Masayuki Hori1, Takahiro Yasumi2, Saeko Shimodera1, Hirofumi Shibata1, Eitaro Hiejima1, Hirotsugu Oda1,3, Kazushi Izawa1, Tomoki Kawai1, Masataka Ishimura4, Naoko Nakano5, Ryutaro Shirakawa6, Ryuta Nishikomori1, Hidetoshi Takada4, Satoshi Morita7, Hisanori Horiuchi6, Osamu Ohara3,8, Eiichi Ishii5, Toshio Heike1. 1. Department of Pediatrics, Kyoto University Graduate School of Medicine, 54 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto, 606-8507, Japan. 2. Department of Pediatrics, Kyoto University Graduate School of Medicine, 54 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto, 606-8507, Japan. yasumi@kuhp.kyoto-u.ac.jp. 3. Laboratory for Integrative Genomics, RIKEN Center for Integrative Medical Sciences, Yokohama, Japan. 4. Department of Pediatrics, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan. 5. Department of Pediatrics, Ehime University Graduate School of Medicine, Toon, Japan. 6. Department of Molecular and Cellular Biology, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan. 7. Department of Biomedical Statistics and Bioinformatics, Kyoto University Graduate School of Medicine, Kyoto, Japan. 8. KAZUSA DNA Research Institute, Kisarazu, Japan.
Abstract
PURPOSE: Familial hemophagocytic lymphohistiocytosis type 3 (FHL3) is a genetic disorder that results in immune dysregulation. It requires prompt and accurate diagnosis. A natural killer (NK) cell degranulation assay is often used to screen for FHL3 patients. However, we recently encountered two cases of late-onset FHL3 carrying novel UNC13D missense mutations: in these cases, the degranulation assays using freshly isolated and interleukin (IL)-2-activated NK cells yielded contradictory results. Since the defective degranulation of CD57+ cytotoxic T lymphocytes (CTLs) in these cases was helpful for making the diagnosis, we assessed whether the CD57+ CTL degranulation assay more effectively identified FHL3 patients than the NK cell assays. METHODS: Forty additional patients with hemophagocytic lymphohistiocytosis were prospectively screened for FHL3 by measuring the perforin expression in NK cells and the expression of Munc13-4, syntaxin-11, and Munc18-2 in platelets and by performing NK cell and CTL degranulation assays. The results were confirmed by genetic analysis. RESULTS: The freshly isolated NK cell degranulation assay detected FHL3 patients with high sensitivity (100%) but low specificity (71%). The IL-2-stimulated NK cell assay had improved specificity, but 3 out of the 31 non-FHL3 patients still showed degranulation below the threshold level. The CD57+ CTL degranulation assay identified FHL3 patients with high sensitivity and specificity (both 100%). CONCLUSIONS: The CD57+ CTL degranulation assay more effectively identified FHL3 patients than the NK cell-based assays.
PURPOSE:Familial hemophagocytic lymphohistiocytosis type 3 (FHL3) is a genetic disorder that results in immune dysregulation. It requires prompt and accurate diagnosis. A natural killer (NK) cell degranulation assay is often used to screen for FHL3 patients. However, we recently encountered two cases of late-onset FHL3 carrying novel UNC13D missense mutations: in these cases, the degranulation assays using freshly isolated and interleukin (IL)-2-activated NK cells yielded contradictory results. Since the defective degranulation of CD57+ cytotoxic T lymphocytes (CTLs) in these cases was helpful for making the diagnosis, we assessed whether the CD57+ CTL degranulation assay more effectively identified FHL3 patients than the NK cell assays. METHODS: Forty additional patients with hemophagocytic lymphohistiocytosis were prospectively screened for FHL3 by measuring the perforin expression in NK cells and the expression of Munc13-4, syntaxin-11, and Munc18-2 in platelets and by performing NK cell and CTL degranulation assays. The results were confirmed by genetic analysis. RESULTS: The freshly isolated NK cell degranulation assay detected FHL3 patients with high sensitivity (100%) but low specificity (71%). The IL-2-stimulated NK cell assay had improved specificity, but 3 out of the 31 non-FHL3 patients still showed degranulation below the threshold level. The CD57+ CTL degranulation assay identified FHL3 patients with high sensitivity and specificity (both 100%). CONCLUSIONS: The CD57+ CTL degranulation assay more effectively identified FHL3 patients than the NK cell-based assays.
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