Automatic quantitative micro-computed tomography evaluation of angiogenesis in an axially vascularized tissue-engineered bone construct.
BACKGROUND
We invented an automatic observer-independent quantitative method to analyze vascularization using micro-computed tomography (CT) along with three-dimensional (3D) reconstruction in a tissue engineering model.


METHODS
An arteriovenous loop was created in the medial thigh of 30 rats and was placed in a particulated porous hydroxyapatite and beta-tricalcium phosphate matrix, filled with fibrin (10 mg/mL fibrinogen and 2 IU/mL thrombin) without (group A) or with (group B) application of fibrin-gel-immobilized angiogenetic growth factors vascular endothelial growth factor (VEGF¹⁶⁵) and basic fibroblast growth factor (bFGF). The explantation intervals were 2, 4, and 8 weeks. Specimens were investigated by means of micro-CT followed by an automatic 3D analysis, which was correlated to histomorphometrical findings.


RESULTS
In both groups, the arteriovenous loop led to generation of dense vascularized connective tissue with differentiated and functional vessels inside the matrix. Quantitative analysis of vascularization using micro-CT showed to be superior to histological analysis. The micro-CT analysis also allows the assessment of different other, more complex vascularization parameters within 3D constructs, demonstrating an early improvement of vascularization by application of fibrin-gel-immobilized VEGF¹⁶⁵ and bFGF.


CONCLUSIONS
In this study quantitative analysis of vascularization using micro-CT along with 3D reconstruction and automatic analysis exhibit to be a powerful method superior to histological evaluation of cross sections.
Arkudas
Andreas
A
Department of Plastic and Hand Surgery, University of Erlangen Medical Center, Erlangen, Germany.
Beier
Justus Patrick
JP
Pryymachuk
Galyna
G
Hoereth
Tobias
T
Bleiziffer
Oliver
O
Polykandriotis
Elias
E
Hess
Andreas
A
Gulle
Heinz
H
Horch
Raymund E
RE
Kneser
Ulrich
U
D000818:Animals; D001331:Automation; D001807:Blood Vessel Prosthesis; D001808:Blood Vessels / Q000502:physiology; D001861:Bone Regeneration / Q000502:physiology; D001842:Bone and Bones / Q000098:blood supply* / Q000000981:diagnostic imaging* / Q000502:physiology*; D016764:Cell Polarity; D048091:Guided Tissue Regeneration / Q000379:methods*; D008297:Male; D018919:Neovascularization, Physiologic / Q000502:physiology*; D051381:Rats; D011917:Rats, Inbred Lew; D023822:Tissue Engineering / Q000379:methods; D054457:Tissue Scaffolds / Q000737:chemistry; D055114:X-Ray Microtomography / Q000379:methods*