Protein kinase c-α mediates cigarette smoke extract- and complement factor 5a-stimulated interleukin-8 release in human bronchial epithelial cells

Ravindra Kashyap, Anthony A. Floreani, Arthur J. Heires, Sam D. Sanderson, Todd A Wyatt

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

Background: Cigarette smoke extract (CSE) activates protein kinase C (PKC) and augments complement factor 5a (C5a)-stimulated release of the proinflammatory cytokine IL-8 in human bronchial epithelial cells (HBEC). We hypothesized that PKC activation by alternative PKC activators will also mediate C5a-stimulated IL-8 release in HBEC. Methods: HBEC were treated with phorbol myristate acetate (100 ng/mL), calcium ionophore A23187 (2 nM), or 10 nM cholesterol-3-sulfate in the presence or absence of C5a. Interleukin-8 (IL-8) release was measured by enzyme-linked immunoadsorbent assay. Results: IL-8 release by PKC activators alone was significantly higher than in unstimulated cells and was further augmented in the presence of C5a. Preincubation with the PKC inhibitor calphostin C (1 μM) significantly suppressed IL-8 release in HBEC treated with CSE and C5a. Preincubation with 10 μM TMB-8 (an intracellular calcium sequester) also significantly suppressed IL-8 release in CSE- and C5a-treated HBEC, suggesting that intracellular calcium is required for CSE- and C5a-mediated IL-8 release. When HBEC were preincubated with 30 nM of the PKCβ-specific inhibitor LY363196, CSE- and C5a-mediated IL-8 release was not inhibited. However, with higher concentrations of LY363196 (>600 nM), which exceeds the IC50 for PKCβ greater than 100 fold, CSE- and C5a-mediated IL-8 release was significantly suppressed. Preincubation of HBEC with 100 nM of Gö 6976, a specific PKCα inhibitor, significantly inhibited CSE- and C5a-mediated stimulation of IL-8 release. Conclusions: Collectively, these data suggest that PKC activators in addition to CSE augment C5a-stimulated IL-8 release from HBEC and that CSE and C5a stimulate IL-8 release in HBEC by activating the calcium-dependent PKCα isoform.

Original languageEnglish (US)
Pages (from-to)46-53
Number of pages8
JournalJournal of Investigative Medicine
Volume50
Issue number1
StatePublished - Jan 1 2002

Fingerprint

Complement C5a
Interleukin-8
Smoke
Tobacco Products
Protein Kinases
Protein Kinase C
Epithelial Cells
Protein C Inhibitor
Protein Kinase Inhibitors
Calcium
Immunosorbents
Calcium Ionophores
Calcimycin
Tetradecanoylphorbol Acetate
Inhibitory Concentration 50
Sulfates

Keywords

  • Airway epithelium
  • C5a
  • Cigarette smoke
  • IL-8
  • PKC

ASJC Scopus subject areas

  • Medicine(all)
  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Protein kinase c-α mediates cigarette smoke extract- and complement factor 5a-stimulated interleukin-8 release in human bronchial epithelial cells. / Kashyap, Ravindra; Floreani, Anthony A.; Heires, Arthur J.; Sanderson, Sam D.; Wyatt, Todd A.

In: Journal of Investigative Medicine, Vol. 50, No. 1, 01.01.2002, p. 46-53.

Research output: Contribution to journalArticle

@article{0819851c9aab487281595ea00b006ffa,
title = "Protein kinase c-α mediates cigarette smoke extract- and complement factor 5a-stimulated interleukin-8 release in human bronchial epithelial cells",
abstract = "Background: Cigarette smoke extract (CSE) activates protein kinase C (PKC) and augments complement factor 5a (C5a)-stimulated release of the proinflammatory cytokine IL-8 in human bronchial epithelial cells (HBEC). We hypothesized that PKC activation by alternative PKC activators will also mediate C5a-stimulated IL-8 release in HBEC. Methods: HBEC were treated with phorbol myristate acetate (100 ng/mL), calcium ionophore A23187 (2 nM), or 10 nM cholesterol-3-sulfate in the presence or absence of C5a. Interleukin-8 (IL-8) release was measured by enzyme-linked immunoadsorbent assay. Results: IL-8 release by PKC activators alone was significantly higher than in unstimulated cells and was further augmented in the presence of C5a. Preincubation with the PKC inhibitor calphostin C (1 μM) significantly suppressed IL-8 release in HBEC treated with CSE and C5a. Preincubation with 10 μM TMB-8 (an intracellular calcium sequester) also significantly suppressed IL-8 release in CSE- and C5a-treated HBEC, suggesting that intracellular calcium is required for CSE- and C5a-mediated IL-8 release. When HBEC were preincubated with 30 nM of the PKCβ-specific inhibitor LY363196, CSE- and C5a-mediated IL-8 release was not inhibited. However, with higher concentrations of LY363196 (>600 nM), which exceeds the IC50 for PKCβ greater than 100 fold, CSE- and C5a-mediated IL-8 release was significantly suppressed. Preincubation of HBEC with 100 nM of G{\"o} 6976, a specific PKCα inhibitor, significantly inhibited CSE- and C5a-mediated stimulation of IL-8 release. Conclusions: Collectively, these data suggest that PKC activators in addition to CSE augment C5a-stimulated IL-8 release from HBEC and that CSE and C5a stimulate IL-8 release in HBEC by activating the calcium-dependent PKCα isoform.",
keywords = "Airway epithelium, C5a, Cigarette smoke, IL-8, PKC",
author = "Ravindra Kashyap and Floreani, {Anthony A.} and Heires, {Arthur J.} and Sanderson, {Sam D.} and Wyatt, {Todd A}",
year = "2002",
month = "1",
day = "1",
language = "English (US)",
volume = "50",
pages = "46--53",
journal = "Journal of Investigative Medicine",
issn = "1081-5589",
publisher = "Lippincott Williams and Wilkins",
number = "1",

