Inhibition of the activation of Hageman factor (factor XII) by complement subcomponent C1q.

Rehmus, Esther H.; Greene, Bruce M.; Everson, Barbara; Ratnoff, Oscar D.

doi:10.1172/jci113100

Cited by 9 publications

(10 citation statements)

References 27 publications

(21 reference statements)

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“…It is a protein of 38 kDa when isolated from plasma and is distinct from the inhibitory property found in human endothelial cells described herein. A P2-glycoprotein in plasma (25) and the Clq subcomponent of the first component of complement (26) can inhibit the activation of HF. A substance elaborated by bovine aortic endothelial cells that inhibited procoagulant activity generated by human blood monocytes exposed to lipopolysaccharide was heat stable and had a much lower molecular mass (<1.5 kDa) (27).…”

Section: Discussionmentioning

confidence: 99%

Endothelial cells produce a substance that inhibits contact activation of coagulation by blocking the activation of Hageman factor.

Kleniewski

Donaldson²

1993

Proc. Natl. Acad. Sci. U.S.A.

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Human umbilical vein endothelial cells (HUVECs) produce a property that impairs the generation of coagulant and amidolytic activity initiated when normal human plasma is exposed to glass. This inhibitory property blocks the adsorption of Hageman factor (factor XII) to glass, thereby preventing the activation of Hageman factor, but does not impair the coagulant or amidolytic activity of already activatedHageman factor (factor XIIa). This property in HUVEC lysates could be neutralized by a purified preparation of Hageman factor but not by purified prekaflikrein or high molecular mass kininogen. A partially purified inhibitory fraction from cell lysates exhibited a single homogeneous band in SDS/PAGE of -22.5 kDa. Inhibitory activity was also found in concentrates of conditioned media from HUVECs, which also impaired the binding of Hageman factor to a surface; it may not be identical with that found in cell lysates.Human blood plasma contains several proteins that can regulate the coagulation mechanism, thus helping to maintain the fluidity of the blood in vivo. Hageman factor (HF; coagulation factor XII) can generate coagulant activity and promote the release of vasoactive polypeptides. When HF is activated (factor XIIa) this balance may be upset, but the endothelial lining of blood vessels can facilitate the action of antithrombin III and limit the generation of clot-promoting activity (1-3). In addition, thrombomodulin, a component of endothelial cell membranes (4), can modify the action of thrombin so that it activates protein C and promotes fibrinolysis. In the experiments to be described a property was found in endothelial cells that impairs the activation of HF upon a glass surface, thereby blocking contact activation of the coagulation mechanism. A fraction of endothelial cell lysates that contained this property exhibited a single homogeneous band in SDS/PAGE analysis. The property was also found in conditioned media from endothelial cell cultures and may provide another mechanism for the regulation of clotpromoting activity.MATERIALS AND METHODS Materials. Materials for culturing and harvesting endothelial cells, including type I collagenase, preservative-free heparin (H-3125 at 10,000 units/ml), soybean trypsin inhibitor, and p-nitrophenyl phosphate (PNP), were from Sigma; L-glutamine, Hanks' balanced salt solution (HBSS), NuSerum, and endothelial cell growth supplement were from Collaborative Research. Hepes buffer came from GIBCO; penicillin/streptomycin mixtures were from Flow Laboratories; gentamicin was from Elkins-Sinn (Cherry Hill, NJ); trypsin was from Worthington. D-Prolyl-L-phenylalanyl-Larginine p-nitroanilide dihydrochloride (S-2302), from KabiVitrum (Stockholm) and Helena Laboratories, was dissolved in 0.02 M Tris buffered saline (TBS), pH 7.4, containing 0.15 M sodium chloride. Amidolytic activity was quantified by a minor adaptation of the method of Svendsen et al. (5), as described in the legends for figures and table.Antibodies. Antibodies to purified human HF were produced in a ...

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Section: Discussionmentioning

confidence: 99%

Endothelial cells produce a substance that inhibits contact activation of coagulation by blocking the activation of Hageman factor.

Kleniewski

Donaldson²

1993

Proc. Natl. Acad. Sci. U.S.A.

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“…Inhibition of the activation of HF by Clq, platelet factor 4, and 82-glycoprotein I has been related to collagen-like sequences in their structure and, indeed, purified placental collagens inhibit activation of HF (11,15,(19)(20)(21)(22). But treatment of HUVEC culture supernates with collagenase did not decrease their inhibitory activity.…”

Section: Discussionmentioning

confidence: 99%

“…The effects of bovine trypsin and collagenase on HUVEC supernates were examined as described (1, 2). The possible presence of Clq, j2-glycoprotein I, platelet factor 4, or thrombospondin was examined by immunodiffusion and by testing the effect of specific heterologous antisera against these proteins on the capacity of HUVEC supernates to inhibit activation of HF, modifying earlier techniques by using sulfatides to activate HF (1,11).…”

Section: Methodsmentioning

confidence: 99%

“…In earlier experiments, activation of HF by ellagic acid was inhibited by the Clq subcomponent of the first component of complement (Cl) (11), by platelet factor 4 (19,20), by ,32-glycoprotein 1 (21,22), and by several species of collagen (15). A concentrate of HUVEC supernate containing 324 ,ug of protein per ml did not form a precipitin line upon immunodiffusion against antisera directed against Clq, platelet factor 4, thrombospondin, or 82-glycoprotein I.…”

Section: Properties Of the Inhibitory Activity Of Huvec Culturementioning

confidence: 99%

“…These antisera did not neutralize the inhibitory properties of HUVEC supernates against activation of HF (data not shown). Collagenase, which destroys the inhibitory properties of Clq (11) …”

Section: Properties Of the Inhibitory Activity Of Huvec Culturementioning

confidence: 99%

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Inhibition of the activation of Hageman factor (factor XII) by human vascular endothelial cell culture supernates.

