Arterial gene transfer of the TGF-beta signalling protein Smad3 induces adaptive remodelling following angioplasty: a role for CTGF.
OBJECTIVE
Although transforming growth factor-beta (TGF-beta) is believed to stimulate intimal hyperplasia after arterial injury, its role in remodelling remains unclear. We investigate whether Smad3, a TGF-beta signalling protein, might facilitate its effect on remodelling.


RESULTS
Using the rat carotid angioplasty model, we assess Smad3 expression following arterial injury. We then test the effect of arterial Smad3 overexpression on the response to injury, and use a conditioned media experimental design to confirm an Smad3-dependent soluble factor that mediates this response. We use small interfering RNA (siRNA) to identify this factor as connective tissue growth factor (CTGF). Finally, we attempt to replicate the effect of medial Smad3 overexpression through adventitial application of recombinant CTGF. Injury induced medial expression of Smad3; overexpression of Smad3 caused neointimal thickening and luminal expansion, suggesting adaptive remodelling. Smad3 overexpression, though exclusively medial, caused adventitial changes: myofibroblast transformation, proliferation, and collagen production, all of which are associated with adaptive remodelling. Supporting the hypothesis that Smad3 initiated remodelling and these adventitial changes via a secreted product of medial smooth muscle cells (SMCs), we found that media conditioned by Smad3-expressing recombinant adenoviral vector (AdSmad3)-infected SMCs stimulated adventitial fibroblast transformation, proliferation, and collagen production in vitro. This effect was attenuated by pre-treatment of SMCs with siRNA specific for CTGF, abundantly produced by AdSmad3-infected SMCs, and significantly up-regulated in Smad3-overexpressing arteries. Moreover, periadventitial administration of CTGF replicated the effect of medial Smad3 overexpression on adaptive remodelling and neointimal hyperplasia.


CONCLUSIONS
Medial gene transfer of Smad3 promotes adaptive remodelling by indirectly influencing the behaviour of adventitial fibroblasts. This arterial cell-cell communication is likely to be mediated by Smad3-dependent production of CTGF.
Kundi
Rishi
R
Division of Vascular Surgery, Weill Medical College of Cornell University, Columbia College of Physicians and Surgeons, New York Presbyterian Hospital, New York, NY, USA.
Hollenbeck
Scott T
ST
Yamanouchi
Dai
D
Herman
Brad C
BC
Edlin
Rachel
R
Ryer
Evan J
EJ
Wang
Chunjie
C
Tsai
Shirling
S
Liu
Bo
B
Kent
K Craig
KC
D000800:Angioplasty, Balloon / Q000009:adverse effects*; D000818:Animals; D020212:Carotid Artery Injuries / Q000235:genetics / Q000378:metabolism / Q000473:pathology / Q000628:therapy*; D017536:Carotid Artery, Common / Q000378:metabolism* / Q000473:pathology; D049109:Cell Proliferation; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis; D055513:Connective Tissue Growth Factor / Q000235:genetics / Q000378:metabolism*; D017077:Culture Media, Conditioned / Q000378:metabolism; D004195:Disease Models, Animal; D005347:Fibroblasts / Q000378:metabolism; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000378:metabolism; D019899:Paracrine Communication; D034622:RNA Interference; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000378:metabolism; D051900:Smad3 Protein / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*
C526316:CCN2 protein, rat; D017077:Culture Media, Conditioned; D011994:Recombinant Proteins; D051900:Smad3 Protein; C497787:Smad3 protein, rat; D053773:Transforming Growth Factor beta1; D055513:Connective Tissue Growth Factor; D003094:Collagen