Yuko Miwa1, Satoko Yazaki, Masaki Iwamoto, Shunichi Suzuki, Kenta Iwasaki, Masataka Haneda, Koji Yamamoto, Shoichi Maruyama, Akira Onishi, Takaaki Kobayashi. 1. 1 Department of Transplant Immunology, Nagoya University Graduate School of Medicine, Nagoya, Japan. 2 Prime Tech Ltd, Tsuchiura, Japan. 3 Department of Developmental Biology, National Institute of Agrobiological Sciences, Tsukuba, Japan. 4 Department of Transfusion Medicine, Nagoya University School of Medicine, Nagoya, Japan. 5 Department of Nephrology, Nagoya University Graduate School of Medicine, Nagoya, Japan. 6 Department Animal Science and Resources, Nihon University of College of Bioresource Sciences, Fujisawa, Japan. 7 Department of Surgery II, Nagoya University Graduate School of Medicine, Nagoya, Japan.
Abstract
BACKGROUND: For successful xenotransplantation, in addition to α1,3-galactosyltransferase gene-knockout and human complement regulatory protein (CD46, CD55, CD59) gene insertion, cloned pigs expressing human thrombomodulin (hTM) have been produced to solve the problem of molecular incompatibility in their coagulation system. Recombinant soluble hTM (S-hTM) which has been recently approved for treatment of disseminated intravascular coagulation might be potentially available. The purpose of this study is to examine the functional difference in endothelial cells between membrane-bound hTM (MB-hTM) and S-hTM and to elucidate effective strategy using both types of hTM. METHODS: The following factors regarding coagulation and inflammation were compared between hTM-expressing pig aortic endothelial cells (PAEC) derived from cloned pig and nontransgenic PAEC in the presence of S-hTM under tumor necrosis factor-α-activated conditions; (i) clotting time (ii) pig tissue factor (TF), (iii) pig E-selectin, (iv) direct prothrombinase activity, (v) activated protein C (APC), and (vi) prothrombinase activity. RESULTS: The MB-hTM significantly suppressed the expression of pig TF and E-selectin and direct prothrombinase activity in tumor necrosis factor-α-activated PAEC, suggesting strong anti-inflammatory effect, compared to S-hTM. In contrast, S-hTM had more potent capacity to inhibit thrombin generation and to produce APC than MB-hTM, although MB-hTM had the same level of capacity as human endothelial cells. CONCLUSIONS: It was speculated that S-hTM treatment would be of assistance during high-risk periods for excessive thrombin formation (e.g., ischemia reperfusion injury or severe infection/rejection). Considering the properties of MB-hTM exhibiting anti-inflammatory function as well as APC production, hTM-expressing cloned pigs might be indispensible to long-term stabilization of graft endothelial cells.
BACKGROUND: For successful xenotransplantation, in addition to α1,3-galactosyltransferase gene-knockout and human complement regulatory protein (CD46, CD55, CD59) gene insertion, cloned pigs expressing humanthrombomodulin (hTM) have been produced to solve the problem of molecular incompatibility in their coagulation system. Recombinant soluble hTM (S-hTM) which has been recently approved for treatment of disseminated intravascular coagulation might be potentially available. The purpose of this study is to examine the functional difference in endothelial cells between membrane-bound hTM (MB-hTM) and S-hTM and to elucidate effective strategy using both types of hTM. METHODS: The following factors regarding coagulation and inflammation were compared between hTM-expressing pig aortic endothelial cells (PAEC) derived from cloned pig and nontransgenic PAEC in the presence of S-hTM under tumor necrosis factor-α-activated conditions; (i) clotting time (ii) pigtissue factor (TF), (iii) pigE-selectin, (iv) direct prothrombinase activity, (v) activated protein C (APC), and (vi) prothrombinase activity. RESULTS: The MB-hTM significantly suppressed the expression of pigTF and E-selectin and direct prothrombinase activity in tumor necrosis factor-α-activated PAEC, suggesting strong anti-inflammatory effect, compared to S-hTM. In contrast, S-hTM had more potent capacity to inhibit thrombin generation and to produce APC than MB-hTM, although MB-hTM had the same level of capacity as human endothelial cells. CONCLUSIONS: It was speculated that S-hTM treatment would be of assistance during high-risk periods for excessive thrombin formation (e.g., ischemia reperfusion injury or severe infection/rejection). Considering the properties of MB-hTM exhibiting anti-inflammatory function as well as APC production, hTM-expressing cloned pigs might be indispensible to long-term stabilization of graft endothelial cells.
Authors: Parth M Patel; Margaret R Connolly; Taylor M Coe; Anthony Calhoun; Franziska Pollok; James F Markmann; Lars Burdorf; Agnes Azimzadeh; Joren C Madsen; Richard N Pierson Journal: Front Immunol Date: 2021-09-09 Impact factor: 7.561
Authors: Hidetaka Hara; Hayato Iwase; Huy Nguyen; Yuko Miyagawa; Kasinath Kuravi; Jeremy B Foote; Will Eyestone; Carol Phelps; David Ayares; David K C Cooper Journal: Cytokine Date: 2021-06-04 Impact factor: 3.861