The optimal fraction of cholesterol in lung surfactants remains controversial;the role of cholesterol in lung surfactant function is still unknown. Small amounts of cholesterol, when added to DPPC or to clinical lung surfactants, reduce the surface shear viscosity by orders of magnitude, without altering the minimum surface tension. We have found that cholesterol separates into a disordered "interphase" that reduces the line tension between semi-crystalline DPPC-rich domains, which, in turn, dramatically alters domain morphology. This hypothesize that this interphase "lubricates" flow, causing the reductions in monolayer viscosity and elasticity, thereby enhancing the surfactant's ability to flow and cover the interface. At higher cholesterol concentrations, we hypothesize that the interphase properties eliminate the monolayer necessary monolayer cohesion so that collapse occurs at higher surface tensions. These observations suggest an optimal cholesterol content exists for a replacement lung surfactant. We will determine this optimal cholesterol content by measuring the shear viscosity and elasticity of clinical and model lung surfactants as a function of cholesterol composition using macro- and micro- rheology instruments unique to our laboratory. These mechanical properties will be correlated with isotherms, fluorescence and atomic force microscopy, and grazing incidence synchrotron X-ray diffraction to determine how cholesterol alters the molecular packing of lung surfactant lipids, which determines the mechanical properties of monolayers necessary for low surface tensions and rapid respreading and adsorption. Our goal is to determine the physiologically optimal viscosity and elasticity for rapid spreading and low surface tension and how best to achieve this optimum by controlling the cholesterol, lipid and protein fractions of a synthetic replacement lung surfactant for respiratory distress syndrome. In addition to an optimal composition, sufficient surfactant must be adsorbed to the interface from the alveolar fluid during the respiratory cycle. The lung surfactant specific proteins SP-A, B and C, along with lipids such as phosphatidylglycerol and cholesterol, are hypothesized to enhance exchange between surfactant in the subphase and the interface. However, little quantitative evidence for specific lipid and/or protein exchange exists. Also unknown is the surface pressures at what adsorption occurs, or if adsorption occurs during compression or expansion of the interface. To address this hypothesis, we will map out the three- dimensional distribution of lung surfactant components from the interface to the subphase using vertical and horizontal optical sectioning with a confocal microscope and multiple fluorescent dyes. We expect that SP-A, B and C promote adsorption;however, we do not know if specific lipids or proteins are adsorbed preferentially to the interface to optimize the monolayer composition that collapses at high surface pressures. Native SP-A, B and C will be compared to peptide mimics to evaluate the efficacy of the peptides.

Public Health Relevance

A lack of lung surfactant, often due to premature delivery, is responsible for neonatal respiratory distress syndrome (NRDS). NRDS affected an estimated 24,000 newborns in the US in 2002, and remains a complication in about 1% of all pregnancies to this day. Current treatments utilize replacement surfactants derived from animal sources, which vary in lipid and protein composition depending on the source, and from batch to batch depending on processing conditions and the types of animals used. The goal of this research is to develop a fundamental understanding of composition-function relationships to determine the composition of an optimized synthetic replacement lung surfactant. Such a surfactant should reduce the costs of NRDS treatment, improve product uniformity, and eliminate the possibility of contamination with infectious agents, thereby improving the efficacy of treatment of NRDS. Our work is also designed to understand the origins of surfactant inhibition in meconium aspiration or acute lung injury, and devise surfactant treatments to better address these conditions.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
5R01HL051177-20
Application #
8719150
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Lin, Sara
Project Start
Project End
Budget Start
Budget End
Support Year
20
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Engineering (All Types)
Type
Biomed Engr/Col Engr/Engr Sta
DUNS #
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
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Zasadzinski, Joseph A; Stenger, Patrick C; Shieh, Ian et al. (2010) Overcoming rapid inactivation of lung surfactant: analogies between competitive adsorption and colloid stability. Biochim Biophys Acta 1798:801-28

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