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Dynamic compression counteracts IL-1B inducible nitric synthase and cyclo-oxygenese-2 expression in chondrocyte/agarose constructs

Dynamic compression counteracts IL-1B inducible nitric synthase and cyclo-oxygenese-2 expression in chondrocyte/agarose constructs
Dynamic compression counteracts IL-1B inducible nitric synthase and cyclo-oxygenese-2 expression in chondrocyte/agarose constructs
Background: Nitric oxide and prostaglandin E2 (PGE2play pivotal roles in both the pathogenesis of osteoarthritis and catabolic processes in articular cartilage. These mediators are influenced by both IL-1? and mechanical loading, and involve alterations in the inducible nitric oxide synthase (iNOS) and cyclo-oxygenase (COX)-2 enzymes. To identify the specific interactions that are activated by both types of stimuli, we examined the effects of dynamic compression on levels of expression of iNOS and COX-2 and involvement of the p38 mitogen-activated protein kinase (MAPK) pathway.

Methods: Chondrocyte/agarose constructs were cultured under free-swelling conditions with or without IL-1? and/or SB203580 (inhibitor of p38 MAPK) for up to 48 hours. Using a fully characterized bioreactor system, constructs were subjected to dynamic compression for 6, 12 and 48 hours under similar treatments. The activation or inhibition of p38 MAPK by IL-1? and/or SB203580 was analyzed by western blotting. iNOS, COX-2, aggrecan and collagen type II signals were assessed utilizing real-time quantitative PCR coupled with molecular beacons. Release of nitrite and PGE2 was quantified using biochemical assays. Two-way analysis of variance and the post hoc Bonferroni-corrected t-test were used to examine data.

Results: IL-1? activated the phosphorylation of p38 MAPK and this effect was abolished by SB203580. IL-1? induced a transient increase in iNOS expression and stimulated the production of nitrite release. Stimulation by either dynamic compression or SB203580 in isolation reduced the IL-1? induced iNOS expression and nitrite production. However, co-stimulation with both dynamic compression and SB203580 inhibited the expression levels of iNOS and production of nitrite induced by the cytokine. IL-1? induced a transient increase in COX-2 expression and stimulated the cumulative production of PGE2 release. These effects were inhibited by dynamic compression or SB203580. Co-stimulation with both dynamic compression and SB203580 restored cytokine-induced inhibition of aggrecan expression. This is in contrast to collagen type II, in which we observed no response with the cytokine and/or SB203580.

Conclusion: These data suggest that dynamic compression directly influences the expression levels of iNOS and COX-2. These molecules are current targets for pharmacological intervention, raising the possibility for integrated pharmacological and biophysical therapies for the treatment of cartilage joint disorders.
1478-6354
R35-[13pp]
Chowdhury, T.T.
3969bc1d-acec-4cad-b523-c8b4885fcb0a
Arghandawi, S.
8f25fb82-48ae-4851-b07f-563fa45eb6aa
Brand, J.
819943f0-2c7f-495f-bc84-962f278d748e
Akanji, O.O.
49ec1295-ca48-4e46-bc0d-fe9d3efd5e14
Bader, D.L.
9884d4f6-2607-4d48-bf0c-62bdcc0d1dbf
Salter, D.M.
94485705-ebda-4ddf-96c9-55fc6c3fe073
Lee, D.A.
fbbf7169-d08b-4deb-ae87-a2cbd97c58e7
Chowdhury, T.T.
3969bc1d-acec-4cad-b523-c8b4885fcb0a
Arghandawi, S.
8f25fb82-48ae-4851-b07f-563fa45eb6aa
Brand, J.
819943f0-2c7f-495f-bc84-962f278d748e
Akanji, O.O.
49ec1295-ca48-4e46-bc0d-fe9d3efd5e14
Bader, D.L.
9884d4f6-2607-4d48-bf0c-62bdcc0d1dbf
Salter, D.M.
94485705-ebda-4ddf-96c9-55fc6c3fe073
Lee, D.A.
fbbf7169-d08b-4deb-ae87-a2cbd97c58e7

Chowdhury, T.T., Arghandawi, S., Brand, J., Akanji, O.O., Bader, D.L., Salter, D.M. and Lee, D.A. (2008) Dynamic compression counteracts IL-1B inducible nitric synthase and cyclo-oxygenese-2 expression in chondrocyte/agarose constructs. Arthritis Research & Therapy, 10 (2), R35-[13pp]. (doi:10.1186/ar2389). (PMID:18348730)

Record type: Article

Abstract

Background: Nitric oxide and prostaglandin E2 (PGE2play pivotal roles in both the pathogenesis of osteoarthritis and catabolic processes in articular cartilage. These mediators are influenced by both IL-1? and mechanical loading, and involve alterations in the inducible nitric oxide synthase (iNOS) and cyclo-oxygenase (COX)-2 enzymes. To identify the specific interactions that are activated by both types of stimuli, we examined the effects of dynamic compression on levels of expression of iNOS and COX-2 and involvement of the p38 mitogen-activated protein kinase (MAPK) pathway.

Methods: Chondrocyte/agarose constructs were cultured under free-swelling conditions with or without IL-1? and/or SB203580 (inhibitor of p38 MAPK) for up to 48 hours. Using a fully characterized bioreactor system, constructs were subjected to dynamic compression for 6, 12 and 48 hours under similar treatments. The activation or inhibition of p38 MAPK by IL-1? and/or SB203580 was analyzed by western blotting. iNOS, COX-2, aggrecan and collagen type II signals were assessed utilizing real-time quantitative PCR coupled with molecular beacons. Release of nitrite and PGE2 was quantified using biochemical assays. Two-way analysis of variance and the post hoc Bonferroni-corrected t-test were used to examine data.

Results: IL-1? activated the phosphorylation of p38 MAPK and this effect was abolished by SB203580. IL-1? induced a transient increase in iNOS expression and stimulated the production of nitrite release. Stimulation by either dynamic compression or SB203580 in isolation reduced the IL-1? induced iNOS expression and nitrite production. However, co-stimulation with both dynamic compression and SB203580 inhibited the expression levels of iNOS and production of nitrite induced by the cytokine. IL-1? induced a transient increase in COX-2 expression and stimulated the cumulative production of PGE2 release. These effects were inhibited by dynamic compression or SB203580. Co-stimulation with both dynamic compression and SB203580 restored cytokine-induced inhibition of aggrecan expression. This is in contrast to collagen type II, in which we observed no response with the cytokine and/or SB203580.

Conclusion: These data suggest that dynamic compression directly influences the expression levels of iNOS and COX-2. These molecules are current targets for pharmacological intervention, raising the possibility for integrated pharmacological and biophysical therapies for the treatment of cartilage joint disorders.

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e-pub ahead of print date: 18 March 2008
Published date: 2008
Organisations: Health Sciences

Identifiers

Local EPrints ID: 169057
URI: https://eprints.soton.ac.uk/id/eprint/169057
ISSN: 1478-6354
PURE UUID: f9c4cdca-9108-4059-8e5d-2503921eacb6
ORCID for D.L. Bader: ORCID iD orcid.org/0000-0002-1208-3507

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Date deposited: 09 Dec 2010 09:45
Last modified: 29 Oct 2019 01:42

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