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DOI: 10.1148/rg.25si055512
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RadioGraphics 2005;25:S85-S97
© RSNA, 2005

MR Angiography of Tumor-related Vasculature: From the Clinic to the Micro-environment1

Marion van Vliet, MD, Cornelis F. van Dijke, MD, PhD, Piotr A. Wielopolski, PhD, Timo L. M. ten Hagen, PhD, Jifke F. Veenland, PhD, Anda Preda, MD, Antonius J. Loeve, BSc, Alexander M. M. Eggermont, MD, PhD and Gabriel P. Krestin, MD, PhD

1 From the Departments of Radiology (M.v.V., C.F.v.D., P.A.W., J.F.V., A.P., A.J.L., G.P.K.), Surgical Oncology (T.L.M.t.H., A.M.M.E.), and Medical Informatics (J.F.V.), Erasmus MC–University Medical Center Rotterdam, Dr Molewaterplein 40, 3015 GD Rotterdam, the Netherlands. Presented as an education exhibit at the 2004 RSNA Annual Meeting. Received February 28, 2005; revision requested April 11 and received May 25; accepted May 31. All authors have no financial relationships to disclose. Address correspondence to C.F.v.D. (e-mail: c.vandijke{at}erasmusmc.nl).


   Abstract
Top
 Abstract
Introduction
 Assessment of Angiogenesis
 Conclusions
 References
 
Angiogenesis is a very important process for tumor growth and proliferation. Given its high temporal and spatial resolution, magnetic resonance (MR) imaging is well suited for use in the assessment of angiogenesis. MR angiography can be used clinically and experimentally for identification of tumor feeding and draining vessels, for tumor characterization, and for treatment planning. The morphologic structure of tumor vessels can be investigated in relation to tumor vessel permeability with use of specific contrast agents. To gain insight into tumor angiogenesis in vivo, the authors compared images obtained with digital photography, high-resolution MR angiography, and intravital microscopy through a dorsal skin-fold window in a rodent model. The close correlation between images obtained with these various modalities, with regard to the depiction of the developing tumor vasculature, indicates that noninvasive quantification of angiogenesis may be possible with MR imaging. Future directions in tumor imaging may include so-called four-dimensional MR angiography, in which high-resolution three-dimensional MR angiography is combined with dynamic contrast-enhanced MR imaging.

© RSNA, 2005


   Introduction
Top
 Abstract
 Introduction
Assessment of Angiogenesis
 Conclusions
 References
 
For cells to survive and proliferate, a sufficient supply of oxygen and nutrients is needed. During the first stages of tumor growth, tumor cells obtain their nutrition and oxygen by means of diffusion. However, this is possible only over a distance of a few millimeters. It has been shown that tumors do not grow beyond 2 mm3 in size without the development of new capillaries from surrounding blood vessels, a process called angiogenesis (1). In angiogenesis, new vessels develop from preexisting ones through sprouting or intussusception rather than vasculogenesis, the de novo generation of blood vessels from endothelial precursors that occurs during embryogenesis. Angiogenesis may be observed in a physiologic setting in the uterus and in a reparative setting in myocardial ischemia and wound healing. In various pathologic conditions, such as tumor growth, synovial proliferation, and diabetic retinopathy, angiogenesis is assumed to be the basis for new vessel development. With regard to cancer, angiogenesis allows tumors to grow and metastasize. Angiogenesis is controlled by a balance of circulating proangiogenic and antiangiogenic factors that is sometimes referred to as the "angiogenic switch" (2). When there is equilibrium between pro- and antiangiogenic factors, the switch is turned off. With regard to tumors, this condition results in a so-called dormant state characterized by the absence of tumor growth. The switch is turned on when there is a surplus of proangiogenic factors, a condition that triggers new vessel formation and thus allows the tumor to grow.

Growth factors produced by the tumor, such as vascular endothelial growth factor, activate endothelial cells in vessels in the vicinity of the tumor. These activated endothelial cells produce several enzymes (proteases and collagenases) that cause the destruction of the extracellular matrix between the vessel and the tumor and allow activated endothelial cells to proliferate and migrate to the tumor. Connections then are established between the migratory endothelial cells, and the cells are structurally reorganized to form new vessels (3,4) (Fig 1). In a normal physiologic situation, the vascular bed does not long remain in a state of dynamic remodeling; with maturation, the newly formed vessels enter a quiescent state. In tumors, however, angiogenesis is continuously active, and this activity leads to a relatively high fraction of immature blood vessels. As a result, tumor vessels are structurally and functionally abnormal (5). Abnormalities in various components of the vessel wall have been described. These include changes in the hierarchy of arterioles, capillaries, and venules, as well as other structural changes that result in the hyperpermeability of tumor vessels. Tumor vessels are tortuous, vary in diameter, and tend toward excessive branching (Fig 2) and shunt formation, processes that result in a heterogeneous vascular network and that may produce both hypovascular and hypervascular regions in the same tumor.



