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Inhibition of calcium‐independent phospholipase A impairs agonist‐induced calcium entry in keratinocytes
Author(s) -
Ross K.,
Parker G.,
Whitaker M.,
Reynolds N.J.
Publication year - 2008
Publication title -
british journal of dermatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.304
H-Index - 179
eISSN - 1365-2133
pISSN - 0007-0963
DOI - 10.1111/j.1365-2133.2007.08298.x
Subject(s) - newcastle upon tyne , medicine , library science , classics , art history , art , computer science
Summary Background In many cells, depletion of intracellular calcium (Ca 2+ ) reservoirs triggers Ca 2+ entry through store‐operated Ca 2+ channels in the plasma membrane. However, the mechanisms of agonist‐induced calcium entry (ACE) in keratinocytes are not fully understood. Objectives This study was designed to determine if pharmacological inhibition of calcium‐independent phospholipase A (iPLA 2 ) impairs ACE in normal human epidermal keratinocytes. Methods Confocal laser scanning microscopy was used to monitor the dynamics of Ca 2+ signalling in keratinocytes loaded with the calcium‐sensitive dye Fluo‐4. Cells were stimulated with extracellular nucleotides [adenosine triphosphate (ATP) or uridine triphosphate (UTP)] or with lysophosphatidic acid (LPA), a bioactive lipid that regulates keratinocyte proliferation and differentiation. Results Both ATP and UTP induced Ca 2+ release in primary human keratinocytes. This was not followed by robust Ca 2+ influx when the experiments were performed in low Ca 2+ (70 μmol L −1 ) medium. Upon elevation of extracellular Ca 2+ to 1·2 mmol L −1 , however, a biphasic response consisting of an initial Ca 2+ peak followed by an elevated plateau was observed. The plateau phase was inhibited when cells were treated with bromoenol lactone, a specific pharmacological inhibitor of iPLA 2 . These findings indicate that iPLA 2 activity is required for ACE in keratinocytes. LPA also evoked Ca 2+ release in keratinocytes but failed to induce sustained Ca 2+ entry even when extracellular Ca 2+ was elevated to 1·2 mmol L −1 . Conclusion Our results demonstrate for the first time an important role for iPLA 2 in regulating ACE in primary human keratinocytes.