Divisions of 1 Rheumatology and Immunology and 3 Nephrology, 2 Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, South Carolina 29425; and 4 Department of Cell Biology and Anatomy, University of Miami School of Medicine, Miami, Florida 33136
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ABSTRACT |
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Transforming growth factor-
(TGF-
) and connective tissue growth factor (CTGF) are ubiquitously
expressed in various forms of tissue fibrosis, including fibrotic
diseases of the kidney. To clarify the common and divergent roles
of these growth factors in the cells responsible for pathological
extracellular matrix (ECM) deposition in renal fibrosis, the effects of
TGF-
and CTGF on ECM expression in primary human mesangial (HMCs)
and human proximal tubule epithelial cells (HTECs) were studied. Both
TGF-
and CTGF significantly induced collagen protein expression with similar potency in HMCs. Additionally,
2(I)-collagen
promoter activity and mRNA levels were similarly induced by TGF-
and
CTGF in HMCs. However, only TGF-
stimulated collagenous protein
synthesis in HTECs. HTEC expression of tenascin-C (TN-C) was increased
by TGF-
and CTGF, although TGF-
was the more potent inducer. Thus both growth factors elicit similar profibrogenic effects on ECM production in HMCs, while promoting divergent effects in HTECs. CTGF
induction of TN-C, a marker of epithelial-mesenchymal
transdifferentiation (EMT), with no significant induction of
collagenous protein synthesis in HTECs, may suggest a more predominant
role for CTGF in EMT rather than induction of excessive collagen
deposition by HTECs during renal fibrosis.
mesangial cells; tubule epithelial cells; collagen; tenascin; epithelial-mesenchymal transdifferentiation; connective tissue growth
factor; transforming growth factor-
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INTRODUCTION |
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PROGRESSIVE RENAL DISEASE can be caused by multiple pathogenetic mechanisms, leading to end-stage renal failure. The progressive renal diseases predominantly originate in the glomerulus, causing glomerular injury and subsequent sclerosis characterized by prominent intraglomerular accumulation of extracellular matrix (ECM) components. Among the resident cells of the glomerulus, mesangial cells are primarily responsible for excessive ECM deposition (55). In recent years, tubulointerstitial damage has been recognized as an equally important component of progressive renal disease (57). It is now believed that glomerular injury triggers tubular cell activation, leading to tubulointerstitial inflammation and fibrosis. Activated tubular cells produce a number of cytokines, chemokines, and profibrogenic growth factors, which act in an autocrine and paracrine fashion (60). Increasing evidence also suggests that tubular epithelial cells have the capacity to undergo epithelial-mesenchymal transdifferentiation (EMT), thereby becoming interstitial fibroblasts (43), a major cell type responsible for interstitial fibrosis (1).
Transforming growth factor- (TGF-
) is implicated as a key
mediator in the development of kidney fibrosis. Elevated expression of
TGF-
isoforms has been demonstrated in the glomeruli and
tubulointerstitium of patients with renal diseases characterized by
excessive ECM accumulation (63). Elevated TGF-
has also
been observed in various animal models of kidney fibrosis (11,
21, 27, 46, 53). Moreover, blocking TGF-
via neutralizing
antibodies resulted in reduction of glomerulosclerosis in experimental
models (6, 54, 69). More recent studies have shown that
connective tissue growth factor (CTGF) expression is strongly
upregulated in diabetic nephropathy and other progressive renal
diseases (33). A strong correlation between the number of
CTGF-positive cells and degree of injury at sites of chronic
tubulointerstitial damage was also observed (33). However,
in contrast to TGF-
, the significance of CTGF in development of
renal fibrosis is not fully defined.
The relationship between TGF- and CTGF in the stimulation of ECM
synthesis has been characterized recently. TGF-
-induced collagen
synthesis is blocked with specific anti-CTGF antibodies or antisense
CTGF oligonucleotides in NRK-fibroblasts in vitro and in wound healing
in vivo (15). Based on these observations, it has been
proposed that CTGF is a downstream mediator of the fibrogenic effect of
TGF-
. In support of these studies, NRK-49F cells treated with CTGF
antisense oligonucleotides showed significant inhibition of fibronectin
and
1(I)-collagen mRNA upon TGF-
stimulation (66). However, additional studies using rat mesangial
cells demonstrated that neutralizing antibodies to CTGF inhibit
TGF-
-mediated fibronectin synthesis but not TGF-
induction of
-smooth muscle actin (4), indicating differential
effects of TGF-
and CTGF on the mesangial cell response to injury in
vitro. Furthermore, recent studies performed in human dermal
fibroblasts suggest an additional divergent role for CTGF and the
closely related CCN family member Cyr61 as matrix remodeling factors
(7, 8). Thus CTGF may be involved in matrix deposition or
matrix degradation, depending on either the cell type or the specific
experimental conditions.