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  		Figure 1.  Diagrams show the stages of tumor-induced angiogenesis. When the tumor reaches a diameter of 1–2 mm, its growth no longer can be sustained by the diffusion of nutrients from external vessels (top left). Growth factors (GF) are then produced by the tumor (top right) that induce external vessels to bud and form new branches (bottom left), which grow toward and into the tumor (bottom right).

 


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  		Figure 2a.  Digital photographs, obtained through a dorsal skin-fold window approximately 3 mm thick, show highly vascularized high-grade nonimmunogenic BN-175 soft-tissue sarcoma. (a) Dotted line indicates tumor circumference after 2 weeks of growth. (b) Magnified gray-scale view shows chaotic intratumoral vasculature and feeding vessels.

 


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  		Figure 2b.  Digital photographs, obtained through a dorsal skin-fold window approximately 3 mm thick, show highly vascularized high-grade nonimmunogenic BN-175 soft-tissue sarcoma. (a) Dotted line indicates tumor circumference after 2 weeks of growth. (b) Magnified gray-scale view shows chaotic intratumoral vasculature and feeding vessels.

 

   Assessment of Angiogenesis
Top
 Abstract
 Introduction
 Assessment of Angiogenesis
Conclusions
 References
 
The most widely used method for assessing tumor angiogenesis is quantification of the microvessel density with microscopy of a biopsy specimen, which is considered the standard of reference. A high correlation between the microvessel density and angiogenic factor expression, tumor growth, occurrence of distant metastasis, and prognosis has been reported (6). Surrogate markers for changes in angiogenesis have been proposed and include changes in plasma levels of vascular endothelial growth factor and in endothelial cell proliferation markers such as CD105. However, there is no definitive measure of the degree of angiogenesis (7). Several imaging methods (eg, magnetic resonance [MR] imaging, Doppler ultrasonography, computed tomography, positron emission tomography, and spectroscopy) may be used to evaluate the degree of angiogenesis indirectly by measuring a parameter or a combination of parameters such as vascular density, blood flow, blood volume, and/or vascular permeability. Given its high temporal and spatial resolution, MR imaging is well suited for use in the assessment of angiogenesis and monitoring of treatment response. Dynamic contrast material–enhanced MR imaging is currently used for this purpose, and MR angiography may be of additional value (Fig 3).



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  		Figure 3a.  Contrast-enhanced MR angiograms obtained for monitoring of treatment response in experimental (a, b) and clinical (c, d) settings. (a, b) Volumetric reconstructions of MR image data obtained with a T1-weighted 3D sequence after the administration of a blood pool contrast agent show sarcoma in a rat limb before (a) and after (b) administration of tumor necrosis factor {alpha} combined with melphalan in a closed-loop system to destroy the vascular bed. Note the considerably decreased enhancement in the tumor in b. (c, d) Arterial phase MR angiograms show tumor vessels in the ankle of a patient before (c) and after (d) systemic treatment similar to that administered in the rat tumor model. Note the enhancement of the tumor vessels in c (arrowhead), and the absence of enhancement in d despite heating of the foot in a warm bath before imaging. The treatment was considered successful.

 


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  		Figure 3b.  Contrast-enhanced MR angiograms obtained for monitoring of treatment response in experimental (a, b) and clinical (c, d) settings. (a, b) Volumetric reconstructions of MR image data obtained with a T1-weighted 3D sequence after the administration of a blood pool contrast agent show sarcoma in a rat limb before (a) and after (b) administration of tumor necrosis factor {alpha} combined with melphalan in a closed-loop system to destroy the vascular bed. Note the considerably decreased enhancement in the tumor in b. (c, d) Arterial phase MR angiograms show tumor vessels in the ankle of a patient before (c) and after (d) systemic treatment similar to that administered in the rat tumor model. Note the enhancement of the tumor vessels in c (arrowhead), and the absence of enhancement in d despite heating of the foot in a warm bath before imaging. The treatment was considered successful.

 


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  		Figure 3c.  Contrast-enhanced MR angiograms obtained for monitoring of treatment response in experimental (a, b) and clinical (c, d) settings. (a, b) Volumetric reconstructions of MR image data obtained with a T1-weighted 3D sequence after the administration of a blood pool contrast agent show sarcoma in a rat limb before (a) and after (b) administration of tumor necrosis factor {alpha} combined with melphalan in a closed-loop system to destroy the vascular bed. Note the considerably decreased enhancement in the tumor in b. (c, d) Arterial phase MR angiograms show tumor vessels in the ankle of a patient before (c) and after (d) systemic treatment similar to that administered in the rat tumor model. Note the enhancement of the tumor vessels in c (arrowhead), and the absence of enhancement in d despite heating of the foot in a warm bath before imaging. The treatment was considered successful.

