过氧化氢和次氯酸盐对大鼠心肌细胞线粒体膜电位的影响

Effects of hydrogen peroxide and hypochlorite on membrane potential of mitochondria in situ in rat heart cells

 

 

　

摘要

 

过氧化氢（H2O2）和次氯酸盐（HOCl）引起各种细胞功能障碍。在这项研究中，我们检查了这些药物对离体大鼠心肌细胞中原位线粒体内膜电位梯度的影响。通过胶原酶消化制备肌细胞，并在H 2 O 2或HOCl存在下孵育。通过亲脂阳离子[3H]三苯基甲基鏻+的分布测量跨膜电梯度。首先通过使用洋地黄皂苷进行透化，然后通过溴十二烷层快速离心沉降，将颗粒部分与细胞溶质隔室分离。

 

我们发现线粒体膜电位（161±7mV，内部负性）在氧化应激下相对较好地保持，即，电位仅在高浓度的HOCl和H2O2下降低并且随时间逐渐降低。分离的大鼠心脏线粒体的膜电位受到浓度和时间依赖性的H2O2和HOC1的影响。高浓度的氧化剂也降低了细胞ATP水平，但没有显着改变基质体积。

 

当透化的肌细胞中线粒体外游离钙浓度增加时，跨膜电位成比例地降低，并且这种降低通过H2O2进一步增强。

 

这些结果支持心脏线粒体具有良好发展的抗氧化剂防御机制的观点，但H2O2对跨膜电梯度的作用因细胞溶质钙的增加而加剧。

　

　

Effects of hydrogen peroxide and hypochlorite on membrane potential of mitochondria in situ in rat heart cells

 

 

ABSTRACT

 

Hydrogen peroxide (H2O2) and hypochlorite (HOCl) cause a variety of cellular dysfunctions. In this study we examined the effects of these agents on the electrical potential gradient across the inner membrane of mitochondria in situ in isolated rat heart myocytes. Myocytes were prepared by collagenase digestion and incubated in the presence of H2O2 or HOCl. Transmembrane electrical gradients were measured by distribution of [3H]triphenylmethylphosphonium+, a lipophilic cation. The particulate fraction was separated from the cytosolic compartment first by permeabilization using digitonin, followed by rapid centrifugal sedimentation through a bromododecane layer.

　

We found that the mitochondrial membrane potential (161 ± 7 mV, negative inside) was relatively well maintained under oxidant stress, i.e., the potential was decreased only at high concentrations of HOCl and H2O2 and gradually with time. The membrane potential of isolated rat heart mitochondria was affected similarly by H2O2 and HOCl in a concentration- and time-dependent manner. High concentrations of oxidants also reduced the cellular ATP level but did not significantly change the matrix volume.

　

When the extra-mitochondrial free calcium concentration was increased in permeabilized myocytes, the transmembrane potential was decreased proportionally, and this decrease was potentiated further by H2O2.

　

These results support the view that heart mitochondria are equipped with well-developed defense mechanisms against oxidants, but the action of H2O2 on the transmembrane electrical gradient is exacerbated by an increase in cytosolic calcium.

　

Keywords: ATP, calcium, cardiomyocyte, cell defense, mitochondrial membrane potential, oxidant, triphenylmethylphosphonium.

　

Noburu Konno and , K. J. Kako

 

Canadian Journal of Physiology and Pharmacology, 1991, 69(11): 1705-1712, https://doi.org/10.1139/y91-253

　

 

Effects of hydrogen peroxide and hypochlorite on membrane potential of mitochondria in situ in rat heart cells - Canadian Journal of Physiology and Pharmacology  http://www.nrcresearchpress.com/doi/abs/10.1139/y91-253#.W-KG2fkdgbw