Hasan Bayram 1,2 , Bülent Göğebakan 2 , Öner Dikensoy 1 , Erhan Ekinci 1 .

1. Effects of Diesel Exhaust Particles on Cell Viability and Release of Inflammatory Cytokines from Human Lung Epithelial Cells* Hasan Bayram1,2, Bülent Göğebakan2, Öner Dikensoy1, Erhan Ekinci1. Department of Respiratory Medicine1, Respiratory Cell Culture Laboratory2, School of Medicine, University of Gaziantep, Gaziantep *Supported by the Scientific Research Fund of Gaziantep University.

2. Introduction-1 • Association between particulate matter 10µm (PM10) pollution and respiratory morbidity and cardiopulmonary mortality (McConnell R et al, 2003,Pope CA et al, 2004 ) • Airway epithelial cells may play role in PM10-induced respiratory morbidity • Diesel exhaust particles (DEP) induce release of inflammatory mediators from airway epithelial cells (Bayram H et al, 1998)

3. Introduction-2 • DEP, under serum free condition, increase A549 cell viability by inducing cell cycle and suppressing apoptosis of these cells • DEP exert these effects by inducing oxidative stress, JNK and NF-B pathways, while inhibiting p21CIP1/WAF1 expression (Bayram H et al, 2006)

4. Hypothesis • DEP-induced A549 cell proliferation may be associated with IL-8 and GM-CSF release from these cells

5. Objectives • To investigate effects of DEP on A549 cell viability • To investigate effects of N-acetylcysteine, JNK inhibitor (SP600125) and ERK inhibitor (PD 98059) on this phenomenon • To investigate whether there is an association with IL-8 and GM-CSF release

6. Methods • A549 cell culture • Incubation for 24, 48 and 72 hrs with DEP (0, 5, 10, 50, 100, 200, 400, 1000 and 2000g/ml) • Incubation for 48 hrs with 50g/ml DEP in the absence or presence of N-acetylcysteine (3.3-10mM), JNK inhibitor (SP600125, 3.3-33 M) and ERK inhibitor (PD 98059, 3.3-33M) • MTT Staining: A549 cell viability • ELISA: IL-8 and GM-CSF analysis

7. Results

8. 24h Effects of DEP on A549 Cell Viability ***p<0.0001vs0g/ml DEP 72h 48h

9. Effect of N-Acetylcysteine (NAC) on Viability of A549 Cells Following 48 hrs’ incubation with DEP (50g/ml)

10. Effect of JNK inh. (SP600125) on Viability of A549 Cells Following 48 hrs’ incubation with DEP (50g/ml)

11. Effect of ERK inh. (PD98059) on Viability of A549 Cells Following 48 hrs’ Incubation with DEP (50 g/ml)

12. 24h Effects of DEP on IL-8 Release from A549 Cells *p<0.05 ***p<0.0001 vs 0g/ml DEP 48h 72h

13. Effects of DEP on Release of GM-CSF from A549 Cells Following 72hrs’ Incubation *p<0.05 -0g/ml DEP

14. Summary-1 • DEP at doses of 200-400µg/ml induced A549 cell viability following 24hrs’ incubation, whereas higher doses (1000-2000µg/ml) decreased cell viability. • DEP induced A549 cell viability after 48hrs’ (10-400µg/ml) and 72hrs’ (50-400µg/ml) incubation • 10-33mM NAC, 33µM JNK inhibitor and 3.3-10µM ERK inhibitor inhibited DEP-induced cell viability

15. Summary-2 • Although 50-400µg/ml DEP inhibited IL-8 release following 48hrs’ incubation, 5µg/ml DEP induced release of this cytokine after 72hrs. • 400µg/ml DEP increased release of GM-CSF following 72hrs’ incubation.

16. Conclusion • DEP induce airway epithelial cell viability • DEP exert this effect by inducing oxidative stress and cell signalling pathways (JNK and ERK) known to be sensitive to the oxidative stress • The role of DEP-induced cytokine release in cell proliferation need to be investigated further.

17. Acknowledgements

18. Acknowledgements

19. Thank You