• 대한수의학회지
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• 제39권5호
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• pp.878-895
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• 1999

Ischemic preconditioing (IPC), i.e., a preliminary brief episode of ischemia and reperfusion, has been shown to reduce the cell damage induced by long ischemia and reperfusion. Superoxide radical which is produced during reperfusion after ischemia was recognized as a factor of the ischemic injury and it is dismutated into $H_2O_2$ and $O_2$ by two types of intracellular superoxide dismutase (SOD), Cu,Zn-SOD in cytoplasm and Mn-SOD in mitochondria. Recently oxygen free radicals are suggested to induce the apoptosis, however mechanism of the reduced apoptosis by ischemic preconditioing was unknown, while many studies performed in mammalian heart indicated that ATP-sensitive $K^+$ ($K_{APT}$) channel activation related with the protective effects. The aim of present study is to investigate 1) whether IP upregulate the Cu,Zn-SOD and Mn-SOD activities, and 2) whether ischemic preconditioning decreases apoptosis via $K_{APT}$ channel activation in timely reperfused skeletal muscle after long ishemia. The experimental animals, Sprague-Dawley rats weighing 250~300g, were divided into 8 groups; 1) control group, 2) ischemic preconditioning only groups, 3) pinacidil, a $K_{APT}$ channel opener, treatment only groups, 4) glibenclamide, a $K_{APT}$ channel blocker, treatment only groups, 5) ischemia groups, 6) ischemia after IPC groups, 7) ischemia and pinacidil treatment groups, and 8) IP and ischemia after glibenclamide pretreatment groups. Animals of the control group were administered with the vehicle (DMSO) alone. Pinacidil (1mg/kg) was administered intravenously 5 minutes after initiation of ischemia, and glibenclamide (0.5mg/kg) was injected intravenously 20 minutes before IPC. In rats that were ischemic preconditioned, the left common iliac artery was occluded for 5 minutes followed by 5 minutes of reperfusion by three times using vascular clamp. Ischemia was done by occlusion of the same artery for 4 hours. The specimens of left rectus femoris muscle were obtained immediately (0 hour), 12 hours, 24 hours after drug administrations, IP or ischemia and reperfusion. The immunoreactivities of SOD and its alterations were observed by use of sheep antihuman Cu,Zn-SOD and Mn-SOD antibodies on the $10{\mu}m$ cryosections. The incidencies of apoptosis were observed by TUNEL methods with in situ apoptosis detection kit on $6{\mu}m$ paraffine section. The results obtained were as follows : 1. After IPC, immunoreactivities of Cu,Zn-SOD mainly in the small-sized fibers were increased by 24 hours, that of Mn-SOD at 0 hour and 24 hours. 2. No significant changes in immunoreactivities of SOD was observed in the pinacidil and in the glibenclamide treatment only groups, and in the ischemia only groups. 3. The immunoreactivities of the Cu,Zn-SOD were increased in the ischemia after IPC groups and the ischemia and pinacidil treatment groups. 4. The immunoreactivities of the Cu,Zn-SOD in the IPC and ischemia after glibenclamide pretreatment groups were not increased except for the 12 hours reperfusion group. But, Mn-SOD immunoreactivities were increased in the 0 hours, 12 hours and 24 hours after reperfusion. 5. In the control group, the IPC only groups, and the pinacidil treatment only groups, negative or trace apoptotic reactions were observed, but the positive apoptotic reaction occured in the glibenclamide treatment groups. 6. Moderate or many number of apoptosis were revealed in the ischemia groups, and also the IPC and ischemia after glibenclamide pretreatment group except for 12 hours and 24 hours after reperfusion. However, the incidence of apoptosis was decreased in the ischemia after IPC groups and in the ischemia and pinacidil treatment groups. 7. There is a coincidence between the increase of Cu,Zn-SOD immunoreactivities and the decrease of apoptosis in the presence of ischemia and reperfusion. These results suggest that the protective effects of ishemic preconditioing may related to the SOD activation, and the ischemic preconditioning decreases the apoptosis partially via $K_{APT}$ channel activation in timely reperfused rat skeletal muscle. It is also suggested that inhibition of apoptosis by IPC may related with the SOD activation.

