Barber Amorette, Zhang Tong, Megli Christina J, Wu Jillian, Meehan Kenneth R, Sentman Charles L
Department of Microbiology and Immunology, Dartmouth Medical School, Dartmouth Hitchcock Medical Center, Lebanon, NH 03756, USA.
Exp Hematol. 2008 Oct;36(10):1318-28. doi: 10.1016/j.exphem.2008.04.010. Epub 2008 Jul 2.
Most myeloma tumor cells from patients express NKG2D ligands. We have reported the development of a chimeric NKG2D receptor (chNKG2D), which consists of the NKG2D receptor fused to the CD3zeta chain. T cells expressing this receptor kill and produce cytokines in response to NKG2D-ligand+ tumor cells. Therefore, we investigated whether human chNKG2D T cells respond against human myeloma cells.
ChNKG2D T cells were generated from healthy donors and myeloma patients. The effector phase of chNKG2D T cells was analyzed by cell-surface marker expression and human myeloma cell lines were tested for expression of NKG2D ligands. Lysis of myeloma cell lines and cytokine secretion by chNKG2D T cells was determined. ChNKG2D T cells grown in serum-free media, or cyropreserved, were assessed for effector cell functions.
Myeloma cell lines expressed NKG2D ligands. ChNKG2D T cells from healthy donors and myeloma patients lysed myeloma cells, and secreted proinflammatory cytokines when cultured with myeloma cells or patient bone marrow, but not with peripheral blood mononuclear cells or normal bone marrow. Lysis of myeloma cells was dependent on chNKG2D T-cell expression of NKG2D and perforin. Additionally, chNKG2D T cells upregulated CD45RO, did not express CD57, and maintained expression of CD27, CD62L, and CCR7, indicating that the T cells were at an early effector stage. Finally, we showed that chNKG2D T cells generated with serum-free media, or when cryopreserved, maintained effector functions.
ChNKG2D T cells respond to human myeloma cells and can be generated using clinically applicable cell culture techniques.
多数骨髓瘤患者的肿瘤细胞表达NKG2D配体。我们已报道了一种嵌合NKG2D受体（chNKG2D）的研发，该受体由与CD3ζ链融合的NKG2D受体组成。表达此受体的T细胞在遇到表达NKG2D配体的肿瘤细胞时会杀伤肿瘤细胞并分泌细胞因子。因此，我们研究了人chNKG2D T细胞是否对人骨髓瘤细胞有反应。
从健康供体和骨髓瘤患者中产生chNKG2D T细胞。通过细胞表面标志物表达分析chNKG2D T细胞的效应阶段，并检测人骨髓瘤细胞系中NKG2D配体的表达。测定chNKG2D T细胞对骨髓瘤细胞系的杀伤作用及细胞因子分泌情况。评估在无血清培养基中培养或冻存的chNKG2D T细胞的效应细胞功能。
骨髓瘤细胞系表达NKG2D配体。来自健康供体和骨髓瘤患者的chNKG2D T细胞可杀伤骨髓瘤细胞，并且在与骨髓瘤细胞或患者骨髓共同培养时分泌促炎细胞因子，但与外周血单个核细胞或正常骨髓共同培养时则不分泌。骨髓瘤细胞的杀伤作用依赖于chNKG2D T细胞对NKG2D和穿孔素的表达。此外，chNKG2D T细胞上调CD45RO的表达，不表达CD57，并维持CD27、CD62L和CCR7的表达，表明这些T细胞处于早期效应阶段。最后，我们发现用无血清培养基产生的或冻存后的chNKG2D T细胞维持了效应功能。
chNKG2D T细胞对人骨髓瘤细胞有反应，并且可以使用临床适用的细胞培养技术产生。
