title: Quantitative analysis of thrombomodulin-mediated conversion of protein C to APC: translation from in vitro to in vivo.
abstract: Thrombomodulin-bound thrombin cleaves protein C (PC) zymogen in blood plasma producing activated protein C (APC), which exerts anti-coagulant, anti-inflammatory, anti-apoptotic and CNS-protective effects. Recombinant APC and thrombomodulin (TM) are both in clinical studies for management of acute conditions including sepsis. Methods that permit accurate measurement of APC in plasma are needed for clinical monitoring and mechanistic studies in animal models. However, the two existing methods require either long incubation periods with substrate, resulting in high background or they also recognize protein C inhibitor (PCI) complexed with APC (APC:PCI), which convolutes analysis of the amount of APC generated. Here we describe a robust quantitative in vivo assay that measures APC generation at both low levels of human protein C seen in chronic inflammatory disease and at physiological levels that shows a >99% fit with in vitro data.
authors: Department of Pharmacology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA. rcarne@mail.med.upenn.edu
authors: Ronald
authors: R
authors: Carnemolla
authors: Kruti Rajan
authors: KR
authors: Patel
authors: Sergei
authors: S
authors: Zaitsev
authors: Douglas B
authors: DB
authors: Cines
authors: Charles T
authors: CT
authors: Esmon
authors: Vladimir R
authors: VR
authors: Muzykantov
mesh_terms: D000818:Animals; D002417:Cattle; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000097:blood / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007700:Kinetics; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D011485:Protein Binding; D011486:Protein C / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism* / Q000494:pharmacology; D018180:Thrombomodulin / Q000378:metabolism*
chemical_list: D004792:Enzyme Precursors; D011486:Protein C; D018180:Thrombomodulin; D013917:Thrombin