High-Resolution X-Ray Techniques as New Tool to Investigate the 3D Vascularization of Engineered-Bone Tissue.

Inna Bukreeva,Maddalena Mastrogiacomo,Raffaele Spanò,Marco Grilli,Manfred Burghammer,Gaetano Campi,Daniele Pelliccia,Alessia Cedola,Giuliana Tromba,Michela Fratini,Francesco Brun,Ranieri Cancedda

doi:10.3389/fbioe.2015.00133

Abstract

The understanding of structure–function relationships in normal and pathologic mammalian tissues is at the basis of a tissue engineering (TE) approach for the development of biological substitutes to restore or improve tissue function. In this framework, it is interesting to investigate engineered bone tissue, formed when porous ceramic constructs are loaded with bone marrow stromal cells (BMSC) and implanted in vivo. To monitor the relation between bone formation and vascularization, it is important to achieve a detailed imaging and a quantitative description of the complete three-dimensional vascular network in such constructs. Here, we used synchrotron X-ray phase-contrast micro-tomography to visualize and analyze the three-dimensional micro-vascular networks in bone-engineered constructs, in an ectopic bone formation mouse-model. We compared samples seeded and not seeded with BMSC, as well as samples differently stained or unstained. Thanks to the high quality of the images, we investigated the 3D distribution of both vessels and collagen matrix and we obtained quantitative information for all different samples. We propose our approach as a tool for quantitative studies of angiogenesis in TE and for any pre-clinical investigation where a quantitative analysis of the vascular network is required.

Highlights

Tissue Engineering (TE) is the biotechnology that combines aspects of medicine, biology, and engineering to generate, repair, or replace human tissues
In sample A, no bone marrow stromal cells (BMSC) were seeded on the scaffold, whereas the scaffold of sample B was seeded with BMSC
Sample C was seeded with BMSC, but, after its recovery from the animal, it was stained with phosphotungstic acid (PTA)

Summary

Introduction

Tissue Engineering (TE) is the biotechnology that combines aspects of medicine, biology, and engineering to generate, repair, or replace human tissues. The TE approach may be used to regenerate bone by implanting a porous ceramic scaffold combined with bone marrow stromal cells (BMSC) in vivo. The scaffold plays a fundamental role since it acts as a guide and it Vascularization of engineered bone tissue stimulates the growth creating TE constructs or living biocomposites (Hench and Polak, 2002; Cancedda et al, 2003). The efficiency of an artificially implanted construct depends on the timely delivery and exchange of nutrients (oxygen, glucose, amino acids, etc.) from surrounding blood vessels to the BMSC, and the contemporary removal of the metabolism waste products (CO2, lactate, and urea) (Carano and Filvaroff, 2003; Jain, 2003). A deeper understanding of the developmental neo-vascularization is necessary for a better treatment of many pathological conditions, including cancer, diabetes, psoriasis, and articular degeneration

Objectives

Methods

Results

Conclusion