• Journal of Ginseng Research
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• 제45권6호
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• pp.642-653
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• 2021

Background: Effective strategies are dramatically needed to prevent and improve the recovery from myocardial ischemia and reperfusion (I/R) injury. Direct interactions between the mitochondria and endoplasmic reticulum (ER) during heart diseases have been recently investigated. This study was designed to explore the cardioprotective effects of gypenoside XVII (GP-17) against I/R injury. The roles of ER stress, mitochondrial injury, and their crosstalk within I/R injury and in GP-17einduced cardioprotection are also explored. Methods: Cardiac contractility function was recorded in Langendorff-perfused rat hearts. The effects of GP-17 on mitochondrial function including mitochondrial permeability transition pore opening, reactive oxygen species production, and respiratory function were determined using fluorescence detection kits on mitochondria isolated from the rat hearts. H9c2 cardiomyocytes were used to explore the effects of GP-17 on hypoxia/reoxygenation. Results: We found that GP-17 inhibits myocardial apoptosis, reduces cardiac dysfunction, and improves contractile recovery in rat hearts. Our results also demonstrate that apoptosis induced by I/R is predominantly mediated by ER stress and associated with mitochondrial injury. Moreover, the cardioprotective effects of GP-17 are controlled by the PI3K/AKT and P38 signaling pathways. Conclusion: GP-17 inhibits I/R-induced mitochondrial injury by delaying the onset of ER stress through the PI3K/AKT and P38 signaling pathways.

• Journal of Chest Surgery
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• 제21권5호
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• pp.803-815
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• 1988

The present experiments were performed to confirm the hypothesis that xanthine oxidase[XOD], as a source and mechanism of oxygen radical production, plays an important role in the genesis of the reperfusion injury of ischemic myocardium. The experimental ischemic-reperfusion injury was induced in isolated, Langendorff preparations of rat hearts by 60 min. Of global ischemia with aortic clamping followed by 20 min. of reperfusion with oxygenated Krebs-Henseleit solution[pH 7.4, 37*C]. The results were as follows: 1. The releases of creatine phosphokinase and a lipid peroxidation product, malondialdehyde[MDA] into the coronary effluent were abruptly increased upon reperfusion of ischemic hearts. The increases of the enzyme and MDA were suppressed significantly in the hearts removed from rats pretreated with allopurinol, a specific XOD inhibitor[20mg/kg, oral, 24 hrs and 2 hrs before study]. This effect of allopurinol was comparable to that of oxygen radical scavengers, superoxide dismutase[5, 000U] and catalase[12, 500 U]. 2. The increased SOD-inhibitable reduction of ferricytochrome C, which was infused to the hearts starting with reperfusion, was significantly suppressed in allopurinol pretreated hearts. 3. Activities of myocardial XOD were compared in the normal control hearts and the ischemic ones. Total enzyme activities were not different in both hearts. However, comparing with the control, the ischemic ones showed higher activity in 0-form and lower activities in D-form and D/O-form. 4. In the ischemic hearts, phenylmethylsulfonyl fluoride, a serine protease inhibitor, prevented significantly the increase of 0-form and the decreases of D and D/O-form, while thiol reagents did not affect the changes of the enzyme. 5. The increase of 0-form and the decreases of D and D/0-form were not significant in both calcium-free perfused and pimozide, a calmodulin inhibitor, treated ischemic hearts. 6. The SOD-inhibitable reduction of ferricytochrome C were suppressed by PMSF and pimozide treatment as well as by calcium-free perfusion. It is suggested from these results that in the ischemic rat myocardium, xanthine oxidase is converted to oxygen radical producing 0-form by calcium, calmodulin-dependent proteolysis and plays a contributing role in the genesis of ischemic-reperfusion injury by producing oxygen free radicals.

• Journal of Chest Surgery
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• 제32권11호
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• pp.971-977
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• 1999

