The Role of Pulmonary Surfactant Phospholipids in Fibrotic Lung Diseases.

Diffuse parenchymal lung diseases (DPLD) or Interstitial lung diseases (ILD) are a heterogeneous group of lung conditions with common characteristics that can progress to fibrosis. Within this group of pneumonias, idiopathic pulmonary fibrosis (IPF) is considered the most common. This disease has no known cause, is devastating and has no cure. Chronic lesion of alveolar type II (ATII) cells represents a key mechanism for the development of IPF. ATII cells are specialized in the biosynthesis and secretion of pulmonary surfactant (PS), a lipid-protein complex that reduces surface tension and minimizes breathing effort. Some differences in PS composition have been reported between patients with idiopathic pulmonary disease and healthy individuals, especially regarding some specific proteins in the PS; however, few reports have been conducted on the lipid components. This review focuses on the mechanisms by which phospholipids (PLs) could be involved in the development of the fibroproliferative response.

Phosphatidylethanolamine Induces an Antifibrotic Phenotype in Normal Human Lung Fibroblasts and Ameliorates Bleomycin-Induced Lung Fibrosis in Mice.

Lung surfactant is a complex mixture of phospholipids and specific proteins but its role in the pathogenesis of interstitial lung diseases is not established. Herein, we analyzed the effects of three representative phospholipid components, that is, dipalmitoilphosphatidylcoline (DPPC), phosphatidylglycerol (PG) and phosphatidylethanolamine (PE), on collagen expression, apoptosis and Ca2+ signaling in normal human lung fibroblasts (NHLF) and probed their effect in an experimental model of lung fibrosis. Collagen expression was measured with RT-PCR, apoptosis was measured by using either the APOPercentage assay kit (Biocolor Ltd., Northern Ireland, UK) or the Caspase-Glo 3/7 assay (Promega, Madison, WI, USA) and Ca2+ signaling by conventional epifluorescence imaging. The effect in vivo was tested in bleomycin-induced lung fibrosis in mice. DPPC and PG did not affect collagen expression, which was downregulated by PE. Furthermore, PE promoted apoptosis and induced a dose-dependent Ca2+ signal. PE-induced Ca2+ signal and apoptosis were both blocked by phospholipase C, endoplasmic reticulum pump and store-operated Ca2+ entry inhibition. PE-induced decrease in collagen expression was attenuated by blocking phospholipase C. Finally, surfactant enriched with PE and PE itself attenuated bleomycin-induced lung fibrosis and decreased the soluble collagen concentration in mice lungs. This study demonstrates that PE strongly contributes to the surfactant-induced inhibition of collagen expression in NHLF through a Ca2+ signal and that early administration of Beractant enriched with PE diminishes lung fibrosis in vivo.

Organic synthesis of 1,2-dipalmitoyl-rac-glycero-3-phosphatidylethanolamine and its effect on the induction of apoptosis in normal human lung fibroblasts.

BACKGROUND /OBJECTIVE: The phospholipid 1,2-dipalmitoyl-rac-glycero-3-phosphatidylethanolamine (PE) comprises two fatty acid chains: glycerol, phosphate, and ethanolamine. PE participates in critical cellular processes such as apoptosis and autophagy, which places it as a target for designing new therapeutic alternatives in diseases such as pulmonary fibrosis. Therefore, this study aimed obtain PE through a six-step organic synthesis pathway and determine its biological effect on apoptosis induction in normal human lung fibroblasts (NHLF). METHODOLOGY: The first step of the organic synthesis route began with protected glycerol that was benzylated at sn-3; later, it was deprotected to react with palmitic acid at sn-1, sn-2. To remove the benzyl group, hydrogenation was performed with palladium on carbon (Pd/C); subsequently, the molecule was phosphorylated in sn-3 with phosphorus oxychloride and triethylamine, and the intermediate was hydrolyzed in an acid medium to obtain the final compound. After PE synthesis, apoptosis assessment was performed: apoptosis was induced using exposure to annexin V-FITC/propidium iodide-ECD (PI) and quantified using flow cytometry. The experiments were performed in three NHLF cell lines with different concentrations of PE 10, 100 and 1000 µg/mL for 24 and 48 h. RESULTS: The PE obtained by organic synthesis presented a melting point of 190-192 °C, a purity of 95%, and a global yield of 8%. The evaluation of apoptosis with flow cytometry showed that at 24 h, exposure to PE 10, 100, and 1000 µg/mL induces early apoptosis in 19.42%- 25.54%, while late apoptosis was only significant P < 0.05 in cells challenged with 100 µg/mL PE. At 48 h, NHLF exposed to PE 10, 100, and 1000 µg/mL showed decreasing early apoptosis: 28.69-32.16%, 12.59-18.84%, and 10.91-12.61%, respectively. The rest of the NHLF exposed to PE showed late apoptosis: 12.03-16-42%, 11.04-15.94%, and 49.23-51.28%. Statistical analysis showed a significance P < 0.05 compared to the control. CONCLUSION: The organic synthesis route of PE allows obtaining rac-1,2-O-Dipalmitoyl-glycero-3-phosphoethanolamine (1), which showed an apoptotic effect on NHLF.