the effect of the inflammatory process in the lungs in bringing about dysfunction of intrinsic surfactant has been recognized for years. A variety of factors have been implicated, including the increased presence of a variety of proteins such as those in edema fluid, damage to the surfactant phospholipids with resulting increased presence of lysophospholipids and fatty acids, hydrolytic activity on surfactant proteins by proteolytic enzymes, decreased surfactant synthesis by type II epithelial cells after damage by oxidant, etc. In cases where the effect of the inflammatory exudate is primarily alveolar, the result of surfactant dysfunction is collapse of the alveoli and atelectasis. This is observed in several disease states such as acute lung injury/acute respiratory distress syndrome. More recently, the question has been repeatedly raised that when the inflammatory process is primarily in the bronchial tree, dysfunction of the surfactant occurs in the bronchioles, leading to their collapse and hyperexpansion of alveoli rather than collapse. It has been proposed for the past 15 years that this pathogenic mechanism may participate in asthma.
In the current articles in focus, Hite and colleagues (Refs. 7 and 8, see pp. L610 and L618 in this issue) have extended significantly this latter theory with the observation that allergen challenge of patients with asthma results in a decrement of surfactant function not observed with saline challenge. Of particular note, the dysfunction of the surfactant could be attributed most clearly to a diminution in phosphatidylglycerol (PG) [and perhaps phosphatidylinositol (PI)] rather than the presence of surfactant-bound protein, loss of phosphatidylcholine (PC), increased levels of lysophospholipids, or the presence of eosinophils. Secretory phospholipase A2 (sPLA2) activity was observed in lavage fluid from the antigen-challenged patients that, in the surfactant-dysfunctional subgroup, exhibited a preference for PG rather than PC. The hydrolysis of PG most closely correlated with sPLA2 of groups IIA and IID.
Although these studies cast light on the relationship of allergic inflammation to the loss of surfactant function in the asthmatic process, the finding that the surfactant dysfunction is most accurately related to a specific loss of the acidic phospholipids PG and PI and not to PC (or other potential factors noted above) opens further questions regarding the molecular mechanisms involved in surfactant function. The widely held theory is that surfactant decreases surface tension during pulmonary expiration by achieving a greatly enriched, nearly crystalline monolayer of dipalmatoylphosphatidylcholine (DPPC) as the surface area of the alveolus is reduced in the expiratory phase of normal tidal ventilation. This is presumably achieved by removal from the monolayer non-DPPC phospholipids by surfactant protein B (SP-B), which bonds preferentially the acidic phospholipids. Then, during inspiration, alveoli are stated to increase in size and surface area with accompanying return of acidic phospholipids into the monolayer. However, this theory has been brought into question by a number of studies:
1) Mixtures of SP-B with DPPC and PG in a Langmuir trough using concentrations of SP-B at or above physiological levels revealed at most two to three molecules of phospholipid removed during compression, without preference for PG or PI (14, 15). In other studies, compression of monolayers of calf lung surfactant extract (CLSE) resulted in the formation of disk-like structures without loss of individual phospholipid components from the monolayer (3, 12).
Possible enrichment of the monolayer by DPPC from the subphase during compression was examined during surface compression in a Langmuir trough (9, 10) using increasing concentrations of SP-B and subphase vesicles of DPPC/PG. Levels of SP-B twice those at physiological concentration were required to obtain transfer (measured indirectly), but a preference for DPPC exchange into the monolayer was not recorded.
2) If, during compression, according to the “squeeze-out” hypothesis, low levels of acidic phospholipids in the monolayer and enriched DPPC are required for high levels of activity, then lower concentrations of PG and PI would be associated with increased activity, not a decrease in surfactant activity as observed in the present studies (7, 8).
