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INTRODUCTION |
Surfactant protein-D
(SP-D)1 is a member of a
family of collagenous host defense lectins termed collectins (1, 2).
SP-D is secreted into the distal airways and alveoli of the lung (3-5) but is expressed also in other tissues (6-8). Each 43-kDa SP-D monomer
consists of an NH2-terminal domain containing two
conserved cysteine residues (Cys-15 and Cys-20), a collagenous domain,
a short neck sequence, and a COOH-terminal carbohydrate recognition domain (CRD) (9-12). Electron microscopy and proteinase digestion studies demonstrated that SP-D monomers are assembled into tetramers of
trimeric subunits (dodecamers) (13, 14). The two
NH2-terminal cysteine residues of SP-D are critical in the
formation of interchain disulfide bonds that stabilize the dodecameric
structure (13).
Substitution of serine for cysteine at positions 15 and 20 of the
mature protein results in the efficient secretion of trimeric subunits
corresponding to a single arm of the SP-D dodecamer (15). Mutant
recombinant rat protein migrates as a monomer on SDS-polyacrylamide gel
electrophoresis in the absence of sulfhydryl reduction and elutes as a
trimer rather than as a dodecamer from gel filtration columns under
nondenaturing conditions (16). The collagen domain forms a triple helix
as assessed by protease digestion (16). The trimeric protein is also a
functional lectin with the same saccharide selectivity as the wild-type
protein. RrSP-Dser15/20 binds to the hemagglutinin of the influenza
virus in a CRD-dependent manner (15, 16). However, the
mutant SP-D does not aggregate particulate ligands such as viral
particles and competitively inhibits aggregation mediated by wild-type
SP-D (15, 16).
High affinity binding of SP-D to various ligands in vitro
requires a trimeric CRD (17-19). However, previous studies using trimeric CRDs, RrSP-Dser15/20, and variably multimerized fractions of
recombinant human SP-D suggest that trimers are functionally univalent
and have a restricted range of biological activities. For example,
trimeric CRDs elicit the chemotaxis of neutrophils and mononuclear
phagocytes in vitro (20) and neutralize the respiratory
syncytial virus in vivo (21). In addition,
RrSP-Dser15/20, similar to wild-type SP-D, inhibits stimulated
proliferation of T-lymphocytes (22). However, other activities of SP-D
involve ligand aggregation and require higher orders of covalently
stabilized oligomerization. For example, trimeric CRDs inhibit the
hemagglutinin of the influenza virus, but unlike SP-D dodecamers, they
cannot mediate viral aggregation, enhance viral binding to neutrophils, or enhance the oxidative response to a bound virus (16, 19, 23).
Analyses of the SP-D gene-targeted mice indicated an important role of
SP-D in the regulation of surfactant lipid homeostasis and macrophage
function (24). Lungs of SP-D null mice have increased surfactant
phospholipid pools associated with abnormally enlarged and foamy
alveolar macrophages and hyperplastic type II cells with an increased
size of lamellar bodies (25, 26). SP-D
/
mice also develop
progressive pulmonary emphysema and subpleural fibrosis, associated
with chronic inflammation and increased matrix metalloproteinase and
increased oxidant production by alveolar macrophages (27).
Expression of rat SP-D (rSP-D) in distal respiratory epithelium of the
lung completely corrected the pulmonary lipid accumulation and
abnormalities in alveolar macrophages seen in SP-D null mice, whereas
the targeted increased expression of wild-type rat SP-D in normal mice
had no effect on endogenous SP-D, lung morphology, or surfactant
phospholipid content (28).
SP-D dodecamers bind to specific surfactant lipids and influence their
state of aggregation (29) and alter the presentation of various
particles to phagocytic cells in vitro (30-32). Based in
part on these observations, we hypothesized that the covalent oligomerization of trimeric subunits is required to mediate the effects
of SP-D on surfactant homeostasis and macrophage function in
vivo. For the present studies, transgenic mice were generated that
expressed single trimeric subunits by targeting the expression of
RrSP-Dser15/20 to peripheral respiratory epithelial cells of the lung
using the human surfactant protein-C promoter (33). The RrSP-Dser15/20
mutant protein did not correct lung phospholipids, alveolar macrophage
abnormalities, or emphysema in SP-D
/
null mice. Expression of the
mutant protein in wild-type mice interfered with the normal covalent
oligomerization of the endogenous mSP-D and caused emphysema and the
accumulation of foamy alveolar macrophages similar to that observed in
SP-D null animals.
