Development and long-term in vivo evaluation of a biodegradable urethane-doped polyester elastomer.

We have recently reported upon the development of crosslinked urethane-doped polyester (CUPE) network elastomers, which was motivated by the desire to overcome the drawbacks presented by crosslinked network polyesters and biodegradable polyurethanes for soft tissue engineering applications. Although the effect of the isocyanate content and post-polymerization conditions on the material structure-property relationship was examined in detail, the ability of the diol component to modulate the material properties was only studied briefly. Herein, we present a detailed report on the development of CUPE polymers synthesized using diols 4, 6, 8, 10, or 12 methylene units in length in order to investigate what role the diol component plays on the resulting material's physical properties, and assess their long-term biological performance in vivo. An increase in the diol length was shown to affect the physical properties of the CUPE polymers primarily through lowered polymeric crosslinking densities and elevated material hydrophobicity. The use of longer chain diols resulted in CUPE polymers with increased molecular weights resulting in higher tensile strength and elasticity, while also increasing the material hydrophobicity to lower bulk swelling and prolong the polymer degradation rates. Although the number of methylene units largely affected the physical properties of CUPE, the choice of diol did not affect the overall polymer cell/tissue-compatibility both in vitro and in vivo. In conclusion, we have established the diol component as an important parameter in controlling the structure-property relationship of the polymer in addition to diisocyanate concentration and post-polymerization conditions. Expanding the family of CUPE polymers increases the choices of biodegradable elastomers for tissue engineering applications.

Crosslinked urethane doped polyester biphasic scaffolds: Potential for in vivo vascular tissue engineering.

In vivo tissue engineering uses the body as a bioreactor for tissue regeneration, thus placing stringent requirements on tissue scaffolds, which should be mechanically robust for immediate implantation without a long in vitro cell culture time. In addition to mechanical strength, vascular grafts fabricated for in vivo tissue engineering approach must have matching mechanical properties to the target tissues to avoid compliance mismatch, which is one of the reasons for graft failure. We recently synthesized a new generation of strong and elastic biodegradable crosslinked urethane-doped polyesters (CUPE) to address the challenge of developing soft, elastic yet strong biodegradable polymers. This study evaluated the tensile strength, burst pressure, and suture retention of CUPE biphasic scaffolds to determine if the scaffolds met the requirements for immediate implantation in an in vivo tissue engineering approach. In addition, we also examined the hemocompatibility and inflammatory potential of CUPE to demonstrate its potential in serving as a blood-contacting vascular graft material. Tensile strength of CUPE biphasic scaffolds (5.02 ± 0.70 MPa) was greater than native vessels (1.43 ± 0.60 MPa). CUPE scaffolds exhibited tunable burst pressure ranging from 1500 mmHg to 2600 mmHg, and adequate suture retention values (2.45 ± 0.23 N). CUPE showed comparable leukocyte activation and whole blood clotting kinetics to poly(L-lactic acid) PLLA. However, CUPE incited a lesser release of inflammatory cytokines and was found to be non hemolytic. Combined with the mechanical properties and previously demonstrated anti-thrombogenic nature, CUPE may serve as a viable graft material for in vivo blood vessel tissue engineering.

Novel polymeric scaffolds using protein microbubbles as porogen and growth factor carriers.

Polymeric tissue engineering scaffolds prepared by conventional techniques like salt leaching and phase separation are greatly limited by their poor biomolecule-delivery abilities. Conventional methods of incorporation of various growth factors, proteins, and/or peptides on or in scaffold materials via different crosslinking and conjugation techniques are often tedious and may affect scaffold's physical, chemical, and mechanical properties. To overcome such deficiencies, a novel two-step porous scaffold fabrication procedure has been created in which bovine serum albumin microbubbles (henceforth MB) were used as porogen and growth factor carriers. Polymer solution mixed with MB was phase separated and then lyophilized to create porous scaffold. MB scaffold triggered substantially lesser inflammatory responses than salt-leached and conventional phase-separated scaffolds in vivo. Most importantly, the same technique was used to produce insulin-like growth factor-1 (IGF-1)-eluting porous scaffolds, simply by incorporating IGF-1-loaded MB (MB-IGF-1) with polymer solution before phase separation. In vitro such MB-IGF-1 scaffolds were able to promote cell growth to a much greater extent than scaffold soaked in IGF-1, confirming the bioactivity of the released IGF-1. Further, such MB-IGF-1 scaffolds elicited IGF-1-specific collagen production in the surrounding tissue in vivo. This novel growth factor-eluting scaffold fabrication procedure can be used to deliver a range of single or combination of bioactive biomolecules to substantially promote cell growth and function in degradable scaffold.

