Early steps in hematogenous metastasis of B16F1 melanoma cells in chick embryos studied by high-resolution intravital videomicroscopy

Abstract

Background: There are few techniques that permit direct observation of tumor metastasis. The ability to observe steps in this process as they occur in experimental animals would complement studies on molecular mechanisms.

Purpose: We have developed a novel procedure using high-resolution intravital videomicroscopy to permit direct observation of cells as they arrest in the microcirculation, extravasate, and form micrometastases. We used this procedure to study early steps in experimental metastasis in immune-deficient chick embryos, permitting us to develop this technique in a relatively accessible respiratory organ and in the absence of host immune responses. Our goals were to develop techniques applicable to this host and to other hosts and to clarify the process of hematogenous tumor spread in this host.

Methods: We injected fluorescently labeled B16F1 melanoma cells into the circulation of 11- to 13-day chick embryos, and using intravital videomicroscopy, we observed the cells in the chorioallantoic membrane over time.

Results: The majority of injected cells were trapped initially in orifices to the chorioallantoic membrane capillary plexus or in tapering ends of arterioles leading to the plexus. During the first 2 hours, cells were found only in vessel lumina. After 8 hours, 83% of cells had extravasated, and the rest were in the process of extravasation. Cell shape changes and pseudopodial extensions were seen during extravasation and tumor development. Tumor cell division was seen only after extravasation. Tumors tended to develop near microvessels and were often wrapped around them.

Conclusions: Intravital videomicroscopy can provide new information about steps in metastasis. This procedure is applicable to other hosts and can be used in future studies to test hypotheses about molecular mechanisms of tumor spread.