Characterization of the association of tissue factor pathway inhibitor with human placenta.
OBJECTIVE
Tissue factor pathway inhibitor (TFPI) is an endothelial-associated inhibitor of blood coagulation. Because the mechanism for attachment of TFPI to endothelium is not clear, we investigated its association with human placenta.


RESULTS
Western blots demonstrate that treatment with phosphatidylinositol-specific phospholipase C (PIPLC) removes more placental TFPI than either PBS or heparin, a finding confirmed by immunohistochemistry. The amounts of heparin-releasable and PIPLC-releasable TFPI activity on placental endothelium were measured in placentas from 5 individuals. PIPLC removes >10-fold more TFPI activity from the placental fragments than 10 U/mL heparin and >100-fold more than 1 U/mL heparin. Pretreatment of the placental fragments with PIPLC increases the amount of heparin-releasable TFPI by approximately 3-fold. An antibody specific for the C-terminal region of TFPI recognizes PIPLC-releasable TFPI in Western blots.


CONCLUSIONS
GPI-anchored TFPI is the predominant form on placental endothelium. Heparin-releasable TFPI likely represents only a small portion of the total TFPI on endothelium that remains attached to cell-surface glycosaminoglycans after cleavage of the GPI anchor by endogenous enzymes. The predominance of GPI-anchored TFPI suggests that heparin infusion does not significantly redistribute TFPI within the vasculature. The intact C-terminus in GPI-anchored TFPI indicates it is not directly attached to a GPI anchor. Rather, it most likely associates with endothelium by binding to a GPI-anchored protein.
Mast
Alan E
AE
Research and Pathology Services, Department of Veterans Affairs, Memphis, Tenn 38104, USA. alan.mast3@med.va.gov
Acharya
Nayana
N
Malecha
Mark J
MJ
Hall
Connie L
CL
Dietzen
Dennis J
DJ
D000911:Antibodies, Monoclonal / Q000378:metabolism; D015153:Blotting, Western / Q000379:methods; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000737:chemistry; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D006493:Heparin / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008074:Lipoproteins / Q000032:analysis* / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D010920:Placenta / Q000737:chemistry* / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000032:analysis / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D013925:Thromboplastin / Q000037:antagonists & inhibitors* / Q000502:physiology*; D010738:Type C Phospholipases / Q000378:metabolism
D000911:Antibodies, Monoclonal; D017261:Glycosylphosphatidylinositols; D008074:Lipoproteins; D011994:Recombinant Proteins; C051928:lipoprotein-associated coagulation inhibitor; D006493:Heparin; D013925:Thromboplastin; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; D043265:Phosphatidylinositol Diacylglycerol-Lyase