 


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  		Figure 3d.  Contrast-enhanced MR angiograms obtained for monitoring of treatment response in experimental (a, b) and clinical (c, d) settings. (a, b) Volumetric reconstructions of MR image data obtained with a T1-weighted 3D sequence after the administration of a blood pool contrast agent show sarcoma in a rat limb before (a) and after (b) administration of tumor necrosis factor {alpha} combined with melphalan in a closed-loop system to destroy the vascular bed. Note the considerably decreased enhancement in the tumor in b. (c, d) Arterial phase MR angiograms show tumor vessels in the ankle of a patient before (c) and after (d) systemic treatment similar to that administered in the rat tumor model. Note the enhancement of the tumor vessels in c (arrowhead), and the absence of enhancement in d despite heating of the foot in a warm bath before imaging. The treatment was considered successful.

 
Dynamic Contrast-enhanced MR Imaging
Dynamic contrast-enhanced MR imaging can be used to assess functional abnormalities of tumors, such as the hyperpermeability of tumor vessels. Data obtained with this modality correlate well with the results of histopathologic analysis of microvessel density (8) and indicate the usefulness of dynamic contrast-enhanced MR imaging for monitoring tumor response to antivascular and antiangiogenic treatments both in experimental models and in humans (9–16). For the quantification of findings on dynamic contrast-enhanced MR images, empirical parameters such as wash-in, washout, maximal enhancement, and time to peak enhancement are used. These parameters are to some extent correlated with the presence of a malignancy (17–19).

The MR contrast media most frequently used in humans are extracellular agents, also known as small-molecular agents. Gadopentetate dimeglumine and similar small-molecular contrast media quickly equilibrate between the intravascular and interstitial spaces. Because of the high rate of their vascular extraction, even in normal vessels, small-molecular contrast agents have inherent disadvantages for use in estimating blood volume and capillary permeability exclusive of the central nervous system. To overcome this problem, several macromolecular contrast agents, also called blood pool agents, have been developed, such as gadopentetate dimeglumine–labeled albumin (20). Blood pool contrast agents have a higher molecular weight and a longer intravascular circulation time than do small-molecular agents and thus permit a longer acquisition time, with advantages for both spatial resolution and signal-to-noise ratio at vascular imaging. In addition, it is thought that macromolecular agents are better than small-molecular ones for differentiating between benign and malignant tumors (21). This difference is due to the fact that blood pool agents, because of the large size of their molecules, can permeate only pathologically leaky vessel walls, whereas small-molecular agents can also permeate normal vessel walls (Figs 4, 5). The direct clinical applicability of blood pool agents is limited, however, because of the slow rate of their glomerular filtration. Several blood pool agents with intermediate molecular sizes are currently under investigation. These include contrast agents with weak and reversible protein-binding capacities, such as gadobenate dimeglumine (Multihance; Bracco Diagnostics, Milan, Italy), as well as agents with strong protein-binding capacities, such as MS-325 (Angiomark; Epix Medical, Cambridge, Mass) (22). Blood pool agents with intermediate molecular size have a longer intravascular circulation time compared with that of small-molecular agents, without posing problems for elimination. However, because of the reversibility of the interaction, bound and unbound fractions of the contrast agent are present in the blood, which may create problems with regard to quantification.



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  		Figure 4.  Diagrams show the difference in permeability between normal vessels (left) and tumor vessels (right). Extracellular contrast agents (top row, small dots) leak into the interstitium from both normal vessels and tumor vessels because of the small size of the molecules in these agents, and this leakage causes contrast enhancement in the vessels and surrounding tissues. Blood pool contrast agents (bottom row, large dots), with a larger molecular size, remain inside normal vessels. Tumor vessel walls, which contain larger gaps than do normal vessel walls, allow blood pool contrast agents to leak into the interstitium.

 


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  		Figure 5a.  Comparison of tumor delineation and vessel enhancement between an MR image obtained with a macromolecular contrast agent, albumin-(Gd-DTPA)45 (a), and an MR image obtained with a combination of albumin-(Gd-DTPA)45 and a small-molecular agent, gadopentetate dimeglumine (Magnevist; Schering) (b). In b, massive extravasation of gadopentetate dimeglumine is visible, and the depiction of the internally fragmented tumor is improved.

 


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  		Figure 5b.  Comparison of tumor delineation and vessel enhancement between an MR image obtained with a macromolecular contrast agent, albumin-(Gd-DTPA)45 (a), and an MR image obtained with a combination of albumin-(Gd-DTPA)45 and a small-molecular agent, gadopentetate dimeglumine (Magnevist; Schering) (b). In b, massive extravasation of gadopentetate dimeglumine is visible, and the depiction of the internally fragmented tumor is improved.

