Richard D Kenagy1, Mete Civelek2, Shinsuke Kikuchi3, Lihua Chen4, Anthony Grieff4, Michael Sobel5, Aldons J Lusis6, Alexander W Clowes4. 1. Department of Surgery, University of Washington, Seattle, Wash. Electronic address: rkenagy@u.washington.edu. 2. Department of Medicine, University of California, Los Angeles, Los Angeles, Calif. 3. Department of Surgery, University of Washington, Seattle, Wash; Department of Vascular Surgery, Asahikawa Medical University, Asahikawa, Japan. 4. Department of Surgery, University of Washington, Seattle, Wash. 5. Department of Surgery, University of Washington, Seattle, Wash; Division of Vascular Surgery, VA Puget Sound Health Care System and University of Washington, Seattle, Wash. 6. Department of Medicine, University of California, Los Angeles, Los Angeles, Calif; Department of Microbiology, University of California, Los Angeles, Los Angeles, Calif; Department of Immunology and Molecular Genetics, University of California, Los Angeles, Los Angeles, Calif; Department of Human Genetics, University of California, Los Angeles, Los Angeles, Calif.
Abstract
OBJECTIVE: Approximately 30% of autogenous vein grafts develop luminal narrowing and fail because of intimal hyperplasia or negative remodeling. We previously found that vein graft cells from patients who later develop stenosis proliferate more in vitro in response to growth factors than cells from patients who maintain patent grafts. To discover novel determinants of vein graft outcome, we have analyzed gene expression profiles of these cells using a systems biology approach to cluster the genes into modules by their coexpression patterns and to correlate the results with growth data from our prior study and with new studies of migration and matrix remodeling. METHODS: RNA from 4-hour serum- or platelet-derived growth factor (PDGF)-BB-stimulated human saphenous vein cells obtained from the outer vein wall (20 cell lines) was used for microarray analysis of gene expression, followed by weighted gene coexpression network analysis. Cell migration in microchemotaxis chambers in response to PDGF-BB and cell-mediated collagen gel contraction in response to serum were also determined. Gene function was determined using short-interfering RNA to inhibit gene expression before subjecting cells to growth or collagen gel contraction assays. These cells were derived from samples of the vein grafts obtained at surgery, and the long-term fate of these bypass grafts was known. RESULTS: Neither migration nor cell-mediated collagen gel contraction showed a correlation with graft outcome. Although 1188 and 1340 genes were differentially expressed in response to treatment with serum and PDGF, respectively, no single gene was differentially expressed in cells isolated from patients whose grafts stenosed compared with those that remained patent. Network analysis revealed four unique groups of genes, which we term modules, associated with PDGF responses, and 20 unique modules associated with serum responses. The "yellow" and "skyblue" modules, from PDGF and serum analyses, respectively, correlated with later graft stenosis (P = .005 and P = .02, respectively). In response to PDGF, yellow was also associated with increased cell growth. For serum, skyblue was also associated with inhibition of collagen gel contraction. The hub genes for yellow and skyblue (ie, the gene most connected to other genes in the module), scavenger receptor class A member 5 (SCARA5) and suprabasin (SBSN), respectively, were tested for effects on proliferation and collagen contraction. Knockdown of SCARA5 increased proliferation by 29.9% ± 7.8% (P < .01), whereas knockdown of SBSN had no effect. Knockdown of SBSN increased collagen gel contraction by 24.2% ± 8.6% (P < .05), whereas knockdown of SCARA5 had no effect. CONCLUSIONS: Using weighted gene coexpression network analysis of cultured vein graft cell gene expression, we have discovered two small gene modules, which comprise 42 genes, that are associated with vein graft failure. Further experiments are needed to delineate the venous cells that express these genes in vivo and the roles these genes play in vein graft healing, starting with the module hub genes SCARA5 and SBSN, which have been shown to have modest effects on cell proliferation or collagen gel contraction.
