Normal human colonic subepithelial myofibroblasts enhance
epithelial migration (restitution) via TGF-
3
Brian C.
McKaig1,
Shavcharn S.
Makh1,
Christopher J.
Hawkey1,
Daniel K.
Podolsky2, and
Yashwant R.
Mahida1
1 Division of Gastroenterology,
University Hospital, Nottingham NG7 2UH, United Kingdom; and
2 Gastrointestinal Unit,
Massachusetts General Hospital, Boston, Massachusetts 02114-2696
 |
ABSTRACT |
-After injury
and loss of epithelial cells, intestinal barrier function is
reestablished by migration of viable epithelial cells from the wound
edge (restitution). Myofibroblasts are located close to the basal
surface of epithelial cells. This study aimed to investigate the role
of human colonic subepithelial myofibroblasts in epithelial
restitution. Primary cultures of subepithelial myofibroblasts were
established. Monolayers of the epithelial cell lines IEC-6 and T84 were
"wounded" in a standard manner to create an in vitro model of
restitution. Migration of epithelial cells across the wound edge was
assessed following culture in myofibroblast-conditioned medium.
Myofibroblast expression of transforming growth factor (TGF)-
isoforms was examined using RT-PCR, and TGF-
isoform bioactivity was assessed using Mv 1 Lu bioassay.
Myofibroblast-conditioned medium, via a TGF-
-dependent pathway,
significantly enhanced migration of epithelial cells across the wound
edge and significantly inhibited cell proliferation in wounded
monolayers. Messenger RNA for TGF-
1, -
2, and -
3 was detected
in the myofibroblasts, and Mv 1 Lu bioassay showed the presence of
predominantly bioactive TGF-
3. This study shows that human colonic
subepithelial myofibroblasts secrete predominantly bioactive TGF-
3
and enhance restitution in wounded epithelial monolayers
via a TGF-
-dependent pathway.
transforming growth factor-
; wound repair
 |
INTRODUCTION |
THE GASTROINTESTINAL TRACT performs the critical
functions of extraction of nutrients, minerals, and electrolytes but
excludes luminal microorganisms and toxic molecules. These essential
functions are mediated by a monolayer of epithelial cells that lines
the intestinal lumen. The epithelial monolayer provides a barrier to
luminal toxic molecules and microorganisms via intercellular tight
junctions. This barrier is disrupted following injury and loss of
epithelial cells, directly exposing the lamina propria cells to luminal
contents. In the normal intestinal mucosa, epithelial continuity and
barrier function are rapidly reestablished following the loss of
injured epithelial cells. This occurs via a process designated
restitution, in which viable epithelial cells migrate from the wound
edge to reestablish epithelial continuity (15, 16, 21, 29, 31, 37, 43,
47). This process can be complete within minutes to hours, depending on
the extent of epithelial injury. Cell proliferation over the subsequent
24-48 h allows the replacement of the lost epithelial cells. There
is increasing appreciation for the fact that epithelial restitution in
vivo involves complex interaction between epithelial cells, the
underlying basement membrane, as well as cells within the lamina
propria matrix. The cells in the lamina propria may interact with the epithelial cells via pores in the basement membrane (26).
Intestinal subepithelial myofibroblasts are present immediately
subjacent to the basement membrane and close to the basal surface of
epithelial cells. Ultrastructural studies have shown that these cells
share characteristics of both fibroblasts (7, 14, 19) and smooth muscle
cells and have therefore been designated myofibroblasts (18, 20, 36,
38). Their location below the basement membrane suggests that these
cells may play an important role in the regulation of a number of
epithelial cell functions (45). We have recently established an ex vivo
model that allows, for the first time, pure populations of
subepithelial myofibroblasts from adult human intestinal mucosa to be
available (24). The isolated myofibroblasts retain a representative and
differentiated phenotype despite prolonged culture and passage (24).
Myofibroblasts have been shown to have an important role in wound
healing and fibrosis (10), but there have been no studies to date
investigating the role of human intestinal myofibroblasts in the
initial wound healing process of restitution.
In this study, we have investigated the role of isolated normal primary
adult human colonic subepithelial myofibroblasts in the regulation of
epithelial restitution. We demonstrate that the myofibroblasts enhance
epithelial cell migration via secreted bioactive transforming growth
factor (TGF)-
and that the predominant bioactive isoform is
TGF-
3.
 |
MATERIALS AND METHODS |
Cell culture.
