Locomotion of lymphocytes towards melanoma cells treated with tumor necrosis factor in a syngeneic in vitro model

Tumor necrosis factor (TNF) causes cell necrosis in vivo by damaging the en dothelium of the neovasculature. However, its mechanism of action is not we ll understood. We hypothesized that TNF affects the tumor microenvironment even before neovascularization occurs, thereby increasing lymphocyte locomo tion through the peritumoral matrix, a crucial step in tumor cell killing. The effect of TNF on lymphocytes was tested with the type I rat-tail collag en mini-assay in peripheral blood lymphocytes (PBL) from normal donors, a n on-migratory PBL cell line (HPB), and a C3H mice splenic lymphocytes. Melan oma cell line (k1735p) was treated with TNF alpha /TNF beta 10 or 20 pg/mul . The syngeneic splenic lymphocytes were layered on top of the collagen, an d their migration into the collagen towards the tumor cells was assessed. T umor cell viability was evaluated before and after TNF treatment. Paired tw o-tailed Student's t-test was used for statistical analysis. TNF alpha and TNF beta had no significant direct effect on locomotion of PBL or HPB. Lymp hocyte locomotion was inhibited in the presence of untreated melanoma cells in 7 of 9 assays (statistically significant in four), and it was significa ntly increased towards TNF alpha- or beta -treated melanoma cells, compared to untreated condition, in 7 of 9 assays (p=0.05 to p=0.0001). The number of viable tumor cells was not significantly different before and after trea tment. In conclusion, treatment of tumor cells with TNF alpha or TNF beta s ignificantly enhances lymphocyte locomotion through the matrix. The effect of TNF is not the result of a direct influence on the lymphocytes, and is n ot associated with a decrease in the number of viable tumor cells. These fi ndings suggest that TNF interaction with the cell microenvironment induces a change in lymphocyte locomotion.