3) Observations have been made on the development of DPPC-rich domains, the so-called liquid-condensed (LC) domains in the surfactant monolayer by epifluorescence or Brewster angle microscopy during surface compression at 20°C. With CLSE, known to contain normal or near-normal quantities of the various components of native pulmonary surfactant, the LC zones at surface pressures similar to those in the alveolus occupied no more than 5% of the surface area especially at 37°C and in the presence of neutral lipids (4, 5, 11). The presence of acidic phospholipids, especially PG and PI, shown in the two articles in focus, are present with all the other components of the surfactant in the so-called liquid-expanded (LE) zone, i.e., the region of the monolayer around the LC zones. Because the LE zones in whole surfactant preparations have been shown to represent by far the greatest portion of the surface area at surface pressures equal to those in the alveolus, the molecular actions accounting for the surfactant function of alveolar stability most certainly reside in this, the LE zone.
4) The assumption that the surfactant monolayer of the alveolus undergoes compression and expansion during normal tidal ventilation has been questioned. Mathematical modeling and studies of the surface area of the branching bronchial airways indicate that the enormous increase in surface area achieved after the 16th branching segment, i.e., at the beginning of the alveolar or respiratory zone, is sufficiently great that gas molecules move not by bulk movement, i.e., convection, but by diffusion (17, 18). This indicates that in normal tidal ventilation, there is negligible change in alveolar size. This conclusion has received direct support from microscopic observations of alveoli in situ (13). The greatest change in normal alveolar areas was found to be 2.4%. Removal of surfactant, however, resulted in large changes in alveolar size during respiration. The lack of changes of alveolar size in the intact lung during normal respiration is incompatible with the theoretical requirement for DPPC enrichment.
These varied studies are in direct conflict with the squeeze-out hypothesis.
A second mechanism of the molecular mechanism of pulmonary surfactant function has gained interest. This may be called the theory of SP-B-induced lateral stability. The theory evolved from studies of peptides synthesized according to sequences of SP-B amino acids or mimicking the sequences (1). When combined with DPPC, PG, neutral lipids, and fatty acids, surfactant activity was achieved (1, 6). Such synthetic SP-B peptide surfactants have been studied in vivo extensively as exogenous surfactant (2) and have successfully undergone clinical trials through the final, phase 3, trials in premature infants in Europe and the Americas (8a, 13a).
The activity of the SP-B peptides requires a combination of stretches of hydrophobic amino acid residues and intermittent basic or positively charged residues. Like SP-B, the peptides have overall hydrophobicity and lie in the acyl side chains of the phospholipids, with strong electrostatic interactions between the positively charged amino acids and, presumably, the negatively charged phosphates of the polar head groups (1). This bonding of peptide and phospholipid molecules produces a cohesiveness or lateral stability to the phospholipid monolayer, essential for holding phospholipid molecules in place in the surfactant monolayer of the alveolus. The lateral stability produced in phospholipid layers by SP-B or SP-B mimic peptides is supported by the findings of an increase in phospholipid head group packing order by Raman spectroscopy and an increase in melting temperature of the phospholipids (16) induced by the presence of SP-B peptides. A role of SP-B and possibly the PG and PI may also account for the formation and folding of flexible phospholipid sheets that form in the monolayer in compression isotherms (6). Without SP-B, fracturing and loss of fragments of the monolayer occurred into the subphase. Thus SP-B provided a cohesiveness to molecules of phospholipids. In other studies, SP-B was observed to induce large sheets from DPPC:PG liposomes which were further enlarged in the presence of Ca2+ (11a). Thus, in the stable alveolus, a combination of DPPC, acidic phospholipids, neutral lipids, fatty acids, and SP-B or SP-B peptides appear to exist in such a manner that the SP-B or SP-B peptides bind together the phospholipids with their rigid acyl side chains, and by virtue of this lateral stability, the cohesive monolayer is able to prevent collapse of the alveolus.
The contributions by Hite et al. (7, 8) thus shed light on the requirement of acidic phospholipids in surfactant function and point to their participation in the interaction of SP-B with all phospholipid classes in the surfactant monolayer. This will be a stimulus for future studies focusing on the role of these acidic phospholipids.
The references provided are not intended to be comprehensive, but for purposes of illustration. Readers may contact the author for more complete referential material.
- Copyright © 2005 the American Physiological Society