Am. J. Respir. Cell Mol. Biol., Vol 12, No. 2, Feb 1995, 238-249.
Analysis of fused-membrane structures in bronchoalveolar lavage fluid from patients with alveolar proteinosis
Y Suzuki, HQ Shen, A Sato and S Nagai
Department of Molecular Pathology, Kyoto University, Japan.
Alveolar macrophages of patients with alveolar proteinosis, obtained by
lung lavage, contain a large number of intracytoplasmic inclusions with
mutlilamellar membranous structures, called fused-membrane structures,
having a periodicity of 4.7 nm. To analyze the composition of these
structures, we concentrated them from the bronchoalveolar lavage fluid of
patients by a combination of sucrose density-gradient ultracentrifugation
and enzyme hydrolysis using proteinase K. Fused- membrane structures were
most numerous (26.2%) in a fraction obtained at the interface between 1.0
and 1.1 M sucrose solutions separated after proteinase K hydrolysis. This
fraction was rich in acidic phospholipids. Hydrophobic surfactant
apoproteins constituted about 77% of the proteins in this fraction, and a
remarkable increase in the content of surfactant-associated protein C
(SP-C) was found. The phospholipid-to-protein ratio was 0.25:1. Based on
these results, we tried to reconstitute fused-membrane structures from
purified lipids and hydrophobic proteins isolated from the patients' lavage
fluid or from pig lungs. Only in the presence of both phospholipids and
SP-C were similar multilamellar structures, having a periodicity of 4.3 to
4.5 nm, formed. These results suggest that fused-membrane structures have a
close relationship to a hydrophobic surfactant-associated protein, SP-C,
which accumulates in alveolar macrophages, possibly by incomplete
digestion.