Given the fact that CTGF is prominently expressed in both glomerular
and tubular compartments in progressive human renal diseases, the goal
of this study was to systematically compare the profibrogenic effects
of CTGF vis a vis TGF-. Because of the possible cell-type-specific effects of CTGF, we used human renal cells for our studies, including human mesangial cells (HMC) and human proximal tubular epithelial cells (HTEC).
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MATERIALS AND METHODS |
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Cell Culture
Human kidney tissues were obtained from surgical nephrectomy specimens and donor transplant kidneys, which were determined unsuitable for transplant in accordance with Institutional Review Board guidelines. HMCs were isolated from glomeruli by collagenase treatment and differential sieving of minced cortical areas of the renal tissue (34). Cells were confirmed to be mesangial by morphological criteria and by the presence of staining toRecombinant CTGF/CTGF Antibody
Human recombinant CTGF (rCTGF) was generated using a baculovirus system to infect Drosophila cells (19). rCTGF was purified using a heparin-Sepharose affinity column, and rCTGF fractions were analyzed via Western blot analysis for the absence of contamination by other growth factors. Goat anti-human CTGF antibody was generated as previously described by Duncan et al. (15).Adenoviral Constructs
Adenoviral vectors expressing CTGF and green fluorescent protein (GFP) were generated using the method described by He et al. (26). Briefly, the cDNA encoding full-length human CTGF (provided by Gary Grotendorst) was cloned in the shuttle vector pAdTRACK-CMV, which contains a GFP expression cassette driven by a separate CMV promoter. The shuttle vector containing CTGF was cotransformed into Escherichia coli BJ5183 cells with the AdEasy-1 adenoviral backbone plasmid, which lacks the E1 and E3 regions of the adenoviral genome. Linearized recombinant plasmid DNA was then transfected into 293 cells, an adenoviral packaging cell line, using the FuGENE 6 transfection reagent (Roche Applied Science, Indianapolis, IN) to generate the recombinant adenovirus expressing CTGF and GFP from separate CMV promoters (AdCTGF). An adenovirus expressing GFP alone (AdGFP) was generated via the same method for use as a control vector.[3H]Proline Incorporation Assay
Cells were plated in 12-well plates and grown to visual confluence, followed by incubation in SFM containing 0.1% BSA supplemented with ascorbic acid (50 µg/ml) andRNA Isolation and Northern Blot Analysis
Confluent cultures of HMCs and HTECs were serum starved for 24 h, followed by stimulation with TGF-Measurement of Spliced and Unspliced COL1A2 by RT-PCR
HMCs were grown to confluence in 10-cm2 dishes and serum starved for 24 h, followed by incubation with TGF-Plasmid Constructs, Transient Transfection, and Luciferase Assay
The COL1A2-luciferase construct contains sequences fromWestern Blot Analysis
TGFRI/TGF
RII.
Confluent cultures of HMCs were serum starved for 24 h and then
stimulated with growth factors [TGF-
, rCTGF, platelet-derived growth factor (PDGF)-AB] at the indicated concentrations for an additional 24 h. Cells were collected and lysed in 50 mM
Tris · Cl (pH 8.0), 150 mM NaCl, 0.02% sodium azide, 0.1%
SDS, 1% Nonidet P-40, 0.5% sodium deoxycholate, and 1 mM
phenylmethylsulfonyl fluoride followed by bicinchoninic acid protein
assay (Pierce, Rockford, IL) to determine total protein concentration.
Equal amounts (50 µg) of total proteins per sample were separated via 8% SDS-PAGE (T
RII) or 10% SDS-PAGE (T
RI) and transferred to nitrocellulose membranes for Western blotting. Polyclonal rabbit antibodies against T
RI (R-20; Santa Cruz Biotechnology, Santa Cruz,
CA) and T
RII (Medical University of South Carolina Antibody Facility) were incubated at dilutions of 1:500 in 3%
milk/Tris-buffered saline (TBS) overnight at 4°C. After washes in
Tween-TBS (TTBS), membranes were probed with anti-rabbit-horseradish
peroxidase (HRP) secondary antibodies (Amersham Pharmacia, Piscataway,
NJ) in 3% milk/TBS at a dilution of 1:3,000 for 1.5 h at room
temperature. After washes in TTBS, proteins were visualized using
enhanced chemiluminescence (ECL) reagents (Amersham). Protein levels
were quantitated using NIH Image densitometry software.
