Tuberculosis and Respiratory Diseases

The Korean Academy of Tuberculosis and Respiratory Diseases (대한결핵및호흡기학회)
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Inhibition of PKC Epsilon Attenuates Cigarette Smoke Extract-Induced Apoptosis in Human Lung Fibroblasts (MRC-5 Cells)

  • Kang, Shin-Myung (Department of Pulmonary and Critical Care Medicine, Gachon University Gil Hospital, Gachon University of Medicine and Science) ;
  • Yoon, Jin-Young (Department of Pulmonary and Critical Care Medicine, Gachon University Gil Hospital, Gachon University of Medicine and Science) ;
  • Kim, Yu-Jin (Department of Pulmonary and Critical Care Medicine, Gachon University Gil Hospital, Gachon University of Medicine and Science) ;
  • Lee, Sang-Pyo (Department of Pulmonary and Critical Care Medicine, Gachon University Gil Hospital, Gachon University of Medicine and Science) ;
  • Jeong, Sung-Hwan (Department of Pulmonary and Critical Care Medicine, Gachon University Gil Hospital, Gachon University of Medicine and Science) ;
  • Park, Jeong-Woong (Department of Pulmonary and Critical Care Medicine, Gachon University Gil Hospital, Gachon University of Medicine and Science)
  • Received : 2011.07.09
  • Accepted : 2011.07.25
  • Published : 2011.08.30
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Abstract

Background: It is known that cigarette smoke (CS) causes cell death. Apoptotic cell death is involved in the pathogenesis of CS-related lung diseases. Some members of the protein kinase C (PKC) family have roles in cigarette smoke extract (CSE)-induced apoptosis. This study was conducted to investigate the role of PKC epsilon in CSE-induced apoptosis in human lung fibroblast cell line, MRC-5. Methods: Lactate dehydrogenase release was measured using a cytotoxicity detection kit. The MTT assay was used to measure cell viability. Western immunoblot, Hoechst 33342 staining and flow cytometry were used to demonstrate the effect of $PKC{\varepsilon}$. Caspase-3 and caspase-8 activities were determined using a colorimetric assay. To examine $PKC{\varepsilon}$ activation, Western blotting was performed using both fractions of membrane and cytosol. Results: We showed that CSE activated $PKC{\varepsilon}$ by demonstrating increased expression of $PKC{\varepsilon}$ in the plasma membrane fraction. Pre-treatment of $PKC{\varepsilon}$ peptide inhibitor attenuated CSE-induced apoptotic cell death, as demonstrated by the MTT assay (13.03% of control, 85.66% of CSE-treatment, and 53.73% of $PKC{\varepsilon}$ peptide inhibitor-pre-treatment, respectively), Hoechst 33342 staining, and flow cytometry (85.64% of CSE-treatment, 53.73% of $PKC{\varepsilon}$ peptide inhibitor-pre-treatment). Pre-treatment of $PKC{\varepsilon}$ peptide inhibitor reduced caspase-3 expression and attenuated caspase-3, caspase-8 activity compared with CSE treatment alone. Conclusion: $PKC{\varepsilon}$ seem to have pro-apoptotic function and exerts its function through the extrinsic apoptotic pathway in CSE-exposed MRC-5 cells. This study suggests that $PKC{\varepsilon}$ inhibition may be a therapeutic strategy in CS-related lung disease such as chronic obstructive pulmonary disease.

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References

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