Apoptosis and the Activity of Ceramide, Bax and Bcl-2 in the Lungs of Neonatal Rats Exposed to Limited and Prolonged Hyperoxia

Overview

Journal Respir Res

Specialty Pulmonary Medicine

Date 2006 Jul 28

PMID 16869980

Citations 16

Authors

Ahmad W Husari

Ghassan S Dbaibo

Hala Bitar

Aline Khayat

Shoghag Panjarian

Michel Nasser

Fadi F Bitar

Marwan El-Sabban

Ghazi Zaatari

Salman M Mroueh

Affiliations

Soon will be listed here.

Abstract

Background: The aim of the study is to examine the effect of limited and prolonged hyperoxia on neonatal rat lung. This is done by examining the morphologic changes of apoptosis, the expression of ceramide, an important mediator of apoptosis, the expression of inflammatory mediators represented by IL-1beta and the expression of 2 proto-oncogenes that appear to modulate apoptosis (Bax and Bcl-2).

Methods: Newborn rats were placed in chambers containing room air or oxygen above 90% for 7 days. The rats were sacrificed at 3, 7 or 14 days and their lungs removed. Sections were fixed, subjected to TUNEL, Hoechst, and E-Cadherin Staining. Sections were also incubated with anti-Bcl-2 and anti-Bax antisera. Bcl-2 and Bax were quantitated by immunohistochemistry. Lipids were extracted, and ceramide measured through a modified diacylglycerol kinase assay. RT-PCR was utilized to assess IL-1beta expression.

Results: TUNEL staining showed significant apoptosis in the hyperoxia-exposed lungs at 3 days only. Co-staining of the apoptotic cells with Hoechst, and E-Cadherin indicated that apoptotic cells were mainly epithelial cells. The expression of Bax and ceramide was significantly higher in the hyperoxia-exposed lungs at 3 and 14 days of age, but not at 7 days. Bcl-2 was significantly elevated in the hyperoxia-exposed lungs at 3 and 14 days. IL-1beta expression was significantly increased at 14 days.

Conclusion: Exposure of neonatal rat lung to hyperoxia results in early apoptosis documented by TUNEL assay. The early rise in Bax and ceramide appears to overcome the anti-apoptotic activity of Bcl-2. Further exposure did not result in late apoptotic changes. This suggests that apoptotic response to hyperoxia is time sensitive. Prolonged hyperoxia results in acute lung injury and the shifting balance of ceramide, Bax and Bcl-2 may be related to the evolution of the inflammatory process.

Citing Articles

Postnatal hypoxic preconditioning attenuates lung damage from hyperoxia in newborn mice.

Millan I, Perez S, Rius-Perez S, Asensi M, Vento M, Garcia-Verdugo J Pediatr Res. 2024; .

PMID: 39317699 DOI: 10.1038/s41390-024-03457-0.

Hyperoxia-Induced miR-195 Causes Bronchopulmonary Dysplasia in Neonatal Mice.

Philpot P, Graumuller F, Melchiorre N, Prahaladan V, Takada X, Chandran S Biomedicines. 2024; 12(6).

PMID: 38927415 PMC: 11201213. DOI: 10.3390/biomedicines12061208.

Synergistic Pulmonoprotective Effect of Natural Prolyl Oligopeptidase Inhibitors in In Vitro and In Vivo Models of Acute Respiratory Distress Syndrome.

Zerikiotis S, Efentakis P, Dapola D, Agapaki A, Seiradakis G, Kostomitsopoulos N Int J Mol Sci. 2023; 24(18).

PMID: 37762537 PMC: 10531912. DOI: 10.3390/ijms241814235.

Heat shock protein 70 protects the lungs from hyperoxic injury in a neonatal rat model of bronchopulmonary dysplasia.

Lee C, Su T, Lee M, Hsu C, Yang R, Kao J PLoS One. 2023; 18(5):e0285944.

PMID: 37200358 PMC: 10194897. DOI: 10.1371/journal.pone.0285944.

The Role of Sphingolipid Signaling in Oxidative Lung Injury and Pathogenesis of Bronchopulmonary Dysplasia.

Thomas J, Sudhadevi T, Basa P, Ha A, Natarajan V, Harijith A Int J Mol Sci. 2022; 23(3).

PMID: 35163176 PMC: 8835774. DOI: 10.3390/ijms23031254.

