Gene expression alterations induced by low molecular weight heparin during bowel anastomosis healing in rats
Languages of publication
Colon anastomosis is therapeutically challenging because multiple, usually undetectable factors influence a spectrum of repair mechanisms. We hypothesized that low molecular weight heparins, routinely administered perioperatively, may differentially affect gene expression related to colon healing. Twenty pairs of untreated and enoxaparin-treated rats underwent left-side hemicolectomy with a primary end-to-end anastomosis. Normal colon and anastomotic bowel segments were resected on day 0 and on days 1, 3, 5, and 7 after surgery, respectively. Serial anastomosis transverse cross-sections were evaluated microscopically and by microarray (Rat Genome 230 2.0, Affymetrix). Differentially expressed probe sets were annotated with Gene Ontology. We also examined the influence of enoxaparin on fibroblast proliferation and viability in vitro. Among the 5476 probe sets, we identified differential expression at each healing time point, yielding 79 subcategories. Most indicated genes were involved in wound healing, including multicellular organismal development, locomotory behavior, immune response, cell adhesion, inflammatory response, cell-cell signaling, blood vessel development, and tissue remodeling. Although we found no intensity differences in histological features of healing between enoxaparin-treated and control rats, treatment did induce significant expression changes during early healing. Of these changes, 83 probe sets exhibited at least twofold changes and represented different functional annotations, including inflammatory response, regulation of transcription, regulation of apoptosis, and angiogenesis. Fibroblast culture confirmed an anti-viability effect of enoxaparin. Enoxaparin affects colon wound-related gene expression profiles, but further studies will resolve whether heparin treatment is a risk factor after intestinal surgery, at least in some patients.
- Affymetrix (2002) GeneChip® Expression Analysis - Data Analysis Fundamentals.
- Arikan S, Adas G, Barut G, Toklu AS, Kocakusak A, Uzun H, Kemik O, Daduk Y, Aydin S, Purisa S (2005) An evaluation of low molecular weight heparin and hyperbaric oxygen treatment in the prevention of intra-abdominal adhesions and wound healing. Am J Surg 189: 155-160.
- Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, Davis AP, Dolinski K, Dwight SS, Eppig JT, Harris MA, Hill DP, Issel-Tarver L, Kasarskis A, Lewis S, Matese JC, Richardson JE, Ringwald M, Rubin GM, Sherlock G (2000) Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nat Genet 25: 25-29.
- Basson MD (2003) Invited research review: Cell-matrix interactions in the gut epithelium. Surgery 133: 263-267.
- Benjamini Y, Hochberg Y (1995) Controlling the false discovery rate: a practical and powerful approach to multiple testing. J Royal Statistical Society 57: 289-300.
- Carroll LA, Koch RJ (2003) Heparin stimulates production of bFGF and TGF-beta 1 by human normal, keloid, and fetal dermal fibroblasts. Med Sci Monit 9: BR97-BR108.
- Cole J, Tsou R, Wallace K, Gibran N, Isik F (2001) Early gene expression profile of human skin to injury using high-density cDNA microarrays. Wound Repair Regen 9: 360-370.
- Collen A, Smorenburg SM, Peters E, Lupu F, Koolwijk P, Van Noorden C, van Hinsbergh VW (2000) Unfractionated and low molecular weight heparin affect fibrin structure and angiogenesis in vitro. Cancer Res 60: 6196-6200.
- Colwell AS, Longaker MT, Peter Lorenz H (2008) Identification of differentially regulated genes in fetal wounds during regenerative repair. Wound Repair Regen 16: 450-459.
- Cooper L, Johnson C, Burslem F, Martin P (2005) Wound healing and inflammation genes revealed by array analysis of 'macrophageless' PU.1 null mice. Genome Biol 6: R5.
- Dayem MA, Moreilhon C, Turchi L, Magnone V, Christen R, Ponzio G, Barbry P (2003) Early gene expression in wounded human keratinocytes revealed by DNA microarray analysis. Comp Funct Genomics 4: 47-55.
- Eberl T, Jagoditsch M, Klingler A, Tschmelitsch J (2008) Risk factors for anastomotic leakage after resection for rectal cancer. Am J Surg 196: 592-598.
- Falcon S, Gentleman R (2007) Using GOstats to test gene lists for GO term association. Bioinformatics 23: 257-258.
- Ferrao AV, Mason RM (1993) The effect of heparin on cell proliferation and type-I collagen synthesis by adult human dermal fibroblasts. Biochim Biophys Acta 1180: 225-230.
- Gentleman RC, Carey VJ, Bates DM, Bolstad B, Dettling M, Dudoit S, Ellis B, Gautier L, Ge Y, Gentry J, Hornik K, Hothorn T, Huber W, Iacus S, Irizarry R, Leisch F, Li C, Maechler M, Rossini AJ, Sawitzki G, Smith C, Smyth G, Tierney L, Yang JY, Zhang J (2004) Bioconductor: open software development for computational biology and bioinformatics. Genome Biol 5: R80.
