The ideal engineered vascular graft would utilize human-derived materials to minimize foreign body response and tissue rejection. Current biological engineered blood vessels (BEBVs) inherently lack the structure required for implantation. We hypothesized that an ECM material would provide the structure needed. Skin dermis ECM is commonly used in reconstructive surgeries, is commercially available and FDA-approved. We evaluated the commercially-available decellularized skin dermis ECM Alloderm for efficacy in providing structure to BEBVs. Alloderm was incorporated into our lab’s unique protocol for generating BEBVs, using fibroblasts to establish the adventitia. To assess structure, tissue mechanics were analyzed. Standard BEBVs without Alloderm exhibited a tensile strength of 67.9 ± 9.78 kPa, whereas Alloderm integrated BEBVs showed a significant increase in strength to 1500 ± 334 kPa. In comparison, native vessel strength is 1430 ± 604 kPa. Burst pressure reached 51.3 ± 2.19 mmHg. Total collagen and fiber maturity were significantly increased due to the presence of the Alloderm material. Vessels cultured for 4 weeks maintained mechanical and structural integrity. Low probability of thrombogenicity was confirmed with a negative platelet adhesion test. Vessels were able to be endothelialized. These results demonstrate the success of Alloderm to provide structure to BEBVs in an effective way.
Introduction: Of the three layers of a mammalian artery, the tunica adventitia provides the most strength to prevent rupture of blood vessels under high pressures. The structural network of this outermost layer mainly consists of fibroblasts and collagen. Many groups have improved strength of their tissue engineered (TE) vessels by conditioning with pulsatile flow in bioreactors. Conditioning stimulates cellular collagen production; however, it is time-consuming ranging from 3-8 weeks. Dermis is comprised primarily of collagen providing skin with considerable strength. Hypothesis: We hypothesized inclusion of decellularized dermis, Alloderm, into TE vessels can increase the strength and thus decrease production time. Methods: To generate a TE adventitia vessel, we adapted our lab’s ring stacking method which creates tissue rings that are stacked to form a tubular, vascular constructs. Human dermal fibroblasts (HDFs) were used to form a cellular fibrin hydrogel in a petri dished with a central post. Alloderm was cut into a donut shape and placed on top of the hydrogel around the central post (Figure 1). Additional HDFs were placed on top of the hydrogel and Alloderm. Over 14 days the HDF monolayer rolled around the post into a ring. These Alloderm rings were stacked and adhered using fibrin glue to create a TE vessel. Results: TE vessel tensile strength was significantly higher with Alloderm than without: 1500 ± 262 kPa (n = 5) versus 67.9 ± 9.78 kPa (n = 5). Histology indicated Alloderm significantly increased total collagen content and fiber maturity compared to TE vessels without. Mechanical integrity of Alloderm rings was maintained for 4 weeks: tensile strength 2760 ± 1130 kPa. In comparison, native adventitia strength is 1430 ± 604 kPa. Conclusions: This work shows that decellularized dermis can successfully be incorporated into TE vessels. Additionally, inclusion of Alloderm significantly increases tissue strength and collagen content that is maintained over time.
Tissue engineering has the advantage of replicating soft tissue mechanics to better simulate and integrate into native soft tissue. However, soft tissue engineering has been fraught with issues of insufficient tissue strength to withstand physiological mechanical requirements. This factor is due to the lack of strength inherent in cell-only constructs and in the biomaterials used for soft tissue engineering and limited extracellular matrix (ECM) production possible in cell culture. To address this issue, we explored the use of an ECM-based hydrogel coating to serve as an adhesive tool, as demonstrated in vascular tissue engineering. The efficacy of cells to supplement mechanical strength in the coating was explored. Specifically, selected coatings were applied to an engineered artery tunica adventitia to accurately test their properties in a natural tissue support structure. Multiple iterations of three primary hydrogels with and without cells were tested: fibrin, collagen, and gelatin hydrogels with and without fibroblasts. The effectiveness of a natural crosslinker to further stabilize and strengthen the hydrogels was investigated, namely genipin extracted from the gardenia fruit. We found that gelatin crosslinked with genipin alone exhibited the highest tensile strength; however, fibrin gel supported cell viability the most. Overall, fibrin gel coating without genipin was deemed optimal for its balance in increasing mechanical strength while still supporting cell viability and was used in the final mechanical and hydrodynamic testing assessments. Engineered vessels coated in fibrin hydrogel with cells resulted in the highest tensile strength of all hydrogel-coated groups after 14 d in culture, demonstrating a tensile strength of 11.9 ± 2.91 kPa, compared to 5.67 ± 1.37 kPa for the next highest collagen hydrogel group. The effect of the fibrin hydrogel coating on burst pressure was tested on our strongest vessels composed of human aortic smooth muscle cells. A significant increase from our previously reported burst pressure of 51.3 ± 2.19 mmHg to 229 ± 23.8 mmHg was observed; however, more work is needed to render these vessels compliant with mechanical and biological criteria for blood vessel substitutes.
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