Abstract W MP51: Insulin-like Growth Factor (igf-1) Enhances The Angiogenic Response Of Aging Endothelial Cells
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Background and Purpose: IGF-1 is recognized as a neuroprotective agent in ischemic stroke models. Our microRNA profiling and KEGG analysis studies identified brain endothelial cells as principal target of IGF-1 action in post-stroke middle-aged females (Bake et al., PLOS One, 2014). Although IGF-1 is known to promote cell survival, the mode of action on injured brain endothelial survival is poorly understood. The present study specifically tested the hypothesis that IGF-1 prevents cell death and enhances angiogenic response of endothelial cells ex vivo. Additionally, since clinical evidence show that older women are at increased risk for stroke occurrence and greater severity, the present study used middle-aged female rats. Methods: Brain microvascular endothelial cells were harvested from acyclic Sprague-Dawley female rats (9-10 mo old). Confluent cultures grown on 24 well plates were exposed to hypoxia (1%O2) in glucose free media for 6h with and without IGF-1 (10ng/ml), followed by re-oxygenation in growth media. Cell death was analyzed by LDH assay in media. For invasion analysis, cells were suspended in media supplemented with VEGF, FGF and reduced serum and were plated onto 96 well plates containing collagen matrix with 1um sphingosine-1-phosphate. Cells were treated as before for hypoxia/aglycemia with and without IGF-1. Gels were fixed (3% glutaraldehyde), stained (toluidine blue in 30% methanol) and sectioned (1mm slices). Invasion density was quantified using Olympus CK2 inverted microscope. Results: Hypoxia resulted in 2-fold increase in media LDH, however, IGF-1 did not alter LDH levels after hypoxia or hypoxia/re-oxygenation, indicating that the peptide does not regulate ischemic cell death in the short-term. However, IGF-1 treatment resulted in a 50% increase in the number of sprouting endothelial cells after hypoxia/re-oxygenation (p<0.05), as compared to the vehicle treated group. Conclusions: These findings support the hypothesis that endothelial cells are target for IGF-1-induced stroke neuroprotection and that IGF-1 promotes angiogenic capacity in aging endothelial cells.
