The Anticoagulant Effect of PGI2S and tPA in Transgenic Umbilical Vein Endothelial Cells Is Linked to Up-Regulation of PKA and PKC

Jian-Hua Wang,Zhi-Min Zhong,Rong-Xin Liang,Min Zheng,Jian-Ming Deng,Zhe-Sheng Wen,Lin-Jing Yuan

doi:10.3390/ijms15022826

Abstract

The selection of vascular grafts for coronary artery bypass surgery is crucial for a positive outcome. This study aimed to establish a novel line of vascular endothelial cells with a potent anticoagulant effect. A lentiviral vector was used to stably transfect human umbilical vein endothelial cells (HUVECs) with PGI2S alone (HUVEC-PGI2S) or both PGI2S and tPA (HUVEC-PGI2S-tPA). Both HUVEC-PGI2S and HUVEC-PGI2S-tPA cells over-expressing PGI2S and tPA were compared to mock-transfected cells. The enzyme-linked immuno sorbent assay (ELISAs) demonstrated that the anticoagulation components, ATIII and PLG, were up-regulated and coagulation factor FVIII was down-regulated in both cell lines. QRT-PCR and western blotting demonstrated the vasodilation and platelet disaggregation proteins PKA, PKC, and PTGIR were up-regulated in both cell lines, but MAPK expression was not altered in either cell line. However, cell viability and colony formation assays and cell cycle analysis demonstrated that both cell lines had a lower rate of cell growth and induced G1 phase arrest. HUVEC-PGI2S and HUVEC-PGI2S-tPA cells have a potent anticoagulant effect and their use in vascular heterografts may decrease the risk of thrombosis.

Highlights

Since the first successful coronary artery bypass operation was performed by Goetz et al in 1960 [1,2], coronary artery bypass surgery, known as coronary artery bypass graft (CABG) surgery, has become the most common surgical method of treating coronary heart disease [3]
The protein expression levels of tPA were not increased in human umbilical vein endothelial cells (HUVECs)-PGI2 Synthase (PGI2S) cells but were 2.0-fold higher in HUVEC-PGI2S-tPA cells compared to HUVEC-mock cells (Figure 1B)
Protein expression was confirmed by Western blotting (Figure 4B). These results indicate that over-expression of PGI2S improves the anticoagulant effect of HUVEC cells, and that these effects were more potent when PGI2S was over-expressed in combination with tPA

Summary

Introduction

Since the first successful coronary artery bypass operation was performed by Goetz et al in 1960 [1,2], coronary artery bypass surgery, known as coronary artery bypass graft (CABG) surgery, has become the most common surgical method of treating coronary heart disease [3]. Four main classes of coronary artery bypass conduits are used: veins or arteries from autografts, allografts, heterografts, or artificial blood vessels [4,5]. Vascular grafts using autologous blood vessels are not ideal, due to the damage that the operation can cause and the potential for formation of vascular graft lesions. Allograft conduits have limited sources and introduce the problem of rejection by the recipient. Heterografts are the most favorable option, as they possess physically suitable structures and are readily available. Unprocessed heterografts can induce a strong rejection reaction in the recipient.

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Conclusion