生理学报Acta Physiologica Sinica, April 25, 2003, 55(2): 171-176
Received 2002-07-02
Accepted 2002-10-17
This study was supported by grants from the Major State Basic Research Development Program of the People′s Republic of China (No.G2000056905) and the National Natural Science Foundation of China (No. 30070306).
*Corresponding author. Tel: +86-21-25074367; E-mail: yuanwj@smmu.edu.cn
研究论文
内皮素-1预处理对培养乳鼠心肌细胞低氧损伤的保护作用
潘燕霞1,2， 林丽1， 袁文俊1，*, 吴国宏1, 唐朝枢3
1第二军医大学生理学教研室, 上海 200433; 2福建医科大学生理学教研室, 福州 350004;
3北京大学医学部生理和病理生理学系， 北京 100083
摘要: 实验观察了0.01-1 nmol/L内皮素-1 (ET-1)预处理对低氧孵育(3% O2-5% CO2 , 12 h) 的培养乳鼠心肌细胞乳酸脱氢酶(LDH)释放量、 培养液上清超氧化物歧化酶(SOD)活性以及丙二醛(MDA)含量的影响。用Fluo-3/AM负载培养的心肌细胞, 在激光扫描共聚焦显微镜下监测急性低氧的心肌细胞[Ca2+]i的变化和ET-1预处理对低氧所致[Ca2+]i变化的影响。结果如下: (1) 心肌细胞低氧孵育12 h后, 培养液上清LDH活力和MDA含量较常氧对照组明显升高, 分别为43.33±1.21 U/L vs 19.33±1.03 U/L和1.71±0.02 nmol/L vs 0.91±0.03 nmol/L (P<0.01), SOD活性为16.93±1.11 U/ml明显低于常氧对照组的33.48±1.15 U/ml (P<0.01); 0.01-1 nmol/L ET-1预处理呈浓度依赖性抑制低氧培养心肌细胞LDH释放, 减少培养液上清MDA含量、 提高SOD活性(P<0.01)。(2) 低氧灌流后29±1.5 s (n=23)心肌细胞自发性钙瞬变完全终止, [Ca2+]i升高了107±13.2% (P<0.001); 0.01-1 nmol/L ET-1能明显加快心肌细胞钙瞬变的频率(P<0.01); ET-1预处理后低氧所致钙瞬变终止的时间较单纯低氧组明显推迟, [Ca2+]i 过度升高被明显减轻 (P<0.01)。上述结果表明, 0.01-1 nmol/L ET-1预处理可减轻培养乳鼠心肌细胞的低氧损伤和抑制低氧所致[Ca2+]i的变化, 具有一定的细胞保护作用。
关键词: 内皮素; 低氧; 心肌细胞; 细胞内钙
中图分类号: Q463; R331.31
Preventive effect
of endothelin-1 pretreatment on hypoxia-induced injury in cultured neonatal rat
cardiomyocytes
PAN Yan-Xia1,2, LIN Li1, YUAN Wen-Jun1，*, WU Guo-Hong1, TANG Chao-Shu3
1Department of Physiology, Second Military Medical University, Shanghai 200433;
2Department of Physiology, Fujian Medical University, Fuzhou 350004;
3Department of Physiology and Pathophysiology, Health Sciences Center, Peking University, Beijing 100083
Abstract: This study was designed to observe the effects of endothelin-1 (ET-1) pretreatment on hypoxia-induced injury and changes in [Ca2+]i in cultured neonatal rat cardiomyocytes. The activity of lactate dehydrogenase (LDH) and superoxide dismutase (SOD), and the content of malondialdehyde (MDA) in the supernatant were determined in the cultured cardiomyocytes subjected to a 12-h hypoxia induced by a 3% O2-5% CO2 atmosphere at 37℃ with or without ET-1 pretreatment. [Ca2+]i was measured with Ca2+-sensitive fluorescent probe fluo-3/AM under a laser scanning confocal microscope. Fluorescence