Comparing subharmonic imaging and immunohistochemical markers of angiogenesis

Gupta, Aditi; Forsberg, Mark A.; Dulin, Kelly; Jaffe, Samantha; Dave, Jaydev K.; Dahibawkar, Manasi; Halldorsdottir, Valgerdur G.; Marshall, Andrew; Forsberg, Anya I.; Machado, Priscilla; Fox, Traci B.; Liu, Ji‐Bin; Forsberg, Flemming

doi:10.1109/ultsym.2014.0284

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“…The relative expression values obtained for all 3 immunohistochemical markers (ie, VEGF, CD31, and COX-2) were then organized into marker-specific pathologic maps. Since the nature of tumor vasculature is heterogeneous, 1 tumor neovascularity was assessed in 4 different ways as follows 26 : 1. Over the entire tumor-In this case, the average relative expression over the entire pathological map of the tumor was calculated.…”

Section: Discussionmentioning

confidence: 99%

“…Images obtained after subtraction were subjected to 3 × 3 open and sharpen filters for enhancement 24 . The red‐green‐blue channels were then used to count the number of stained pixels ( s i ) in each image, and the ratio to the total number of pixels ( s i + x i ), representing the percent stained area or relative expression ( RE ), was calculated according to the following formula 24 – 26 :

R E = \frac{\sum s_{i}}{\sum s_{i} + x_{i}} .

…”

Section: Methodsmentioning

confidence: 99%

“…Since the nature of tumor vasculature is heterogeneous, 1 tumor neovascularity was assessed in 4 different ways as follows 26 : Over the entire tumor—In this case, the average relative expression over the entire pathological map of the tumor was calculated. In small sub‐ROIs—In this case, the pathologic map of the tumor was divided into smaller ROIs (3–4 mm) based on the size of the tumor, and the average relative expression for each ROI was calculated. This process was done to account for the heterogeneity of the tumor and examine different regions of the tumor separately 25 …”

Section: Methodsmentioning

confidence: 99%

“…Data Analysis A semiautomated histomorphometry system was used to obtain and analyze digital images of each histologic slide. [24][25][26] A DXC-970MD color charge-coupled device camera (Sony Corporation, Tokyo, Japan) was used in conjunction with a Labophot-2 microscope (Nikon, Melville, NJ) set to a ×10 ocular magnification with a ×10 objective (total magnification ×100). Between 20 and 130 images were acquired to encompass the entire tumor.…”

Section: Pathologic Data Acquisitionmentioning

confidence: 99%

“…Since the nature of tumor vasculature is heterogeneous, 1 tumor neovascularity was assessed in 4 different ways as follows 26 : 1. Over the entire tumor-In this case, the average relative expression over the entire pathological map of the tumor was calculated.…”

Section: Pathologic Data Acquisitionmentioning

confidence: 99%

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Comparing Quantitative Immunohistochemical Markers of Angiogenesis to Contrast‐Enhanced Subharmonic Imaging

Gupta

Forsberg

Dulin

et al. 2016

J of Ultrasound Medicine

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ngiogenesis is the growth of new blood vessels and is a fundamental physiologic process for reproduction, development, and wound repair. 1,2 Under normal circumstances, both the proangiogenic and antiangiogenic factors of normal cells strike a perfect balance for controlled angiogenesis. Abnormal rapid proliferation of blood vessels (so-called pathologic angiogenesis)Aditi Gupta, MS, Mark A. Forsberg, BS, Kelly Dulin, BS, Samantha Jaffe, BS, Jaydev K. Dave, PhD, Valgerdur G. Halldorsdottir, PhD, Andrew Marshall, MS, Anya I. Forsberg, John R. Eisenbrey, PhD, Priscilla Machado, MD, Traci B. Fox, EdD, Ji-Bin Liu, MD, Flemming Forsberg, PhD Received May 11, 2015, Electrical and Electronics Engineers;:1156-1159 Gupta A, Dulin K, Jaffe S, et al. Prog RSNA 2014; abstract PHS13616; and Gupta A, Jaffe S, Forsberg MA, et al. J Ultrasound Med 2014; 33(suppl) ORIGINAL RESEARCHObjectives-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.

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Section: Discussionmentioning

confidence: 99%