Tumor necrosis factor-alpha (TNF-(alpha)) is an important mediator during the inflammatory phase of wound healing. Excessive amounts of pro-inflammatory cytokines such as TNF-(alpha) are associated with inflammatory diseases including chronic wounds. Matrix metalloproteinases (MMPs) are involved in matrix re-modeling during wound healing, angiogenesis and tumor metastasis. As with pro-inflammatory cytokines, high levels of MMPs have been found in inflammatory states such as chronic wounds. In this report we relate these two phenomena. TNF-(alpha) stimulates secretion of active MMP-2, a type IV collagenase, in organ-cultured full-thickness human skin. This suggests a mechanism whereby excess inflammation affects normal wound healing. To investigate this observation at the cellular and molecular levels, we examined TNF-(alpha) mediated activation of pro-MMP-2, induction of MT1-MMP, and the intracellular signaling pathways that regulate the proteinase in isolated human dermal fibroblasts. We found that TNF-(alpha) substantially promoted activation of pro-MMP-2 in dermal fibroblasts embedded in type-I collagen. In marked contrast, collagen or TNF-(alpha) individually had little influence on the fibroblast-mediated pro-MMP-2 activation. One well-characterized mechanism for pro-MMP-2 activation is through a membrane type matrix metalloproteinase, such as MT1-MMP. We report that TNF-(alpha) significantly induced MT1-MMP at the mRNA and protein levels when the dermal fibroblasts were grown in collagen. Although the intracellular signaling pathway regulating mt1-mmp gene expression is still obscure, both TNF-(alpha) and collagen activate the NF-(kappa)B pathway. In this report we provide three sets of evidence to support a hypothesis that activation of NF-(kappa)B is essential to induce MT1-MMP expression in fibroblasts after TNF-(alpha) exposure. First, SN50, a peptide inhibitor for NF-(kappa)B nuclear translocation, simultaneously blocked the TNF-(alpha) and collagen mediated MT1-MMP induction and pro-MMP-2 activation. Secondly, TNF-(alpha) induced I(kappa)B to breakdown in fibroblasts within the collagen lattice, a critical step leading to NF-(kappa)B activation. Lastly, a consensus binding site for p65 NF-(kappa)B (TGGAGCTTCC) was found in the 5′-flanking region of human mt1-mmp gene. Based on these results and previous reports, we propose a model to explain TNF-(alpha) activation of MMP-2 in human skin. Activation of NF(kappa)B signaling in fibroblasts embedded in collagen induces mt1-mmp gene expression, which subsequently activates the pro-MMP-2. The findings provide a specific mechanism whereby TNF-(alpha) may affect matrix remodeling during wound healing and other physiological and pathological processes.

REFERENCES

REFERENCES
Arumugam
S.
,
Jang
Y. C.
,
Chen-Jensen
C.
,
Gibran
N. S.
,
Isik
F. F.
(
1999
).
Temporal activity of plasminogen activators and matrix metalloproteinases during cutaneous wound repair.
Surgery
125
,
587
593
Birkedal-Hansen
H.
(
1995
).
Proteolytic remodeling of extracellular matrix.
Curr. Opin. Cell Biol
7
,
728
735
Birkedal-Hansen
H.
,
Moore
W. G.
,
Bodden
M. K.
,
Windsor
L. J.
,
Birkedal-Hansen
B.
,
DeCarlo
A.
,
Engler
J. A.
(
1993
).
Matrix metalloproteinases: a review.
Crit. Rev. Oral Biol. Med
4
,
197
250
Bullen
E. C.
,
Longaker
M. T.
,
Updike
D. L.
,
Benton
R.
,
Ladin
D.
,
Hou
Z.
,
Howard
E. W.
(
1995
).
Tissue inhibitor of metalloproteinases-1 is decreased and activated gelatinases are increased in chronic wounds.
J. Invest. Dermatol
104
,
236
240
Cooney
R.
,
Iocono
J.
,
Maish
G.
,
Smith
J. S.
,
Ehrlich
P.
(
1997
).
Tumor necrosis factor mediates impaired wound healing in chronic abdominal sepsis.
J. Trauma
42
,
415
420
Ehrlich
H. P.
,
Rajaratnam
J. B.
(
1990
).
Cell locomotion forces versus cell contraction forces for collagen lattice contraction: an in vitro model of wound contraction.
Tissue Cell
22
,
407
417
Fajardo
L. F.
,
Kwan
H. H.
,
Kowalski
J.
