• Molecules and Cells
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• 제21권1호
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• pp.1-6
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• 2006

Bcl-2/adenovirus E1B 19 kDa-interacting protein 3 (BNIP3) is a mitochondrial pro-apoptotic protein that has a single Bcl-2 homology 3 (BH3) domain and a COOH-terminal transmembrane (TM) domain. Although it belongs to the Bcl-2 family and can heterodimerize with Bcl-2, its pro-apoptotic activity is distinct from those of other members of the Bcl-2 family. For example, cell death mediated by BNIP3 is independent of caspases and shows several characteristics of necrosis. Furthermore, the TM domain, but not the BH3 domain, is required for dimerization, mitochondrial targeting and pro-apoptotic activity. BNIP3 plays an important role in hypoxia-induced death of normal and malignant cells. Its expression is markedly increased in the hypoxic regions of some solid tumors and appears to be regulated by hypoxia-inducible factor (HIF), which binds to a site on the BNIP3 promoter. Silencing, followed by methylation, of the BNIP3 gene occurs in a significant proportion of cancer cases, especially in pancreatic cancers. BNIP3 also has a role in the death of cardiac myocytes in ischemia. Further studies of BNIP3 should provide insight into hypoxic cell death and may contribute to improved treatment of cancers and cardiovascular diseases.

• 대한수의학회지
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• 제34권1호
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• pp.25-36
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• 1994

The inward tail current after a short depolarizing pulse has been known as Na-Ca exchange current activated by intracellular calcium which forms late plateau of the action potential in rabbit atrial myocytes. Chloride conductance which is also dependent upon calcium concentration has been reported as a possible tail current in many other excitable tissues. Thus, in order to investigate the exsitance of the calcium activated chloride current and its contribution to tail current, whole cell voltage clamp measurement has been made in single atrial cells of the rabbit. The current was recorded during repolarization following a brief 2 ms depolarizing pulse to +40mV from a holding potential of -70mV. When voltage-sensitive transient outward current was blocked by 2 mM 4-aminopyridine or replacement potassium with cesium, the tail current were abolished by ryanodine$(1{\mu}M)$ or diltiazem$(10{\mu}M)$ and turned out to be calcium dependent. The magnitudes of the tail currents were increased when intracellular chloride concentration was increased to 131 mM from 21 mM. The current was decreased by extracellular sodium reduction when intracellular chloride concentration was low(21 mM), but it was little affected by extracellular sodium reduction when intracellual chloride concentration was high(131 mM). The current-voltage relationship of the difference current before and after extracellular sodium reduction, shows an exponential voltage dependence with the largest magnitude of the current occurring at negative potentials, with is similar to current-voltage relationship at negative potentials, which is similar to current-voltage relationship of Na-Ca exchange current. The current was also decreased by $10{\mu}M$ niflumic acid and 1 mM bumetanide, which is well known anion channel blockers. The reversal potentials shifted according to changes in chloride concentration. The current-voltage relationships of the niflumic acid-sensitive currents in high and low concentration of chloride were well fitted to those predicted as chloride current. From the above results, it is concluded that calcium activated chloride component exists in the tail current with Na-Ca exchange current and it shows the reversal of tail current. Therefore it is thought that in the physiologic condition it leads to rapid end of action potential which inhibits calcium influx and it contributes to maintain the low intracellular calcium concentration with Na-Ca exchange mechanism.

• Journal of Chest Surgery
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• 제31권8호
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• pp.739-748
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• 1998