intensity emitted from fluo-3/AM-loaded cells reflected the concentration of [Ca2+]i. The hypoxia model used in [Ca2+]i measurement was established by continously perfusing cardiomyocytes for 30 min with 95% N2-5% CO2 saturated DMEM solution containing 1 mmol/L Na2S2O4. Pretreatment with ET-1 consisted of three cycles of ET-1 perfusion (5 min for each) followed by ET-1-free DMEM solution (10 min for each) prior to hypoxia. The results showed that (1) after incubation in a 3% O2-5% CO2 hypoxic atmosphere for 12 h, the activity of LDH and the content of MDA in the supernatant significantly increased from 19.33±1.03 U/L to 43.33±1.21 U/L and from 0.91±0.03 nmol/L to 1.71±0.02 nmol/L, respectively, whereas the activity of SOD decreased from 33.48±1.15 U/ml to 16.93±1.11 U/ml (P<0.01). In hypoxic cardiomyocytes pretreated with 0.01-1 nmol/L ET-1, LDH release and supernatant MDA content were decreased, while SOD activity was enhanced dose-dependently (P<0.01). (2) The spontaneous calcium transient in cultured cardiomyocytes terminated at 29±1.5 s and [Ca2+]i increased by 107±13.2% during perfusion of hypoxic solution (P<0.001) at the end of 30 min. ET-1 (0.01-1 nmol/L) increased the frequency of [Ca2+]i transient in cultured cardiomyocytes in a dose-dependent manner (P<0.01). The termination of [Ca2+]i transient and the elevation of [Ca2+]i caused by hypoxia were postponed by pretreatment with 0.01-1 nmol/L ET-1 (P<0.01). These results show that pretreatment with 0.01-1 nmol/L ET-1 attenuates hypoxia-induced injury and [Ca2+]i changes in cultured neonatal cardiomyocytes, indicating a cyto-protective role of ET-1 pretreatment.
Key words: ET-1; hypoxia; cardiomyocytes; intracellular calcium
内皮素-1(endothelin-1, ET-1)是主要分布于心血管系统的缩血管肽, 外源性ET-1可以引起心肌缺血性损伤和心律失常[1,2 ], 内源性ET-1在心肌缺血/ 缺氧时合成与释放增加, ET-1单抗、 ETA受体拮抗剂或ETA/ETB受体拮抗剂及ET-1反义寡核苷酸可有效缩小心肌梗死面积, 促进心功能的恢复, 减轻心肌缺血/再灌注损伤和缺血性心律失常[3-5], 表明ET-1在心肌缺血或缺血/再灌注损伤病理过程中具有重要作用。ET-1预处理对心肌缺血或缺血/再灌注损伤是加重抑或保护作用尚有争议, Bugge[6] 等报道, 离体心脏心肌缺血前短暂的ET-1灌流能减小心肌梗死面积, 非选择性ET受体拮抗剂Bosentan和ETA特异性受体拮抗剂BQ123可阻断这一作用。Wang[7]和Hide[8]认为ET-1预处理也可模拟缺血预适应产生保护效应。但Mark[9]报道, 离体灌流的心脏ET-1预处理反而加重心肌缺血/再灌注损伤。在整体或离体水平, ET-1预处理对心脏的保护作用尚不能区分是ET-1的直接效应或继发于ET-1缩血管产生的缺血预适应。因此, 本研究旨在探讨ET-1预处理对培养心肌细胞低氧损伤和低氧时[Ca2+]i变化的影响。
1 材料和方法
1.1 主要仪器、 试剂与材料