,
Prionas
S. D.
,
Allison
A. C.
(
1992
).
Dual role of tumor necrosis factor-alpha in angiogenesis.
Am. J. Pathol
140
,
539
544
Forsyth
P. A.
,
Wong
H.
,
Laing
T. D.
,
Rewcastle
N. B.
,
Morris
D. G.
,
Muzik
H.
,
Leco
K. J.
,
Johnston
R. N.
,
Brasher
P. M.
,
Sutherland
G.
,
Edwards
D. R.
(
1999
).
Gelatinase-A (MMP-2), gelatinase-B (MMP-9) and membrane type matrix metalloproteinase-1 (MT1-MMP) are involved in different aspects of the pathophysiology of malignant gliomas.
Br. J. Cancer
79
,
1828
1835
Fortunato
S. J.
,
Menon
R.
,
Lombardi
S. J.
(
1998
).
The effect of transforming growth factor and interleukin-10 on interleukin-8 release by human amniochorion may regulate histologic chorioamnionitis.
Am. J. Obstet. Gynecol
179
,
794
799
Garner
W. L.
,
Karmiol
S.
,
Rodriguez
J. L.
,
Smith
D. J.
Jr.
,
Phan
S. H.
(
1993
).
Phenotypic differences in cytokine responsiveness of hypertrophic scar versus normal dermal fibroblasts.
J. Invest. Dermatol
101
,
875
879
Ghosh
S.
,
May
M. J.
,
Kopp
E. B.
(
1998
).
NF-kappa B and Rel proteins: evolutionarily conserved mediators of immune responses.
Annu. Rev. Immunol
16
,
225
260
Haas
T. L.
,
Davis
S. J.
,
Madri
J. A.
(
1998
).
Three-dimensional type I collagen lattices induce coordinate expression of matrix metalloproteinases MT1-MMP and MMP-2 in microvascular endothelial cells.
J. Biol. Chem
273
,
3604
3610
Haas
T. L.
,
Stitelman
D.
,
Davis
S. J.
,
Apte
S. S.
,
Madri
J. A.
(
1999
).
Egr-1 mediates extracellular matrix-driven transcription of membrane type 1 matrix metalloproteinase in endothelium.
J. Biol. Chem
274
,
22679
22685
Kahari
V. M.
,
Saarialho-Kere
U.
(
1999
).
Matrix metalloproteinases and their inhibitors in tumour growth and invasion.
Ann. Med
31
,
34
45
Kitzis
V.
,
Engrav
L. H.
,
Quinn
L. S.
(
1999
).
Transient exposure to tumor necrosis factor-alpha inhibits collagen accumulation by cultured hypertrophic scar fibroblasts.
J. Surg. Res
87
,
134
141
Leibovich
S. J.
,
Polverini
P. J.
,
Shepard
H. M.
,
Wiseman
D. M.
,
Shively
V.
,
Nuseir
N.
(
1987
).
Macrophage-induced angiogenesis is mediated by tumour necrosis factor-alpha.
Nature
329
,
630
632
Lin
Y. Z.
,
Yao
S. Y.
,
Veach
R. A.
,
Torgerson
T. R.
,
Hawiger
J.
(
1995
).
Inhibition of nuclear translocation of transcription factor NF-kappa B by a synthetic peptide containing a cell membrane-permeable motif and nuclear localization sequence.
J. Biol. Chem
270
,
14255
14258
Lohi
J.
,
Lehti
K.
,
Valtanen
H.
,
Parks
W. C.
,
Keski-Oja
J.
(
2000
).
Structural analysis and promoter characterization of the human membrane-type matrix metalloproteinase-1 (MT1-MMP) gene.
Gene
242
,
75
86
Madtes
D. K.
,
Raines
E. W.
,
Sakariassen
K. S.
,
Assoian
R. K.
,
Sporn
M. B.
,
Bell
G. I.
,
Ross
R.
(
1988
).
Induction of transforming growth factor-alpha in activated human alveolar macrophages.
Cell
53
,
285
293
Martin
P.
(
1997
).
Wound healing—aiming for perfect skin regeneration.
Science
276
,
75
81
Mercurio
F.
,
Manning
A. M.
(
1999
).
Multiple signals converging on NF-kappaB.
Curr. Opin. Cell Biol
11
,
226
232
Migita
K.
,
Eguchi
K.
,
Kawabe
Y.
,
Ichinose
Y.
,
Tsukada
T.
