RESPONSE OF MULTICELLULAR TUMOR SPHEROIDS TO LIPOSOMES CONTAINING TNF-ALPHA

This publication describes a new model to investigate the influence of tumor necrosis factor-alpha (TNF-alpha) on a three-dimensional glial cell aggregate under defined, standardized, reproducible conditions us ing the glioma cell line A 172. The cells are initially grown as norma l monolayer culture until they reach a cell density of up to 1 x 10(6) . Subsequently they are grown as spheroids by the liquid overlay techn ique. Spheroids grown in this way were divided into ten groups of more than 50 cell aggregates. Three groups were coincubated with free TNF- alpha in increasing dosages (100 ng/ml, 200 ng/ml and 1000 ng/ml); thr ee groups were incubated with empty liposomes (0.2 mg/ml, 0.4 mg/ml an d 2 mg/ml); three groups received liposomes which had been loaded with TNF-alpha, and one group, which received no treatment, served as cont rol. The diameter of the spheroids ranged from 80 mu m to 350 mu m. Th ere was no significant difference in growth between the 3 groups treat ed with 'free' TNF-alpha. Comparing spheroids treated with TNF-alpha w ith those which had been coincubated with empty liposomes, there was a significant difference (p < 0.001) in growth, which correlated with t he amount of liposomes. Similarly, free TNF-alpha had a significantly (P < 0.001) stronger growth-inhibiting effect as compared to liposomes loaded with TNF-alpha. Comparing the groups treated with liposomes on ly to those treated with liposomes loaded with TNF-alpha, the latter e xhibited a more marked (although not significantly) growth-inhibiting effect. The preliminary conclusion is that the major growth-inhibiting effect seems to be mediated by the liposomes. This phenomenon is in a greement with results obtained in monolayer cultures.