CO2孵箱(Forma -3131, Sci. Inc.)、 激光共聚焦显微镜(LSM510, Zeiss Inc.)、全自动生化分析仪(Hitachi 7150, Japan)、 紫外分光光度计(Ultrospec 2000, Pharmacia Biotech Inc.)、 DMEM (Gibco)、 BrdU、 Fluo-3/AM (Sigma)、 ET-1 (Peptide Institute, Osaka, Japan)、 MDA试剂盒(南京建成生物工程公司)和SD乳鼠(上海西普尔-毕凯公司)。
1.2 心肌细胞原代培养
取1-3日龄Sprague-Dawley大鼠心室肌, 0.2% 胰酶消化分离心肌细胞, 以含10%小牛血清和10%胎牛血清的DMEM培养液制成密度为3×105-4×105/ml的单细胞悬液, 接种于24孔培养板以测定乳酸脱氢酶(lactate dehydrogenase, LDH)、 超氧化物歧化酶(superoxide dismutase, SOD)和脂质过氧化产物丙二醛(malondialdehyde, MDA), 或接种于盖玻片上以测定胞浆钙离子浓度。置于5% CO2孵箱37℃孵育, 24 h后换液并加入0.1 mmol/L 5-溴-2′脱氧尿苷(5-Bromo-2′-deoxyuridine, BrdU)以抑制成纤维细胞生长。各项实验前24 h换成无血清DMEM培养液, 实验用培养3-4 d的细胞。
1.3 实验分组
实验分为常氧对照组、 低氧对照组和ET-1预处理＋低氧组。ET-1 预处理浓度分别为 0.01、 0.1、 1 nmol/L。
1.4 ET-1预处理对低氧心肌细胞培养液上清LDH、 SOD和MDA含量的影响
0.01-1 nmol/L ET-1预先孵育心肌细胞10 min, 换正常DMEM溶液孵育10 min, 再换以正常DMEM溶液在3% O2-5% CO2低氧环境(CO2孵箱控制)中孵育12 h, 测定细胞上清液LDH、 SOD和MDA。低氧对照组以DMEM替代ET-1进行预处理; 常氧对照组换以正常DMEM溶液后在含5% CO2 的常氧环境中孵育12 h。
采用全自动生化分析仪测定培养液上清LDH, 微量邻苯三酚自氧化法测定SOD活性[10], 硫代巴比妥酸比色法测定MDA含量(MDA试剂盒)。
1.5 ET-1预处理对低氧所致心肌细胞[Ca2+]i 变化的影响
采用钙敏感荧光探针Fluo-3/AM结合激光扫描共聚焦显微镜测定胞浆游离钙[11]。将生长有心肌细胞的盖玻片移入特制的小室中, 加入终浓度为5 μmol/L 的Fluo-3/AM 37℃避光负载细胞30 min, 然后用DMEM清洗, 除去细胞外Fluo-3/AM。细胞负载Fluo-3/AM后, 将小室置于共聚焦显微镜恒温载物台上, 与灌流系统相接, 用37℃ DMEM溶液恒速(1 ml/min)灌流以维持细胞功能状态。激光共聚焦显微镜采用激发光波长λ＝488 nm, 发射光波长λ＝510 nm, 每200 ms采样1 次, 其中采样时间为196 ms, 间隔时间4 ms, 连续记录给药或低氧前后同一视野细胞内钙荧光强度的动态变化, 该荧光强度的强弱即反映胞浆游离钙的相对浓度。
以95% N2-5% CO2饱和的含1 mmol/L Na2S2O4的DMEM低氧溶液(氧分压为1-2 mmHg)[12], 经灌流系统持续灌入放置细胞的小室(小室内液体氧分压为10-15 mmHg)制备心肌细胞低氧灌流模型。低氧之前以ET-1(0.01-1 nmol/L)灌流细胞5 min, 然后换以正常DMEM灌流10 min, 重复3次, 连续监测ET-1处理前后同一视野中的心肌细胞[Ca2+]i 动态变化, 并观察ET-1预处理对低氧心肌细胞[Ca2+]i变化的影响。低氧对照组以DMEM替代ET-1进行预处理; 常氧对照组以正常DMEM 溶液模拟ET-1预处理, 按相同程序灌流细胞。
1.6 统计学处理
数据均以mean±SD表示, 方差齐性的多组数据采用方差分析, 方差不齐的数据则采用秩和检验, P<0.05 表示差异有统计学意义。
钙荧光强度变化的百分数(%)=(Fmax-F0)/ F0 ×100% Fmax为低氧后荧光强度的峰值, F0为低氧前钙荧光强度。
2结果
2.1 ET-1预处理对低氧心肌细胞培养液上清LDH、 SOD和MDA含量的影响
心肌细胞经3% O2-5% CO2低氧孵育12 h后, 培养液上清 LDH活性和MDA含量较常氧对照组明显升高, 分别为43.33±1.21 U/L vs 19.33±1.03 U/L和1.71±0.02 nmol/L vs 0.91±0.03 nmol/L (P<0.01); SOD活性则显著低于常氧对照组(16.93±1.11 U/ml vs 33.48±1.15 U/ml, P<0.01)。0.01-1 nmol/L ET-1预处理呈浓度依赖性降低低氧孵育心肌细胞的培养液上清LDH和MDA含量, 提高SOD活性(表1), 提示ET-1预处理能增强心肌细胞SOD活性, 减少脂质过氧化物的生成, 提高心肌细胞抗氧化能力。
表1. ET-1预处理对低氧心肌细胞培养液上清LDH、 SOD 活性和MDA含量变化的影响
Table 1. Effects of ET-1 pretreatment on hypoxia-induced changes in supernatant LDH, MDA and SOD in cultured neonatal rat cardiomyocytes (mean±SD)
