1 Department of Pathology and Molecular Medicine and Centre for Gene Therapeutics, McMaster University, Hamilton, Ontario, Canada L8N 3Z5; 2 Medizinische Klinik, Julius-Maximilians-Universität, 97080 Würzburg, Germany; and 3 University of Rochester School of Medicine, Rochester, New York 14642-8692
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ABSTRACT |
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Transforming growth factor (TGF)- is a key cytokine in
the pathogenesis of pulmonary fibrosis, and pharmacological
interference with TGF-
can ameliorate the fibrotic tissue response.
The small proteoglycans decorin and biglycan are able to bind and
inhibit TGF-
activity in vitro. Although decorin has anti-TGF-
properties in vivo, little is known about the physiological role of
biglycan in vivo. Adenoviral gene transfer was used to overexpress
active TGF-
, decorin, and biglycan in cell culture and in murine
lungs. Both proteoglycans were able to interfere with TGF-
bioactivity in vitro in a dose-dependant manner. In vivo,
overexpression of TGF-
resulted in marked lung fibrosis, which was
significantly reduced by concomitant overexpression of decorin.
Biglycan, however, had no significant effect on lung fibrosis induced
by TGF-
. The data suggest that differences in tissue distribution
are responsible for the different effects on TGF-
bioactivity in
vivo, indicating that decorin, but not biglycan, has potential
therapeutic value in fibrotic disorders of the lung.
pulmonary fibrosis; extracellular matrix; treatment
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INTRODUCTION |
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FIBROSIS IS A MAIN FEATURE of various human organ disorders, from lung and kidney fibrosis to liver cirrhosis and atherosclerosis. It is commonly present in chronic and end stages of disease processes. Morphologically, fibrosis is characterized by a disproportionate increase and disordered deposition of extracellular matrix (ECM), resulting in distortion and irreversible loss of organ function. Once present, there is currently no curative treatment for fibrosis (19).
Cytokines have a crucial role in the pathophysiology of fibrosis
independent of the system involved. Fibrotic diseases are likely the
product of an overwhelming repair process after tissue injury (5,
28). The "normal scenario" after injury shows initial
expression of proinflammatory cytokines such as interleukin-1, interleukin-6, and tumor necrosis factor-, which help the immune system to eliminate the injurious agent. Shortly afterward, other cytokines and growth factors, among them transforming growth factor (TGF)-
, connective tissue growth factor, and platelet-derived growth
factor, are expressed to limit the inflammation and repair the damage.
Usually, all those cytokines are in a subtle balance. During the
development of fibrosis, however, profibrotic cytokines are
overexpressed and induce overwhelming repair and accumulation of ECM
(5, 18, 28).
One of the key profibrotic cytokines is TGF-, which is
chemotactic for fibroblasts, induces the synthesis of matrix proteins and glycoproteins, and inhibits collagen degradation by induction of
protease inhibitors and reduction of metalloproteases (16, 28). We have previously shown that transient overexpression of
TGF-
by adenoviral gene transfer induces a severe fibrotic reaction
in the lungs (25). On the other hand, neutralization of
TGF-
using antibodies significantly reduced experimental lung and
kidney fibrosis (4, 10).
ECM components such as collagen, fibronectin, and proteoglycans also
have effects on stromal cell growth and are able to stimulate protein
synthesis. These effects can be either pro- or antifibrotic. Collagen
and fibronectin are known to act as chemoattractants for fibroblasts
and enhance connective tissue synthesis (11). Other
components such as decorin have opposite effects on the ECM, acting
likely through interference with profibrotic cytokines (22). Decorin belongs to the group of small, leucine-rich
proteoglycans and is thought to be a natural inhibitor of TGF-,
capable of binding and neutralizing significant amounts of the cytokine
(12, 29). Decorin has been successfully employed to reduce
tissue fibrosis in different disease models in kidney, lung, and
vasculature (8, 9, 14, 17). Biglycan is another small
proteoglycan related to decorin, and its ability to interfere with
TGF-
has been demonstrated in vitro (12). To date,
biglycan has not been shown to interfere with the activity of TGF-
in vivo.
