The (beta)1-null fibroblastic cell line GD25 and its derivatives were studied to gain an understanding of the roles of (beta)1 and (beta)3 integrins in the initial (1-hour) contraction of collagen gels. Stable transfectants of GD25 cells expressing the (beta)1A splice variant of (beta)1 ((beta)1A-GD25) did not express (alpha)2(beta)1A and did not adhere to collagen. After transfection of (alpha)2 into (beta)1A-GD25 cells, the (alpha)2(beta)1A-GD25 transfectants contracted collagen gels in the presence of serum, whereas (beta)1A-GD25 cells did not. The GD25 parental cells, however, also contracted collagen gels. Collagen gel contraction by GD25 cells was blocked by antibodies to (alpha)v(beta)3 or a RGD-containing peptide, indicating that (alpha)v(beta)3 is the integrin responsible for mediation of contraction by GD25 cells. Collagen gel contraction by (alpha)2(beta)1A-GD25 cells was not inhibited by antibodies to (alpha)v(beta)3 or RGD-containing peptide, but was inhibited by anti-(alpha)2 antibody. Flow cytometry demonstrated negligible expression of (alpha)v(beta)3 by (beta)1A-GD25 and (alpha)2(beta)1A-GD25 cells when compared to GD25 cells. Platelet derived growth factor (PDGF) and sphingosine-1-phosphate (S1P) enabled gel contraction by (alpha)2(beta)1A-GD25 and GD25 cells, respectively, in the absence of serum. PDGF-stimulated contraction by (alpha)2(beta)1A-GD25 cells was attenuated in the presence of inhibitors of phosphatidylinositol-3-kinase whereas such inhibitors had no effect on S1P-stimulated contraction by GD25 cells. These experiments using the (beta)1-null GD25 cells and (beta)1A and (alpha)2(beta)1A transfectants demonstrate that (alpha)2(beta)1A and (alpha)v(beta)3 independently mediate collagen gel contraction and are regulated by different serum factors and signaling pathways.

REFERENCES

Bell
E.
,
Ivarsson
B.
,
Merrill
C.
(
1979
).
Production of a tissue-like structure by contraction of collagen lattices by human fibroblasts of different proliferative potential in vitro.
Proc. Nat. Acad. Sci. USA
76
,
1274
1278
Brown
R. A.
,
Talas
G.
,
Porter
R. A.
,
McGrouther
D. A.
,
Eastwood
M.
(
1996
).
Balanced mechanical forces and microtubule contribution to fibroblast contraction.
J. Cell. Physiol
169
,
439
447
Carver
W.
,
Molano
I.
,
Reaves
T. A.
,
Borg
T. K.
,
Terracio
L.
(
1995
).
Role of the1 1integrin complex in collagen gel contraction in vitro by fibroblasts.
J. Cell. Physiol
165
,
425
437
Chen
Y.
,
O'Toole
T. E.
,
Leong
L.
,
Liu
B.
,
Diaz-Gonzalez
F.
,
Ginsberg
M. H.
(
1995
).
Beta 3 integrin-mediated fibrin clot retraction by nucleated cells: differing behavior ofIIb 3andv 3.
Blood
86
,
2606
2615
Chiquet-Ehrismann
R.
,
Tannheimer
M.
,
Koch
M.
,
Brunner
A.
,
Spring
J.
,
Martin
D.
,
Baumgartner
S.
,
Chiquet
M.
(
1994
).
Tenascin-C expression by fibroblasts is elevated in stressed collagen gels.
J. Cell Biol
127
,
2093
2101
Clark
R. A.
,
Folkvord
J. M.
,
Hart
C. E.
,
Murray
M. J.
,
McPherson
J. M.
(
1989
).
Platelet isoforms of platelet-derived growth factor stimulate fibroblasts to contract collagen matrices.
J. Clin. Invest
84
,
1036
1040
Clark
R. A.
(
1993
).
Regulation of fibroplasia in cutaneous wound repair.
Am. J. Med. Sci
306
,
42
48
Clark
R. A.
,
Nielsen
L. D.
,
Welch
M. P.
,
McPherson
J. M.
(
1995
).
Collagen matrices attenuate the collagen-synthetic response of cultured fibroblasts to TGF-beta.
J. Cell Sci
108
,
1251
1261
Davies
P. F.
(
1995
).
Flow-mediated endothelial mechanotransduction.
Physiol. Rev.
75
,
519
560
Davis
G. E.
(
1992
).
Affinity of integrins for damaged extracellular matrix:v 3 binds to denatured collagen type Ic through RGD sites.
