Intimal hyperplasia (IH) in arterialized vein bypass grafts is a significant cause of vein graft (VG) stenosis and delayed graft failure. Injury at the time of implantation or as a consequence of transplantation into the high- pressure arterial system contributes to these delayed events. In a canine model, we have identified alterations in the transcriptome following implantation/arterialization injury, and have separated genetic events in the endothelium from those in medial smooth muscle cells (SMC). Using Systems Biology, we have identified the upregulated genes that were most essential to the injury response. Through back propagation, an integrated network was built starting with genes differentially expressed at the latest time-points i.e. 30 days (D), followed by adding upstream interactive genes from each prior time-point. This identified collagen 1A1 (Col1A1) at 30D, as a central cornerstone of back propagation and dominant contributor to IH lesions, as well as Interleukin (IL)- 6, IL-8, and PKC? as focus hub genes that were differentially upregulated across all time-points, starting at 2 hours (H) -12H post-surgery. These results establish causality relationships clarifying the pathogenesis of VG implantation injury, and identifying novel targets for its prevention. It is our hypothesis that silencing of focal hub and final lesion genes will diminish processes associated with VG implantation injury and thereby IH. Toward this goal, we have devised and refined methodology for silencing one or more genes under operating room constrains. In the proposed study we will apply siRNA technology to (i) systematically evaluate effectiveness and durability of silencing target genes (IL-6, IL-8, and PKC?, and Col1A1) and achieving protein knockdown in human saphenous vein endothelial cell (EC) and SMC cultures and confirming this effectiveness ex vivo in the intact wall of the human saphenous vein, (ii) determine the most successful siRNA cocktail that can prevent IH using a mouse vein bypass graft model and (iii) test the most effective siRNA cocktail and determine the genetic sequelae in a canine vein bypass graft translational model. State-of the art microarrays, Laser Capture Microdissection (LCM), as well as sophisticated and innovative global transcriptome analysis using Systems Biology will be employed. In addition, standardized immunohistochemistry, cellular, biochemical and molecular techniques will be used. Our investigative team has demonstrated the multidisciplinary collaboration essential to successful conduct of this proposal. We strongly believe this work will greatly strengthen the application of gene silencing to VG in patients, forecasting its expansion to other clinical problems in vascular surgery. In addition, this work will undoubtedly broaden our understanding of vascular wall biology.

Public Health Relevance

Scar tissue formation due to surgical injury is a major cause for failure of heart bypass grafts and bypass grafts for peripheral vascular disease. The investigators propose to prevent formation of this scar tissue from forming by controlling expression of the genes that cause it, using techniques that can be applied in the operating room.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
2R01HL086741-05
Application #
8437529
Study Section
Bioengineering, Technology and Surgical Sciences Study Section (BTSS)
Program Officer
Lundberg, Martha
Project Start
2007-02-01
Project End
2017-12-31
Budget Start
2013-01-08
Budget End
2013-12-31
Support Year
5
Fiscal Year
2013
Total Cost
$467,385
Indirect Cost
$198,773
Name
Beth Israel Deaconess Medical Center
Department
Type
DUNS #
071723621
City
Boston
State
MA
Country
United States
Zip Code
02215
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Nabzdyk, Christoph S; Chun, Maggie C; Oliver-Allen, Hunter S et al. (2014) Gene silencing in human aortic smooth muscle cells induced by PEI-siRNA complexes released from dip-coated electrospun poly(ethylene terephthalate) grafts. Biomaterials 35:3071-9
Pradhan-Nabzdyk, Leena; Huang, Chenyu; LoGerfo, Frank W et al. (2014) Current siRNA targets in atherosclerosis and aortic aneurysm. Discov Med 17:233-46
Nabzdyk, Christoph S; Chun, Maggie; Pradhan, Leena et al. (2011) High throughput RNAi assay optimization using adherent cell cytometry. J Transl Med 9:48
Andersen, Nicholas D; Chopra, Atish; Monahan, Thomas S et al. (2010) Endothelial cells are susceptible to rapid siRNA transfection and gene silencing ex vivo. J Vasc Surg 52:1608-15
Monahan, Thomas S; Andersen, Nicholas D; Martin, Michelle C et al. (2009) MARCKS silencing differentially affects human vascular smooth muscle and endothelial cell phenotypes to inhibit neointimal hyperplasia in saphenous vein. FASEB J 23:557-64