연구배경: 재관류 없이 허혈 심근세포의 기능이나 형태학적 변화를 원래의 상태로 회복시킬 수 없음은 주 지의 사실이나 재관류가 반드시 유익하지만은 않다는 실험결과들이 있으므로 국소적으로 일과성 허혈을 유도하여 심근세포에서 일어나는 형태학적 변화를 관찰하고 재관류가 허혈심근에 미치는 영향을 알아보고자 하였다. 재료 및 방법: 생후 12개월 내외의 홀슈타인종 소를 사용하여 정맥 마취하에서 흉부를 열고 왼쪽 관상동맥 의 전하방 가지를 20분 동안 결찰하여 국소허혈을 유도하고 결찰을 풀어 재관류를 유도하였다. 위험부위의 심근조직 을 재관류 직후, 재관류후 1, 2, 3, 6, 12시간 및 재관류 12시간후 1시간동안 보조호흡과 수액공급만을 한 후 각각 생검하여 통상적인 방법으로 처리하여 투과전자현미경으로 관찰하였다. 결과: 20분 동안의 국소허혈로 심근세포는 경도에서 중등도에 이르는 미세구조적 변화들이 나타났는데 특히 세포막하, 핵, 사립체, 심근원섬유 등의 소기관에서 많은 변화들이 관찰되었다. 그러나 재관류를 시작하여 1시간이 지나면 허혈심근세포에서는 회복을 시사하는 소견들이 나타나기 시작하였으며 미세혈관 내에서 혈전형성이나 내강의 협착 등이 관찰되었으나 재관류가 계속됨에 따라 심근세포의 미세구조적 변화들은 서서히 회복되어 가는 양상을 나 타내었다. 그러나 시간이 경과함에 따라 심내막하 심근세포의 일부에서는 재관류 손상으로 추측되는 미세구조적 변 화들이 관찰되었다. 결론: 이상의 결과로 미루어 미세혈관은 허혈에 대한 저항력이 심근세포보다 강하며 허혈 심근세포는 재관류 없이는 회복될 수 없고, 회복에는 비교적 장시간이 요구되며, 따라서 혈관폐쇄로 인한 허혈시 혈관성형술이나 혈전 용해를 촉진하는 물질을 이용한 치료는 타당성이 있는 것으로 생각되나 재관류 유발성 세포손상에 대한 주의가 요망 된다.

• The Korean Journal of Physiology and Pharmacology
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• 제2권6호
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• pp.725-732
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• 1998

In order to elucidate the molecular mechanism of the intracellular $Ca^{2+}$ overload frequently reported from diabetic heart, diabetic rats were induced by the administration of streptozotocin, the membrane vesicles of junctional SR (heavy SR, HSR) were isolated from the ventricular myocytes, and SR $Ca^{2+}$ uptake and SR $Ca^{2+}$ release were measured. The activity of SR $Ca^{2+}-ATPase$ was $562{\pm}14$ nmol/min/mg protein in control heart. The activity was decreased to $413{\pm}30$ nmol/min/mg protein in diabetic heart and it was partially recovered to $485{\pm}18$ nmol/min/mg protein in insulin-treated diabetic heart. A similar pattern was observed in SR $^{45}Ca^{2+}$ uptakes; the specific uptake was the highest in control heart and it was the lowest in diabetic heart. In SR $^{45}Ca^{2+}$ release experiment, the highest release, 45% of SR $^{45}Ca^{2+}$, was observed in control heart. The release of diabetic heart was 20% and it was 30% in insulin-treated diabetic heart. Our results showed that the activities of both SR $Ca^{2+}-ATPase$ and SR $Ca^{2+}$ release channel were decreased in diabetic heart. In order to evaluate how these two factors contribute to SR $Ca^{2+}$ storage, the activity of SR $Ca^{2+}-ATPase$ was measured in the uncoupled leaky vesicles. The uncoupling effect which is able to increase the activity of SR $Ca^{2+}-ATPase$ was observed in control heart; however, no significant increments of SR $Ca^{2+}-ATPase$ activities were measured in both diabetic and insulin-treated diabetic rats. These results represent that the $Ca^{2+}$ storage in SR is significantly depressed and, therefore, $Ca^{2+}-sequestering$ activity of SR may be also depressed in diabetic heart.

• The Korean Journal of Physiology and Pharmacology
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• 제7권6호
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• pp.333-339
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• 2003

It has been demonstrated that an unidentified cytosolic factor(s) reduces $K_{ACh}$ channel function. Therefore, this study attempted to elucidate the cytosolic factor. Fresh cytosol isolated from normal heart (FC) depressed the $K_{ACh}$ channel activity, but cytosol isolated from the ischemic hearts (IC) did not modulate the channel function. Electrophorectic analysis revealed that a protein of ${\sim}80 kDa was markedly reduced or even lost in IC. By using peptide sequencing analysis and Western blot, this 80 kDa protein was identified as transferrin (receptor-mediated $Fe^{3+}$ transporter, 76 kDa). Direct application of transferrin (100 nM) to the cytoplasmic side of inside-out patches decreased the open probability ($P_o$, 12.7${\pm}6.4%, n=4) without change in mean open time (${\tau}_o$, $98.5{\pm}1.3$%, n=4). However, the equimolar apotransferrin, which is free of $Fe^{3+}$, had no effect on the channel activity (N*$P_o$, $129.1{\pm}13.5$%, n=3). Directly applied $Fe^{3+}$ (100 nM) showed results similar to those of transferrin (N*$P_o$: $21.1{\pm}3.9$%, n=5). However $Fe^{2+}$ failed to reduce the channel function (N*$P_o$, $106.3{\pm}26.8$%, n=5). Interestingly, trivalent cation La3+ inhibited N*$P_o$ of the channel ($6.1{\pm}3.0$%, n=3). Taken together, these results suggest that $Fe^{3+}$ bound to transferrin can modulate the $K_{ACh}$ channel function by its electrical property as a polyvalent cation.

