Dynamic remodeling of the vascular bed precedes tumor growth: MLS ovarian carcinoma spheroids implanted in nude mice

Abstract

The goal of this study was to monitor the vascular bed during the lag phase in growth of implanted spheroids as a model of tumor dormancy. Vascular development and tumor growth were followed up by magnetic resonance imaging in a model system of MLS ovarian carcinoma spheroids implanted subcutaneously in female nude mice. Apparent vessel density in a 1-mm rim surrounding the spheroid was evaluated by gradient echo imaging as a measure of the angiogenic potential of the tumor. Vascular functionality and maturation were assessed by signal intensity changes in response to hyperoxia (elevated oxygen) and hypercapnia (elevated carbon dioxide), respectively. Tumor growth was delayed by 12 to 57 days after implantation. During this long period in which tumor volume did not change, up to 6 cycles of vascular development and regression were observed. We propose here that dynamic remodeling of the vascular bed may precede exit of tumors from dormancy. The sustained oscillations in the angiogenic response to the implanted spheroid are consistent with hypoxic regulation of vascular endothelial growth factor (VEGF), combined with the role of VEGF as an essential survival factor for newly formed blood vessels. Vascular maturation, manifested by physiological vasodilatory response to carbon dioxide, may be important for conferring vascular stability and exit from dormancy.

Figure 1

Kinetics of tumor growth and vascularity. MLS spheroids, 1.2 mm in diameter, were implanted subcutaneously in female CD-1 nude mice at the age of 12 weeks. Up to 22 measurements were taken over 87 days for each mouse. (A, B) Lag phase and exponential growth of two different tumors. (C, D) Oscillations in AVD observed during the lag phase. Data were derived from analysis of gradient echo MRI as reported previously (18).

Figure 2

Oscillations in vascularity during the tumor lag phase. (A) Summary of number of oscillations in the lag phase (n=9). (B) Correlation between the duration of the lag phase and the number of oscillations (r=0.68, P=0.03, n=9).

Figure 3

Analysis of vascular function and maturation. Images were acquired 18 days after spheroid implantation and 3 days before the initiation of tumor growth. (A) Gradient echo image. The tumor in the center of the image is surrounded by a hypointense rim corresponding to neovasculature induced by the tumor. Gradient echo images were acquired during inhalation of air, air-carbon dioxide (95% air, 5% carbon dioxide) and carbogen (95% oxygen, 5% carbon dioxide). (B) Vascular functionality was assessed by signal changes in response to hyperoxia (VF). (C) Changes in signal intensity between air and air-carbon dioxide, corresponding to vasodilation (VD).

Figure 4

Model of dynamic vascular remodeling during tumor dormancy. Tumor growth leads to hypoxia, thus inducing expression of VEGF. This however does not lead inevitably to tumor exit from dormancy. The neovasculature can restore oxygenation and suppress VEGF expression. Acute VEGF withdrawal results in vascular collapse and tumor hypoxia. Such a dynamic cycle can trap the tumor in extended dormancy. Exit of tumors from dormancy will depend on stabilization of the vascular bed either by an exogenous source of VEGF (such as a local injury) or by vascular maturation.