Small proteoglycans have important structural and functional roles in
tissue remodeling and fibrosis (13, 22). We report here
about the ability of two different proteoglycans, human decorin and
human biglycan, to interfere with TGF- in vitro and in vivo in a
model of TGF-
-mediated fibrogenesis. Proteoglycans and active TGF-
were delivered using adenoviral gene transfer. Decorin was able
to block TGF-
in vitro and in vivo as we have previously shown in a
model of bleomycin-induced lung fibrosis. In contrast, biglycan was
effective in vitro to inhibit TGF-
but failed to reduce the fibrotic
tissue response in vivo.
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METHODS AND MATERIALS |
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Recombinant adenovirus.
The construction of adenoviral vectors is described in detail elsewhere
(2, 17, 25). For the current study, full-length human
decorin and human biglycan cDNAs (gift of L. W. Fisher, National
Institutes of Health, Bethesda, MD) were cloned into shuttle vectors
with a human CMV promoter and cotransfected with a virus-rescuing
vector. The resulting replication-deficient virus (AdDec and AdBig) was
amplified and purified by CsCl gradient centrifugation and PD-10
Sephadex chromatography, and finally plaque titered on 293 cells.
AdTGF223/225 (a mutant TGF-
1 which was translated
into spontaneously bioactive TGF-
) and control vectors (AdDL) with
no insert in the E1 region were produced in the same way.
TGF- bioassay.
Murine lung fibroblasts (ATCC CCL-206) were plated in a
100-cm2 flask and left in MEM supplemented with 1%
L-glutamine, 1% penicillin-streptomycin, 0.4%
amphotericin, and 10% newborn calf serum until confluent. The cells
were infected with AdDec, AdBig, AdTGF
223/225, or AdDL
at a multiplicity of infection of 20 pfu/cell. After 14 h,
supernatants were removed and cells were washed six times with
phosphate-buffered saline (PBS). Medium containing 1% serum was added,
and supernatants generated over 48 h were saved for further
analysis. After the supernatant was removed, cells were washed twice
with PBS and lysed in Trizol for RNA extraction.
Animal treatment.
The 6-wk-old female C57BL/6 mice were obtained from Charles River
Laboratories (Montreal, PQ, Canada) and housed under specific pathogen-free conditions. Rodent laboratory food and water were provided ad libitum. The animals were treated in accordance with the
guidelines of the Canadian Council of Animal Care. All animal procedures were performed with inhalation anesthesia with isoflurane (MTC Pharmaceuticals, Cambridge, ON, Canada). Then 4 × 108 pfu of AdTGF223/225 were given together
with 4 × 108 pfu of AdDec, AdBig, or AdDL by
intranasal injection, suspended with PBS in a total volume of 25 µl.
Different control groups of animals received AdDec plus AdDL, AdBig
plus AdDL, or AdDL only (8 × 108 pfu total dose).
Mice were killed by abdominal aortic bleeding at days 3,
7, and 21 after injection of adenovectors.
Bronchoalveolar lavage.
After the chest cavity was opened, the lungs were removed and rinsed
with PBS. Bronchoalveolar lavage (BAL) was performed as described
earlier (17). A total of 0.5 ml of PBS was injected intratracheally and retrieved. The fluid was centrifuged at 1,500 rpm
for 10 min, and the supernatant was taken and frozen at 70°C for
determination of TGF-
1. BAL cells were counted with a hemocytometer, centrifuged in a cytospin, and stained for differential cytology (Hema3 solution, Biochemical Sciences, Swedesboro, NJ). A
total of 300 cells per sample were counted for differentials.
RNA extraction and mRNA analysis. Frozen lung samples were homogenized in Trizol with a tissue homogenizer. Chloroform was added, and the samples were centrifuged at 3,000 rpm for 30 min. The aqueous layer was aspirated, and RNA was precipitated with isopropanol. After centrifugation at 9,000 rpm for 10 min and washing the pellet with 75% ethanol, total RNA was dissolved in RNase-free water and the concentration was determined with a spectrophotometer.
Northern blot technique was used to detect mRNA specific for human decorin and human biglycan in treated cells and lungs. Then 15 µg of total RNA extracts from CCL-206 cells or total lung homogenate were separated on a 1% formaldehyde gel and transferred to a nylon membrane (ICN Pharmaceuticals, Montreal, Canada). Blots were hybridized with a 1.6-kb cDNA probe for human decorin or a 1.6-kb cDNA probe for human biglycan, both EcoRI fragments of the original cDNA used for the construction of the adenovectors. Blots were stringently washed and exposed to film for 1-3 days (Kodak XAR, Rochester, NY). Equal loading was confirmed by hybridization with 300-bp cDNA probe for GAPDH.Determination of TGF- levels in BAL fluid.
