Cyclic AMP-dependent protein kinase A plays a role in the directed migration of human keratinocytes in a DC electric field

Christine E. Pullar, Roslyn Rivkah Isseroff, Richard Nuccitelli

Research output: Contribution to journalArticle

57 Citations (Scopus)

Abstract

Skin wound healing requires epithelial cell migration for re-epithelialization, wound closure, and re-establishment of normal function. We believe that one of the earliest signals to initiate wound healing is the lateral electric field generated by the wound current. Normal human epidermal keratinocytes migrate towards the negative pole, representing the center of the wound, in direct currents of a physiological strength, 100 mV/mm. Virtually nothing is known about the signal transduction mechanisms used by these cells to sense the endogenous electric field. To elucidate possible protein kinase (PK) involvement in the process, PK inhibitors were utilized. Two important findings have been described. Firstly, addition of 50 nM KT5720, an inhibitor of PKA, resulted in a 53% percent reduction in the directional response of keratinocytes in the electric field, while not significantly affecting general cell motility. The reduction was dose-dependent, there was a gradual decrease in the directional response from 5 to 50 nM. Secondly, addition of 1 μM ML-7, a myosin light chain kinase inhibitor, resulted in an approximate 31% decrease in the distance the cells migrated without affecting directional migration. The PKC inhibitors GF109203X at 4 μM and H-7 at 20 μM and W-7, a CaM kinase inhibitor, did not significantly alter either directed migration or cell migration, although they all resulted in a slight reduction in directional migration. D-erythro-sphingosine at 15 μM, a PKC inhibitor, had virtually no effect on either migration distance or directed migration. These findings demonstrate that divergent kinase signaling pathways regulate general cell motility and sustained directional migration and highlight the complexity of the signal transduction mechanisms involved. The inhibitor studies described in this paper implicate a role for PKA in the regulation of the directional migratory response to applied electric fields, galvanotaxis.

Original languageEnglish (US)
Pages (from-to)207-217
Number of pages11
JournalCell Motility and the Cytoskeleton
Volume50
Issue number4
DOIs
StatePublished - 2001

Fingerprint

Cyclic AMP-Dependent Protein Kinases
Keratinocytes
Cell Movement
Wound Healing
Signal Transduction
Wounds and Injuries
Phosphotransferases
Re-Epithelialization
1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine
Myosin-Light-Chain Kinase
Protein Kinase Inhibitors
Protein Kinases
Epithelial Cells
Skin

Keywords

  • Galvanotaxis
  • Keratinocyte
  • Motility
  • Protein kinase A
  • Wound healing

ASJC Scopus subject areas

  • Cell Biology

Cite this

Cyclic AMP-dependent protein kinase A plays a role in the directed migration of human keratinocytes in a DC electric field. / Pullar, Christine E.; Isseroff, Roslyn Rivkah; Nuccitelli, Richard.

In: Cell Motility and the Cytoskeleton, Vol. 50, No. 4, 2001, p. 207-217.

Research output: Contribution to journalArticle

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AB - Skin wound healing requires epithelial cell migration for re-epithelialization, wound closure, and re-establishment of normal function. We believe that one of the earliest signals to initiate wound healing is the lateral electric field generated by the wound current. Normal human epidermal keratinocytes migrate towards the negative pole, representing the center of the wound, in direct currents of a physiological strength, 100 mV/mm. Virtually nothing is known about the signal transduction mechanisms used by these cells to sense the endogenous electric field. To elucidate possible protein kinase (PK) involvement in the process, PK inhibitors were utilized. Two important findings have been described. Firstly, addition of 50 nM KT5720, an inhibitor of PKA, resulted in a 53% percent reduction in the directional response of keratinocytes in the electric field, while not significantly affecting general cell motility. The reduction was dose-dependent, there was a gradual decrease in the directional response from 5 to 50 nM. Secondly, addition of 1 μM ML-7, a myosin light chain kinase inhibitor, resulted in an approximate 31% decrease in the distance the cells migrated without affecting directional migration. The PKC inhibitors GF109203X at 4 μM and H-7 at 20 μM and W-7, a CaM kinase inhibitor, did not significantly alter either directed migration or cell migration, although they all resulted in a slight reduction in directional migration. D-erythro-sphingosine at 15 μM, a PKC inhibitor, had virtually no effect on either migration distance or directed migration. These findings demonstrate that divergent kinase signaling pathways regulate general cell motility and sustained directional migration and highlight the complexity of the signal transduction mechanisms involved. The inhibitor studies described in this paper implicate a role for PKA in the regulation of the directional migratory response to applied electric fields, galvanotaxis.

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