Department of Toxicology, Hannover Medical School, Carl Neuberg Str. 1, 30625 Hannover, Germany; Department of Neurology, Hannover Medical School, Carl Neuberg Str. 1, 30625 Hannover, Germany.
Department of Cellular Imaging and Electron Microscopy, Leibniz Institute of Molecular Pharmacology, Robert Rössle Str. 10, 13125 Berlin, Germany.
Eur J Pharmacol. 2013 Nov 15;720(1-3):227-36. doi: 10.1016/j.ejphar.2013.10.022. Epub 2013 Oct 24.
An increase in cytosolic free calcium concentration [Ca(2+)]i initiates the exocytotic activity in various types of secretory cells. The guanosine 5'-O-[3-thio]triphosphate (GTPγS), a non-hydrolysable analogue of GTP (guanosine 5'-triphosphate), is an effective secretagogue for different cell types of different species, like mast cells, neutrophils or eosinophils. Consequently, the internal administration of GTPγS causes degranulation of mouse and rat mast cells. Regarding rat mast cells, it is proved that Ca(2+) can cooperate with GTP or GTPγS in accelerating and increasing amplitude of the secretory response. All the previous studies with respect to capacitance recordings and mast cells were performed using mouse or rat mast cells, usually derived from peritoneum or the rat basophilic leukaemia cell line RBL. In this study, we applied the capacitance measurement technique to the human mast cell line-1 (HMC-1) cells, an immature cell line established from a patient with mast cell leukaemia. Patch-clamp dialysis experiments revealed that high intracellular free Ca(2+) and GTPγS concentrations are both required for considerable capacitance increases in HMC-1 cells. During degranulation of HMC-1 cells, the total membrane capacitance (Cm) increase appeared continuously and, in some cases, as a discrete capacitance change, developing in a stepwise manner. Then, we tested the effect of latrunculin B upon HMC-1 cell capacitance increase as well as of some classic mast cell stimulators like PMA, A23187 and IL-1β in hexosaminidase release. Finally, we could conclude that the HMC-1 cell line represents a suitable model for the study of human mast cell degranulation.
细胞溶质游离钙浓度 [Ca(2+)]i 的增加启动了各种类型分泌细胞的胞吐活性。鸟苷 5'-O-[3-硫]三磷酸 (GTPγS),一种 GTP(鸟苷 5'-三磷酸)的非水解类似物,是不同物种的不同细胞类型的有效分泌激动剂,如肥大细胞、嗜中性粒细胞或嗜酸性粒细胞。因此,GTPγS 的内源性给药会导致小鼠和大鼠肥大细胞脱颗粒。关于大鼠肥大细胞,已经证明 Ca(2+)可以与 GTP 或 GTPγS 协同作用,加速和增加分泌反应的幅度。所有以前关于电容记录和肥大细胞的研究都是使用小鼠或大鼠肥大细胞进行的,通常来源于腹膜或大鼠嗜碱性白血病细胞系 RBL。在这项研究中,我们将电容测量技术应用于人类肥大细胞系-1 (HMC-1) 细胞,这是一种从肥大细胞白血病患者中建立的未成熟细胞系。膜片钳透析实验表明,高细胞内游离 Ca(2+)和 GTPγS 浓度都是 HMC-1 细胞中相当大的电容增加所必需的。在 HMC-1 细胞脱颗粒过程中,总膜电容 (Cm) 的增加持续出现,在某些情况下,作为离散的电容变化,以逐步的方式发展。然后,我们测试了 latrunculin B 对 HMC-1 细胞电容增加的影响,以及 PMA、A23187 和 IL-1β 等一些经典肥大细胞刺激物对 hexosaminidase 释放的影响。最后,我们可以得出结论,HMC-1 细胞系是研究人类肥大细胞脱颗粒的合适模型。