Angiogenesis: noninvasive quantitative assessment with contrast-enhanced functional US in murine model.
OBJECTIVE
To evaluate quantitative functional ultrasonography (US) in a murine gel model by using microbubble destruction kinetics to determine whether parametric indices provided with US could help assess angiogenesis.


METHODS
Institutional Animal Subjects Committee approved experiments and procedures. In 36 normal mice, two 0.4-mL gel implants were placed subcutaneously on either side of spine. One implant contained 0.5, 1.0, or 1.5 microg human basic fibroblast growth factor (bFGF) per milliliter of gel. Functional US quantitative analysis of angiogenesis with microbubble contrast agent was performed on days 3, 6, 9, and 12; histologic data were collected. Time-intensity curve of implant was fitted to mathematic decay model to calculate fractional blood volume and fraction of blood replaced per unit of time. Microvascular density (MVD) and percentage of microvascular area (MVA) were measured after anti-CD31 staining. Spearman rank order correlation was used in analyses.


RESULTS
bFGF-containing implants induced MVD of eight, 35, 42, and 42 vessels per square millimeter on days 3, 6, 9, and 12, respectively; in controls, MVD was four vessels/mm2 (P<.05 on days 6, 9, and 12). bFGF-containing implants induced percentage MVA of 2%, 5%, 20%, and 27%, respectively; in controls, it was 0.5% (P<.05). Maximum enhancement was significantly increased in bFGF implants (23.3 gray level+/-14.1 [standard deviation]) compared with controls (11.0+/-5.5, P<.001). Implants containing bFGF showed poor correlations between fractional blood volume and MVD (r2=0.42) or percentage MVA (r2=0.51) at US. There was no correlation between microbubble velocity and MVD (r2<0.05) or percentage MVA (r2<0.13).


CONCLUSIONS
Functional US perfusion parameters do not correlate with current histologic indices for quantifying angiogenesis. MVD, as a histologic quantitative measurement of angiogenesis, may not be an appropriate standard for contrast-enhanced imaging that relies on perfused neovessels.
Lucidarme
Olivier
O
Department of Radiology, University of California, San Diego, CA 92103, USA.
Kono
Yuko
Y
Corbeil
Jacqueline
J
Choi
Sang-Hee
SH
Golmard
Jean-Louis
JL
Varner
Judith
J
Mattrey
Robert F
RF
D000818:Animals; D001810:Blood Volume / Q000187:drug effects; D003287:Contrast Media; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D005466:Fluorocarbons; D005782:Gels; D006801:Humans; D007089:Image Enhancement / Q000379:methods; D051379:Mice; D045423:Microbubbles; D008833:Microcirculation / Q000187:drug effects; D023421:Models, Animal; D018919:Neovascularization, Physiologic / Q000502:physiology*; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000032:analysis; D019736:Prostheses and Implants; D011897:Random Allocation; D012867:Skin / Q000098:blood supply / Q000000981:diagnostic imaging; D013997:Time Factors; D014463:Ultrasonography
D003287:Contrast Media; D005466:Fluorocarbons; D005782:Gels; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D016222:Fibroblast Growth Factor 2; C078626:perflexane