Abstract
Surfactant protein A (SP-A) is the most abundant protein associated with phospholipids in pulmonary surfactant. There are several lines of evidence that pulmonary and gastrointestinal epithelium produce closely related surface-active materials, although the presence of SP-A in gastrointestinal tract has so far not been reported. Indirect immunofluorescence experiments using different antibodies raised against rat pulmonary SP-A showed that some jejunal and colonic but not gastric epithelial cells positively stained for SP-A. Analysis of the proteins in cell lysates from rat small intestine and colon studied by Western blot revealed several immuno-reactive bands, including the characteristic triplet of 26-, 32-, and 38-kDa monomeric proteins, less strongly labeled than in lung cells, and higher molecular mass forms of 66 and 120 kDa also present in lung cells. The 66- and 120-kDa bands displayed the expected isoelectric pH of SP-A after two-dimensional electrophoresis. Alkylation induced conversion of the 120-kDa form (almost completely) and the 66-kDa form (partly) into the 26-38-kDa monomeric species. The presence of SP-A mRNA in rat stomach, small intestine, and colon was then searched for by conventional cDNA/reverse transcriptase-polymerase chain reaction. Products of appropriate size (372 base pairs) identical to that of pulmonary tissue were amplified in small intestine and colon but not in stomach or in other tissues used as controls. Cloning and sequencing of rat colon SP-A cDNA revealed the same sequence as the one reported for rat lung SP-A. Furthermore, analysis of the transcriptional initiation site of SP-A gene in colon by anchored-polymerase chain reaction showed that transcription was initiated at the same site in both colon and lung. These data, which demonstrate that small intestine and colon express SP-A constitutively and that this protein is present in some epithelial cells, extend the concept of intestinal surfactant and underline its close relationships to pulmonary surfactant.
Highlights
Rat surfactant-associated protein A (SP-A),1 an octadecameric sialoglycoprotein, binds strongly to surfactant glycerophospholipids and acts in a calcium-dependent manner to promote the transformation of the secreted lamellar bodies to tubular myelin within the pulmonary alveolus [1]
Whereas the presence of pulmonary surfactant proteins SP-B and SP-D has been reported in intestinal lumen [11] and SP-D in the stomach [39], SP-A has not yet been identified elsewhere than in pulmonary tissue
The upper half of rat small intestine and colon was flushed free of content with cold phosphate-buffered saline (PBS) (150 mM NaCl, 30 mM KCl, 10 mM Na2HP04, 15 mM KH2P04 ), pH 6, everted, and washed twice in the same solution
Summary
Gastrointestinal surfactant has been described as a hydrophobic layer of surface-active phospholipids between the apical border of epithelial cells and the luminal contents. In the rat small intestine, Eliakim and co-workers [8] described the presence oflamellar bodies morphologically similar to those in epithelial alveolar cells, in the enterocytes themselves, in the intercellular spaces, and adjacent to the apical membrane. To those in lung epithelium, these lamellar bodies contain saturated phosphatidylcholine and possess the ability to lower surface tension [4, 8]. We here report evidence for the expression of pulmonary surfactant protein A gene in small intestine and colon
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