Effects of 1α,25-(OH)2D3 on rat growth zone chondrocytes are mediated via cyclooxygenase-1 and phospholipase A2

V. L. Sylvia, F. Del Toro, D. D. Dean, R. R. Hardin, Z. Schwartz, B. D. Boyan

Research output: Contribution to journalArticle

26 Scopus citations

Abstract

1α,25-(OH)2D3 mediates its effects on growth zone chondrocytes via rapid membrane-associated events as well as through traditional nuclear receptor mechanisms. The membrane-associated signaling pathways include rapid production of diacylglycerol and activation of protein kinase C (PKC), as well as activation of phospholipase A2 (PLA2), increased production of arachidonic acid, and increased production of prostaglandins. This study examined the roles of PLA2 and cyclooxygenase (Cox) in the mechanism of action of 1α,25-(OH)2D3 in these cells to determine whether one or both enzymes catalyze the rate limiting step and whether constitutive or inducible Cox is involved. Cultures were incubated with 1α,25-(OH)2D3 for 9 min to measure PKC or for 24 h to measure physiological responses ([3H]-thymidine incorporation, alkaline phosphatase specific activity, [35S]-sulfate incorporation). Based on RT-PCR and Northern blot analysis, growth zone chondrocytes expressed mRNAs for both Cox-1 and Cox-2 and neither Cox was modulated by 1α,25-(OH)2D3. To examine the role of Cox, the cultures were also treated with resveratrol (a specific inhibitor of Cox-1), NS-398 (a specific inhibitor of Cox-2), or indomethacin (a general Cox inhibitor). The results showed that Cox-1 inhibition reduced the 1α,25-(OH)2D3-dependent effects on proliferation, differentiation, and matrix production, whereas inhibition of Cox-2 only had an effect on proliferation. The effects of Cox inhibition were not rate limiting, based on experiments in which PLA2 was activated with melittin or inhibited with quinacrine. However, at least part of the action of 1α,25-(OH)2D3 was regulated by metabolism of arachidonic acid to prostaglandins. This supports the hypothesis that 1α,25-(OH)2D3 exerts its effects via more than one signaling pathway and that these pathways are interrelated via the modulation of PLA2 as a rate-limiting step. PKC regulation may occur at multiple stages in the signal transduction cascade.

Original languageEnglish (US)
Pages (from-to)32-45
Number of pages14
JournalJournal of Cellular Biochemistry
Volume81
DOIs
StatePublished - Jan 1 2001

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Keywords

  • 1,25-(OH)D
  • Alkaline phosphatase
  • Chondrocyte cultures
  • Cox-1
  • Cox-2
  • Cyclooxygenase
  • Membrane receptor
  • PKC
  • Protein kinase C

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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