Benzo[a]pyrene inhibits protein kinase C activity in subcultured rat aortic smooth muscle cells

Xiaolan Ou, Kenneth Ramos

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

Recent studies in this laboratory have shown that benzo[a]pyrene (BaP) interferes with protein kinase C (PKC)-mediated phosphorylation of aortic smooth muscle cell (SMC) proteins in vivo. To evaluate the biochemical basis of this response, the present studies have been conducted to examine the time- and concentration-dependent effects of BaP on PKC activity in vitro. Growth-arrested subcultures of rat aortic SMCs were exposed to 0.3, 3, or 30 μM BaP in the presence of fetal bovine serum for various times and then processed for measurements of exogenous histone Type III-S phosphorylation under PKC-activating conditions. Challenge of SMCs with BaP for 8 h was associated with a concentration-dependent inhibition of PKC activity in both cytosolic and particulate fractions. While no changes of enzymatic activity were observed in either fraction following exposure of SMCs to 0.3 μM BaP, higher concentrations of BaP inhibited PKC in both cytosolic and particulate fractions. A 49% and 68% reduction of cytosolic PKC activity was observed in SMCs treated with 3 and 30 μM BaP, respectively. The inhibitory response elicited by BaP was more pronounced in the particulate fraction where 61% and 89% decreases in PKC activity were observed in cultures treated with 3 and 30 μM BaP, respectively. Time course studies revealed that inhibition of PKC activity by 30 μM BaP occurred as early as 30 min and was sustained for up to 24 h in both fractions. Benzo[a]pyrene (30 μM) did not interfere with the ability of phorbol-12-myristate-13-acetate to induce PKC translocation from the cytosolic to particulate compartment since maximal translocation occurred by 5 min and lasted for up to 60 min in both control and BaP-treated cultures. The inhibitory effects of BaP were independent of new protein or RNA synthesis, but appear to involve oxidative metabolism of the parent compound since 3-hydroxy-BaP, the major P450-derived BaP metabolite in SMCs, also inhibited cytosolic and particulate PKC activity. Collectively, these data demonstrate that BaP and its 3-hydroxy metabolite inhibit PKC activity in rat aortic SMCs and raise the possibility that interference with PKC-mediated protein phosphorylation participates in the deregulation of SMC growth and differentiation induced by BaP.

Original languageEnglish (US)
Pages (from-to)29-40
Number of pages12
JournalChemico-Biological Interactions
Volume93
Issue number1
DOIs
StatePublished - 1994
Externally publishedYes

Fingerprint

Benzo(a)pyrene
Protein Kinase C
Smooth Muscle Myocytes
Muscle
Rats
Cells
Phosphorylation
Metabolites
Time and motion study
Proteins
Deregulation
Muscle Proteins
Cell growth
Growth
Metabolism
Histones
Cell Differentiation
Acetates
RNA

Keywords

  • Aortic smooth muscle cells
  • Benzo[a]pyrene
  • Protein kinase C

ASJC Scopus subject areas

  • Toxicology

Cite this

Benzo[a]pyrene inhibits protein kinase C activity in subcultured rat aortic smooth muscle cells. / Ou, Xiaolan; Ramos, Kenneth.

In: Chemico-Biological Interactions, Vol. 93, No. 1, 1994, p. 29-40.

Research output: Contribution to journalArticle

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abstract = "Recent studies in this laboratory have shown that benzo[a]pyrene (BaP) interferes with protein kinase C (PKC)-mediated phosphorylation of aortic smooth muscle cell (SMC) proteins in vivo. To evaluate the biochemical basis of this response, the present studies have been conducted to examine the time- and concentration-dependent effects of BaP on PKC activity in vitro. Growth-arrested subcultures of rat aortic SMCs were exposed to 0.3, 3, or 30 μM BaP in the presence of fetal bovine serum for various times and then processed for measurements of exogenous histone Type III-S phosphorylation under PKC-activating conditions. Challenge of SMCs with BaP for 8 h was associated with a concentration-dependent inhibition of PKC activity in both cytosolic and particulate fractions. While no changes of enzymatic activity were observed in either fraction following exposure of SMCs to 0.3 μM BaP, higher concentrations of BaP inhibited PKC in both cytosolic and particulate fractions. A 49{\%} and 68{\%} reduction of cytosolic PKC activity was observed in SMCs treated with 3 and 30 μM BaP, respectively. The inhibitory response elicited by BaP was more pronounced in the particulate fraction where 61{\%} and 89{\%} decreases in PKC activity were observed in cultures treated with 3 and 30 μM BaP, respectively. Time course studies revealed that inhibition of PKC activity by 30 μM BaP occurred as early as 30 min and was sustained for up to 24 h in both fractions. Benzo[a]pyrene (30 μM) did not interfere with the ability of phorbol-12-myristate-13-acetate to induce PKC translocation from the cytosolic to particulate compartment since maximal translocation occurred by 5 min and lasted for up to 60 min in both control and BaP-treated cultures. The inhibitory effects of BaP were independent of new protein or RNA synthesis, but appear to involve oxidative metabolism of the parent compound since 3-hydroxy-BaP, the major P450-derived BaP metabolite in SMCs, also inhibited cytosolic and particulate PKC activity. Collectively, these data demonstrate that BaP and its 3-hydroxy metabolite inhibit PKC activity in rat aortic SMCs and raise the possibility that interference with PKC-mediated protein phosphorylation participates in the deregulation of SMC growth and differentiation induced by BaP.",
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