禁食会增强缺氧对离体大鼠肝细胞中三磷酸腺苷（ATP）、钙离子（Ca2+）及细胞损伤的影响。

Fasting enhances the effects of anoxia on ATP, Cai2+ and cell injury in isolated rat hepatocytes.

作者信息

Gasbarrini A, Borle A B, Farghali H, Caraceni P, Van Thiel D

机构信息

Department of Physiology, University of Pittsburgh School of Medicine, PA.

出版信息

Biochim Biophys Acta. 1993 Jul 28;1178(1):9-19. doi: 10.1016/0167-4889(93)90105-x.

DOI:10.1016/0167-4889(93)90105-x

PMID:8329459

Abstract

The effect of fasting and anoxia on the intracellular concentration of ATP, Na+, Ca2+, Mg2+, and H+ was studied in isolated perfused rat hepatocytes. ATP and intracellular Mg2+ were measured by 31P-NMR spectroscopy, cytosolic free calcium was measured with aequorin, intracellular Na+ with SBFI, intracellular pH with BCECF, lactic dehydrogenase by NADH absorbance. In hepatocytes from fasted rats, intracellular ATP was depressed 52% (P < 0.001), Nai+ was increased 70% from 16.9 to 27.7 mM (P < 0.02), and Cai2+ was increased 79% from 137 to 245 nM (P < 0.05) when compared to fed rats. Mgi2+ and pHi were unchanged. During anoxia, ATP and the cell phosphorylation potential decreased 90% to practically the same low levels in both fed and fasted groups. On the other hand, in hepatocytes from fasted animals, Cai2+ increased faster and to significantly higher levels than in hepatocytes from fed rats: Cai2+ reached 2.19 microM in 10 min compared to 1.45 microM in 1 h, respectively (P < 0.05). Cell injury assessed by LDH release and trypan blue exclusion also occurred earlier and was more severe in hepatocytes from fasted rats. Fructose and Ca(2+)-free perfusion reduced the rise in Cai2+, abolished LDH release and significantly improved the cell viability measured by Trypan blue exclusion. The data demonstrate that fasting decreases the hepatocytes energy potential and increases Nai+ and Cai2+ which are inversely related to the cell energy potential. Consequently, in hepatocytes isolated from fasted rats, the increase in Cai2+ and the resulting cell injury evoked by anoxia occur earlier and are more severe than in fed rats. These results suggest that Ca2+ plays a crucial role in the development of anoxic cell injury.

摘要

在分离的灌注大鼠肝细胞中研究了禁食和缺氧对细胞内ATP、Na⁺、Ca²⁺、Mg²⁺和H⁺浓度的影响。通过³¹P-NMR光谱法测量ATP和细胞内Mg²⁺，用水母发光蛋白测量胞质游离钙，用SBFI测量细胞内Na⁺，用BCECF测量细胞内pH，通过NADH吸光度测量乳酸脱氢酶。与喂食大鼠相比，禁食大鼠的肝细胞中，细胞内ATP降低了52%（P < 0.001），细胞内Na⁺从16.9 mM增加到27.7 mM，增加了70%（P < 0.02），细胞内Ca²⁺从137 nM增加到245 nM，增加了79%（P < 0.05）。细胞内Mg²⁺和细胞内pH未发生变化。在缺氧期间，喂食组和禁食组的ATP和细胞磷酸化电位均降低了90%，降至几乎相同的低水平。另一方面，在禁食动物的肝细胞中，细胞内Ca²⁺升高得更快，且显著高于喂食大鼠的肝细胞：细胞内Ca²⁺在10分钟内达到2.19 μM，而喂食大鼠的肝细胞在1小时内达到1.45 μM（P < 0.05）。通过乳酸脱氢酶释放和台盼蓝排斥评估的细胞损伤在禁食大鼠的肝细胞中也更早发生且更严重。果糖和无钙灌注减少了细胞内Ca²⁺的升高，消除了乳酸脱氢酶释放，并显著提高了通过台盼蓝排斥测量的细胞活力。数据表明，禁食降低了肝细胞的能量潜力，增加了与细胞能量潜力呈负相关的细胞内Na⁺和细胞内Ca²⁺。因此，在从禁食大鼠分离的肝细胞中，缺氧引起的细胞内Ca²⁺增加及由此导致的细胞损伤比喂食大鼠更早发生且更严重。这些结果表明Ca²⁺在缺氧性细胞损伤的发生发展中起关键作用。