Tissue Engineering of in vivo vascularized soft tissue free flaps in a large animal model

Status:

completed 12/2022

Aims:

The arteriovenous loop (AVL) technique enables the in vivo generating of axially vascularized connective tissue flaps as an alternative to conventional free flaps. However, AVL soft-tissue flaps have never been investigated in standardized large animal models for their suitability in reconstruction of critical defects as a free flap.

The main objective of this study was to generate axially vascularized connective tissue flap grafts in clinically relevant dimensions in a large animal model, which can be autotransplanted and used for free defect reconstruction.

The secondary objective of this study was to examine the vascularization kinetics and connective tissue maturation process of AVL flap grafts.

The secondary objective of this study was to investigate genes and MicroRNA (miRNAs) that regulate AVL-associated angiogenesis.

Activities/Methods:

Each two AVLs (n=12) were created in 6 sheep and embedded in a bovine collagen/elastin scaffold within a subcutaneous polytetrafluoroethylene chamber. The tissue volume, extent of angiogenesis, and proportion of proliferating cells were examined on postoperative day (POD) 28 using immunohistochemistry (ki67, CD31, Factor VIII) and micro-computed tomography (pCT). Additionally, the biomechanical properties of the AVL flaps were examined using dynamic mechanical analysis. Four AVL flaps were microsurgically anastomosed to the neck vessels in a standardized defect model as a free flap. Postoperative defect closure and flap graft perfusion were examined using angiography and histology. The gene expression of angiogenesis-associated genes was examined using qRT-PCR from venous grafts of the AVL.

Results:

On POD 28, the AVL flaps completely filled the isolation chamber and showed histologically and in pCT homogeneous microvascular networks. The mean number of microvessels and vessel volume, as well as the percentage of proliferating cells, increased significantly over time. Dynamic mechanical analysis showed a greater stiffness of the AVL flap compared to fasciocutaneous flaps from patients. QRT-PCR showed analogous regulation of angiogenesis-associated gene expression compared to previous studies on patients who underwent two-stage defect reconstruction with a free flap anastomosed to an AVL. In the defect model, stable defect closure with homogeneous tissue integration into the surrounding tissue was clinically, angiographically, and histologically observed 10 days after transplantation in all flaps.

Conclusion and clinical relevance: The AVL flap graft is suitable for free transplantation and defect reconstruction with minimal donor site morbidity and can potentially be applied as a surgical.

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