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EXPERIMENTAL PROCEDURES |
Generation of Transgenic Mice--
The 1.3-kb mutant cDNA of
rat SP-D (RrSP-Dser15/20) was generated previously (16) and inserted
into the EcoRI site of the BGI-hSPC expression vector (34)
(Fig. 1). Restriction enzyme digestion
and partial DNA sequencing confirmed the orientation of the insert. The
vector contained the 3.7-kb human SP-C promoter, which drives
cell-specific expression in the bronchiolar and alveolar respiratory
epithelial cells (33). The transgene was microinjected into fertilized
FVB/N oocytes by the Children's Hospital Transgenic Core facility, and
founders were identified either by Southern blot analysis using a
32P-radiolabeled probe that recognized the RrSP-Dser15/20
cDNA or by transgene-specific PCR using upstream primer 5'-ATA GGA
CCC CAA GGC AAA CCAG-3' and downstream primer 5'-CAC CCC CCA GAA TAG AAT GAC AC-3'. Transgenic animals were crossed with SP-D null mice
(mSP-D
/
) (25) to generate heterozygous mice. Heterozygous mice were
bred to generate transgenic mice in either a wild-type (RrSP-Dser15/20+, mSP-D+/+) or SP-D null background (RrSP-Dser15/20+, mSP-D
/
). Transgenic mice were compared biochemically and
histologically with nontransgenic littermates (mSP-D+/+ or mSP-D
/
).
The absence of a PCR product for the mSP-D gene and the presence of a
438-base pair PCR product for the neomycin resistance gene confirmed
the mSP-D
/
genotype.

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Fig. 1.
Schematic representation of the
RrSP-Dser15/20 transgene. The construct was generated by inserting
the 1.3-kb RrSP-Dser15/20 cDNA into the EcoRI site of
the BGI-hSPC expression vector. The entire 1.3-kb cDNA fragment was
radiolabeled with [ -32P]dCTP and used as a probe in
the Southern blot analysis. A transgenic-specific 1,035-base pair PCR
product confirms the transgenic positive genotype using an upstream
primer in the RrSP-Dser15/20 sequence and a downstream primer in the
bovine growth hormone poly(A) sequence.
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Animal Husbandry--
The mice described under "Experimental
Procedures" were handled in accordance with approved protocols of the
Institutional Animal Care and Use Committee at Children's Hospital
(Cincinnati). All mice had been maintained in the vivarium under
barrier containment facilities. At the time of study, all mice appeared
healthy and were without evidence of infection. Sentinel mice in the
colony were serologically negative for common murine pathogens.
Western Blot Analysis--
Animals were weighed, anesthetized by
intraperitoneal injection of pentobarbital, and exsanguinated by
severing the distal aorta. Bronchoalveolar lavage was performed five
times with saline for each lung, and the volume was measured (35).
Bronchoalveolar lavage fluid (BALF) (25 µl) from each mouse was dried
and reconstituted in 15 µl of nonreduced Laemmli sample buffer
(Bio-Rad). After resolution with a 10-20%
SDS-Tris/glycine-polyacrylamide gel (NOVEX, San Diego, CA) and transfer
to a nitrocellulose membrane (Bio-Rad), the blots were blocked with 5%
nonfat milk and then incubated at room temperature overnight with
rabbit anti-mouse SP-D antiserum diluted 1:5,000 in Tris-buffered
saline with 0.1% Tween. Blots were washed with Tris-buffered
saline/Tween and incubated at room temperature for 4 h with a
1:10,000 dilution of peroxidase-conjugated goat anti-rabbit IgG
antibody (Calbiochem). After washing, blots were developed with a
chemiluminescence detection system (Amersham Pharmacia Biotech).
Generally, 4-8 mice from each genotype were analyzed for each mouse
line and representative results shown. Protein bands on a Western gel
were quantified using the computer program ImageQuant 1.2. The density
volume of each band is the sum of pixel density above background in the
band area.
