Wharton M, Granger D L, Durack D T
Department of Medicine, Duke University Medical Center, Durham, NC 27710.
J Immunol. 1988 Aug 15;141(4):1311-7.
Activated macrophages inhibit replication of murine lymphoblastic leukemia L1210 cells without lysis. This inhibition of replication is associated with abnormalities of mitochondrial electron transport at the level of NADH dehydrogenase (NADH-DH) and succinate dehydrogenase (SDH). The mechanism of inhibition is unknown, although it has been demonstrated that as NADH-DH and SDH activity is lost, iron is released from cells. Because both NADH-DH and SDH contain numerous iron-sulfur clusters, damage to these structures may be one result of injury by activated macrophages. L1210 cells were labeled with 55Fe and co-cultivated with activated murine peritoneal macrophages (injured L1210 cells). At 48 h, injured L1210 cells had released 83 +/- 8% (mean +/- SEM of 55Fe activity into the media, compared with 25 +/- 4% release from control and 37 +/- 7% from nondividing mitomycin C-treated control cells. All cells were greater than 90% viable. These differences were also reflected in the iron content of the cells. Mitochondria were then separated by centrifugation after cell disruption and 55Fe activity was found to be similarly decreased in both mitochondrial and nonmitochondrial fractions of injured L1210 cells. To further characterize the changes in mitochondrial iron content, mitochondrial proteins from injured and control L1210 cells were separated by IEF and 55Fe activity of gel slices was determined. There was selective loss of 55Fe activity in the area of the gel corresponding to SDH and NADH-DH, suggesting that iron loss from iron-sulfur clusters may occur in L1210 cells injured by activated macrophages. Iron uptake into L1210 cells after removal from macrophages showed a rapid large influx of radioactive iron. L1210 cells in contact with macrophages appear to develop an iron-depleted state, which is dependent on the continued presence of macrophages.
活化的巨噬细胞可抑制鼠淋巴细胞白血病L1210细胞的复制,且不发生裂解。这种对复制的抑制与线粒体电子传递在NADH脱氢酶(NADH-DH)和琥珀酸脱氢酶(SDH)水平上的异常有关。抑制机制尚不清楚,尽管已证明随着NADH-DH和SDH活性丧失,铁从细胞中释放出来。由于NADH-DH和SDH都含有大量铁硫簇,这些结构的损伤可能是活化巨噬细胞造成损伤的一个结果。用55Fe标记L1210细胞,并与活化的鼠腹腔巨噬细胞(损伤的L1210细胞)共培养。48小时后,损伤的L1210细胞已将83±8%(55Fe活性的平均值±标准误)释放到培养基中,相比之下,对照细胞释放25±4%,丝裂霉素C处理的非分裂对照细胞释放37±7%。所有细胞的活力均大于90%。这些差异也反映在细胞的铁含量上。然后在细胞破碎后通过离心分离线粒体,发现损伤的L1210细胞的线粒体和非线粒体部分中的55Fe活性均同样降低。为了进一步表征线粒体铁含量的变化,通过等电聚焦分离损伤的和对照的L1210细胞的线粒体蛋白,并测定凝胶切片的55Fe活性。在凝胶上对应于SDH和NADH-DH的区域有55Fe活性的选择性丧失,这表明在被活化巨噬细胞损伤的L1210细胞中可能发生铁硫簇中铁的丢失。从巨噬细胞中移出后,L1210细胞对铁的摄取显示放射性铁迅速大量流入。与巨噬细胞接触的L1210细胞似乎会形成铁缺乏状态,这取决于巨噬细胞的持续存在。