Purpose: Guanine nucleotide binding protein (G-protein) coupled receptors are involved in smooth muscle cell proliferation, but the role of G-proteins in arterial intimal hyperplasia has not been defined. This study examines the expression of G-proteins in the developing intimal hyperplasia after balloon injury of the rat carotid artery and specifically tests the hypothesis that the pertussis toxin sensitive G(i) G-protein subunit plays a role in the initiation of intimal hyperplasia.
Methods: In vitro responses to serum stimulation (10% fetal bovine serum) were examined in the presence and absence of pertussis toxin (PTx). After a standard balloon injury in male Sprague-Dawley rats, the expression of G-protein subunits (alpha(o), alpha(i), alpha(q), alpha(s), and betagamma) was determined by means of Western blotting in the first 28 days. Thereafter, a second set of animals was allocated to control and PTx-treated (a Galpha(i) inhibitor; 500 ng/mL in an externally applied 30% pluronic gel) groups. Smooth muscle cell proliferation was estimated by means of thymidine analogue 5-bromo-2' deoxyuridine incorporation 2 days after injury, and vessel dimensions were determined by means of videomorphometry 14 days after injury.
Results: There was inhibition of DNA synthesis and smooth muscle cell proliferation in response to serum with an IC(50) of 100 ng/mL. Three days after balloon injury, there was an increase in Galpha(i3) expression, which decreased at days 7, 14, and 28, compared with the uninjured carotid. Galpha(q) expression increased in a time-dependent manner. There was a marked time-dependent increase in Gbetagamma in the 28 days. Galpha(i2) and Galpha(s) isoforms (45 and 52 kDa) did not change significantly with time. There was no major change in Galpha(i1) and Galpha(o) in the study period. At 14 days, PTx treatment reduced intimal hyperplasia by 52% (63 +/- 4 microm vs. 30 +/- 5 microm, control vs. PTx; P <.001). Medial smooth muscle cell proliferation at day 2 was decreased in the PTx group, compared with that in the gel-coated group (15% +/- 2% and 26% +/- 3%; P = .02).
Conclusion: After balloon injury, there is a time-dependent increase in G-protein expression, which is subunit specific. Activation of PTx sensitive G-proteins (Galpha(i)) is involved during the initiation of intimal hyperplasia after arterial injury, and their inhibition results in a decrease in early medial cell proliferation. This acute interruption of G(i) signaling produces a long-term decrease in intimal hyperplasia.