Tenascin-C.
Confluent cultures of HMC or HTEC in 12-well plates were serum starved
for 24 h, followed by stimulation with TGF- or rCTGF (2.5, 5, and 10 ng/ml) for 72 h. To inhibit rCTGF stimulation, cells were
pretreated with neutralizing CTGF antibody (50 µg/ml) or normal goat
IgG as a control for 30 min, followed by addition of rCTGF (5 ng/ml)
for 72 h. Aliquots of conditioned media normalized for cell number
were analyzed for tenascin protein levels via 6% SDS-PAGE followed by
transfer to a nitrocellulose membrane. The membrane was probed with a
mouse monoclonal antibody against human tenascin-C (TN-C; 1:1,000,
kindly provided by Dr. Wolfgang Rettig, Boehringer Ingelheim Pharma)
used in a previously published study by our laboratory
(36). After washes in TTBS, the blot was incubated with
anti-mouse-HRP secondary antibody (1:3,000). After final washes in
TTBS, tenascin protein levels were visualized with ECL and quantitated
as above.
Statistical Analysis
Student's t-test analysis using GraphPad InStat Statistics Software (version 1.12) was performed to determine statistical significance. Values of ![]() |
RESULTS |
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TGF- and CTGF are Inducers of ECM Proteins in HMC
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Because induction of collagen type I is specifically associated with
renal fibrosis, we next investigated the effects of TGF- and CTGF on
COL1A2 promoter activity using a COL1A2 promoter/reporter construct as
described in MATERIALS AND METHODS. Recent studies have
determined that TGF-
stimulates COL1A2 gene transcription via the
Smad3 and Sp1 response elements located in the proximal region of the
COL1A2 promoter (9, 47, 68). Furthermore, it has been
reported that CTGF stimulates the COL1A2 promoter in human fibroblasts
(56). We observed that both TGF-
and CTGF significantly
stimulate the COL1A2 promoter activity in HMCs with similar potency,
suggesting that the CTGF response element is also located in the
proximal promoter region (Fig.
2A). The specific cis elements and cognate transcription factors mediating
CTGF stimulation of the COL1A2 promoter remain to be characterized.
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To further confirm the effects of CTGF on COL1A2 transcription, we used
a previously described method based on the PCR (51). RT-PCR using primers designed for spliced and unspliced (newly transcribed) collagen type I RNA was performed to analyze the effect of
CTGF on COL1A2 mRNA expression. HMCs were stimulated with CTGF for
24 h or left unstimulated as a control. Figure 2B, left, demonstrates that CTGF induced COL1A2 steady-state
mRNA levels (spliced) with a 1.33-fold increase in COL1A2 spliced mRNA when normalized for 18S RNA levels (compare lanes 1 and
3). Furthermore, CTGF increased COL1A2 heterogeneous nuclear
RNA (unspliced RNA) levels (Fig. 2B, right,
compare lanes 2 and 4) consistent with CTGF
stimulation of COL1A2 promoter activity (Fig. 2A), which collectively suggests induction of collagen gene transcription by CTGF.
Additionally, endogenously expressed CTGF in HMCs transduced with
AdCTGF also induced COL1A2 transcription compared with HMCs transduced
with AdGFP as a control (Fig. 2B, right). The
average degree of increase in unspliced/newly transcribed COL1A2 by
rCTGF and endogenously expressed CTGF (AdCTGF) was 1.72 when normalized for 18S RNA levels. Taken together, these observations (Figs. 1 and 2)
suggest that TGF- and CTGF have similar effects on collagen expression in HMCs.
TGF- Receptor Expression Is Not Modulated by Either TGF-
or
CTGF in HMCs
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TGF- and CTGF Have Divergent Effects on ECM Production by HTECs
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To determine if the differential effects of TGF- and CTGF in HTECs
were specific for collagen type IV or whether other ECM proteins were
also differentially regulated by these growth factors, we analyzed TN-C
expression. In healthy kidney, TN-C is diffusely expressed in the
medulla, but it is ubiquitously overexpressed in areas of
tubulointerstitial damage, regardless of diagnosis (59).
Therefore, TN-C expression may serve as an early marker of fibrosis.