Background: Immunologic and inflammatory responses of cardiopulmonary bypass(CPB) influence postoperative mortality and morbidity with multiple organ injury. It has been reported that ischemia/reperfusion induced-myocardial injury during CPB is causative of release of inflammatory cytokines such as interleukin-6(IL-6) and tumor necrosis factor-$\alpha$ (TNF-$\alpha$). The purpose of this study was to detect the time course of the activated cytokine and troponin-T(TnT), and to examine the correlation between such parameters during CPB. Material and Method: The serial samples were collected from arterial blood via radial arterial catheter in 23 patients who are underwent open heart surgery (OHS) with CPB, the IL-6, TNF-$\alpha$ and TnT were checked. Result: \circled1 IL-6, TNF$\alpha$- and TnT concentration increased significantly during CPB with a peaking level of CPB-off (p 0.05). \circled2 IL-6 had highly positive correlation with aortic cross clamping time and total bypass time(r=0.80, 0.78; p 0.05, respectively). \circled3 There was no correlation among IL-6, TNF-$\alpha$ and TnT. Conclusion: In conclusion, these data showed that elevated production of serum IL-6 during CPB was attributable to ischemia/reperfusion induced-myocardial damage. IL-6 will become a new and sensitive biological marker in assessment of myocardial damage during OHS with CPB. However, further studies will be needed to apply IL-6 in more patient population.

• 대한약리학회지
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• 제30권3호
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• pp.321-330
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• 1994

허혈전처치(ischemic preconditioning)의 재관류심근손상 보호작용과 그 기전을 규명하기 위한 연구의 일환으로 허혈전처치가 심근세포의 산소라디칼 생성능력에 미치는 영향을 검토하였다. 흰쥐 적출심장의 Langendorff 관류표본에서 실험적인 허혈(30분)-재관류(20분)손상을 유도하였고, 허혈전처치는 재관류손상 유도전에 5분 허혈-5분 재관류를 3회 반복하여 시행하였다. 허혈심근 재관류손상의 지표로 심수축기능, 세포질효소 유출, 칼슘 유입 및 미세형태학적 변화를, 그리고 심근세포의 산소라디칼 생성기전으로 xanthine oxidase system의 변동을 허혈전처치와 비전처치 재관류 심장들에서 비교검토하였다. 연구 성적은 다음과 같다. 1. 허혈전처치는 허혈-재관류 심장의 관상혈류량, 심박수, 좌심실압 등 심기능의 저하를 현저히 회복시켰다(회복률; 91%) 2. 허혈-재관류 심장에서 lactate dehydrogenase 유출증가는 허혈전처치에 의해 현저히 저하되었다. 3. 허혈-재관류 심근세포의 전자현미경상 미세구조는 허혈전처치시 비교적 잘 보존되었으며, 특히 myofibril 구조의 보존이 매우 뚜렷하였다. 4. 허혈-재관류시 산화성 조직손상 척도의 하나인 malondialdehyde 생성이 허혈전처치에 의하여 현저히 감소되었다. 5. 허혈전처치 심장에서 xanthine oxidase(D,O 및 D/O형)활성은 변화되지 않았으나 그 기질인 hypoxanthine의 조직함량은 현저히 감소되었다. 이상의 결과들로 부터 허혈전처치는 세포수준에서 허혈심근의 재관류손상을 방지하며, 허혈전처치에 따른 산소라디칼 생성 감소가 재관류손상 방지에 일부 기여할 수 있으리라 사료된다.an을 2주간 처치한 경우에도 영향을 받지 않았다. 이상의 결과로 미루어 아마도 losartan의 내피세포에 대한 작용은 constitutive NO 생성을 증가시키나 inducible NO 생성에는 영향을 미치지 않을 것으로 여겨진다.cium ion이 상당히 중요한 역할을 하는데, 특히 소뇌에서의 NO생성의 감소는 이들 약물들의 치명적 부작용인 tardive dyskinesia와 매우 깊은 관련을 추측할 수 있다. 그러나, 더 많은 약물들의 검색으로 일관적인 기본 결과가 필요 되고 또 각개 약물의 특정적 기전이 연구되기 위하여 현재 실험중이다.(신칭(新稱), Cystostereum subabruptum), 털융단버섯(신칭(新稱), Tomentella pilosa), 노란소나무무늬버섯(신칭(新稱), Asterostroma laxum), 붉은소나무비늘버섯(신칭(新稱), Hymenochaete cruenta), 가루소나무비늘버섯(신칭(新稱), Hymenochaete fuliginosa), 소나무비늘버섯(신칭(新稱), Hymenochaete tabacina), 갈색시루삔버섯(신칭(新稱), Inonotus radiatus), 벚나무진흙버섯(신칭(新稱), Phellinus pomaceus), 회주름구멍버섯(신칭(新稱), Antrodia crassa), 층주름구멍버섯(신칭(新稱), Antrodia serialis), 흰그물구멍버섯(신칭(新稱), Ceriporia reticulata), 겹친손등버섯(신칭(新稱), Oligoporus balsameus), 점박이손등버섯(신칭(新稱), Oligoporus guttulatus), 무른흰살버섯(신칭(新稱), Oxyporus cuneatus), 각목버섯(신칭(新稱),