Binding to Maltosyl-Sepharose--
BALF from 6-week-old mice
were incubated with maltose immobilized on 4% beaded agarose (Sigma)
overnight at 4 °C in TCB (20 mM Tris, pH 7.4, 10 mM CaCl2). After washing three times with TCB,
equal fractions of the maltosyl-Sepharose beads were incubated with 0, 2, 5, 10, and 25 mM maltose in TCB overnight at 4 °C. The maltosyl-Sepharose beads then were suspended in 15 µl of reduced Laemmli sample buffer (Bio-Rad) and resolved on a 10-20%
SDS-Tris/glycine-polyacrylamide gel. Western analysis with rabbit
anti-mouse SP-D antiserum was performed to detect the SP-Ds that bound
to the maltosyl-Sepharose beads.
Phospholipid Analysis--
Bronchoalveolar lavage was performed
five times with saline on each lung for 8-10-week-old mice. Lung
tissue was homogenized in saline after lavage. The amount of saturated
phosphatidylcholine (Sat PC) in BALF and lung homogenate was measured
as described previously (25). Animals from each genotype were analyzed,
and the differences between genotypes were evaluated by ANOVA Fisher analysis. Differences of p < 0.05 were considered significant.
Lung Morphology--
Mouse lungs (12 weeks old) were fixed at 25 cm of water pressure with 4% paraformaldehyde in phosphate-buffered
saline and processed into paraffin blocks. Seven-micrometer sections
from each lobe were stained with hematoxylin and eosin. Morphometric measurements on the ratio of alveolar parenchyma to airspace areas were
performed as described previously (36). Animals (4-6) from each
genotype were analyzed in each mouse line, and the differences between
genotypes were evaluated by ANOVA Fisher analysis. Differences of
p < 0.05 were considered significant.
Immunostaining of endogenous mSP-D and transgenic RrSP-Dser15/20 was
performed in lung tissue using an avidin-biotin-peroxidase technique
(Vectastain Elite ABC kit, Vector Laboratories). The rabbit anti-SP-D
antibody was generated against purified mouse SP-D and affinity
absorbed against lung homogenates from SP-D
/
mice (8).
Immunostaining was blocked completely by co-incubation with purified
mouse SP-D.
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RESULTS |
Transgenic Mouse Lines--
Eleven founder mice were identified by
Southern blot analysis of tail-clip DNA using 1.3-kb RrSP-Dser15/20
cDNA as a probe (data not shown). Germ line transmission was
demonstrated in eight founders using transgene-specific PCR on
offspring tail DNA (data not shown). The expression of transgenic
protein was confirmed in these eight mouse lines by Western blot
analysis of mouse BALF using a rabbit anti-mouse SP-D antibody (data
not shown). Two mouse lines, one with moderate (line 52) and the other
with high (line 75) levels of RrSP-Dser15/20 protein expression were
selected for further breeding and analysis.
Expression of RrSP-Dser15/20 Protein--
Mouse and rat SP-Ds are
92% identical, and the rabbit anti-mouse SP-D antibody recognized both
endogenous mSP-D and transgenic RrSP-Dser15/20 protein by Western blot
and immunohistochemistry. However, RrSP-Dser15/20 protein did not form
interchain disulfide bonds because of the substitution of serine for
cysteine at positions 15 and 20 (16). Therefore, under nonreducing
conditions on SDS gels, endogenous mSP-D was detected as an oligomer,
whereas RrSP-Dser15/20 protein migrated as a monomer. In line 52 mice,
the concentration of the mutant protein in BALF was similar to that of
endogenous mSP-D (Fig. 2A,
density volume ratio between RrSP-Dser15/20 in lane 3 and
mSP-D in lane 2 is 2.1). The monomeric form of SP-D was
detected in transgenic mice of both wild-type (mSP-D+/+) and null
(mSP-D
/
) backgrounds (Fig. 2A, lanes 1 and
3). No oligomeric forms of SP-D were detected in the null
(mSP-D
/
) background (Fig. 2A, lanes 3 and
4). In wild-type mice the larger oligomeric form of mSP-D
was decreased consistently in the transgenic mice (RrSP-Dser15/20+,
mSP-D+/+) compared with nontransgenic mice (mSP-D+/+) (Fig.
2A, lanes 1 and 2). In mice from line
75, the concentration of RrSP-Dser15/20 was increased markedly compared
with that of the endogenous mSP-D (Fig. 2B, density volume
ratio between RrSP-Dser15/20 in lane 3 and mSP-D in
lane 2 is 24). The slower mobility oligomeric forms of mSP-D
typical of wild-type mice were not detectable in line 75 transgenic
mice (RrSP-Dser15/20+, mSP-D+/+) or in mSP-D
/
null mice (Fig.