The nontransformed rat small intestinal epithelial cell line IEC-6 was
obtained from the European Collection of Animal Cell Cultures (ECACC;
Porton Down, UK) and studied at passages
26-31. The cells were maintained in DMEM (GIBCO
BRL, Gaithersburg, MD) supplemented with 5% FCS (GIBCO), 2 mM
glutamine (Sigma Chemical, St. Louis, MO), 100 U/ml penicillin
(Britannia Pharmaceuticals, Surrey, UK), 0.1 mg/ml streptomycin (Evans
Medical, Surrey, UK), and insulin (final concentration 4 µg/ml; Sigma).
The human colon cancer cell line T84 was obtained from the ECACC and
studied at passages 70-75. The
cells were maintained in DMEM-Ham's F-12 medium (GIBCO) supplemented
with 10% FCS (GIBCO), 2 mM glutamine (Sigma), 100 U/ml penicillin
(Britannia Pharmaceuticals), and 0.1 mg/ml streptomycin (Evans Medical).
Human colonic subepithelial myofibroblasts were isolated from normal
colonic mucosal samples obtained from six colonic resection specimens.
The normal colonic mucosal samples were obtained >5 cm from the
tumor, and myofibroblasts were isolated as previously described (24).
In brief, the mucosal samples were completely denuded of epithelial
cells by three 30-min periods of incubation (at 37°C) in 1 mmol/l
EDTA (Sigma). The deepithelialized mucosal samples were subsequently
cultured (at 37°C in 5%
CO2) in RPMI 1640 (GIBCO)
containing 10% FCS. The cells in suspension were removed after every
24- to 72-h culture period, and the denuded tissue was maintained in
culture for up to 6 wk. Established colonies of myofibroblasts were
cultured in DMEM supplemented with 10% FCS and 1% nonessential amino
acids (GIBCO), penicillin (100 U/ml), and streptomycin (0.1 mg/ml). The
cells were passaged using 0.1% (wt/vol) trypsin-0.2% (vol/wt) EDTA in
a 1:2 to 1:3 split ratio. Studies were carried out at
passages 2-7 of myofibroblasts
isolated from six resection specimens.
For myofibroblast-conditioned medium, subconfluent and confluent
monolayers of cells were washed with DMEM and subsequently cultured in
0.1% FCS-DMEM for 24 h. The conditioned medium was stored at
70°C until use in studies on wounded epithelial monolayers and TGF-
bioassay (see below).
Wounding assays.
Wound assays were performed in multiples of six, using a previously
described method (27) with modification. Confluent monolayers of IEC-6
and T84 cells in six-well tissue culture plates (Nunc, GIBCO) were
wounded under microscopic vision using a razor blade and a Gilson
"p2" pipette tip. Cells were washed three times with fresh
serum-deprived medium (0.1% FCS-DMEM), and the wounded monolayers were
further cultured in fresh serum-deprived medium in the presence or
absence of recombinant human TGF-
1 (5 ng/ml; R&D Systems, Minneapolis, MN) or myofibroblast-conditioned medium.
Wound assays were also performed in the presence of polyclonal
neutralizing antibody to recombinant TGF (rTGF)-
1, -
2, and -
3
(1 µg/ml; R&D Systems).
As previously described, migration of IEC-6 cells was assessed in a
blinded fashion by the determination of the mean number of cells found
across the wound border in a standardized wound area (12), and
migration of T84 cells was assessed in a blinded fashion by
determination of the reduction in wound width over 24 h (34). Wound
areas were standardized by taking photographs at 100-fold magnification
using an Olympus CK-2 inverted microscope and an Olympus OM-1 camera.
Experiments were performed in quadruplicate, and data are presented as
means ± SD.
Cell proliferation in wounded epithelial monolayers.
"Wounded" monolayers of IEC-6 cells in six-well plates were
incubated as described in wounding assays. After culture for 20 h,
[3H]thymidine
(Amersham International, Buckinghamshire, UK) was added to each well (1 µCi/well). After a further incubation of 4 h, the cells were
subsequently fixed with methanol-acetic acid (vol/vol, 3:1) at room
temperature for 1 h, washed twice with 80% methanol, and lysed with 1 M NaOH. Uptake of
[3H]thymidine was
determined using an LKB (Wallac, Milton Keynes, UK) beta counter.