• The Korean Journal of Physiology and Pharmacology
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• 제11권2호
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• pp.45-55
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• 2007

We elucidated the effects of various components of ischemic medium on the outcome of simulated ischemia-reperfusion injury. Hypoxia for up to 12 hours induced neither apoptotic bodies nor LDH release. However, reoxygenation after 6 or 12 hours of hypoxia resulted in a marked LDH release along with morphological changes compatible with oncotic cell death. H9c2 cells were then subjected to 6 hours of simulated ischemia by exposing them to modified hypoxic glucose-free Krebs-Henseleit buffer. Lowered pH (pH 6.4) of simulated-ischemic buffer resulted in the generation of apoptotic bodies during ischemia, with no concomitant LDH release. The degree of reperfusion-induced LDH release was not affected by the pH of ischemic buffer. Removal of sodium bicarbonate from the simulated ischemic buffer markedly increased cellular damages during both the simulated ischemia and reperfusion. Addition of lactate to the simulated ischemic buffer increased apoptotic cell death during the simulated ischemia. Most importantly, concomitant acidosis and high lactate concentration in ischemic buffer augmented the reperfusion-induced oncotic cell death. These results confirmed the influences of acidosis, bicarbonate deprivation and lactate on the progression and outcome of the simulated ischemia-reperfusion, and also demonstrated that concomitant acidosis and high lactate concentration in simulated ischemic buffer contribute to the development of reperfusion injury.

• Biomolecules & Therapeutics
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• 제1권1호
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• pp.9-19
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• 1993

DA-125, a new anthracycline antibiotic, showed antitumor activity against animal tumors and human tumors. Therefore we studied the cardiotoxic potential of DA-125 in hamsters and rats as a part of safety research, and compared it with that of doxorubicin(DXR). In acute cardiotoxicity test model used hamsters DA-125 was administered intravenously at a dose of 6, 9, 12 mg/kg, and DXR at 3 mg/kg was given. The electrocardiogram(ECG) of hamsters was recorded for 30 minutes after administration. The DA-125 caused slight ECG alterations at a dose of 6 mg/kg. At a dose of 12 mg/kg DA-125 induced moderate to remarkable changes in ECG like decrease of heart rate, widening of PR interval and 07 interval, and A-V block in 3 out of 5 animals. The severity of ECG alteration at 12 mg/kg of DA-125 was similar to that at 3mg/kg of DXR and these changes caused by DA-125 and DXR recovered within 10 minutes after injection. In chronic cardiotoxicity test model used rats, DA-125 was administered intravenously once a week for three weeks at a dose of 6, 9mg/kg and DXR was given at a dose of 6mg/kg. Electrocardiogram was recorded every week from the start of administration to 2 weeks after the last administration and the animals were sacrificed for histological heart examination at 1 week or 2 weeks after the last administration. DA-125 did not cause any abnormal changes in ECG and in histological heart examination due to administration, but DXR caused widening of ST segment, QRS complex, and QT interval from 1 week after administration and these changes were continued to necropsy. These alterations in ECG were accompanied by cardiac histological lesions such as vacuolation in myocardiac cells, interstitial edema and necrosis of myocytes. These results suggest that DA-125 is less cardiotoxic than DXR.

