Toward a patient-specific tissue engineered vascular graft

Abstract

Integrating three-dimensional printing with the creation of tissue-engineered vascular grafts could provide a readily available, patient-specific, autologous tissue source that could significantly improve outcomes in newborns with congenital heart disease. Here, we present the recent case of a candidate for our tissue-engineered vascular graft clinical trial deemed ineligible due to complex anatomical requirements and consider the application of three-dimensional printing technologies for a patient-specific graft. We 3D-printed a closed-disposable seeding device and validated that it performed equivalently to the traditional open seeding technique using ovine bone marrow-derived mononuclear cells. Next, our candidate's preoperative imaging was reviewed to propose a patient-specific graft. A seeding apparatus was then designed to accommodate the custom graft and 3D-printed on a commodity fused deposition modeler. This exploratory feasibility study represents an important proof of concept advancing progress toward a rationally designed patient-specific tissue-engineered vascular graft for clinical application.

Keywords: 3D-printing; Fontan operation; Tissue-engineered vascular graft; cell seeding; patient-specific modeling.

Figure 1.

Flowchart of the current clinical TEVG fabrication protocol. Boxed items indicate process components, diamonds highlight a critical process decision, and circles indicate release testing. Briefly, 5 mL/kg whole bone marrow is aspirated from the iliac crests of the Fontan patient prior to operation. After aspiration, the patient is prepped for Fontan completion and dissection is begun. The bone marrow is delivered to an ISO Class 7 Clean room where mononuclear cells are enriched via density gradient centrifugation. Samples for in-process testing are collected and include cell viability and CD45 flow cytometry. The seeding team waits for a call from the operating room, which communicates the appropriate graft diameter based on intraoperative conduit sizing. The graft is selected and seeded via an operator independent vacuum method. Samples are collected for release testing and include Gram staining of a graft sample, dsDNA assay, and seeding efficiency. Sterility cultures are also collected for post-process monitoring. If all tests pass the criteria, then the seeded TEVG scaffold is delivered to the operating room for implantation.

Figure 2.

(a) Rendering of 3D-printed closed, disposable seeding system for preparation of currently utilized TEVG scaffolds. (b) Photographs of 3D-printed prototype utilized in this report. (c) 20× photomicrograph of seeded scaffold section stained with DAPI visualized with polarized transmitted (top) and fluorescent reflected light (middle) to identify birefringent scaffold polymer and seeded cell nuclei, respectively. The merged image (bottom) reveals seeded BM-MNCs embedded within the porous scaffold wall and confirms dsDNA assay results.

Figure 3.

Preoperative CMR reconstructions of a candidate Fontan TEVG patient in the (a) coronal plane viewed from the (a) anterior-posterior and (b) posterior–anterior perspectives. The patient presented with heterotaxy syndrome, asplenia, associated dextrocardia, right dominant atrioventricular canal defect with a severely hypoplastic left ventricle, double outlet right ventricle with malposed great arteries with subpulmonary stenosis, and total anomalous pulmonary venous return (TAPVR) to the superior vena cava status post TAPVR repair and bidirectional Glenn shunting. CAD was used to propose a patient-specific TEVG conduit, which was superimposed on the CMR reconstructions. The proposed Fontan conduit is highlighted in green and viewed in the coronal plane from the (c) anterior–posterior and (d) posterior–anterior perspectives.

Figure 4.

(a) Design of patient-specific TEVG conduit demonstrating a compound curve and lumen diameter changing 3.0 mm from the proximal to distal anastomoses. (b) 3D renderings of (a). (c) A custom fenestrated seeding mandrel was reverse-engineered from the proposed scaffold. (d) Intraoperative photograph from the proposed TEVG candidate after implantation of a standard-of-care Gore-Tex conduit. Note that the linear conduit adopted a nonlinear shape in response to patient anatomy, which closely resembles the geometry of the proposed patient-specific TEVG scaffold.

Figure 5.

A custom patient-specific seeding apparatus was designed to accommodate the custom seeding mandrel. (a) Rendering of 3D-printed patient-specific closed-disposable seeding system. (b) Photograph of 3D-printed apparatus fabricated as a proof of concept.

Figure 6.

Incorporation of fluid dynamic and mechanobiologic computational modeling with additive manufacturing would permit a revised clinical protocol for design and preparation of the Fontan TEVG. Boxed items indicate process components, diamonds highlight a critical process decision, and circles indicate “go/no go” testing. Briefly, preoperative CMR would inform computational models to predict the optimal scaffold design. This design would be used to create a custom scaffold and 3D-printed seeding device. Bone marrow would be aspirated prior to start of the Fontan operation and a closed, disposable system for bone marrow mononuclear cell enrichment would be connected to the closed custom seeding apparatus. Vacuum seeding would be completed and samples collected for release testing prior to being implanted as a patient-specific Fontan conduit.