References

Mantell L, Horowitz S, Davis J, Kazzaz J . Hyperoxia-induced cell death in the lung--the correlation of apoptosis, necrosis, and inflammation. Ann N Y Acad Sci. 2000; 887:171-80. DOI: 10.1111/j.1749-6632.1999.tb07931.x. View

Barazzone C, White C . Mechanisms of cell injury and death in hyperoxia: role of cytokines and Bcl-2 family proteins. Am J Respir Cell Mol Biol. 2000; 22(5):517-9. DOI: 10.1165/ajrcmb.22.5.f180. View

Sawada M, Nakashima S, Banno Y, Yamakawa H, Hayashi K, Takenaka K . Ordering of ceramide formation, caspase activation, and Bax/Bcl-2 expression during etoposide-induced apoptosis in C6 glioma cells. Cell Death Differ. 2000; 7(9):761-72. DOI: 10.1038/sj.cdd.4400711. View

McGrath-Morrow S, Stahl J . Apoptosis in neonatal murine lung exposed to hyperoxia. Am J Respir Cell Mol Biol. 2001; 25(2):150-5. DOI: 10.1165/ajrcmb.25.2.4362. View

Manji J, OKelly C, Leung W, Olson D . Timing of hyperoxic exposure during alveolarization influences damage mediated by leukotrienes. Am J Physiol Lung Cell Mol Physiol. 2001; 281(4):L799-806. DOI: 10.1152/ajplung.2001.281.4.L799. View

Homaidan F, El-Sabban M, Chakroun I, El-Sibai M, Dbaibo G . IL-1 stimulates ceramide accumulation without inducing apoptosis in intestinal epithelial cells. Mediators Inflamm. 2002; 11(1):39-45. PMC: 1781644. DOI: 10.1080/09629350210313. View

Von Haefen C, Wieder T, Gillissen B, Starck L, Graupner V, Dorken B . Ceramide induces mitochondrial activation and apoptosis via a Bax-dependent pathway in human carcinoma cells. Oncogene. 2002; 21(25):4009-19. DOI: 10.1038/sj.onc.1205497. View

Hannun Y, Obeid L . The Ceramide-centric universe of lipid-mediated cell regulation: stress encounters of the lipid kind. J Biol Chem. 2002; 277(29):25847-50. DOI: 10.1074/jbc.R200008200. View

Pagano A, Donati Y, Metrailler I, Barazzone Argiroffo C . Mitochondrial cytochrome c release is a key event in hyperoxia-induced lung injury: protection by cyclosporin A. Am J Physiol Lung Cell Mol Physiol. 2003; 286(2):L275-83. DOI: 10.1152/ajplung.00181.2003. View

10.

Buccellato L, Tso M, Akinci O, Chandel N, Budinger G . Reactive oxygen species are required for hyperoxia-induced Bax activation and cell death in alveolar epithelial cells. J Biol Chem. 2003; 279(8):6753-60. DOI: 10.1074/jbc.M310145200. View

11.

Yamamoto K, Tomita N, Yoshimura S, Nakagami H, Taniyama Y, Yamasaki K . Hypoxia-induced renal epithelial cell death through caspase-dependent pathway: role of Bcl-2, Bcl-xL and Bax in tubular injury. Int J Mol Med. 2004; 14(4):633-40. View

12.

Rouser G, Fkeischer S, Yamamoto A . Two dimensional then layer chromatographic separation of polar lipids and determination of phospholipids by phosphorus analysis of spots. Lipids. 1970; 5(5):494-6. DOI: 10.1007/BF02531316. View

13.

Merrill Jr A, Wang E, Stevens J, Brumley G . Activities of the initial enzymes of glycerolipid and sphingolipid synthesis in lung microsomes from rats exposed to air or 85% oxygen. Biochem Biophys Res Commun. 1984; 119(3):995-1000. DOI: 10.1016/0006-291x(84)90872-6. View

14.

Randell S, Mercer R, Young S . Postnatal growth of pulmonary acini and alveoli in normal and oxygen-exposed rats studied by serial section reconstructions. Am J Anat. 1989; 186(1):55-68. DOI: 10.1002/aja.1001860105. View

15.

Randell S, Mercer R, Young S . Neonatal hyperoxia alters the pulmonary alveolar and capillary structure of 40-day-old rats. Am J Pathol. 1990; 136(6):1259-66. PMC: 1877590. View

16.

Gavrieli Y, Sherman Y, Ben-Sasson S . Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol. 1992; 119(3):493-501. PMC: 2289665. DOI: 10.1083/jcb.119.3.493. View

17.

Obeid L, Linardic C, Karolak L, Hannun Y . Programmed cell death induced by ceramide. Science. 1993; 259(5102):1769-71. DOI: 10.1126/science.8456305. View

18.

Dbaibo G, Obeid L, Hannun Y . Tumor necrosis factor-alpha (TNF-alpha) signal transduction through ceramide. Dissociation of growth inhibitory effects of TNF-alpha from activation of nuclear factor-kappa B. J Biol Chem. 1993; 268(24):17762-6. View

19.

Reed J . Bcl-2 and the regulation of programmed cell death. J Cell Biol. 1994; 124(1-2):1-6. PMC: 2119888. DOI: 10.1083/jcb.124.1.1. View

20.

Fernandes R, Cotter T . Apoptosis or necrosis: intracellular levels of glutathione influence mode of cell death. Biochem Pharmacol. 1994; 48(4):675-81. DOI: 10.1016/0006-2952(94)90044-2. View