Ratnoff

Everson

Embury

et al. 1991

Proc. Natl. Acad. Sci. U.S.A.

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The supernatant fluid (conditioned medium) of cultured human vascular endothelial cells inhibits activation of Hageman factor (factor XII), whether by ellagic acid, bovine brain sulfatides, or bismuth subgallate; inhibition appears to be a property of one or more proteins in the culture supernates.This phenomenon may contribute to maintaining the fluidity of circulating blood by inhibiting surface activation of the intrinsic pathway of coagulation.Circulating blood remains unclotted despite contact with the surfaces of blood cells and vascular endothelium. Suspensions of nucleated peripheral blood cells or platelets and their cell-depleted supernates inhibit activation of Hageman factor (HF; factor XII), phenomena associated with as yet unidentified proteins (1, 2). Several lines of evidence suggest that normal vascular endothelial cells, too, may actively impede blood clotting (3-6). The experiments to be described demonstrate that the supernatant fluids (conditioned media) of cultured human vascular endothelial cells inhibit activation of HF. Whether the responsible agent(s) is identical to those associated with peripheral blood cells is uncertain. MATERIALS AND METHODSHuman umbilical vein endothelial cells (HUVECs) and saphenous vein endothelial cells (Endotech, Indianapolis) were used at passages 1-4. The endothelial cells were plated onto T-75 tissue culture flasks (Corning) coated with human fibronectin (5-10 ,tg/cm2; Chemicon) and maintained in a medium of M-199 (Whittaker Bioproducts) containing 20% heat-inactivated fetal bovine serum (HyClone) supplemented with mixtures of porcine insulin (5 ,g/ml), transferrin (5 ng/ml), and selenium (5 ,g/ml) (Sigma); retinal-derived growth factor (80 ,ug/ml); and heparin (15 units/ml) (EndoRet HGF; Endotech) (7). Cells were refed every 3-4 days and grown to confluence, with a typical confluent density of 3-4 x 106 cells per flask. Identification of endothelial cells was confirmed by immunohistochemical staining for von Willebrand factor (8).The media from four flasks of confluent endothelial cells were removed and discarded. The cell monolayers were rinsed once with 25 ml of Dulbecco's phosphate-buffered saline (Whittaker Bioproducts) followed by two or three 5-min rinses with M-199 medium without serum. Ten milliliters of serum-free M-199 was added to each flask. Incubation was continued in humidified 5% C02/95% air for 24 ± 1 hr at 37°C, after which the medium of the replicate flasks was pooled and centrifuged for 10 min at 450 x g. The supernate was removed and stored at -70°C until used. Before testing, the culture supernates were dialyzed against 300 vol of barbital/saline buffer (BS) for 24 hr at 40C. In some experiments, the supernatant fluid was filtered through a Millex GV 0.22-,um filter (Millipore) unit attached to a polypropylene syringe as described (1).HUVEC supemates were concentrated 10-to 20-fold either by filtration through Centricon 30 or Centriprep 30 concentrators (Amicon) or by precipitation of the fraction soluble at 33% and insoluble a...

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Inhibition of the activation of hageman factor (factor XII) by eosinophils and eosinophilic constituents

et al. 1993

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Several syndromes characterized by striking eosinophilia may be complicated by thrombosis. The experiments described indicate that, paradoxically, eosinophils and certain of their constituents inhibit the activation of Hageman factor (HF, factor XII). In earlier studies, suspensions of mixed types of granulocytes, other nucleated peripheral blood cells, and platelets inhibited activation of Hageman factor by ellagic acid, glass, and sulfatides. After these cells were sedimented by centrifugation, the supernatant fluids were also inhibitory. No attempt had been made earlier to distinguish among different granulocytic species. In the present study, suspensions of eosinophils and the supernatant fluid after eosinophils had been separated by centrifugation inhibited activation of Hageman factor by ellagic acid. The protein concentration of that amount of supernatant fluid that inhibited activation by about half was 16 micrograms/ml, approximately the same as had been described for suspensions of peripheral blood mononuclear cells. Activation of Hageman factor by ellagic acid was also inhibited by certain constituents of eosinophils, including eosinophil peroxidase, eosinophil major basic protein and eosinophil cationic protein. Inhibition was not specific for ellagic acid-induced activation of Hageman factor, as inhibition was also observed with sulfatide-induced activation. Inhibition was presumably related to neutralization of the negative charge of activators of Hageman factor. Thus, bismuth subgallate, a particulate activator of Hageman factor, was no longer effective after it had been exposed to eosinophil cationic protein. The observations reported here raise the question of whether in vivo eosinophils modulate certain of the defense reactions ascribed to Hageman factor.

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Inhibition of the activation of Hageman factor (factor XII) by complement subcomponent C1q.

Cited by 9 publications

References 27 publications

Endothelial cells produce a substance that inhibits contact activation of coagulation by blocking the activation of Hageman factor.

Endothelial cells produce a substance that inhibits contact activation of coagulation by blocking the activation of Hageman factor.

Inhibition of the activation of Hageman factor (factor XII) by human vascular endothelial cell culture supernates.

Inhibition of the activation of hageman factor (factor XII) by eosinophils and eosinophilic constituents

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