|
Group |
n |
LDH (U/L) |
SOD (U/ml) |
MDA (nmol/L) |
|
Normal control |
6 |
19.33±1.03 |
33.48±1.15 |
0.91±0.03 |
|
Hypoxia control |
6 |
43.33±1.21** |
16.93±1.11** |
1.71±0.02** |
|
0.01 nmol/L ET-1+hypoxia |
6 |
34.00±1.21**## |
22.67±1.33**## |
1.33±0.04**## |
|
0.1 nmol/L ET-1+hypoxia |
6 |
28.67±1.03**## |
27.37±1.21**## |
1.16±0.02**## |
|
1 nmol/L ET-1+hypoxia |
6 |
25.50±1.05**## |
24.83±0.89**## |
1.22±0.04**## |
** P<0.01 vs normal control; ##P<0.01 vs hypoxia control.
2.2 正常心肌细胞负载Fluo-3后[Ca2+]i 的变化
培养的乳鼠心肌细胞在倒置相差显微镜下可观察到心肌细胞的自发性、 节律性搏动。钙敏感荧光探针Fluo-3/AM负载心肌细胞后, 在激光共聚焦显微镜下可见心肌细胞内钙荧光强度呈自发性、 瞬时性和周期性升高, 即为钙瞬变(calcium transient), 其节律与细胞的自发搏动同步(图1)。
2.3 急性低氧心肌细胞[Ca2+]i 的变化
培养心肌细胞经灌流低氧液造成急性低氧, 在低氧后29±1.5 s (n=23), 心肌细胞钙瞬变完全停止, 此后1-3 min内舒张期末[Ca2+]i先短暂轻微下降, 继而[Ca2+]i持续上升(图2A)。低氧灌流30 min 后, [Ca2+]i 升高了107±13.2% (n=12)。
图1.培养的新生大鼠心肌细胞自发性钙瞬变波形
Fig. 1.A representative trace showing spontaneous transient of [Ca2+]i in cultured neonatal rat cardiomyocytes loaded with Fluo-3.
2.4 ET-1引起的心肌细胞[Ca2+]i 变化
0.01-1 nmol/L ET-1灌流心肌细胞, [Ca2+]i 瞬变频率均明显加快, ET-1重复3次灌流, 每次间隔以正常DMEM溶液灌流10 min, [Ca2+]i 瞬变的频率仍高于给药前(表2)。ET-1 (0.1、 0.01 nmol/L)预处理对心肌细胞收缩期末和舒张期末[Ca2+]i 均无明显影响, [Ca2+]i瞬变幅度也无明显变化; 而1 nmol/L ET-1预处理后, 收缩期末和舒张期末[Ca2+]i 均升高, 舒张期末[Ca2+]i的增加大于收缩期末, 钙瞬变幅度减小(表3)。单纯低氧组和ET-1预处理后低氧组钙瞬变消失前的舒张期末[Ca2+]i 之间无显著差异。对照组采用正常DMEM 溶液替代ET-1, 按相同程序灌流3 个循环, 心肌细胞[Ca2+]i 瞬变幅度轻度减小, 但无统计学意义(表3), [Ca2+]i 瞬变频率无明显变化(表2)。
表2. ET-1 预处理对培养心肌细胞(低氧前)胞浆钙瞬变频率的影响
Table 2. Effects of repeated ET-1 administration on the frequency of calcium transient in pre-hypoxia cultured neonatal rat cardiomyocytes (mean±SD)
|
Group |
Frequency of [Ca2+]i transient (spikes/min) |
|||
|
Prior to ET-1 |
ET-11 |
ET-12 |
ET-13 |
|
|
DMEM (n=12) |
24.3±3.0 |
23.0±1.5 |
21.2±2.5 |
21.3±1.0 |
|
0.01 nmol/L ET-1 (n=12) |
24.8±1.0 |
34.7±2.0**## |
33.8±2.0**## |
32.0±1.7**## |
|
0.1 nmol/L ET-1 (n=15) |
24.7±2.1 |
42.6±2.3**## |
42.6±2.3**## |
38.0±2.9**## |
|
1 nmol/L ET-1 (n=12) |
25.5±1.9 |
57.5±3.1**## |
53.0±1.8**## |
48.8±2.6**## |
**P<0.01 vs prior to ET-1; ##P<0.01 vs DMEM. ET-11: perfusing ET-1 for 5 min the first time; ET-12: perfusing ET-1 for 5 min the second time; ET-13: perfusing ET-1 for 5 min the third time. There was a 10-min washout with normal DMEM solution between ET-1 administrations.
