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Inhibition of interferon-gamma-induced intercellular adhesion molecule-1 expression on human keratinocytes by
M Hertl1, L M Neckers, S I Katz
1Dermatology Branch, National Cancer Institute, Bethesda, Maryland 20892, USA.
Abstract:
Expression of intercellular adhesion molecule-1 (ICAM-1) by keratinocytes is an important event in the pathogenesis of T-cell-mediated inflammatory skin diseases. To determine if ICAM-1 expression could be selectively modulated, two antisense phosphorothioate oligonucleotides (S-ODN) targeting the translation initiation and 3' untranslated regions of ICAM-1 mRNA were added as lipid complexes to cultures of keratinocytes. Interferon-gamma was added after 24 h to induce ICAM-1 expression, which was quantitated by flow cytometry after 48 h. The S-ODN targeting the translation initiation site did not inhibit ICAM-1 expression at 0.2-20.0 microM. In contrast, 0.2-1.0 microM of the S-ODN targeting a site in the 3' untranslated region abrogated ICAM-1 expression in up to 75% of the keratinocytes; this inhibition was reversible when complementary sense S-ODN was added. Phosphodiester ODN (PD-ODN) targeting the same sites did not inhibit ICAM-1 expression on keratinocytes, most likely as a consequence of rapid degradation. Inhibition of ICAM-1 by the antisense S-ODN was selective; expression of beta 2-microglobulin, alpha 3-integrin, and beta 1-integrin remained largely unaffected and interferon-gamma-induced HLA-DR expression was inhibited to a much lesser extent than ICAM-1. Antisense-non-specific inhibition was also noted in that two scrambled S-ODN with an identical nucleotide (14 of 20 cytosines) composition inhibited ICAM-1 expression in up to 44% of the keratinocytes, whereas a degenerate S-ODN did not. The data demonstrate the complex effects exerted by antisense S-ODN in that ICAM-1 expression was inhibited via antisense-non-specific mechanisms probably due to the intrinsic properties of the S-ODN as well as via the anticipated sequence-specific mechanisms.
Insights
Antisense oligonucleotides targeting intercellular adhesion molecule-1 (ICAM-1) mRNA can inhibit its expression in skin cells. A specific S-ODN in the 3' untranslated region effectively reduced ICAM-1, demonstrating potential for treating inflammatory skin diseases.
Area of Science:
- Dermatology
- Molecular Biology
- Immunology
Background:
- Intercellular adhesion molecule-1 (ICAM-1) expression on keratinocytes is crucial in T-cell-mediated inflammatory skin diseases.
- Targeting ICAM-1 offers a potential therapeutic strategy for these conditions.
Purpose of the Study:
- To investigate the selective modulation of ICAM-1 expression in keratinocytes using antisense oligonucleotides (S-ODN).
- To assess the efficacy of S-ODN targeting different regions of ICAM-1 mRNA.
Main Methods:
- Two S-ODN targeting ICAM-1 mRNA (translation initiation and 3' untranslated regions) were introduced into keratinocyte cultures via lipid complexes.
- Interferon-gamma was used to induce ICAM-1 expression, followed by quantification using flow cytometry.
- Control experiments included phosphodiester ODN (PD-ODN), sense ODN, and scrambled S-ODN.
Main Results:
- An S-ODN targeting the 3' untranslated region of ICAM-1 mRNA inhibited expression in up to 75% of keratinocytes.
- This inhibition was sequence-specific and reversible, with minimal impact on other cell surface molecules like beta 2-microglobulin and integrins.
- Non-specific inhibition was observed with some scrambled S-ODN, indicating complex mechanisms of action.
Conclusions:
- Antisense S-ODN can selectively inhibit ICAM-1 expression in keratinocytes, particularly when targeting the 3' untranslated region.
- The findings suggest potential for developing targeted therapies for inflammatory skin conditions by modulating ICAM-1.
- Both sequence-specific and non-specific mechanisms contribute to the observed effects of S-ODN.