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Roles of human β-defensins in innate immune defense at the ocular surface: arming and alarming corneal and conjunctival epithelial cells

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Abstract

Human β-defensins are cationic peptides produced by epithelial cells that have been proposed to be an important component of immune function at mucosal surfaces. In this study, the expression and inducibility of β-defensins at the ocular surface were investigated in vitro and in vivo. Expression of human β-defensins (hBD) was determined by RT-PCR and immunohistochemistry in tissues of the ocular surface and lacrimal apparatus. Cultured corneal and conjunctival epithelial cells were stimulated with proinflammatory cytokines and supernatants of different ocular pathogens. Real-time PCR and ELISA experiments were performed to study the effect on the inducibility of hBD2 and 3. Expression and inducibility of mouse β-defensins-2, -3 and -4 (mBD2–4) were tested in a mouse ocular surface scratch model with and without treatment of supernatants of a clinical Staphylococcus aureus (SA) isolate by means of immunohistochemistry. Here we show that hBD1, -2, -3 and -4 are constitutively expressed in conjunctival epithelial cells and also partly in cornea. Healthy tissues of the ocular surface, lacrimal apparatus and human tears contain measurable amounts of hBD2 and -3, with highest concentrations in cornea and much lower concentrations in all other tissues, especially tears, suggesting intraepithelial storage of β-defensins. Exposure of cultured human corneal and conjunctival epithelial cells to proinflammatory cytokines and supernatants of various bacteria revealed that IL-1β is a very strong inductor of hBD2 and Staphylococcus aureus increases both hBD2 and hBD3 production in corneal and conjunctival epithelial cells. A murine corneal scratch model demonstrated that β-defensins are only induced if microbial products within the tear film come into contact with a defective epithelium. Our finding suggests that the tear film per se contains so much antimicrobial substances that epithelial induction of β-defensins occurs only as a result of ocular surface damage. These findings widen our knowledge of the distribution, amount and inducibility of β-defensins at the ocular surface and lacrimal apparatus and show how β-defensins are regulated specifically.

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Abbreviations

HCE:

Human corneal epithelial cells

HCjE:

Human conjunctival epithelial cells

IL:

Interleukin

TNF:

Tumor necrosis factor

hBD1:

Human β-defensin 1

hBD2:

Human β-defensin 2

hBD3:

Human β-defensin 3

hBD4:

Human β-defensin 4

mBD2:

Mouse β-defensin 2

mBD3:

Mouse β-defensin 3

mBD4:

Mouse β-defensin 4

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Acknowledgments

We thank Ute Beyer, Stephanie Beilecke, Susann Möschter, Karin Stengel and Regine Worm for excellent technical assistance and Dr. Bernhard Nölle (Department of Ophthalmology, Christian Albrecht University of Kiel) for providing us with clinical samples. Moreover, we are grateful to Deike Varoga (Department of Trauma Surgery, University Hospital of Schleswig-Holstein, Campus Kiel, Germany) for helpful advice and discussion with regard to the corneal scratch model as well as Ute Schulze and Sylvia Dyczek for their help with the translation. This work was supported by the Deutsche Forschungsgemeinschaft (DFG)–program grants PA 738/9-1 and PA 738/9-2, BMBF–Wilhelm Roux Program, Halle, Germany–program grants FKZ 09/16, 14/25, and 16/35, as well as Sicca Forschungsförderung of the professional Association of German Ophthalmologists.

Conflict of interest statement

The authors have no financial conflict of interest.

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Authors and Affiliations

  1. Department of Anatomy and Cell Biology, Martin Luther University of Halle-Wittenberg, Große Steinstraße 52, 06108, Halle (Saale), Germany

    Fabian Garreis, Thomas Schlorf, Lars Bräuer, Kristin Jäger & Friedrich P. Paulsen

  2. Department of Hygiene, Martin Luther University of Halle-Wittenberg, Große Steinstraße 52, 06108, Halle (Saale), Germany

    Dieter Worlitzsch

  3. Department of Ophthalmology, University Medical Center Schleswig-Holstein, Campus Lübeck, Ratzeburger Allee 160, 23538, Lübeck, Germany

    Philipp Steven

  4. Department of Anatomy II, Friedrich Alexander University Erlangen-Nürnberg, Universitätsst. 19, 91054, Erlangen, Germany

    Fabian Garreis & Friedrich P. Paulsen

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  1. Fabian Garreis
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Correspondence to Fabian Garreis or Friedrich P. Paulsen.

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Garreis, F., Schlorf, T., Worlitzsch, D. et al. Roles of human β-defensins in innate immune defense at the ocular surface: arming and alarming corneal and conjunctival epithelial cells. Histochem Cell Biol 134, 59–73 (2010). https://doi.org/10.1007/s00418-010-0713-y

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