The Collagen-like Region of Surfactant Protein A (SP-A) Is Required for Correction of Surfactant Structural and Functional Defects in the SP-A Null Mouse

Machiko Ikegami,Baher M Elhalwagi,Nades Palaniyar,Krista Dienger,Thomas Korfhagen,Jeffrey A Whitsett,Francis X Mccormack

doi:10.1074/jbc.m102054200

Machiko Ikegami, Baher M Elhalwagi + Show 5 more

Open Access

https://doi.org/10.1074/jbc.m102054200

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Abstract

Pulmonary surfactant isolated from gene-targeted surfactant protein A null mice (SP-A(-/-)) is deficient in the surfactant aggregate tubular myelin and has surface tension-lowering activity that is easily inhibited by serum proteins in vitro. To further elucidate the role of SP-A and its collagen-like region in surfactant function, we used the human SP-C promoter to drive expression of rat SP-A (rSPA) or SP-A containing a deletion of the collagen-like domain (DeltaG8-P80) in the Clara cells and alveolar type II cells of SP-A(-/-) mice. The level of the SP-A in the alveolar wash of the SP-A(-/-,rSP-A) and SP-A(-/-,DeltaG8-P80) mice was 6.1-and 1.3-fold higher, respectively, than in the wild type controls. Tissue levels of saturated phosphatidylcholine were slightly reduced in the SP-A(-/-,rSP-A) mice compared with SP-A(-/-) littermates. Tubular myelin was present in the large surfactant aggregates isolated from the SP-A(-/-,rSP-A) lines but not in the SP-A(-/-,DeltaG8-P80) mice or SP-A(-/-) controls. The equilibrium and minimum surface tensions of surfactant from the SP-A(-/-,rSP-A) mice were similar to SP-A(-/-) controls, but both were markedly elevated in the SP-A(-/-,DeltaG8-P80) mice. There was no defect in the surface tension-lowering activity of surfactant from SP-A(+/+,DeltaG8-P80) mice, indicating that the inhibitory effect of DeltaG8-P80 on surface activity can be overcome by wild type levels of mouse SP-A. The surface activity of surfactant isolated from the SP-A(-/-,rSP-A) but not the SP-A(-/-,DeltaG8-P80) mice was more resistant than SP-A(-/-) littermate control animals to inhibition by serum proteins in vitro. Pressure volume relationships of lungs from the SP-A(-/-), SP-A(-/-,rSP-A), and SP-A(-/-,DeltaG8-P80) lines were very similar. These data indicate that expression of SP-A in the pulmonary epithelium of SP-A(-/-) animals restores tubular myelin formation and resistance of isolated surfactant to protein inhibition by a mechanism that is dependent on the collagen-like region.

Highlights

Pulmonary surfactant isolated from gene-targeted surfactant protein A null mice (SP-A؊/؊) is deficient in the surfactant aggregate tubular myelin and has surface tension-lowering activity that is inhibited by serum proteins in vitro
The data presented here indicate that replacement of SP-A in the SP-AϪ/Ϫ mouse restores tubular myelin formation and corrects the defect in the resistance of isolated surfactant to protein inhibition
The collagen-like region/and or full oligomeric assembly plays an important role in the accommodation of SP-A in the alveolar lining fluid, because the ⌬G8-P80 mutation converts SP-A into an inhibitor of surfactant function

Summary

Introduction

Pulmonary surfactant isolated from gene-targeted surfactant protein A null mice (SP-A؊/؊) is deficient in the surfactant aggregate tubular myelin and has surface tension-lowering activity that is inhibited by serum proteins in vitro. Pressure volume relationships of lungs from the SP-A؊/؊, SP-A؊/؊,rSP-A, and SP-A؊/؊,⌬G8-P80 lines were very similar These data indicate that expression of SP-A in the pulmonary epithelium of SP-A؊/؊ animals restores tubular myelin formation and resistance of isolated surfactant to protein inhibition by a mechanism that is dependent on the collagen-like region. The ⌬G8-P80 mutation does not affect aggregation of lipid vesicles by the protein or the enhancement of the surface tension-lowering properties of surfactant lipids in the bubble surfactometer [14] These in vitro data suggested that a partial SP-A molecule containing a deletion of the collagen-like domain retained the structural elements that are required for the surface tension enhancing properties of SP-A. The objectives of this study were 2-fold: 1) to determine whether genetic replacement of SP-A in the SP-AϪ/Ϫ mouse corrects surfactant structural and functional defects and 2) to determine whether the collagen-like region of SP-A is required for tubular myelin formation and resistance of surfactant to protein inhibitors

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