Total TGF-
1 in BAL fluid was determined after acid activation using
ELISA (R&D Systems). Level of active TGF-
1 was measured using the
assay without acid activation. The sensitivity of the assay is 7 pg/ml.
Histology and immunohistochemistry. After fixation in 10% buffered formalin for 24 h, a longitudinal section of the lung was paraffin embedded, sectioned, and stained with hematoxylin and eosin and Masson-Trichrome.
Immunohistochemistry was performed to stain cells and structures positive forHydroxyproline assay. Frozen lung samples were homogenized in 5 ml of deionized water. The homogenate (1 ml) was hydrolyzed in 2 ml of 6 N HCl for 16 h at 110°C. Hydroxyproline content was determined by a colorimetric assay described earlier (27). Briefly, the reaction was started by adding 1 ml of chloramine T solution to 400 µl of sample (diluted with 1.6 ml of water after the pH was adjusted to 7.0). The reaction was stopped with 1 ml of 70% perchloric acid, and 1 ml of dimethylbenzaldehyde solution was added. After an incubation period of 20 min at 60°C, optical density was determined within 30 min at a wavelength of 557 nm. The results were calculated as micrograms of hydroxyproline per milligram of wet lung weight with hydroxyproline standards (Sigma Chemicals).
Statistical analysis. Data are shown as means ± SE unless otherwise mentioned. For evaluation of group differences, we used the Student t-test assuming unequal variances. P < 0.05 was considered significant.
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RESULTS |
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Gene expression of human decorin and human biglycan in cell culture
and lungs.
Murine lung fibroblasts infected with AdDec and AdBig and
incubated overnight showed a strong positive signal for human
decorin and biglycan mRNAs by Northern gel analysis (Fig.
1). In cells infected with control virus
AdDL, no mRNA for human decorin or biglycan was detected. In animals
treated with AdDec or AdBig, mRNA signals for human decorin or biglycan
were found in total lung homogenates 3 days after infection (Fig. 1).
Again, no mRNA for human decorin or biglycan was detectable in lungs
treated with control virus AdDL.
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Bioactivity of TGF- in vitro and abrogation by
adenoviral-derived decorin and biglycan.
To demonstrate the inhibitory effect of transgene-derived
proteoglycans on TGF-
, we used an established bioassay for
active TGF-
using MLEC transfected with the luciferase gene under
control of a PAI-1 promoter. With infection of murine lung fibroblasts with AdTGF
223/225, we generated supernatants
containing bioactive TGF-
(Fig.
2A). This supernatant diluted
1:1 and 1:4 in medium showed the same bioactivity as 1.0 and 0.5 ng/ml
recombinant human TGF-
, respectively. Supernatants generated
by infecting cells with AdDec, AdBig, and AdDL resulted in luciferase
activities similar to supernatants without virus treatment
(negative control). When supernatants of
AdTGF
223/225-infected cells (1:4 diluted) were
combined with supernatants of AdDec- or AdBig-infected cells, a
significant and dose-dependent reduction of TGF-
-induced luciferase
activity was observed (Fig. 2B). However, the
concentration of transgene-derived proteoglycan in the supernatant was
not sufficient to completely block TGF-
activity. With anti-TGF-
antibodies, complete abrogation of TGF-
activity was achievable.
Supernatants of AdDL-infected or noninfected cells did not alter
TGF-
bioactivity in this assay.
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Effect of AdDec and AdBig on TGF- in BAL fluid of
AdTGF
223/225-injected mice.
In AdDL/AdTGF
223/225- and
AdBig/AdTGF
223/225-treated animals, total and active
TGF-
in BAL fluid (measured by ELISA) was significantly increased by
day 3 and further increased by day 7 compared
with AdDL/AdDL-treated mice (total 4,752 ± 1,211 and 3,571 ± 1,222 vs. 119 ± 32 pg/ml, active 480 ± 174 and 294 ± 106 vs. <10 pg/ml, P < 0.0001, Fig.
3).
