A Porcine Heterotopic Heart Transplantation Protocol for Delivery of Therapeutics to a Cardiac Allograft

Abstract

Cardiac transplantation is the gold standard treatment for end-stage heart failure. However, it remains limited by the number of available donor hearts and complications such as primary graft dysfunction and graft rejection. The recent clinical use of an ex vivo perfusion device in cardiac transplantation introduces a unique opportunity for treating cardiac allografts with therapeutic interventions to improve function and avoid deleterious recipient responses. Establishing a translational, large-animal model for therapeutic delivery to the entire allograft is essential for testing novel therapeutic approaches in cardiac transplantation. The porcine, heterotopic heart transplantation model in the intraabdominal position serves as an excellent model for assessing the effects of novel interventions and the immunopathology of graft rejection. This model additionally offers long-term survival for the pig, given that the graft is not required to maintain the recipient's circulation. The aim of this protocol is to provide a reproducible and robust approach for achieving ex vivo delivery of a therapeutic to the entire cardiac allograft prior to transplantation and provide technical details to perform a survival heterotopic transplant of the ex vivo perfused heart.

Figure 1:. Porcine heterotopic heart model in the intra-abdominal position.

Diagram of the heterotopic heart model where the allograft is transplanted in the intra-abdominal position while the recipient’s native heart remains in its natural location. The pulmonary artery of the allograft is anastomosed to the infra-renal inferior vena cava while the aorta of the allograft is anastomosed to the infra-renal aorta of the recipient.

Figure 2:. Protocol schematic for therapeutic delivery to an entire cardiac allograft using normothermic ex vivo sanguinous perfusion.

(A) The heart and blood are procured from the donor pig. (B) The blood is washed using a cell saver device to remove any therapeutic neutralizing components from the donor serum. (C) The cardiac allograft is mounted onto the normothermic ex vivo perfusion device and perfused for 2 h. (D) Soon after the allograft is mounted, the therapeutic of interest is added to the perfusate. (E) After the allotted ex vivo perfusion period, the allograft is transplanted into the recipient pig in the intra-abdominal, heterotopic position. This figure has been modified from .

Figure 3:. Cardiac allograft on ex vivo perfusion device.

The cardiac allograft mounted on a normothermic, ex vivo perfusion device where it is perfused with therapeutic-infused perfusate for 2 h prior to implantation into the recipient.

Figure 4:. Representative ex vivo perfusion parameters.

(A) Circulatory flow rates measured from the pulmonary artery (blue), the aorta (green), and the coronary arteries (red). (B) Representative aortic pressure measurements: mean pressure (blue), systolic pressure (red), diastolic pressure (green). (C) Heart rate of a cardiac allograft during ex vivo perfusion. (D) Recorded temperature of the cardiac allograft during ex vivo perfusion. (E) demonstrates the values of mixed venous oxygen saturation (SvO₂) measured from the perfusate during the perfusion period. (F) Hematocrit (hct) values measured from the perfusate during the perfusion period.

Figure 5:. Cardiac allograft transplanted in the recipient.

A cardiac allograft on post-operative day 35 that was treated with therapeutic at the time of implantation. The donor was selected to be a perfect SLA match with the recipient. Abbreviation: SLA = Swine Leukocyte Antigen.

Figure 6:. Luciferase activity after transduction of cardiac allografts.

Presented are the results of three cardiac allografts that were transduced with adenoviral vectors carrying a luciferase transgene. Demonstrated is the average fold-change in luciferase protein activity in each area of the cardiac allograft. This figure has been modified from Bishawi et al.