G-protein expression and intimal hyperplasia after arterial injury: A role for Gα_i proteins

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 (α _o, α _i, α _q, α _s, and βγ) 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 Gα _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 Gα _i3 expression, which decreased at days 7, 14, and 28, compared with the uninjured carotid. Gα _q expression increased in a time-dependent manner. There was a marked time-dependent increase in Gβγ in the 28 days. Gα _i2 and Gα _s isoforms (45 and 52 kDa) did not change significantly with time. There was no major change in Gα _i1 and Gα _o in the study period. At 14 days, PTx treatment reduced intimal hyperplasia by 52% (63 ± 4 μm vs 30 ± 5 μm, 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 (Gα _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.