Alveolar Epithelial cell differentiation and response to injury.
Fiaturi, Najla.
2014
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Abstract: Lung immaturity is the major cause of morbidity and mortality in premature infants, especially those born <28 weeks gestation. Proper lung development from 23-28 weeks requires coordinated cell proliferation and differentiation. Infants born at this age are at high risk for respiratory distress syndrome (RDS), a lung disease characterized by insufficient surfactant production due to imma... read moreturity of the saccular/alveolar type II epithelial cells in the lung. Neuregulin (NRG) stimulation of ErbB4 signaling is important for development of type II cell surfactant synthesis. ErbB4 may mediate gene expression via a non-canonical pathway involving enzymatic cleavage releasing its intracellular domain (4ICD) for nuclear trafficking and gene regulation. The accepted model for release of 4ICD is consecutive cleavage by Tumor necrosis factor alpha Converting Enzyme (TACE) and γ-secretase enzymes. Here we hypothesize that cleavage of ErbB4 by the two enzymes TACE and gamma secretase is necessary for surfactant production in type 2 epithelial cells. Our data show that Presenilin-1 (PSEN-1) is the active enzyme in the γ-secretase complex which cleaves ErbB4 to release 4ICD. We further show that release of 4ICD by PSEN-1 cleavage is not dependent on previous TACE cleavage. We used siRNA to silence PSEN-1 expression in a mouse lung type II epithelial cell line (MLE12 cells), and both siRNA knockdown and chemical inhibition of TACE. Knockdown of PSEN-1 significantly decreased baseline and NRG-stimulated surfactant phospholipid synthesis, expression of the surfactant proteins SP-B and SP-C, as well as 4ICD levels, with no change in ErbB4 ectodomain shedding. Neither siRNA knockdown nor chemical inhibition of TACE inhibited 4ICD release or surfactant synthesis. PSEN-1 cleavage of ErbB4 for non-canonical signaling through 4ICD release does not require prior cleavage by TACE. Preterm babies with RDS are treated with oxygen and supportive breathing using ventilators. Higher inspired oxygen levels improve preterm infants' survival but increase the risk for a condition of chronic lung injury, called Bronchopulmonary Dysplasia (BPD). BPD is characterized by arrested alveolarization and airway hyperreactivity. The mechanisms regulating normal alveolar development and BPD are not well understood. Of novel interest is the potential role of the matricellular protein CCN5 (Cysteine-rich protein 61/Connective tissue growth factor/Nephroblastoma-overexpressed protein), which the Castellot laboratory previously reported is present in developing alveolar epithelium. The mechanism of action and biological function of CCN5 in smooth muscle cells (SMCs) is well-studied. CCN5 is a cell cycle regulator that inhibits SMC proliferation and promotes SMC differentiation. The cell-specific expression and function of CCN5 in alveolar development and injury is unknown. Here we hypothesize that CCN5 is a negative regulator of type2 cell proliferation and motility during alveolar development and response to injury. Our data show that CCN5 knockdown in MLE12 cells reduces proliferation and motility in these cells. A mouse model of BPD, in which neonatal pups are exposed to 90% oxygen from post natal (P) 5 through P13 was used to study CCN5 and alveolar epithelial cell fate. Western Blot analysis of whole lung lysates showed that CCN5 expression is significantly reduced in hyperoxic versus room air P13 lungs. Using immunofluorescence labeling CCN5 was not expressed in alveolar epithelial type II cells in RA lungs. In oxygen-exposed lungs an intermediate cell type that expressed markers for both type II and I epithelial cells was observed. These intermediate cells showed proliferation activity and labeled positive for CCN5. Type I cells also showed proliferation activity. Overall, only proliferating alveolar epithelial cells demonstrated CCN5 expression. In conclusion, CCN5 expression correlates positively with proliferation in alveolar epithelial cells in response to neonatal oxygen injury. This contrasts strongly with studies in SMCs in which CCN5 is highly expressed in non-proliferating cells and is down regulated in proliferating SMCs. Neonatal hyperoxic lung injury induces a transdifferentiation of type II epithelial cells to type I epithelial cells, which is accompanied by both cell proliferation and by CCN5 expression. We propose that CCN5 is a functional regulator of alveolar epithelial proliferation during development and during the response of the neonatal lung to oxygen injury.
Thesis (Ph.D.)--Tufts University, 2014.
Submitted to the Dept. of Pharmacology & Experimental Therapeutics.
Advisors: John Castellot, and Heber Nielsen.
Committee: Theoharis Theoharides, Martin Bienborn, MaryAnn Volpe, and Maria Ramirez.
Keyword: Biology.read less - ID:
- 1z40m510r
- Component ID:
- tufts:20329
- To Cite:
- DCA Citation Guide EndNote
