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Theranostics 2019; 9(13):3903-3917. doi:10.7150/thno.31037 This issue Cite

Research Paper

Cysteine-type cathepsins promote the effector phase of acute cutaneous delayed-type hypersensitivity reactions

Johannes Schwenck^1,2, Andreas Maurer^1,3, Birgit Fehrenbacher⁴, Roman Mehling¹, Philipp Knopf¹, Natalie Mucha¹, Dennis Haupt¹, Kerstin Fuchs¹, Christoph M. Griessinger¹, Daniel Bukala¹, Julia Holstein⁴, Martin Schaller⁴, Irene Gonzalez Menendez⁵, Kamran Ghoreschi^4,5, Leticia Quintanilla-Martinez⁶, Michael Gütschow⁷, Stefan Laufer⁸, Thomas Reinheckel^9,10, Martin Röcken⁴, Hubert Kalbacher³, Bernd J Pichler^1,11, Manfred Kneilling^1,4✉

1. Werner Siemens Imaging Center, Department of Preclinical Imaging and RadiopharmacyEberhard Karls University, 72076 Tübingen, Germany
2. Department of Nuclear Medicine and Clinical Molecular Imaging, Eberhard Karls University, 72076 Tübingen, Germany
3. Interfaculty Institute of Biochemistry, Eberhard Karls University, 72076 Tübingen, Germany
4. Department of Dermatology, Eberhard Karls University, 72076 Tübingen, Germany
5. Department of Dermatology, Venereology and Allergology, Charité - Universitätsmedizin Berlin, 10117 Berlin, Germany
6. Department of Pathology, Eberhard Karls University, 72076 Tübingen, Germany
7. Pharmaceutical Institute, Pharmaceutical Chemistry I, University of Bonn, 53121 Bonn, Germany
8. Department of Pharmaceutical and Medicinal Chemistry, Institute of Pharmacy, Eberhard Karls University, 72076 Tübingen, Germany
9. Institute of Molecular Medicine and Cell Research, Medical Faculty, Albert-Ludwigs University, 79085 Freiburg, Germany
10. German Cancer Consortium (DKTK), partner site Freiburg and German Cancer Research Center (DKFZ), Heidelberg, Germany
11. German Cancer Consortium (DKTK), partner site Tübingen and German Cancer Research Center (DKFZ), Heidelberg, Germany

✉ Corresponding author: email: manfred.kneillinguni-tuebingen.de; telephone: +49-7071-29-83427; fax: +49-7071-29-4451

Abstract

Cysteine-type cathepsins such as cathepsin B are involved in various steps of inflammatory processes such as antigen processing and angiogenesis. Here, we uncovered the role of cysteine-type cathepsins in the effector phase of T cell-driven cutaneous delayed-type hypersensitivity reactions (DTHR) and the implication of this role on therapeutic cathepsin B-specific inhibition.

Methods: Wild-type, cathepsin B-deficient (Ctsb^-/-) and cathepsin Z-deficient (Ctsz^-/-) mice were sensitized with 2,4,6-trinitrochlorobenzene (TNCB) on the abdomen and challenged with TNCB on the right ear to induce acute and chronic cutaneous DTHR. The severity of cutaneous DTHR was assessed by evaluating ear swelling responses and histopathology. We performed fluorescence microscopy on tissue from inflamed ears and lymph nodes of wild-type mice, as well as on biopsies from psoriasis patients, focusing on cathepsin B expression by T cells, B cells, macrophages, dendritic cells and NK cells. Cathepsin activity was determined noninvasively by optical imaging employing protease-activated substrate-like probes. Cathepsin expression and activity were validated ex vivo by covalent active site labeling of proteases and Western blotting.

Results: Noninvasive in vivo optical imaging revealed strong cysteine-type cathepsin activity in inflamed ears and draining lymph nodes in acute and chronic cutaneous DTHR. In inflamed ears and draining lymph nodes, cathepsin B was expressed by neutrophils, dendritic cells, macrophages, B, T and natural killer (NK) cells. Similar expression patterns were found in psoriatic plaques of patients. The biochemical methods confirmed active cathepsin B in tissues of mice with cutaneous DTHR. Topically applied cathepsin B inhibitors significantly reduced ear swelling in acute but not chronic DTHR. Compared with wild-type mice, Ctsb^-/- mice exhibited an enhanced ear swelling response during acute DTHR despite a lack of cathepsin B expression. Cathepsin Z, a protease closely related to cathepsin B, revealed compensatory expression in inflamed ears of Ctsb^-/- mice, while cathepsin B expression was reciprocally elevated in Ctsz^-/- mice.

Conclusion: Cathepsin B is actively involved in the effector phase of acute cutaneous DTHR. Thus, topically applied cathepsin B inhibitors might effectively limit DTHR such as contact dermatitis or psoriasis. However, the cathepsin B and Z knockout mouse experiments suggested a complementary role for these two cysteine-type proteases.

Keywords: inflammation, proteases, cathepsin B, optical imaging, delayed-type hypersensitivity

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