• Toxicological Research
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• 제4권1호
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• pp.23-35
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• 1988

The role of calcium in the production of oxygen radical which causes reperfusion damage of ischemic heart has been examined. The reperfusion damage was indrced in isolated Langendorff perfused rat hearts by aortic clamping for 60 min followed by reperfusion with oxygenated Krebs-Henseleit solution with or without 1.25 mM $CaCl_2.$ On reperfusion of the ischemic hearts with the calcium containing solution, the release of cytosolic enzymes (LDH and CPK) increased abruptly. These increased release of enzymes were significantly inhibited by additions of oxygen radical scavengers (SOD, 5,000 U; catalase, 12,500 U) into the reperfusion solution. In the hearts isolated from rats pretreated with allopurinol(20 mg/kg orally, 24 hr and 2 hr prior to the experiments), the levels of enzymes being released during reperfusion were significantly lower than that of the control. However, in the hearts perfused with the calcium-free but oxygenated solution, the increase in the release of cytosolic enzymes during reperfusion was neither inhibited by oxygen radical scavengers nor by allopurinol pretreatment. For providing the evidence of oxygen radical generation during the reperfusion of ischemic hearts in situ, the SOD-inhibitable reduction of exogenously administered ferricytochrome C was measured. In the hearts perfused with the calcium containing solution, the SOD-inhibitable ferricytochrome C reduction increased within the first minute of reperfusion, and was almost completely inhibited by allopurinol pretreatment. When the heart was perfused with the calcium free solution, however, the reduction of ferricytochrome C was not only less than that in the calcium containing condition, but also was not so completely inhibited by allopurinol pretreatment. By ischemia, xanthine oxidase (XOD) in the ventricular tissue was changed qualitatively, but not quantitatively. In the heart made ischemic with the calcium containing condition, the oxygen radical producing O-form of XOD increased, while the D- and D/O-form decreased. However, in the ischemic heart reperfused with the calcium free condition, the D/O-form of XOD was elevated without significant increase in O-form of the enzyme. It is suggested from these results that the calclum may play a contributing role in the genesis of reperfusion damage by promoting the conversion of xanthine oxidase from the D/O-form to the oxygen radical producing O-form in the ischemic myocardium.

• Journal of Chest Surgery
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• 제29권6호
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• pp.593-598
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• 1996

과거에 심근보호에 대한 많은 조사는 저온의 고칼릅 심정지액 및 국소적 냉각에 의한 방법으로는한 계가 있다는 지적이 있었다. 따라서 최근의 실험들은 재관류의 방법에 따른 허혈 후 심근회복에 대하여 촛점이 맞춰 지고 있다. 재관류 시 산소에 의한 심근손상이 밝혀 짐으로써 oxygen free radical scavenger에 대한 관심이 높아지고 있다. 따라서 본 교실에서는 쥐에게 항산화제로서 비타민 C를 먹인 후 Langendorf'r system을 이용하여 허혈 및 재관류 시 좌심실 기능의 변화를 관찰하였다 대상은 체중 190-))Og의 Sprague-Dawley쥐를 암수 구별없이 사용하였다. 편의상 비타민을 먹이지 않 은 대조군을 Group A (n=10)라 하였고 200mg의 비타민을 먹인 실험군을 Group R (n=10)라 하였다. 실 험군의 경우는 비타민 C 200mg을 경구투여한 후 24시간에 시행하였다. 방법은 언저 복강을 통해 헤파 린과 펜토탈을 주입한 후 심장을 적출하여 Langendorff system에 거치하고 비운동성 역관류를 시켰다. 관류액은 변형된 Krebs-Henseleit solution을 사용하였다. 좌심실내에 풍선을 삽입하여 polygraph를 통해 좌심실의 혈역학적 기능을 관찰하였다. 먼저 20분간 심 揚\ulcorner안정될 때까지 기다린 다음 51. Thomas심 정 지액으로 심정 지를 시킨후 30분간 허 혈시키고 다시 20분간 재관류시켰다. 각 Group에서 허혈전후의 좌심실압의 비, dp/dt의 비,박동수의 비, 재관류 후 첫 박동 및 안정될 때 까지의 시간을Group간에 비 교하였다. 좌심실압의 비는 Group A가 평균 88.9%, Group B가 114%로 실험군이 의의있게 높았으며 역시 dp/dt도 Group A가 89.6%, Group B가 1)2.9%로 실험군이 의의 있게 높았다 그러나 박동수의 비, 재관류 후 첫 박동 및 안정될 때 까지의 시간의 비교는 통계 학적 의의가 없었다. 결론적으로 항산화제로서 비타민 C는 허혈 및 재관류 시 좌심실 기능회복에 도움이 되었다. 그러나 향후 임상적 적용을 위해서는 더욱 자세하고 많은 실험 이 요구된다.