 
MR Angiography
In addition to functional measurements, information about neovascularization can be obtained with high-resolution three-dimensional (3D) MR angiography. Of the three main techniques of MR angiography—the time-of-flight method based on flow-related enhancement of vessels, the phase-contrast method based on phase shifts created by flowing blood, and the contrast-enhanced method based on the T1-shortening effect of gadolinium chelates—the last is the most sensitive for assessing tumor feeding vessels and intratumoral vessels (23). MR angiograms obtained with this method, which generally depict vessels with high signal intensity because of T1 shortening, are displayed by using maximum intensity projection or 3D volume rendering to show the entire vascular tree (Fig 6). Targeted reconstructions with fewer sections may provide better depiction of the vasculature within the tumor.



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  		Figure 6a.  Tumor at 2 weeks after implantation in a rat limb. (a) High-resolution T2-weighted fat-suppressed MR image depicts only the mass. (b) Three-dimensional volume-rendered MR image, obtained with a T1-weighted sequence (field of view, 4 cm) after administration of the blood pool contrast agent albumin-(Gd-DTPA)45, shows high-signal-intensity vessels surrounding the tumor (arrow), which was artificially enhanced with postprocessing to better show its position with regard to the vasculature.

 


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  		Figure 6b.  Tumor at 2 weeks after implantation in a rat limb. (a) High-resolution T2-weighted fat-suppressed MR image depicts only the mass. (b) Three-dimensional volume-rendered MR image, obtained with a T1-weighted sequence (field of view, 4 cm) after administration of the blood pool contrast agent albumin-(Gd-DTPA)45, shows high-signal-intensity vessels surrounding the tumor (arrow), which was artificially enhanced with postprocessing to better show its position with regard to the vasculature.

 
Clinical Applications.— The diagnostic applications of MR angiography have increased considerably in the past few years. MR angiography is used in the diagnosis of peripheral vascular ischemic disease, arterial stenosis, aneurysms, and peripheral vascular malformations. In tumors, the degree of vessel tortuosity and the vessel density, diameter, and branching pattern are important indicators of malignancy (24). These characteristics not only have diagnostic value but also may be monitored to detect tumor response to antiangiogenic therapy (25).

Malignant tumors often are surrounded by a region of higher vascularity that contains dilated vessels. This is thought to be caused by the high metabolic demand of the tumor and/or the diminished resistance to blood flow in tumor vessels. With gadopentetate dimeglumine–enhanced MR angiography, a statistically significant correlation was found between malignant breast cancer and increased vascularity in the ipsilateral breast. Breasts with malignant neoplasms had greater vascularity than did the contralateral breasts (26). Although the study demonstrated an association between breast cancer and greater vascularity, the sensitivity and specificity of this isolated sign for the diagnosis of malignancy were limited. Therefore, this sign cannot be used as the sole criterion for diagnosis; it can, however, be used as an additional sign of malignancy. Increased vascularity also was found in the tissues surrounding other types of tumors: A diffuse increase in vascularity in tissues surrounding soft-tissue sarcomas in the human limb and neck was found at MR angiography by using gadopentetate dimeglumine (Figs 7, 8), and a similar increase was found in tissues surrounding soft-tissue sarcomas in a rat model by using albumin-(Gd-DTPA)45 (a formula in which 45 molecules of gadopentetate dimeglumine are bound to one molecule of albumin) (Fig 9).



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  		Figure 7a.  Increased vascularity associated with soft-tissue sarcoma in the left leg of a patient. (a–e) Sequential 3D MR angiograms obtained with gadopentetate dimeglumine show enhancement of the vasculature and the tumor at five time points from the peak arterial phase to the venous phase. Greater vessel density around the tumor borders in the affected leg is attributable to vessel recruitment by the tumor. (f) Subtraction image (unenhanced image minus late-phase contrast-enhanced image) from T1-weighted turbo spin-echo MR imaging shows marked enhancement of the tumor border and minimal enhancement of the tumor center.

 


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  		Figure 7b.  Increased vascularity associated with soft-tissue sarcoma in the left leg of a patient. (a–e) Sequential 3D MR angiograms obtained with gadopentetate dimeglumine show enhancement of the vasculature and the tumor at five time points from the peak arterial phase to the venous phase. Greater vessel density around the tumor borders in the affected leg is attributable to vessel recruitment by the tumor. (f) Subtraction image (unenhanced image minus late-phase contrast-enhanced image) from T1-weighted turbo spin-echo MR imaging shows marked enhancement of the tumor border and minimal enhancement of the tumor center.