}

TY - JOUR

T1 - Protein kinase c-α mediates cigarette smoke extract- and complement factor 5a-stimulated interleukin-8 release in human bronchial epithelial cells

AU - Kashyap, Ravindra

AU - Floreani, Anthony A.

AU - Heires, Arthur J.

AU - Sanderson, Sam D.

AU - Wyatt, Todd A

PY - 2002/1/1

Y1 - 2002/1/1

N2 - Background: Cigarette smoke extract (CSE) activates protein kinase C (PKC) and augments complement factor 5a (C5a)-stimulated release of the proinflammatory cytokine IL-8 in human bronchial epithelial cells (HBEC). We hypothesized that PKC activation by alternative PKC activators will also mediate C5a-stimulated IL-8 release in HBEC. Methods: HBEC were treated with phorbol myristate acetate (100 ng/mL), calcium ionophore A23187 (2 nM), or 10 nM cholesterol-3-sulfate in the presence or absence of C5a. Interleukin-8 (IL-8) release was measured by enzyme-linked immunoadsorbent assay. Results: IL-8 release by PKC activators alone was significantly higher than in unstimulated cells and was further augmented in the presence of C5a. Preincubation with the PKC inhibitor calphostin C (1 μM) significantly suppressed IL-8 release in HBEC treated with CSE and C5a. Preincubation with 10 μM TMB-8 (an intracellular calcium sequester) also significantly suppressed IL-8 release in CSE- and C5a-treated HBEC, suggesting that intracellular calcium is required for CSE- and C5a-mediated IL-8 release. When HBEC were preincubated with 30 nM of the PKCβ-specific inhibitor LY363196, CSE- and C5a-mediated IL-8 release was not inhibited. However, with higher concentrations of LY363196 (>600 nM), which exceeds the IC50 for PKCβ greater than 100 fold, CSE- and C5a-mediated IL-8 release was significantly suppressed. Preincubation of HBEC with 100 nM of Gö 6976, a specific PKCα inhibitor, significantly inhibited CSE- and C5a-mediated stimulation of IL-8 release. Conclusions: Collectively, these data suggest that PKC activators in addition to CSE augment C5a-stimulated IL-8 release from HBEC and that CSE and C5a stimulate IL-8 release in HBEC by activating the calcium-dependent PKCα isoform.

AB - Background: Cigarette smoke extract (CSE) activates protein kinase C (PKC) and augments complement factor 5a (C5a)-stimulated release of the proinflammatory cytokine IL-8 in human bronchial epithelial cells (HBEC). We hypothesized that PKC activation by alternative PKC activators will also mediate C5a-stimulated IL-8 release in HBEC. Methods: HBEC were treated with phorbol myristate acetate (100 ng/mL), calcium ionophore A23187 (2 nM), or 10 nM cholesterol-3-sulfate in the presence or absence of C5a. Interleukin-8 (IL-8) release was measured by enzyme-linked immunoadsorbent assay. Results: IL-8 release by PKC activators alone was significantly higher than in unstimulated cells and was further augmented in the presence of C5a. Preincubation with the PKC inhibitor calphostin C (1 μM) significantly suppressed IL-8 release in HBEC treated with CSE and C5a. Preincubation with 10 μM TMB-8 (an intracellular calcium sequester) also significantly suppressed IL-8 release in CSE- and C5a-treated HBEC, suggesting that intracellular calcium is required for CSE- and C5a-mediated IL-8 release. When HBEC were preincubated with 30 nM of the PKCβ-specific inhibitor LY363196, CSE- and C5a-mediated IL-8 release was not inhibited. However, with higher concentrations of LY363196 (>600 nM), which exceeds the IC50 for PKCβ greater than 100 fold, CSE- and C5a-mediated IL-8 release was significantly suppressed. Preincubation of HBEC with 100 nM of Gö 6976, a specific PKCα inhibitor, significantly inhibited CSE- and C5a-mediated stimulation of IL-8 release. Conclusions: Collectively, these data suggest that PKC activators in addition to CSE augment C5a-stimulated IL-8 release from HBEC and that CSE and C5a stimulate IL-8 release in HBEC by activating the calcium-dependent PKCα isoform.

KW - Airway epithelium

KW - C5a

KW - Cigarette smoke

KW - IL-8

KW - PKC

UR - http://www.scopus.com/inward/record.url?scp=0036155794&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0036155794&partnerID=8YFLogxK

M3 - Article

VL - 50

SP - 46

EP - 53

JO - Journal of Investigative Medicine

JF - Journal of Investigative Medicine

SN - 1081-5589

IS - 1

ER -