title: Exploring smooth muscle phenotype and function in a bioreactor model of abdominal aortic aneurysm.
abstract: BACKGROUND
  Vascular smooth muscle cells (SMC) are central to arterial structure and function yet their involvement in the progression of abdominal aortic aneurysm (AAA) disease is not well studied. The progressive and silent nature of AAA in man essentially restricts research to the use of "end-stage" tissue recovered during surgical repair. This study aimed to generate an ex vivo model of AAA using protease-treated porcine carotid arteries maintained in a novel bioreactor, and to compare the structural and functional changes in SMC cultured from the recovered vessels with those from human tissue acquired at elective surgical repair.
  
  
  METHODS
  Freshly isolated porcine arteries were pretreated with collagenase and/or elastase before culturing under flow in a bioreactor for 12 days. Human end-stage aneurysmal tissue and saphenous veins from age-matched controls were collected from patients undergoing surgery. SMC were cultured and characterised (immunocytochemistry, measurement of spread cell area) and assessed functionally at the level of proliferation (cell-counting) and matrix-metalloproteinase (MMP) secretion (gelatin zymography). Cellular senescence was investigated using β-galactosidase staining and apoptosis was quantified using a fluorescence-based caspase 3 assay.
  
  
  RESULTS
  Co-expression of alpha-smooth muscle actin and smooth muscle myosin heavy chain confirmed all cell populations as SMC. Porcine SMC harvested and cultivated after collagenase/elastase pretreatment displayed a prominent "rhomboid" morphology, increased spread area (32%, P < 0.01), impaired proliferation (47% reduction, P < 0.05), increased senescence (52%, P < 0.001), susceptibility to apoptosis and reduced MMP-2 secretion (60% decrease, P < 0.01) compared with SMC from vehicle, collagenase or elastase pre-treated vessels. Notably, these changes were comparable to those observed in human AAA SMC which were 2.4-fold larger than non-aneurysmal SMC (P < 0.001) and exhibited reduced proliferation (39% reduction, P < 0.001), greater apoptosis (4-fold increase, P < 0.001), and increased senescence (61%, P < 0.05).
  
  
  CONCLUSIONS
  Combined collagenase/elastase exposure of porcine artery maintained in a bioreactor under flow conditions induced a SMC phenotype characteristic of those cultured from end-stage AAA specimens. This model has potential and versatility to examine temporal changes in SMC biology and to identify the molecular mechanisms leading to early aberrancies in SMC function. In the longer term this may inform new targets to maintain aortic SMC content and drive cells to a "reparative" phenotype at early stages of the disease.
authors.affiliation: Division of Cardiovascular and Diabetes Research, Leeds Institute of Genetics, Health and Therapeutics (LIGHT), University of Leeds, Leeds, UK. medkep@leeds.ac.uk.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Riches
authors.forename: Timothy G
authors.initials: TG
authors.lastname: Angelini
authors.forename: Gurprit S
authors.initials: GS
authors.lastname: Mudhar
authors.forename: Jean
authors.initials: J
authors.lastname: Kaye
authors.forename: Emily
authors.initials: E
authors.lastname: Clark
authors.forename: Marc A
authors.initials: MA
authors.lastname: Bailey
authors.forename: Soroush
authors.initials: S
authors.lastname: Sohrabi
authors.forename: Sotirios
authors.initials: S
authors.lastname: Korossis
authors.forename: Peter G
authors.initials: PG
authors.lastname: Walker
authors.forename: D Julian A
authors.initials: DJ
authors.lastname: Scott
authors.forename: Karen E
authors.initials: KE
authors.lastname: Porter
mesh_terms: D000818:Animals; D017544:Aortic Aneurysm, Abdominal / Q000473:pathology*; D017209:Apoptosis / Q000187:drug effects; D019149:Bioreactors; D002339:Carotid Arteries / Q000187:drug effects / Q000473:pathology; D002452:Cell Count; D049109:Cell Proliferation / Q000187:drug effects; D048430:Cell Shape / Q000187:drug effects; D016922:Cellular Senescence / Q000187:drug effects; D017364:Collagenases / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D008297:Male; D020782:Matrix Metalloproteinases / Q000378:metabolism; D008954:Models, Biological; D009130:Muscle, Smooth / Q000187:drug effects / Q000473:pathology*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D010196:Pancreatic Elastase / Q000494:pharmacology; D010641:Phenotype; D034421:Sus scrofa
chemical_list: D010196:Pancreatic Elastase; D017364:Collagenases; D020782:Matrix Metalloproteinases
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