Caco-2 cell transfection by rat intestinal alkaline phosphatase cDNA increases surfactant-like particles.
The rat enterocyte produces a particle with surfactant-like properties (including a whorled appearance, enrichment for dipalmitoyl phosphatidylcholine, and ability to lower surface tension) that also is enriched for intestinal alkaline phosphatase. Human Caco-2 cells grown on polycarbonate filters were utilized to study the secretion of these particles and exhibited whorls and strands of unilamellar membranes, particularly concentrated at the apical pole or near junctional complexes. Concentrated culture medium from these cells separated on continuous NaBr gradients revealed a fraction at density = 1.07 g/l enriched for phosphatidylcholine and intestinal alkaline phosphatase. This fraction contained membranous sheets containing alkaline phosphatase, detected by immunolocalization. Phosphatidylcholine comprised 54% of phospholipid in this fraction, compared with 20% in brush borders. When Caco-2 cells were transfected with cDNA encoding rat intestinal alkaline phosphatase, cellular phosphatase activity increased twofold, but activity in the medium increased 14-fold to > 200 (average 32)-fold. Ultrastructurally, compared with mock-transfected cells or cells transfected with human placental alkaline phosphatase, transfection with rat intestinal alkaline phosphatase cDNA led to intracellular and extracellular accumulation of surfactant-like particles. We conclude that surfactant-like particles are produced by Caco-2 cells, and their production can be enhanced by transfection with a cDNA encoding a protein known to be associated with such particles.
Tietze
C C
CC
Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
Becich
M J
MJ
Engle
M
M
Stenson
W F
WF
Mahmood
A
A
Eliakim
R
R
Alpers
D H
DH
D000469:Alkaline Phosphatase / Q000235:genetics*; D000818:Animals; D002461:Cell Line, Transformed; D002499:Centrifugation, Density Gradient; D004247:DNA; D007413:Intestinal Mucosa / Q000378:metabolism*; D007422:Intestines / Q000166:cytology / Q000201:enzymology; D008854:Microscopy, Electron; D016253:Microscopy, Immunoelectron; D008871:Microvilli / Q000378:metabolism; D011663:Pulmonary Surfactants / Q000378:metabolism*; D051381:Rats; D014162:Transfection
D011663:Pulmonary Surfactants; D004247:DNA; D000469:Alkaline Phosphatase