McClane B A
Department of Microbiology, Biochemistry and Molecular Biology, University of Pittsburgh School of Medicine, PA 15261.
Microb Pathog. 1989 Jan;6(1):17-28. doi: 10.1016/0882-4010(89)90004-1.
This report characterizes the involvement of Ca2+ in the release of nucleotides from Vero cells caused by Clostridium perfringens enterotoxin (CPE). A positive linear correlation was observed between increased CPE-induced nucleotide-release and increased extracellular calcium over the range 0.01 to 10 mM calcium. Above 5 mM Ca2+, CPE-specific lysis (i.e. disintegration of cells as monitored by light microscopy) was observed. Addition of 1.7 mM Ca2+ to Vero cells previously CPE-treated in Ca2+-free buffer rapidly increased nucleotide-release, even when cells had been previously incubated for 1 h at 37 degrees C in Ca2+-free buffer. Withdrawal of Ca2+, even after the onset of nucleotide-release, halted further CPE-induced nucleotide-release. These results indicate that Ca2+ must be continuously present for significant CPE-induced nucleotide-release. However, withdrawal of Ca2+ did not reverse membrane bleb formation by CPE. This differentiates bleb formation and nucleotide-release (both Ca2+-dependent CPE effects) and suggests that nucleotide-release does not result simply from bleb formation. Lastly, it was shown that other ions besides physiologic Ca2+ (1.7 mM) are required for CPE-induced nucleotide-release. Interestingly, a role for other ions (but not physiologic Ca2+) is also shown for 86Rb-release by CPE (an early Ca2+-independent CPE effect). This indicates that extracellular ions other than physiologic Ca2+ can be required for both Ca2+-independent and Ca2+-dependent CPE effects.
本报告描述了钙离子（Ca2+）在产气荚膜梭菌肠毒素（CPE）引起的Vero细胞核苷酸释放过程中的作用。在0.01至10 mM钙浓度范围内，观察到CPE诱导的核苷酸释放增加与细胞外钙增加呈正线性相关。当Ca2+浓度高于5 mM时，可观察到CPE特异性裂解（即通过光学显微镜监测的细胞解体）。向先前在无钙缓冲液中用CPE处理过的Vero细胞中添加1.7 mM Ca2+，即使细胞先前已在无钙缓冲液中于37℃孵育1小时，也会迅速增加核苷酸释放。即使在核苷酸释放开始后去除Ca2+，也会阻止CPE进一步诱导的核苷酸释放。这些结果表明，必须持续存在Ca2+才能实现显著的CPE诱导的核苷酸释放。然而，去除Ca2+并不会逆转CPE引起的膜泡形成。这区分了膜泡形成和核苷酸释放（两者均为Ca2+依赖性CPE效应），并表明核苷酸释放并非简单地由膜泡形成导致。最后，研究表明，除了生理浓度的Ca2+（1.7 mM）外，CPE诱导的核苷酸释放还需要其他离子。有趣的是，对于CPE诱导的86Rb释放（一种早期的非Ca2+依赖性CPE效应），也显示了其他离子（而非生理浓度的Ca2+）的作用。这表明除生理浓度的Ca2+外，细胞外离子对于非Ca2+依赖性和Ca2+依赖性CPE效应可能都是必需的。
