IMMUNOCYTOCHEMICAL LOCALIZATION OF ENDOGENOUS ANTI-THROMBIN-III IN THE VASCULATURE OF RAT-TISSUES REVEALS LOCATIONS OF ANTICOAGULANTLY ACTIVE HEPARAN-SULFATE PROTEOGLYCANS

We localized endogenous anti-thrombin III (ATIII) by light and electro n microscopic immunocytochemical staining in cryostat and ultra-thin f rozen sections of 10 different rat tissues, using rabbit alpha-human A TIII antibody that was shown to crossreact strongly with rat ATIII. EM immunocytochemical methods revealed discrete deposits of endogenous A TIII (absent after heparinase treatment), and thus by inference antico agulantly active heparan sulfate proteoglycans (HSPGs) at a resolution of 10-20 nm, or an order of magnitude better than autoradiography or LM. ATIII was found in variable amounts almost entirely in the subendo thelial space of blood vessels in various rat tissues. In kidney, ATII I was found immediately beneath the endothelium, in concentrated duste rs associated with the vascular basement membrane. Equally important i s the observed variation in expression of ATIII in the various tissues studied (i.e., kidney > liver, aorta, lung, spleen, adrenal > intesti ne, muscle, brain). On the basis of these observations, we confirm a m odel in which vascular abluminal and, perhaps to a much smaller extent , luminal anticoagulantly active HSPGs regulate coagulation mechanism activity, either by serving as a reserve of anticoagulant or by modula ting the ambient function of the coagulation cascade.-