物理化学学报 >> 2018, Vol. 34 >> Issue (1): 73-80.doi: 10.3866/PKU.WHXB201707043

论文 上一篇    下一篇

不同条件下离体大鼠肝脏线粒体能量代谢微量热分析

袁莲1,刘玉娇1,何欢1,蒋风雷1,李会荣2,刘义1,*()   

  1. 1 武汉大学化学与分子科学学院,武汉430072
    2 武汉东湖学院生命科学与化学学院,武汉430212
  • 收稿日期:2017-06-13 发布日期:2017-11-01
  • 通讯作者: 刘义 E-mail:yiliuchem@whu.edu.cn
  • 基金资助:
    国家自然科学基金(21673166);湖北省自然科学基金项目(2015CFC892)

Microcalorimetric Analysis of Isolated Rat Liver Mitochondrial Metabolism under Different Conditions

Lian YUAN1,Yu-Jiao LIU1,Huan HE1,Feng-Lei JIANG1,Hui-Rong LI2,Yi LIU1,*()   

  1. 1 College of Chemistry and Molecular Sciences, Wuhan University, Wuhan 430072, P. R. China
    2 College of Life Science and Chemistry, Wuhan Donghu University, Wuhan 430212, P. R. China
  • Received:2017-06-13 Published:2017-11-01
  • Contact: Yi LIU E-mail:yiliuchem@whu.edu.cn
  • Supported by:
    the National Natural Science Foundation of China(21673166);Natural Science Foundation of Hubei Province, China(2015CFC892)

摘要:

为了探究线粒体的能量代谢过程,本文以离体大鼠肝脏线粒体为模型,利用多通道、高灵敏度的热活性检测仪TAM Ⅲ,实时监测了不同线粒体浓度、不同底物、不同缓冲液、几种呼吸抑制剂以及Ca2+和线粒体渗透转换孔抑制剂CsA存在时线粒体的能量代谢,获得了完整的热功率―时间曲线,并通过计算得到了线粒体能量代谢的热动力学参数。通过分析发现:(1)线粒体浓度越大,代谢越快;(2)直接底物琥珀酸钠使线粒体代谢更快;(3)高浓度Ca2+能够刺激线粒体快速产热,且在长期代谢进程中,线粒体渗透转换孔抑制剂CsA并不能改变Ca2+造成的影响;(4)不同缓冲液对线粒体代谢的影响基于其组分的不同,缓冲液中含有呼吸底物;(5)呼吸抑制剂都能抑制线粒体的能量代谢,尤其是复合物IV的抑制剂NaN3,高浓度下使代谢停止。

关键词: 线粒体, 微量热, 代谢速率, 呼吸链抑制剂, 线粒体渗透转换

Abstract:

Isolated rat liver mitochondria were proposed as a model to monitor real-time heat metabolism.A high-throughput and sensitive thermal activity monitor Ⅲ (TAM Ⅲ) was used to detect the P-t curves of mitochondria under different conditions, including different mitochondrial concentrations, different substrates, different buffers, respiratory inhibitors, Ca2+, and CsA.We determined the thermokinetic parameters through calculation.The results showed that:(1) higher concentration of mitochondria led to faster energetic metabolism; (2) when succinate was the direct respiratory substrate, it promoted mitochondrial metabolism, in contrast to the condition when an indirect substrate, pyruvate, was used; (3) high concentration of Ca2+(2.5 mmol·L-1) stimulated mitochondrial metabolism, however CsA, an inhibitor of mitochondrial permeability transition pores, could not reverse the Ca2+-induced mitochondrial alteration; (4) mitochondria in various buffers displayed different rates of heat metabolism, because of the different composition of the buffers; (5) mitochondrial metabolism was inhibited by respiratory inhibitors, especially NaN3, which is an inhibitor of Complex Ⅳ and which completely stopped the mitochondrial heat release.

Key words: Mitochondria, Microcalorimetry, Metabolic rate, Respiratory inhibitor, Mitochondrial permeability transition

MSC2000: 

  • O642.3