쥐 간세포 NA+-Ca2+ 교환이동체의 생리적 역할

초록

Na+-Ca2+ exchanger (NCX) plays an important role in regulating the intracellular Ca2+ concentration ([Ca2+]i) and is expressed in various excitable cells as well as non-excitable cells. However, the role of the NCX in the regulation of [Ca2+]i in hepatocytes is not understood clearly. The purpose of this study is to examine the presence of NCX and determine the NCX activity in hepatocytes. In addition, the role of NCX in isolated rat hepatocytes exposed to ROS was investigated. The immunoblot results showed that the 70 kDa NCX specific band was detected and the expression of the NCX was decreased in As-oligos treated cells, when compared to control cells. Immunofluorescence imaging of permeabilized cultured hepatocytes indicates that NCX was distributed throughout the cytoplasm and an intense labeling near the perinuclear region was observed. In the As-oligos treated cells, the labeling was weak in comparison to that observed both in the control and lipofectamine treated cells. To examine the membrane current conducted via NCX, the conventional whole-cell patch clamp method was performed. The current shift was triggered by ramp voltage pulses in the Na+-free solution and also in the normal Na+ solution. The intersection point of I-V curves was observed between -50 and -40 mV. The inward current mediated by NCX was blocked in hepatocytes treated with As-oligos. The H2O2 treatment increased NCX current, when compared to the normal condition. The NCX activity was investigated by measuring [Ca2+]i using Fura-2. When hepatocytes were perfused with 0 Na+/2.5 mM Ca2+ and 140 mM Na+/0 Ca2+ normal Tyrode solution containing thapsigargin (1 uM), caffeine (5 mM), and La3+ (250 uM), the changes in [Ca2+]i observed upon treatment with these agents were similar to that observed with the As-oligos treated cells. The treatment of H2O2 enhanced an increase in [Ca2+]i significantly. The expression of NCX These results imply that NCX plays an important role in modulating intrac