Biochem. Biophys. Res. Commun
182
,
1025
1031
Eckes
B.
,
Msauch
C.
,
Huppe
G.
,
Krieg
T.
(
1993
).
Downregulation of collagen synthesis in fibroblasts within three-dimensional collagen lattices involves transcription mechanisms.
FEBS Lett
318
,
129
133
Edelman
J.
,
Chan
M.
,
Uniyal
B. M. C.
,
Onodera
S.
,
Wang
H.
,
Damjanovich
D. Z.
,
Latzer
L.
,
Finberg
D. B.
,
Bergelson
J. M.
(
1994
).
The mouse VLA-2 homologue supports collagen and laminin adhesion but not virus binding.
Cell Adhes. Commun
2
,
131
143
Ehrlich
H. P.
,
Wyler
D. J.
(
1983
).
Fibroblast contraction of collagen lattices in vitro: Inhibition by chronic inflammatory cell mediators.
J. Cell. Physiol
116
,
345
351
Eicholtz
T.
,
Jalink
K.
,
Fahrenfort
I.
,
Moolenaar
W. H.
(
1993
).
The bioactive phospholipid lysophosphatidic acid is released from activated platelets.
Biochem. J
291
,
677
680
Elsdale
T.
,
Bard
J.
(
1972
).
Collagen substrata for studies on cell behavior.
J. Cell Biol
54
,
626
637
Fässler
R.
,
Pfaff
M.
,
Murphy
J.
,
Noegel
A. A.
,
Johansson
S.
,
Timpl
R.
,
Albrecht
R.
(
1995
).
Lack of1 integrin gene in embryonic stem cells affects morphology, adhesion, and migration but not integration into the inner cell mass of blastocysts.
J. Cell Biol
128
,
979
988
Greiling
D.
,
Clark
R. A.
(
1997
).
Fibronectin provides a conduit for fibroblast transmigration from collagenous stroma into fibrin clot provisional matrix.
J. Cell Sci
110
,
861
870
Grinnell
F.
(
1994
).
Fibroblasts, myofibroblasts, and wound contraction.
J. Cell Biol
124
,
401
404
Grinnell
F.
,
Ho
C. H.
,
Lin
Y. C.
,
Skuta
G.
(
1999
).
Differences in the regulation of fibroblast contraction of floating versus stress collagen matrices.
J. Biol. Chem
274
,
918
923
Guidry
C.
,
Grinnell
F.
(
1985
).
Studies on the mechanism of hydrated collagen gel reorganization by human skin fibroblasts.
J. Cell Sci
79
,
67
81
Gullberg
D.
,
Tingsrom
A.
,
Thuresson
A.
,
Olsson
L.
,
Terracio
L.
,
Borg
T. K.
,
Rubin
K.
(
1990
).
Beta 1 integrin-mediated collagen gel contraction is stimulated by PDGF.
Exp. Cell. Res
186
,
264
272
Halliday
N. L.
,
Tomasek
J. J.
(
1995
).
Mechanical properties of the extracellular matrix influence fibronectin fibril assembly invitro.
Exp. Cell Res
217
,
109
117
Ingber
D.
(
1991
).
Integrins as mechanochemical transducers.
Curr. Opin. Cell Biol
3
,
841
848
Katagiri
Y.
,
Hiroyama
T.
,
Akamatsu
N.
,
Suzuki
H.
,
Yamazaki
H.
,
Tanoue
K.
(
1995
).
Involvement ofv 3integrin in mediating fibrin gel retraction.
J. Biol. Chem
270
,
1785
1790
Kitamura
M.
,
Maruyama
N.
,
Mitarai
T.
,
Nagasawa
R.
,
Yoshida
H.
,
Sakai
O.
(
1992
).
Extracellular matrix contraction by cultured mesangial cells: modulation by transforming growth factor-and matrix components.
Exp. Mol. Path
56
,
132
143
Klein
C. E.
,
Dressel
D.
,
Steinmayer
T.
,
Mauch
C.
,
Eckes
B.
,
Krieg
T.
,
Bankert
R. B.
,
Weber
L.
(
1991
).
Integrin2 1 is upregulated in fibroblasts and highly aggressive melanoma cells in three-dimensional collagen lattices and mediates the reorganization of collagen I fibrils.
J. Cell Biol
115
,
1427
1436
Kolodney
M. S.
,
Wysolmerski
R. B.
(
1992
).
Isometric contraction by fibroblasts and endothelial cells in tissue culture: a quantitative study.
J. Cell Biol
11
,
73
82
Koyoma
H.
