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Journal of Clinical Immunology

, Volume 37, Issue 1, pp 92–99 | Cite as

A CD57+ CTL Degranulation Assay Effectively Identifies Familial Hemophagocytic Lymphohistiocytosis Type 3 Patients

  • Masayuki Hori
  • Takahiro YasumiEmail author
  • Saeko Shimodera
  • Hirofumi Shibata
  • Eitaro Hiejima
  • Hirotsugu Oda
  • Kazushi Izawa
  • Tomoki Kawai
  • Masataka Ishimura
  • Naoko Nakano
  • Ryutaro Shirakawa
  • Ryuta Nishikomori
  • Hidetoshi Takada
  • Satoshi Morita
  • Hisanori Horiuchi
  • Osamu Ohara
  • Eiichi Ishii
  • Toshio Heike
Original Article

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.

Keywords

Familial hemophagocytic lymphohistiocytosis type 3 lysosomal degranulation defect functional screening assay UNC13D 

Notes

Acknowledgements

The authors are grateful to all of the participating patients, their families, and the referring physicians for their generous cooperation.

Authorship Contributions

Contribution: T.Y., R.N., and T.H. designed the research; M.I., N.N., H.T., and E.I. treated patients 1 and 2; M.H., S.S., H.S., E.H., K.I., and T.K. performed the degranulation and protein expression assays; H.O. and O.O. performed the genetic analyses; R.S. and H.H. prepared the anti-Munc13-4 and anti-Syntaxin11 antibodies; M.H., T.Y., K.I., T.K., R.N., S.M., and T.H. analyzed and discussed the results; T.Y. and S.M. performed the statistical analysis; and M.H. and T.Y. wrote the paper.

Compliance with Ethical Standards

Informed consent was obtained from the patients and their parents in accordance with the institutional review board of Kyoto University Hospital and the Declaration of Helsinki.

Conflict of Interest

The authors declare that they have no competing interests.

Funding

This work was supported by JSPS KAKENHI (Grant Numbers 26461582 and 25670475) and by grants from the “Research on Measures for Intractable Diseases” Project: matching fund subsidy from the Japanese Ministry of Health, Labor, and Welfare.

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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Masayuki Hori
    • 1
  • Takahiro Yasumi
    • 1
    Email author
  • Saeko Shimodera
    • 1
  • Hirofumi Shibata
    • 1
  • Eitaro Hiejima
    • 1
  • Hirotsugu Oda
    • 1
    • 2
  • Kazushi Izawa
    • 1
  • Tomoki Kawai
    • 1
  • Masataka Ishimura
    • 3
  • Naoko Nakano
    • 4
  • Ryutaro Shirakawa
    • 5
  • Ryuta Nishikomori
    • 1
  • Hidetoshi Takada
    • 3
  • Satoshi Morita
    • 6
  • Hisanori Horiuchi
    • 5
  • Osamu Ohara
    • 2
    • 7
  • Eiichi Ishii
    • 4
  • Toshio Heike
    • 1
  1. 1.Department of PediatricsKyoto University Graduate School of MedicineKyotoJapan
  2. 2.Laboratory for Integrative GenomicsRIKEN Center for Integrative Medical SciencesYokohamaJapan
  3. 3.Department of Pediatrics, Graduate School of Medical SciencesKyushu UniversityFukuokaJapan
  4. 4.Department of PediatricsEhime University Graduate School of MedicineToonJapan
  5. 5.Department of Molecular and Cellular Biology, Institute of Development, Aging and CancerTohoku UniversitySendaiJapan
  6. 6.Department of Biomedical Statistics and BioinformaticsKyoto University Graduate School of MedicineKyotoJapan
  7. 7.KAZUSA DNA Research InstituteKisarazuJapan

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