Proliferation was also assessed by determination of bromodeoxyuridine
(BrdU) uptake, as reflected in the mean number of BrdU-positive cells
observed per low-power field as previously described (4). Briefly, BrdU
(final concentration of 10 µmol/l; Sigma) was added to the wounded
monolayer. BrdU uptake was assessed after culture for 24 h in either
serum-deprived medium or myofibroblast-conditioned medium by addition
of mouse IgG anti-BrdU antibody (final concentration of 200 mU/ml;
Sigma) and detected using a universal Vectastain ABC detection kit
(Vector Laboratories, Peterborough, UK).
TGF-
bioassay.
The presence of bioactive TGF-
in myofibroblast-conditioned medium
was determined using a specific bioassay, which is based on the ability
of TGF-
to inhibit proliferation of the mink lung epithelial cell
line Mv 1 Lu (ECACC) (25). Latent TGF-
present in the
myofibroblast-conditioned medium was activated by the addition of
concentrated HCl to pH 2 and left to stand at room temperature for 60 min, followed by neutralization with NaOH and HEPES (to a final
concentration of 16 mmol/l).
Mv 1 Lu cells (in 0.2% FCS-DMEM) were seeded at 5 × 104 cells/well in 24-well cell
culture plates (Nunc, GIBCO). Four hours after cells were seeded,
acid-treated and untreated myofibroblast-conditioned medium and
standardized concentrations of rTGF-
1 were added and incubated for
20 h at 37°C (95% O2, 5%
CO2). After 20 h,
[3H]thymidine (1 µCi/well) was added, and the incubation was continued for an
additional 4 h. The cells were then fixed with methanol-acetic acid
(3:1), washed twice with 80% methanol, and lysed with 1 M NaOH. Uptake
of [3H]thymidine was
determined as before. From the standard curve obtained, the
concentration of total and biologically active TGF-
present in the
myofibroblast-conditioned medium could be calculated.
The contribution of each TGF-
isoform to total bioactivity was
assessed using Mv 1 Lu bioassay performed in the presence of TGF-
isoform-specific monoclonal antibodies (1 µg/ml; R&D Systems).
Conditioned medium from confluent and wounded IEC-6 and T84 monolayers
was also assessed for TGF-
bioactivity.
RNA isolation and reverse transcription.
RNA was isolated from myofibroblasts using RNAzolB (Biogenesis, Poole,
UK). Random hexamer primer (Pharmacia Biotech, Brussels, Belgium) was
mixed with 10 µg RNA (final volume of 37.5 µl) and heated to
70°C for 10 min and allowed to cool on ice. Reverse transcription
to cDNA was performed by adding the following and incubating at
37°C for 60 min: 5 µl of 10× PCR buffer [0.5 M Tris (pH 8.3), 0.75 M KCl, and 30 mM
MgCl2 (Stratagene, La Jolla,
CA)], 1.5 µl of 5 mM 2'-deoxyribonucleotide
5'-triphosphate mix (containing dATP, dCTP, dGTP, and dTTP each
at 25 mM; Ultrapure dNTP set, Pharmacia), 1 µl of Moloney murine
leukemia virus RT (200 U/µl; GIBCO), and 5 µl of 0.1 M DTT.
Subsequent enzyme deactivation was performed by heating to 90°C for
5 min, and the cDNA was stored at
20°C.
Polymerase chain reaction.
The following reaction mixture was added to 5 µl of the cDNA product:
5 µl of enzyme buffer [0.5 mM KCl, 0.1 M
Tris · HCl (pH 9.0), and 1% Triton X-100 (Promega,
Madison, WI)], 2 µl of 5 mM dNTPs, 0.5 µl of
Taq DNA polymerase (5 U/µl;
Promega), and sterile water to make a final solution of 50 µl. The
following primer pairs were used (to a final concentration of 5 µM)
based on published nucleotide sequences: 5'-CAG AAA TAC AGC AAC
AAT TCC TGG-3' (sense) and 5'-TTG CAG TGT GTT ATC CGT GCT
GTC-3' (antisense) to amplify 187-bp human TGF-
1 product (8);
5'-TCC AAA GAT TTA ACA TCT CCA ACC-3' (sense) and
5'-TCC CAC TGT TTT TTT TCC TAG TGG-3' (antisense) to
amplify 310-bp human TGF-
2 product (23); 5'-ACA TTT CTT TCT
TGC TGG-3' (sense) and 5'-GGG GAA GAA CCC ATA ATG-3'
(antisense) to amplify 685-bp human TGF-
3 product (9); and
5'-GGT GAA GGT CGG AGT CAA CGG A-3' (sense) and
5'-GAG GGA TCT CGC TCC TGG AAG A-3' (antisense) to amplify
240-bp human glyceraldehyde-6-phosphate dehydrogenase product.