,
Raines
E. W.
,
Bornfeldt
K. E.
,
Roberts
J. M.
,
Ross
R.
(
1996
).
Fibrillar collagen inhibits arterial smooth muscle proliferation through regulation of cdk2 inhibition.
Cell
87
,
1069
1078
Lambert
C. A.
,
Soudant
E. P.
,
Nusgens
B. V.
,
Lapiere
C. M.
(
1998
).
Pretranslational regulation of extracellular matrix macromolecules and collagenase expression in fibroblasts by mechanical forces.
Lab. Invest
66
,
444
451
Langholz
O.
,
Rockel
D.
,
Mauch
C.
,
Kozlowska
E.
,
Bank
I.
,
Krieg
T.
,
Eckes
B.
(
1995
).
Collagen and collagenase gene expression in three-dimensional collagen lattices are differentially regulated by1 1and2 1integrins.
J. Cell Biol
131
,
1903
1915
Lee
M.-J.
,
Van Brocklyn
J. R.
,
Thangada
S.
,
Liu
C. H.
,
Hand
A. R.
,
Menzeleev
R.
,
Spiegel
S.
,
Hla
T.
(
1998
).
Sphingosine-1-phosphate as a ligand for the G-protein-coupled receptor EDG-1.
Science
279
,
1552
1555
Lee
Y. R.
,
Oshita
Y.
,
Tsuboi
R.
,
Ogawa
H.
(
1996
).
Combination of insulin-like growth factor (IGF)-1 and IGF-binding protein-1 promotes fibroblast-embedded collagen gel contraction.
Endocrinology
137
,
5278
5283
Maniotis
A. J.
,
Chen
C. S.
,
Ingber
D. E.
(
1997
).
Demonstration of mechanical connections between integrins, cytoskeletal filaments, and nucleoplasm that stabilize nuclear structure.
Proc. Nat. Acad. Sci. USA
94
,
849
854
Mochitate
K.
,
Pawelek
P.
,
Grinnell
F.
(
1991
).
Stress relaxation of contracted collagen gels: disruption of actin filament bundles, release of cell surface fibronectin, and down-regulation of DNA and protein synthesis.
Exp. Cell Res
193
,
198
207
Moulin
V.
,
Castilloux
G.
,
Auger
F. A.
,
Garrel
D.
,
O'Conner-McCourt
M. D.
,
Germain
L.
(
1998
).
Modulated response to cytokines of human wound healing myofibroblasts compared to dermal fibroblasts.
Exp. Cell Res
238
,
283
293
Moroi
M.
,
Jung
S. M.
(
1997
).
Platelet receptors for collagen.
Thromb. & Haemostas
78
,
439
444
Nakagawa
S.
,
Pawelek
P.
,
Grinnell
F.
(
1989
).
Long-term culture of fibroblasts in contracted collagen gels: effects on cell growth and biosynthetic activity.
J. Invest. Dermatol
93
,
792
798
Pierschbacher
M. D.
,
Ruoslahti
E.
(
1984
).
The cell attachment activity of fibronectin can be duplicated by small fragments of the molecule.
Nature
309
,
30
33
Pilcher
B. K.
,
Levine
N. S.
,
Tomasek
J. J.
(
1995
).
Thrombin promotion of isometric contraction in fibroblasts: its extracellular mechanism of action.
Plast. Reconstr. Surg
96
,
1188
1195
Rayan
G. M.
,
Parizi
M.
,
Tomasek
J. J.
(
1995
).
Pharmacologic regulation of Dupuytren's fibroblast contraction in vitro.
J. Hand Surg
21
,
1065
1070
Riikonen
T.
,
Koivisot
L.
,
Viihinen
P.
,
Heino
J.
(
1995
).
Transforming growth factor-regulates collagen gel contraction by increasing 21integrin expression in osteogenic cells.
J. Biol. Chem
270
,
376
382
Rosenfeldt
H.
,
Lee
D. J.
,
Grinnell
F.
(
1998
).
Increased c-fos mRNA expression by human fibroblasts contracting stressed collagen matrices.
Mol. Cell. Biol
18
,
2659
2667
Sakai
T.
,
Peyruchaud
O.
,
Fässler
R.
,
Mosher
D. F.
(
1998
).
Restoration of1A integrins is required for lysophosphatidic acid-induced migration of 1-null mouse fibroblastic cells.
J. Biol. Chem
273
,
19378
19382
Schneller
M.
,
Vuori
K.
,
Ruoslahti
E.
(
1997
).
v 3integrin associates with activated insulin and PDGPreceptors and potentiates the biological activity of PDGF.
