Calreticulin, a potential vascular regulatory protein, reduces intimal hyperplasia after arterial injury

Both thrombotic and inflammatory responses to arterial injury have been implicated in atherosclerotic plaque growth. Calreticulin is a ubiquitous calcium-binding protein with antithrombotic activity and, in addition, is associated with leukocyte activation. We are investigating calreticulin as a potential vascular regulatory protein. The development of intimal hyperplasia was studied at sites of balloon injury in iliofemoral arteries from 91 rats. Calreticulin was infused directly into the artery immediately before balloon injury, and plaque growth was then assessed at 4 weeks' follow-up. Parallel studies of the effects of each calreticulin domain as well as a related calcium-binding protein, calsequestrin, were examined. The effects of calreticulin on platelet activation, clot formation, and mononuclear cell migration were also studied. When infused before balloon injury in rat iliofemoral arteries, calreticulin, or its high-capacity Ca(2+)-binding C domain, significantly reduces plaque development, whereas calsequestrin, a related calcium-binding protein that lacks the multifunctional nature of calreticulin, does not decrease plaque area (saline: 0.037 +/- 0.007 mm2, calsequestrin: 0.042 +/- 0.021 mm2, calreticulin: 0.003 +/- 0.002 mm2, n = 46, P < .04). The N domain and more specifically the P domain, a low-capacity, high-affinity calcium-binding domain in calreticulin, do not reduce intimal hyperplasia (N + P domain: 0.038 +/- 0.012 mm2, C domain: 0.003 +/- 0.002 mm2, n = 45 rats, P < .0001). Calreticulin reduces macrophage and T cell staining in the arterial wall after injury but has no direct effect on monocyte migration in vitro (percent medial area staining positive for macrophage 24 hours after injury (N + P: 4.06 +/- 1.42, calreticulin: 0.273 +/- 0.02; n = 26, P < .009). Calreticulin does, however, reduce platelet-dependent whole blood clotting time, in vitro (baseline: 78.23 +/- 2.04 seconds, calreticulin: 113.5 +/- 1.95 seconds; n = 5, P < .002). We conclude that calreticulin significantly reduces intimal hyperplasia after arterial injury, potentially acting as a vascular regulatory protein.