 


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  		Figure 7c.  Increased vascularity associated with soft-tissue sarcoma in the left leg of a patient. (a–e) Sequential 3D MR angiograms obtained with gadopentetate dimeglumine show enhancement of the vasculature and the tumor at five time points from the peak arterial phase to the venous phase. Greater vessel density around the tumor borders in the affected leg is attributable to vessel recruitment by the tumor. (f) Subtraction image (unenhanced image minus late-phase contrast-enhanced image) from T1-weighted turbo spin-echo MR imaging shows marked enhancement of the tumor border and minimal enhancement of the tumor center.

 


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  		Figure 7d.  Increased vascularity associated with soft-tissue sarcoma in the left leg of a patient. (a–e) Sequential 3D MR angiograms obtained with gadopentetate dimeglumine show enhancement of the vasculature and the tumor at five time points from the peak arterial phase to the venous phase. Greater vessel density around the tumor borders in the affected leg is attributable to vessel recruitment by the tumor. (f) Subtraction image (unenhanced image minus late-phase contrast-enhanced image) from T1-weighted turbo spin-echo MR imaging shows marked enhancement of the tumor border and minimal enhancement of the tumor center.

 


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  		Figure 7e.  Increased vascularity associated with soft-tissue sarcoma in the left leg of a patient. (a–e) Sequential 3D MR angiograms obtained with gadopentetate dimeglumine show enhancement of the vasculature and the tumor at five time points from the peak arterial phase to the venous phase. Greater vessel density around the tumor borders in the affected leg is attributable to vessel recruitment by the tumor. (f) Subtraction image (unenhanced image minus late-phase contrast-enhanced image) from T1-weighted turbo spin-echo MR imaging shows marked enhancement of the tumor border and minimal enhancement of the tumor center.

 


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  		Figure 7f.  Increased vascularity associated with soft-tissue sarcoma in the left leg of a patient. (a–e) Sequential 3D MR angiograms obtained with gadopentetate dimeglumine show enhancement of the vasculature and the tumor at five time points from the peak arterial phase to the venous phase. Greater vessel density around the tumor borders in the affected leg is attributable to vessel recruitment by the tumor. (f) Subtraction image (unenhanced image minus late-phase contrast-enhanced image) from T1-weighted turbo spin-echo MR imaging shows marked enhancement of the tumor border and minimal enhancement of the tumor center.

 


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  		Figure 8a.  Increased vascularity related to soft-tissue sarcoma. (a) Arterial phase MR angiogram obtained with gadopentetate dimeglumine shows enhanced vessels that surround a large unenhanced tumor in the neck of a patient. (b) T1-weighted image, obtained after MR angiography, shows the enhanced tumor.

 


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  		Figure 8b.  Increased vascularity related to soft-tissue sarcoma. (a) Arterial phase MR angiogram obtained with gadopentetate dimeglumine shows enhanced vessels that surround a large unenhanced tumor in the neck of a patient. (b) T1-weighted image, obtained after MR angiography, shows the enhanced tumor.

 


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  		Figure 9a.  MR angiograms obtained after the administration of albumin-(Gd-DTPA)45 show sarcomas in the thigh in two different rats 2 weeks after implantation. The tumor size and contrast enhancement pattern in a differs markedly from that in b, although the same tumor model was used.

 


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  		Figure 9b.  MR angiograms obtained after the administration of albumin-(Gd-DTPA)45 show sarcomas in the thigh in two different rats 2 weeks after implantation. The tumor size and contrast enhancement pattern in a differs markedly from that in b, although the same tumor model was used.

 
MR angiography can be performed to obtain information regarding the possibilities for embolotherapy of tumors. Indications for embolotherapy include preoperative devascularization of lesions and therapeutic or palliative treatment of malignant and benign tumors, including vascular malformations and neoplasms. Embolization before surgical resection of primary and secondary masses (eg, metastatic renal cell carcinoma) has proved successful for the reduction of blood loss and of subsequent metastatic spread.

Tumors have a very inhomogeneous blood supply, and both hyper- and hypovascular regions may coexist in the same tumor (27). This variation may influence the success of intravenously administered therapy, as the therapeutic agent may not reach hypovascular regions. The supply of the therapeutic agent to some regions also may be limited because of partial clotting or destruction of tumor feeding vessels during therapy. The tumor blood supply and, therefore, therapeutic access to tumors can be assessed with MR angiography.

