给予供体氯膦酸盐和 DDAVP 治疗后，hEPCR.hTBM.hCD47.hHO-1 可改善经人血灌注的 GalTKO.hCD46 猪肝脏的灌注和功能。
hEPCR.hTBM.hCD47.hHO-1 with donor clodronate and DDAVP treatment improves perfusion and function of GalTKO.hCD46 porcine livers perfused with human blood.
机构信息
Department of Surgery, University of Maryland School of Medicine, Baltimore, Maryland, USA.
Revivicor, Inc., Blacksburg, Virginia, USA.
出版信息
Xenotransplantation. 2022 Mar;29(2):e12731. doi: 10.1111/xen.12731. Epub 2022 Feb 15.
INTRODUCTION
Platelet sequestration, inflammation, and inappropriate coagulation cascade activation are prominent in liver xenotransplant models and are associated with poor outcomes. Here, we evaluate a cassette of six additional genetic modifications to reduce anti-pig antibody binding (α-1,3-galactosyl transferase knockout [GalTKO]) and target coagulation dysregulation (human endothelial protein C receptor [hEPRC] and thrombomodulin [hTBM]), complement pathway regulation (human membrane cofactor protein, hCD46), inflammation heme oxygenase 1 [hHO-1]), and a self-recognition receptor (integrin-associated protein [hCD47]), as well as donor pharmacologic treatments designed to blunt these phenomena.
METHODS
Livers from GaltKO.hCD46 pigs ("2-gene," n = 3) and GalTKO.hCD46 pigs also transgenic for hEPRC, hTBM, hCD47, and hHO-1 ("6-gene," n = 4) were perfused ex vivo with whole human blood. Six-gene pigs were additionally pretreated with desmopressin (DDAVP) and clodronate liposomes to deplete vWF and kupffer cells, respectively.
RESULTS
The average perfusion times increased from 304 (±148) min in the 2-gene group to 856 (±61) min in the 6-gene group (p = .010). The average heparin administration was decreased from 8837 U/h in the 2-gene to 1354 U/h in the 6-gene group (p = .047). Platelet sequestration tended to be delayed in the 6-gene group (p = .070), while thromboxane B2 (TXB2, a platelet activation marker) levels were lower over the first hour (p = .044) (401 ± 124 vs. 2048 ± 712 at 60 min). Thrombin production as measured by F1+2 levels tended to be lower in the 6-gene group (p = .058).
CONCLUSIONS
The combination of the hEPCR.hTBM.hCD47.hHO-1 cassette along with donor pig DDAVP and clodronate liposome pretreatment was associated with prolonged function of xenoperfused livers, reduced coagulation pathway perturbations, and decreased TXB2 elaboration, and reflects significant progress to modulate liver xenograft injury in a pig to human model.
简介
血小板隔离、炎症和不适当的凝血级联激活在肝异种移植模型中很明显，与不良结局相关。在这里，我们评估了一组六个额外的基因修饰，以减少抗猪抗体结合（α-1,3-半乳糖基转移酶敲除[GalTKO]）和靶向凝血失调（人内皮蛋白 C 受体[hEPRC]和血栓调节蛋白[hTBM]）、补体途径调节（人膜辅因子蛋白，hCD46）、炎症血红素加氧酶 1 [hHO-1]）和自我识别受体（整合素相关蛋白[hCD47]），以及旨在减轻这些现象的供体药物治疗。
方法
用全人血对 GalTKO.hCD46 猪（“2 基因”，n=3）和 GalTKO.hCD46 猪的肝脏进行离体灌注，这些猪还过表达 hEPRC、hTBM、hCD47 和 hHO-1（“6 基因”，n=4）。用去氨加压素（DDAVP）和氯膦酸盐脂质体预处理 6 基因猪，分别耗尽 vWF 和枯否细胞。
结果
2 基因组的平均灌注时间从 304（±148）分钟增加到 6 基因组的 856（±61）分钟（p=0.010）。2 基因组的肝素平均给药量从 8837 U/h 减少到 6 基因组的 1354 U/h（p=0.047）。血小板隔离在 6 基因组中延迟（p=0.070），而血栓烷 B2（TXB2，血小板活化标志物）水平在第一个小时内较低（p=0.044）（60 分钟时 401±124 与 2048±712）。6 基因组的凝血酶生成量（用 F1+2 水平测量）趋于较低（p=0.058）。
结论
hEPCR.hTBM.hCD47.hHO-1 盒与供体猪 DDAVP 和氯膦酸盐脂质体预处理相结合，与延长异种灌注肝脏的功能、减少凝血途径紊乱和减少 TXB2 产生有关，并反映了在猪到人模型中调节肝异种移植物损伤的显著进展。
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