• The Korean Journal of Physiology and Pharmacology
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• 제25권4호
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• pp.281-296
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• 2021

The beneficial effects of hypoxic preconditioning are abolished in the diabetes. The present study was designed to investigate the protective effects and mechanisms of repeated episodes of whole body hypoxic preconditioning (WBHP) in db/db mice. The protective effects of preconditioning were explored on diabetes-induced vascular dysfunction, cognitive impairment and ischemia-reperfusion (IR)-induced increase in myocardial injury. Sixteen-week old db/db (diabetic) and C57BL/6 (non-diabetic) mice were employed. There was a significant impairment in cognitive function (Morris Water Maze test), endothelial function (acetylcholine-induced relaxation in aortic rings) and a significant increase in IR-induced heart injury (Langendorff apparatus) in db/db mice. WBHP stimulus was given by exposing mice to four alternate cycles of low (8%) and normal air O₂ for 10 min each. A single episode of WBHP failed to produce protection; however, two and three episodes of WBHP significantly produced beneficial effects on the heart, brain and blood vessels. There was a significant increase in the levels of brain-derived neurotrophic factor (BDNF) and nitric oxide (NO) in response to 3 episodes of WBHP. Moreover, pretreatment with the BDNF receptor, TrkB antagonist (ANA-12) and NO synthase inhibitor (L-NAME) attenuated the protective effects imparted by three episodes of WBHP. These pharmacological agents abolished WBHP-induced restoration of p-GSK-3β/GSK-3β ratio and Nrf2 levels in IR-subjected hearts. It is concluded that repeated episodes of WHBP attenuate cognitive impairment, vascular dysfunction and enhancement in IR-induced myocardial injury in diabetic mice be due to increase in NO and BDNF levels that may eventually activate GSK-3β and Nrf2 signaling pathway to confer protection.

• The Korean Journal of Physiology and Pharmacology
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• 제25권3호
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• pp.239-249
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• 2021

The present study explored the therapeutic potential of hydrogen sulfide (H2S) in restoring aging-induced loss of cardioprotective effect of remote ischemic preconditioning (RIPC) along with the involvement of signaling pathways. The left hind limb was subjected to four short cycles of ischemia and reperfusion (IR) in young and aged male rats to induce RIPC. The hearts were subjected to IR injury on the Langendorff apparatus after 24 h of RIPC. The measurement of lactate dehydrogenase, creatine kinase and cardiac troponin served to assess the myocardial injury. The levels of H2S, cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), nuclear factor erythroid 2-related factor 2 (Nrf2), and hypoxia-inducible factor (HIF-1α) were also measured. There was a decrease in cardioprotection in RIPC-subjected old rats in comparison to young rats along with a reduction in the myocardial levels of H2S, CBS, CSE, HIF-1α, and nuclear: cytoplasmic Nrf2 ratio. Supplementation with sodium hydrogen sulfide (NaHS, an H2S donor) and l-cysteine (H2S precursor) restored the cardioprotective actions of RIPC in old hearts. It increased the levels of H2S, HIF-1α, and Nrf2 ratio without affecting CBS and CSE. YC-1 (HIF-1α antagonist) abolished the effects of NaHS and l-cysteine in RIPC-subjected old rats by decreasing the Nrf2 ratio and HIF-1α levels, without altering H2S. The late phase of cardioprotection of RIPC involves an increase in the activity of H2S biosynthetic enzymes, which increases the levels of H2S to upregulate HIF-1α and Nrf2. H2S has the potential to restore aging-induced loss of cardioprotective effects of RIPC by upregulating HIF-1α/Nrf2 signaling.

• 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.