AdDec/AdTGF
223/225-treated mice showed a similar
increase of TGF-
by day 3. However, by day 7,
these animals had significantly lower TGF-
concentration in BAL
fluid compared with AdDL/AdTGF
223/225-treated animals
(total 1,170 ± 565 pg/ml, active 61 ± 43 pg/ml, P < 0.05). AdBig/AdDL and AdDec/AdDL groups showed no
significant changes of TGF-
in BAL fluid. By day 21,
TGF-
was not found elevated in any group compared with control.
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Effect of TGF- overexpression on inflammation, tissue fibrosis,
and hydroxyproline content in the lung in the presence of AdDec and
AdBig.
Administration of AdTGF
223/225 induced a transient
inflammatory reaction in the lung present as early as 3 days after
injection and most pronounced after 7 days. Total cells in BAL were
significantly elevated in AdDL/AdTGF
223/225-,
AdDec/AdTGF
223/225-, and
AdBig/AdTGF
223/225-treated animals compared with
untreated control animals: approximately threefold at day
3 and fivefold at day 7 (Table
1). Injection of AdDL/AdDL resulted in a
twofold increase of total cells, likely an acute inflammatory reaction
against adenovectors. Throughout the experiment, no significant
differences were observed in total cell counts between
AdDec/AdTGF
223/225- and
AdBig/AdTGF
223/225-treated animals. Cell differentials
showed mainly alveolar macrophages and an increased percentage of
neutrophil granulocytes. No difference in cell differentials between
treatment groups was observed. AdDec/AdDL and AdBig/AdDL had no further
effect on BAL cells compared with AdDL/AdDL (data not shown).
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DISCUSSION |
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Fibrotic diseases are characterized by a disproportionate accumulation of ECM after tissue injury. The major component of the matrix is collagen, predominantly types I and III (20). Others are proteoglycans and glycoproteins. In the context of tissue injury and loss of organ-specific cells, ECM proteins are used to restructure the defect (21). Other, less-recognized functions of ECM molecules are to participate actively in intercellular communication (13, 22). This can happen either by direct chemotactic stimulation or by interference with cytokines and thus affect cell traffic and function.
Decorin, biglycan, fibromodulin, and lumican are small proteoglycans
with leucine-rich repeat structures (13, 22). Decorin carries a single glycosaminoglycan chain and is widely distributed in
mesenchymal tissues, associated and bound to collagen, which gains
stability through this interaction (3, 13). One of the key
features of decorin knockout mice is fragile skin, probably due to
irregularly shaped collagen (6). Biglycan has two
glycosaminoglycan chains and is localized closely around cells
(3, 13). The precise physiological role of biglycan is
still under discussion; various interactions with collagen and
glycoproteins have been suggested (23). The distribution
of fibromodulin and lumican is somewhat more restricted, with
fibromodulin present mainly in cartilage and tendons and lumican in the
cornea (12). However, recent reports about the composition
of pulmonary proteoglycan matrix showed significant quantities of
lumican in normal lung and fibromodulin in bleomycin-injured lung,
which brought these molecules more into the focus of pulmonary matrix
research (7, 26). It has been demonstrated that decorin,
biglycan, and fibromodulin are able to interact in vitro with TGF-,
which is a profibrotic key mediator in tissue fibrosis (12,
28). The affinity to TGF-
is similar for all, and it has been
suggested that these proteoglycans may be able to sequester an
overwhelming amount of TGF-
into the matrix and thus control its
biological effects (12, 22). For decorin, the in vitro
data have been confirmed in different disease models in animals, all of
them TGF-
mediated. Decorin was successful in reduction of
experimental pulmonary fibrosis induced with bleomycin given either
repeatedly as proteoglycan or once as gene using adenoviral gene
transfer (9, 17). Furthermore, decorin was employed to
reduce fibrotic kidney disease and neointimal proliferation in arteries
after balloon angioplasty (8, 14). Current data imply a
positive role of biglycan in the course of fibrosis and potential
therapeutic value; however, the application of biglycan in an animal
model of fibrotic disease has not been reported.