2B, lanes 1, 3, and 4).

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Fig. 2.
Western blot analyses of SP-D in
bronchoalveolar lavage fluid from line 52 (A) and line
75 (B) mice. The primary antibody is rabbit
anti-mouse SP-D, and the secondary antibody is goat anti-rabbit IgG.
Numbers on the right are molecular mass
markers in kDa (MW KD). The monomeric form of SP-D migrates
at 43 kDa, and the oligomeric form of SP-D migrates slower than 200 kDa.
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RrSP-Dser15/20 Binds Maltosyl-Sepharose--
To assess whether the
mutant SP-D maintained lectin activity, binding of wild-type SP-D and
RrSP-Dser15/20 to maltosyl-Sepharose was assessed. Both wild-type and
mutant SP-D in the mouse BALF maintained the ability to bind to maltose
in vitro (Fig. 3). Increasing concentrations of maltose inhibited binding of wild-type and mutant SP-D to the maltosyl-Sepharose beads (Fig. 3). The affinity of wild-type SP-D and mutant SP-D to maltosyl-Sepharose was similar.

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Fig. 3.
Maltosyl-Sepharose binding activities of SP-D
and RrSP-Dser15/20 proteins. Maltosyl-Sepharose beads were
incubated with BALF from wild-type, line 52, and line 75 (RrSP-Dser15/20+, mSP-D / ) mice, washed extensively, incubated again
with an increased concentration of maltose, and resolved on reduced
SDS-polyacrylamide gel electrophoresis. Endogenous mSP-D and mutant
RrSP-Dser15/20 proteins bound to the maltosyl-Sepharose beads were
detected by Western blot analysis using rabbit anti-mouse SP-D
antibody. WT, wild type.
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RrSP-Dser15/20 Does Not Correct Lung Phospholipids--
Sat PC in
BALF and lung homogenates was assessed in transgenic (RrSP-Dser15/20+,
mSP-D+/+ and RrSP-Dser15/20+, mSP-D
/
), null (mSP-D
/
), and
wild-type (mSP-D+/+) mice. After normalization for body weight, there
were no statistically significant differences in lung phospholipid
levels between lines 52 and 75 mice. Therefore, data from lines 52 and
75 mice were pooled together and presented in Fig.
4. Although statistically significant
increased Sat PC pool sizes were observed in mSP-D
/
mice compared
with wild-type mice (Fig. 4, compare bars 1 and 2 with bars 3 and 4, respectively), RrSP-Dser15/20 did not correct Sat PC pool sizes in SP-D null mice
(Fig. 4, bars 3 and 4) and did not perturb Sat PC
levels in wild-type mice (Fig. 4, bars 1 and
2).

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Fig. 4.
Saturated phosphatidylcholine pool
sizes. Alveolar tissue and total lung Sat PC concentrations were
determined in wild-type (mSP-D+/+), null (mSP-D / ), and transgenic
(RrSP-Dser15/20+, mSP-D+/+ and RrSP-Dser15/20+, mSP-D / ) mice and
normalized for body weight. Data from lines 52 and 75 were not
different and therefore were pooled. Values are mean ± S.E. ANOVA
Fisher analysis showed that Sat PC in mice from the mSP-D+/+ background
were significantly lower than in mice with mSP-D / background
(p < 0.0001). There were no statistically
significant differences in Sat PC in RrSP-Dser15/20+ and
RrSP-Dser15/20 mice whether in mSP-D+/+ (wild-type) or mSP-D /
(null) backgrounds.
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Lung Morphology--
Expression of the RrSP-Dser15/20 transgene in
the lung did not correct the emphysema or foamy macrophage accumulation
typical of the mSP-D
/
mouse lungs. Enlarged foamy alveolar
macrophages were detected in both transgenic line 52 (Fig.
5E) and line 75 (Fig.
5F) in the mSP-D
/
background. Morphometric analysis
showed that the RrSP-Dser15/20 protein did not correct emphysema as
determined by the lack of correction in the relative proportion of
respiratory parenchyma to airspace areas in either transgenic mouse
line (Fig. 6, bars 3 and
4).

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Fig. 5.
Lung histology. Lungs were fixed and
stained with hematoxylin and eosin. A, lungs from wild-type
mice; B, RrSP-Dser15/20 (line 52) in wild-type background;
C, RrSP-Dser15/20 (line 75) in wild-type background;
D, SP-D / mice; E, RrSP-Dser15/20 (line 52) in
SP-D / background; F, RrSP-Dser15/20 (line 75) in
SP-D / background. The arrows point to enlarged foamy
alveolar macrophages. The arrowheads point to normal
macrophages.