Amplification was performed using a Trio-Thermoblock (Biometra). PCR
cycles consisted of denaturation for 45 s at 95°C, annealing at
54°C for 90 s, and extension at 72°C for 90 s. A
total of 30 cycles were used and completed by extension for 2 min at
54°C followed by 3 min at 72°C.
PCR products were analyzed by adding 1 µl of ethidium bromide (10 mg/ml) and 5 µl of gel-loading buffer (Sigma Chemical) to 15 µl PCR
product and electrophoresis on a 2% agarose gel. The specificity of
RT-PCR for TGF-
1, -
2, and -
3 was confirmed by hybridization of
the PCR product using specific digoxigenin-labeled internal
oligonucleotide probes and subsequent detection using alkaline
phosphatase-labeled rabbit anti-digoxigenin antibody (28) and by DNA
sequence analysis. The following digoxigenin-labeled probes were used:
5'-GTT GTG CGG CAG TGG-3' (to identify TGF-
1 product),
5'-GAG CAG AAG GCG AAT-3' (to identify TGF-
2 product), and 5'-GGG AGA AGG AAG GGC-3' (to identify TGF-
3 product).
Statistical analysis.
Results are expressed as means ± SD. Statistical analyses were
performed using one-way ANOVA and Student's
t-test.
 |
RESULTS |
Myofibroblast-conditioned medium enhances epithelial migration in
wounded IEC-6 and T84 monolayers.
The results illustrated in Table 1 and
Figs. 1 and
2 show that conditioned medium of
myofibroblast cultures derived from mucosal samples of six different
colons consistently enhanced repair of wounded IEC-6 and T84
monolayers. As expected (4, 12) and shown in Table 1, rTGF-
1 also
enhanced wound repair.

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Fig. 1.
Anti-transforming growth factor (TGF)- antibody (Ab) significantly
inhibits myofibroblast-conditioned medium (MFCM)-mediated enhancement
of epithelial restitution in wounded IEC-6
(A) and T84
(B) monolayers. "Wounded"
monolayers were cultured in control medium (0.1%), MFCM, or
recombinant TGF (rTGF)- in the presence or absence of neutralizing
antibody to TGF- . For both cell lines, in the presence of
anti-TGF- antibody, there was a significant
(* P < 0.01 for IEC-6 and
* P = 0.02 for T84 vs. MFCM
alone) reduction in wound repair. MFCM and rTGF- significantly
enhanced cell migration compared with control medium
(# P < 0.01 vs. control
medium).
|
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Fig. 2.
Acid-treated (acid Rx) and untreated (UnRx) MFCM enhance epithelial
cell migration in wounded IEC-6 monolayers to the same extent
(P < 0.01), confirming that in
untreated MFCM majority of TGF- is in the bioactive form.
|
|
Repair of wounded epithelial monolayers may occur as a consequence of
cell migration (restitution) or proliferation. To investigate the
mechanism by which myofibroblast-conditioned medium enhanced epithelial
wound repair, studies were performed using
[3H]thymidine
incorporation and BrdU uptake as indexes of cell proliferation. As
demonstrated in Table 2,
myofibroblast-conditioned medium consistently inhibited cell
proliferation in the wounded IEC-6 monolayers. This was also the case
with rTGF-
1 (data not shown). In wounded monolayers cultured in
control medium, scattered BrdU-labeled epithelial cells were seen in
the remaining monolayer, as previously described (5). In wounded
monolayers cultured in myofibroblast-conditioned medium or rTGF-
1,
there was a significant reduction in the number of BrdU-labeled cells
(data not shown).
Myofibroblasts enhance epithelial migration via secreted
TGF-
.
The similarity with rTGF-
1 in the effect on cell migration and
proliferation in wounded epithelial monolayers suggested that the
myofibroblast-conditioned medium contained bioactive TGF-
. This was
confirmed by inhibition of myofibroblast-conditioned medium-mediated
epithelial migration by polyclonal neutralizing antibody to TGF-
(Fig. 1A). The
myofibroblast-mediated enhanced restitution in wounded T84 cells was
also shown to be partially dependent on bioactive TGF-
(Fig.
1B). Addition of neutralizing antibody alone to wounded IEC-6 monolayers (cultured in control medium)
did not significantly inhibit cell migration (data not shown).
