Premium
LRP‐1 Deficiency in Type II Pneumocytes Decreased Surfactant but Increased Intracellular Lipids
Author(s) -
GarciaArcos Itsaso,
Goldberg Ira J.,
Foronjy Robert F.
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.1132.13
Subject(s) - chemistry , pulmonary surfactant , small hairpin rna , phosphatidylcholine , phospholipid , lipid droplet , biochemistry , microbiology and biotechnology , biology , gene knockdown , apoptosis , membrane
Pulmonary surfactant is synthesized by type II pneumocytes and is mainly composed of phospholipids and minor amounts of cholesterol and specific proteins. Surfactant allows the lungs to inflate by decreasing alveolar surface tension and the work needed for inspiration in each breathing cycle. A delicate balance between synthesis, secretion, recycling and degradation of the lipids is required to maintain surfactant function and lung performance. Single nucleotide polymorphisms in the LDL receptor associated protein 1 (LRP‐1) are associated with decreased lung function in patients with chronic obstructive pulmonary disease (COPD). But the function of LRP1 in the lung is not known. Amongst other roles, LRP‐1 participates in liver lipoprotein metabolism, in adipocyte cell differentiation and in macrophage inflammatory cascades. We deleted LRP‐1 in type II pneumocyte‐derived A549 cells using stable transfection. LRP1 shRNA‐treated cells and scrambled shRNA‐treated cells were cultured in collagen coated transwell inserts at the air‐liquid interphase, exposing the cellular basal side to the culture media and the apical side to the air. LRP‐1 shRNA decreased surfactant phospholipid, cholesterol and cholesteryl esters on the apical surface of the cells. LRP‐1 shRNA cells showed reduced gene expression of the rate limiting enzymes for fatty acid synthesis FAS, ACAT1 and SCD1, and of phosphatidylcholine synthesis enzymes choline transferase and choline kinase. Surprisingly, triglycerides and cholesteryl esters accumulated in intracellular lipid droplets in LRP‐1 shRNA cells, while phospholipids and unesterified cholesterol were not changed when compared to scrambled shRNA cells. LRP‐1 shRNA cells showed >3 fold overexpression of the fatty acid transporter CD36 and reduced gene expression of acyl‐CoA oxidase. After an overnight incubation with fresh media in the basal side, LRP‐1 shRNA cells showed higher media depletion of phospholipid, cholesterol esters and unesterifed cholesterol. Uptake of LDL‐derived phosphatidylcholine was not altered by LRP‐1 deletion, but the gene expression of phospholipid and cholesterol exporters ABCA1 and ABCG1 was reduced. Insulin signaling was affected by LRP‐1 deletion. LRP‐1 shRNA cells showed decreased protein expression of insulin receptor substrates 1 and 2 (IRS‐1 and IRS‐2) and increased phospho‐serine IRS‐1/total IRS‐1 ratio. After IGF‐1 treatment, LRP‐1 shRNA cells failed to Tyr‐phosphorylate the IGF‐1 receptor b, and phosphorylation of the downstream target Akt was lower than in scrambled shRNA cells These data suggest that LRP‐1 in type II pneumocytes is involved in lipid export, intracellular surfactant lipid metabolism and insulin signaling. Support or Funding Information FAMRI Young Clinical Scientist Award (to I.G.A); AHA postdoctoral fellowship (to I.G.A), and NHLBI (to R.F.F).