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Thy28 protects against anti-CD3-mediated thymic cell death in vivo

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Abstract

Apoptotic cell death plays a pivotal role in the development and/or maintenance of several tissues including thymus. Deregulated thymic cell death is associated with autoimmune diseases including experimental autoimmune encephalomyelitis (EAE), a prototype murine model for analysis of human multiple sclerosis. Because Thy28 expression is modulated during thymocyte development, we tested whether Thy28 affects induction of EAE as effectively as antigen-induced thymocyte deletion using Thy28 transgenic (TG) mice. Thy28 TG mice showed partial resistance to anti-CD3 monoclonal antibody (mAb)-induced thymic cell death in vivo, as assessed by annexin V-expression and loss of mitochondrial membrane potential. The resistance to anti-CD3 mAb-induced cell death in Thy28 TG mice appeared to correlate with a decreased c-Jun N-terminal kinase phosphorylation and reduced down-regulation of Bcl-xL. Moreover, thymic hyperplasia was detected in Thy28 TG mice, although thymocyte development was unaltered. Development of peripheral lymphoid tissues including spleen and lymph nodes was also unaltered. Thy28 TG spleen T cells showed an increased production of IFN-γ, but not IL-17, in response to both anti-CD3 and anti-CD28 mAbs. Finally, Thy28 TG mice displayed accelerated induction of EAE as assessed by disease incidence, clinical score, and pathology following immunization with myelin oligodendrocyte glycoprotein compared with control WT mice. These findings suggest that modulation of Thy28 expression plays a crucial role in the determination of thymic cell fate, which may contribute to the development of EAE through proinflammatory cytokine production.

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Abbreviations

DP:

Double positive

JNK:

c-Jun N-terminal kinase

EAE:

Experimental autoimmune encephalomyelitis

MS:

Multiple sclerosis

MOG:

Myelin oligodendrocyte glycoprotein

Th :

Helper T cell

TG:

Transgenic

WT:

Wild type

MAb:

Monoclonal antibody

MMP:

Mitochondrial membrane potential

AP-1:

Activator protein 1

CFA:

Complete Freund’s adjuvant

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Acknowledgments

We thank Dr. K. Miyake (Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan) for pCAGGS vector. This work was supported by a Grant from the Intractable Immune System Disease Center of Tokyo Medical University, which is supported by the Ministry of Education, Culture, Sports, Science, and Technology of Japan. We also thank Zenji Sakamoto for his support.

Conflict of interest

We declare no conflicts of interest.

Author information

Authors and Affiliations

  1. Department of Immunology and Intractable Immunology Research Center, Tokyo Medical University, 6-1-1 Shinjuku, Shinjuku-ku, Tokyo, 160-8402, Japan

    H. Toyota, N. Yanase & J. Mizuguchi

  2. Animal Research Center, Tokyo Medical University, 6-1-1 Shinjuku, Shinjuku-ku, Tokyo, 160-8402, Japan

    K. Sudo & K. Kojima

  3. Department of Anatomic Pathology, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku-ku, Tokyo, 160-0023, Japan

    T. Nagao, R. H. Takahashi & H. Iobe

  4. Department of Immunology, Toho University School of Medicine, 5-21-16, Omori-nishi, Ota-ku, Tokyo, 143-8540, Japan

    T. Kuwabara & T. Kakiuchi

  5. Department of Advanced and Integrated Analysis of Infectious Diseases, Toho University School of Medicine, 5-21-16, Omori-nishi, Ota-ku, Tokyo, 143-8540, Japan

    T. Kakiuchi

Authors
  1. H. Toyota
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  2. K. Sudo
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Corresponding author

Correspondence to H. Toyota.

Electronic supplementary material

Below is the link to the electronic supplementary material.

S1

Thy28 expression in Thy28-121 TG mice. Thy28 mRNA expression levels from Thy28 TG and control individual mice were determined by RT-PCR. Expression levels of GAPDH were also determined as control. The data are representative of three independent experiments. (PPTX 12 kb)

S2

Anti-CD3 mAb-induced reduction in proportion of DP thymocytes is abrogated in Thy28-121 TG mice. The percentage of DP cells from Thy28-121 or control mice after administration of 10 μg anti-CD3 mAb or control PBS was enumerated. The data are representative of three independent experiments. (PPTX 109 kb)

S3

Thy28-121 TG mice demonstrate an augmented clinical score, accompanied by increased cell infiltration and demyelination relative to control mice following MOG administration. (A) Thy28-121 TG (n=10) and control WT mice (n=10) were immunized with MOG and assessed by clinical score every 1–3 day up to day 25. (B) In a separate experiment, Thy28 TG (b and d, n=5) and WT mice (a and c, n=5) were assessed by disease incidence, clinical score, and pathology on day 25 after MOG immunization. Spinal sections stained with H & E (a and b) and Luxol fast blue (c and d). The location of cellular infiltration (b) and myelin sheath (d) was shown by arrows. (PPTX 128 kb)

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Toyota, H., Sudo, K., Kojima, K. et al. Thy28 protects against anti-CD3-mediated thymic cell death in vivo. Apoptosis 20, 444–454 (2015). https://doi.org/10.1007/s10495-014-1082-0

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