Endothelial cell protection and complement inhibition in xenotransplantation: a novel in vitro model using whole blood

Banz, Yara; Cung, Trinh; Korchagina, Elena Y; Bovin, Nicolai V; Haeberli, André; Rieben, Robert (2005). Endothelial cell protection and complement inhibition in xenotransplantation: a novel in vitro model using whole blood. Xenotransplantation, 12(6), pp. 434-43. Hoboken, N.J.: Wiley 10.1111/j.1399-3089.2005.00239.x

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BACKGROUND: Studying the interactions between xenoreactive antibodies, complement and coagulation factors with the endothelium in hyperacute and acute vascular rejection usually necessitates the use of in vivo models. Conventional in vitro or ex vivo systems require either serum, plasma or anti-coagulated whole blood, making analysis of coagulation-mediated effects difficult. Here a novel in vitro microcarrier-based system for the study of endothelial cell (EC) activation and damage, using non-anticoagulated whole blood is described. Once established, the model was used to study the effect of the characterized complement- and coagulation inhibitor dextran sulfate (DXS, MW 5000) for its EC protective properties in a xenotransplantation setting. METHODS: Porcine aortic endothelial cells (PAEC), grown to confluence on microcarrier beads, were incubated with non-anticoagulated whole human blood until coagulation occurred or for a maximum of 90 min. PAEC-beads were either pre- or co-incubated with DXS. Phosphate buffered saline (PBS) experiments served as controls. Fluid phase and surface activation markers for complement and coagulation were analyzed as well as binding of DXS to PAEC-beads. RESULTS: Co- as well as pre-incubation of DXS, followed by washing of the beads, significantly prolonged time to coagulation from 39 +/- 12 min (PBS control) to 74 +/- 23 and 77 +/- 20 min, respectively (P < 0.005 vs. PBS). DXS treatment attenuated surface deposition of C1q, C4b/c, C3b/c and C5b-9 without affecting IgG or IgM deposition. Endothelial integrity, expressed by positivity for von Willebrand Factor, was maintained longer with DXS treatment. Compared with PBS controls, both pre- and co-incubation with DXS significantly prolonged activated partial thromboplastin time (>300 s, P < 0.05) and reduced production of thrombin-antithrombin complexes and fibrinopeptide A. Whilst DXS co-incubation completely blocked classical pathway complement activity (CH50 test) DXS pre-incubation or PBS control experiments showed no inhibition. DXS bound to PAEC-beads as visualized using fluorescein-labeled DXS. CONCLUSIONS: This novel in vitro microcarrier model can be used to study EC damage and the complex interactions with whole blood as well as screen ''endothelial protective'' substances in a xenotransplantation setting. DXS provides EC protection in this in vitro setting, attenuating damage of ECs as seen in hyperacute xenograft rejection.

Item Type:	Journal Article (Original Article)
Division/Institute:	04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > Forschungsbereich Mu50 > Forschungsgruppe Herz und Gefässe 04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > Unit Childrens Hospital > Thromboselabor Kinderklinik [discontinued]
UniBE Contributor:	Banz Wälti, Yara Sarah, Cung, Thi Thu Trinh, Häberli, André, Rieben, Robert
ISSN:	0908-665X
Publisher:	Wiley
Language:	English
Submitter:	Factscience Import
Date Deposited:	04 Oct 2013 15:12
Last Modified:	02 Mar 2023 23:23
Publisher DOI:	10.1111/j.1399-3089.2005.00239.x
PubMed ID:	16202066
Web of Science ID:	000232802000003
URI:	https://boris.unibe.ch/id/eprint/31663 (FactScience: 196311)