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Use of human reconstructed epidermis to analyze the regulation of -defensin hBD-1, hBD-2, and hBD-3 expression in response to LPS

Abstract : Defensins have been identified as key elements of innate immunity against microbial infections. In the present study, human beta-defensin-2 (hBD-2) mRNA and peptide expression were evaluated by RT-PCR and Western blotting in normal human keratinocytes, in function of their stage of differentiation. In proliferating, non-differentiating keratinocytes generated in serum-free, low-calcium medium, a very low hBD-2 mRNA expression was found. A significantly higher expression was detected in high-calcium cultivated keratinocytes grown either as monolayers or as multilayers under submerged conditions. In an air-liquid interface culture of keratinocytes, allowing epidermis to be reconstructed, hBD-2 mRNA expression level was significantly higher than in the other conditions and displayed inter-individual variability as observed in native epidermis. The peptide was detected only in reconstructed epidermis. These results indicate that hBD-2 gene expression in normal human keratinocytes is dependent upon their stage of differentiation. The level of expression of hBD-1 mRNA was lower and that of hBD-3 was higher than that of hBD-2 in reconstructed epidermis. Exposure of reconstructed epidermis to bacterial lipopolysaccharide (LPS) resulted in an average 4-fold increase in hBD-2 mRNA 18 h after challenge, but not of hBD-1 and hBD-3 gene expression. These results show the selective regulation of hBD-2-encoding gene in an organotypic epidermal model, in response to LPS. They also provide evidence that in vitro reconstructed epidermis represents a useful model for studying regulation of expression of beta-defensins after skin challenge with pathogenic microorganisms in conditions as close as possible to the in vivo situation.
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Contributor : Philippe CHADEBECH Connect in order to contact the contributor
Submitted on : Friday, May 28, 2021 - 5:52:53 PM
Last modification on : Wednesday, June 2, 2021 - 3:03:11 AM

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P. Chadebech, D. Goidin, C. Jacquet, J. Viac, D. Schmitt, et al.. Use of human reconstructed epidermis to analyze the regulation of -defensin hBD-1, hBD-2, and hBD-3 expression in response to LPS. Cell Biology and Toxicology, Springer Verlag, 2003, 19 (5), pp.313-324. ⟨10.1023/b:cbto.0000004975.36521.c8⟩. ⟨inserm-03241614⟩



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