2.5 ET-1预处理对心肌细胞急性低氧[Ca2+]i 变化的影响
0.01、 0.1和1 nmol/L ET-1预处理均可明显推迟低氧引起的[Ca2+]i 瞬变终止, 低氧5-8 min后, [Ca2+]i开始上升。0.1 nmol/L ET-1预处理后, 低氧引起心肌细胞 [Ca2+]i 瞬变的终止时间推迟到低氧后60 s左右, 随后舒张期末[Ca2+]i先轻微下降而后恢复到低氧前水平, 低氧5 min, 舒张期末[Ca2+]i 未见明显上升(图2B)。 在0.01、 0.1和1 nmol/L ET-1预处理组, 低氧引起的[Ca2+]i 瞬变终止时间分别为51±1.0 s (n=12)、 58±1.3 s (n=26)和43±2.6 s (n=23), 较单纯低氧组显著推迟(P<0.001)。
表3. ET-1 预处理对培养心肌细胞(低氧前)胞浆钙瞬变的影响
Table 3. Effects of repeated ET-1 administration on the calcium transient in pre-hypoxia cultured neonatal rat cardiomyocytes (mean±SD)
|
Group |
Prior to ET-1 |
After ET-1 |
||||
|
Fmax |
Fmin |
Amp |
Fmax |
Fmin |
Amp |
|
|
DMEM (n=12) |
140.7±12.4 |
52.9±11.6 |
88.7±12.2 |
137.5±11.3 |
55.0±6.7 |
82.5±9.6 |
|
0.01 nmol/L ET-1(n=12) |
138.0±9.3 |
51.3±4.8 |
87.6±9.6 |
137.5±7.7 |
57.5±9.6 |
80.4±13.1 |
|
0.1 nmol/L ET-1(n=15) |
141.9±12.1 |
53.0±9.3 |
88.5±11.4 |
144.0±11.8 |
60.3±10.0 |
83.8±10.4 |
|
1 nmol/L ET-1(n=12) |
144.2±8.1 |
53.3±8.7 |
90.8±9.0 |
148.8±9.2 |
67.9±9.3* |
80.0±7.7* |
*P<0.05 vs pre-ET-1; Fmax, the systolic value of Ca2+ fluorescence intensity; Fmin, the diastolic value of Ca2+ fluorescence intensity; Amp, amplitude is the difference between the systolic and diastolic values.
低氧30 min后, 0.01、 0.1和1 nmol/L ET-1预处理组心肌细胞[Ca2+]i分别升高了60±7.6%、 58±6.8% 和70.9±6.0%, 均显著低于单纯低氧组(P<0.001)(图3)。上述结果表明, 0.01-1 nmol/L ET-1预处理可有效推迟低氧所致[Ca2+]i 瞬变终止的发生, 并延缓[Ca2+]i升高 。
图2. 新生大鼠培养心肌细胞低氧时[Ca2+]i的变化
Fig. 2.Effects of hypoxia on [Ca2+]i in cultured neonatal rat cardiomyocytes. A: Pretreatment with normal DMEM solution. B: Pretreatment with 0.1 nmol/L ET-1. Arrows indicate the onset of hypoxia.
图3.ET-1 预处理对低氧所致培养心肌细胞[Ca2+]i升高的抑制效应
Fig. 3.Suppressive effects of ET-1 pretreatment on [Ca2+]i elevation caused by 30-min hypoxia in cultured neonatal rat cardiomyocytes. Values are expressed as an elevation percentage of the baseline [Ca2+]i (pre-hypoxia). n=12 in each group. *** P<0.001 vs 0 concentration of ET-1.