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Fig. 6.
Effects of mutant SP-D on the relative
proportions of lung parenchyma to airspace areas.
Parenchyma/airspace ratios were determined by morphometric
analysis. Values are mean ± S.E. and were compared by
ANOVA Fisher analysis. The upper panel represents data from
line 52 mice, and the lower panel represents data from line
75 mice. Parenchyma/airspace ratios were similar in RrSP-Dser15/20+,
mSP-D+/+ mice compared with mSP-D+/+ mice in line 52. In line 75 mice,
parenchyma/airspace ratios were decreased significantly in
RrSP-Dser15/20+, mSP-D+/+ mice compared with wild-type mice (*,
p = 0.0114). Parenchyma/airspace ratios were decreased
in both RrSP-Dser15/20+, mSP-D / and mSP-D / mice, reflecting
emphysema in both mice.
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Pulmonary morphology in wild-type mice expressing the RrSP-Dser15/20
transgene differed in the two mouse lines examined. In line 52, which
had a moderate RrSP-Dser15/20 protein expression (Fig. 2A),
alveolar macrophages were normal in appearance (Fig. 5B),
and the relative proportion of respiratory parenchyma to airspace areas
was not different from that in the wild-type mice (Fig. 6, upper
panel, bars 1 and 2). Surprisingly, in
RrSP-Dser15/20+, mSP-D+/+ transgenic mouse line 75, in which high
levels of RrSP-Dser15/20 protein were expressed, lung morphology was
similar to that in the mSP-D
/
null mice. Enlarged foamy macrophages
were detected readily (Fig. 5C), and the enlargement of
airspace was shown by the significant decrease in the relative
proportion of respiratory parenchyma to airspace areas (Fig. 6,
lower panel, bars 1 and 2). The
RrSP-Dser15/20 transgene did not correct the abnormal alveolar
parenchyma/airspace ratio in mSP-D
/
mice (Fig. 6, bars 3 and 4).
The expression pattern of SP-D in lines 52 and 75 was evaluated in
lungs by immunohistochemistry. Weak SP-D staining was observed in
alveolar type II cells in the lungs of wild-type mice (Fig. 7A), whereas no SP-D staining
was detected in the lungs of mSP-D
/
mice (Fig. 7D). In
line 52, expression of SP-D was detected readily in both alveolar type
II cells and bronchiolar epithelial cells (Fig. 7B). In line
75 transgenic mouse lungs, SP-D was detected not only in type II cells
and bronchiolar epithelial cells but also in foamy alveolar macrophages
(Fig. 7C). This pattern of SP-D staining was similar in line
75 in both wild-type and mSP-D
/
backgrounds. In both transgenic
mouse lines expressing RrSP-Dser15/20 in the mSP-D
/
background,
SP-D immunostaining was detected in type II cells, bronchiolar
epithelial cells, and foamy alveolar macrophages (Fig. 7, E
and F).

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Fig. 7.
Immunostaining of SP-D in lungs from adult
mice. The panel represents data from wild-type mice
(A), RrSP-Dser15/20 (line 52) in wild-type background
(B), RrSP-Dser15/20 (line 75) in wild-type background
(C), SP-D / mice (D), RrSP-Dser15/20 (line 52)
in SP-D / background (E), and RrSP-Dser15/20 (line) 75 in
SP-D / background (F). Thin arrows point to
SP-D staining in type II cells, thick arrows point to SP-D
staining in bronchiolar epithelial cells, and arrowheads
point to SP-D staining in macrophages.
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DISCUSSION |
The expression of a mutant RrSP-Dser15/20 protein was directed to
bronchiolar and alveolar epithelial cells with the human SP-C promoter
in wild-type and SP-D
/
null mice. Mutations of the two
amino-terminal cysteine residues resulted in the secretion of a protein
that bound to maltose and migrated exclusively as monomers on
SDS-polyacrylamide gel electrophoresis in the absence of reduction.
