Hepatic natural killer cells exclusively kill splenic/blood natural killer‐resistant tumor cells by the perforin/granzyme pathway
Hepatic natural killer cells exclusively kill splenic/blood natural killer‐resistant tumor cells by the perforin/granzyme pathway
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DOI：
10.1189/jlb.72.4.668
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发表时间：
2002-10
影响因子：
5.5
通讯作者：
D. Vermijlen;D. Luo;C. Froelich;J. Medema;J. Kummer;E. Willems;F. Braet;E. Wisse
中科院分区：
文献类型：
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作者：
D. Vermijlen;D. Luo;C. Froelich;J. Medema;J. Kummer;E. Willems;F. Braet;E. Wisse
肝脏自然杀伤 (NK) 细胞位于附着于内皮的肝窦中。人类和大鼠肝脏 NK 细胞会诱导对脾脏或血液 NK 细胞有抵抗力的肿瘤细胞发生细胞溶解。为了研究细胞死亡的机制，我们检测了分离的纯 (90%) 大鼠肝 NK 细胞杀死脾/血 NK 抗性肥大细胞瘤细胞系 P815 的能力。通过荧光和透射电子显微镜、DNA 碎片和 51Cr 释放来观察和量化细胞死亡。通过实时逆转录聚合酶链反应和蛋白质印迹法测定RNA和蛋白质表达。与脾NK细胞相比，肝NK细胞表达更高水平的穿孔素和颗粒酶B，并且容易诱导P815细胞凋亡。尽管 P815 细胞会屈服于重组 Fas 配体 (FasL) 或分离的穿孔素/颗粒酶 B，但肝 NK 细胞仅使用颗粒途径来杀死该靶标。此外，肝NK细胞和肝窦内皮细胞强烈表达颗粒酶B抑制剂、蛋白酶抑制剂9（PI-9）/丝氨酸PI-6（SPI-6），而P815细胞和肝细胞呈阴性。用这种抑制剂转染靶细胞可完全抵抗肝 NK 细胞诱导的细胞凋亡。总之，肝脏 NK 细胞仅通过穿孔素/颗粒酶途径杀死脾/血 NK 耐药/FasL 敏感肿瘤细胞。丝氨酸蛋白酶抑制剂 PI-9/SPI-6 在肝窦内皮细胞中的表达可能会保护肝脏微环境免受这种用于杀死转移癌细胞的高活性穿孔素/颗粒酶途径的影响。
Hepatic natural killer (NK) cells are located in the liver sinusoids adherent to the endothelium. Human and rat hepatic NK cells induce cytolysis in tumor cells that are resistant to splenic or blood NK cells. To investigate the mechanism of cell death, we examined the capacity of isolated, pure (90%) rat hepatic NK cells to kill the splenic/blood NK‐resistant mastocytoma cell line P815. Cell death was observed and quantified by fluorescence and transmission electron microscopy, DNA fragmentation, and 51Cr release. RNA and protein expression were determined by real time reverse transcription‐polymerase chain reaction and Western blotting. Compared with splenic NK cells, hepatic NK cells expressed higher levels of perforin and granzyme B and readily induced apoptosis in P815 cells. Although P815 cells succumbed to recombinant Fas ligand (FasL) or isolated perforin/granzyme B, hepatic NK cells used only the granule pathway to kill this target. In addition, hepatic NK cells and sinusoidal endothelial cells strongly expressed the granzyme B inhibitor, protease inhibitor 9 (PI‐9)/serine PI‐6 (SPI‐6), and P815 cells and hepatocytes were negative. Transfection of target cells with this inhibitor resulted in complete resistance to hepatic NK cell‐induced apoptosis. In conclusion, hepatic NK cells kill splenic/blood NK‐resistant/FasL‐sensitive tumor cells exclusively by the perforin/granzyme pathway. Serine protease inhibitor PI‐9/SPI‐6 expression in liver sinusoidal endothelial cells may protect the liver microenvironment from this highly active perforin/granzyme pathway used to kill metastasizing cancer cells.