Hosono Yuki, Tomiyasu Noriyuki, Kasai Hayato, Ishikawa Eri, Takahashi Masatomo, Imamura Akihiro, Ishida Hideharu, Compostella Federica, Kida Hiroshi, Kumanogoh Atsushi, Bamba Takeshi, Izumi Yoshihiro, Yamasaki Sho
Department of Molecular Immunology, Research Institute for Microbial Diseases, Osaka University, Suita, Japan.
Laboratory of Molecular Immunology, Immunology Frontier Research Center, Osaka University , Suita, Japan.
J Exp Med. 2025 Feb 3;222(2). doi: 10.1084/jem.20240728. Epub 2024 Dec 20.
Invariant natural killer T (iNKT) cells are unconventional T cells recognizing lipid antigens in a CD1d-restricted manner. Among these lipid antigens, α-galactosylceramide (α-GalCer), which was originally identified in marine sponges, is the most potent antigen. Although the presence of α-anomeric hexosylceramide and microbiota-derived branched α-GalCer is reported, antigenic α-GalCer has not been identified in mammals. Here, we developed a high-resolution separation and detection system, supercritical fluid chromatography tandem mass spectrometry (SFC/MS/MS), that can discriminate hexosylceramide diastereomers (α-GalCer, α-GlcCer, β-GalCer, or β-GlcCer). The B16 melanoma tumor cell line does not activate iNKT cells; however, ectopic expression of CD1d was sufficient to activate iNKT cells without adding antigens. B16 melanoma was unlikely to generate iNKT cell antigens; instead, antigen activity was detected in cell culture serum. Activity-based purification and SFC/MS/MS identified dihydrosphingosine-based saturated α-GalCer as an antigenic component in serum, bile, and lymphoid tissues. These results show the first evidence for the presence of potent antigenic α-GalCer in mammals.
不变自然杀伤T（iNKT）细胞是一类非传统的T细胞，以CD1d限制性方式识别脂质抗原。在这些脂质抗原中，最初在海洋海绵中发现的α-半乳糖神经酰胺（α-GalCer）是最有效的抗原。尽管有报道称存在α-异头物己糖神经酰胺和微生物群衍生的分支α-GalCer，但在哺乳动物中尚未鉴定出具有抗原性的α-GalCer。在此，我们开发了一种高分辨率分离和检测系统，即超临界流体色谱串联质谱（SFC/MS/MS），它可以区分己糖神经酰胺非对映异构体（α-GalCer、α-GlcCer、β-GalCer或β-GlcCer）。B16黑色素瘤肿瘤细胞系不会激活iNKT细胞；然而，CD1d的异位表达足以在不添加抗原的情况下激活iNKT细胞。B16黑色素瘤不太可能产生iNKT细胞抗原；相反，在细胞培养血清中检测到了抗原活性。基于活性的纯化和SFC/MS/MS鉴定出基于二氢鞘氨醇的饱和α-GalCer是血清、胆汁和淋巴组织中的一种抗原成分。这些结果首次证明了哺乳动物中存在具有强抗原性的α-GalCer。