3讨论
心肌缺血/缺氧时,
ET-1合成与释放增加, ET-1可引起冠状动脉收缩, 加重心肌缺血性损伤并参与诱发缺血性心律失常[3,4,13]; 抑制ET-1作用可减轻心肌缺血性损伤和缺血性心律失常[4,5,14,15],
提示ET-1在心肌缺血或缺血/再灌注损伤病理过程中具有重要作用。对ET-1 预处理的心肌保护效应目前仍存在争议。 在整体[16]或离体灌流心脏[6,7],
ET-1预处理可减小缺血/再灌注心脏心肌梗死面积、 减少缺血性心律失常[16], 抑制PKC活性或阻断KATP通道则取消ET-1预处理的保护作用[6,7], 提示ET-1预处理可模拟心肌缺血预适应产生心脏保护效应,
其保护机制与激活PKC和KATP通道有关。但Mark等[9]报道, ET-1预处理可加重离体灌流的心脏的缺血/再灌注损伤。ETA受体拮抗剂可阻断ET-1预处理的保护作用,
但不能取消缺血预适应的保护作用[6,17] 。
预适应是机体的一种强而有效的内源性保护机制。药物预适应能否产生保护效应与预处理方式、
动物种属、 实验模型密切相关。由于ET-1强烈的缩血管作用和对心肌的原发性致缺血效应(proischemic effect)[3,9],
在整体或离体心脏, ET-1预处理的保护作用尚不能区分是ET-1的直接效应抑或是继发于ET-1缩血管产生的缺血预适应, 还是ET-1引起心肌释放其他内源性保护物质。本工作选用培养心肌细胞模型,
排除整体或离体心脏实验中ET-1收缩血管等因素的影响, 在细胞水平观察ET-1预处理对心肌细胞低氧损伤的直接影响。结果表明, 0.01-1 nmol/L
ET-1预处理可有效抑制低氧心肌细胞LDH释放的增加, 提示ET-1预处理可减轻心肌细胞的低氧损伤; 低氧还可引起培养心肌细胞脂质过氧化产物MDA含量升高、
SOD活性降低, 而0.01-1 nmol/L ET-1预处理可减少MDA含量, 提高SOD活性, 表明上述浓度的ET-1预处理可增强低氧心肌细胞抗自由基和脂质过氧化损伤的能力。
低氧引起[Ca2+]i过度升高是造成细胞损伤的重要机制之一。本实验观察到,
培养的乳鼠心肌细胞急性低氧后, 胞内自发性钙瞬变迅速终止, 随后舒张期[Ca2+]i 持续升高, 与Quaife等[18]报道一致。在低氧灌流之前, 上述三种浓度的ET-1预处理均可明显加快培养心肌细胞钙瞬变的频率,
0.1和0.01 nmol/L ET-1预处理对心肌细胞收缩期末和舒张期末[Ca2+]i 及[Ca2+]i瞬变幅度均无明显影响; 1 nmol/L ET-1预处理后,
收缩期末和舒张期末[Ca2+]i 均升高, 但舒张期末[Ca2+]i的增加更明显, 因而钙瞬变幅度减小。单纯低氧组和和ET-1预处理低氧组钙瞬变终止前的舒张期末[Ca2+]i
无显著差异。ET-1预处理虽然不能完全取消低氧引起的[Ca2+]i 变化, 但明显推迟自发性钙瞬变终止的发生, 延缓舒张期[Ca2+]i 的升高。提示ET-1预处理对维持低氧心肌细胞[Ca2+]i
稳定具有一定的保护作用, 这可能既是细胞耐受低氧的表现, 也是细胞耐受低氧的机制之一, 而跨膜钙离子内流、 胞内钙贮释放钙以及肌浆网重摄取钙等钙活动在ET-1预处理作用下所发生的变化尚需进一步研究。ET-1预处理的保护作用可能涉及ETA受体[6]、
Gαq/11[16]、 PKC、 KATP等信号转导通路中的多个环节[6-8]。本实验在培养乳鼠心肌细胞首次观察到ET-1预处理可抑制低氧所致[Ca2+]i
过度升高, 其机制尚待进一步研究, 但ET-1预处理很有可能成为一种有效的药物预适应的保护措施。综上, 本实验在培养的乳鼠心肌细胞证明一定浓度的ET-1预处理可提高心肌细胞抗氧化损伤的能力、
抑制低氧所致胞浆游离钙的过度升高, 减轻细胞低氧损伤, 维持[Ca2+]i稳态, 具有一定的细胞保护作用。
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