EMBO J
18
,
5600
5607
Shiro
J. A.
,
Chan
B. M. C.
,
Roswit
W. T.
,
Kassner
P. D.
,
Pentland
A. P.
,
Hemler
M. D.
,
Eisen
A. Z.
,
Kupper
T. S.
(
1991
).
Integrins2 1(VLA-2) mediates reorganization and contraction of collagen matrices by human cells.
Cell
67
,
403
410
Shyy
J.
,
Chien
S.
(
1997
).
Role of integrins in cellular responses to mechanical stress and adhesion.
Curr. Opin. Cell Biol
9
,
707
713
Stopak
D.
,
Harris
A. K.
(
1982
).
Connective tissue morphogenesis by fibroblast traction.
Dev. Biol
90
,
383
398
Thoreson
W. B.
,
Khandalavala
B. N.
,
Manahan
R. G.
,
Polyak
I. A.
,
Liu
J. L.
,
Chacko
D. M.
(
1997
).
Lysophosphatidic acid stimulates proliferation of human retinal pigment epithelial cells.
Curr. Eye Res
16
,
698
702
Tomasek
J. J.
,
Hay
E. D.
,
Fujiwara
K.
(
1982
).
Collagen modulates cell shape and cytoskeleton of embryonic corneal and fibroma fibroblasts: distribution of actin,-actin, and myosin.
Dev. Biol
92
,
107
122
Tomasek
J. J.
,
Haaksma
C. J.
,
Eddy
R. J.
,
Vaughan
M. B.
(
1992
).
Fibroblast contraction occurs on release of tension in attached collagen lattices: Dependency on an organized actin cytoskeleton and serum.
Anat. Record
232
,
359
368
Vernon
R. B.
,
Sage
E. H.
(
1996
).
Contraction of fibrillar type 1 collagen by endothelial cells: A study in vitro.
J. Cell. Biochem
60
,
185
197
Wakita
H.
,
Matsushita
K.
,
Nishimura
K.
,
Tokura
Y.
,
Furukawa
F.
,
Takigawa
M.
(
1998
).
Sphingosylphosphorylcholine stimulates proliferation and upregulates cell surface-associated plasminogen activator activity in cultured human keratinocytes.
J. Invest. Dermatol
110
,
253
258
Wennerberg
K.
,
Lohikangas
L.
,
Gullberg
D.
,
Pfaff
M.
,
Johansson
S.
,
Fässler
R.
(
1996
).
1 Integrin-dependent and-independent polymerization of fibronectin.
J. Cell Biol
132
,
227
238
Wennstrom
S.
,
Siegbahn
A.
,
Yokote
K.
,
Arvidsson
A. K.
,
Heldin
C. H.
,
Moris
S.
,
Claesson-Welsh
L.
(
1994
).
Membrane ruffling and chemotaxis transduced by the PDGF beta-receptor require the binding site for phosphatidylinositol 3kinase.
Oncogene
9
,
651
660
Werb
Z.
,
Tremble
P.
,
Damsky
C.
(
1990
).
Regulation of extracellular matrix degradation by extracellular matrix interactions.
Cell Diff. Dev
32
,
299
306
Xu
J.
,
Clark
R. A. F.
(
1996
).
Extracellular matrix alters PDGF regulation of fibroblast integrins.
J. Cell Biol
132
,
239
249
Xu
J.
,
Zutter
M. M.
,
Santoro
S. A.
,
Clark
R. A.
(
1998
).
A three-dimensional collagen lattice activates NF-B in human fibroblasts: role in integrin 2 gene expression and tissue remodeling.
J. Cell Biol
140
,
709
719
Yatomi
Y.
,
Yamamura
S.
,
Ruan
F.
,
Igarashi
Y.
(
1997
).
Sphingosine 1-phosphate induces platelet activation through an extracellular action and shares a platelet surface receptor with lysophosphatidic acid.
J. Biol. Chem
272
,
5291
5297
Zhang
Q.
,
Peyruchaud
O.
,
French
K. J.
,
Magnusson
M. K.
,
Mosher
D. F.
(
1999
).
Sphingosine 1-phosphate stimulates fibronectin matrix assembly through a rho-dependent signal pathway.
Blood
93
,
2984
2990
Zondag
G. C. M.
,
Postma
F. R.
,
Van Etten
I.
,
Verlaan
I.
,
Moolenaar
W. H.
(
1998
).
Sphingosine-1-phosphate signaling through the G-protein-coupled receptor Edg-1.
Biochem. J
333
,
605
609
This content is only available via PDF.