- Gurtner GC, Werner S, Barrandon Y, Longaker MT (2008) Wound repair and regeneration. Nature 453: 314-321.
- Khatri P, Draghici S (2005) Ontological analysis of gene expression data: current tools, limitations, and open problems. Bioinformatics 21: 3587-3595.
- Khorana AA, Sahni A, Altland OD, Francis CW (2003) Heparin inhibition of endothelial cell proliferation and organization is dependent on molecular weight. Arterioscler Thromb Vasc Biol 23: 2110-2115.
- Kock HJ, Werther S, Uhlenkott H, Taeger G (2002) [Influence of unfractionated and low-molecular-weight heparin on bone healing: an animal model]. Unfallchirurg 105: 791-796.
- Krzesniak-Wszola N, Bielecki K, Ostrowski J (2007) Do low-molecular-weight heparins influence the healing process in colon anastomosis? Colorectal Dis 9: 489-493.
- Kuzu MA, Kuzu L, Koksov C, Akryl FH, Uzol D, Kale IT, Orhan D, Terzi C (1998) Histological evaluation of colonic anastomotic healing in the rat following preoperative 5-fluorouracil, fractionated irradiation, and combined treatment. Int J Colorectal Dis 13: 235-240.
- Lake AC, Castellot JJ Jr (2003) CCN5 modulates the antiproliferative effect of heparin and regulates cell motility in vascular smooth muscle cells. Cell Commun Signal 1: 5.
- Lane DA, Lindahl U (1989) Heparin: chemical and biological properties, clinical applications. CRC Press Boca Raton.
- Lustosa SA, Matos D, Atallah AN, Castro AA (2002) Stapled versus handsewn methods for colorectal anastomosis surgery: a systematic review of randomized controlled trials. Sao Paulo Med J 120: 132-136.
- Makela JT, Kiviniemi H, Laitinen S (2003) Risk factors for anastomotic leakage after left-sided colorectal resection with rectal anastomosis. Dis Colon Rectum 46: 653-60.
- Matzsch T, Bergqvist D, Blomquist P, Jiborn H (1987) Influence of standard heparin or low molecular weight heparin on healing of abdominal wounds and colonic anastomoses in rats. Acta Chir Scand 153: 593-8.
- Ostrowski J, Mikula M, Karczmarski J, Rubel T, Wyrwicz LS, Bragoszewski P, Gaj P, Dadlez M, Butruk E, Regula J (2007) Molecular defense mechanisms of Barrett's metaplasia estimated by an integrative genomics. J Mol Med 85: 733-743.
- Schmidt O, Merkel S, Hohenberger W (2003) Anastomotic leakage after low rectal stapler anastomosis: significance of intraoperative anastomotic testing. Eur J Surg Oncol 29: 239-243.
- Singer AJ, Clark RA (1999) Cutaneous wound healing. N Engl J Med 341: 738-746.
- Steffensen B, Hakkinen L, Larjava H (2001) Proteolytic events of wound-healing - coordinated interactions among matrix metalloproteinases (MMPs), integrins, and extracellular matrix molecules. Crit Rev Oral Biol Med 12: 373-398.
- Street J, Power C, Wakai A, Wang JH, McGuinness A, Redmond HP (2000a) Low molecular weight heparins induce osteoblast apoptosis: a putative mechanism for iatrogenic osteopenia. J Bone Joint Surgery 84-B: 7.
- Street JT, McGrath M, O'Regan K, Wakai A, McGuinness A, Redmond HP (2000b) Thromboprophylaxis using a low molecular weight heparin delays fracture repair. Clin Orthop Relat Res (381) 278-289.
- Sturm A, Dignass AU (2008) Epithelial restitution and wound healing in inflammatory bowel disease. World J Gastroenterol 14: 348-353.
- Stumpf M, Klinge U, Wilms A, Zabrocki R, Rosch R, Junge K, Krones C, Schumpelick V (2005) Changes of the extracellular matrix as a risk factor for anastomotic leakage after large bowel surgery. Surgery 137: 229-234.
- Tiozzo R, Reggiani D, Cingi MR, Bianchini P, Osima B, Calandra S (1991) Effect of heparin derived fractions on the proliferation and protein synthesis of cells in culture. Thromb Res 62: 177-188.
- Wu Z, Irizarry RA, Gentleman R, Martinez-Murillo F, Spencer F (2004) A model based background adjustment for oligonucleotide expression arrays. J Am Stat Ass 99: 909-917.
- Verhofstad MH, Lange WP, van der Laak JA, Verhofstad AA, Hendriks T (2001) Microscopic analysis of anastomotic healing in the intestine of normal and diabetic rats. Dis Colon Rectum 44: 423-431.
Publication order reference