• The Korean Journal of Physiology and Pharmacology
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• 제5권5호
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• pp.367-371
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• 2001

The chromosome 7-linked long QT syndrome (LQT2) is caused by mutations in the human ether-a- go-go-related gene (HERG) that encodes the rapidly activating delayed rectifier $K^+$ current, $I_{Kr},$ in cardiac myocytes. Different types of mutations have been identified in various locations of HERG channel. One of the mechanisms for the loss of normal channel function is due to membrane trafficking of channel protein. The decreased channel function in some deletion mutants appears to be due to loss of coupling with wild type HERG to form the functional channel as the tetramer. Most of missense mutants with few exceptions could interact with wild type HERG to form functional tetramer and caused dominant negative suppression with co-injection with wild type HERG showing variable effects on current amplitude, voltage dependence, and kinetics of activation and inactivation. Two missense mutants at pore regions of HERG found in Japanese LQT2 (A614V and V630L) showed accentuated inward rectification due to a negative shift in steady-state inactivation and fast inactivation. One mutation in S4 region (R534C) produced a negative shift in current activation, indicating the S4 serving as the voltage sensor and accelerated deactivation. The C-terminus mutation, S818L, could not express the current by mutant alone and did not show dominant negative suppression with co-injection of equal amount of wild type cRNA. Co-injection of excess amount of mutant with wild type produced dominant negative suppression with a shift in voltage dependent activation. Therefore, multiple mechanisms are involved in different mutations and functional abnormality in LQT2. Further characterization with the interactions between various mutants in HERG and the regulatory subunits of the channels (MiRP1 and minK) is to be clarified.

• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 2003년도 정기총회 및 학술발표회
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• pp.29-29
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• 2003

Compartmentation of intracellular signaling pathways serves as an important mechanism conferring the specificity of G protein-coupled receptor (GPCR) signaling. In the heart, stimulation of $\beta$$_2$-adrenoceptor ($\beta$$_2$-AR), a prototypical GPCR, activates a tightly localized protein kinase A (PKA) signaling, which regulates substrates at cell surface membranes, bypassing cytosolic target proteins (eg, phospholamban). Although a concurrent activation of $\beta$$_2$-AR-coupled $G_{i}$ proteins has been implicated in the functional compartmentation of PKA signaling, the exact mechanism underlying the restriction of the $\beta$$_2$-AR-PKA pathway remains unclear. In the present study, we demonstrate that phosphatidylinositol 3-kinase (PI3K) plays an essential role in confining the $\beta$$_2$-AR-PKA signaling. Inhibition of PI3K with LY294002 or wortmannin enables $\beta$$_2$-AR-PKA signaling to reach intracellular substrates, as manifested by a robust increase in phosphorylation of phospholamban, and markedly enhances the receptor-mediated positive contractile and relaxant responses in cardiac myocytes. These potentiating effects of PI3K inhibitors are not accompanied by an increase in $\beta$$_2$-AR-induced cAMP formation. Blocking $G_{i}$ or $G_{$\square$$\square$}$ signaling with pertussis toxin or $\beta$ARK-ct, a peptide inhibitor of $G_{$\square$$\square$}$, completely prevents the potentiating effects induced by PI3K inhibition, indicating that the pathway responsible for the functional compartmentation of $\beta$$_2$-AR-PKA siglaling sequentially involves $G_{i}$, $G_{$\square$$\square$}$, and PI3K. Thus, PI3K constitutes a key downstream event of $\beta$$_2$-AR- $G_{i}$ signaling, which confines and negates the concurrent $\beta$$_2$-AR/Gs-mediated PKA signaling.gnaling.

• Archives of Pharmacal Research
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• 제28권8호
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• pp.930-935
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• 2005

We have investigated the effects of relatively high concentration of carbachol (CCh), an agonist of muscarinic acetylcholine receptor (mAChR), on cardiac automaticity in mouse heart. Action potentials from automatically beating right atria of mice were measured with conventional microelectrodes. When atria were treated with $100{\mu}M$ CCh, atrial beating was immediately arrested and diastolic membrane potential (DMP) was depolarized. After exposure of the atria to CCh for $\~4 min$, action potentials were regenerated. The regenerated action potentials had lower frequency and shorter duration when compared with the control. When atria were pre-exposed to pirenzepine $(1{\mu}M)$, an $M_1$ mAChR antagonist, there was complete inhibition of CCh-induced depolarization of DMP and regeneration of action potentials. Pre-exposure to AFDX-116 (11 ({2-[(diethylamino)-methyl]-1-piperidyl}acetyl)-5, 11-dihydro-6H-pyridol[2,3-b][1,4] benzodiazepine-6-one base, $1{\mu}M$), an $M_2$ mAChR antagonist, failed to block CCh-induced arrest of the beating. However, prolonged exposure to CCh elicited gradual depolarization of DMP and slight acceleration in beating rate. Our data indicate that high concentration of CCh depolarizes membrane potential and recovers right atrial automaticity via $M_1$ mAChR, providing functional evidence for the role of $M_1$ mAChR in the atrial myocytes.