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

Bukreeva I, Fratini M, Campi G, Pelliccia D, Spanò R, Tromba G, Brun F, Burghammer M, Grilli M, Cancedda R, Cedola A, Mastrogiacomo M - Front Bioeng Biotechnol (2015)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Consiglio Nazionale delle Ricerche - Istituto NANOTEC, c/o Dipartimento di Fisica, Università Sapienza , Rome , Italy.

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.

No MeSH data available.


Related in: MedlinePlus

(A) Table of quantitative information obtained for the A, B, C, and D samples. Vm and VM are the total numbers of small and large vessels, respectively (see text). (B) Vascularization factor (see text) for the four samples.
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Figure 4: (A) Table of quantitative information obtained for the A, B, C, and D samples. Vm and VM are the total numbers of small and large vessels, respectively (see text). (B) Vascularization factor (see text) for the four samples.

Mentions: To get higher level of detail on how vessels of different size are spatially distributed, we divided the vessels into two categories: small vessels with a diameter between 10 and 15 μm and large vessels with a diameter between 15 and 20 μm. Plot 2 reports the number Vm of small vessels (represented in gray) and the number VM of large vessels (represented in black) at different depths inside the samples. Clearly, in the whole space occupied by the vascular trees of B, C, and D samples, small vessels were by far more numerous and a significant branching occurred. This analysis emphasizes even more strikingly the difference with respect to the A sample, where only one large vessel was present, which intersected the sample, crossing it for about 1 mm, and abruptly disappearing. Few small vessels occupied the first part of the sample, from 400 to 800 μm depth, and then disappeared outside this interval. The table shown in Figure 4A extends and summarizes the quantitative comparison of vessels in the four samples. The individual structure of the vessels was similar in the three samples because the maximal sections and the minimal diameters were comparable. Again the major difference laid in the average number of branches forming the vascular trees. The A sample was poorly ramified, while B, C, and D samples displayed thriving trees with many branches. Finally the overall flow rate of the vascular networks for the four samples, represented by the vascularization factor (VF) reported in the last line of the Table in Figure 4A and visualized in Figure 4B, showed a major difference between sample A and the other three samples. Assuming that for each vessel the blood flow is proportional to its section, this quantity is calculated as the integral of small vessels weighted by their average section (~123 μm2) plus the integral of large vessels weighted by their average section (~240 μm2) (the sum is then normalized by the total depth of 1400 μm).


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

Bukreeva I, Fratini M, Campi G, Pelliccia D, Spanò R, Tromba G, Brun F, Burghammer M, Grilli M, Cancedda R, Cedola A, Mastrogiacomo M - Front Bioeng Biotechnol (2015)

(A) Table of quantitative information obtained for the A, B, C, and D samples. Vm and VM are the total numbers of small and large vessels, respectively (see text). (B) Vascularization factor (see text) for the four samples.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4561513&req=5

Figure 4: (A) Table of quantitative information obtained for the A, B, C, and D samples. Vm and VM are the total numbers of small and large vessels, respectively (see text). (B) Vascularization factor (see text) for the four samples.
Mentions: To get higher level of detail on how vessels of different size are spatially distributed, we divided the vessels into two categories: small vessels with a diameter between 10 and 15 μm and large vessels with a diameter between 15 and 20 μm. Plot 2 reports the number Vm of small vessels (represented in gray) and the number VM of large vessels (represented in black) at different depths inside the samples. Clearly, in the whole space occupied by the vascular trees of B, C, and D samples, small vessels were by far more numerous and a significant branching occurred. This analysis emphasizes even more strikingly the difference with respect to the A sample, where only one large vessel was present, which intersected the sample, crossing it for about 1 mm, and abruptly disappearing. Few small vessels occupied the first part of the sample, from 400 to 800 μm depth, and then disappeared outside this interval. The table shown in Figure 4A extends and summarizes the quantitative comparison of vessels in the four samples. The individual structure of the vessels was similar in the three samples because the maximal sections and the minimal diameters were comparable. Again the major difference laid in the average number of branches forming the vascular trees. The A sample was poorly ramified, while B, C, and D samples displayed thriving trees with many branches. Finally the overall flow rate of the vascular networks for the four samples, represented by the vascularization factor (VF) reported in the last line of the Table in Figure 4A and visualized in Figure 4B, showed a major difference between sample A and the other three samples. Assuming that for each vessel the blood flow is proportional to its section, this quantity is calculated as the integral of small vessels weighted by their average section (~123 μm2) plus the integral of large vessels weighted by their average section (~240 μm2) (the sum is then normalized by the total depth of 1400 μm).

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Consiglio Nazionale delle Ricerche - Istituto NANOTEC, c/o Dipartimento di Fisica, Università Sapienza , Rome , Italy.

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.

No MeSH data available.


Related in: MedlinePlus