The presence of bioactive TGF-
in myofibroblast-conditioned medium
was confirmed using the Mv 1 Lu bioassay (Table
3). After acidification, there was only a
small increase in TGF-
bioactivity detected in the conditioned
medium, implying that the majority of TGF-
secreted by the
myofibroblasts is in the biologically active form. The predominance of
bioactive TGF-
in myofibroblast-conditioned medium was confirmed by
lack of a difference between acid-treated and untreated samples in the
enhancement of restitution in wounded IEC-6 monolayers (Fig. 2).
TGF-
bioactivity present in conditioned medium of IEC-6 and T84
epithelial monolayers was negligible in comparison to that present in
myofibroblast-conditioned medium. After acid treatment, the TGF-
concentration of IEC-6- and T84-conditioned medium was 60 and 6 pg/ml,
respectively (TGF-
concentration range in myofibroblast-conditioned
medium was 600-1,500 pg/ml). There was no significant difference
in total TGF-
bioactivity in conditioned medium collected from
confluent vs. subconfluent myofibroblast monolayers or from wounded vs.
confluent IEC-6 and T84 monolayers (data not shown).
The contribution of each TGF-
isoform to total bioactivity was
assessed by the use of monoclonal isoform-specific neutralizing antibodies. As shown in Fig. 3, TGF-
3
was the predominant bioactive isoform secreted by the myofibroblasts.
Neutralizing antibodies to TGF-
1 and -
2 reduced the overall
percentage of TGF-
bioactivity by 16.2 ± 6.7% and 3.2 ± 3.8%, respectively. In contrast, anti-TGF-
3 antibody reduced total
bioactivity by 81.0 ± 17.6%.

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Fig. 3.
Relative TGF- isoform bioactivity in MFCM. Neutralizing antibodies
(Ab) to TGF- 1 or - 2 did not significantly reduce overall TGF-
bioactivity. In contrast, neutralizing antibody to TGF- 3
significantly reduces bioactivity by 81.0 ± 17.6%
(n = 5, * P < 0.05).
|
|
Studies using RT-PCR demonstrated that mRNA transcripts for TGF-
1,
-
2, and -
3 are expressed by myofibroblasts (Fig.
4). These PCR products were confirmed to be
those of the individual isoforms of TGF-
by hybridization using
digoxigenin-labeled internal oligonucleotide probes and by DNA
sequencing as described in MATERIALS AND
METHODS.

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Fig. 4.
Expression of mRNA transcripts for glyceraldehyde-3-phosphate
dehydrogenase (GAPDH) and TGF- 1, - 2, and - 3
(lanes
1-4,
respectively) in human colonic myofibroblasts. RNA was isolated from a
confluent monolayer of cells and reverse transcribed, and relevant
transcripts were amplified by PCR using designed primers and
electrophoresed on a 2% agarose gel. DNA size markers are indicated on
left. TGF- PCR products were
isolated and gel purified. Specificity of the TGF- PCR products was
determined as described in MATERIALS AND
METHODS.
|
|
 |
DISCUSSION |
In the intestine, the surface monolayer of epithelial cells plays a
critical role in the maintenance of mucosal integrity, providing an
essential barrier to penetration by luminal microorganisms and their
products. Injury to the epithelial cells results in a loss of monolayer
continuity and barrier function and allows penetration by luminal
contents into the lamina propria. It is therefore of importance to the
host to repair any such breach in the barrier to avoid or limit an
inflammatory response. Epithelial continuity and barrier function is
normally rapidly restored by migration of viable cells adjacent to the
wound edge to cover the mucosal defect, a process termed restitution
(15, 16, 21, 29, 31, 37, 43, 47). This process is independent of
proliferation and begins within minutes of the initial injury. Proliferation of epithelial cells, required to make up for the lost
cells, occurs over 24-48 h after the injury (16). Previous studies
of epithelial cell restitution have shown that a large number of
peptide growth factors (interleukin-1
, interferon-
, epidermal
growth factor, TGF-
, hepatocyte growth factor, basic fibroblast
growth factor, intestinal trefoil factor) can influence this process
(11-13, 32). However, TGF-
plays a central role in mediating
the effects of many of these peptide growth factors (12).