Knowledge of the status of tumor feeding vessels before and during therapy may also increase our understanding of the MR imaging findings. Changes in tumor enhancement may be due not only to actual tumor destruction but also to partial clotting of proximal vessels that results in a diminished supply of the contrast agent to the tumor (Fig 10). The use only of dynamic contrast-enhanced MR imaging to assess the effect of therapy may result in overestimation of the treatment effect, as the slope of the contrast enhancement–time curve for tumor tissue may be reduced if one or more tumor feeding vessels are obstructed. Tumor blood supply and, therefore, also the therapeutic access to tumors may be more accurately assessed with a combination of dynamic contrast-enhanced MR imaging and MR angiography. Different techniques have been developed to obtain so-called four-dimensional MR angiograms, which have the combined features of dynamic contrast-enhanced MR images and 3D MR angiograms. Time-resolved MR angiographic sequences have been developed for the rapid acquisition of multiple 3D volumes throughout the passage of the contrast agent bolus. The resultant high-resolution 3D angiograms provide dynamic information from both the arterial and the venous phases (28) (Fig 11). Both small-molecular and macromolecular contrast agents can be used with time-resolved sequences. Zhu et al reported the use of high-resolution 3D MR angiography to obtain dynamic angiograms with a temporal resolution of 1–2 seconds (29).



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  		Figure 10a.  (a) Arterial phase MR angiogram, obtained after administration of gadopentetate dimeglumine, shows a soft-tissue sarcoma and related vasculature. Partial blockage of a venous segment (arrow) just below the enhancing tumor may be due to clotting. An arterial blockage proximal to the tumor (obscured in this projection) also was observed. (b) Axial T2-weighted MR image provides better delineation of the tumor.

 


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  		Figure 10b.  (a) Arterial phase MR angiogram, obtained after administration of gadopentetate dimeglumine, shows a soft-tissue sarcoma and related vasculature. Partial blockage of a venous segment (arrow) just below the enhancing tumor may be due to clotting. An arterial blockage proximal to the tumor (obscured in this projection) also was observed. (b) Axial T2-weighted MR image provides better delineation of the tumor.

 


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  		Figure 11a.  MR angiograms obtained in the arm of a patient during the arterial phase (a) and the venous phase (b) illustrate the advantages of performing MR angiography at different time points after contrast material administration. Whereas a shows no tumor, b clearly depicts a tumor (arrow) in the forearm.

 


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  		Figure 11b.  MR angiograms obtained in the arm of a patient during the arterial phase (a) and the venous phase (b) illustrate the advantages of performing MR angiography at different time points after contrast material administration. Whereas a shows no tumor, b clearly depicts a tumor (arrow) in the forearm.

 
Experimental Applications.— Intratumoral vasculature in rodent models has been visualized by using high-resolution 3D MR angiography. Fink et al (30) demonstrated that differentiation between tumors with different degrees of vascularity was possible by performing MR angiography with SH L 643 A (Gadomer-17; Schering, Berlin, Germany). Intratumoral blood vessels with diameters ranging from 150 to 200 µm could be depicted with MR angiography at a spatial resolution of 210 x 170 x 320 µm. Tumor vessels were located mainly in the periphery of the tumor. Also, dilatation of subcutaneous blood vessels was observed. It was assumed that this dilatation was caused either by local growth of the tumor, with resultant vessel compression and stasis of blood, or by secretion of angiogenic factors. Kobayashi and colleagues, with the use of MR contrast agents with a polyamidoamine den-drimer core, demonstrated the ability of MR angiography to depict intratumoral vessels with a diameter of approximately 100 µm, and they proposed that MR angiography could be used for estimating the effect of antiangiogenic therapy (31,32). Schwickert et al compared the usefulness of albumin-(Gd-DTPA)30 with that of gadopentetate dimeglumine at MR imaging for visualization of peritumoral vessels in a rat model. They concluded that the use of albumin-(Gd-DTPA)30 with high-resolution MR imaging produced a time-persistent and detailed angiographic depiction of peritumoral vessels that was of higher quality than that achieved with use of gadopentetate dimeglumine (33). It was not possible in these earlier studies to perform in vivo microscopy for comparison with high-resolution MR angiography of intratumoral vessels and surrounding extratumoral vessels. Currently, however, with the development of a dorsal skin-fold window model, the combined use of in vivo microscopy and MR imaging is possible.

In the department of surgical oncology at our institution, an MR-compatible dorsal skin-fold viewing chamber was developed by using an adaptation of the procedure described by Papenfuss et al (34) (Fig 12). After the anesthetization and fixation of the animal on the stage of the microscope, the tumor vasculature can be monitored over time through the dorsal skin-fold window. With the use of optical or fluorescence intravital microscopy, the features of the microvascular architecture, including self-loops, feeding and draining vessels, bifurcations, venous convolutions, arterioles, capillaries, venules, vessel wall characteristics, tumor vessel budding and growth, blood flow, vessel dimensions, and the extravasation of agents can be assessed and monitored over time by using specific contrast agents singly or in combination. In addition, the vascular effects of therapeutic agents can be monitored in vivo.