The effects of TGF-
and CTGF on TN-C steady-state mRNA and protein
expression in HTECs were determined by Northern blot and Western blot
analysis, respectively. TGF-
significantly induced TN-C protein
levels (Fig. 5, A and
B), which correlated with
induction of TN-C mRNA levels (Fig. 5D). CTGF also
significantly stimulated production of TN-C protein, although with
lesser potency than TGF-
(Fig. 5, A and B).
The increase in TN-C protein was abolished in HTECs treated with
neutralizing CTGF antibody before addition of rCTGF, demonstrating the
specificity of the stimulatory effect of CTGF (Fig. 5C). A
more modest stimulation of TN-C mRNA levels was observed after addition
of various doses of CTGF compared with TGF-
(Fig. 5D).
Collectively, these data suggest that under the experimental conditions
used in our study, TGF-
and CTGF have opposing effects on collagen
IV synthesis in HTECs but similar effects on TN-C synthesis in HTECs.
We also analyzed the effects of TGF-
and CTGF on TN-C production in
HMCs. Cultured HMCs synthesized TN-C, but TN-C levels were not
modulated by either TGF-
or CTGF (data not shown).
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DISCUSSION |
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The profibrogenic role of TGF- in various fibrotic diseases,
including renal fibrosis, is well established. More recently, CTGF has
also been proposed as a key cytokine responsible for elevated
deposition of ECM proteins in vivo. Both TGF-
and CTGF are
universally overexpressed in various diseases characterized by tissue
fibrosis (5, 22, 31, 42). However, although the role of
TGF-
as a potent inducer of ECM synthesis in a variety of cells,
including renal cells, is well documented, little is known regarding
the mechanisms by which CTGF affects ECM deposition. Original studies
by Duncan et al. (15) using embryonic rat kidney cells
(NRK fibroblasts) have demonstrated that CTGF is a mediator of the
fibrogenic effects of TGF-
, including proliferation and ECM
production. Subsequent studies have shown induction of collagen type I
and fibronectin by CTGF in rat mesangial cells (48, 49). Additionally, CTGF has been shown to induce expression of fibronectin in mouse tubule epithelial cells (61). On the other hand,
Chen et al. (7) have demonstrated that adhesion of human
fibroblasts to CTGF resulted in the formation of focal adhesion
complexes and activation of FAK, paxillin, and Rac kinases, which
correlated with prolonged mitogen-activated protein kinase activation
and induction of metalloproteinases MMP-1 and MMP-3 (7).
The latter study suggests a role for CTGF as a matrix-remodeling
factor. Finally, there is increasing evidence that CTGF and a closely related factor, Cyr61, have proangiogenic functions (2,
3). These studies underscore the cell-type-specific effects of
this pleiotropic growth factor. Our study was undertaken to
characterize the possible profibrogenic functions of CTGF compared with
TGF-
in human renal cells.
HMCs in culture constitutively produce ECM proteins, mainly collagen
types I, III, and IV (47). ECM synthesis can be further upregulated by low concentrations (2.5-5 ng/ml) of TGF-. Under our experimental conditions, similar upregulation of ECM was also observed with low doses of CTGF (2.5-5 ng/ml). In addition, the pattern of stimulated ECM proteins was indistinguishable between TGF-
and CTGF, suggesting that both factors induce a similar profibrogenic program in HMCs. In previous studies using rat mesangial cells, stimulation of collagen type I and fibronectin was also observed
after addition of either TGF-
or CTGF (49). However, in
contrast to our study, stimulation of matrix proteins was achieved with
much higher doses of CTGF (20 ng/ml). It is possible that HMCs are more
responsive than rat cells to CTGF. However, we cannot exclude the
possibility that the difference in CTGF potency was the result of the
different sources of CTGF, which is not commercially available. To
further compare the effects of TGF-
and CTGF on ECM synthesis, we
focused on collagen type I, which is highly upregulated in fibrotic
conditions. Furthermore, TGF-
regulation of collagen type I is
relatively well defined. In agreement with previous studies, CTGF and
TGF-
stimulated COL1A2 mRNA levels in HMCs (40, 47). In
addition, our studies demonstrate that similar to TGF-
, CTGF
expressed either exogenously or endogenously (via adenoviral vector)
upregulates the collagen type I gene at the level of transcription. It
is well documented that TGF-
upregulation of the COL1A2 promoter in
HMCs and fibroblasts occurs via Smad3 and Sp1. Although TGF-
and
CTGF response elements map to the proximal promoter region, it is
unlikely that Smad3 is directly involved in the CTGF response. However,
TGF-
stimulation of the CTGF promoter is Smad3 dependent
(28). In the future, we may characterize the specific
transcription factors that mediate CTGF stimulation of the collagen
type I gene. Together, these studies demonstrate that TGF-
and CTGF
can similarly stimulate ECM production by HMCs in vitro, suggesting
that both growth factors may contribute to glomerulosclerosis.
