Enhanced cell volume regulation: a key protective mechanism of ischemic preconditioning in rabbit ventricular myocytes
Enhanced cell volume regulation: a key protective mechanism of ischemic preconditioning in rabbit ventricular myocytes
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DOI：
10.1016/s0022-2828(02)00277-8
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发表时间：
2003-01-01
影响因子：
5
通讯作者：
Wilson, GJ
中科院分区：
文献类型：
--
作者：
Diaz, RJ;Armstrong, SC;Wilson, GJ
代谢活性代谢物的蓄积（其产生的渗透压梯度估计约为60 mCsM）和细胞肿胀是缺血性心肌细胞死亡的显著特征。本研究验证了这样一个假设，即通过增强细胞体积调节来减少缺血性肿胀是缺血预处理（IPC）延迟缺血性心肌细胞死亡的关键机制。实验方案解决了以下问题：（i）IPC是否触发细胞体积调节机制，其在随后的指数缺血期间减少心肌细胞肿胀;（ii）缺血细胞肿胀的这种减少在幅度上足以解释IPC保护;（iii）介导IPC的分子机制也介导细胞体积调节。用兔心室肌细胞研究了两种实验模型：新鲜分离的颗粒状心肌细胞和培养48小时的心肌细胞。通过不同的短模拟缺血（SI）/模拟再灌注方案（IPC）或通过使肌细胞经受药理学预处理（PPC）方案（1 μ M calyculin A或1 μ M N-6-2-（4-氨基苯基）乙基腺苷（APNEA），然后使它们经受不同持续时间的长SI或30分钟低渗应激）来预处理肌细胞。通过台盼蓝染色监测细胞死亡（蓝色正方形肌细胞百分比）。通过溴代十二烷细胞浮选试验（定性）或视频/共聚焦显微镜（定量）测定细胞肿胀。模拟缺血引起两种模型的心肌细胞肿胀。在沉淀的肌细胞中，IPC或PPC与calyculin A或APNEA一起产生了显著减少的缺血细胞肿胀，如通过细胞漂浮测定所确定的。在培养的心肌细胞中，IPC显著减少缺血细胞肿胀（P < 0.001）。IPC对缺血性细胞肿胀的影响与IPC和PPC（伴APNEA）介导的细胞体积调节性减少（RVD）触发有关。IPC和APNEA也能显著减轻低渗细胞肿胀（P < 0.001）。这种IPC和APNEA效应可被腺苷受体、PKC或Cl-通道抑制剂阻断。IPC保护的渗透当量约为50-60 mOsM，渗透梯度类似于预处理和非预处理心肌细胞的估计缺血渗透负荷。结果表明，细胞体积调节是心肌细胞IPC保护的主要机制。(C)2003爱思唯尔科技有限公司版权所有。
Accumulation of osmotically active metabolites, which create an osmotic gradient estimated at similar to60 mCsM, and cell swelling are prominent features of ischemic myocardial cell death. This study tests the hypothesis that reduction of ischemic swelling by enhanced cell volume regulation is a key mechanism in the delay of ischemic myocardial cell death by ischemic preconditioning (IPC). Experimental protocols address whether: (i) IPC triggers a cell volume regulation mechanism that reduces cardiomyocyte swelling during subsequent index ischemia; (ii) this reduction in ischemic cell swelling is sufficient in magnitude to account for the IPC protection; (iii) the molecular mechanism that mediates IPC also mediates cell volume regulation. Two experimental models with rabbit ventricular myocytes were studied: freshly isolated pelleted myocytes and 48-h cultured myocytes. Myocytes were preconditioned either by distinct short simulated ischemia (SI)/simulated reperfusion protocols (IPC), or by subjecting myocytes to a pharmacological preconditioning (PPC) protocol (1 muM calyculin A, or 1 muM N-6-2-(4-aminophenyl)ethyladenosine (APNEA), prior to subjecting them to either different durations of long SI or 30 min hypo-osmotic stress. Cell death (percent blue square myocytes) was monitored by trypan blue staining. Cell swelling was determined by either the bromododecane cell flotation assay (qualitative) or video/confocal microscopy (quantitative). Simulated ischemia induced myocyte swelling in both the models. In pelleted myocytes, IPC or PPC with either calyculin A or APNEA produced a marked reduction of ischemic cell swelling as determined by the cell floatation assay. In cultured myocytes, IPC substantially reduced ischemic cell swelling (P < 0.001). This IPC effect on ischemic cell swelling was related to an IPC and PPC (with APNEA) mediated triggering of cell volume regulatory decrease (RVD). IPC and APNEA also significantly (P < 0.001) reduced hypo-osmotic cell swelling. This IPC and APNEA effect was blocked by either adenosine receptor, PKC or Cl- channel inhibition. The osmolar equivalent for IPC protection approximated 50-60 mOsM, an osmotic gradient similar to the estimated ischemic osmotic load for preconditioned and non-preconditioned myocytes. The results suggest that cell volume regulation is a key mechanism that accounts for most of the IPC protection in cardiomyocytes. (C) 2003 Elsevier Science Ltd. All rights reserved.