abstract43 - GENE EXPRESSION, FIBRONECTIN AND SCARRING PATTERNS IN HUMAN FIBROBLASTS IN RESPONSE TO MESH AND CATHETER MATERIALS USING A NOVEL 3D COLLAGEN MODEL
GENE EXPRESSION, FIBRONECTIN ANDSCARRING PATTERNS IN HUMAN FIBROBLASTS IN RESPONSE TO MESH ANDCATHETER MATERIALS USING A NOVEL 3D COLLAGEN MODEL
Y. LI, L. STOTHERS, A. GHAHARY;
Univ. of British Columbia, Vancouver, Canada.
Introduction: Factors regulatingscarring secondary to mesh and prosthetic materials remain poorlyunderstood within the genitourinary tract. Fibrotic matrices containfibronectin; and alpha-smooth muscle actin contributes contraction.Metalloproteinases (MMPs) such as MMP-1 and -3 can modulate matrixprotein accumulation through degradation. Results from ourinstitution have shown that silica based materials can directlyinduce scarring through the interaction with tissue fibroblasts invitro. By extension, we hypothesized that other materials may inducefibrotic changes through cellular matrix gene expression.
Objective:The objectives of this study were: 1) to establish a 3D model ofhuman fibroblasts to study patterns of fibroblast response tomaterials and 2) measure gene expression in human fibroblasts exposedto prosthetic and mesh materials compared to a control.
Methods:Collagen gel was prepared by using 3 mg/ml in final concentrationwith 0.5% of polyvinyl alcohol (PVA). Mesh or catheter materials andhuman dermal fibroblasts (70,000 /ml) were added to a collagen geland seeded in a 24-well plate (0.5 ml of gel in each well) to createa 3D environment for fibroblast response. After polymerization ofcollagen, another 250,000 cells in 0.5 ml medium were added on thetop of gel. Cells were cultured at 37 0C, 5% CO2 forindicated time point. Images of cells were taken under reversemicroscopy to determine the pattern of the scarring contraction. Gelcell matrix was harvested and digested with 1 mg/ml of collagenasefor 15 minutes, pelleted by centrifugation and RNA was extracted.RT-PCR was performed for 32 cycles to analyze geneexpression.
Results: After 5 days, fibroblast contractionswere identified surrounding prosthetic materials but not within thecontrol. There were increases in type 1 collagen, α-smooth muscleactin and fibronectin expression in fibroblasts exposed to prostheticmaterials compared to fibroblasts grown in collagen gel alone. MMP-1and MMP-3 were also detected.
Conclusions: Fibroblastsexposed to mesh and catheter materials responded with an increase infibronectin, alpha smooth muscle actin and type 1 collagen that isincreased compared to controls. This may indicate why in vivo thesematerials induce fibrosis. Because fibronectin, type 1 collagen andalpha smooth muscle actin are main components of scarring and theirgene expression is elevated, future directions include development ofmedical devices that could induce downregulation of thesegenes.