Abstract
Efforts for tissue engineering vascular grafts focuses on the tunica media and intima, although the tunica adventitia serves as the primary structural support for blood vessels. In surgery, during endarterectomies, surgeons can strip the vessel, leaving the adventitia as the main strength layer to close the vessel. Here, we adapted our recently developed technique of forming vascular tissue rings then stacking the rings into a tubular structure, to accommodate human fibroblasts to create adventitia vessels in 8 days. Collagen production and fibril cross-linking was augmented with TGF-β and ascorbic acid, significantly increasing tensile strength to 57.8 ± 3.07 kPa (p = 0.008). Collagen type I gel was added to the base fibrin hydrogel to further increase strength. Groups were: Fibrin only; 0.7 mg/ml COL; 1.7 mg/ml COL; and 2.2 mg/ml COL. The 0.7 mg/ml collagen rings resulted in the highest tensile strength at 77.0 ± 18.1 kPa (p = 0.015). Culture periods of 1–2 weeks resulted in an increase in extracellular matrix deposition and significantly higher failure strength but not ultimate tensile strength. Histological analysis showed the 0.7 mg/ml COL group had significantly more, mature collagen. Thus, a hydrogel of 0.7 mg/ml collagen in fibrin was ideal for creating and strengthening engineered adventitia vessels.
Highlights
There have been a limited number of engineered vascular grafts utilizing fibroblast cells
We found that the fibroblast rings improved strength with supplementation with collagen production stimulating factors
Growth factor stimulation is a common tool used in the tissue engineering domain, we discovered that the growth factor stimulation approach for inducing collagen production in fibroblast cells was not alone sufficient to induce adequate collagen to create a robust engineered adventitia
Summary
There have been a limited number of engineered vascular grafts utilizing fibroblast cells. L’Heureux et al created autologous human dermal fibroblast cell sheets and wrapped them around a mandrel to create a tube structure[11] To strengthen their adventitia vessel, a 10 week maturation period under perfusion flow in a bioreactor was required. We modified the RSM to facilitate formation of human dermal fibroblasts rings and stacks Base parameters such as cell seeding number, plate size and post dimensions were re-optimized to accommodate fibroblast cell size and growth rates. We hypothesized that the addition of type I collagen in a fibrin matrix would increase the strength of engineered adventitia rings and enhance collagen deposition compared to growth factor stimulation alone. Collagen gel was effective in strengthening the rings, and allowed us to achieve significantly higher strength in the engineered adventitia rings and vessels. In trialing Fibrin only; 0.7 mg/ml COL; 1.7 mg/ml COL; and 2.2 mg/ml COL gel hydrogels, the 0.7 mg/ml collagen gel rings were the most robust, consistently formed, strongest rings with the highest collagen content and most mature collagen
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