OBJECTIVE: Approximately 30% of autogenous vein grafts develop luminal narrowing and fail because of intimal hyperplasia or negative remodeling. We previously found that vein graft cells from patients who later develop stenosis proliferate more in vitro in response to growth factors than cells from patients who maintain patent grafts. To discover novel determinants of vein graft outcome, we have analyzed gene expression profiles of these cells using a systems biology approach to cluster the genes into modules by their coexpression patterns and to correlate the results with growth data from our prior study and with new studies of migration and matrix remodeling. METHODS: RNA from 4-hour serum- or platelet-derived growth factor (PDGF)-BB-stimulated human saphenous vein cells obtained from the outer vein wall (20 cell lines) was used for microarray analysis of gene expression, followed by weighted gene coexpression network analysis. Cell migration in microchemotaxis chambers in response to PDGF-BB and cell-mediated collagen gel contraction in response to serum were also determined. Gene function was determined using short-interfering RNA to inhibit gene expression before subjecting cells to growth or collagen gel contraction assays. These cells were derived from samples of the vein grafts obtained at surgery, and the long-term fate of these bypass grafts was known. RESULTS: Neither migration nor cell-mediated collagen gel contraction showed a correlation with graft outcome. Although 1188 and 1340 genes were differentially expressed in response to treatment with serum and PDGF, respectively, no single gene was differentially expressed in cells isolated from patients whose grafts stenosed compared with those that remained patent. Network analysis revealed four unique groups of genes, which we term modules, associated with PDGF responses, and 20 unique modules associated with serum responses. The "yellow" and "skyblue" modules, from PDGF and serum analyses, respectively, correlated with later graft stenosis (P = .005 and P = .02, respectively). In response to PDGF, yellow was also associated with increased cell growth. For serum, skyblue was also associated with inhibition of collagen gel contraction. The hub genes for yellow and skyblue (ie, the gene most connected to other genes in the module), scavenger receptor class A member 5 (SCARA5) and suprabasin (SBSN), respectively, were tested for effects on proliferation and collagen contraction. Knockdown of SCARA5 increased proliferation by 29.9% ± 7.8% (P < .01), whereas knockdown of SBSN had no effect. Knockdown of SBSN increased collagen gel contraction by 24.2% ± 8.6% (P < .05), whereas knockdown of SCARA5 had no effect. CONCLUSIONS: Using weighted gene coexpression network analysis of cultured vein graft cell gene expression, we have discovered two small gene modules, which comprise 42 genes, that are associated with vein graft failure. Further experiments are needed to delineate the venous cells that express these genes in vivo and the roles these genes play in vein graft healing, starting with the module hub genes SCARA5 and SBSN, which have been shown to have modest effects on cell proliferation or collagen gel contraction.
Authors: George T Lau; Lloyd J Ridley; Paul G Bannon; Louise A Wong; Joseph Trieu; David B Brieger; Harry C Lowe; Ben S Freedman; Leonard Kritharides Journal: Circulation Date: 2006-07-04 Impact factor: 29.690
Authors: Xue Ma; Jeffrey D Pearce; David B Wilson; William P English; Matthew S Edwards; Randolph L Geary Journal: J Vasc Surg Date: 2013-06-12 Impact factor: 4.268
Authors: Richard D Kenagy; Nozomi Fukai; Seung-Kee Min; Florencia Jalikis; Ted R Kohler; Alexander W Clowes Journal: J Vasc Surg Date: 2009-05 Impact factor: 4.268
Authors: Hisham Bazzi; Katherine A Fantauzzo; Gavin D Richardson; Colin A B Jahoda; Angela M Christiano Journal: Dev Dyn Date: 2007-04 Impact factor: 3.780
Authors: Jonathan P Rehfuss; Kenneth M DeSart; Jared M Rozowsky; Kerri A O'Malley; Lyle L Moldawer; Henry V Baker; Yaqun Wang; Rongling Wu; Peter R Nelson; Scott A Berceli Journal: Circ Genom Precis Med Date: 2018-03
Authors: Richard D Kenagy; Shinsuke Kikuchi; Lihua Chen; Errol S Wijelath; Andrew B Stergachis; John Stamatoyannopoulos; Gale L Tang; Alexander W Clowes; Michael Sobel Journal: J Vasc Surg Date: 2017-05-16 Impact factor: 4.268
Authors: Michael Sobel; Shinsuke Kikuchi; Lihua Chen; Gale L Tang; Tom N Wight; Richard D Kenagy Journal: J Vasc Surg Date: 2018-06-15 Impact factor: 4.268
Authors: Shinsuke Kikuchi; Lihua Chen; Kevin Xiong; Yukihiro Saito; Nobuyoshi Azuma; Gale Tang; Michael Sobel; Thomas N Wight; Richard D Kenagy Journal: J Vasc Surg Date: 2017-06-21 Impact factor: 4.268
Authors: Rohaina Che Man; Nadiah Sulaiman; Mohamad Fikeri Ishak; Ruszymah Bt Hj Idrus; Mohd Ramzisham Abdul Rahman; Muhammad Dain Yazid Journal: Int J Environ Res Public Health Date: 2020-10-26 Impact factor: 3.390