Project Name

Primary authors: Robert Orr

Motivation:

Tissue Engineering is not a novel field; however, the applicability of designed scaffolds is limited.  Constraints on the engineered product exist due to poor nutrient supply for seeded cells and adjacent native cells.  Initial nutrients cultured on the scaffold before implantation endows some support to cells, but diffusion confines the distance of nutrient exchange—approximately 100-200μm.  Host angiogenesis occurs independent of any specific signal provided by the implanted tissue, although passive hypoxia influences host response. 

This mechanism is slow, with the proliferation rate equaling tenths of micrometers per day.  Preliminary nutrients will be exhausted before complete vascularization; therefore, an ideal system is one that immediately stimulates vascular propagation and accelerates growth.  These problems are obvious when examining current engineered tissues—they are either avascular or thin constructs where the host vascularization mechanism is sufficient.  Such modest requirements complement skin and cartilage, but novel designs for bone and muscle require a vascular component.

Vascularization In Tissue Engineering:

Methods for stimulating vascularization in implanted tissue exist, these strategies are:  physical properties of the scaffold, supplying angiogenic factors, and initiating vascularization before final implantation.  These techniques do assist in creating vascular networks; however, variance in effectiveness of these methods is ubiquitous.  Simply stimulating the host response results in a marginal decrease of time for complete tissue vascularization, whereas prevascularization creates a instant vascularized tissue upon implantation, assuming correct procedure.  Figure 1 illustrates the aformentioned methods for instigating vascularization in engineered scaffolds. 

"Figure 1. Different strategies for improving vascularization in tissue engineering. (a) Scaffold design. Panel (i) shows a scaffold that was prepared with compression

molding and salt leaching. The scaffold in panel (ii) was obtained by 3D fiber deposition. Panels (iii) and (iv) schematically illustrate the scaffold geometries of (i) and (ii),

respectively. Note that in the irregular scaffold (iii), some pores (depicted in red) cannot be reached from the outside, so vascular ingrowth will be prevented in these pores.

Partly adapted, with permission, from Ref. [51]. (b) Growth factor delivery. Fibrin gel matrices were placed on a chicken chorioallantoic membrane (a membrane of the

chicken egg). Panel (i) shows the effects of freely diffusible VEGF121, which resulted in the formation of vessels with a disturbed morphology. Many of the newly formed

vessel branches were characterized by malformed, corkscrew-like structures (indicated by the arrowheads). Furthermore, many of those branches appeared to abruptly

drain into zones of irregular capillary enlargement and growth (indicated by the arrows). In panel (ii), VEGF121 was released enzymatically by MMPs in a cell-demanded

release. Note that upon cell-demanded release, a more regular organization of the vascular structures can be observed. Adapted, with permission, from Ref. [32]. (c) In vivo

prevascularization. An artery (A) and a vein (V) were joined via a loop, which was then placed around a bone-tissue-engineered scaffold and implanted. Panel (i) shows the

construct before implantation with plastic tubing instead of the arteriovenous loop for illustration. Panel (ii) shows the highly vascularized construct that was obtained eight

weeks after implantation. Panel (iii) schematically depicts in vivo prevascularization. 1: Tissue construct preparation in vitro. 2: Implantation at the prevascularization site,

supplied by a vascular axis. 3: Formation of a microvascular network by vessel ingrowth from the vascular axis. 4: Explantation of the prevascularized construct with the

vascular axis. 5: Implantation of the construct at the defect site and surgical connection of the vascular axis to the vasculature. Partly adapted, with permission, from Ref.

[34]. (d) In vitro prevascularization. Mouse myoblast cells (C2C12) were combined with human umbilical vein endothelial cells (HUVECs) and mouse embryonic fibroblasts

(MEFs) and seeded on a scaffold, resulting in the formation of a 3D prevascular network. After implantation, the network anastomosed to the mouse vasculature. The

prevascular network that is formed in vitro is shown in (i). This picture shows a cross section of the scaffold after in vitro culture in which endothelial cells are stained brown

and muscle cells are stained blue. Note the presence of cross sections of tubular structures in brown, which shows that the endothelial cells have organized into vascular

structures. The anastomosis of the prevascular network after implantation is illustrated in (ii), which shows a cross section of the scaffold after implantation. The vascular

network that was formed in vitro is stained in green and all vessels that were perfused with blood at the time of explantation are stained red. The double staining

demonstrates that the preformed vessels connected to the host vasculature and were perfused with blood. Panel (iii) schematically depicts in vitro vascularization. 1: A

tissue construct containing endothelial cells is prepared in vitro. 2: The endothelial cells organize into a vascular network (blue). 3: The tissue construct is implanted and

host vessels (red) grow into the construct. 4: When the host vessels reach the precultured vascular network, the vessels connect and the entire construct becomes perfused."

Future Strategies

A 3-4 paragraph summary describing future strategies that are being used to treat the disease or disorder (this should include the technology that you are researching). Be sure to emphasize the strengths of future strategies. Provide cost estimates of these treatment strategies if you can find this information.

Citations

Rouwkema, Jeroen. et al. "Vascularization in Tissue Engineering."  Trends in Biotechnology, Vol. 26, No.8