It is recognized that the process of restitution in vivo involves
complex interactions between epithelial cells, cells within the lamina
propria, and components of the extracellular matrix. Previous studies
have shown that components of the extracellular matrix can regulate
epithelial restitution (2, 3, 17); however, there is little information
on the role of individual cell types within the lamina propria in the
regulation of epithelial restitution. In this study, we have examined
the role of primary adult human colonic subepithelial myofibroblasts in
epithelial cell migration. Cultures of subepithelial myofibroblasts
were established as recently described (24). During culture of mucosal samples denuded of epithelial cells, myofibroblasts migrate out of the
lamina propria via pores in the basement membrane. These cells
subsequently become adherent to the culture dish and proliferate to
form monolayers of myofibroblasts. We have previously shown that,
despite prolonged culture and passage, the myofibroblasts retain a
representative and differentiated phenotype (24). Myofibroblast cultures used in this study were established as previously reported, and their phenotypes were confirmed by demonstration of the expression of
-smooth muscle actin and vimentin (data not shown). The use of
the IEC-6 and T84 cell lines to study restitution in vitro is well
recognized. Extensive studies have shown the similarity between the
untransformed small intestinal rat epithelial IEC-6 cell line and the
normal rat crypt intestinal epithelial cell (35). These cell lines
allow the study of epithelial cell responses without the ambiguity of
contamination inherent to studies with preparations of primary
epithelial cells.
The studies in this report show that myofibroblasts secrete a factor or
factors that enhance cell migration in wounded epithelial monolayers while they paradoxically inhibit epithelial cell
proliferation, suggesting that this effect was mediated by TGF-
.
This was confirmed by the addition of neutralizing antibody to TGF-
,
which abrogated the enhanced epithelial cell migration seen in response
to the myofibroblast-conditioned medium. This response was more
pronounced in the nontransformed IEC-6 cell line compared with the
transformed T84 cell line. This could be explained by the relative
resistance of transformed cell lines to the effect of TGF-
, one of
the postulated mechanisms for the development of colonic carcinoma.
Previous reports have shown that cells can produce latent TGF-
as
well as plasminlike protease activity required to effect bioactivation
through liberation of the mature TGF-
dimer (39, 40, 44). We have
shown that myofibroblasts themselves express mRNA transcripts for all
three human TGF-
isoforms (TGF-
1, -
2, and -
3). The secreted
TGF-
is biologically active, suggesting that the myofibroblasts also
produce plasminlike protease activity. This finding differs from that
of TGF-
secretion by platelets (33) or other cell types (6, 22, 46).
In previous studies, TGF-
mRNA in the normal colon has been shown to
be expressed predominantly in the lamina propria closest to the surface
epithelium (1). From our current studies, we postulate that TGF-
derived from myofibroblasts may play an important role in epithelial
restitution in vivo. The myofibroblasts lie close to the epithelium,
and the secreted bioactive TGF-
could reach the epithelial cells
both by passive diffusion and via pores in the basement membrane (26). It is also possible that there are other myofibroblast-derived factors
that enhance epithelial restitution by inducing secretion of TGF-
by
the epithelial cells (4, 5). In addition, the subepithelial network of
myofibroblasts may facilitate repair of the injured intestinal
epithelium by contraction, thereby reducing the denuded surface area to
be reepithelialized (30).
It is recognized that myofibroblasts also participate in the formation
of the extracellular matrix and granulation tissue leading to fibrosis
during repair (10), processes that appear to be influenced to a large
extent by the TGF-
superfamily. This superfamily is one of the most
complex groups of cytokines, with widespread effects on many aspects of
growth, development, and wound healing. The three mammalian isoforms of
TGF-
(TGF-
1, -
2, and -
3) have been localized in healing
wounds, and there is evidence that manipulation of the ratios of the
TGF-
isoforms may influence the latter stages of the repair process,
namely scarring and fibrosis (41). Thus recent studies maintain that either neutralization of TGF-
1 and -
2 or exogenous addition of
TGF-
3 reduces scarring in cutaneous rat wounds (42). In our study,
we have shown for the first time that primary cultures of normal human
colonic subepithelial myofibroblasts release predominantly bioactive
TGF-
3. Such predominant expression of bioactive TGF-
3 in vivo
would imply a capacity for wound healing without fibrosis. Myofibroblasts may therefore play a pivotal role in regulating several
stages of wound repair from restitution to end-stage fibrosis.
 |
ACKNOWLEDGEMENTS |
This work was supported by the Digestive Disorders Foundation (UK) and
The National Association for Colitis and Crohn's Disease (UK).
 |
FOOTNOTES |
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. §1734 solely to indicate this fact.
Address for reprint requests and other correspondence: Y. R. Mahida,
Division of Gastroenterology, Univ. Hospital, Queen's Medical Centre,
Nottingham NG7 2UH, UK.
Received 25 September 1998; accepted in final form 4 February
1999.
 |
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