,
Aoyagi
T.
,
Nakamura
H.
,
Nagataki
S.
(
1996
).
TNF-alpha-mediated expression of membrane-type matrix metalloproteinase in rheumatoid synovial fibroblasts.
Immunology
89
,
553
557
Mohan
R.
,
Rinehart
W. B.
,
Bargagna-Mohan
P.
,
Fini
M. E.
(
1998
).
Gelatinase B/lacZ transgenic mice, a model for mapping gelatinase B expression during developmental and injury-related tissue remodeling.
J. Biol. Chem
273
,
25903
25914
Moses
M. A.
,
Marikovsky
M.
,
Harper
J. W.
,
Vogt
P.
,
Eriksson
E.
,
Klagsbrun
M.
,
Langer
R.
(
1996
).
Temporal study of the activity of matrix metalloproteinases and their endogenous inhibitors during wound healing.
J. Cell Biochem
60
,
379
386
Murphy
G.
,
Stanton
H.
,
Cowell
S.
,
Butler
G.
,
Knauper
V.
,
Atkinson
S.
,
Gavrilovic
J.
(
1999
).
Mechanisms for pro matrix metalloproteinase activation.
APMIS
107
,
38
44
Reno
C.
,
Boykiw
R.
,
Martinez
M. L.
,
Hart
D. A.
(
1998
).
Temporal alterations in mRNA levels for proteinases and inhibitors and their potential regulators in the healing medial collateral ligament.
Biochem. Biophys. Res. Commun
252
,
757
763
Salomon
G. D.
,
Kasid
A.
,
Cromack
D. T.
,
Director
E.
,
Talbot
T. L.
,
Sank
A.
,
Norton
J. A.
(
1991
).
The local effects of cachectin/tumor necrosis factor on wound healing.
Ann. Surg
214
,
175
180
Sato
H.
,
Takino
T.
,
Okada
Y.
,
Cao
J.
,
Shinagawa
A.
,
Yamamoto
E.
,
Seiki
M.
(
1994
).
A matrix metalloproteinase expressed on the surface of invasive tumour cells [see comments].
Nature
370
,
61
65
Sunderkotter
C.
,
Steinbrink
K.
,
Goebeler
M.
,
Bhardwaj
R.
,
Sorg
C.
(
1994
).
Macrophages and angiogenesis.
J. Leuko. Biol
55
,
410
422
Thompson
E. W.
,
Yu
M.
,
Bueno
J.
,
Jin
L.
,
Maiti
S. N.
,
Palao-Marco
F. L.
,
Pulyaeva
H.
,
Tamborlane
J. W.
,
Tirgari
R.
,
Wapnir
I.
, et al. 
. (
1994
).
Collagen induced MMP-2 activation in human breast cancer.
Breast Cancer Res. Treat
31
,
357
370
Tuan
T. L.
,
Keller
L. C.
,
Sun
D.
,
Nimni
M. E.
,
Cheung
D.
(
1994
).
Dermal fibroblasts activate keratinocyte outgrowth on collagen gels.
J. Cell Sci
107
,
2285
2289
Werb
Z.
,
Chin
J. R.
(
1998
).
Extracellular matrix remodeling during morphogenesis.
Ann. NY Acad. Sci
857
,
110
118
Wysocki
A. B.
,
Staiano-Coico
L.
,
Grinnell
F.
(
1993
).
Wound fluid from chronic leg ulcers contains elevated levels of metalloproteinases MMP-2 and MMP-9.
J. Invest. Dermatol
101
,
64
68
Xu
J.
,
Zutter
M. M.
,
Santoro
S. A.
,
Clark
R. A.
(
1998
).
A three-dimensional collagen lattice activates NF-kappaB in human fibroblasts: role in integrin alpha2 gene expression and tissue remodeling.
J. Cell Biol
140
,
709
719
Yammamoto
H.
,
Itoh
F.
,
Adachi
Y.
,
Fukushima
H.
,
Itoh
H.
,
Sasaki
S.
,
Hinoda
Y.
,
Imai
K.
(
1999
).
Messenger RNA expression of matrix metalloproteinase and tissue inhibitors of metalloproteinase in human hepatocellular carcinoma.
Jpn J. Clin. Oncol
29
,
58
62
Young
P. K.
,
Grinnell
F.
(
1994
).
Metalloproteinase activation cascade after burn injury: a longitudinal analysis of the human wound environment.
J. Invest. Dermatol
103
,
660
664
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