过氧化氢对大鼠肝卵圆细胞内游离钙含量的影响

Effects of Hydrogen Peroxide on Intracellular Free Ca ^(2+) Content in Rat Liver Oval Cells

目的研究小剂量(800nmol/L)过氧化氢H2O2诱导培养的大鼠肝卵圆细胞(WB细胞)胞质内游离钙犤Ca2+犦i含量的变化及其机制。方法以800nmol/LH2O2刺激WB细胞,采用Fluo-3/AM作为荧光指示剂,以激光共聚焦扫描显微镜测定不同条件下犤Ca2+犦i荧光值的改变以反映犤Ca2+犦i含量的变化。结果(1)小剂量H2O2可引起犤Ca2+犦i升高,约300s后达到高峰,800s后恢复正常,过氧化氢酶(CAT)可抑制此变化;(2)细胞在无钙外环境下,相同浓度的H2O2不引起犤Ca2+犦i变化;(3)钙拮抗剂心痛定(nifedipine)不能抑制犤Ca2+犦i荧光值的升高,而蒽-9-羧酸(A9C)则可抑制此变化。结论小剂量H2O2刺激WB细胞可诱使犤Ca2+犦i含量升高,其机制可能是H2O2引起细胞膜上非选择性阳离子通道开放使胞外Ca2+进入胞内,造成胞浆内犤Ca2+犦i含量增高。

Objective To study the effects and mechanism of hydrogen peroxide(H 2 O 2 )of low concentration on dynamic changes of intracellular free calcium contents(Ca 2+ i )in cultural rat liver oval cells(WB -F344cells).Methods Using Fluo-3/Am as fluorescent indicator of Ca 2+ i and it was measured by laser scanning confocal microscope system.Results The results showed that:(1)A rapid transient spiking of Ca 2+ i occurred after the stimulation of H 2 O 2 of low concentration(800nmol/L).(2)The Ca 2+ i increase was abolished by pretreated with catalase(CAT)or by incubated in D-Hank's solution containing EGTA,the chelate of extracellular Ca 2+ .(3)The Ca 2+ i increase was not inhibited by pretreated nifedipine,Ca 2+ channel blocker,but was abolished by pretreated with anthracere-9-cardoxylic acid(A9C),the Cl -channel blocker and which also blocked calcium activated non-selective cation channel(CAN).Conclusions These results suggest that the increase of Ca 2+ i induced by H 2 O 2 of low concentration may be due to the influx of extracellular Ca 2+ through CAN.