Recent studies showed that soluble annexin A2 dramatically increases tissue plasminogen activator (tPA)-mediated plasmin generation in vitro, and reduces thrombus formation in vivo. Here, we hypothesize that combining annexin A2 with tPA can significantly enhance thrombolysis efficacy, so that lower doses of tPA can be applied in ischemic stroke to avoid neurotoxic and hemorrhagic complications. In vitro activity assays confirmed tPA-specific amplification of plasmin generation by recombinant annexin A2. In a rat focal embolic stroke model, combination therapy with tPA and recombinant annexin A2 protein at 2 h post-ischemia decreased the effective dose required for tPA by four-fold and reduced brain infarction. Combining annexin A2 with tPA also lengthened the time window for thrombolysis. Compared with tPA (10 mg/kg) alone, the combination of annexin A2 (5 mg/kg) plus low-dose tPA (2.5 mg/kg) significantly enhanced fibrinolysis, attenuated mortality, brain infarction, and hemorrhagic transformation, even when administered at 4 h post-ischemia. Combination with recombinant annexin A2, the effective thrombolytic dose of tPA can be decreased. As a result, brain hemorrhage and infarction are reduced, and the time window for stroke reperfusion prolonged. Our present findings provide a promising new approach for enhancing tPA-based thrombolytic stroke therapy.

Objective-To determine the role of the cardiovascular-restricted, hairy-related bHLH transcription factor, CHF1/Hey2, in the biological response to vascular injury. Methods and Results-We investigated the response of CHF1/Hey2-deficient mice to vascular injury in vivo and the response of primary cultured vascular smooth muscle cells (VSMCs) from these mice to growth factors in vitro. Neointima formation after arterial wire injury is decreased in knockout (KO) compared with wild-type (WT) mice (0.025Ϯ0.011 mm 2 in WT [nϭ13]) versus 0.016Ϯ0.008 mm 2 in KO (nϭ12; PϽ0.05) and is accompanied by reduced cellular proliferation. CHF1/Hey2-deficient VSMCs proliferate slowly compared with WT VSMCs and also show decreased migration in response to platelet-derived growth factor (PDGF) (62.6Ϯ10.3 CPF versus 37.2Ϯ13.5 CPF; PϽ0.01) and heparin-binding epidermal growth factor-like growth factor (HB-EGF) (27.4Ϯ7.7 CPF versus 6.4Ϯ3.7 CPF, PϽ0.05). Furthermore, lamellipodia formation and membrane ruffling induced by these chemoattractants are diminished in KO VSMCs, which is correlated with decreased activation of the small GTPase Rac1. Although total Rac1 protein was not changed in KO VSMCs, the level of the Rac guanine exchange factor (GEF), Sos1, was decreased. Key Words: genetically altered mice Ⅲ growth factors Ⅲ vascular biology Ⅲ vascular muscle T he vascular smooth muscle cell (VSMC) is a major component of the arterial wall and plays a critical role in the development of occlusive vascular lesions. 1 In normal vessels, VSMCs are quiescent, differentiated, contractile, and function to maintain vascular tone and blood pressure. In response to injury, VSMCs undergo a phenotypic transition whereby they proliferate, migrate from the medial layer to the intima, secrete matrix metalloproteinases, decrease their expression of contractile proteins, and increase their expression of extracellular matrix molecules. The critical steps that regulate this phenotypic transition are not completely understood. Conclusions-CHF1/Hey2Previously, we and other groups cloned the bHLH protein CHF1/Hey2. 2-7 CHF1/Hey2 (also known as Hesr-2, HRT2, HERP1, and gridlock) is one of only a few tissue-restricted bHLH transcription factors, along with relatives CHF2/Hey1 and CHF3/HeyL, expressed in VSMCs. Studies of the zebrafish homologue of CHF1/Hey2, gridlock, suggested a role in development of the aorta 6 and in patterning of the arterial and venous systems. 8 Other studies have suggested that CHF1/Hey2 gene expression decreases after vascular injury and growth factor stimulation, and that immortalized smooth muscle cell lines overexpressing the related gene CHF2/Hey1 (also known as HRT1 and Hesr-1) show enhanced smooth muscle cell growth and protection against apoptosis. 9,10 CHF2/Hey1 has also been suggested to play a role in vascular endothelial cell differentiation. 11 Despite these interesting findings about CHF1/Hey2 and its relative CHF2/Hey1, the function of CHF1/Hey2 in the adult vasculature has not been demonstrated in vivo by loss of funct...

No abstract