|Eduardo R Lazarowski, PhD, Research Associate Professor|
Dr. Lazarowski studies the molecular basis of nucleotide release. Extracellular nucleotides and nucleosides regulate the mucociliary clearance (MCC) process that removes noxious materials from the airways. Nucleotide actions in the airways are mediated by several G protein-coupled purinergic receptors, mainly the ATP/UTP-sensing P2Y2 receptor and the A2B adenosine receptor (Fig. 1). By developing enzymatic- and HPLC-based assays that accurately quantify adenine and uridine nucleotides, the Lazarowski lab has played an important role in defining the contribution of purinergic receptors to MCC responses initiated by the release of ATP and UTP into the airway surface liquid.
Airway epithelial cells release nucleotides via vesicular and conductive mechanisms. Our recent studies indicate that:
- Release of nucleotides, in part, reflects the contribution of nucleotides entering the constitutive secretory pathway via Golgi-resident SLC35 nucleotide transporters (Sesma et al. J. Biol. Chem. 2009) (Fig. 2).
- Ciliated cell-rich airway epithelia exhibit robust release of ATP in response to environmental challenges, including mechanical, osmotic, and biochemical stimuli. Our recent work, using fluorescence microscopy imaging, dominant negative mutants, small interference RNA techniques, and gene targeting approaches, indicates that pannexin 1 (Panx1) represents a major conductive pathway for ATP release from non-mucous cells (Figs. 1 and 2). We are currently investigating the consequences of Panx1 gene deletion to lung physiology and pathophysiology, using the Panx1-/- mouse recently generated by our group (Seminario-Vidal et al, J. Biol. Chem. 2011). An additional focus of these studies is on the involvement of the transient receptor potential vanilloid 4 (TRPV4), which transduces osmotic stress into RhoA activation, upstream of Panx1-mediated ATP release (Seminario-Vidal et al, 2011).
- Airway mucin secreting goblet cells release nucleotides from mucin granules (Kreda et al. J. Physiol. 2010). We are currently investigating the molecular mechanisms by which cytosolic nucleotides are taken and stored into mucin granules (Fig. 2).
- UDP-glucose and other UDP-sugars acting on the P2Y14 receptor are released from airway epithelial cells and accumulate in (patho)physiologically relevant concentrations in lung secretions from patients with chronic lung inflammation (e.g., cystic fibrosis). We are investigating the hypothesis that UDP-sugars released into the airway lumen play pro-inflammatory roles via activation of the P2Y14 receptor expressed in inflammatory cells (Sesma et al. FASEB J. 25:751.16 -abstract, 2011).
|Figure 1. Model of purinergic control of airway epithelial MCC functions. A2B adenosine receptors (A2B-R) on ciliated cells impart cyclic AMP-regulated CFTR Cl- channel activity, while ATP/UTP-sensing P2Y2 receptors (P2Y2-R) promote Cl- secretion (via both CaCC/TMEM16 and PKC-mediated CFTR activation) and inhibit Na+ absorption. P2Y2-Rs also are expressed on goblet cells, promoting mucin secretion. ATP, ADP, and AMP, are stored within mucin granules (MGs). Goblet cell MGs and pannexin 1 (Px1) channels on ciliated cells are major contributors to ASL nucleotides. Ecto-nucleotidases sequentially convert ATP to ADP, AMP, and ADO. See full-size image...||Figure 2. Pathways for airway epithelial nucleotide release. Several scenarios possibly account for the constitutive and stimulated release of nucleotides from airway epithelial cells. (i) Nucleotides entering the Golgi lumen via specific transporters are released as a residual cargo product of the constitutive secretory pathway. (ii) Mucin granules containing ATP are competent for Ca2+-regulated exocytosis. (iii) Cytosolic ATP (and other NTPs) is released via the plasma membrane pannenxin1 channel. See full-size image...|
Directed by Dr. Eduardo Lazarowski, the Nucleotide Measurement Laboratory was established as Core lab in 2000. The function of the Core is to assist investigators within the UNC CF Center and other UNC faculties to analyze and quantify the nucleotide and nucleoside content of cell/tissue samples derived from studies in their labs. This Core has adapted and modified published techniques for quantifying purine nucleotides and nucleosides with nanomolar sensitivity. Using the etheno-derivatization technique, adenine-containing species are modified to fluorescent etheno-derivatives, e.g., etheno-adenosine (e-ADO), e-ADP, e-AMP, and e-ATP, which are separated by HPLC and analyzed/quantified by high sensitive fluorescence detectors. The core also provides unique methodology for quantifying uridine nucleotides (UTP and UDP) and nucleotide sugars (UDP-glucose, UDP-N-acetylgucosamine, and UDP-galactose) in biological samples, by combining highly selective radio-enzymatic assays with HPLC analysis of radioactive species.
Eduardo R Lazarowski, PhD, Research Associate Professor
Education and Training
(1975) BS, Clinical Chemistry, University of Buenos Aires, Argentina.
