Chaban Ryan, McGrath Gannon, Habibabady Zahra, Rosales Ivy, Burdorf Lars, Ayares David L, Rybak Elana, Zhang Tianshu, Harris Donald G, Dahi Siamak, Ali Franchesca, Parsell Dawn M, Braileanu Gheorghe, Cheng Xiangfei, Sievert Evelyn, Phelps Carol, Azimzadeh Agnes M, Pierson Richard N
Center for Transplantation Sciences and Department of Surgery, Massachusetts General Hospital and Harvard School of Medicine, Boston, Massachusetts, USA.
Department of Cardiac and Vascular Surgery, University Hospital of Johannes Gutenberg University Mainz, Mainz, Germany.
Xenotransplantation. 2023 Jul-Aug;30(4):e12812. doi: 10.1111/xen.12812. Epub 2023 Jul 28.
Expression of human complement pathway regulatory proteins (hCPRP's) such as CD46 or CD55 has been associated with improved survival of pig organ xenografts in multiple different models. Here we evaluate the hypothesis that an increased human CD46 gene dose, through homozygosity or additional expression of a second hCPRP, is associated with increased protein expression and with improved protection from injury when GTKO lung xenografts are perfused with human blood.
Twenty three GTKO lungs heterozygous for human CD46 (GTKO.heteroCD46), 10 lungs homozygous for hCD46 (GTKO.homoCD46), and six GTKO.homoCD46 lungs also heterozygous for hCD55 (GTKO.homoCD46.hCD55) were perfused with human blood for up to 4 h in an ex vivo circuit.
Relative to GTKO.heteroCD46 (152 min, range 5-240; 6/23 surviving at 4 h), survival was significantly improved for GTKO.homoCD46 (>240 min, range 45-240, p = .034; 7/10 surviving at 4 h) or GTKO.homoCD46.hCD55 lungs (>240 min, p = .001; 6/6 surviving at 4 h). Homozygosity was associated with increased capillary expression of hCD46 (p < .0001). Increased hCD46 expression was associated with significantly prolonged lung survival (p = .048),) but surprisingly not with reduction in measured complement factor C3a. Hematocrit, monocyte count, and pulmonary vascular resistance were not significantly altered in association with increased hCD46 gene dose or protein expression.
Genetic engineering approaches designed to augment hCPRP activity - increasing the expression of hCD46 through homozygosity or co-expressing hCD55 with hCD46 - were associated with prolonged GTKO lung xenograft survival. Increased expression of hCD46 was associated with reduced coagulation cascade activation, but did not further reduce complement activation relative to lungs with relatively low CD46 expression. We conclude that coagulation pathway dysregulation contributes to injury in GTKO pig lung xenografts perfused with human blood, and that the survival advantage for lungs with increased hCPRP expression is likely attributable to improved endothelial thromboregulation.
在多种不同模型中，人类补体途径调节蛋白（hCPRP's）如CD46或CD55的表达与猪器官异种移植的存活改善相关。在此，我们评估这样一种假设，即通过纯合性或第二种hCPRP的额外表达使人类CD46基因剂量增加，与蛋白表达增加以及当GTKO肺异种移植物用人类血液灌注时对损伤的保护改善相关。
23个携带人类CD46杂合基因的GTKO肺（GTKO.杂合CD46）、10个hCD46纯合的肺（GTKO.homoCD46）以及6个同时携带hCD55杂合基因的GTKO.homoCD46肺（GTKO.homoCD46.hCD55）在体外循环中用人类血液灌注长达4小时。
相对于GTKO.杂合CD46（152分钟，范围5 - 240分钟；4小时时6/23存活），GTKO.homoCD46（>240分钟，范围45 - 240分钟，p = 0.034；4小时时7/10存活）或GTKO.homoCD46.hCD55肺（>240分钟，p = 0.001；4小时时6/6存活）的存活显著改善。纯合性与hCD46的毛细血管表达增加相关（p < 0.0001）。hCD46表达增加与肺存活显著延长相关（p = 0.048），但令人惊讶的是与测量的补体因子C3a的降低无关。血细胞比容、单核细胞计数和肺血管阻力与hCD46基因剂量或蛋白表达增加无显著改变。
旨在增强hCPRP活性的基因工程方法——通过纯合性增加hCD46的表达或与hCD46共表达hCD55——与GTKO肺异种移植存活延长相关。hCD46表达增加与凝血级联激活减少相关，但相对于CD46表达相对较低的肺，并未进一步降低补体激活。我们得出结论，凝血途径失调导致用人类血液灌注的GTKO猪肺异种移植损伤，并且hCPRP表达增加的肺的存活优势可能归因于改善的内皮血栓调节。