These findings are consistent with the results described by
Brown-Augsburger et al. (15) for transfected CHO-K1 cells. Given that interchain bonds are required for the stability of dodecamers at 37 °C in vitro (16), the absence of
disulfide cross-linked oligomers is consistent with the absence of
dodecamers in vivo. Because expression of the RrSP-Dser15/20
mutant protein failed to correct phospholipid accumulation, foamy
macrophage production, or emphysema, the findings strongly suggest that
oligomerization of trimeric subunits is required for these
SP-D-dependent functions in vivo.
In both transgenic mouse lines, expression of RrSP-Dser15/20 failed to
rescue the histological and biochemical phenotypes of SP-D
/
mice.
The characteristic phenotype of mSP-D null mice, with the increased
pool size of alveolar phospholipids, abnormal foamy macrophages, and
emphysema, persisted despite high levels of RrSP-Dser15/20 proteins. In
contrast, a previous study demonstrated that expression of the normal
rSP-D at high levels rescued all aspects of the pulmonary abnormalities
seen in mSP-D
/
null mice using the same human SP-C promoter used in
this study (28). Although the findings suggest that multivalency of
trimeric subunits is required, it remains possible that cysteines 15 and 20 or more subtle local conformational perturbations render the
protein defective with respect to its ability to rescue the SP-D null
phenotype. In this regard, thermal stability of the collagen domain of
RrSP-Dser15/20 is decreased, presumably secondary to the absence of
interchain disulfide cross-links (16).
In RrSP-Dser15/20 transgenic mouse line 75, in which normal disulfide
cross-linked oligomers of SP-D were undetectable by Western blot
analysis, lung morphology was similar to that characteristic of
mSP-D
/
null mice, with enlarged foamy macrophages and emphysema. The expression of RrSP-Dser15/20 protein did not disrupt lung morphology in line 52, in which the mutant protein was expressed at
levels that did not eliminate the formation of disulfide cross-linked oligomers of the endogenous mSP-D. These results suggest that overexpression of chains lacking the capacity to participate in interchain disulfide bonds may interfere with the function of SP-D,
causing a dominant negative effect through formation of heteropolymers
of wild-type mouse and mutant rat chains that are unable to participate
in the formation of stable dodecamers. Given that the mouse and rat
proteins are identical in length and domain structure, it is likely
that folding of the carboxyl-terminal CRDs and neck domains of the
heteropolymers is unaltered. Indeed, the lectin activity of the mutant
SP-D was retained, supporting the likelihood that the folding of the
CRDs was maintained. However, it is unclear whether differences in
functions of the mutant SP-D molecules are related to activities
determined by the serine at positions 15 and 20 or to the effect of the
mutant protein on oligomerization of the wild-type SP-D, leaving the
lung relatively deficient in functional disulfide-linked oligomeric
forms. Alternatively, heteropolymeric SP-D could be degraded, resulting
in selective secretion of mutant SP-D. Although the lungs of wild-type
mice expressing the mutant SP-D at high levels (line 75) contained numerous foamy macrophages and developed emphysema, pulmonary Sat PC
concentrations were not increased significantly, suggesting that some
function of the endogenous mSP-D was maintained. It was possible that a
very small amount of appropriately assembled endogenous mSP-D was
present in the BALF but was undetectable by Western blot. It remains
possible that various activities of SP-D may be
concentration-dependent. For example, the concentrations of
SP-D inducing neutrophil chemotaxis (20) are 100-1,000 times lower
than those required for aggregating particles (16).
In summary, the present findings strongly support the concept that the
dodecameric structure of SP-D is essential for some of its homeostatic
functions in vivo. Increased expression of the mutant SP-D
in wild-type mice decreased the concentration of the normal disulfide
cross-linked oligomeric forms of mouse SP-D and caused emphysema and
foamy macrophage accumulation in the absence of alterations in alveolar
phospholipid concentrations. Thus, some aspects of the SP-D null
phenotype, namely the pulmonary emphysema and production of foamy
alveolar macrophages, are not likely caused by increased surfactant
phospholipids. Although the human lung contains a predominance of
highly oligomerized SP-D, trimeric species have been identified in the
lungs of some patients with alveolar proteinosis (37), and less highly
aggregated SP-D forms may be generated by proteolytic degradation in
the setting of infection or lung injury. Polymorphisms in nucleotide sequences of the collagen domain of the serum mannose binding lectin
altered oligomerization that was associated with defects in its host
defense function (38). We therefore speculate that increased levels of
mutant SP-D forms that interfere with higher order oligomerization may
influence the function of SP-D that might contribute to the
pathogenesis of certain lung diseases.