Development of aliphatic biodegradable photoluminescent polymers.

None of the current biodegradable polymers can function as both implant materials and fluorescent imaging probes. The objective of this study was to develop aliphatic biodegradable photoluminescent polymers (BPLPs) and their associated cross-linked variants (CBPLPs) for biomedical applications. BPLPs are degradable oligomers synthesized from biocompatible monomers including citric acid, aliphatic diols, and various amino acids via a convenient and cost-effective polycondensation reaction. BPLPs can be further cross-linked into elastomeric cross-linked polymers, CBPLPs. We have shown representatively that BPLP-cysteine (BPLP-Cys) and BPLP-serine (BPLP-Ser) offer advantages over the traditional fluorescent organic dyes and quantum dots because of their preliminarily demonstrated cytocompatibility in vitro, minimal chronic inflammatory responses in vivo, controlled degradability and high quantum yields (up to 62.33%), tunable fluorescence emission (up to 725 nm), and photostability. The tensile strength of CBPLP-Cys film ranged from 3.25 +/- 0.13 MPa to 6.5 +/- 0.8 MPa and the initial Modulus was in a range of 3.34 +/- 0.15 MPa to 7.02 +/- 1.40 MPa. Elastic CBPLP-Cys could be elongated up to 240 +/- 36%. The compressive modulus of BPLP-Cys (0.6) (1:1:0.6 OD:CA:Cys) porous scaffold was 39.60 +/- 5.90 KPa confirming the soft nature of the scaffolds. BPLPs also possess great processability for micro/nano-fabrication. We demonstrate the feasibility of using BPLP-Ser nanoparticles ("biodegradable quantum dots") for in vitro cellular labeling and noninvasive in vivo imaging of tissue engineering scaffolds. The development of BPLPs and CBPLPs represents a new direction in developing fluorescent biomaterials and could impact tissue engineering, drug delivery, bioimaging.

Method to analyze three-dimensional cell distribution and infiltration in degradable scaffolds.

Effective cell seeding throughout the tissue scaffold often determines the success of tissue-engineering products, although most current methods focus on determining the total number, not the distribution, of the cells associated with tissue-engineering constructs. The purpose of this investigation was to establish a quick, convenient, and efficient method to quantify cell survival, distribution, and infiltration into degradable scaffolds using a combination of fluorescence cell staining and cryosectioning techniques. After cell seeding and culture for different periods of time, seeded scaffolds were stained with a live cell dye and then cryosectioned. Cryosectioned scaffolds were then recompiled into a three-dimensional (3D) image to visualize cell behavior after seeding. To test the effectiveness of this imaging method, four common seeding methods, including static surface seeding, cell injection, orbital shaker seeding, and centrifuge seeding, were investigated for their seeding efficacy. Using this new method, we were able to visualize the benefits and drawbacks of each seeding method with regard to the cell behavior in 3D within the scaffolds. This method is likely to provide useful information to assist the development of novel materials or cell-seeding methods for producing full-thickness tissue grafts.

Development of biodegradable crosslinked urethane-doped polyester elastomers.

Traditional crosslinked polyester elastomers are inherently weak, and the strategy of increasing crosslink density to improve their mechanical properties makes them brittle materials. Biodegradable polyurethanes, although strong and elastic, do not fare well in dynamic environments due to the onset of permanent deformation. The design and development of a soft, strong and completely elastic (100% recovery from deformation) material for tissue engineering still remains a challenge. Herein, we report the synthesis and evaluation of a new class of biodegradable elastomers, crosslinked urethane-doped polyesters (CUPEs), which is able to satisfy the need for soft, strong, and elastic biomaterials. Tensile strength of CUPE was as high as 41.07+/-6.85 MPa with corresponding elongation at break of 222.66+/-27.84%. The initial modulus ranged from 4.14+/-1.71 MPa to 38.35+/-4.5 MPa. Mechanical properties and degradation rates of CUPE could be controlled by varying the choice of diol used for synthesis, the polymerization conditions, as well as the concentration of urethane bonds in the polymer. The polymers demonstrated good in vitro and in vivo biocompatibilities. Preliminary hemocompatibility evaluation indicated that CUPE adhered and activated lesser number of platelets compared to PLLA. Good mechanical properties and easy processability make these materials well suited for soft tissue engineering applications. The introduction of CUPEs provides new avenues to meet the versatile requirements of tissue engineering and other biomedical applications.