Effects of local cytochalasin D delivery on smooth muscle cell migration and on collar-induced intimal hyperplasia in the rabbit carotid artery

Abstract

1. Smooth muscle cell (SMC) migration has been implicated in neointima formation after angioplasty. Therefore, we investigated whether cytochalasin D, a fungal metabolite that inhibits actin filament formation, suppressed SMC migration and collar-induced intimal hyperplasia in the rabbit carotid artery. 2. To establish effective concentrations, contractions of carotid artery rings to phenylephrine were determined after incubation with cytochalasin D (10(-8) - 10(-6) M) for 30 min or 3 days. In vitro cell migration was studied using carotid artery explants and a modified Boyden chamber with SMCs isolated from the rabbit aorta. The in vivo effect was tested after infusion of 10(-8) - 10(-4) M cytochalasin D into collars placed around the left carotid artery; collars placed around the right artery served as controls. 3. Contractions to phenylephrine decreased after 30 min or 3 days exposure to 10(-7) and 10(-6) M cytochalasin D; the effect was partly reversible. These concentrations also inhibited cellular outgrowth and SMC migration in the in vitro assays. 4. Immunohistochemistry showed that local delivery of 10(-5) or 10(-4) M cytochalasin D for 2 weeks suppressed collar-induced alpha-SMC actin expression in the intima by 68% and 84% respectively. However, the cross-sectional area of the intima was not reduced due to an influx of T-lymphocytes and macrophages. 5. It is concluded that cytochalasin D suppressed SMC contractility and migration in vitro. Although perivascular infusion of cytochalasin D inhibited collar-induced SMC migration from media to intima in vivo as well, the intimal hyperplasia was not reduced due to concomitant development of an inflammatory response.

Figure 1

Phenylephrine contractions in segments of the rabbit carotid artery after in vitro exposure to cytochalasin for 30 min (A, acute) or 3 days (B, chronic). Data represent mean±s.e.mean, n=6 – 7. *P<0.05, **P<0.01, different from control (0 M), Dunnett's test.

Figure 2

Cytochalasin D inhibits cellular outgrowth from rabbit carotid artery segments in culture. *P<0.05, different from control (0 M), Dunnett's test.

Figure 3

Photomicrographs of cross-sections of rabbit carotid arteries infused for 14 days (B, D, F, H) or 5 days (J) with 10⁻⁴ M cytochalasin D (right panels) and their contra-lateral controls (A, C, E, G, I, left panels). Phalloidin: Oregon green 488-phalloidin; α-actin: α-smooth muscle actin; RAM11: macrophage marker; CD43 T-lymphocyte marker. ∧: internal elastic lamina. Bar=10 μm.

Figure 4

Effect of local cytochalasin D infusion for 14 days on absolute (A) and relative (B) α-smooth muscle actin immunoreactive areas in the intima. Data represent mean±s.e.mean, n=8 (10⁻⁸ – 10⁻⁶ M) or n=4 (10⁻⁵ – 10⁻⁴ M). Control: contralateral artery with a collar not connected to an osmotic minipump. * Different from control: P<0.05, Student's t-test.

Figure 5

Effect of local cytochalasin D infusion for 14 days on α-smooth muscle actin, RAM11 and CD43 immunoreactive areas in the intima of rabbit carotid arteries. Data represent mean±s.e.mean, n=4. Control: contralateral artery with a collar not connected to an osmotic minipump.