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  		Figure 12a.  Diagrams show the position (a) and use (b) of the skin-fold window model in a rat. In b, the processes of tumor implantation (left), tumor growth (middle), and coil positioning for MR imaging (right) are shown. A flap of skin was removed and sandwiched between two frames, and the skin and frames were fixed in place with sutures. Before the surgical site was closed, a small piece of tumor (0.1 mm3) was implanted. The window was then closed with the insertion of an 18-mm-diameter layer of microscopic cover glass on both sides of the implantation site, and the tumor was allowed to grow. For high-resolution MR imaging (MR angiography), a 1- or 2-cm loop receiver antenna was placed on the window to obtain resolution of 60 x 60 x 100-µm and 40 x 40 x 100-µm interpolated voxels, respectively, with examination times of less than 20 minutes on a clinical 3.0-T MR imager. In this way, high-resolution MR angiography with use of a blood pool contrast agent can be performed over the course of several days to monitor tumor growth and angiogenesis.

 


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  		Figure 12b.  Diagrams show the position (a) and use (b) of the skin-fold window model in a rat. In b, the processes of tumor implantation (left), tumor growth (middle), and coil positioning for MR imaging (right) are shown. A flap of skin was removed and sandwiched between two frames, and the skin and frames were fixed in place with sutures. Before the surgical site was closed, a small piece of tumor (0.1 mm3) was implanted. The window was then closed with the insertion of an 18-mm-diameter layer of microscopic cover glass on both sides of the implantation site, and the tumor was allowed to grow. For high-resolution MR imaging (MR angiography), a 1- or 2-cm loop receiver antenna was placed on the window to obtain resolution of 60 x 60 x 100-µm and 40 x 40 x 100-µm interpolated voxels, respectively, with examination times of less than 20 minutes on a clinical 3.0-T MR imager. In this way, high-resolution MR angiography with use of a blood pool contrast agent can be performed over the course of several days to monitor tumor growth and angiogenesis.

 
With use of the dorsal skin-fold window model, contrast-enhanced MR images can be obtained with a resolution of 40 x 40 x 100 µm on a clinical 3.0-T MR imager (Fig 13). Tumor growth can be monitored over several days by using a blood pool agent (Fig 14). When fluorescently labeled contrast agents with various molecular sizes are administered, the combined use of fluorescence microscopy and high-resolution MR imaging can provide insight into the structural and functional characteristics of the tumor vasculature (Figs 15, 16) and can be used to assess the sensitivity of this method for detecting changes induced by therapeutic agents. The combination of the two techniques also may be used to develop and evaluate methods for quantifying findings on dynamic contrast-enhanced MR images, because direct visualization of the agent is possible with confocal microscopy.



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  		Figure 13a.  MR and optical images of a tumor in a skin-fold window model in a rat. Digital photograph (a) and comparable high-resolution MR angiogram obtained with the blood pool contrast agent albumin-(Gd-DTPA)45 (b) show tumor vasculature 2 weeks after implantation. Often, as in b, the tumor is not optimally depicted against the background on MR angiograms obtained with a blood pool agent, and gadopentetate dimeglumine is added to improve tumor delineation, as in c.

 


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  		Figure 13b.  MR and optical images of a tumor in a skin-fold window model in a rat. Digital photograph (a) and comparable high-resolution MR angiogram obtained with the blood pool contrast agent albumin-(Gd-DTPA)45 (b) show tumor vasculature 2 weeks after implantation. Often, as in b, the tumor is not optimally depicted against the background on MR angiograms obtained with a blood pool agent, and gadopentetate dimeglumine is added to improve tumor delineation, as in c.

 


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  		Figure 13c.  MR and optical images of a tumor in a skin-fold window model in a rat. Digital photograph (a) and comparable high-resolution MR angiogram obtained with the blood pool contrast agent albumin-(Gd-DTPA)45 (b) show tumor vasculature 2 weeks after implantation. Often, as in b, the tumor is not optimally depicted against the background on MR angiograms obtained with a blood pool agent, and gadopentetate dimeglumine is added to improve tumor delineation, as in c.

 


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  		Figure 14a.  Image series obtained with high-resolution MR angiography and the blood pool agent albumin-(Gd-DTPA)45 over the course of 2 weeks in the skin-fold window model in a rat. Maximum intensity projections show development of the tumor and tumor feeding vessels at day 1 (tumor diameter, approximately 3 mm) (a), day 3 (tumor diameter, approximately 7 mm) (b), and day 7 (tumor diameter, approximately 11 mm) (c).

 


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  		Figure 14b.  Image series obtained with high-resolution MR angiography and the blood pool agent albumin-(Gd-DTPA)45 over the course of 2 weeks in the skin-fold window model in a rat. Maximum intensity projections show development of the tumor and tumor feeding vessels at day 1 (tumor diameter, approximately 3 mm) (a), day 3 (tumor diameter, approximately 7 mm) (b), and day 7 (tumor diameter, approximately 11 mm) (c).

