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• W2007129229 endingPage "495" @default.
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• W2007129229 abstract "Petersen B, Ramackers W, Tiede A, Lucas-Hahn A, Herrmann D, Barg-Kues B, Schuettler W, Friedrich L, Schwinzer R, Winkler M, Niemann H. Pigs transgenic for human thrombomodulin have elevated production of activated protein C.Xenotransplantation 2009; 16: 486–495. © 2009 John Wiley & Sons A/S. Abstract: Background: The inability of porcine thrombomodulin (TM) to activate human anticoagulant protein C after pig-to-human xenotransplantation may lead to an aberrant activation of coagulation with microthrombosis and ultimately failure of the transplanted organ. Here, we describe the production of triple-transgenic pigs expressing hCD59/DAF and human thrombomodulin (hTM) and tested hTM-transgenic fibroblasts obtained from these pigs for their ability to activate human protein C in a new in vitro assay. Methods: Fibroblast cell cultures were established from a hCD59/DAF transgenic pig and transfected with a vector coding for hTM under transcriptional control of the CMV promoter. Transfected cells were analyzed for integration and expression of the hTM vector by PCR and RT-PCR. One cell clone was used as donor for somatic cell nuclear transfer to produce triple transgenic (CD59/DAF/hTM) pigs. Pigs were characterized in detail with regard to hTM integration and expression by PCR, RT-PCR, Northern blot, Western blot, immunostaining, and FACS analysis. Fibroblasts from hTM-transgenic pigs were analyzed in a new in vitro hTM coactivity assay to assess the production of activated protein C (aPC) and results were compared to those from wild-type controls. Results: In total, 1040 cloned transgenic embryos were transferred to eight recipients. Five recipients remained pregnant and delivered 22 piglets. Expression of hTM was detected in all xenorelavant organs including heart, liver, kidney, lung, and pancreas. The lowest levels of expression were found in lung and liver while all animals showed a strong, but frequently patchy expression pattern of hTM in heart, kidney, and pancreas. The hTM cofactor activity (ranging on a scale from 5–18 U/105 cells/2h) was significantly higher in fibroblasts of hTM-transgenic clones compared to wild-type porcine fibroblasts (1.7 U/105 cells/2h). Conclusions: For the first time, healthy hTM-transgenic pigs could be successfully generated by somatic cell nuclear transfer. hTM can be expressed in porcine organs without perturbation of the porcine coagulation system. hTM-transgenic porcine fibroblasts showed elevated aPC production in an in vitro hTM coactivity assay. These findings warrant further work on the control of the xenogenic activation of coagulation by transgenic approaches." @default.
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• W2007129229 date "2009-11-01" @default.
• W2007129229 modified "2023-10-02" @default.
• W2007129229 title "Pigs transgenic for human thrombomodulin have elevated production of activated protein C" @default.
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• W2007129229 doi "https://doi.org/10.1111/j.1399-3089.2009.00537.x" @default.
• W2007129229 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/20042048" @default.
• W2007129229 hasPublicationYear "2009" @default.
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