Covalent Surface Heparinization Potentiates Porous Polyurethane Scaffold Vascularization

Abstract

Porous scaffolds play an integral role in many tissue-engineering approaches, and the ability to improve vascularization, without eliciting an excessive inflammatory response, would constitute an important step towards achieving long-term healing and function of devices made from these materials. After having previously optimized the dimensional requirements of the well-defined pores, the present study aimed at a further shift from inflammation to vascularization via surface immobilization with heparin. Porous polyurethane disks were produced to contain well-defined pores (147 +/- 2 microm) with abundant interconnecting windows (67 +/- 2 microm). After heparinization via copolymer grafting and amination to contain 32 microg of heparin, the modification appeared as a uniform layer on all exposed surfaces, with no signs of pore obliteration or significant changes in pore size. After 28 days implantation in a rat subcutaneous model, the scaffolds were assessed for vascularization/arteriolization and inflammation using CD31/actin and ED-1 staining, respectively. Heparinization resulted in a significant increase in vascularization: capillaries increased by 62% in number (66.2 +/- 0.8 to 107.3 +/- 1.4 vessels/mm( 2); p < 0.03) and 56% in total area (0.9 +/- 0.1 to 1.4 +/- 0.02%; p<0.02). Arteriolization also increased in absolute terms (200% in number; p<0.03), but did not change significantly when normalized to capillary number. Heparinization did not significantly affect the inflammatory response at this time-point, as quantified by ED-1 positive macrophage and foreign body giant cell (FBGC) content. Thus, the in vivo vascularization of porous scaffolds could be increased without concomitant increase in the inflammatory response by employing a simple surface modification technique. This could be a valuable tool for in vivo tissue engineering applications where enhanced vascularization is required.

Fast-degrading elastomer enables rapid remodeling of a cell-free synthetic graft into a neoartery

“…Heparin may also promote cellular infiltration by binding cell adhesion proteins such as PECAM-1 and L-selectin, adhesive matrix proteins, and chemokines 38 . Previous work demonstrated that heparinization improves implant cellularity and angiogenesis 39 . Heparin can also promote remodeling by binding remodeling factors such as tissue plasminogen activator 38 .…”

The performance of cross-linked acellular arterial scaffolds as vascular grafts; pre-clinical testing in direct and isolation loop circulatory models

“…The heparin content of ETVGs (n=3) was quantitatively determined by 3-methyl-2-benzothiazoninone hydrazone (MBTH) assay adapted from Risenfeld and Roden [18] similar to that described before by Bezuidenhout et al [19]. Heparin content (mg/g tissue) was calculated from standard curves.…”

Induced chronic hypoxia negates the pro‐angiogenic effect of surface immobilized heparin in a polyurethane porous scaffold

“…To evaluate ingrowth of granulation tissue including neo‐angiogenesis into the pores of the PU discs, a subcutaneous implantation model was utilized 4. The anaesthesia and surgical procedure for implantation of three round, flexible PU discs was as follows:…”

“…It is generally accepted today that enhancing vascularization of matrices and scaffolds is of utmost importance for the survival and proper function of such constructs 2, 3. We recently demonstrated that angiogenesis into a porous polyurethane (PU) scaffold could be significantly increased by heparin surface modification 4. Vascular endothelial growth factor (VEGF), a major long‐term angiogenic stimulus5 has been reported to further increase heparin augmented angiogenesis 6, 7.…”

“…The current scaffolds have the advantage that they contain well‐defined, interconnecting pores that allow for unobstructed tissue ingrowth 26. Covalent coupling of heparin to the porous surface was found to be inherently pro‐angiogenic 4. The aim of this study was to evaluate, whether chronic whole body hypobaric hypoxia would influence angiogenesis into a heparin surface modified PU in a wound environment.…”