In this study, we used adenoviral transient gene transfer to generate
prolonged production and presence of active TGF-, decorin, and
biglycan in a model of pulmonary fibrosis. AdTGF
223/225
induces the synthesis of a mutated TGF-
molecule, which is
spontaneously active (25). Murine lung fibroblasts
infected with AdTGF
223/225 produce a high amount of
active TGF-
as shown in a bioassay using MLEC transfected with the
luciferase gene under control of the PAI-1 promoter. When injected
intratracheally into rat lungs, this adenovector leads to severe
interstitial fibrosis (25). AdDec and AdBig encode the
gene for human decorin and biglycan, respectively. We infected murine
lung fibroblasts with these vectors, which transcribed the foreign cDNA
into mRNA in vitro. Supernatants of cells infected with either AdDec
and AdBig, but not with control vector AdDL, were able to inhibit the
activity of AdTGF
223/225 supernatants in a
dose-dependent manner. The ability of both proteoglycans to interfere
with TGF-
was comparable, which is consistent with earlier reports.
To further investigate, if decorin and biglycan are able to interfere
with TGF- in vivo, we used a mouse model of pulmonary fibrosis.
Intranasal injection of AdTGF
223/225 resulted in
transient overexpression of TGF-
in the lung as measured by elevated
cytokine concentration in BAL fluid of treated mice after 3 and 7 days.
When AdDec was administered simultaneously with
AdTGF
223/225, the increase of TGF-
in BAL fluid was
reduced substantially compared with the combination
AdDL/AdTGF
223/225; although total and active TGF-
were still elevated above control, the concentration was four and eight
times lower than in AdDL/AdTGF
223/225. This observation
is in agreement with an earlier report, which has shown that decorin
mainly binds the active form of TGF-
(12). In contrast,
AdBig given together with AdTGF
223/225 did not affect
cytokine concentration in BAL fluid. After 21 days, considerable
interstitial fibrosis was present in mice treated with
AdDL/AdTGF
223/225. The fibrotic response was
significantly reduced when AdDec was administered together with
AdTGF
223/225, whereas AdBig had no positive
effect. Lung fibrosis was determined by histology and hydroxyproline
concentrations in lung homogenates and was preceded by accumulation of
myofibroblasts in the tissue 1 wk after administration. Although we did
not measure proteoglycans in BAL fluid or lung tissue, we attribute the
effects of AdDec and AdBig on TGF-
concentration in BAL fluid and on
fibrotic tissue responses to the presence of transgene product in the
lungs. We demonstrated strong mRNA signals in the tissue of treated
mice 3 days after infection. In previous experiments, we have shown that mRNA of decorin persists at least for 7 days in mouse lungs after
injection (17).
These data show that the previously documented inhibitory effect of
decorin on TGF- in vitro can be successfully transferred to an in
vivo model of disease in which tissue fibrosis and matrix accumulation
are induced by transient overexpression of active TGF-
. The data
also support earlier reports that biglycan is able to interfere with
TGF-
function in vitro (12). However, biglycan did not
affect TGF-
activity in the animal model. The failure of biglycan to
induce an antifibrotic tissue effect in this model could be explained
by differential binding of the TGF-
molecule through either lowered
affinity or altered site specificity. However, earlier reports and the
data presented here showing in vitro anti-TGF-
properties of both
proteoglycans suggest that biglycan and decorin have similar binding
properties to TGF-
(12). Therefore, we speculate that
the tissue localization of the two proteoglycans might account for
differences in their biological effect in vivo. Decorin is bound to
collagen and is probably able to bind TGF-
and prohibit its
interaction with cellular receptors, thus controlling its biological
effects on matrix-producing cells. Biglycan in contrast is more closely
associated to the pericellular space and cell surface (3,
22). In this localization, it could bind TGF-
but still allow
interaction with the receptor. This hypothesis is strengthened by the
observation that AdDec/AdTGF
223/225-treated animals had
a significantly lower concentration of TGF-
in BAL fluid compared
with AdBig/AdTGF
223/225-treated animals. It suggests
that the decorin-bound transgene TGF-
is fixed to collagen, whereas
biglycan-bound TGF-
remains in the pericellular space where it is
more easily liberated by BAL procedure and released into BAL fluid. In
previous experiments with rats, we observed that biglycan can induce
transient accumulation of myofibroblasts and ECM in the lung after
intratracheal injection without resulting in persistent tissue fibrosis
(24). It is possible that under certain circumstances
biglycan could even act as a profibrotic agent by binding TGF-
and
presenting it to the receptor similar to the related molecule
betaglycan. Betaglycan is the TGF-
receptor III, which can bind and
store TGF-
and eventually presents it to the signal transducing
receptors I and II (15).
