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    Temporal and spatial angiogenesis-associated gene expression in rat ischemic skin flaps

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    Authors
    Johnson, Thomas
    Issue Date
    2008-04
    URI

    http://hdl.handle.net/10675.2/623784
    
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    Abstract
    Background: Emerging therapies designed to improve ischemic soft tissue flap survival include the use of angiogenic factors. However, endogenous expression patterns for these factors have not been characterized. The purpose of this study was to identify variations in gene expression patterns for angiogenesis-associated genes in ischemic rat skin flaps with respect to post operative time and distance from flap base. Methods: Caudally based dorsal random pattern skin flaps measuring 3x10 cm were created on fifteen male Sprague-Dawley rats. The rats were sacrificed at one, three, or seven days post surgery. Following euthanasia, flaps were harvested and sectioned into four 3x2.5 cm zones, with zone I located furthest from the flap base. Total RNA was isolated from each zone, amplified, labeled with biotin, and hybridized with oligoDNA microarrays containing probes for 113 angiogenesis-associated genes. Microarrays were imaged using chemiluminescence, and signal intensities were analyzed using web based software. In a previously published report, flap survival was assessed using orthogonal polarization spectral imaging, and the areas of the flap characterized by blood flow versus stasis ( absent capillary blood flow) were identified. Results: Orthogonal polarization spectral imaging results showed the transition of tissue from stasis to necrosis over time. The area of necrosis increased from day one to day seven as necrotic tissue replaced viable tissue in the stasis zone. Rat angiogenesis microarray analysis revealed significant modulation in the expression of multiple genes (p<0.05) in the areas of the skin flap characterized by advancing necrosis and stasis. A burst of gene modulation occurred from day one to day three in the most distal portion of the flap, and a similar burst occurred in the area characterized by stasis from day three to day seven. Six patterns of gene expression were defined. Patterns of angiogenesisassociated gene expression were different in the areas of blood flow compared to the areas of necrosis and stl!5is. Modulation of various gene functional groups also varied temporally and spatially. Conclusions: This study represents the first attempt to characterize endogenous spatial and temporal angiogenesis-associated gene expression patterns in ischemic skin flaps. The molecular analysis performed in this study correlates with the hemodynamic profile previously published. The gene expression patterns observed in this study may represent responses to hypoxia and necrosis as these processes proceeded proximally from the distal flap margin. In the future, molecular analysis of ischemic tissue may allow clinicians to distinguish hopeless tissue from a flap that may be rescued via pharmacological or surgical intervention. Additionally, molecular analysis in treated and untreated tissue may offer insight into the mechanisms by which pharmacological agents enhance flap survival.
    Affiliation
    Medical College of Georgia
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