Initial Stages of Tumor Cell-Induced Angiogenesis: Evaluation Via Skin Window Chambers in Rodent Models

TLDR

There is a paucity of information about the immediate events following tumor‑cell initiation of angiogenesis, which is relevant to the timing of micrometastatic vascularization and treatment accessibility. The study aimed to monitor the earliest stage of tumor‑induced angiogenesis in vivo and to identify chemotactic signals that drive tumor cells toward existing vasculature, offering potential targets to prevent progression and metastasis. GFP‑labeled R3230Ac or 4T1 cells (20–50 cells) were implanted into dorsal skinfold window chambers of Fischer 344 rats or BALB/c mice, and tumor angiogenesis was serially and noninvasively monitored for up to four weeks. Angiogenesis began when the tumor mass reached roughly 100–300 cells, with host vasculature modification evident at 60–80 cells; tumor cells displayed chemotaxis toward existing vessels, and ex‑flk1 injection markedly inhibited early angiogenic and tumor growth activities.

Abstract

There is a paucity of information about events that follow immediately after tumor cells are triggered to initiate the process of angiogenesis (the formation of new blood vessels). Such information is relevant to the issue of when micrometastases vascularize and has implications for the accessibility of micrometastases to various treatments. In this study, we attempted to monitor events at the initiation of angiogenesis at the earliest possible stage of tumor growth in vivo.Two different rodent mammary tumor cell lines, R3230Ac from the Fischer 344 rat and 4T1 from the BALB/c mouse, were stably transfected with a gene that encodes an enhanced version of green fluorescence protein (GFP). GFP-labeled R3230Ac or 4T1 cells (about 20-50 cells) were implanted into dorsal skinfold window chambers of Fischer 344 rats or BALB/c mice, respectively. Tumor angiogenesis was then monitored serially and noninvasively for up to 4 weeks.Clear evidence of modification of the host vasculature was observed when tumor mass reached approximately 60-80 cells, and functional new blood vessels were seen when tumor mass reached roughly 100-300 cells. Individual tumor cells exhibited a chemotaxis-like growth pattern toward the pre-existing host vasculature. When ex-flk1 (a soluble, truncated vascular endothelial cell growth factor receptor protein known to be antiangiogenic) was injected with the tumor cells, the initial angiogenic and tumor growth activities were inhibited considerably, indicating that angiogenesis inhibitors may halt tumor growth even before the onset of angiogenesis.Angiogenesis induced by tumor cells after implantation in the host begins at a very early stage, i.e., when the tumor mass contains roughly 100-300 cells. Identification of chemotactic signals that initiate tumor cell migration toward the existing vasculature may provide valuable targets for preventing tumor progression and/or metastases.