TY - JOUR

T1 - Automatic quantification of angiogenesis in 2D sections

T2 - a precise and timesaving approach

AU - Weis, C

AU - Covi, J M

AU - Hilgert, J G

AU - Leibig, N

AU - Arkudas, A

AU - Horch, R E

AU - Kneser, U

AU - Schmidt, V J

N1 - © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

PY - 2015/9

Y1 - 2015/9

N2 - INTRODUCTION: The standardized characterization of angiogenesis is crucial in the field of tissue engineering as sufficient blood supply is the limiting factor of mass transfer. However, reliable algorithms that provide a straight forward and observer-independent assessment of new vessel formation are still lacking. We propose an automatic observer-independent quantitative method (including downloadable source code) to analyze vascularization using two-dimensional microscopic images of histological cross-sections and advanced postprocessing, based on a 'positive- and negative-experts' model and a (corrected) nearest neighbour classification, in a vascularized tissue engineering model.MATERIALS AND METHODS: An established angioinductive rat arteriovenous loop model was used to compare the new automatic analysis with a common 2D method and a μCT algorithm. Angiogenesis was observed at three different time points (5, 10 and 15 days).RESULTS: In line with previous results, formation of functional new vessels that arose from the venous graft was evident within the three-dimensional construct and a significant (p < 0.05) increase in vessel count and area was observed over time. The proposed automatic analysis obtained precise values for vessel count and vessel area that were similar to the manually gained data. The algorithm further provided vectorized parameterization of the newly formed vessels for advanced statistical analysis. Compared to the μCT-based three-dimensional analyses, the presented two-dimensional algorithm was superior in terms of small vessel detection as well as cost and time efficiency.CONCLUSIONS: The quantitative evaluation method, using microscopic images of stained histological sections, 'positive- and negative-experts'-based vessel segmentation, and nearest neighbour classification, provides a user-independent and precise but also time- and cost-effective tool for the analysis of vascularized constructs. Our algorithm, which is freely available to the public, outperforms previous approaches especially in terms of unambiguous vessel classification and statistical analyses.

AB - INTRODUCTION: The standardized characterization of angiogenesis is crucial in the field of tissue engineering as sufficient blood supply is the limiting factor of mass transfer. However, reliable algorithms that provide a straight forward and observer-independent assessment of new vessel formation are still lacking. We propose an automatic observer-independent quantitative method (including downloadable source code) to analyze vascularization using two-dimensional microscopic images of histological cross-sections and advanced postprocessing, based on a 'positive- and negative-experts' model and a (corrected) nearest neighbour classification, in a vascularized tissue engineering model.MATERIALS AND METHODS: An established angioinductive rat arteriovenous loop model was used to compare the new automatic analysis with a common 2D method and a μCT algorithm. Angiogenesis was observed at three different time points (5, 10 and 15 days).RESULTS: In line with previous results, formation of functional new vessels that arose from the venous graft was evident within the three-dimensional construct and a significant (p < 0.05) increase in vessel count and area was observed over time. The proposed automatic analysis obtained precise values for vessel count and vessel area that were similar to the manually gained data. The algorithm further provided vectorized parameterization of the newly formed vessels for advanced statistical analysis. Compared to the μCT-based three-dimensional analyses, the presented two-dimensional algorithm was superior in terms of small vessel detection as well as cost and time efficiency.CONCLUSIONS: The quantitative evaluation method, using microscopic images of stained histological sections, 'positive- and negative-experts'-based vessel segmentation, and nearest neighbour classification, provides a user-independent and precise but also time- and cost-effective tool for the analysis of vascularized constructs. Our algorithm, which is freely available to the public, outperforms previous approaches especially in terms of unambiguous vessel classification and statistical analyses.

KW - Algorithms

KW - Animals

KW - Automation

KW - Blood Vessels/anatomy & histology

KW - Humans

KW - Models, Theoretical

KW - Neovascularization, Physiologic

KW - Rats

KW - Tissue Engineering/methods

U2 - 10.1111/jmi.12252

DO - 10.1111/jmi.12252

M3 - Journal article

C2 - 25882279

VL - 259

SP - 185

EP - 196

JO - Journal of Microscopy

JF - Journal of Microscopy

SN - 0022-2720

IS - 3

ER -