Influencing the interaction between human blood and porcine endothelial cells using gene knockouts and human transgenes in the context of pulmonary xenotransplantation.

Background The limited availability of human organs and the lack of artificial alternatives for transplantation in end-stage lung diseases promotes the research on xenogenic organ transplantation. Although great progress has been made for heart and kidney xenotransplantation in recent years, there are still some difficulties for other organs, such as the lung. In 2022, Burdorf et al. published a survival of porcine lungs in primates of up to one month [1], compared to 499 days for kidneys [2] and 945 days for non-live-supporting hearts [3] in primates.

The aim of the research project is to identify gene knockouts and human transgenes in donor pigs that have a positive impact on tolerance in porcine-to-human lung xenotransplantation. Therefore, an in vitro model will be established to analyze the contact between immortalized porcine lung endothelial cells (EC) and human blood components.

Methods & Materials In order to obtain porcine ECs from porcine lungs, different isolation protocols were tested and compared, allowing the isolation of ECs from pulmonary arteries or veins, lung tissue sections and the pulmonary microvasculature.

Results By perfusing larger vessels and the microvasculature, ECs with different microscopic appearance were identified. Different antibody combinations were used to further characterize the ECs by flow cytometry: CD31 is used to confirm the endothelial phenotype, while different sets of surface markers are used to describe the phenotypic origin of the cells (e.g. EphrinB2 or GJA5 for capillary ECs).

Conclusion Various subtypes of ECs have been obtained by isolating ECs from different vessels. These subtypes can be distinguished by morphological characteristics as well as by different sets of cell surface markers. The immortalized cells generated for this project will now be compared to the primary cells to ensure that the specific endothelial phenotype is maintained.

• Literatur

• 1 Burdorf L. et al. Pig-to-baboon lung xenotransplantation: Extended survival with targeted genetic modifications and pharmacologic treatments. Am J Transplant 2022; 1: 28-45
• 2 Kim SC. et al Long-term survival of pig-to-rhesus macaque renal xenografts is dependent on CD4 T cell depletion. Am J Transplant 2019; 19: 2174-2185
• 3 Mohiuddin MM. et al. Chimeric 2C10R4 anti-CD40 antibody therapy is critical for long-term survival of GTKO.hCD46.hTBM pig-to-primate cardiac xenograft. Nature Communications 2016; 7: 1-10

Publikationsverlauf

Artikel online veröffentlicht:
13. August 2024