(1978) MS, Biochemistry, University of Buenos Aires, Argentina.
(1989) PhD, Biochemistry, University of Buenos Aires, Argentina.
(1986-990) Visiting Fellow, The Wellcome Research Laboratories, NC, USA.
(1990-1994) Post-doctoral fellow. Department of Medicine. UNC Chapel Hill.
(1994-1998) Research Associate. Cystic Fibrosis Center and Department of Pharmacology, UNC Chapel Hill.
(1998-2004) Research Assistant Professor. Department of Medicine, UNC Chapel Hill.
(2004-present) Research Associate Professor. Department of Medicine, UNC Chapel Hill.
Professional Societies, Editorial Appointments, and Committees
(2004- present) Editorial Board of Purinergic Signaling. Kluwer Academic Publishers.
(2004- present) American Society of Biochemistry and Molecular Biology.
(2004) Organizing Committee of the 4th International Symposium of Nucleosides and Nucleotides. Purines 2004. Chapel Hill, NC.
(2011) Associate Faculty Member of Faculty of 1000.
Selected Recent Publications
Please see Pubmed feed in the righthand column for more links to current publications.
Seminario-Vidal L, Okada S. F., Sesma J.I., Kreda S.M., van Heusden C.A., Zhu Z., Jones L., O'Neal W. K., Penuela S., Laird D. W., Boucher R. C., and Lazarowski E. R. Rho signaling regulates pannexin 1-mediated ATP release from airway epithelia. J Biol Chem. 286:26277–26286 (2011).
Okada S. F., Zhang L., Kreda S. M., Absullah L. H., Davis C. W., Pickels R. J., Lazarowski E. R., Boucher R. C. Coupled nucleotide and mucin hypersecretion from goblet cell metaplasic human airway epithelium. J Respir Cell Mol Biol. (in press, 2011).
Seminario-Vidal L., van Heusden C., Mugesh G., Lazarowski E. R., Ebselen is a potent non-competitive inhibitor of extracellular nucleoside diphosphokinase. Purinergic Signaling 6:383-391 (2011).
Chekeni F. B., Elliot M. R, Sandilos J. K., Walk S. F., Kinchen J. M., Lazarowski E. R., Armstrong A. J., Penuela S., Laird D. W., Salvesen S., Isakson B. E., Bayliss D. A., Ravichandran K. S. Pannexin 1 channels mediate ‘find-me’ signal release and membrane permeability during apoptosis. Nature 467:863-867 (2010).
Kreda S. M., Seminario-Vidal L, van Heusden C. A., O’Neal W., Jones L., Boucher R. C., and Lazarowski E. R. Receptor-promoted exocytosis of airway epithelial mucin granules containing a spectrum of adenine nucleotides. J. Physiol. 588:2255-2267 (2010).
Lazarowski, E. R. Quantification of extracellular UDP-galactose. Anal. Biochem. 396 23–29 (2010).
Sesma, J. I., Esther Jr, C. R., Kreda, S. M., Jones L., O’Neal, W., Nishihara, S., Nicholas, R. A., Lazarowski, E. R. ER/Golgi nucelotide sugar transporters contribute to the cellular release of UDP-sugar signaling molecules. J. Biol. Chem. 284:12572-12583 (2009).
Lazarowski E. R., Sesma J., Seminario-Vidal L., Kreda S. M. Release and interconversion of purine and pyrimidine nucleotides. Advances in Pharmacology 61:221-61 (2011).
Harden T. K., Sesma J. I., Fricks I.P., and Lazarowski E. R. Signaling and Pharmacological Properties of the P2Y14 Receptor. Acta Physiol. 199:149-60 (2010).
Elliott, M. R., Chekeni, F. B., Trampont, P. C., Lazarowski, E. R., Kadl, A., Walk, S. F., Park, D., Woodson, R. I., Ostankovich, M., Sharma, P., Lysiak, J. J., Harden, T. K., Leitinger, N., Ravichandran, K. S. Nucleotides released by apoptotic cells act as a find-me signal to promote phagocytic clearance. Nature 461:282-6 (2009).
Seminario-Vidal, L., Kreda, S., Jones, L., O’Neal, W., Trejo, J., Boucher, R. C., and Lazarowski, E. R. Thrombin promotes release of ATP from lung epithelial cells through coordinated activation of Rho- and Ca2+-dependent signaling pathways. J. Biol. Chem. 284:20638-20648 (2009).
Lazarowski, E. R., and Boucher, R. C. Purinergic receptors in airway epithelia. Current Opinion in Pharmacology 9:262-267 (2009).
National Institutes of Health (NIH)
|Eduardo Lazarowski, PhD, Research Associate Profesor||Juliana Sesma, PhD, Postdoctoral Research Associate||Catja VanHeusen, Research Specialist and Laboratory Manager|
The University of North Carolina at Chapel Hill
Campus Box #7248
Chapel Hill, NC 27599-7248
Phone: (919) 966-0991
Phone: (919) 309-4403