Expression of TGF- receptors is elevated in human renal diseases
(64) and in various animal models of renal fibrosis
(27, 29, 53, 58). However, relatively little is known
about the factors that upregulate TGF-
receptor expression in
general and in renal cells in particular. Some of the factors that have
been previously implicated in modulating TGF-
receptor levels in
renal cells include ANG II (62), high glucose
(32), and leptin (23). On the other hand,
TGF-
has been shown to downregulate its receptor expression in rat
kidney fibroblasts (20). In this study, there was no
effect of either CTGF or TGF-
on type I and type II TGF-
receptor
protein and mRNA levels in HMCs, suggesting that these profibrotic
growth factors do not enhance ECM production via upregulation of the
classic transmembrane TGF-
signaling receptors. In contrast to our
recent studies using dermal fibroblasts, which demonstrated selective
upregulation of TGF-
receptor type II by PDGF-AB (13), PDGF-AB selectively reduced TGF-
type II receptor levels in normal adult HMCs. The relevance of this observation is presently unknown. However, differential regulation of TGF-
receptor subunits has been
noted in other experimental models. For example, TGF-
type II
receptor is exclusively induced by glucose treatment in mouse mesangial
cells (32), whereas both receptor mRNA and protein levels
were elevated by either high glucose or mechanical stretch in rat
mesangial cells (50). We have also observed discordant expression of TGF-
receptor subunits in vivo in the remnant model of
chronic renal failure (Gore-Hyer E and Trojanowska M, unpublished observations). In addition, altered ratios of TGF-
receptors have been reported in atherosclerotic lesions (37) and
liver cirrhosis (52). These observations may suggest that
alterations in the TGF-
signaling pathway via modulation of receptor
levels are common features of various fibrotic diseases. Further
studies are necessary to understand the implications of altered TGF-
receptor ratios in the context of fibrosis.
Tubulointerstitial injury correlates with the decline in renal function
and progression of renal disease (57). Furthermore, elevated levels of TGF- and CTGF correlate with tubulointerstitial fibrosis (21, 33, 61). Therefore, we analyzed whether
HTECs may directly contribute to tubulointerstitial fibrosis by
responding to profibrogenic growth factors such as TGF-
and CTGF.
Our data demonstrate that ECM protein synthesis by HTECs is
significantly elevated in response to TGF-
. Elevated collagen
synthesis in response to TGF-
was previously observed in rat tubular
cells (12). In contrast to HMCs, HTECs do not express
increased collagenous proteins in response to CTGF. In fact, the lowest
doses of CTGF used in our study decreased the levels of secreted
collagenous proteins. This decrease may be either due to inhibition of
ECM synthesis or due to stimulation of MMPs, as previously shown in dermal fibroblasts (7). These data also suggest that
unlike NRK cells, in which CTGF was shown to mediate the profibrogenic effects of TGF-
, the profibrogenic effects of TGF-
may be CTGF independent in HTECs. Further studies are needed to resolve these issues. Although our studies suggest that CTGF is not a potent stimulator of collagenous protein synthesis in HTECs, it has been demonstrated that CTGF can induce fibronectin expression in mouse proximal tubule epithelial cells (61). Thus CTGF may act
as a fibrogenic stimulus in HTECs to induce fibronectin accumulation in
renal fibrosis.
There is increasing evidence that EMT is one of the mechanisms
contributing to the pathogenesis of tubulointerstitial fibrosis (41). EMT, the process whereby epithelial cells transform
into mesenchymal cells, occurs during development and in pathological processes such as tumorigenesis (24). Recent studies
suggest that the increased numbers of interstitial fibroblasts, which are the primary cell type responsible for ECM deposition in
tubulointerstitial fibrosis (1), may be derived in part by
the transdifferentiation of tubular epithelial cells into
fibroblasts/myofibroblasts (39, 41, 44). During renal
fibrosis, EMT is thought to occur when tubular epithelial cells acquire
mesenchymal/fibroblast characteristics, including expression of
vimentin (38), -smooth muscle actin (17),
and fibroblast specific protein-1 (36), which allow the
cells to migrate through their basement membrane into the interstitium
where the cells are then identified as fibroblasts/myofibroblasts. It
has been demonstrated that HTECs undergoing EMT acquire a migratory phenotype in vitro, which likely facilitates their translocation from
the tubular basement membrane to the renal interstitium in vivo
(65, 67). Furthermore, alterations in the basement
membrane composition, including downregulation of collagen type IV, are important for epithelial cell migration during EMT (67).
