title: Comparing Quantitative Immunohistochemical Markers of Angiogenesis to Contrast-Enhanced Subharmonic Imaging.
abstract: OBJECTIVE
  Different methods for obtaining tumor neovascularity parameters based on immunohistochemical markers were compared to contrast-enhanced subharmonic imaging (SHI).
  
  
  METHODS
  Eighty-five athymic nude female rats were implanted with 5 × 10(6) breast cancer cells (MDA-MB-231) in the mammary fat pad. The contrast agent Definity (Lantheus Medical Imaging, North Billerica, MA) was injected, and SHI was performed using a modified Sonix RP scanner (Analogic Ultrasound, Richmond, British Columbia, Canada) with a L9-4 linear array (transmitting/receiving frequencies, 8/4 MHz). Afterward, specimens were stained for endothelial cells (CD31), vascular endothelial growth factor (VEGF), and cyclooxygenase 2 (COX-2). Tumor neovascularity was assessed in 4 different ways using a histomorphometry system (×100 magnification: (1) over the entire tumor; (2) in small sub-regions of interest (ROIs); (3) in the tumor periphery and centrally; and (4) in 3 regions of maximum marker expression (so-called hot spots). Results from specimens and from SHI were compared by linear regression.
  
  
  RESULTS
  Fifty-four rats (64%) showed tumor growth, and 38 were successfully imaged. Subharmonic imaging depicted the tortuous morphologic characteristics of tumor neovessels and delineated small areas of necrosis. The immunohistochemical markers did not correlate with SHI measures over the entire tumor area or over small sub-ROIs (P > .18). However, when the specimens were subdivided into central and peripheral regions, COX-2 and VEGF correlated with SHI in the periphery (r = -0.42; P = .005; and r = -0.32; P = .049, respectively).
  
  
  CONCLUSIONS
  When comparing quantitative contrast measures of tumor neovascularity to immunohistochemical markers of angiogenesis in xenograft models, ROIs corresponding to the biologically active region should be used to account for tumor heterogeneity.
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA, School of Biomedical Engineering, Sciences, and Health Systems, Drexel University, Philadelphia, Pennsylvania USA.
authors.forename: Aditi
authors.initials: A
authors.lastname: Gupta
authors.affiliation: Yale University, New Haven, Connecticut USA.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Forsberg
authors.affiliation: University of Pittsburgh, Pittsburgh, Pennsylvania USA.
authors.forename: Kelly
authors.initials: K
authors.lastname: Dulin
authors.affiliation: University of Pittsburgh, Pittsburgh, Pennsylvania USA.
authors.forename: Samantha
authors.initials: S
authors.lastname: Jaffe
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA.
authors.forename: Jaydev K
authors.initials: JK
authors.lastname: Dave
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA, School of Biomedical Engineering, Sciences, and Health Systems, Drexel University, Philadelphia, Pennsylvania USA.
authors.forename: Valgerdur G
authors.initials: VG
authors.lastname: Halldorsdottir
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA, School of Biomedical Engineering, Sciences, and Health Systems, Drexel University, Philadelphia, Pennsylvania USA.
authors.forename: Andrew
authors.initials: A
authors.lastname: Marshall
authors.affiliation: Plymouth-Whitemarsh High School, Plymouth Meeting, Pennsylvania USA.
authors.forename: Anya I
authors.initials: AI
authors.lastname: Forsberg
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA.
authors.forename: John R
authors.initials: JR
authors.lastname: Eisenbrey
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA.
authors.forename: Priscilla
authors.initials: P
authors.lastname: Machado
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA, Department of Radiologic Sciences, College of Health Professions, Thomas Jefferson University, Philadelphia, Pennsylvania USA.
authors.forename: Traci B
authors.initials: TB
authors.lastname: Fox
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA.
authors.forename: Ji-Bin
authors.initials: JB
authors.lastname: Liu
authors.affiliation: Department of Radiology, Thomas Jefferson University, Philadelphia, Pennsylvania USA flemming.forsberg@jefferson.edu.
authors.forename: Flemming
authors.initials: F
authors.lastname: Forsberg
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D001943:Breast Neoplasms / Q000098:blood supply* / Q000000981:diagnostic imaging / Q000378:metabolism; D003287:Contrast Media; D051546:Cyclooxygenase 2 / Q000378:metabolism; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000473:pathology; D005260:Female; D007089:Image Enhancement / Q000379:methods*; D008819:Mice, Nude; D009389:Neovascularization, Pathologic / Q000000981:diagnostic imaging* / Q000378:metabolism*; D051381:Rats; D014463:Ultrasonography; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
chemical_list: D015415:Biomarkers; D003287:Contrast Media; D042461:Vascular Endothelial Growth Factor A; D051546:Cyclooxygenase 2
keywords: breast cancer; immunohistochemistry; murine xenografts; subharmonic imaging