Retinoic Acid Inhibition of Human Bladder Tumor Cell Invasion in Membrane Invasion Culture System

Kim, Choung Soo

Abstract

During the development of invasive tumor, tumor cells penetrate the basement membrane and enter the underlying interstitial stroma to gain access to lymphatics and blood vessel for further dissemination. Invasion of the basement membrane is one of the most important steps. The basement membrane of bladder is composed of laminin and type IV collagen primarily and plasminogen activator and cathepsin B are increased in bladder carcinoma. Retinoic acid which is vitamin A metabolite suppresses carcinogenesis and promotes the differentiation of several types of tumor cells. Membrane invasion culture system (MICS) is an in vitro invasion assay using artificial basement membrane, Matrigel. Using this in vitro chemoinvasion assay, we evaluated the invasion of human invasive bladder cancer cell J82 and the ability of retinoic acid to inhibit the invasion of J82 quantitatively. The invaded tumor cells were 38.1+/-9.0 cells/field, 24.4+/-13.3 cells/field, 20.6+/-6.5 cells/field and 16.0+/-8.2 cells/field after adding retinoic acid 0uM, 0.1uM, 1.0uM and 10.0uM respectively and the decrease was statistically significant(p <0.05). Thereafter, the possible mechanisms of action responsible for the anti-invasive effect were further investigated, comparing retinoic acid-treated cells with untreated cells. The survival fractions were 100%, 82.4%, 65.3% and 46% with retinoic acid concentration 0uM, 0.1uM, 1.0uM and 10.0uM respectively on MTT test. Type IV collagenolytic activities were measured by type IV collagenolysis assay using [3H] proline labeled collagen film. Type IV collagenolysis activities were significantly decreased to 100%, 82.4 %, 65.3 % and 46.0% after adding retinoic acid, 0 M, 0.1 M, 1.0 M and 10.0 M respectively. But retinoic acid could not inhibit either the activity of plasminogen activator nor activity of cathepsin B enzyme significantly. These results suggest that retinoic acid inhibit the invasion of human invasive bladder cancer cell J82 and the inhibitory effect of retinoic acid may be caused by suppression of type IVcollagenolysis and direct cytotoxicity.