Berthou C, Marolleau J P, Lafaurie C, Soulié A, Dal Cortivo L, Bourge J F, Benbunan M, Sasportes M
INSERM U 93, Centre de Transfusion sanguine, St Louis Hospital, Paris, France.
Blood. 1995 Nov 1;86(9):3500-6.
Granzyme B and perforin are cytoplasmic granule-associated proteins used by cytotoxic T lymphocytes and natural killer (NK) cells to kill their targets. However, granzyme B gene expression has also been detected in a non-cytotoxic hematopoietic murine multipotent stem cell line, FDCP-Mix. The objective of the present study was to investigate whether granzyme B and perforin could be expressed in human hematopoietic CD34+ cells and if present, discover what their physiologic relevance could be. The primitive CD34+ human cell line KG1a was investigated first and was found to express granzyme B and perforin. Highly purified hematopoietic stem/progenitor cells were then selected using the CD34 surface antigen as marker. Steady-state bone marrow (BM) CD34+ cells did not contain these proteins. Peripheral blood (PB) CD34+ cells, which had been induced to circulate, were also analyzed. After chemotherapy (CT) and granulocyte colony-stimulating factor (G-CSF) treatment, CD34+ cells strongly expressed mRNAs and proteins of granzyme B and perforin. In contrast, CD34+ cells mobilized by G-CSF alone were negative. Western blot analysis further showed that granzyme B and perforin proteins were identical in CD34+ cells and activated PBLs. Such proteins might be implicated in the highly efficient migration of CD34+ stem/progenitor cells from BM to PB after CT and G-CSF treatment. The cellular adhesion mechanisms involved in the BM homing of CD34+ cells are disrupted at least temporarily after CT. The Asp-ase proteolytic activity of granzyme B on extracellular matrix proteins could be used by progenitor cells for their rapid detachment from BM stromal cells and perforin might facilitate their migration across the endothelial cell barrier.
颗粒酶B和穿孔素是细胞毒性T淋巴细胞和自然杀伤（NK）细胞用来杀死靶标的细胞质颗粒相关蛋白。然而，在一种非细胞毒性的造血小鼠多能干细胞系FDCP-Mix中也检测到了颗粒酶B基因表达。本研究的目的是调查颗粒酶B和穿孔素是否能在人造血CD34+细胞中表达，如果存在，探究它们的生理相关性可能是什么。首先研究了原始的人CD34+细胞系KG1a，发现其表达颗粒酶B和穿孔素。然后使用CD34表面抗原作为标志物选择高度纯化的造血干/祖细胞。稳态骨髓（BM）CD34+细胞不含这些蛋白质。还分析了被诱导循环的外周血（PB）CD34+细胞。化疗（CT）和粒细胞集落刺激因子（G-CSF）治疗后，CD34+细胞强烈表达颗粒酶B和穿孔素的mRNA和蛋白质。相比之下，仅由G-CSF动员的CD34+细胞为阴性。蛋白质印迹分析进一步表明，CD34+细胞和活化的外周血淋巴细胞（PBL）中的颗粒酶B和穿孔素蛋白是相同的。这些蛋白质可能与CT和G-CSF治疗后CD34+干/祖细胞从骨髓高效迁移到外周血有关。CT后，参与CD34+细胞骨髓归巢的细胞黏附机制至少会暂时受到破坏。颗粒酶B对细胞外基质蛋白的天冬氨酸酶蛋白水解活性可被祖细胞用于使其快速从骨髓基质细胞脱离，而穿孔素可能有助于它们穿过内皮细胞屏障迁移。