In summary, the current study confirms the antifibrotic properties of
the proteoglycan decorin in an animal model of pulmonary fibrosis in
which fibrosis was induced by transient overexpression of active
TGF- using adenoviral gene transfer. The data demonstrate that
decorin binds and inhibits TGF-
in vitro and in vivo. Biglycan, another proteoglycan with the capability of binding TGF-
in vitro, failed to inhibit the fibrogenic effects of the cytokine in vivo. We
conclude that decorin but not biglycan has a potential role in the
future treatment of fibrotic disorders.
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ACKNOWLEDGEMENTS |
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We thank Duncan Chong, Xueya Feng, and Mary Jo Smith for outstanding technical help, and Tom Galt, Zhou Xing, and Michael Schmidt for helpful discussions and advice during the course of the experiments.
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FOOTNOTES |
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The work was supported by the Medical Research Council (MRC) of Canada. P. J. Sime is supported by James P. Wilmot Foundation and P. J. Margetts by MRC and Kidney Foundation of Canada.
Address for reprint requests and other correspondence: J. Gauldie, Dept. of Pathology and Molecular Medicine and Centre for Gene Therapeutics, McMaster Univ., 1200 Main St., West Hamilton, Ontario, Canada L8N 3Z5 (E-mail: gauldie{at}mcmaster.ca).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 27 November 2000; accepted in final form 11 January 2001.
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REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Abe, M,
Harpel JG,
Metz CN,
Nunes I,
Loskutoff DJ,
and
Rifkin DB.
An assay for transforming growth factor-beta using cells transfected with a plasminogen activator inhibitor-1 promoter-luciferase construct.
Anal Biochem
216:
276-284,
1994[ISI][Medline].
2.
Bett, AJ,
Haddara W,
Prevec L,
and
Graham FL.
An efficient and flexible system for construction of adenoviral vectors with insertions or deletions in early regions 1 and 3.
Proc Natl Acad Sci USA
91:
8802-8806,
1994[Abstract].
3.
Bianco, P,
Fisher LW,
Young MF,
Termine JD,
and
Robey PG.
Expression and localization of the two small proteoglycans biglycan and decorin in developing human skeletal and nonskeletal tissues.
J Histochem Cytochem
38:
1549-1563,
1990[Abstract].
4.
Border, WA,
Okuda S,
Languino LR,
Sporn MB,
and
Ruoslahti E.
Suppression of experimental glomerulonephritis by antiserum against transforming growth factor 1.
Nature
346:
371-374,
1990[ISI][Medline].
5.
Coker, RK,
and
Laurent GJ.
Pulmonary fibrosis: cytokines in the balance.
Eur Respir J
11:
1218-1221,
1998
6.
Danielson, KG,
Baribault H,
Holmes DF,
Graham H,
Kadler KE,
and
Iozzo RV.
Targeted disruption of decorin leads to abnormal collagen fibril morphology and skin fragility.
J Cell Biol
136:
729-743,
1997
7.
Dolhnikoff, M,
Morin J,
Roughley PJ,
and
Ludwig MS.
Expression of lumican in human lungs.
Am J Respir Cell Mol Biol
19:
582-587,
1998
8.
Fischer, JW,
Kinsella MG,
Clowes MM,
Lara S,
Clowes AW,
and
Wight TN.
Local expression of bovine decorin by cell-mediated gene transfer reduces neointimal formation after balloon injury in rats.
Circ Res
86:
676-683,
2000
9.
Giri, SN,
Hyde DM,
Braun RK,
Gaarde W,
Harper JR,
and
Pierschbacher MD.
Antifibrotic effect of decorin in a bleomycin hamster model of lung fibrosis.
Biochem Pharmacol
54:
1205-1216,
1997[ISI][Medline].
10.
Giri, SN,
Hyde DM,
and
Hollinger MA.
Effect of antibody to TGF on bleomycin-induced accumulation of lung collagen in mice.
Thorax
48:
959-966,
1993[Abstract].
11.
Grotendorst, GR,
Paglia L,
McIvor C,
Barsky S,
Martinet Y,
and
Pencev D.
Chemoattractants in fibrotic disorders.
Ciba Found Symp
114:
150-163,
1985[ISI][Medline].
12.