Interestingly, this study demonstrates the novel finding that CTGF
induces TN-C in HTECs, albeit with lesser potency than TGF-
. The
matricellular protein TN-C is significantly elevated in
tubulointerstitial lesions of various kidney diseases (45)
and is expressed during development at the sites of EMT
(16). TN-C possesses both adhesive and anti-adhesive properties, which may promote cell migration in tumor metastasis, embryonic development, and wound healing (35). Thus it is
possible that TN-C is one of the factors facilitating EMT in vitro and in vivo during renal fibrosis. Our data demonstrating the ability of
CTGF to induce TN-C and also diminish collagen IV protein synthesis in
HTECs may indicate a predominant role for CTGF in the induction of EMT
during the progression of tubulointerstitial fibrosis.
Several studies have demonstrated a pivotal role for TGF- in EMT in
various physiological processes and in kidney fibrosis (17, 43,
67). The divergent effects of TGF-
and CTGF on collagen IV
expression in HTECs suggest that these growth factors may direct
different HTEC functions in the progression of EMT and
tubulointerstitial fibrosis. Although TGF-
significantly induced
collagenous proteins in HTECs, CTGF diminished collagen protein
expression, which may elicit the changes in basement membrane composition necessary to facilitate EMT. Thus, in addition to TGF-
,
CTGF may be an important and distinct mediator of HTEC transdifferentiation in interstitial fibroblasts. In support of the
potential role of CTGF in EMT in vivo, Frazier et al. (18) observed CTGF, TGF-
, and PDGF expression in close proximity to epithelial cells in transition to the myofibroblast phenotype in the
remnant model of renal fibrosis. It is tempting to speculate that CTGF
may contribute to EMT during renal fibrosis by stimulating the
migratory phenotype of HTECs via alterations in ECM composition (decreasing collagen type IV expression) and via its induction of TN-C,
which may further promote cell migration. Further studies are required
to test the specific mechanisms of CTGF in EMT.
In conclusion, our study demonstrates that the profibrogenic potential
of CTGF is equal to that of TGF- in HMCs, with both growth factors
exhibiting similar stimulation of collagen gene expression at the
promoter, mRNA, and protein levels. To date, various profibrotic
effects of CTGF have been demonstrated predominantly using rat or mouse
cell lines. However, given the cell type- and/or cell line-dependent
properties of in vitro cultures, it is necessary to fully characterize
the effects of CTGF in primary human cells. These findings are also
important to clarify the profibrotic actions of CTGF in human renal
cells in view of the emerging pleiotropic nature of CTGF as a
matrix-remodeling factor capable of inducing matrix degradation in
human dermal fibroblasts (7). Furthermore, this study
reveals distinct effects of TGF-
and CTGF on ECM expression by
HTECs. Both growth factors induce expression of TN-C, a marker of EMT,
which is overexpressed in renal tubulointerstitial disease (45,
59). However, opposite to TGF-
, CTGF did not significantly induce collagen protein synthesis but slightly decreased collagen type
IV protein levels in HTECs, which may indicate a distinct role for CTGF
in EMT. TGF-
has recently been shown to have both CTGF-dependent and
-independent effects in various cell types and experimental systems.
Therefore, it will be of great interest to determine the overlapping
and distinct roles of TGF-
and CTGF in human tubular epithelial
cells undergoing EMT, a potential mechanism contributing to renal
tubulointerstitial fibrosis.
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ACKNOWLEDGEMENTS |
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Current address for E. L. Greene: Dept. of Internal Medicine, Nephrology Div., Mayo Clinic, Rochester, MN 55905.
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FOOTNOTES |
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Address for reprint requests and other correspondence: M. Trojanowska, Div. of Rheumatology and Immunology, Medical Univ. of South Carolina, 96 Jonathan Lucas St., Suite 912, P.O. Box 250623, Charleston, SC 29425 (E-mail: trojanme{at}musc.edu).
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.
May 14, 2002;10.1152/ajprenal.00007.2002
Received 7 January 2002; accepted in final form 6 May 2002.
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