Tønnessen T I, Aas A T, Ludt J, Blomhoff H K, Olsnes S
Institute for Cancer Research, Norwegian Radium Hospital, Montebello, Oslo.
J Cell Physiol. 1990 Apr;143(1):178-87. doi: 10.1002/jcp.1041430125.
The effect of serum, phorbol-12-myristate-13-acetate (TPA), and forskolin on the activity Na+/H+ antiport and the Na(+)-coupled and Na(+)-independent Cl-/HCO3- antiport was studied in Vero cells by measuring 22Na+ and 36Cl- fluxes and changes in cytosolic pH (pHi). The Na(+)-independent Cl-/HCO3- antiport, which acts as an acidifying mechanism, is strongly pH-sensitive. In serum-starved cells it is activated at alkaline cytosolic pH, with a half-maximal activity at pHi approximately 7.20. Incubation with serum increased the activity of the Na(+)-independent Cl-/HCO3- antiport at pHi values from 6.8 to 7.2. Thus serum appeared to alter the pHi sensitivity of this antiporter such that the threshold value for activation of the antiport was shifted to a more acidic value. Na+/H+ antiport was somewhat stimulated initially by addition of serum, but further incubation with serum (greater than 45 min) decreased its activity. The activity of the Na(+)-coupled Cl-/HCO3- antiport, which is the major alkalinizing antiport in Vero cells, was not altered by short-term incubation with serum (less than 10 min) but decreased after prolonged incubation (greater than 45 min). Our findings with TPA and forskolin indicate that the effect of serum is partly mediated by the protein kinase C pathway, whereas the cyclic adenosine monophosphate pathway does not appear to play an important role. The net effect of serum on the pHi-regulating antiports was a slight decrease in intracellular pH.
通过测量22Na+和36Cl-通量以及胞质pH（pHi）的变化，研究了血清、佛波醇-12-肉豆蔻酸酯-13-乙酸酯（TPA）和毛喉素对Vero细胞中Na+/H+逆向转运体活性以及Na(+)-偶联和Na(+)-非依赖性Cl-/HCO3-逆向转运体的影响。作为一种酸化机制的Na(+)-非依赖性Cl-/HCO3-逆向转运体对pH非常敏感。在血清饥饿的细胞中，它在碱性胞质pH下被激活，在pHi约为7.20时活性达到最大值的一半。用血清孵育可增加pHi值在6.8至7.2之间时Na(+)-非依赖性Cl-/HCO3-逆向转运体的活性。因此，血清似乎改变了这种逆向转运体的pHi敏感性，使得逆向转运体激活的阈值向更酸性的值移动。加入血清最初对Na+/H+逆向转运体有一定刺激作用，但继续用血清孵育（大于45分钟）会降低其活性。Na(+)-偶联的Cl-/HCO3-逆向转运体是Vero细胞中主要的碱化逆向转运体，短期用血清孵育（小于10分钟）对其活性没有影响，但长时间孵育（大于45分钟）后活性降低。我们关于TPA和毛喉素的研究结果表明，血清的作用部分是由蛋白激酶C途径介导的，而环磷酸腺苷途径似乎没有发挥重要作用。血清对pHi调节逆向转运体的净效应是细胞内pH略有降低。
