Thymosin β4 improves functional neurological outcome in a rat model of embolic stroke

Thymosin β4 (Tβ4) is a developmentally expressed 43-amino acid peptide that was originally isolated from thymic extract and exists in numerous tissues and isoforms (Goldstein et al., 1966). Tβ4 inhibits organization of the actin-cytoskeleton by sequestration of G-actin monomers thereby enabling cells to migrate (Goldstein et al., 2005). Additionally, Tβ4 may regulate nuclear actin thereby influencing chromatin remodeling and ultimately gene expression (Huff et al., 2004). Tβ4 has multiple biological functions in addition to its actin binding properties. Tβ4 is an angiogenic promoting molecule which stimulates endothelial cell migration and tubule formation (Malinda et al., 1999, Smart et al., 2007). Tβ4 also promotes healing properties in both wounds and corneal injury by increasing cell migration and reducing inflammation. (Malinda et al., 1999, Sosne et al., 2002). Clinically, Tβ4 promotes wound healing and is being tested in a clinical trial of wound healing (Guarnera et al., 2007). Tβ4 promotes cardiomyocyte survival and improves cardiac function after myocardial infarction (MI) in experimental adult mice (Bock-Marquette et al., 2004). Tβ4 administration reduces left ventricular scar volume and promotes cardiomyocyte cell survival within 24 hours of coronary ligation. Tβ4 activation of Akt has been proposed as a potential mechanism that increases cell survival after acute MI. Hinkel et al, demonstrated in a pig MI model that downregulation of Tβ4 in embryonic endothelial progenitor cells used to treat experimental MI confers a loss of cardiomyocyte survival, increases infarct volume and reduces left ventricle function (Hinkel et al., 2008). Furthermore, Smart et al, demonstrated that epicardial progenitor cells isolated from wild type adult hearts differentiate into smooth muscle and endothelial cells when cultured in the presence of Tβ4 (Smart et al., 2007). Collectively, Tβ4 may improve cardiac function by promoting cardiomyocyte survival and may stimulate epicardial progenitor cells to differentiate into smooth muscle and endothelial cell types to repair damaged myocardium. Over the past two decades, research on treatment of stroke has focused on neuroprotection and revascularization strategies. The only successful clinical trial that has resulted from this research is the use of tissue plasminogen activator (rt-PA) within 4.5 hours of symptom onset (The NINDS rt-PA Stroke Study Group 1995, Hacke et al., 2008). Use of rt-PA has been limited because of its narrow time-dependent treatment window as most stroke patients present to the Emergency Department well beyond six hours of symptom onset (California Acute Stroke Pilot Registry (CASPR) Investigators, 2005, Kleindorfer et al., 2007) Moreover, use of rt-PA is complicated by a 6.4% symptomatic intracerbral hemmorhage (ICH) rate which has caused considerable controversy regarding its practical use, particularly amongst Emergency physicians. The restriction on time and potential adverse effects have limited the use of rt-PA to approximately 3% of stroke patients (California Acute Stroke Pilot Registry (CASPR) Investigators, 2005, Kleindorfer et al., 2009). Thus, it is imperative to develop therapies for ischemic stroke designed specifically to reduce neurological deficits, which can be employed to treat the vast majority of patients. Tβ4 is expressed in both the developing and adult brain (Carpintero et al., 1999, Gomez-Marquez and Anadon, 2002). Specifically, Tβ4 is localized in growing neurites of neurons (van Kesteren et al., 2006, Yang et al., 2008). In many disease states such as focal ischemia and Huntington’s disease, gene expression of Tβ4 is upregulated (Vartiainen et al., 1996, Sapp et al., 2001). Moreover, Tβ4 is upregulated 12 hours to 7 days after focal ischemia (Vartiainen et al., 1996). Since actin dynamics contribute to cell migration and synaptogenesis, we speculate that administration of Tβ4 could enhance recovery. Using a rat model of embolic stroke, we tested the hypothesis that treatment with Tβ4 improves neurological functional outcome when administered 24 hours after onset of ischemia.