 


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  		Figure 14c.  Image series obtained with high-resolution MR angiography and the blood pool agent albumin-(Gd-DTPA)45 over the course of 2 weeks in the skin-fold window model in a rat. Maximum intensity projections show development of the tumor and tumor feeding vessels at day 1 (tumor diameter, approximately 3 mm) (a), day 3 (tumor diameter, approximately 7 mm) (b), and day 7 (tumor diameter, approximately 11 mm) (c).

 


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  		Figure 15a.  (a) High-resolution MR angiogram (maximum intensity projection) obtained with the blood pool agent albumin-(Gd-DTPA)45 and with a 1-cm loop receiver coil on a 3.0-T clinical MR imager shows a tumor 11/2 weeks after implantation in a skin-fold window model in a mouse. (b–d) Magnification of the maximum intensity projection image enables a comparison of vessels depicted on the MR angiogram (b) with those depicted on an optical photomicrograph (c) and fluorescence photomicrograph (d).

 


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  		Figure 15b.  (a) High-resolution MR angiogram (maximum intensity projection) obtained with the blood pool agent albumin-(Gd-DTPA)45 and with a 1-cm loop receiver coil on a 3.0-T clinical MR imager shows a tumor 11/2 weeks after implantation in a skin-fold window model in a mouse. (b–d) Magnification of the maximum intensity projection image enables a comparison of vessels depicted on the MR angiogram (b) with those depicted on an optical photomicrograph (c) and fluorescence photomicrograph (d).

 


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  		Figure 15c.  (a) High-resolution MR angiogram (maximum intensity projection) obtained with the blood pool agent albumin-(Gd-DTPA)45 and with a 1-cm loop receiver coil on a 3.0-T clinical MR imager shows a tumor 11/2 weeks after implantation in a skin-fold window model in a mouse. (b–d) Magnification of the maximum intensity projection image enables a comparison of vessels depicted on the MR angiogram (b) with those depicted on an optical photomicrograph (c) and fluorescence photomicrograph (d).

 


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  		Figure 15d.  (a) High-resolution MR angiogram (maximum intensity projection) obtained with the blood pool agent albumin-(Gd-DTPA)45 and with a 1-cm loop receiver coil on a 3.0-T clinical MR imager shows a tumor 11/2 weeks after implantation in a skin-fold window model in a mouse. (b–d) Magnification of the maximum intensity projection image enables a comparison of vessels depicted on the MR angiogram (b) with those depicted on an optical photomicrograph (c) and fluorescence photomicrograph (d).

 


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  		Figure 16a.  Digital photograph (a), confocal fluorescence photomicrograph (b), and volume-rendered MR angiogram (c) show similar morphologic features of tumor-related vasculature at the center (solid arrow) and periphery (open arrow) of a skin-fold window model. The close correlation between images from the different modalities indicates that noninvasive quantification of angiogenesis may be possible in the future with MR imaging.

 


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  		Figure 16b.  Digital photograph (a), confocal fluorescence photomicrograph (b), and volume-rendered MR angiogram (c) show similar morphologic features of tumor-related vasculature at the center (solid arrow) and periphery (open arrow) of a skin-fold window model. The close correlation between images from the different modalities indicates that noninvasive quantification of angiogenesis may be possible in the future with MR imaging.

 


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  		Figure 16c.  Digital photograph (a), confocal fluorescence photomicrograph (b), and volume-rendered MR angiogram (c) show similar morphologic features of tumor-related vasculature at the center (solid arrow) and periphery (open arrow) of a skin-fold window model. The close correlation between images from the different modalities indicates that noninvasive quantification of angiogenesis may be possible in the future with MR imaging.

 

   Conclusions
Top
 Abstract
 Introduction
 Assessment of Angiogenesis
 Conclusions
References
 
Tumor-related vasculature can be well assessed in vivo both macroscopically and microscopically by using high-resolution contrast-enhanced MR angiography and dynamic contrast-enhanced MR imaging. The imaging potential of these modalities may be used to expand our understanding of the pathophysiology of angiogenesis and, therefore, to increase the applicability of these technologies for the detection, treatment, and monitoring of human disease.


   Acknowledgments
 
The authors thank Dirk Verver, Karin ten Wolde, and Andries Zwamborn for the preparation of the images; Gisela Ambagtsheer, BSc, and Sandra van Tiel, BSc, for the animal handling; and Lejla Alic, MSc, and Linda Everse, PhD, for their help with the preparation of the manuscript.


   Footnotes
 

Abbreviations: 3D = three-dimensional

See the commentary by Szklaruk and Murthy following this article.


   References
Top
 Abstract
 Introduction
 Assessment of Angiogenesis
 Conclusions
 References
 

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