013 Variable p53 Response in a 3D Matrix

Detachment of an attached (stressed) fibroblast‐populated collagen matrix (FPCM) induces fibroblast apoptosis; this may be secondary to increased p53 after detachment. We have noted, however, that the p53 response after detachment of an attached FPCM (i.e., stress‐release) varies among cells strains. We hypothesized that modulation of p53 after FPCM stress‐release may be related to the fibroblast population density at the time of release. Human foreskin fibroblasts (initially seeded at 0.5 × 10 6 /ml in 0.2 ml) were cultured for 48 hr in attached collagen matrices prior to detachment for 0–6 hr; p53 levels and lysate DNA concentration (used as a measure of fibroblast population density) were determined with immunoblot densitometry and a spectrophotometer, respectively. The experiment was performed with 15 different strains of fibroblasts. The p53 level increased (defined as ≥100% increase in baseline p53 level) during the 6 hr detachment period in 6/15 (40%) of fibroblast strains (responders), and downregulated or unchanged in the remaining strains (nonresponders). The mean DNA concentration in the responders vs . nonresponders was 1.56 μg/ml (median 1.63, range 0.89–2.25) vs . 2.57 (median 1.94, range 1.38–5.09), respectively (p < 0.05, Wilcoxon rank sum test). Stress‐release of the FPCM resulted in a variable p53 response which appeared to have a relationship with the fibroblast population density; a lower density was associated with p53 upregulation after detachment. This variable p53 response may in turn be related to varying proliferative capacity in the collagen matrix, as there was a broad range (nearly an order of magnitude) of DNA concentration among the 15 cell strains after 48 hr of attached FPCM cuture.