Hildebrand, AM,
Romaris M,
Rasmussen LM,
Heinegard D,
Twardzik DR,
Border WA,
and
Ruoslahti E.
Interaction of the small interstitial proteoglycans biglycan, decorin and fibromodulin with TGF.
Biochem J
302:
527-534,
1994[ISI][Medline].
13.
Iozzo, RV.
Matrix proteoglycans: from molecular design to cellular function.
Annu Rev Biochem
67:
609-652,
1998[ISI][Medline].
14.
Isaka, Y,
Brees DK,
Ikegaya K,
Kaneda Y,
Imai E,
Noble NA,
and
Border WA.
Gene therapy by skeletal muscle expression of decorin prevents fibrotic disease in rat kidney.
Nat Med
2:
418-423,
1996[ISI][Medline].
15.
Kaname, S,
and
Ruoslahti E.
Betaglycan has multiple binding sites for transforming growth factor-beta 1.
Biochem J
315:
815-820,
1996[ISI][Medline].
16.
Kelley, J.
TGF.
In: Cytokines of the Lung, edited by Kelley J.. New York: Dekker, 1993, p. 101-137.
17.
Kolb M, Margetts PJ, Galt T, Sime PJ, Xing Z, Schmidt M, and Gauldie
J. Transient transgene expression of decorin in the lung reduces
the fibrotic response to bleomycin. Am J Respir Crit Care
Med. In press.
18.
Lasky, JA,
and
Brody AR.
Interstitial fibrosis and growth factors.
Environ Health Perspect
108, Suppl 4:
751-762,
2000[ISI][Medline].
19.
Mason, RJ,
Schwarz MI,
Hunninghake GW,
and
Musson RA.
NHLBI Workshop Summary. Pharmacological therapy for idiopathic pulmonary fibrosis. Past, present, and future.
Am J Respir Crit Care Med
160:
1771-1777,
1999
20.
McAnulty, RJ,
and
Laurent GJ.
Collagen and its regulation in pulmonary fibrosis.
In: Pulmonary Fibrosis, edited by Phan SH,
and Thrall RS.. New York: Dekker, 1995, p. 135-171.
21.
McAnulty, RJ,
and
Laurent GJ.
Pathogenesis of lung fibrosis and potential new therapeutic strategies.
Exp Nephrol
3:
96-107,
1995[ISI][Medline].
22.
Roberts, CR,
Wight TN,
and
Hascall VC.
Proteoglycans.
In: The Lung: Scientific Foundations (2nd ed.), edited by Crystal RG,
West JB,
Barnes PJ,
and Weibel ER.. Philadelphia, PA: Lippincott-Raven, 1997, p. 757-767.
23.
Schonherr, E,
Witsch-Prehm P,
Harrach B,
Robenek H,
Rauterberg J,
and
Kresse H.
Interaction of biglycan with type I collagen.
J Biol Chem
270:
2776-2783,
1995
24.
Sime, PJ,
Sarnstrand B,
Xing Z,
Graham F,
Fisher L,
and
Gauldie J.
Adenovirus-mediated gene transfer of the proteoglycan biglycan induces fibroblastic responses in the lung.
Chest
111, Suppl:
137S,
1997
25.
Sime, PJ,
Xing Z,
Graham FL,
Csaky KG,
and
Gauldie J.
Adenovector mediated gene transfer of active TGF1 induces prolonged severe fibrosis in rat lung.
J Clin Invest
100:
768-776,
1997
26.
Venkatesan, N,
Ebihara T,
Roughley PJ,
and
Ludwig MS.
Alterations in large and small proteoglycans in bleomycin-induced pulmonary fibrosis in rats.
Am J Respir Crit Care Med
161:
2066-2073,
2000
27.
Woessner, JF, Jr.
The determination of hydroxyproline in tissue and protein samples containing small proportions of this imino acid.
Arch Biochem Biophys
93:
440-447,
1961[ISI].
28.
Xing, Z,
Jordana M,
Gauldie J,
and
Wang J.
Cytokines and pulmonary inflammatory and immune diseases.
Histol Histopathol
14:
185-201,
1999[ISI][Medline].
29.
Yamaguchi, Y,
Mann DM,
and
Ruoslahti E.
Negative regulation of TGF by the proteoglycan decorin.
Nature
346:
281-284,
1990[ISI][Medline].