experimental preparation in studying the fatigue phenomena in skeletal muscle. In addition, the possibility of activation of delayed rectifier during the fatigue development remains to be studied further.