从血清学阳性或阴性的健康受试者或干细胞移植受者中诱导巨细胞病毒特异性CD4 + 细胞毒性T淋巴细胞。

Induction of cytomegalovirus-specific CD4+ cytotoxic T lymphocytes from seropositive or negative healthy subjects or stem cell transplant recipients.

作者信息

Tazume Kei, Hagihara Masao, Gansuvd Balgansuren, Higuchi Ayako, Ueda Yoko, Hirabayashi Kaoru, Hojo Motoharu, Tanabe Akiko, Okamoto Ayako, Kato Shunichi, Hotta Tomomitsu

机构信息

Department of Hematology, Tokai University School of Medicine, Kanagawa, Japan.

出版信息

Exp Hematol. 2004 Jan;32(1):95-103. doi: 10.1016/j.exphem.2003.10.008.

DOI:10.1016/j.exphem.2003.10.008

PMID:14725906

Abstract

OBJECTIVE

We generated cytomegalovirus (CMV)-specific cytotoxic T lymphocytes (CTL) in vitro using dendritic cells (DC) pulsed with crude CMV antigens (Ag).

PATIENTS AND METHODS

Mononuclear cells from healthy CMV-seropositive or seronegative volunteers and from stem cell transplant (SCT) recipients were cultured with CD14(+) monocyte-derived DC prepulsed with CMV Ag and then matured in vitro with lipopolysaccharide and tumor necrosis factor-alpha. After proliferation, cells were checked for phenotype (CD4/CD8), while killing activity was measured by 51Cr-release assay.

RESULTS

CD4(+) T cells, the main proliferating cells from both seropositive and seronegative individuals, killed autologous Ag-pulsed DC but not vehicle-pulsed autologous DC or CMV-pulsed allogeneic DC. Similar CTL induction was accomplished from SCT recipients. Significant killing of autologous CMV-infected fibroblasts required 16-hour incubation as opposed to the standard 4-hour incubation, which was prevented by either a perforin inhibitor or anti-Fas ligand monoclonal antibody. CTL enhanced surface HLA-DR expression of CMV-infected fibroblasts, and their activity was neutralized by anti-HLA-DR monoclonal antibody.

CONCLUSION

CMV-specific CD4(+) CTL were inducible with or without antiviral humoral immunity, even from immunosuppressed SCT recipients. These CTL showed perforin- and Fas/Fas ligand-mediated cytotoxicity after long-term (16-hour) contact with CMV-infected targets.

摘要

目的

我们使用经粗制巨细胞病毒（CMV）抗原（Ag）脉冲处理的树突状细胞（DC）在体外产生了CMV特异性细胞毒性T淋巴细胞（CTL）。

患者与方法

将来自健康CMV血清阳性或血清阴性志愿者以及干细胞移植（SCT）受者的单核细胞与预先用CMV Ag脉冲处理的CD14(+)单核细胞衍生DC一起培养，然后在体外使用脂多糖和肿瘤坏死因子-α使其成熟。增殖后，检查细胞的表型（CD4/CD8），同时通过51Cr释放试验测量杀伤活性。

结果

CD4(+) T细胞是血清阳性和血清阴性个体的主要增殖细胞，可杀伤自体Ag脉冲处理的DC，但不杀伤载体脉冲处理的自体DC或CMV脉冲处理的同种异体DC。从SCT受者中也实现了类似的CTL诱导。与标准的4小时孵育不同，对自体CMV感染的成纤维细胞进行显著杀伤需要16小时的孵育，而穿孔素抑制剂或抗Fas配体单克隆抗体可阻止这种杀伤。CTL增强了CMV感染的成纤维细胞表面HLA-DR的表达，并且其活性被抗HLA-DR单克隆抗体中和。