Stereological quantification of carboxyfluorescein-labeled rat lung metastasis: a new method for the assessment of natural killer cell activity and tumor adhesion in vivo and in situ

Abstract

The function of natural killer (NK) cells is often studied by assessing in vitro levels of NK cell mediated lysis of target cells, or by assessing in vivo levels of lung tumor cell retention or metastatic colonization of intravenously injected tumor cells. However, these methods do not permit direct quantification and visualization of NK cells and their targets in vivo and in situ. Here, a new approach is described to visualize effector-to-target interactions as well as to estimate total numbers of targets in the lung, in vivo and in situ. MADB106 tumor cells were vitally labeled using carboxyfluorescein (CFSE) and intravenously (i.v.) injected into Fischer 344 rats (10 6 cells/rat). This mammary adenocarcinoma derived cell line is syngeneic to the inbred Fischer 344 rat and highly sensitive to NK cell activity in vivo. Effector-to-target interactions were visualized by immunostaining. Using the optical fractionator method, total numbers of CFSE-labeled MADB106 tumor cells were estimated in the left lung of the animals 5 min after tumor inoculation. To further demonstrate the usefulness of this approach in reflecting in vivo processes, rats were inoculated with MADB106 cells and simultaneously with a single i.v. bolus of either 1 μg/kg adrenaline or saline. Both lungs were removed 5 min later. Adrenaline caused a significant 80% reduction in the total number of lung CFSE-labeled MADB106 tumor cells, suggesting a rapid modulation of metastasis by stress hormones. This new approach facilitates the monitoring of effector-to-target interactions and the quantification of immune cell function or tumor adhesion in vivo and in situ.