Hidetaka Hara1, Hayato Iwase2, Huy Nguyen3, Yuko Miyagawa4, Kasinath Kuravi5, Jeremy B Foote6, Will Eyestone5, Carol Phelps5, David Ayares5, David K C Cooper3. 1. Xenotransplantation Program, Department of Surgery, University of Alabama at Birmingham, Birmingham, AL, USA; Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh, Pittsburgh, PA, USA. Electronic address: harahjp@icloud.com. 2. Xenotransplantation Program, Department of Surgery, University of Alabama at Birmingham, Birmingham, AL, USA; Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh, Pittsburgh, PA, USA. 3. Xenotransplantation Program, Department of Surgery, University of Alabama at Birmingham, Birmingham, AL, USA. 4. Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh, Pittsburgh, PA, USA. 5. Revivicor, Blacksburg, VA, USA. 6. Department of Microbiology and Animal Resources Program, University of Alabama at Birmingham, Birmingham, AL, USA.
Abstract
BACKGROUND: Xenotransplantation is associated with an inflammatory response. The proinflammatory cytokine, TNF-α, downregulates the expression of thrombomodulin (TBM), and induces coagulation dysfunction. Although human (h) TBM-transgenic pigs (p) have been developed to reduce coagulation dysfunction, the effect of TNF-α on the expression of hTBM and its functional activity has not been fully investigated. The aims of this study were to investigate (i) whether the expression of hTBM on pig (p) cells is down-regulated during TNF-α stimulation, and (ii) whether cells from hTBM pigs regulate the inflammatory response. METHODS: TNF-α-producing T, B, and natural killer cells in blood from baboons with pig heart or kidney xenografts were investigated by flow cytometry. TNF-α staining in the grafts was detected by immunohistochemistry. Aortic endothelial cells (AECs) from GTKO/CD46 and GTKO/CD46/hTBM pigs were stimulated by hTNF-α, and the expression of the inflammatory/coagulation regulatory protein, TBM, was investigated. RESULTS: After pig organ xenotransplantation, there was a trend to increases in TNF-α-producing T and natural killer cells in the blood of baboons. In vitro observations demonstrated that after hTNF-α stimulation, there was a significant reduction in the expression of endogenous pTBM on pAECs, and a significant increase in the expression of inflammatory molecules. Blocking of NF-κB signaling significantly up-regulated pTBM expression, and suppressed the inflammatory response induced by hTNF-α in pAECs. Whereas the expression of pTBM mRNA was significantly reduced by hTNF-α stimulation, hTBM expression on the GTKO/CD46/hTBM pAECs was not affected. Furthermore, after hTNF-α stimulation, there was significant suppression of expression of inflammatory molecules on GTKO/CD46/hTBM pAECs compared to GTKO/CD46 pAECs. CONCLUSIONS: The stable expression of hTBM in pig cells may locally regulate the inflammatory response. This will help suppress the inflammatory response and prevent coagulation dysregulation after xenotransplantation.
BACKGROUND: Xenotransplantation is associated with an inflammatory response. The proinflammatory cytokine, TNF-α, downregulates the expression of thrombomodulin (TBM), and induces coagulation dysfunction. Although human (h) TBM-transgenic pigs (p) have been developed to reduce coagulation dysfunction, the effect of TNF-α on the expression of hTBM and its functional activity has not been fully investigated. The aims of this study were to investigate (i) whether the expression of hTBM on pig (p) cells is down-regulated during TNF-α stimulation, and (ii) whether cells from hTBM pigs regulate the inflammatory response. METHODS: TNF-α-producing T, B, and natural killer cells in blood from baboons with pig heart or kidney xenografts were investigated by flow cytometry. TNF-α staining in the grafts was detected by immunohistochemistry. Aortic endothelial cells (AECs) from GTKO/CD46 and GTKO/CD46/hTBM pigs were stimulated by hTNF-α, and the expression of the inflammatory/coagulation regulatory protein, TBM, was investigated. RESULTS: After pig organ xenotransplantation, there was a trend to increases in TNF-α-producing T and natural killer cells in the blood of baboons. In vitro observations demonstrated that after hTNF-α stimulation, there was a significant reduction in the expression of endogenous pTBM on pAECs, and a significant increase in the expression of inflammatory molecules. Blocking of NF-κB signaling significantly up-regulated pTBM expression, and suppressed the inflammatory response induced by hTNF-α in pAECs. Whereas the expression of pTBM mRNA was significantly reduced by hTNF-α stimulation, hTBM expression on the GTKO/CD46/hTBM pAECs was not affected. Furthermore, after hTNF-α stimulation, there was significant suppression of expression of inflammatory molecules on GTKO/CD46/hTBM pAECs compared to GTKO/CD46 pAECs. CONCLUSIONS: The stable expression of hTBM in pig cells may locally regulate the inflammatory response. This will help suppress the inflammatory response and prevent coagulation dysregulation after xenotransplantation.
Authors: Hayato Iwase; Hidetaka Hara; Mohamed Ezzelarab; Tao Li; Zhongqiang Zhang; Bingsi Gao; Hong Liu; Cassandra Long; Yi Wang; Amy Cassano; Edwin Klein; Carol Phelps; David Ayares; Abhinav Humar; Martin Wijkstrom; David K C Cooper Journal: Xenotransplantation Date: 2017-03-17 Impact factor: 3.907
Authors: S Crikis; X M Zhang; S Dezfouli; K M Dwyer; L M Murray-Segal; E Salvaris; C Selan; S C Robson; H H Nandurkar; P J Cowan; A J F d'Apice Journal: Am J Transplant Date: 2010-01-06 Impact factor: 8.086
Authors: Hayato Iwase; Burcin Ekser; Hidetaka Hara; Carol Phelps; David Ayares; David K C Cooper; Mohamed B Ezzelarab Journal: Xenotransplantation Date: 2013-11-05 Impact factor: 3.907
Authors: Anjan K Bongoni; Nikolai Klymiuk; Eckhard Wolf; David Ayares; Robert Rieben; Peter J Cowan Journal: Transplantation Date: 2016-09 Impact factor: 4.939
Authors: Guoqiang Zhang; Hayato Iwase; Qi Li; Takayuki Yamamoto; Abhijit Jagdale; Mohamed B Ezzelarab; David Ayares; David K C Cooper; Hidetaka Hara; Gangcheng Wang Journal: Front Immunol Date: 2021-12-08 Impact factor: 7.561