Antiinflammatory and Anticoagulant Effects of Transgenic Expression of Human Thrombomodulin in Mice

Abstract

Thrombomodulin (TBM) is an important vascular anticoagulant that has species specific effects. When expressed as a transgene in pigs, human (h)TBM might abrogate thrombotic manifestations of acute vascular rejection (AVR) that occur when GalT-KO and/or complement regulator transgenic pig organs are transplanted to primates. hTBM transgenic mice were generated and characterized to determine whether this approach might show benefit without the development of deleterious hemorrhagic phenotypes. hTBM mice are viable and are not subject to spontaneous hemorrhage, although they have a prolonged bleeding time. They are resistant to intravenous collagen-induced pulmonary thromboembolism, stasis-induced venous thrombosis and pulmonary embolism. Cardiac grafts from hTBM mice to rats treated with cyclosporine in a model of AVR have prolonged survival compared to controls. hTBM reduced the inflammatory reaction in the vein wall in the stasis-induced thrombosis and mouse-to-rat xenograft models and reduced HMGB1 levels in LPS-treated mice. These results indicate that transgenic expression of hTBM has anticoagulant and antiinflammatory effects that are graft-protective in murine models.

ICAM-1–targeted thrombomodulin mitigates tissue factor–driven inflammatory thrombosis in a human endothelialized microfluidic model

“…22,24 TM itself demonstrates species-specific differences in the balance between anti-inflammatory and antithrombotic activity. 26,27 Finally, translation to human pathologies has been limited by the absence of species cross-reactivity of the murine therapeutic.…”

Thrombomodulin Gene Polymorphism (C1418T) is Associated with the Development of Coronary Allograft Vasculopathy

“…The G-33A and C-133A mutations in the promoter region may drive the down-regulation of the TM gene and soluble plasma TM levels, which may result in carotid atherosclerosis (5), venous thromboembolic disease, and the onset of acute myocardial infarction (6) in smokers. Additionally, an experimental murine model demonstrated that the transgenic expression of human TM has anti-coagulant and anti-inflammatory effects, resulting in the protection of allografts (7). In clinical identification, plasma-soluble TM levels are elevated in patients who have undergone hemodialysis, while levels are significantly decreased after kidney transplantation.…”

Protective Effects of Transgenic Human Endothelial Protein C Receptor Expression in Murine Models of Transplantation

“…hEPCR transgenic mice were generated using a mouse H-2K b promoterdriven construct derived from a vector previously used to generate human TBM (hTBM) transgenic mice (10). Total RNA isolated from the human endothelial cell line EAhy926 was reverse transcribed and used for PCR amplification of a 0.7-kbp hEPCR cDNA with primers hEPCR-F (5'-TTCTCGAGCAACTTCAGGATGTTGAC-3') and hEPCR-R (5'-TTGCGGCCGCGGAGAGTAATTAACATCGC-3').…”

“…Total RNA isolated from the human endothelial cell line EAhy926 was reverse transcribed and used for PCR amplification of a 0.7-kbp hEPCR cDNA with primers hEPCR-F (5'-TTCTCGAGCAACTTCAGGATGTTGAC-3') and hEPCR-R (5'-TTGCGGCCGCGGAGAGTAATTAACATCGC-3'). This cDNA was cloned in place of the 3.7-kbp hTBM cDNA in the hTBM vector and the resulting construct was prepared and microinjected into C57BL/6 fertilized oocytes as described (10). Mice were screened by PCR of tail-tip genomic DNA using primers hEPCR-F1 (5'-TTCTCGAGCAACTTCAGGATGTTGA-3') and hEPCR-R1 (5'-TTGCGGCCGCGGAGAGTAATTAAC-3').…”

“…Transgenic expression of human TBM (hTBM) in pigs has therefore been proposed as a protective strategy in solid organ xenotransplantation. We have shown that transgenic mice expressing hTBM possess an anticoagulant phenotype and are protected from transplant-associated thrombotic and inflammatory challenges (10). Pigs expressing hTBM have also been generated (11), but transplant data have not yet been reported.…”