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Granzyme B-dependent and perforin-independent DNA fragmentation in intestinal epithelial cells induced by anti-CD3 mAb-activated intra-epithelial lymphocytes

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Abstract

We previously found that an i.p. injection of anti-CD3 monoclonal antibody (mAb) into mice caused DNA fragmentation in the intestinal villous epithelial cells (IVECs) of the duodenum and the jejunum. In this study, in order to elucidate the mechanism of DNA fragmentation in IVECs, we searched for the inducer(s) of DNA fragmentation by using immunohistochemistry. The release of cytoplasmic granules from intraepithelial lymphocytes (IELs) and the formation of large gaps between IELs and IVECs were observed electron microscopically after antibody administration. The presence and distribution pattern of Granzyme B (GrB), a serine protease in cytolytic granules present in cytotoxic T lymphocytes and natural killer cells and considered to be the responsible molecule for DNA fragmentation in target cells, was examined in detail in intestinal villi by immunohistology. GrB was detected in cytoplasmic granules in nearly all IELs. The time-kinetics of granule release from IELs after mAb injection coincided not only with that of the extracellular diffusion of GrB, but also with that of DNA fragmentation in IVECs. On the other hand, perforin (Pfn), assumed to cooperate with GrB in DNA fragmentation, could not be detected in IELs, and its release was not confirmed after the anti-CD3 mAb injection. Anti-CD3 mAb injection also induced DNA fragmentation in IVECs in Pfn-knockout mice. These results support the notion that DNA fragmentation in IVECs by the stimulated IELs in the present study is induced by a mechanism involving GrB, but independent of Pfn.

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Authors and Affiliations

  1. Division of Immunology and Embryology, Department of Cell Biology, Tohoku University School of Medicine, 980-8575, Sendai, Japan

    Masaki Ogata, Yuta Ota & Tsunetoshi Itoh

  2. Laboratory of Immune Regulation, Wakayama Medical University, Osaka, Japan

    Takaji Matsutani

  3. Department of Applied Research, Division of Immunology, Yakult Central Institute for Microbiological Research, Tokyo, Japan

    Masanobu Nanno

  4. Department of Rheumatology and Clinical Immunology, Clinical Research Center for Allergy and Rheumatology, Sagamihara National Hospital, Sagamihara, Japan

    Ryuji Suzuki

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  1. Masaki Ogata
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  2. Yuta Ota
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  3. Takaji Matsutani
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  4. Masanobu Nanno
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  5. Ryuji Suzuki
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  6. Tsunetoshi Itoh
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Correspondence to Masaki Ogata.

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This work was in part supported by a Grant-in-aid for Scientific Research from the Ministry of Education, Science and Culture, Japan (20590181 to M.O. and 21590207 to T.I.).

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Ogata, M., Ota, Y., Matsutani, T. et al. Granzyme B-dependent and perforin-independent DNA fragmentation in intestinal epithelial cells induced by anti-CD3 mAb-activated intra-epithelial lymphocytes. Cell Tissue Res 352, 287–300 (2013). https://doi.org/10.1007/s00441-012-1549-7

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